Effect of Light Modification by Shading Nets on Yield, Composition, and Antioxidant Activity of Lavandula angustifolia Mill. Essential Oil

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Review of the Manustript entitled:

 Effect of light modification by shading nets on yield, composi-tion, and antioxidant activity of Lavandula angustifolia Mill. es-sential oil

In this manuscript, the authors present the results of a study investigating the influence of different shading nets (pearl, red, and blue) on the yield, composition, and antioxidant activity of Lavandula angustifolia essential oil isolated from leaves and flowers. Plants cultivated in the open field were used as a control. The manuscript is well written, but the following sections have the following shortcomings.

M&M:

The description of the experiment is very unclear and poorly written. The authors should include a schematic diagram to explain the experimental design, treatments, and the layout of the experiment.

The data regarding the experimental design is missing. What was the plot size? How many plants represented each replication of each treatment? Did the authors measure water consumption for irrigation for each treatment? The data on temperature under each shading net is also missing.

Was photosynthetically active radiation measured continuously or every third or fourth day (Figure 1)?

The title of Table 1 is unclear. What does ‘Average, max value’ mean?

The description of the statistical analysis is incorrect. Why did the authors use two-way analysis of variance? How many factors were included in the study?

The authors stated: “All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.” Where are the standard deviations? Only the results of chemical analysis are presented in this form.

The results of the inflorescences harvested do not belong in the Materials and Methods section.

Results: The main drawback of the article is that it does not include a qualitative analysis of the spectrum produced under each individual shade net.

Discussion:

In the Discussion section, the authors only compare their results with previously published findings in the literature. There is a lack of in-depth explanation of the biochemical mechanisms triggered by different light qualities.

The author has drawn the conclusions from the results.

According to the aforementioned findings, I suggest that the manuscript is not accepted for publication in the journal Plants (MDPI) in its current form.

Author Response

 

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13].

 

We exclude old reference [13] with L. dentata, and include new reference [13] with L. angustifolia.

 

Mijatovic, S.; Stankovic, J.A.; Calovski, I.C.; Dubljanin, E.; Pljevljakusic, D.; Bigovic, D.; Dzamic, A. Antifungal activity of Lavandula angustifolia essential oil against Candida albicans: Time-kill study on pediatric sputum isolates. Molecules. 2022, 27(19), 6300. doi: 10.3390/molecules27196300.   

            

Lavender EO has antifungal activities, which supports their promising fungicidal potential. [13].

 

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

We have corrected it. Now stay lavender

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

Yes it is mistake. We have revised it.

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

We accept your recommendation and remove these parts

Inflorescences were harvested once, with fresh yield (spike-like inflorescences) of 1.320 t/ha under the pearl net, 1.302 t/ha under the red net, 0.72 t/ha under the blue net, and 0.78 t/ha in the open field. The fresh-to-dry mass ratio was uniform (1.71–1.81).

In September, when plants had formed more branches (13.7 in the open field and 14.1–16.4 under shading nets), green biomass was harvested once. Yields reached 5.6 t/ha in the open field and 6.1–7.9 t/ha under photoselective nets.   

and replace with new one

Flowering stems developed in late June. Harvesting of spike-like inflorescences was performed once, when the majority of flowers were fully open. The cut spike-like inflorescences were placed in a shaded and well-ventilated area for drying. The vegetative aboveground biomass (leaves and stems) was harvested later, in the last decade of September. The shoots were cut when 13–14 branches had formed in plants grown under the blue net and in the open field (control), and 15–17 branches in plants grown under the pearl and red nets. At this stage, the vegetative biomass of the plants was sufficiently developed to withstand cutting. The apical parts of the branches about10 cm in length, were cut.

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?

 

• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.

We exclude third paragraph and replace with new one.

In July shading with pearl, red, and blue nets decreased the mean PPFD by 45%, 41%, and 50%, respectively, compared to the unshaded (open field) control condition. Also, shading substantially reduced light availability compared to the unshaded open field condition-control (996 W/m²), with mean irradiance decreased by 40% under the pearl net, 35% under the red net, and 44% under the blue net.

Overall, photo-selective netting represents a cost-effective strategy for manipulating crop microclimate conditions, plant grown, allowing the regulation of not only yield but also quality and functional or bioactive plant properties.

 

• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?

ʹHerbs' we replace with 'vegetativeʹ mass (leaves and steams) ……

• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).

We are accepting your suggestion. Significant differences (symbols, a,b, ab)between treatments  incorporated in Table (as in Table 3).

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

 

• First paragraph after table 2 - please move to Discussion section.

Yes, we do it and add totally new paragraph

Harvesting of spike-like lavender inflorescences was carried out in late June. As the plants were in their second year of growth, harvesting was performed once, when the majority of flowers were fully open. The yield of fresh inflorescences obtained from plants grown in the open field (0.78 t·ha⁻¹) and under the blue net (0.72 t·ha⁻¹) was significantly lower compared to the values recorded under the red and pearl nets (1.302-1.320 t·ha⁻¹). In addition to their positive effects on the morphological traits of lavender plants, shading with red and pearl nets also positively affected the yield of fresh and dry inflorescences per unit area. Shading resulted in a more uniform fresh-to-dry mass ratio of spike-like inflorescences, regardless of the color of the applied photoselective nets.

A single harvest of the aboveground vegetative biomass (leaves and stems) of lavender plants was carried out in the last decade of September. The fresh biomass yield of plants grown under the blue net (6.10 t·ha¹)   was very significantly lower (p < 0.01) than that of plants grown under the pearl net (7.90 t·ha⁻¹). The positive effect of the red and pearl nets on lavender was even more pronounced when compared with plants grown without shading in the open field. The use of photoselective nets, especially red and pearl ones, represents an effective agrotechnical approach that enhances both vegetative and generative biomass production in lavender, with potential positive effects on quality. The altered light quality caused by shading affects plant performance, often resulting in higher yields for medicinal plants. However, the specific effects depend on the color and type of the photoselective net, as these determine both the light spectrum and intensity reaching the canopy.

 

• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values

Yes we accept your recommendation and remove standard deviation

• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.

             It’s a typist mistake …. This is correct…….linalool (24.7–27.3%),

 

• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit

RI lit (the retention index) was determined according to the different references (a,b,c,d)

^aAdams RP (2007). Identification of essential oil components by gas chromatography /mass spectrometry, Carol Stream, Allured Publishing Co., Illinois, USA.

^bBenkaci-Ali, F.; Baaliouamer, A.; Meklati, B.Y.; Chemat, F., Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation, Flavour Fragr. J., 2007, 22, 2, 148-153, https://doi.org/10.1002/ffj.1773.

^cSaroglou, V.; Dorizas, N.; Kypriotakis, Z.; Skaltsa, H.D., Analysis of the essential oil composition of eight Anthemis species from Greece, J. Chromatogr. A, 2006, 1104, 1-2, 313-322, https://doi.org/10.1016/j.chroma.2005.11.087 .

^dZouari, N.; Ayadi, I.; Fakhfakh, N.; Rebai, A.; Zouari, S., Variations of chemical composition of essential oils in wild-populations of Thymus algeriensis Boiss et Reut., a North African endemic species, Lipids in Health and Desease, 2012, 11, 1, 28-39, https://doi.org/10.1186/1476-511X-11-28.

 

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

We adopt your recommendation and exclude some irrelevant sentences in discussion and add new one

To the best of our knowledge, studies addressing UVB:PAR or UVA:PAR ratios, as well as plant responses to these ratios under horticultural growing conditions, are lacking or extremely scarce. Even studies that focus exclusively on the effects of UV radiation on the growth of horticultural crops are rare. UV radiation is associated with the accumulation of plant secondary metabolites (phenolic compounds, including flavonoids, etc.), which are related to plant color, taste, and perceived health-promoting attributes. 

High solar radiation and elevated temperatures during the summer months can negatively affect both plant yield and quality. The incorporation of light-dispersive and reflective chromatic additives into photo-selective nets transforms direct sunlight into diffuse radiation, enabling deeper light penetration into the inner plant canopy. The radiometric properties of photo-selective nets are determined by their porosity and color. During cultivation, modifications in light quality beneath photo-selective shade nets can positively influence yield, quality traits, and phytochemical composition of aromatic and medicinal plants [37].

Color shade nets alter light spectra, triggering biochemical changes in medicinal plants by manipulating photoreceptors (like phytochromes and cryptochromes) that regulate secondary metabolite (phenols, flavonoids, essential oils) production, photosynthesis, antioxidant activity, and pigmentation, often boosting beneficial compounds by shifting energy balance and influencing stress response pathways for enhanced medicinal quality [38]. 

 

We are also add new paragraph in discussion

Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications

 

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

You are right; we have removed part of the last paragraph to eliminate any ambiguities.

 

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

We have restructured the Conclusions in accordance with your comments.

Based on the results, shading lavender with pearl and red nets increased essential oil (EO) yield in both plant parts compared to non-shaded plants. The most abundant EO component was 1,8-cineole (eucalyptol); the highest content in flower essential oil (LAFEO, 32.2%) was obtained from plants covered with red nets, whereas the highest content in leaf essential oil (LALEO, 39.8%) was recorded in plants shaded with blue nets. Shading with colored nets differentially affected the content of individual essential oil components. A higher camphor content in flowers, which negatively affects oil quality, was observed in non-shaded plants. However, the essential oil from non-shaded plants was characterized by a higher antioxidant capacity than that from all shaded treatments. Nevertheless, this preliminary study, which evaluated both the yield and quality of lavender essential oil under colored shading nets, highlights the strong potential of this species for expanded production in southern Serbia.

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13].

 

We exclude old reference [13] with L. dentata, and include new reference [13] with L. angustifolia.

 

Mijatovic, S.; Stankovic, J.A.; Calovski, I.C.; Dubljanin, E.; Pljevljakusic, D.; Bigovic, D.; Dzamic, A. Antifungal activity of Lavandula angustifolia essential oil against Candida albicans: Time-kill study on pediatric sputum isolates. Molecules. 2022, 27(19), 6300. doi: 10.3390/molecules27196300.   

            

Lavender EO has antifungal activities, which supports their promising fungicidal potential. [13].

 

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

We have corrected it. Now stay lavender

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

Yes it is mistake. We have revised it.

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

We accept your recommendation and remove these parts

Inflorescences were harvested once, with fresh yield (spike-like inflorescences) of 1.320 t/ha under the pearl net, 1.302 t/ha under the red net, 0.72 t/ha under the blue net, and 0.78 t/ha in the open field. The fresh-to-dry mass ratio was uniform (1.71–1.81).

In September, when plants had formed more branches (13.7 in the open field and 14.1–16.4 under shading nets), green biomass was harvested once. Yields reached 5.6 t/ha in the open field and 6.1–7.9 t/ha under photoselective nets.   

and replace with new one

Flowering stems developed in late June. Harvesting of spike-like inflorescences was performed once, when the majority of flowers were fully open. The cut spike-like inflorescences were placed in a shaded and well-ventilated area for drying. The vegetative aboveground biomass (leaves and stems) was harvested later, in the last decade of September. The shoots were cut when 13–14 branches had formed in plants grown under the blue net and in the open field (control), and 15–17 branches in plants grown under the pearl and red nets. At this stage, the vegetative biomass of the plants was sufficiently developed to withstand cutting. The apical parts of the branches about10 cm in length, were cut.

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?

 

• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.

We exclude third paragraph and replace with new one.

In July shading with pearl, red, and blue nets decreased the mean PPFD by 45%, 41%, and 50%, respectively, compared to the unshaded (open field) control condition. Also, shading substantially reduced light availability compared to the unshaded open field condition-control (996 W/m²), with mean irradiance decreased by 40% under the pearl net, 35% under the red net, and 44% under the blue net.

Overall, photo-selective netting represents a cost-effective strategy for manipulating crop microclimate conditions, plant grown, allowing the regulation of not only yield but also quality and functional or bioactive plant properties.

 

• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?

ʹHerbs' we replace with 'vegetativeʹ mass (leaves and steams) ……

• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).

We are accepting your suggestion. Significant differences (symbols, a,b, ab)between treatments  incorporated in Table (as in Table 3).

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

 

• First paragraph after table 2 - please move to Discussion section.

Yes, we do it and add totally new paragraph

Harvesting of spike-like lavender inflorescences was carried out in late June. As the plants were in their second year of growth, harvesting was performed once, when the majority of flowers were fully open. The yield of fresh inflorescences obtained from plants grown in the open field (0.78 t·ha⁻¹) and under the blue net (0.72 t·ha⁻¹) was significantly lower compared to the values recorded under the red and pearl nets (1.302-1.320 t·ha⁻¹). In addition to their positive effects on the morphological traits of lavender plants, shading with red and pearl nets also positively affected the yield of fresh and dry inflorescences per unit area. Shading resulted in a more uniform fresh-to-dry mass ratio of spike-like inflorescences, regardless of the color of the applied photoselective nets.

A single harvest of the aboveground vegetative biomass (leaves and stems) of lavender plants was carried out in the last decade of September. The fresh biomass yield of plants grown under the blue net (6.10 t·ha¹)   was very significantly lower (p < 0.01) than that of plants grown under the pearl net (7.90 t·ha⁻¹). The positive effect of the red and pearl nets on lavender was even more pronounced when compared with plants grown without shading in the open field. The use of photoselective nets, especially red and pearl ones, represents an effective agrotechnical approach that enhances both vegetative and generative biomass production in lavender, with potential positive effects on quality. The altered light quality caused by shading affects plant performance, often resulting in higher yields for medicinal plants. However, the specific effects depend on the color and type of the photoselective net, as these determine both the light spectrum and intensity reaching the canopy.

 

• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values

Yes we accept your recommendation and remove standard deviation

• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.

             It’s a typist mistake …. This is correct…….linalool (24.7–27.3%),

 

• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit

RI lit (the retention index) was determined according to the different references (a,b,c,d)

^aAdams RP (2007). Identification of essential oil components by gas chromatography /mass spectrometry, Carol Stream, Allured Publishing Co., Illinois, USA.

^bBenkaci-Ali, F.; Baaliouamer, A.; Meklati, B.Y.; Chemat, F., Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation, Flavour Fragr. J., 2007, 22, 2, 148-153, https://doi.org/10.1002/ffj.1773.

^cSaroglou, V.; Dorizas, N.; Kypriotakis, Z.; Skaltsa, H.D., Analysis of the essential oil composition of eight Anthemis species from Greece, J. Chromatogr. A, 2006, 1104, 1-2, 313-322, https://doi.org/10.1016/j.chroma.2005.11.087 .

^dZouari, N.; Ayadi, I.; Fakhfakh, N.; Rebai, A.; Zouari, S., Variations of chemical composition of essential oils in wild-populations of Thymus algeriensis Boiss et Reut., a North African endemic species, Lipids in Health and Desease, 2012, 11, 1, 28-39, https://doi.org/10.1186/1476-511X-11-28.

 

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

We adopt your recommendation and exclude some irrelevant sentences in discussion and add new one

To the best of our knowledge, studies addressing UVB:PAR or UVA:PAR ratios, as well as plant responses to these ratios under horticultural growing conditions, are lacking or extremely scarce. Even studies that focus exclusively on the effects of UV radiation on the growth of horticultural crops are rare. UV radiation is associated with the accumulation of plant secondary metabolites (phenolic compounds, including flavonoids, etc.), which are related to plant color, taste, and perceived health-promoting attributes. 

High solar radiation and elevated temperatures during the summer months can negatively affect both plant yield and quality. The incorporation of light-dispersive and reflective chromatic additives into photo-selective nets transforms direct sunlight into diffuse radiation, enabling deeper light penetration into the inner plant canopy. The radiometric properties of photo-selective nets are determined by their porosity and color. During cultivation, modifications in light quality beneath photo-selective shade nets can positively influence yield, quality traits, and phytochemical composition of aromatic and medicinal plants [37].

Color shade nets alter light spectra, triggering biochemical changes in medicinal plants by manipulating photoreceptors (like phytochromes and cryptochromes) that regulate secondary metabolite (phenols, flavonoids, essential oils) production, photosynthesis, antioxidant activity, and pigmentation, often boosting beneficial compounds by shifting energy balance and influencing stress response pathways for enhanced medicinal quality [38]. 

 

We are also add new paragraph in discussion

Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications

 

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

You are right; we have removed part of the last paragraph to eliminate any ambiguities.

 

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

We have restructured the Conclusions in accordance with your comments.

Based on the results, shading lavender with pearl and red nets increased essential oil (EO) yield in both plant parts compared to non-shaded plants. The most abundant EO component was 1,8-cineole (eucalyptol); the highest content in flower essential oil (LAFEO, 32.2%) was obtained from plants covered with red nets, whereas the highest content in leaf essential oil (LALEO, 39.8%) was recorded in plants shaded with blue nets. Shading with colored nets differentially affected the content of individual essential oil components. A higher camphor content in flowers, which negatively affects oil quality, was observed in non-shaded plants. However, the essential oil from non-shaded plants was characterized by a higher antioxidant capacity than that from all shaded treatments. Nevertheless, this preliminary study, which evaluated both the yield and quality of lavender essential oil under colored shading nets, highlights the strong potential of this species for expanded production in southern Serbia.

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13].

 

We exclude old reference [13] with L. dentata, and include new reference [13] with L. angustifolia.

 

Mijatovic, S.; Stankovic, J.A.; Calovski, I.C.; Dubljanin, E.; Pljevljakusic, D.; Bigovic, D.; Dzamic, A. Antifungal activity of Lavandula angustifolia essential oil against Candida albicans: Time-kill study on pediatric sputum isolates. Molecules. 2022, 27(19), 6300. doi: 10.3390/molecules27196300.   

            

Lavender EO has antifungal activities, which supports their promising fungicidal potential. [13].

 

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

We have corrected it. Now stay lavender

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

Yes it is mistake. We have revised it.

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

We accept your recommendation and remove these parts

Inflorescences were harvested once, with fresh yield (spike-like inflorescences) of 1.320 t/ha under the pearl net, 1.302 t/ha under the red net, 0.72 t/ha under the blue net, and 0.78 t/ha in the open field. The fresh-to-dry mass ratio was uniform (1.71–1.81).

In September, when plants had formed more branches (13.7 in the open field and 14.1–16.4 under shading nets), green biomass was harvested once. Yields reached 5.6 t/ha in the open field and 6.1–7.9 t/ha under photoselective nets.   

and replace with new one

Flowering stems developed in late June. Harvesting of spike-like inflorescences was performed once, when the majority of flowers were fully open. The cut spike-like inflorescences were placed in a shaded and well-ventilated area for drying. The vegetative aboveground biomass (leaves and stems) was harvested later, in the last decade of September. The shoots were cut when 13–14 branches had formed in plants grown under the blue net and in the open field (control), and 15–17 branches in plants grown under the pearl and red nets. At this stage, the vegetative biomass of the plants was sufficiently developed to withstand cutting. The apical parts of the branches about10 cm in length, were cut.

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?

 

• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.

We exclude third paragraph and replace with new one.

In July shading with pearl, red, and blue nets decreased the mean PPFD by 45%, 41%, and 50%, respectively, compared to the unshaded (open field) control condition. Also, shading substantially reduced light availability compared to the unshaded open field condition-control (996 W/m²), with mean irradiance decreased by 40% under the pearl net, 35% under the red net, and 44% under the blue net.

Overall, photo-selective netting represents a cost-effective strategy for manipulating crop microclimate conditions, plant grown, allowing the regulation of not only yield but also quality and functional or bioactive plant properties.

 

• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?

ʹHerbs' we replace with 'vegetativeʹ mass (leaves and steams) ……

• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).

We are accepting your suggestion. Significant differences (symbols, a,b, ab)between treatments  incorporated in Table (as in Table 3).

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

 

• First paragraph after table 2 - please move to Discussion section.

Yes, we do it and add totally new paragraph

Harvesting of spike-like lavender inflorescences was carried out in late June. As the plants were in their second year of growth, harvesting was performed once, when the majority of flowers were fully open. The yield of fresh inflorescences obtained from plants grown in the open field (0.78 t·ha⁻¹) and under the blue net (0.72 t·ha⁻¹) was significantly lower compared to the values recorded under the red and pearl nets (1.302-1.320 t·ha⁻¹). In addition to their positive effects on the morphological traits of lavender plants, shading with red and pearl nets also positively affected the yield of fresh and dry inflorescences per unit area. Shading resulted in a more uniform fresh-to-dry mass ratio of spike-like inflorescences, regardless of the color of the applied photoselective nets.

A single harvest of the aboveground vegetative biomass (leaves and stems) of lavender plants was carried out in the last decade of September. The fresh biomass yield of plants grown under the blue net (6.10 t·ha¹)   was very significantly lower (p < 0.01) than that of plants grown under the pearl net (7.90 t·ha⁻¹). The positive effect of the red and pearl nets on lavender was even more pronounced when compared with plants grown without shading in the open field. The use of photoselective nets, especially red and pearl ones, represents an effective agrotechnical approach that enhances both vegetative and generative biomass production in lavender, with potential positive effects on quality. The altered light quality caused by shading affects plant performance, often resulting in higher yields for medicinal plants. However, the specific effects depend on the color and type of the photoselective net, as these determine both the light spectrum and intensity reaching the canopy.

 

• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values

Yes we accept your recommendation and remove standard deviation

• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.

             It’s a typist mistake …. This is correct…….linalool (24.7–27.3%),

 

• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit

RI lit (the retention index) was determined according to the different references (a,b,c,d)

^aAdams RP (2007). Identification of essential oil components by gas chromatography /mass spectrometry, Carol Stream, Allured Publishing Co., Illinois, USA.

^bBenkaci-Ali, F.; Baaliouamer, A.; Meklati, B.Y.; Chemat, F., Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation, Flavour Fragr. J., 2007, 22, 2, 148-153, https://doi.org/10.1002/ffj.1773.

^cSaroglou, V.; Dorizas, N.; Kypriotakis, Z.; Skaltsa, H.D., Analysis of the essential oil composition of eight Anthemis species from Greece, J. Chromatogr. A, 2006, 1104, 1-2, 313-322, https://doi.org/10.1016/j.chroma.2005.11.087 .

^dZouari, N.; Ayadi, I.; Fakhfakh, N.; Rebai, A.; Zouari, S., Variations of chemical composition of essential oils in wild-populations of Thymus algeriensis Boiss et Reut., a North African endemic species, Lipids in Health and Desease, 2012, 11, 1, 28-39, https://doi.org/10.1186/1476-511X-11-28.

 

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

We adopt your recommendation and exclude some irrelevant sentences in discussion and add new one

To the best of our knowledge, studies addressing UVB:PAR or UVA:PAR ratios, as well as plant responses to these ratios under horticultural growing conditions, are lacking or extremely scarce. Even studies that focus exclusively on the effects of UV radiation on the growth of horticultural crops are rare. UV radiation is associated with the accumulation of plant secondary metabolites (phenolic compounds, including flavonoids, etc.), which are related to plant color, taste, and perceived health-promoting attributes. 

High solar radiation and elevated temperatures during the summer months can negatively affect both plant yield and quality. The incorporation of light-dispersive and reflective chromatic additives into photo-selective nets transforms direct sunlight into diffuse radiation, enabling deeper light penetration into the inner plant canopy. The radiometric properties of photo-selective nets are determined by their porosity and color. During cultivation, modifications in light quality beneath photo-selective shade nets can positively influence yield, quality traits, and phytochemical composition of aromatic and medicinal plants [37].

Color shade nets alter light spectra, triggering biochemical changes in medicinal plants by manipulating photoreceptors (like phytochromes and cryptochromes) that regulate secondary metabolite (phenols, flavonoids, essential oils) production, photosynthesis, antioxidant activity, and pigmentation, often boosting beneficial compounds by shifting energy balance and influencing stress response pathways for enhanced medicinal quality [38]. 

 

We are also add new paragraph in discussion

Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications

 

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

You are right; we have removed part of the last paragraph to eliminate any ambiguities.

 

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

We have restructured the Conclusions in accordance with your comments.

Based on the results, shading lavender with pearl and red nets increased essential oil (EO) yield in both plant parts compared to non-shaded plants. The most abundant EO component was 1,8-cineole (eucalyptol); the highest content in flower essential oil (LAFEO, 32.2%) was obtained from plants covered with red nets, whereas the highest content in leaf essential oil (LALEO, 39.8%) was recorded in plants shaded with blue nets. Shading with colored nets differentially affected the content of individual essential oil components. A higher camphor content in flowers, which negatively affects oil quality, was observed in non-shaded plants. However, the essential oil from non-shaded plants was characterized by a higher antioxidant capacity than that from all shaded treatments. Nevertheless, this preliminary study, which evaluated both the yield and quality of lavender essential oil under colored shading nets, highlights the strong potential of this species for expanded production in southern Serbia.

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13].

 

We exclude old reference [13] with L. dentata, and include new reference [13] with L. angustifolia.

 

Mijatovic, S.; Stankovic, J.A.; Calovski, I.C.; Dubljanin, E.; Pljevljakusic, D.; Bigovic, D.; Dzamic, A. Antifungal activity of Lavandula angustifolia essential oil against Candida albicans: Time-kill study on pediatric sputum isolates. Molecules. 2022, 27(19), 6300. doi: 10.3390/molecules27196300.   

            

Lavender EO has antifungal activities, which supports their promising fungicidal potential. [13].

 

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

We have corrected it. Now stay lavender

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

Yes it is mistake. We have revised it.

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

We accept your recommendation and remove these parts

Inflorescences were harvested once, with fresh yield (spike-like inflorescences) of 1.320 t/ha under the pearl net, 1.302 t/ha under the red net, 0.72 t/ha under the blue net, and 0.78 t/ha in the open field. The fresh-to-dry mass ratio was uniform (1.71–1.81).

In September, when plants had formed more branches (13.7 in the open field and 14.1–16.4 under shading nets), green biomass was harvested once. Yields reached 5.6 t/ha in the open field and 6.1–7.9 t/ha under photoselective nets.   

and replace with new one

Flowering stems developed in late June. Harvesting of spike-like inflorescences was performed once, when the majority of flowers were fully open. The cut spike-like inflorescences were placed in a shaded and well-ventilated area for drying. The vegetative aboveground biomass (leaves and stems) was harvested later, in the last decade of September. The shoots were cut when 13–14 branches had formed in plants grown under the blue net and in the open field (control), and 15–17 branches in plants grown under the pearl and red nets. At this stage, the vegetative biomass of the plants was sufficiently developed to withstand cutting. The apical parts of the branches about10 cm in length, were cut.

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?

 

• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.

We exclude third paragraph and replace with new one.

In July shading with pearl, red, and blue nets decreased the mean PPFD by 45%, 41%, and 50%, respectively, compared to the unshaded (open field) control condition. Also, shading substantially reduced light availability compared to the unshaded open field condition-control (996 W/m²), with mean irradiance decreased by 40% under the pearl net, 35% under the red net, and 44% under the blue net.

Overall, photo-selective netting represents a cost-effective strategy for manipulating crop microclimate conditions, plant grown, allowing the regulation of not only yield but also quality and functional or bioactive plant properties.

 

• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?

ʹHerbs' we replace with 'vegetativeʹ mass (leaves and steams) ……

• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).

We are accepting your suggestion. Significant differences (symbols, a,b, ab)between treatments  incorporated in Table (as in Table 3).

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

 

• First paragraph after table 2 - please move to Discussion section.

Yes, we do it and add totally new paragraph

Harvesting of spike-like lavender inflorescences was carried out in late June. As the plants were in their second year of growth, harvesting was performed once, when the majority of flowers were fully open. The yield of fresh inflorescences obtained from plants grown in the open field (0.78 t·ha⁻¹) and under the blue net (0.72 t·ha⁻¹) was significantly lower compared to the values recorded under the red and pearl nets (1.302-1.320 t·ha⁻¹). In addition to their positive effects on the morphological traits of lavender plants, shading with red and pearl nets also positively affected the yield of fresh and dry inflorescences per unit area. Shading resulted in a more uniform fresh-to-dry mass ratio of spike-like inflorescences, regardless of the color of the applied photoselective nets.

A single harvest of the aboveground vegetative biomass (leaves and stems) of lavender plants was carried out in the last decade of September. The fresh biomass yield of plants grown under the blue net (6.10 t·ha¹)   was very significantly lower (p < 0.01) than that of plants grown under the pearl net (7.90 t·ha⁻¹). The positive effect of the red and pearl nets on lavender was even more pronounced when compared with plants grown without shading in the open field. The use of photoselective nets, especially red and pearl ones, represents an effective agrotechnical approach that enhances both vegetative and generative biomass production in lavender, with potential positive effects on quality. The altered light quality caused by shading affects plant performance, often resulting in higher yields for medicinal plants. However, the specific effects depend on the color and type of the photoselective net, as these determine both the light spectrum and intensity reaching the canopy.

 

• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values

Yes we accept your recommendation and remove standard deviation

• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.

             It’s a typist mistake …. This is correct…….linalool (24.7–27.3%),

 

• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit

RI lit (the retention index) was determined according to the different references (a,b,c,d)

^aAdams RP (2007). Identification of essential oil components by gas chromatography /mass spectrometry, Carol Stream, Allured Publishing Co., Illinois, USA.

^bBenkaci-Ali, F.; Baaliouamer, A.; Meklati, B.Y.; Chemat, F., Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation, Flavour Fragr. J., 2007, 22, 2, 148-153, https://doi.org/10.1002/ffj.1773.

^cSaroglou, V.; Dorizas, N.; Kypriotakis, Z.; Skaltsa, H.D., Analysis of the essential oil composition of eight Anthemis species from Greece, J. Chromatogr. A, 2006, 1104, 1-2, 313-322, https://doi.org/10.1016/j.chroma.2005.11.087 .

^dZouari, N.; Ayadi, I.; Fakhfakh, N.; Rebai, A.; Zouari, S., Variations of chemical composition of essential oils in wild-populations of Thymus algeriensis Boiss et Reut., a North African endemic species, Lipids in Health and Desease, 2012, 11, 1, 28-39, https://doi.org/10.1186/1476-511X-11-28.

 

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

We adopt your recommendation and exclude some irrelevant sentences in discussion and add new one

To the best of our knowledge, studies addressing UVB:PAR or UVA:PAR ratios, as well as plant responses to these ratios under horticultural growing conditions, are lacking or extremely scarce. Even studies that focus exclusively on the effects of UV radiation on the growth of horticultural crops are rare. UV radiation is associated with the accumulation of plant secondary metabolites (phenolic compounds, including flavonoids, etc.), which are related to plant color, taste, and perceived health-promoting attributes. 

High solar radiation and elevated temperatures during the summer months can negatively affect both plant yield and quality. The incorporation of light-dispersive and reflective chromatic additives into photo-selective nets transforms direct sunlight into diffuse radiation, enabling deeper light penetration into the inner plant canopy. The radiometric properties of photo-selective nets are determined by their porosity and color. During cultivation, modifications in light quality beneath photo-selective shade nets can positively influence yield, quality traits, and phytochemical composition of aromatic and medicinal plants [37].

Color shade nets alter light spectra, triggering biochemical changes in medicinal plants by manipulating photoreceptors (like phytochromes and cryptochromes) that regulate secondary metabolite (phenols, flavonoids, essential oils) production, photosynthesis, antioxidant activity, and pigmentation, often boosting beneficial compounds by shifting energy balance and influencing stress response pathways for enhanced medicinal quality [38]. 

 

We are also add new paragraph in discussion

Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications

 

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

You are right; we have removed part of the last paragraph to eliminate any ambiguities.

 

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

We have restructured the Conclusions in accordance with your comments.

Based on the results, shading lavender with pearl and red nets increased essential oil (EO) yield in both plant parts compared to non-shaded plants. The most abundant EO component was 1,8-cineole (eucalyptol); the highest content in flower essential oil (LAFEO, 32.2%) was obtained from plants covered with red nets, whereas the highest content in leaf essential oil (LALEO, 39.8%) was recorded in plants shaded with blue nets. Shading with colored nets differentially affected the content of individual essential oil components. A higher camphor content in flowers, which negatively affects oil quality, was observed in non-shaded plants. However, the essential oil from non-shaded plants was characterized by a higher antioxidant capacity than that from all shaded treatments. Nevertheless, this preliminary study, which evaluated both the yield and quality of lavender essential oil under colored shading nets, highlights the strong potential of this species for expanded production in southern Serbia.

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

- Inconsistency between LAFEO, LAEO, LALEO.

6. The English language revision is strongly recommended.

7. Table 2 is inconsistent.

8. Add the yield calculation formula to the experimental section. 

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

Author Response

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

1. The DPPH EC₅₀ values are extremely high (up to 50 mg/mL): This corresponds to low antioxidant activity, however the text refers to “strong antioxidant activity.”

Abstract (Line 7):

"Flower samples from non-shaded (control) plants showed the strongest antioxidant activity, with the lowest EC₅₀ values (51.97 mg/mL at 20 min, 32.26 mg/mL at 60 min, and 20.26 mg/mL at 120 min)."

Corrected to

"Flower samples from non-shaded (control) plants showed moderate antioxidant activity, with EC₅₀ values decreasing over time, indicating the highest activity among treatments tested."

Conversely, the weakest activity was recorded in plant leaves under pearl nets, which showed the highest EC₅₀ value at 120 min (42.40 mg/mL).

Corrected to

Conversely, plant leaves under pearl nets showed the lowest activity among samples, with an EC₅₀ value of 42.40 mg/mL at 120 min, still within the moderate antioxidant activity range.

 

"Unlike the FRAP results, the DPPH assay showed generally stronger activity in flowers."

Corrected to

Unlike the FRAP results, the DPPH assay showed relatively higher activity in flowers compared to leaves, though all samples exhibited moderate antioxidant capacity.

Results Section (Antioxidant activity paragraph):

Current (WRONG):

"Flower samples generally exhibited lower EC50 values than leaves samples, confirming that floral material contains more potent antioxidant components. Among the flowers samples from plants without shading (control) showed the strongest antioxidant activity, with EC50 values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min), the lowest of all tested samples."

Corrected:

"Flower samples generally exhibited lower EC₅₀values than leaf samples, indicating relatively higher radical scavenging activity in floral material. Among all samples, flowers from non-shaded (control) plants showed the highest antioxidant activity, with EC₅₀values of 51.97 mg/mL (20 min), 32.26 mg/mL (60 min), and 20.26 mg/mL (120 min). According to standard classifications chemes, EC₅₀values between 10-50 mg/mLi ndicate moderate antioxidant activity, while values >50 mg/mL indicate weak activity. Therefore, our samples exhibited primarily moderate (20.26-42.40 mg/mL at 120 min) to weak (51.97 mg/mL at 20 min) antioxidant capacity."

 

DiscussionSection:

Current (WRONG):

"In our study LAFEO samples from non-shaded (control) plants exhibited the strongest antioxidant activity, showing the lowest EC50 values at allincubationtimes. In contrast, LALEO samples covered with pearl nets had thehighest EC50 values, indicating the weakest activity. Overall, flower from non-shadingplantsconsistentlydemonstratedhigherantioxidantcapacity (20.26 mg/mL) thanleaf (24.33 mg/mL) samplesacrossallshadingtreatmentsandincubationtimes (Table 6)."

✓ CORRECTED:

"In our study, LAFEO samples from non-shaded (control) plants exhibited moderate antioxidant activity, showing the lowest EC₅₀ values at all incubation times (20.26-51.97 mg/mL). In contrast, LALEO samples covered with pearl nets had the highest EC₅₀ values (42.40 mg/mL at 120 min), representing the lowest activity among treatments, though still with in the moderate range. Overall, flowers from non-shaded plants showed relatively higher radical scavenging activity (EC₅₀ = 20.26 mg/mL at 120 min) compared to leaves (EC₅₀ = 24.33 mg/mL at 120 min) across all shading treatments and incubation times (Table 6). These EC₅₀values (20-50 mg/mL) classify the antioxidant activity as moderate according to established standards, which is appropriate for essential oils and relevant for cosmetic and pharmaceutical applications."

ADDITIONAL REQUIRED ADDITIONS:

ADD to Results Section (after Table 6, before FRAP discussion):

"Interpretation of DPPH Results:

According to standard classification for DPPH radical scavenging activity [cite appropriate reference, e.g., Molyneux 2004], EC₅₀values<10 mg/mL indicate strong antioxidant activity, 10-50 mg/mL indicate moderate activity, and>50 mg/mL indicate weak activity. Based on this scale, the lavender essential oils tested in this study exhibited primarily moderate antioxidant activity at 120 min incubation (EC₅₀ = 20.26-42.40 mg/mL), with initial weak activity at 20 min (EC₅₀ = 51.97-78.58 mg/mL) improving over extended reaction time. For comparison, synthetic antioxidants such as BHT typically show EC₅₀ values of 5-15 mg/mL, while natural compounds like ascorbic acid exhibit EC₅₀ values of 2-5 mg/mL.

2. The split-plot design is mentioned, but: primary/secondary factors are not defined, and there is no clear description of the plots.

You are right; it is indeed a one-factor experiment, since the different shading nets represent a single factor, and the experiment was conducted over one growing season.

 

3. “Combinations of shaded sage and non-shaded sage control plants.” The work focuses on lavender, not sage.

Its mistake…we change these….

4. The DPPH and FRAP results are contradictory. There is no serious mechanistic discussion to explain these differences. ABTS is listed in the reagents but is never used.

We thank the reviewer for this important observation regarding the apparent contradiction between DPPH and FRAP results.

1. Mechanistic Explanation Added: We agree that our original explanation was superficial. We have substantially expanded the Results and Discussion sections to provide detailed mechanistic interpretation of why flowers excel in DPPH (hydrogen atom transfer, driven by high linalool content) while leaves show superior FRAP activity (electron transfer, likely involving camphor, borneol, and non-volatile phenolic compounds not detected by GC/MS). The revised text now includes:

• Mechanistic basis for each assay (HAT vs. SET)
• Structure-activity relationships of major compounds
• Discussion of non-volatile phenolics in leaves
• Kinetic considerations (120 min vs. 30 min reactions)
• Biological/application relevance of each mechanism

REVISED ResultsSection (after Table 7):

BEFORE (inadequate):

"This pattern differs from the DPPH assay results, where flowers generally showed stronger radical-scavenging activity. Such divergence is notun common, as FRAP and DPPH measure different antioxidant mechanisms, FRAP evaluating ferric ion reduction, while DPPH measures hydrogen atom or electron donation to a stable radical."

 

AFTER (explanation):

The divergence between DPPH and FRAP results requires mechanistic explanation. While flowers exhibited superior DPPH radical scavenging activity, leaves demonstrated stronger ferric-reducing capacity in the FRAP assay. This apparent contradiction reflects fundamental differences in antioxidant mechanisms and the chemical nature of active compounds in each tissue.

DPPH assay measures single-electron transfer (SET) and hydrogen atom transfer (HAT) mechanisms, favoring compounds with phenolic hydroxyl groups capable of donating hydrogen atoms to stabilize free radicals [33]. The higher linalool content in flowers (24.7-27.3%) compared to leaves (6.0-8.6%) likely contributes to superior DPPH activity, as linalool's tertiaryhydroxyl group readily donates hydrogen to DPPH radicals [34].

FRAP assay, conversely, exclusively measures electron-donating capacity under acidic conditions (pH 3.6), reflecting the reductionof Fe³⁺-TPTZ complex to Fe²⁺form. The higher FRAP values in leavesmayresult from: (1) elevated camphor content (11.3-13.9% vs. 7.5-8.6% in flowers), whose carbonyl group participates in electron transfer reactions; (2) greater abundance of borneol (21.9-26.5% vs. 18.0-21.9%), a secondary alcohol with stronger reducing properties than tertiary alcohols; and (3) possibly higher concentrations of non-volatile phenolic compounds (e.g., rosmarinic acid, caffeic acid derivatives) that were not detected by GC/MS but are known to be more abundant in photosynthetic leaf tissue and exhibit exceptional FRAP activity [35].

Additionally, 1,8-cineole, the most abundant compound in both flowers (27.4-32.2%) and leaves (30.4-39.8%), is an ether with minimal antioxidant activity in both assays [cite], explaining why total essential oil content does not directly correlate with antioxidant capacity.

The kinetics also differ: DPPH reached equilibrium after 120 min incubation (allowing slow-reacting compounds to contribute), while FRAP measures instant aneous reducing capacity after 30 min at 37°C, potentially favoring different compoundclasses. Similar tissue-dependent antioxidant activity patterns have been reported in other Lamiaceae species [36].

 

 

33. Gulcin, I.; Alwasel, S.H. DPPH Radical Scavenging Assay. Processes 2023,11, 2248. https://doi.org/10.3390/pr1108 2248
34. Baliyan, S.; Mukherjee, R.; Priyadarshini, A.; Vibhuti, A.; Gupta, A.; Pandey, R.P. Chang CM. Determination of antioxidants by DPPH radical scavenging activity and quantitative phytochemical analysis of Ficus religiosa. Molecules. 2022, 27(4),1326. doi: 10.3390/molecules27041326
35. Proestos, C.; Komaitis, M. Analysis of Naturally Occurring Phenolic Compounds in Aromatic Plants by RP-HPLC Coupled to Diode Array Detector (DAD) and GC-MS after Silylation. Foods. 2013, 2, 90-99.

        doi: 10.3390/foods2010090. 

36. Macedo Arantes, S.; Teresa Caldeira, A.; Rosário Martins, M. Essential oils high in 1,8-Cineole of Mediterranean flavoring plants: Health Benefits. IntechOpen. 2022.  doi: 10.5772/intechopen.103831

ADD to Discussion Section (new paragraph specifically addressing this)

"Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications.

 

2. ABTS Reagent Error Corrected: We apologize for this oversight. ABTS was initially planned for the study but ultimately not used. It has been removed from the reagents list (Section 2.2).

Current Reagents Section:

 

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA)."

 

CORRECTED (Remove ABTS):

"Ethanol 96% p.a. (Reahem d.o.o., Novi Sad, Serbia), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical, 2,4,6-tris(2 pyridyl)-s-triazine (TPTZ), iron (III) chloride hexahydrate (Sigma Chemical Company, St. Louis, MO, USA). All other chemicals are of analytical reagent grade (p.a.)."

5. Repeated errors in nomenclature

- Lavendula instead of Lavandula (repeated error) and “lavander” instead of “lavender,”

Yes it is mistake. We have revised it.

 

- Inconsistency between LAFEO, LAEO, LALEO.

We exclude abbreviation LAEO, it will be used consistently throughout the text lavender EO

 

6. The English language revision is strongly recommended.

We try to improve English language

 

7. Table 2 is inconsistent.

We are accepting your suggestion ….Significant differences (symbols, a,b, ab) between treatments  incorporated in Table 2 (as in Table 3).

 

Table 2.

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

8. Add the yield calculation formula to the experimental section. 

To calculate fresh herb yield (t/ha) from g/plant, first find the average fresh weight per plant, multiply by plant density (plants/m² * 10,000 for m²/ha) to get kg/ha, then divide by 1000 to get t/ha, remembering to sample representative plants, clean them lightly, and weigh them immediately for accuracy in your open-field trial

9. Underutilized ANOVA: lack of F values, degrees of freedom, and factor × factor interaction.

All measurements were conducted in triplicate, and results were expressed as mean ± standard deviation.  For morphological evaluation and biochemical analyses, the data were analyzed using one-way ANOVA, followed by Duncan’s multiple range test (p < 0.05).

 

10. The article needs other statistical analyses to improve its quality, such as principal component analysis.

The manuscript is already quite extensive, and including a PCA analysis would further increase its length. However, if you insist, we will perform the PCA analysis.

 

 

Comments on the Quality of English Language

The manuscript contains numerous grammatical and spelling errors. A thorough professional revision in English is strongly recommended.

We have attempted to improve the English with the help of colleagues who are native speakers, and we hope that the improvement is evident.

 

Author Response File: Author Response.docx

Reviewer 3 Report

Comments and Suggestions for Authors

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

 

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?
• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.
• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?
• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).
• First paragraph after table 2 - please move to Discussion section.
• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values
• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.
• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit column.

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

Comments for author File: Comments.pdf

Author Response

v

Manuscript entitled "Effect of light modification by shading nets on yield, composi tion, and antioxidant activity of Lavandula angustifolia Mill. es sential oil " submitted to the Plants journal is original and interesting. The manuscript concerns the important issue of yielding, chemical composition and biological activities of Lavandula angustifolia flower essential oil (LAFEO) and leaves (LALEO) cultivated under different shade nets compared to non-shading  plants. Studying the impact of growth environmental conditions on plants is important due to changes in the quality of raw materials, so these results deserve publication. Although the manuscript is interesting, it requires consideration of the following comments and the way of presenting and describing the results should be improved according to below:

- Introduction: information about "lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13] to L. dentata, not L. angustioflia as in this study. These are two different products; the effects of EO from one lavender species should not be generalized to others.

lavender EO has antifungal and insecticidal activities, which supports their promising use in agricultural production and storage of leguminous crops" according to the cite literature refers [13].

 

We exclude old reference [13] with L. dentata, and include new reference [13] with L. angustifolia.

 

Mijatovic, S.; Stankovic, J.A.; Calovski, I.C.; Dubljanin, E.; Pljevljakusic, D.; Bigovic, D.; Dzamic, A. Antifungal activity of Lavandula angustifolia essential oil against Candida albicans: Time-kill study on pediatric sputum isolates. Molecules. 2022, 27(19), 6300. doi: 10.3390/molecules27196300.   

            

Lavender EO has antifungal activities, which supports their promising fungicidal potential. [13].

 

- there is a typo in the Latin name of lavender twice, i.e. incorrect LavEndula instead of LavAndula - please correct it (the first sentence of Abstract and the first sentence of Material and methods - unfortunately the lines are not numbered)

We have corrected it. Now stay lavender

Methodology:

- Only one-year of field experiments with the agrotechnical factor raises methodological doubts

- The second paragraph of the Material and Methods describes an experiment with sage, not lavender. Is this a mistake?

Yes it is mistake. We have revised it.

- The 3. and 4. paragraph of the Materials and Methods chapter describes the yield results (move to the Results section). The harvesting methodology should be described here: for example, when and how the shoots were cut, at what height, what stage of plant development the plants were in, etc.

We accept your recommendation and remove these parts

Inflorescences were harvested once, with fresh yield (spike-like inflorescences) of 1.320 t/ha under the pearl net, 1.302 t/ha under the red net, 0.72 t/ha under the blue net, and 0.78 t/ha in the open field. The fresh-to-dry mass ratio was uniform (1.71–1.81).

In September, when plants had formed more branches (13.7 in the open field and 14.1–16.4 under shading nets), green biomass was harvested once. Yields reached 5.6 t/ha in the open field and 6.1–7.9 t/ha under photoselective nets.   

and replace with new one

Flowering stems developed in late June. Harvesting of spike-like inflorescences was performed once, when the majority of flowers were fully open. The cut spike-like inflorescences were placed in a shaded and well-ventilated area for drying. The vegetative aboveground biomass (leaves and stems) was harvested later, in the last decade of September. The shoots were cut when 13–14 branches had formed in plants grown under the blue net and in the open field (control), and 15–17 branches in plants grown under the pearl and red nets. At this stage, the vegetative biomass of the plants was sufficiently developed to withstand cutting. The apical parts of the branches about10 cm in length, were cut.

Results:

• The first paragraph of Results is essentially a literature review (move to the appropriate chapter). It lacks interpretation of the results and indication of differences (including statistical differences) between the various shading nets and the control.
• Figure 1: Why does the chart only compare to the red shade net? And why not the others?

 

• Third paragraph - avoid balding in the text. The text again looks like a literature review. Interpret the results from Table 2 specifically.

We exclude third paragraph and replace with new one.

In July shading with pearl, red, and blue nets decreased the mean PPFD by 45%, 41%, and 50%, respectively, compared to the unshaded (open field) control condition. Also, shading substantially reduced light availability compared to the unshaded open field condition-control (996 W/m²), with mean irradiance decreased by 40% under the pearl net, 35% under the red net, and 44% under the blue net.

Overall, photo-selective netting represents a cost-effective strategy for manipulating crop microclimate conditions, plant grown, allowing the regulation of not only yield but also quality and functional or bioactive plant properties.

 

• Table 2. Explain what "herb" means. The title and methodology refer to obtaining leaves - are they the same?

ʹHerbs' we replace with 'vegetativeʹ mass (leaves and steams) ……

• table footer 2: LSD is unclear which value is assigned to which column - add the next rows in the table and place these values ​​in them. Ideally, mark homologous groups with the same letters (as in Table 3).

We are accepting your suggestion. Significant differences (symbols, a,b, ab)between treatments  incorporated in Table (as in Table 3).

Shade nets	Plant height (cm)	Number of primary branches	Total number of branch	Vegetative biomass g/plant	Vegetative biomass t/ha	Fresh inflorescence mass (g/plant)	Fresh inflorescence mass -g/m²	Inflorescence length (cm)	Dry inflorescence mass (g/plant)	Dry inflorescence mass-g/m²	Fresh-to-dry inflorescence mass ratio
Pearl Red Blue Control	28.6 a a 27.9 ab a 24.2 bc ab 22.1 c b	3.24 a a 3.21 a a 2.98 ab a 2.73 b a	16.4 a a 15.6 a a 14.1 b ab 13.7 b b	284.4 a a 255.3 ab ab 218.4 bc ab 204.6 c b	7.90a a 7.11ab ab 6.10bc b 5.69c b	47.3 a a 46.52 a a 25.79 b b 27.92 b b	132.40 a a 130.28 a a 72.197 b b 78.189 b b	6.7 a a 6.4 ab a 5.9 ab a 5.5 b a	26.740 a a 27.023 a a 14.327 b b 15.393 b b	74.76 a a 75.65 a a 40.08 b b 43.09 b b	1.77 1.72 1.80 1.81

 

 

• First paragraph after table 2 - please move to Discussion section.

Yes, we do it and add totally new paragraph

Harvesting of spike-like lavender inflorescences was carried out in late June. As the plants were in their second year of growth, harvesting was performed once, when the majority of flowers were fully open. The yield of fresh inflorescences obtained from plants grown in the open field (0.78 t·ha⁻¹) and under the blue net (0.72 t·ha⁻¹) was significantly lower compared to the values recorded under the red and pearl nets (1.302-1.320 t·ha⁻¹). In addition to their positive effects on the morphological traits of lavender plants, shading with red and pearl nets also positively affected the yield of fresh and dry inflorescences per unit area. Shading resulted in a more uniform fresh-to-dry mass ratio of spike-like inflorescences, regardless of the color of the applied photoselective nets.

A single harvest of the aboveground vegetative biomass (leaves and stems) of lavender plants was carried out in the last decade of September. The fresh biomass yield of plants grown under the blue net (6.10 t·ha¹)   was very significantly lower (p < 0.01) than that of plants grown under the pearl net (7.90 t·ha⁻¹). The positive effect of the red and pearl nets on lavender was even more pronounced when compared with plants grown without shading in the open field. The use of photoselective nets, especially red and pearl ones, represents an effective agrotechnical approach that enhances both vegetative and generative biomass production in lavender, with potential positive effects on quality. The altered light quality caused by shading affects plant performance, often resulting in higher yields for medicinal plants. However, the specific effects depend on the color and type of the photoselective net, as these determine both the light spectrum and intensity reaching the canopy.

 

• First paragraph after the third table: there is no need to provide the standard deviation in the text next to the result values

Yes we accept your recommendation and remove standard deviation

• The result of the linalool content in LAFEO (2.7-27.3%) is inconsistent with the table - please check it.

             It’s a typist mistake …. This is correct…….linalool (24.7–27.3%),

 

• Table 4 and 5.: Please explain what the letters a and b mean in the RI lit

RI lit (the retention index) was determined according to the different references (a,b,c,d)

^aAdams RP (2007). Identification of essential oil components by gas chromatography /mass spectrometry, Carol Stream, Allured Publishing Co., Illinois, USA.

^bBenkaci-Ali, F.; Baaliouamer, A.; Meklati, B.Y.; Chemat, F., Chemical composition of seed essential oils from Algerian Nigella sativa extracted by microwave and hydrodistillation, Flavour Fragr. J., 2007, 22, 2, 148-153, https://doi.org/10.1002/ffj.1773.

^cSaroglou, V.; Dorizas, N.; Kypriotakis, Z.; Skaltsa, H.D., Analysis of the essential oil composition of eight Anthemis species from Greece, J. Chromatogr. A, 2006, 1104, 1-2, 313-322, https://doi.org/10.1016/j.chroma.2005.11.087 .

^dZouari, N.; Ayadi, I.; Fakhfakh, N.; Rebai, A.; Zouari, S., Variations of chemical composition of essential oils in wild-populations of Thymus algeriensis Boiss et Reut., a North African endemic species, Lipids in Health and Desease, 2012, 11, 1, 28-39, https://doi.org/10.1186/1476-511X-11-28.

 

Discussion: The discussion is very long-winded and sometimes repeats results, for the reader's comfort it can be shortened and focused on the essence, i.e. how these shading nets modify the raw material.

We adopt your recommendation and exclude some irrelevant sentences in discussion and add new one

To the best of our knowledge, studies addressing UVB:PAR or UVA:PAR ratios, as well as plant responses to these ratios under horticultural growing conditions, are lacking or extremely scarce. Even studies that focus exclusively on the effects of UV radiation on the growth of horticultural crops are rare. UV radiation is associated with the accumulation of plant secondary metabolites (phenolic compounds, including flavonoids, etc.), which are related to plant color, taste, and perceived health-promoting attributes. 

High solar radiation and elevated temperatures during the summer months can negatively affect both plant yield and quality. The incorporation of light-dispersive and reflective chromatic additives into photo-selective nets transforms direct sunlight into diffuse radiation, enabling deeper light penetration into the inner plant canopy. The radiometric properties of photo-selective nets are determined by their porosity and color. During cultivation, modifications in light quality beneath photo-selective shade nets can positively influence yield, quality traits, and phytochemical composition of aromatic and medicinal plants [37].

Color shade nets alter light spectra, triggering biochemical changes in medicinal plants by manipulating photoreceptors (like phytochromes and cryptochromes) that regulate secondary metabolite (phenols, flavonoids, essential oils) production, photosynthesis, antioxidant activity, and pigmentation, often boosting beneficial compounds by shifting energy balance and influencing stress response pathways for enhanced medicinal quality [38]. 

 

We are also add new paragraph in discussion

Mechanistic Interpretation of Antioxidant Assay Divergence

The contrasting results between DPPH and FRAP assay slighlight the complexity of antioxidant assessment in essential oils. Our findings—where flowers excelled in DPPH but leaves in FRAP—align with reports in other aromatic plants showing tissue-specific antioxidant profiles [11]. The high linalool content in flowers (up to 27.3%) explains superior hydrogen-donating capacity in DPPH assays, as monoterpene alcohols are effective radical scavengers through hydrogen atom transfer [30].

Conversely, the elevated FRAP activity in leaves likely reflects: (1) synergistic effects of camphor and borneol, whose combined electron-donating capacity exceeds individual contributions [citeifavailable]; (2) the presence of unquanti fied phenolic compounds that accumulate preferentially in photosynthetic tissue and exhibit strong ferric-reducing activity [31]; and (3) potential matrix effects, as the more complex leaf essential oil composition (55-65 compounds vs. 47-59 in flowers) may create favorable conditions for electron transfer reactions.

Importantly, these assays measure complementary rather than redundant aspects of antioxidant capacity. DPPH reflects biological relevance in lipid peroxidation scenarios (relevant for cosmetic applications), while FRAP indicates potential in metal chelation and redox regulation (relevant for pharmaceutical use).

Thus, the apparent contradiction actually demonstrates the multifunctional antioxidant profiles of both flower and leaf essential oils, with each suited to different applications rather than one being universally superior.

The revised manuscript now clearly explains that the DPPH/FRAP divergence reflects complementary antioxidant mechanisms rather than contradictory results, demonstrating that flower and leaf essential oils possess multifunctional antioxidant profiles suited to different applications

 

At the end of the discussion, there's information about the characterization of the EO aroma. This should be in the Results section. The methodology for this characterization is described in the Materials and Methods section.

Overall, the last paragraph of the discussion is a bit of an exaggeration. Neither the profitability of cultivation nor the associated income were studied; these are just assumptions.

You are right; we have removed part of the last paragraph to eliminate any ambiguities.

 

Conclusion: The conclusions include a description confirming already known facts, for example, regarding typical oil components. The reader expects to receive guidance on which color of shading nets is best to use. If it's impossible to identify a single best shading nets, it should be noted which features are enhanced and which are diminished by the shading nets of a particular color.

We have restructured the Conclusions in accordance with your comments.

Based on the results, shading lavender with pearl and red nets increased essential oil (EO) yield in both plant parts compared to non-shaded plants. The most abundant EO component was 1,8-cineole (eucalyptol); the highest content in flower essential oil (LAFEO, 32.2%) was obtained from plants covered with red nets, whereas the highest content in leaf essential oil (LALEO, 39.8%) was recorded in plants shaded with blue nets. Shading with colored nets differentially affected the content of individual essential oil components. A higher camphor content in flowers, which negatively affects oil quality, was observed in non-shaded plants. However, the essential oil from non-shaded plants was characterized by a higher antioxidant capacity than that from all shaded treatments. Nevertheless, this preliminary study, which evaluated both the yield and quality of lavender essential oil under colored shading nets, highlights the strong potential of this species for expanded production in southern Serbia.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Dear authors,

the manuscript has now been corrected according to my suggestions, but some drawbacks remain in the text, which the authors should address.

Please correct the following mistakes:

In Line 246: The first letter of the month names must be written with a capital letter.

In L 278: The legend for Table 2 is missing. What do all the letters beside the values indicate?

In Table 2, the labelling of statistically significant differences is incorrect and misleading.

What does doubled label with character mean??  (16.4 a a; 28.6 a a; 27.9 ab a???)

Line 291: … was very significantly lower …. Please remove 'very'

Author Response

Please correct the following mistakes:

In Line 246: The first letter of the month names must be written with a capital letter.

We correct that

 

In L 278: The legend for Table 2 is missing. What do all the letters beside the values indicate?

Each column in Table 2 represents the value of the corresponding morphological trait, vegetative yield, or generative yield. Letters are shown next to each value in the column. Values followed by different letters are significantly different at p < 0.05 and p < 0.01.

In Table 2, the labelling of statistically significant differences is incorrect and misleading.

The experiment was designed as a monofactortrial with three replications for each value presented in Table 2. Statistical analysis was performed based on these replications. Experimental results were considered statistically significant at P < 0.05 and P < 0.01. Different letters next to each value indicate these significant differences. Data analysis was supported by the MSTAT C software.

What does doubled label with character mean??  (16.4 a a; 28.6 a a; 27.9 ab a???)

Values followed by different letters are significantly different at p < 0.05 and p < 0.01.

Thus, for the value 16 aa, the first “a” refers to a significant difference at p < 0.05, while the second “a” refers to a significant difference at p < 0.01.

Example 2: 27.9 ab a, where the letters “ab” indicate a significant difference at p < 0.05, whereas the letter “a” indicates a significant difference at p < 0.01.

Line 291: … was very significantly lower …. Please remove 'very'

We remove ʹvery'

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

Accept 

Author Response

Thanks a lot for your support and contribution !!!!!!!!

Reviewer 3 Report

Comments and Suggestions for Authors

The authors took my comments into account and significantly improved the manuscript. It may be published in this version.

Author Response

Thanks a lot for your support and contribution !!!!!!!!