Sorbitol-Stabilized Silicon Formulation Improve Root Traits and Antioxidant Response in Drought-Stressed Soybean

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

In this manuscript, the authors analyze“Sorbitol-Stabilized Silicon Formulation Improve Root Traits and Antioxidant Response in Drought-Stressed Soybean”. There are many concerns about the manuscript that the authors need to  be addressed and substantially  revised,  to ensure the scientific rigor and reliability.

 

Question 1:  In line 271-272. It was mentioned that “These findings are  consistent with other studies [28,29].  Please explain with details the  differecne and rewrite it.

 

 Question 2:  Many Figure Legends，coordinate axis  and Annotations were need  to be modified.  For example, in the main text was showed with the Figure 1A,B,C,D, but the figure legends were used figure 1a, b,c,d.  

 

Question 3: The figrue 8 was not cited in the text, as well as the figure7.

Question 4:  In line 249-251，In addition, tyrosine contents also decreased under Si supply, which may reflect an increased requirement for phenolic compound biosynthesis, including lignin and antioxidant metabolites, under these conditions. But， there were not data   of  lignin and antioxidant metabolites showed in the manuscript. Please added and clarified.

 

Question 5: In line 271-272. It was mentioned that “These findings are  consistent with other studies [28,29].  Please explain with details the  differecne and rewrite it.

 

Question 6: In the  discussion and conclusions, the author should provide more constructive suggestions and viewpoints based on the results.

 

Question 7: In text and references, many typographical errors and formatting inconsistencies，species name ( Latin) should be italic font, and the format of references needs to be uniform. Please check carefully and change it.   

 

Question 8: There were many references are outdated, for example, [38，44-49] , please check and update new versions.

 

 

Comments on the Quality of English Language

The English could be improved to more clearly express the research.

Author Response

Question 1:  In line 271-272. It was mentioned that “These findings are consistent with other studies [28,29].  Please explain with details the difference and rewrite it.

Dear reviewer, thank you for your comment. This step was not clearly described in the original version, so we have now expanded the explanation and clarified the rationale for each cited article, detailing how they were used as background knowledge to support and complement the discussion in our manuscript. “Similarly, Abdullah et al. [29] reported a 23% increase in photosynthetic rate in drought-stressed plants treated with 200 kg ha⁻¹ of Si applied to the soil compared to the control. This improvement may be linked to the activation of the phenylpropanoid pathway and stress-responsive enzymes triggered by Si, which reduce the physiological impact of abiotic stress [30].” [lines 305-310]”.

Question 2:  Many Figures Legends coordinate axis and Annotations were need to be modified.  For example, in the main text was showed with the Figure 1A, B, C, D, but the figure legends were used figure 1a, b, c, d.  

Dear reviewer, thank you for your comment. The figure legend coordinates have been standardized in the text as '1a, b, c, d…'.

Question 3: The figrue 8 was not cited in the text, as well as the figure7.

Dear reviewer, thank you for your comment. We included the correspondence of the figure 7 and 8 in the lines 233 and 256, respectively.

Question 4:  In line 249-251，In addition, tyrosine contents also decreased under Si supply, which may reflect an increased requirement for phenolic compound biosynthesis, including lignin and antioxidant metabolites, under these conditions. But there were not data   of lignin and antioxidant metabolites showed in the manuscript. Please added and clarified.

Dear Reviewer, thank you for your comment. The line 249 has been rewritten to read: 'which may reflect an increased requirement for phenolic compound biosynthesis under these conditions.' We have excluded the comment about lignin. Moreover, the introduction cited some studies that analyzed the relationship between silicon and lignin in soybean (lines 92–94). In the study, the lignin content in the plant was not evaluated.

Question 5: In line 271-272. It was mentioned that “These findings are consistent with other studies [28,29].  Please explain with details the difference and rewrite it.

Dear Reviewer, thank you for your comment. Maybe I responded this question in the question 1.

Question 6: In the discussion and conclusions, the author should provide more constructive suggestions and viewpoints based on the results.

Dear reviewer, thank you for your comment. In the discussion the mechanisms involved in the morphological and physiological treats of the plants was more explored by fundamental and recent articles: “These findings are consistent with the study by Sah et al. [28], who observed increased leaf area in silicon-treated plants, particularly under high water availability. Similarly, Abdullah et al. [29] reported a 23% increase in photosynthetic rate in drought-stressed plants treated with 200 kg ha⁻¹ of Si applied to the soil compared to the control. This improvement may be linked to the activation of the phenylpropanoid pathway and stress-responsive enzymes triggered by Si, which reduce the physiological impact of abiotic stress [30].

As expected, drought stress reduced stem diameter and increased leaf temperature. Water deficit is known to disrupt xylem hydraulic continuity by promoting cavitation and the formation of air embolisms, which impair water transport efficiency. Since transpiration-driven water flow is essential for heat dissipation and temperature regulation in leaves, reduced stomatal conductance under drought conditions further limits evaporative cooling. As a result, leaf temperature rises, and metabolic activity becomes restricted, as many enzymatic processes operate within narrow thermal optima. [lines 307-319].”

“These findings align with previous studies [13, 29, 31], which suggest that enhanced root development under Si supply may be linked to the modulation of gene expression associated with root growth—particularly genes involved in auxin biosynthesis and transport, such as YUC, PIN, and WAT. In addition, improved allocation of photoassimilates to belowground organs have been proposed as a complementary mechanism. Supporting this, Tripathi et al. (2022) reported that soil-applied Si in soybean enhances the activity of phenolic compounds, osmoprotectants, and antioxidant enzymes. These biochemical responses help maintain chlorophyll homeostasis during drought stress and ensure a steady supply of sugars to support sustained root growth [lines 331-340]”.

“Previous studies have reported that Si can stimulate ascorbic acid accumulation by enhancing the ascorbate–glutathione cycle, promoting reactive oxygen species (ROS) detoxification, and modulating metabolic pathways that support antioxidant biosynthesis [10]. [lines 350-353]”.

“Interestingly, in the context of drought stress, the observed patterns of proline accumulation suggest that adequate Si supply, particularly in the sorbitol-stabilized form (SiKe), reduced the accumulation of this amino acid, which is commonly recognized as a biochemical marker of water deficit. Si supplementation likely alleviated the perceived stress intensity, enabling plants to maintain metabolic homeostasis. This stress moderation may have resolved an energy allocation trade-off by negating the need for the energetically costly synthesis of osmoprotectants such as proline. As a result, limited carbon and nitrogen resources could be preferentially directed toward vegetative or reproductive growth processes [37]. Additionally, reduced proline levels may reflect a metabolic reallocation, in which this amino acid is diverted toward the biosynthesis of other stress-related compounds that support cellular homeostasis [37].  [lines 397-407]”

The conclusion was more explored and the results/discussions was more descriptive for more clary: “Across two complementary experiments, SiKe consistently promoted root development, increased biomass accumulation, enhanced antioxidant capacity, and modulated key primary and secondary metabolic pathways, particularly isoflavonoid biosynthesis, thereby improving tolerance to oxidative stress. These coordinated responses suggest that SiKe strengthens physiological resilience while maintaining growth and metabolic homeostasis, especially under water deficit. [lines 567-572].

“stabilized Si formulations under diverse soil and climatic field conditions, investigating the temporal dynamics of foliar Si absorption and internal redistribution, and elucidating the molecular pathways modulated by SiKe, particularly those associated with phenylpropanoid metabolism and osmotic adjustment. Additionally, identifying the most effective phenological stages for Si application will be essential to optimizing its benefits. Advancing these research fronts will help establish stabilized Si as a practical and scalable solution for enhancing soybean resilience and productivity in an era of increasing environmental stress [lines 579-586].”

Question 7: In text and references, many typographical errors and formatting inconsistencies, species name ( Latin) should be italic font, and the format of references needs to be uniform. Please check carefully and change it.   

Dear Reviewer, thank you for your comment. We have rechecked the manuscript and corrected all inconsistencies in scientific species names and reference formatting.

Question 8: There were many references are outdated, for example, [38，44-49] , please check and update new versions.

Dear Reviewer, thank you for your comment. I carefully reviewed the entire reference list and replaced several older citations with more recent studies (within the last six years), particularly those used for conceptual background and literature review. However, a few classical references were intentionally retained because they correspond to the original and officially established descriptions of the laboratory methods employed in this study, such as: Chlorophyll and carotenoid determination (Lichtenthaler, 1987); Total phenolics determination (Singleton & Rossi, 1965); Relative water content/turgidity (Barrs & Weatherley, 1962); Silicon digestion procedure (Elliott & Snyder, 1991); Soil chemical analyses for fertility evaluation in tropical soils (Raij et al., 2001).

Reviewer 2 Report

Comments and Suggestions for Authors

• Experimental Aim Clarity
  The objective is stated clearly, but it is somewhat lengthy and repetitive. The authors could succinctly highlight the knowledge gap (insufficient understanding of Si sorbitol stabilization effects) before aiming to improve flow.

• Lines 44–50-The introduction appropriately emphasizes climatic extremes but could better connect global challenges to soybean-specific vulnerabilities and Si’s role in mitigation.

• Lines 85–99-The discussion of ROS generation and antioxidant defense is technically correct, but too descriptive. Suggest linking these mechanisms directly to the Si-induced physiological outcomes reported later.

• Line 107–115-The hypothesis is implicit. Please explicitly state your working hypothesis (e.g., “We hypothesized that sorbitol-stabilized Si improves drought resilience by enhancing root traits and antioxidant activity”).

• Lines 118–128-Figure 1 combines multiple variables (SPAD, leaf area, etc.). Consider splitting into subfigures or enhancing legends for clarity. The differences between water and Si treatments could also be summarized in a supporting table.

• Lines 147–164-While SiKe improved root parameters significantly, the physiological reasons (e.g., hormonal modulation, osmotic adjustment) are not deeply discussed. Recommend expanding the discussion with literature support (Tripathi et al., 2022; Abdullah et al., 2025).

• Lines 188–199-The authors note strong variation but do not explain why proline was reduced under Si or how this links to stress mitigation. Consider relating this to energy trade-offs and osmoprotection balance.

• Lines 217–233-PCA interpretation is sound, but the figure (Figure 7) lacks quantitative clarity. Suggest labeling major contributing variables on the PCA axes or including a loading table in the supplementary material.

• Lines 238–251-The heatmap is visually useful but would benefit from a numerical correlation summary in the text. Explain how decreased amino acid levels under SiKe indicate improved metabolic homeostasis.

• Lines 259–297-The discussion summarizes results effectively but often repeats descriptive findings instead of interpreting the mechanistic basis (e.g., Si-induced gene regulation, antioxidant pathway priming). Expand connections to Si transporters or phenylpropanoid regulation.

• Lines 298–310-Excellent interpretation regarding daidzein induction. Still, the authors should compare these findings with previous molecular-level studies (e.g., La et al., 2023; Trush & Pal'ove-Balang, 2023) to strengthen novelty claims.

Comments on the Quality of English Language

The manuscript reads clearly overall but needs minor grammatical and stylistic corrections, particularly verb tense consistency (past tense for methods/results, present for general discussion). Phrases such as “the Si can regulate” (Line 85) should be revised to “Si regulates.”

Author Response

The objective is stated clearly, but it is somewhat lengthy and repetitive. The authors could succinctly highlight the knowledge gap (insufficient understanding of Si sorbitol stabilization effects) before aiming to improve flow.

Dear Reviewer, thank you for your comment. Some topics of the text was cutted off to evicted the essive repetition of some subject, the phrases that was retired are described above:

“Likewise, Si application via nutrient solution was reported to ameliorate the drought stress in tomato (Solanum lycopersicum L.) [13], sorghum (Sorghum bicolor (L.) Moench) [9] and soybean [14]. [line 66-68]”. “ROS is either produced by redox reactions or by the own metabolism in different cell organelles, e.g., mitochondria, peroxisomes, endoplasmic reticulum, cell wall and apoplast [20]. [lines 75-77]”. “Chloroplast, photosystem I, photosystem II and electron transport chain are producers of ROS, e.g., ¹O₂ and O₂^-. There is also formation of ROS by the reduction of oxygen (O₂) to hydrogen peroxide (H₂O₂) in the complexes I and III from mitochondria by electron transport. [lines 77-80]”

It was created a new paragraph to emphases the question about the use of sorbitol + Si: “Leaf Si absorption can be enhanced by combining it with sorbitol, which lowers the solution’s deliquescence point, reduces droplet evaporation, and facilitates uptake [24]. This improves spray stability and, when combined with Si, can enhance the absorption in soybean, maize and cotton [24]. The central question of this study is how a sorbitol-stabilized Si solution affects various physiological and biometric traits in soybean under both well-watered and drought conditions.  [lines 106-111]”.

Additionally, it was added a phrase in the started-middle of the text to start the reader for the interest topic of the article, the effects of the Si in the soybean plant, and the influence of sorbitol under drought stress: “Additionally, the composition of Si-containing solutions, particularly the inclusion of sorbitol, may enhance their effects on both physiological processes and structural development in soybean plants, thereby improving their ability to withstand drought stress.” [lines 71-74].

Lines 44–50-The introduction appropriately emphasizes climatic extremes but could better connect global challenges to soybean-specific vulnerabilities and Si’s role in mitigation.

Dear Reviewer, thank you for your comment. For an appropriate emphasizes climatic extremes to the challenges to soybean-specific vulnerabilities, it was created a phrase “Its widespread cultivation, both in Brazil and worldwide, exposes it to a wide range of climatic conditions, particularly drought, since soybean is a drought-sensitive herbaceous species. Given the crop’s limited resilience to severe climate events, there is a clear need to enhance its capacity to withstand such stresses.” [lines 56-59] to relate the low resilience of soybean plants to global climatic problems.

Lines 85–99-The discussion of ROS generation and antioxidant defense is technically correct, but too descriptive. Suggest linking these mechanisms directly to the Si-induced physiological outcomes reported later.

Dear Reviewer, thank you for your comment. As described in the question “The objective is stated clearly, but it is somewhat lengthy and repetitive. The authors could succinctly highlight the knowledge gap (insufficient understanding of Si sorbitol stabilization effects) before aiming to improve flow.” Some repetitive lines was cutted off, and more emphasizes was made to the use of silicon to decrease excessive ROS formation in soybean stressed plants.

Moreover, in the lines 79-84 was described the indirect link that Si could be in the accumulation of different antioxidant not enzymatic components for the ameliorate of effects of stress in plants. “Proline reduces the toxic effects of ROS [10], and stabilizes signaling proteins involved in drought tolerance, such as kinases and transcription factors [10]. In common bean (Phaseolus vulgaris L.), Si application has been reported to increase osmolyte and metabolite accumulation, thereby enhancing cell turgor and water uptake [23].” [lines 86-89].

Line 107–115-The hypothesis is implicit. Please explicitly state your working hypothesis (e.g., “We hypothesized that sorbitol-stabilized Si improves drought resilience by enhancing root traits and antioxidant activity”).

Dear Reviewer, thank you for your comment. In the last paragraph of the introduction, we tried to add the hypothesis after the objective without become too repetitive and more clear for the reader the principal idea of the study: “Although numerous studies have demonstrated the benefits of Si in enhancing plant tolerance to abiotic stresses, the underlying physiological and metabolic mechanisms, particularly in soybean, remain insufficiently understood. In this context, the objective of this study is to evaluate how increasing Si content in plant tissues, especially through a sorbitol-stabilized formulation, influences the physiological, biochemical, and biometric traits of soybean under both well-watered and drought conditions. We hypothesize that Si mitigates drought stress, enhances agronomic performance, and stimulates the production of osmolytes and non-enzymatic antioxidants in soybean cells—effects that are expected to be more pronounced when a stabilized Si source is used.” [lines 106-120].

Lines 118–128-Figure 1 combines multiple variables (SPAD, leaf area, etc.). Consider splitting into subfigures or enhancing legends for clarity. The differences between water and Si treatments could also be summarized in a supporting table.

Dear Reviewer, thank you for your comment. As the studies had too variable evaluated we tried to summarize in figures that had related variables, as in figure 1 are demonstrated the results of the “Experiment I”. For evaluate the significant effect of Si and drought in the variables, we created a table in the supplementary material to describe exactly the significance difference inside each factor (A3). It was added a new topic in each figure description, for the reader that want to see the relation of Si and water in the variable check in the supplementary material “Detailed p-values for each variable can be found in supplementary material Table A3.”

Lines 147–164-While SiKe improved root parameters significantly, the physiological reasons (e.g., hormonal modulation, osmotic adjustment) are not deeply discussed. Recommend expanding the discussion with literature support (Tripathi et al., 2022; Abdullah et al., 2025).

Dear Reviewer, thank you for your comment. To better support the discussion regarding the effects of Si on root parameters and its influence on the expression of auxin-related genes, we have included an additional paragraph in the Discussion section: “These findings align with previous studies [13, 29, 31], which suggest that enhanced root development under Si supply may be linked to the modulation of gene expression associated with root growth—particularly genes involved in auxin biosynthesis and transport, such as YUC, PIN, and WAT. In addition, improved allocation of photoassimilates to belowground organs have been proposed as a complementary mechanism. Supporting this, Tripathi et al. (2022) reported that soil-applied Si in soybean enhances the activity of phenolic compounds, osmoprotectants, and antioxidant enzymes. These biochemical responses help maintain chlorophyll homeostasis during drought stress and ensure a steady supply of sugars to support sustained root growth.” [lines 329-338].

Lines 188–199-The authors note strong variation but do not explain why proline was reduced under Si or how this links to stress mitigation. Consider relating this to energy trade-offs and osmoprotection balance.

Dear Reviewer, thank you for your comment. In the Discussion topic, it was added a step in which the decrease of the Proline for the Si, especially for the SiKe treatment demonstrated a decrease in the accumulation of osmoprotector, which can mean a low stress condition: “This stress moderation may have resolved an energy allocation trade-off by negating the need for the energetically costly synthesis of osmoprotectants such as proline. As a result, limited carbon and nitrogen resources could be preferentially directed toward vegetative or reproductive growth processes [37]. Additionally, reduced proline levels may reflect a metabolic reallocation, in which this amino acid is diverted toward the biosynthesis of other stress-related compounds that support cellular homeostasis [37].” [lines 399 – 405].

Lines 217–233-PCA interpretation is sound, but the figure (Figure 7) lacks quantitative clarity. Suggest labeling major contributing variables on the PCA axes or including a loading table in the supplementary material.

Dear Reviewer, thank you for your comment. For a better interpretation of the PCA we have included Table A4 in the Supplementary Material with the loadings for PC1 and PC2. The loadings clarify variable contributions to each component and confirm the vector directions in the biplot. The PC1 (and/or PC2) axis was multiplied by –1 only to ensure graphical consistency; this does not change the data structure. In the article, it was included a description in the legend of the figure 7 to inform the reader for access the Supplementary Material “To detail the contribution of each variable to PC1 and PC2, we have included the full loadings in Table A4 of the Supplementary Material.” [lines 249-250].

Lines 238–251-The heatmap is visually useful but would benefit from a numerical correlation summary in the text. Explain how decreased amino acid levels under SiKe indicate improved metabolic homeostasis.

Dear Reviewer, thank you for your comment. We appreciate the suggestion. We would like to clarify that the heatmap presented in Figure 8 uses z-score standardization, highlighting the relative increase or decrease of each amino acid among treatments rather than the correlation between variables. In this way we included a better discussion in the text to debate the decrease in amino acid content under Si application and the improved metabolic homeostasis [line 261-275]: “Interestingly, under well-watered conditions, control plants exhibited consistently higher z-scores for total amino acids, valine, threonine, serine, proline, histidine, and alanine compared to SiKe-treated plants, indicating a clear reduction in the relative accumulation of these metabolites in the SiKe/Well-watered treatment. Specifically, total amino acid content shifted from a strongly positive z-score in the Control/Well-watered group (0.91) to a markedly negative value in the SiKe/Well-watered group (−0.76). Valine followed a similar trend, decreasing from 0.93 to −0.63, while threonine exhibited an even more pronounced reduction, dropping from 1.12 to −1.13. Serine showed one of the most significant contrasts, declining from 1.31 to −0.91, and proline decreased from 0.98 to −1.04, indicating substantial suppression. Histidine presented the most intense decline among the amino acids analyzed, decreasing from 0.92 to −1.24, while alanine also dropped significantly, from 0.96 to −0.76. Collectively, these results suggest that SiKe application under non-limiting water conditions significantly modulates amino acid metabolism. This likely reflects adjustments in nitrogen assimilation and carbon–nitrogen balance, rather than a stress-induced accumulation pattern.”

In the discussion the impact of use Si in the amino acid homeostasis was discussed in [lines 385 - 409].

Lines 259–297-The discussion summarizes results effectively but often repeats descriptive findings instead of interpreting the mechanistic basis (e.g., Si-induced gene regulation, antioxidant pathway priming). Expand connections to Si transporters or phenylpropanoid regulation.

Thank you for your comment. The Discussion section has been extensively revised to address the issues of repetition and overly expository language. The updated version places greater emphasis on analytical interpretation, integrates mechanistic explanations, and provides a more critical synthesis of the findings, thereby improving the overall clarity and scientific depth of the manuscript.

Lines 298–310-Excellent interpretation regarding daidzein induction. Still, the authors should compare these findings with previous molecular-level studies (e.g., La et al., 2023; Trush & Pal'ove-Balang, 2023) to strengthen novelty claims.

Dear Reviewer, thank you for your comment. We appreciated. To better address the molecular basis underlying isoflavonoid induction, we have now incorporated two additional references and expanded the discussion in the manuscript. These studies provide molecular-level evidence supporting the involvement of key enzymes and regulatory pathways in isoflavonoid biosynthesis. The revised text clarifies how our findings align with and advance previous knowledge in this area: “These results suggest that Si may act as an elicitor of key genes and transcription factors involved in isoflavonoid biosynthesis. This is supported by findings from Chen et al. (2023) [31], Moreover, Si has been reported to enhance the activity of phenylalanine ammonia-lyase (PAL), a key enzyme in the phenylpropanoid pathway, thereby stimulating isoflavonoid synthesis [32; 33].” [lines 357 – 361].

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript explores how a sorbitol-stabilized silicon source (SiKe) influences soybean growth under both drought and well-watered conditions. The authors use two well-structured greenhouse experiments with a 3×2 factorial design and present an extensive set of physiological, biochemical, and root-related measurements. Their results show that SiKe improves Si uptake, root development, antioxidant activity (notably ascorbic acid), and daidzein levels, indicating a potential role in enhancing drought resilience.

The findings are interesting and relevant, but several areas need clarification or improvement before the work can be considered for publication. In particular, the statistical analyses should be described more clearly, some figures require better labeling and readability, and several mechanistic interpretations should be toned down or more carefully framed. The discussion of yield-related parameters also needs refinement, especially since these traits did not show improvement under drought stress.

Overall, the study has merit and could make a useful contribution to the literature on silicon-mediated stress tolerance in soybean, but major revisions are needed to improve clarity, ensure accurate interpretation of results, and strengthen the overall presentation.

Author Response

The manuscript explores how a sorbitol-stabilized silicon source (SiKe) influences soybean growth under both drought and well-watered conditions. The authors use two well-structured greenhouse experiments with a 3×2 factorial design and present an extensive set of physiological, biochemical, and root-related measurements. Their results show that SiKe improves Si uptake, root development, antioxidant activity (notably ascorbic acid), and daidzein levels, indicating a potential role in enhancing drought resilience.

The findings are interesting and relevant, but several areas need clarification or improvement before the work can be considered for publication. In particular, the statistical analyses should be described more clearly, some figures require better labeling and readability, and several mechanistic interpretations should be toned down or more carefully framed. The discussion of yield-related parameters also needs refinement, especially since these traits did not show improvement under drought stress.

Overall, the study has merit and could make a useful contribution to the literature on silicon-mediated stress tolerance in soybean, but major revisions are needed to improve clarity, ensure accurate interpretation of results, and strengthen the overall presentation.

Dear Reviewer, thank you for your comment. Throughout the manuscript, several revisions were implemented, including the addition of more critical and detailed sections to strengthen the discussion and provide a deeper interpretation of our results. These updates were guided by previous studies conducted in similar systems, which allowed us to better explain the morphological (e.g., root traits) and physiological (e.g., isoflavonoid accumulation) responses from a molecular perspective and to more effectively contextualize our findings within the broader literature.

For greater clarity regarding the statistical analyses, the figure legends were updated to indicate where the corresponding p-value tables can be found in the Supplementary Material. In the Results section, we carefully reviewed potential gaps or insufficiently developed explanations and refined the text when necessary. Additionally, we clarified in the figure legends that side-by-side and combined graphical representations denote whether or not factor interactions were detected in the statistical analysis “[Detailed p-values for each factor can be found in supplementary material Table A3.]”.

Regarding the yield experiment, the discussion was not expanded in depth because the results did not show a clear or consistent pattern, limiting the extent to which meaningful interpretations or broader conclusions could be drawn.