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Article

Interspecific Variation in the Antioxidant Potential of Culinary and Medicinal Herbs

by
Anna Rusaczonek
1,*,
Patryk Sankiewicz
1,
Maria Duszyn
2,
Mirosława Górecka
1,
Katarzyna Chwedorzewska
1 and
Ewa Muszyńska
1,*
1
Department of Botany, Institute of Biology, Warsaw University of Life Sciences-SGGW, Nowoursynowska 159, 02-776 Warsaw, Poland
2
Department of Plant Genetics, Breeding and Biotechnology, Institute of Biology, Warsaw University of Life Sciences-SGGW, Nowoursynowska 159, 02-776 Warsaw, Poland
*
Authors to whom correspondence should be addressed.
Agriculture 2025, 15(15), 1586; https://doi.org/10.3390/agriculture15151586
Submission received: 18 June 2025 / Revised: 16 July 2025 / Accepted: 22 July 2025 / Published: 24 July 2025
(This article belongs to the Special Issue Sensory Properties Analysis for Quality Evaluation of Agricultural Product)
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Abstract

Herbs are valued for their antioxidant richness and traditional use in cuisine and medicine. This study analysed wild herbs (e.g., Achillea, Lamium) and cultivated spices (Salvia, Artemisia) for their bioactive compounds. It was found that antioxidant profiles varied notably among species, even within the same family. Helichrysum italicum and Salvia officinalis had the highest polyphenol levels, while Achillea millefolium and Ocimum basilicum had the lowest. Total polyphenols did not always correlate with antioxidant activity. For instance, Petroselinum hortense and Salvia rosmarinus showed high antioxidant activity despite low polyphenol levels, whereas Levisticum officinale and Artemisia dracunculus combined both. Mentha spicata, M. x citrata, Origanum vulgare, and S. officinalis were rich in carotenoids, while H. italicum showed high α-carotene but low levels of other carotenoids. Most Lamiaceae accumulated a high amount of chlorophylls and polyphenols. Cultivated herbs like M. spicata, M. x citrata, and S. officinalis exhibited stronger and more diverse properties than wild species. It can be concluded that taxonomy alone does not predict antioxidant potential. The differences observed may be attributed to species-specific metabolic pathways, ecological adaptations, or environmental factors influencing phytochemical expression. These findings highlight the importance of conducting species-level screenings in the search for plant-derived antioxidants with potential therapeutic applications.

Graphical Abstract

1. Introduction

Reactive oxygen species (ROS) refer to a diverse array of oxidant molecules with varying properties and biological functions, ranging from signalling to inducing cell damage. ROS are continuously generated in plants as by-products of aerobic metabolism [1]. They are produced in various cellular compartments, including mitochondria, chloroplasts, peroxisomes, and the cytosol, where their production is tightly regulated to prevent the release of intermediate products. When plants are under stress induced by biotic or abiotic factors, like heavy metals, oxidising pollutants (like ozone and sulfur dioxide), water deficit, salinity, and temperature fluctuations or pathogens, their production of ROS can exceed their detoxification ability and lead to ROS overproduction [2].
Under unstressed conditions, the formation and removal of ROS are generally in equilibrium. Plants have developed efficient antioxidant defence systems to scavenge ROS and protect against harmful reactions. These systems include enzymatic antioxidants, like superoxide dismutases, peroxidases, catalase, and glutathione reductase, as well as non-enzymatic antioxidants, such as ascorbate, glutathione, tocopherol, carotenoids, polyphenols, and alkaloids [3]. Protective enzymes and low-molecular-weight antioxidants are consistently present in plants. However, their effectiveness can vary in response to internal and external factors, including environmental conditions and developmental processes [4]. Among enzymes, superoxide dismutases (SODs) serve as the first line of defence against ROS and play a significant role in helping plants respond to environmental stresses by dismutation of superoxide radicals to oxygen and hydrogen peroxide (H2O2). Then, catalase (CAT) removes H2O2 generated in peroxisomes by oxidases involved in the β-oxidation of fatty acids, the glyoxylate cycle (which occurs during photorespiration), and purine catabolism [5]. Similarly, peroxidases (POXs) reduce H2O2 and alkyl hydroperoxides by oxidising different substrates in various cellular compartments, including chloroplasts and mitochondria [6]. Therefore, the balance between the activities of the mentioned enzymes is crucial for establishing a stable level of superoxide radicals and hydrogen peroxide.
Polyphenols are a diverse group of antioxidants comprising over 10,000 compounds [7,8]. These phenolic compounds include secondary metabolites, such as flavonoids, tannins, hydroxycinnamate esters, and lignin, which are crucial in the plant cells’ hydrogen peroxide scavenging process. Certain polyphenols, like kaempferol and naringenin, have an ideal structural configuration for radical scavenging, making them more effective than tocopherols and ascorbate [9]. The antioxidant properties of polyphenols stem from several factors: (i) their high reactivity as hydrogen or electron donors, (ii) the capacity of polyphenol-derived radicals to stabilise and delocalise an unpaired electron, which acts as a chain-breaking function, (iii) their ability to chelate transition metal ions, thereby terminating the Fenton reaction, and (iv) the role of flavonoids in altering peroxidation kinetics [10]. Specifically, flavonoids modify lipid packing, reducing membrane fluidity and making it more difficult for free radicals to diffuse, inhibiting peroxidative reactions [11]. These mechanisms contribute to the overall effectiveness of polyphenols as antioxidants.
The second most frequently mentioned group of compounds is carotenoids, liposoluble pigments comprising over 600 different types. These are classified into xanthophylls and carotenes [12]. Carotenoids are found in the fruits and roots of plants and in chloroplasts. They serve as accessory light-harvesting pigments and function as antioxidants. β-carotene, in particular, is known for its ability to quench singlet oxygen and react quickly with free radicals. The resulting radicals are more energetically stable, making them less likely to engage in harmful chain reactions. Additionally, β-carotene interacts with triplet states of specific photosensitising agents, especially ketonic sensitisers, whose excited states resemble diradicals and slow down the initiation of chain reactions [13]. This interaction likely occurs within chloroplasts and helps to quench unwanted excited states of chlorophyll that could otherwise lead to the production of singlet oxygen or other harmful species. Notably, the content of β-carotene increases during severe water drought [14]. Carotenoids protect photosystems through several mechanisms: (i) by reacting with lipid peroxidation products to terminate chain reactions, (ii) by scavenging singlet oxygen and dissipating energy as heat, (iii) by dissipating excess excitation energy via the xanthophyll cycle, (iv) by interacting with triplet or excited chlorophyll molecules to prevent singlet oxygen formation, and (v) by augmenting α-tocopherol in scavenging peroxy-radicals [15].
The least frequently mentioned group capable of scavenging reactive oxygen species is chlorophylls. Chlorophylls can directly scavenge various ROS, including superoxide anions, hydroxyl radicals, and singlet oxygen. This ability is due to their conjugated double bond system, which allows for electron donation to stabilise the radicals [16]. Additionally, chlorophylls and their derivatives can chelate metal ions (such as Fe2+ and Cu2+), which reduces the Fenton reaction and consequently decreases ROS production. This process helps protect cell membranes by inhibiting lipid peroxidation [17,18]. Furthermore, by absorbing ultraviolet and visible light, chlorophylls can reduce light-induced oxidative stress and exhibit activity against mutagens like aflatoxins [19].
Recent studies have increasingly highlighted the antioxidant potential of herbs belonging to the Lamiaceae, Apiaceae, and Asteraceae families due to their richness in bioactive compounds, such as polyphenols, carotenoids, and essential oils [15,20,21]. Species from these families are widely utilised as culinary herbs, traditional medicines, aromatic plants, and ingredients in cosmetic formulations, which underscores their significance not only in phytochemical and pharmacological studies but also in the development of products for nutrition and healthcare. However, comparative studies that integrate both enzymatic and non-enzymatic antioxidant traits across these families remain limited. Therefore, in this study, we selected 15 representative herb species, both wild-growing and commonly cultivated, and evaluated them with respect to total polyphenol content, enzymatic antioxidant activity, and pigment composition to assess their antioxidant potential in a species-specific context. This random choice of taxa included in our investigation allowed for obtaining diverse research material and thus a more objective assessment of the antioxidant potential of plants belonging to different botanical families. We hypothesised that the tested species would exhibit notable interspecific variation in their antioxidant profiles, primarily driven by variation in their metabolic pathways and antioxidant defence strategies, rather than merely their taxonomic classification.

2. Materials and Methods

2.1. Plant Material

Fifteen species of herbs from three botanical families, which are important in terms of their use in phytotherapy and in everyday diet, were randomly chosen for analysis (Table 1). The plants were cultivated in a universal growing substrate based on sphagnum peat moss (pH 5.5–6.5) mixed with perlite in a ratio of 3:1 (v/v), and its humidity was maintained at 60–70% of the maximum water capacity, which corresponds to the optimal conditions for most herbal plants [22]. Fertilisation was applied twice by using Hoagland solution to water the plants at the beginning and after four weeks of growth. The controlled, identical conditions in the climatic chamber were applied (temperature of 24 °C ± 1 °C, and a photoperiod of 16 h:8 h (light:dark)) to eliminate the environmental effects on the accumulation of health-related compounds. After eight weeks of cultivation, the aerial parts of each species were collected separately, ground to prepare a bulk sample, and immediately placed in liquid nitrogen.

2.2. Total Polyphenol Content

The total polyphenol content was measured with the use of the Folin–Ciocâlteu reagent, following the method of Singleton and Rossi [23]. Approximately one hundred milligrams of frozen tissue was homogenised in a Mixer Mill MM 400 (Retsch, Düsseldorf, Germany) (5 min, 30 Hz) with 1 mL of 80% methanol and centrifuged (15 min, 13,000 rpm, RT). The obtained supernatant was mixed with deionised water, Folin–Ciocâlteu reagent, and saturated sodium carbonate (Na2CO3). The absorbance measurement was performed after incubating samples in the dark (45 min, 23 °C) at 765 nm using a microplate reader Multiscan-GO (Thermo Scientific, Waltham, MA, USA). The results were expressed as mg of gallic acid equivalents (GAEs) per 1 g of fresh weight (FW).

2.3. ABTS•+ Scavenging Activity

The antioxidant potential of the methanol extracts was analysed by the scavenging ability of ABTS cation radicals. First, 80–100 mg of frozen tissue was homogenised in a Mixer Mill MM 400 (Retsch, Düsseldorf, Germany) (5 min, 30 Hz) with 1 mL of 80% methanol and centrifuged (15 min, 13,000 rpm, RT). The scavenging activity against ABTS•+ was investigated as previously described, following the original method of Re et al. [24]. To obtain the green-blue ABTS•+ solution, an aqueous mixture of 7 mM ABTS (2,2′-azinobis (3-ethylbenzothiazoline-6-sulfonic acid)) and 2.45 mM potassium persulfate was left to stand in the dark overnight at room temperature. Later, the ABTS•+ solution was diluted with ethanol until the absorbance was 0.7 ± 0.02 at 734 nm. The supernatant of the prepared samples was further mixed with ABTS•+ solution, and after 6 min of reaction, the absorbance was measured at a wavelength of 734 nm using a microplate reader (Multiscan-GO, Thermo Scientific, Waltham, MA, USA). The results were presented as µmoles of Trolox equivalent (TE) per 1 g of FW.

2.4. Antioxidant Enzyme Activity—Catalase (CAT) and Superoxide Dismutase (SOD)

For enzyme extraction, 80–100 mg of frozen tissue was homogenised in a Mixer Mill MM 400 (Retsch, Düsseldorf, Germany) (5 min, 30 Hz, 4 °C) with 1 mL of extraction buffer (100 mM Tricine, 3 mM MgSO4, 3 mM EGTA, 1 mM dithiothreitol, 1 M Tris/HCl, pH 7.5). The samples were incubated on ice for 15 min and centrifuged (13,000 rpm, 20 min, 4 °C). The protein concentration of the enzyme extracts was determined using a Bradford assay kit (Thermo Scientific, Rockford, IL, USA) with bovine serum albumin (BSA) as a standard. The spectrophotometric assay of superoxide dismutase (SOD) activity was conducted following the original method established by Beauchamp and Fridovich [25], incorporating modifications as previously documented [26]. The enzyme assay mixture comprised 0.1 M phosphate buffer at a pH of 7.5, 2.4 μM riboflavin, 840 μM NBT, 150 mM methionine, and 12 mM Na2EDTA at a ratio of 8:1:1:1:1 (v/v/v/v/v). The enzyme extract was combined with the enzyme assay mixture to ensure the inhibition of NBT oxidation within the range of 20% to 80%. Absorbance measurements were taken at 560 nm, −15 min after exposure to either 500 μmol m−2 s−1 illumination (white LED light source, SL 3500, Photon Systems Instruments, Drásov, Czech Republic) or incubation in darkness as a blank sample. The results were expressed in units representing the quantity of enzyme required to inhibit NBT photoreduction to blue formazan by 50% per milligram of protein. Catalase (CAT) activity was determined spectrophotometrically, following the methodology outlined in the original method by Aebi [27], with some adjustments [26]. Perhydrol was diluted with 50 mM phosphate buffer (pH 7.0) to achieve an absorbance of 0.5 (±0.02) at 240 nm, representing an initial H2O2 concentration of approximately 13 mM. The enzyme extract was combined with reaction buffer, resulting in a decrease in absorbance, falling within the 20% to 80% range. CAT activity was quantified based on the rate of H2O2 degradation observed over a 2 min period, utilising the molecular extinction coefficient of H2O2 at 240 nm (ε = 43.6 mol−1 cm−1). The results were expressed in units of micromoles of H2O2 per minute per milligram of protein. All spectrophotometric measurements were performed using a UV-VIS microplate reader Multiscan-GO (Thermo Scientific, Waltham, MA, USA).

2.5. Pigments Analysis

First, 20–50 mg of frozen tissue was homogenised in a Mixer Mill MM 400 (Retsch, Düsseldorf, Germany) (5 min, 30 Hz, 4 °C) with 1 mL of cold acetone (−20 °C). Then, the homogenate was evaporated using Savant DNA120 SpeedVac (Thermo Scientific, Waltham, MA, USA), dissolved in cold solvent A (acetonitrile: methanol; 90:10; v/v), and re-homogenised for 1 min. The extract was filtered through a 0.2 μm nylon filter (Whatman, Marlborough, MA, USA) into an auto-sampler vial, capped, and stored in the dark at −80 °C for HPLC analysis (Shimadzu, Kyoto, Japan). The pigments were separated on a SynergiTM 4 μm MAX-RP 80 Å LC Column 250 × 4.6 mm (Phenomenex, Torrance, CA, USA) at 30 °C. Solvent A was used for 10 min to elute all xanthophylls, followed by solvent B (methanol–ethyl acetate; 68:32; v/v) for 10 min at a flow rate of 1 mL min−1 to elute the rest of the pigments (chlorophylls and carotenes). The results were given as the peak area per μg of FW, according to the previously used protocol [26,28]. Pigments were identified using standards (Sigma (St. Louis, MO, USA), Hoffmann-LaRoche (Basel, Switzerland), Fluka (Buchs, Switzerland)). The total carotenoid content was expressed as the sum of the individual amounts of xanthophylls and carotenes.

2.6. Data Analysis

In all experiments, twelve plants of each species were used in three biological replicates and three technical ones. Statistical analysis was performed in STATISTICA 13.0 software (StatSoft, Inc., Tulsa, OK, USA) using one-way ANOVA and post hoc Tukey honest significant difference (HSD) test at the significance level of 0.05. For all data, a Shapiro–Wilk normality test and Bartlett test of variance homogeneity were performed. For heatmap visualisation, the data were normalised within each experiment (min–max normalisation: 0–100) and plotted using GraphPad Prism version 8.0 software (Dotmatics, San Diego, CA, USA).

3. Results

3.1. Natural ROS Scavengers and Their Antioxidant Potential

This study measured total polyphenol content expressed as GAE (gallic acid equivalent). The accumulation of polyphenols varied from 0.24 to almost 1.0 mg GAE g−1 FW (Table 2). Among the tested herbs, the highest amount of polyphenols was detected in Helichrysum italicum (0.99 mg GAE g−1 FW) and Salvia officinalis (0.92 mg GAE g−1 FW). A statistically lower but still high polyphenol content of about 0.8 mg GAE g−1 FW was noticed in some representatives of the Lamiaceae family, including all Mentha species, Origanum vulgare, and Thymus vulgaris. Although these species showed one of the highest amounts of polyphenols, the antioxidant activity of their methanolic extracts was relatively low and varied between 29 and 44 µmol TE g−1 FW (Table 2). The exception was M. spicata, which reached 66 µmol TE g−1 FW. A higher activity by about 15% was determined only for Salvia rosmarinus. A moderate quantity of polyphenols at the level of 0.7 mg was found in Levisticum officinale, Petroselinum hortense, and Artemisia dracunculus. Despite a comparable amount, their antioxidant activity was strongly diversified. The highest ability to scavenge ABTS radicals was ascertained for a methanolic extract of P. hortense (app. 70 µmol TE g−1 FW), which was more than 4.5- and 2.5-times higher than for L. officinale and A. dracunculus, respectively. In turn, L. album, along with A. graveolens and A. millefolium, contained twice as many phenolic compounds as the lowest amount determined for O. basilicum, which was only about 0.24 mg GAE g−1 FW. Among the mentioned species, there were also those with the lowest antioxidant activity, i.e., A. millefolium (5 µmol TE g−1 FW) and then O. basilicum (15 µmol TE g−1 FW). Interestingly, only the Asteraceae family’s tested representatives showed that the increasing polyphenol content strongly corresponded with the increasing antioxidant activity. These results indicated that the phenolic compounds made a significant contribution to the antioxidant capacity of some of the analysed herbs (Table 2).
In this research, the highest catalase (CAT) activity was determined for L. officinale and Mentha citrata (59 and 55 µmol H2O2 min−1 mg−1 protein, respectively; Table 3). An average enzyme activity of approximately 40 µmol H2O2 min−1 mg−1 protein was observed in A. dracunculus and M. spicata. In contrast, the activity in A. graveolens, H. italicum, T. vulgaris, and S. officinalis was only about 12% lower. In turn, P. hortense, A. millefolium, and M. arvensis showed comparable CAT activity. Additionally, the CAT activity in the last-mentioned herb reached a similar level to O. vulgare and S. rosmarinus. It was more than two times higher than the lowest activity found in L. album and O. basilicum (8–9 µmol H2O2 min−1 mg−1 protein). On the other hand, O. basilicum demonstrated the highest superoxide dismutase (SOD) activity at the level of 127 U mg−1 protein, which was comparable to A. millefolium (Table 3). Significantly reduced SOD activity was noted in A. dracunculus and S. officinalis (app. 100 U mg−1 protein), but also in L. officinale (96 U mg−1 protein), which, in this case, corresponded to the activity in S. rosmarinus (88 U mg−1 protein). H. officinale and O. vulgare maintained SOD activities higher by about 12% than M. x citrata and T. vulgaris. In turn, M. spicata and M. arvensis exhibited enzyme activity similar to L. album and A. graveolens. In these plants, SOD activity reached app. 55 U mg−1 protein, which was higher by about 33% than the lowest one determined for P. hortense. Interestingly, in species from the Apiaceae family, SOD activity changed comparably to CAT. At the same time, in other the families, the tendency was rather opposite, and an increase in the activity of one enzyme seemed to be accompanied by a decrease in the activity of the other.

3.2. Total Carotenoids, Carotene and Xanthophyll Accumulation, and Their Ratios

The lowest amount of total carotenoids was determined in H. italicum (1381 peak area μg−1 FW), which reached a similar level to P. hortense, M. arvensis, O. basilicum, and S. rosmarinus (Figure 1A). In turn, L. officinale, A. dracunculus, L. album, M. spicata, and T. vulgaris accumulated an average content of app. 2300 peak area μg−1 FW. In the last-mentioned one, it was additionally comparable to A. graveolens and A. millefolium (app. 1900 peak area μg−1 FW). A significantly higher amount of total carotenoids was noticed in M. x citrata, S. officinalis, and O. vulgare, which achieved the highest value of app. 2800 peak area μg−1 FW. These herbs were also characterised by the highest amount of total xanthophylls, which was about 57% higher than the lowest one ascertained in H. italicum (883 peak area μg−1 FW; Figure 1B). Similarly, a high accumulation of xanthophylls was established in L. album and L. officinale. Still, in this case, there were no statistical differences with other species like A. dracunculus and M. spicata, which, in turn, amassed the same amount of app. 1750 peak area μg−1 FW as A. graveolens and T. vulgaris. The last group of herbs constituted P. hortense, A. millefolium, M. arvensis, O. basilicum, and S. rosmarinus. They accumulated approximately 1300 peak area μg−1 FW of total xanthophylls, reaching an amount higher by about 34% than the lowest one in H. italicum (Figure 1B).
The lowest amount of total carotenes was noted in A. graveolens (195 peak area μg−1 FW; Figure 1C). In turn, P. hortense, M. arvensis, O. basilicum, and S. rosmarinus accumulated on average 114 peak area μg−1 FW more carotenes than A. graveolens but significantly less, by about 34%, than Levisticum officinale, A. millefolium, H. italicum, L. album, and T. vulgaris. The concentration of total carotenes in A. dracunculus, M. spicata, O. vulgare, and S. officinalis reached 650 peak area μg−1 FW, while in M. x citrata, it was 783 peak area μg−1 FW, which was the highest value among all the tested species (Figure 1C).
The highest share of xanthophylls in the total pool of carotenoids was found in A. graveolens (Figure 2A). A statistically significantly lower but still high value of this ratio was found in L. album (82%), where it was also comparable with L. officinale and P. hortense from the same botanical family, as well as with M. arvensis and S. rosmarinus from Lamiaceae. In general, the representatives of Lamiaceae were characterised by a ratio of total xanthophylls to total carotenoids close to 80%, and the lowest value of this parameter was calculated for M. x citrata (73%) and M. spicata (74%). A similar proportion of xanthophylls and carotenoids, varying between 74 and 76%, was determined for A. millefolium and A. dracunculus. They belong to the Asteraceae family, together with H. italicum, for which the lowest xanthophylls to carotenoids ratio was found among the tested species (64%). On the other hand, H. italicum was characterised by the highest ratio of total carotenes to carotenoids, which was about 70% higher than in A. graveolens, which had the lowest value of this parameter (11%; Figure 2B). For L. officinale, P. hortense, M. arvensis, and S. rosmarinus, the calculated ratio was simultaneously comparable with both L. album and O. basilicum and varied between 0.18 and 0.21%. A statistically higher ratio was determined for O. vulgare and T. vulgaris, forming a homogeneous group with A. millefolium and S. officinalis (24%). These last-mentioned species achieved a similar carotenes/carotenoids ratio to M. spicata and A. dracunculus (26%), which, in turn, were statistically close to M. x citrata (Figure 2B).

3.3. The Content of Lipid-Soluble Pigments from Particular Carotenoid Groups

The lutein content ranged from 498 peak area μg−1 FW to 950 peak area μg−1 FW in the herbs tested (Figure 3A). Its highest amount was detected in M. x citrata (950 peak area μg−1 FW) and Salvia officinalis (896 peak area μg−1 FW). However, the last species accumulated a similar amount of lutein as O. vulgare, which, in turn, did not differ from M. spicata (824 peak area μg−1 FW). The next group of plants with a comparable lutein content at the level of 710–762 peak area μg−1 FW included A. millefolium and T. vulgaris, but also A. graveolens and A. dracunculus. A similar amount of lutein was accumulated by L. album. Despite this, its content was simultaneously statistically the same as in L. officinale, H. italicum, and M. arvensis (on average 599 peak area μg−1 FW), forming a homogeneous group with P. hortense and O. basilicum (app. 570 peak area μg−1 FW). The lowest lutein concentration in S. rosmarinus did not exceed 498 peak area μg−1 FW.
The highest percentage of lutein in the total pool of xanthophylls was found in H. italicum, followed by A. millefolium, where it constituted 67% and 51%, respectively (Figure 3B). These values were statistically higher than in P. hortense, all Mentha species, and T. vulgaris, in which the lutein/xanthophylls ratio was slightly lower than 50%. Except for M. spicata, the mentioned herbs showed comparable values of this parameter to A. graveolens and S. officinalis (44%). In A. dracunculus, O. vulgare, and S. rosmarinus, lutein constituted only 40% of the total xanthophylls. It was about 17% higher than in L. officinale and L. album, where this ratio was the lowest. The highest content of α-carotene was determined in H. italicum, up to 19 times more than in the lowest one described for L. album, M. arvensis, and O. basilicum (app. 5 peak area μg−1 FW; Figure 4A). P. hortense accumulated 10 peak area μg−1 FW, which was statistically lower by about 39% than in A. graveolens and S. rosmarinus. A similar amount of slightly above 20 peak area μg−1 FW was noticed for L. officinale, M. spicata, O. vulgare, S. officinalis, and T. vulgaris. Statistically higher levels of α-carotene were ascertained in A. dracunculus and M. x citrata, while in A. millefolium, it reached 42 peak area μg−1 FW.
Regarding β-carotene, its highest amount was found in M. x citrata (750 peak area μg−1 FW; Figure 4B), then in M. spicata, O. vulgare, and S. officinalis from Lamiaceae, as well as in A. dracunculus from Asteraceae (app 590 peak area μg−1 FW). The other tested species from the last-mentioned family accumulated statistically lower amount of β-carotene, about 160 peak area μg−1 FW, which was also comparable to L. album, T. vulgaris, and L. officinale. Four species, namely, P. hortense, M. arvensis, O. basilicum, and S. rosmarinus, were characterised by a β-carotene content at the level of on average 288 peak area μg−1 FW, which was significantly higher than in A. graveolens, with the lowest amount of this compound (179 peak area μg−1 FW).
The percentage of determined carotenes in the total pool of carotenes was calculated for β-carotene, which constituted the largest part, varying between 82% and 99% (Figure 4C). The lowest ratio was determined in H. italicum, while the highest was in L. album and O. basilicum, as well as in M. arvensis. However, these species contained a moderate amount of β-carotene. In turn, M. spicata, O. vulgare, and S. officinalis had statistically the exact value of this ratio as P. hortense and M. x citrata. Still, the amount of β-carotene in total carotenes in M. x citrata was also comparable to T. vulgaris and L. officinale. Additionally, the calculated ratio in L. officinale put this species in a homogeneous group with S. rosmarinus, in which the proportion of β-carotene in the total pool of carotenes was similar to A. dracunculus (95%). The value of this parameter in A. graveolens and A. millefolium was higher by only about 10% than the lowest one (82%).

3.4. Accumulation of Chlorophylls

The total chlorophyll content (Figure 5A) varied between 1002 and 2413 peak area μg−1 FW. A lower amount was noted in both H. italicum, but also in L. officinale and S. rosmarinus, which accumulated a comparable quantity of pigments as A. graveolens (1343 peak area μg−1 FW). The last species had a similar amount of chlorophylls as L. album. Regarding this value, L. album also belonged to the same larger group, which accumulated on average 1790 peak area μg−1 FW and included P. hortense, A. millefolium, A. dracunculus, O. vulgare, and T. vulgaris. The total chlorophyll content in the two last-mentioned herbs reached the pigment level specified for M. arvensis and O. basilicum (on average 1989 peak area μg−1 FW). In turn, M. x citrata contained the highest total chlorophyll amount (2413 peak area μg−1 FW), which statistically did not differ from M. spicata and S. officinalis.
The tested herbs with the lowest chlorophyll a content, fluctuating between 623 and 1194 peak area μg−1 FW, can be ranked as follows: H. italicum ≤ L. officinale ≤ S. rosmarinus ≤ A. graveolens ≤ L. album (Figure 5B). Nevertheless, the chlorophyll a content in L. album was also close to P. hortense, A. millefolium, A. dracunculus, and T. vulgaris, which accumulated app. 1345 peak area μg−1 FW. Similarly, a high amount of chlorophyll a was determined in M. arvensis, O. basilicum, and O. vulgare (app. 1460 peak area μg−1 FW). In turn, S. officinalis and M. spicata were characterised by a comparable chlorophyll a level of 1680 peak area μg−1 FW, which was only about 200 peak area μg−1 FW lower than in M. x citrata. The chlorophyll b content in L. officinale and A. graveolens was significantly lower, by about 27%, than in P. hortense, A. millefolium, O. vulgare, and T. vulgaris, which achieved app. 429 peak area μg−1 FW (Figure 5C). At the same time, in A. graveolens, it was comparable to S. rosmarinus (360 peak area μg−1 FW) and A. dracunculus, H. italicum, and L. album, containing, on average, 392 peak area μg−1 FW. The highest amount of chlorophyll b was determined in O. basilicum (564 peak area μg−1 FW), M. x citrata (532 peak area μg−1 FW), and S. officinalis (525 peak area μg−1 FW). Still, the last two-mentioned herbs accumulated similar levels of chlorophyll b as M. spicata and M. arvensis, reaching app. 473 peak area μg−1 FW.

4. Discussion

Plants are vital sources of bioactive compounds produced through primary and secondary metabolism. These compounds serve a variety of functions. For instance, some provide colour to petals, attracting pollinators and signalling that fruits are ripe, aiding seed dispersal. Others act as repellents against herbivores while simultaneously playing crucial metabolic roles, such as functioning as effective antioxidants. Antioxidant compounds interact with reactive oxygen species, affecting both plants and their consumers, including humans and animals, as these compounds enter the food chain [29,30]. Reducing oxidative stress in cells can help treat various human diseases, including cancer, cardiovascular diseases, and inflammatory conditions [31]. Among plants, herbs occupy a significant place, as they are rich in antioxidants and have been used for centuries as spices, aromatic plants, or natural medicines. Therefore, we evaluated various wild-growing herbs, such as Achillea and Lamium, and those commonly cultivated as spices, e.g., Salvia or Artemisia. The presented study indicated that the analysed herbs significantly differed in the accumulation of polyphenols, carotenoids, chlorophylls, and their antioxidant activity. To visualise these variations in the examined traits, a heatmap was generated, which enables noticing some regularities (Figure 6).
Our study found that the total polyphenol content did not always correspond to its antioxidant activity, with three notable exceptions. For Helichrysum italicum and Mentha spicata, a high total polyphenol content was associated with high antioxidant activity. Conversely, in Achillea millefolium, we observed that a low total polyphenol content reflected a low antioxidant activity. In the cases of Petroselinum hortense and Salvia rosmarinus, we detected a high ABTS radical scavenging capacity but a relatively low level of polyphenols. It is highly probable that the observed variations in antioxidant responses can be attributed to the endogenous presence of other compounds with a diversified ability to neutralise ROS, which were not directly analysed in our study. Among them, ascorbic acid for parsley or isoprenoid quinones for rosemary could be mentioned as examples [32,33]. This disparity may also arise from quantitative differences in the composition of polyphenols across the analysed species, as well as mutual interactions between various antioxidants, which may modify antioxidant activity in a different way than would be expected from individual components [34]. Polyphenols contain various compounds that exhibit varying antioxidant activities [35]. The mechanism of free radical neutralisation by this group of secondary metabolites includes hydrogen atom transfer via rupturing the hydroxyl bond in their structure, electron donation or metal ion chelation to prevent the Fenton reaction, thereby inhibiting the oxidative cascades responsible for cellular damage [36]. Since polyphenols show a highly diverse chemical structure, particularly in terms of the number and position of hydroxyl groups, degree of conjugation, and potential for metal binding, their ability to modulate oxidative stress differs. For example, a notable distinction exists between regular flavonoids and tannins. Tannins are considered superior antioxidants because of their potential for oxidation, which can lead to oligomerisation through phenolic coupling. This process increases the number of reactive sites. In contrast, this type of reaction has not been observed with flavonoids themselves [37].
To fully understand the defence strategies against oxidative stress, both non-enzymatic and enzymatic antioxidant systems were thoroughly analysed. Polyphenols can collaborate with antioxidant enzymes in plants, such as catalase (CAT) and superoxide dismutase (SOD), working in complementary and synergistic ways [38]. Polyphenols enhance the protective effects of SOD and CAT against oxidative stress in plant cells, while these enzymes may also contribute to the storage capacity of plant products, particularly fruits, vegetables, and seeds. They help slow wilting, reduce spoilage, and improve the retention of colour, firmness, and flavour [39]. Polyphenols assist by scavenging reactive oxygen species (ROS) that these enzymes cannot eliminate. Under stress conditions such as drought, UV exposure, or pathogen attacks, both the levels of polyphenols and the activity of SOD and CAT increase, indicating a coordinated stress response [40]. Some studies suggest that polyphenols can also regulate the expression of genes for antioxidant enzymes, thereby enhancing their production [41]. In species like Levisticum officinale and Artemisia dracunculus, high levels of polyphenols and increased enzymatic activity have been observed, indicating a synergistic interaction between both systems. However, in Achillea millefolium and Ocimum basilicum, relatively low levels of polyphenols are found alongside high and medium enzymatic activity, which likely indicates the dominance of the antioxidant enzyme system in these species. The complex relationships between polyphenols and antioxidant enzymes in plants are highly context-dependent, varying not only among species and genotypes but also with environmental conditions and the specific composition of polyphenolic compounds. The interactions between non-enzymatic and enzymatic antioxidant systems are intricate and far from uniform, highlighting the need for further comparative research to unravel their molecular basis [29,42,43]. For instance, while methyl jasmonate application significantly enhanced antioxidant enzyme activities and improved photosynthetic efficiency in heat-stressed Triticum aestivum [44], the same treatment in Mentha piperita increased phenolic compound accumulation and overall antioxidant activity but simultaneously triggered senescence-like symptoms and reduced photosynthesis and protein content [43], illustrating species- and stress-specific divergences. Furthermore, exposure to heavy metals, such as copper or lead, often promotes phenolic accumulation and upregulation of their biosynthetic enzymes [29,43]. In turn, supra-optimal concentrations or specific metals like arsenite can inhibit key antioxidant enzymes (e.g., SOD, CAT) while leaving others unaffected (e.g., POX) [42], further underscoring the selective and context-dependent nature of polyphenol–enzyme interactions. Our results confirmed this complexity, even under controlled, non-stress laboratory conditions.
In biological systems, exposure to light triggers the formation of ROS, which can damage biomolecules and compromise the integrity and stability of subcellular structures, cells, and tissues [45]. These photooxidative processes contribute to the underlying biochemistry of diseases affecting light-exposed tissues, such as the eye and skin. Protection against these photooxidative effects has been linked to the antioxidant properties of macular carotenoids, which also filter light [46,47]. Humans cannot synthesise carotenoids; instead, they rely on dietary sources to obtain these pigments, either absorbed directly or undergoing slight structural modifications [48]. Carotenoids enhance human health and animal well-being because of their potent antioxidant and anti-inflammatory properties [15]. Among the carotenoids in leafy vegetables, lutein and β-carotene have been extensively studied for their health benefits [46]. Lutein, a yellow pigment found in leafy greens and other colourful vegetables, is essential for maintaining eye health. Furthermore, β-carotene is crucial in reducing the risk of cataract development, which is often linked to oxidative stress and free radical damage. Some research has also shown that lutein significantly accumulates in brain regions responsible for memory, emotion regulation, vision, and hearing [49]. There is also a growing interest in new research regarding carotenoids in edible plants, driven by an increasing focus on incorporating natural compounds into food. This is further supported by European Community directives that favour natural over synthetic ingredients [50]. Among the analysed plants, the highest levels of lutein and β-carotene were found Mentha spicata and M. x citrata, Origanum vulgare, and Salvia officinalis. Those plants also have the highest total carotenes and xanthophylls. An interesting observation was the high level of α-carotene found in the Helichrysum italicum, along with a low content of other carotenoids compared to the other analysed species. This compound is likely responsible for the yellow colour of the flowers in this plant, and its presence is probably abundant not only in the flowers but throughout the plant.
Despite their status as the most abundant pigments in nature, chlorophylls were often excluded from research trials [51]. However, recently, there has been a growing interest in chlorophylls and their bioactive properties, which include antioxidant, antimutagenic, antigenotoxic, and anti-obesity effects [52]. Among these, chlorophyll a exhibited the most potent antioxidant activity, followed by chlorophyll b and their metal-free derivatives, pheophorbide b and pheophytin b [53]. According to studies by Lanfer-Marquez et al. [51], chlorophylls’ antioxidant activities were comparable to those of BHT (butylated hydroxytoluene) when tested using the β-carotene bleaching method. What seems to agree with our studies is that in the Lamiaceae family, the total chlorophylls and both chlorophyll a and b contents were the highest, except for S. rosmarinus, and coincided with the highest accumulation of polyphenols. Furthermore, plant leaves are recognised as rich sources of edible antioxidants. For instance, the chlorophyll (both a and b) content in stem amaranth (Amaranthus lividus) leaves showed a significant positive correlation with the total antiradical activity [54]. A similar relationship was noticed in Abies alba and Nepeta pannonica [55]. Contrarily, in our studies, representatives of the Apiaceae and Asteraceae families, which are commonly used as green leafy herbs, such as Anethum graveolens, Levisticum officinale, or Helichrysum italicum, were relatively poor in chlorophyll content. However, they had a moderate, species-dependent level of other antioxidants, like polyphenols or antioxidant enzymes.
To sum up, one should be aware that our study was conducted under controlled conditions to enable direct species screening and interspecific comparisons. However, field environments, characterised by rapid fluctuations and interactions of multiple abiotic factors—such as drought, temperature and light variations, soil properties, and climate change—may cause substantially different antioxidant responses. Balance between ROS production and scavenging via both enzymatic (e.g., SOD, CAT, APX) and non-enzymatic (e.g., polyphenols, glutathione, carotenoids) mechanisms is highly stress-dependent and can vary significantly between plant species and when stresses act individually or in combination, which controlled conditions may not fully capture in terms of field suitability [56,57]. Therefore, further field studies are needed to validate the antioxidant potential of the investigated species under natural environmental conditions. Additionally, considering that statistical significance provides useful information about differences between species, but it does not reflect how large or biologically meaningful those differences are, future experiments should incorporate effect size statistics alongside p-values. Such an approach could allow for a deeper and more complete understanding of the practical relevance of the observed effects and support stronger biological conclusions, particularly in the context of multiple comparisons across so many species.

5. Conclusions

The outcomes of this experiment indicate significant interspecific variation in both the content of antioxidant compounds and antioxidant activity, even among species within the same family. This suggests that taxonomic relatedness alone is not a reliable predictor of antioxidant potential. The highest amount of polyphenols was detected in Helichrysum italicum and Salvia officinalis, and the lowest was detected in Achillea millefolium and Ocimum basilicum. Our study found that the total polyphenol content did not always correspond to its antioxidant activity, with three notable exceptions. For H. italicum and Mentha spicata, a high total polyphenol content was associated with high antioxidant activity. Conversely, in A. millefolium, a low total polyphenol content reflected a low antioxidant activity. In the cases of Petroselinum hortense and Salvia rosmarinus, there was a high scavenging activity but a relatively low level of polyphenols. In species like Levisticum officinale and Artemisia dracunculus, high levels of polyphenols and increased enzymatic activity were observed, indicating a synergistic interaction between both systems. However, in A. millefolium and O. basilicum, relatively low levels of polyphenols were found alongside high and medium enzymatic activity, which likely indicates the dominance of the antioxidant enzyme system in these species. Among the analysed plants, the highest levels of lutein and β-carotene were found in Mentha spicata and M. x citrata, Origanum vulgare, and Salvia officinalis. Those plants also had the highest total carotenes and xanthophylls. An interesting observation was the high level of α-carotene found in H. italicum, along with a low content of other carotenoids compared to the other analysed species. High chlorophyll and polyphenol levels coincided in the Lamiaceae family (except S. rosmarinus). Moreover, many of the commonly cultivated herbs studied here, such as Mentha spicata, M. x citrata, and Salvia officinalis, exhibited stronger or more functionally diverse antioxidant traits compared to wild-growing species. The observed differences may be attributed to species-specific metabolic pathways, ecological adaptations, or environmental factors influencing phytochemical expression. Therefore, our experiment underscores the importance of species-level screening in the search for plant-derived antioxidants with potential therapeutic applications. It also demonstrates that comparative biochemical profiling can successfully identify high-antioxidant species regardless of taxonomic affiliation and offers a valuable tool for selecting plants for functional use beyond traditional assumptions.

Author Contributions

Conceptualisation, A.R. and E.M.; methodology, A.R., P.S. and M.G.; formal analysis, A.R., E.M., K.C. and M.D.; data curation, A.R., E.M. and M.G.; writing—original draft preparation, E.M., K.C. and A.R.; visualisation, A.R. and M.D. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Institutional Review Board Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in this article. Further inquiries can be directed to the corresponding authors.

Conflicts of Interest

The authors declare no conflicts of interest.

References

  1. Sies, H.; Belousov, V.V.; Chandel, N.S.; Davies, M.J.; Jones, D.P.; Mann, G.E.; Murphy, M.P.; Yamamoto, M.; Winterbourn, C. Defining Roles of Specific Reactive Oxygen Species (ROS) in Cell Biology and Physiology. Nat. Rev. Mol. Cell Biol. 2022, 23, 499–515. [Google Scholar] [CrossRef] [PubMed]
  2. Pandhair, V.; Sekhon, B.S. Reactive Oxygen Species and Antioxidants in Plants: An Overview. J. Plant Biochem. Biotechnol. 2006, 15, 71–78. [Google Scholar] [CrossRef]
  3. Mehla, N.; Sindhi, V.; Josula, D.; Bisht, P.; Wani, S.H. An Introduction to Antioxidants and Their Roles in Plant Stress Tolerance. In Reactive Oxygen Species and Antioxidant Systems in Plants: Role and Regulation under Abiotic Stress; Khan, M.I.R., Khan, N.A., Eds.; Springer: Singapore, 2017; pp. 1–23. ISBN 978-981-10-5254-5. [Google Scholar]
  4. Soares, C.; Carvalho, M.E.A.; Azevedo, R.A.; Fidalgo, F. Plants Facing Oxidative Challenges—A Little Help from the Antioxidant Networks. Environ. Exp. Bot. 2019, 161, 4–25. [Google Scholar] [CrossRef]
  5. García-Caparrós, P.; De Filippis, L.; Gul, A.; Hasanuzzaman, M.; Ozturk, M.; Altay, V.; Lao, M.T. Oxidative Stress and Antioxidant Metabolism under Adverse Environmental Conditions: A Review. Bot. Rev. 2021, 87, 421–466. [Google Scholar] [CrossRef]
  6. Pathak, J.; Rajneesh; Ahmed, H.; Singh, D.K.; Singh, P.R.; Kumar, D.; Kannaujiya, V.K.; Singh, S.P.; Sinha, R.P. Oxidative Stress and Antioxidant Defense in Plants Exposed to Ultraviolet Radiation. In Reactive Oxygen, Nitrogen and Sulfur Species in Plants; John Wiley & Sons, Ltd.: Hoboken, NJ, USA, 2019; pp. 371–420. ISBN 978-1-119-46867-7. [Google Scholar]
  7. Ben Alaya, I.; Alves, G.; Lopes, J.; Silva, L.R. Use of Encapsulated Polyphenolic Compounds in Health Promotion and Disease Prevention: Challenges and Opportunities. Macromol 2024, 4, 805–842. [Google Scholar] [CrossRef]
  8. Carboni Martins, C.; Rodrigues, R.C.; Domeneghini Mercali, G.; Rodrigues, E. New Insights into Non-Extractable Phenolic Compounds Analysis. Food Res. Int. 2022, 157, 111487. [Google Scholar] [CrossRef] [PubMed]
  9. Charlton, N.C.; Mastyugin, M.; Török, B.; Török, M. Structural Features of Small Molecule Antioxidants and Strategic Modifications to Improve Potential Bioactivity. Molecules 2023, 28, 1057. [Google Scholar] [CrossRef] [PubMed]
  10. Belščak-Cvitanović, A.; Durgo, K.; Huđek, A.; Bačun-Družina, V.; Komes, D. 1—Overview of Polyphenols and Their Properties. In Polyphenols: Properties, Recovery, and Applications; Galanakis, C.M., Ed.; Woodhead Publishing: Cambridge, UK, 2018; pp. 3–44. ISBN 978-0-12-813572-3. [Google Scholar]
  11. Sharma, P.; Jha, A.B.; Dubey, R.S.; Pessarakli, M. Reactive Oxygen Species, Oxidative Damage, and Antioxidative Defense Mechanism in Plants under Stressful Conditions. J. Bot. 2012, 2012, 217037. [Google Scholar] [CrossRef]
  12. Young, A.J.; Lowe, G.L. Carotenoids—Antioxidant Properties. Antioxidants 2018, 7, 28. [Google Scholar] [CrossRef] [PubMed]
  13. Khoo, H.-E.; Prasad, K.N.; Kong, K.-W.; Jiang, Y.; Ismail, A. Carotenoids and Their Isomers: Color Pigments in Fruits and Vegetables. Molecules 2011, 16, 1710–1738. [Google Scholar] [CrossRef] [PubMed]
  14. Munné-Bosch, S.; Alegre, L. The Significance of β-Carotene, α-Tocopherol and the Xanthophyll Cycle in Droughted Melissa Officinalis Plants. Funct. Plant Biol. 2000, 27, 139–146. [Google Scholar] [CrossRef]
  15. Crupi, P.; Faienza, M.F.; Naeem, M.Y.; Corbo, F.; Clodoveo, M.L.; Muraglia, M. Overview of the Potential Beneficial Effects of Carotenoids on Consumer Health and Well-Being. Antioxidants 2023, 12, 1069. [Google Scholar] [CrossRef] [PubMed]
  16. Zheng, C.; Chen, J.-P.; Wang, X.-W.; Li, P. Reactive Oxygen Species in Plants: Metabolism, Signaling, and Oxidative Modifications. Antioxidants 2025, 14, 617. [Google Scholar] [CrossRef] [PubMed]
  17. Hsu, C.-Y.; Chao, P.-Y.; Hu, S.-P.; Yang, C.-M. The Antioxidant and Free Radical Scavenging Activities of Chlorophylls and Pheophytins. Food Nutr. Sci. 2013, 4, 1–8. [Google Scholar] [CrossRef]
  18. Turkiewicz, I.P.; Brzezowska, J.; Tkacz, K.; Wojdyło, A. Polysaccharide- and Protein-Based Carriers as a Key Strategy for Obtaining Microencapsulated Chlorophyll-Rich Extracts: UPLC-PDA/ESI-QToF-MS Characterization and Evaluation of Antidiabetic Potential. Food Chem. 2025, 486, 144627. [Google Scholar] [CrossRef] [PubMed]
  19. Yao, J.; Chen, W.; Fan, K. Recent Advances in Light Irradiation for Improving the Preservation of Fruits and Vegetables: A Review. Food Biosci. 2023, 56, 103206. [Google Scholar] [CrossRef]
  20. Thiviya, P.; Gamage, A.; Piumali, D.; Merah, O.; Madhujith, T. Apiaceae as an Important Source of Antioxidants and Their Applications. Cosmetics 2021, 8, 111. [Google Scholar] [CrossRef]
  21. Ekiert, H.; Klimek-Szczykutowicz, M.; Rzepiela, A.; Klin, P.; Szopa, A. Artemisia Species with High Biological Values as a Potential Source of Medicinal and Cosmetic Raw Materials. Molecules 2022, 27, 6427. [Google Scholar] [CrossRef] [PubMed]
  22. Herrera, E.; Tremblay, N.; Desroches, B.; Gosselin, A. Optimization of Substrate and Nutrient Solution for Organic Cultivation of Medicinal Transplants in Multicell Flats. J. Herbs Spices Med. Plants 1997, 4, 69–82. [Google Scholar] [CrossRef]
  23. Singleton, V.L.; Rossi, J.A. Colorimetry of Total Phenolics with Phosphomolybdic-Phosphotungstic Acid Reagents. Am. J. Enol. Vitic. 1965, 16, 144–158. [Google Scholar] [CrossRef]
  24. Re, R.; Pellegrini, N.; Proteggente, A.; Pannala, A.; Yang, M.; Rice-Evans, C. Antioxidant Activity Applying an Improved ABTS Radical Cation Decolorization Assay. Free Radic. Biol. Med. 1999, 26, 1231–1237. [Google Scholar] [CrossRef] [PubMed]
  25. Beauchamp, C.; Fridovich, I. Superoxide Dismutase: Improved Assays and an Assay Applicable to Acrylamide Gels. Anal. Biochem. 1971, 44, 276–287. [Google Scholar] [CrossRef] [PubMed]
  26. Rusaczonek, A.; Czarnocka, W.; Kacprzak, S.; Witoń, D.; Ślesak, I.; Szechyńska-Hebda, M.; Gawroński, P.; Karpiński, S. Role of Phytochromes A and B in the Regulation of Cell Death and Acclimatory Responses to UV Stress in Arabidopsis Thaliana. J. Exp. Bot. 2015, 66, 6679–6695. [Google Scholar] [CrossRef] [PubMed]
  27. Aebi, H. Catalase in Vitro. Methods Enzymol. 1984, 105, 121–126. [Google Scholar] [CrossRef] [PubMed]
  28. Rusaczonek, A.; Czarnocka, W.; Willems, P.; Sujkowska-Rybkowska, M.; Van Breusegem, F.; Karpiński, S. Phototropin 1 and 2 Influence Photosynthesis, UV-C Induced Photooxidative Stress Responses, and Cell Death. Cells 2021, 10, 200. [Google Scholar] [CrossRef] [PubMed]
  29. Zagoskina, N.V.; Zubova, M.Y.; Nechaeva, T.L.; Kazantseva, V.V.; Goncharuk, E.A.; Katanskaya, V.M.; Baranova, E.N.; Aksenova, M.A. Polyphenols in Plants: Structure, Biosynthesis, Abiotic Stress Regulation, and Practical Applications (Review). Int. J. Mol. Sci. 2023, 24, 13874. [Google Scholar] [CrossRef] [PubMed]
  30. Średnicka-Tober, D.; Ponder, A.; Hallmann, E.; Głowacka, A.; Rozpara, E. The Profile and Content of Polyphenols and Carotenoids in Local and Commercial Sweet Cherry Fruits (Prunus avium L.) and Their Antioxidant Activity In Vitro. Antioxidants 2019, 8, 534. [Google Scholar] [CrossRef] [PubMed]
  31. Pizzino, G.; Irrera, N.; Cucinotta, M.; Pallio, G.; Mannino, F.; Arcoraci, V.; Squadrito, F.; Altavilla, D.; Bitto, A. Oxidative Stress: Harms and Benefits for Human Health. Oxid. Med. Cell. Longev. 2017, 2017, 8416763. [Google Scholar] [CrossRef] [PubMed]
  32. Nieto, G.; Ros, G.; Castillo, J. Antioxidant and Antimicrobial Properties of Rosemary (Rosmarinus officinalis, L.): A Review. Medicines 2018, 5, 98. [Google Scholar] [CrossRef] [PubMed]
  33. El-Nakhel, C.; Pannico, A.; Graziani, G.; Giordano, M.; Kyriacou, M.C.; Ritieni, A.; De Pascale, S.; Rouphael, Y. Mineral and Antioxidant Attributes of Petroselinum Crispum at Different Stages of Ontogeny: Microgreens vs. Baby Greens. Agronomy 2021, 11, 857. [Google Scholar] [CrossRef]
  34. Joshi, T.; Deepa, P.R.; Sharma, P.K. Effect of Different Proportions of Phenolics on Antioxidant Potential: Pointers for Bioactive Synergy/Antagonism in Foods and Nutraceuticals. Proc. Natl. Acad. Sci. India Sect. B Biol. Sci. 2022, 92, 939–946. [Google Scholar] [CrossRef] [PubMed]
  35. Lv, Q.; Long, J.; Gong, Z.; Nong, K.; Liang, X.; Qin, T.; Huang, W.; Yang, L. Current State of Knowledge on the Antioxidant Effects and Mechanisms of Action of Polyphenolic Compounds. Nat. Prod. Commun. 2021, 16, 1934578X211027745. [Google Scholar] [CrossRef]
  36. Lang, Y.; Gao, N.; Zang, Z.; Meng, X.; Lin, Y.; Yang, S.; Yang, Y.; Jin, Z.; Li, B. Classification and Antioxidant Assays of Polyphenols: A Review. J. Future Foods 2024, 4, 193–204. [Google Scholar] [CrossRef]
  37. Bors, W.; Michel, C. Chemistry of the Antioxidant Effect of Polyphenols. Ann. N. Y. Acad. Sci. 2002, 957, 57–69. [Google Scholar] [CrossRef] [PubMed]
  38. Zaheer, M.S.; Rizwan, M.; Aijaz, N.; Hameed, A.; Ikram, K.; Ali, H.H.; Niaz, Y.; Usman Aslam, H.M.; Manoharadas, S.; Riaz, M.W.; et al. Investigating the Synergistic Effects of Biochar, Trans-Zeatin Riboside, and Azospirillum Brasilense on Soil Improvement and Enzymatic Activity in Water-Stressed Wheat. BMC Plant Biol. 2024, 24, 314. [Google Scholar] [CrossRef] [PubMed]
  39. Meitha, K.; Pramesti, Y.; Suhandono, S. Reactive Oxygen Species and Antioxidants in Postharvest Vegetables and Fruits. Int. J. Food Sci. 2020, 2020, 8817778. [Google Scholar] [CrossRef] [PubMed]
  40. Mohammadi, M.A.; Cheng, Y.; Aslam, M.; Jakada, B.H.; Wai, M.H.; Ye, K.; He, X.; Luo, T.; Ye, L.; Dong, C.; et al. ROS and Oxidative Response Systems in Plants Under Biotic and Abiotic Stresses: Revisiting the Crucial Role of Phosphite Triggered Plants Defense Response. Front. Microbiol. 2021, 12, 631318. [Google Scholar] [CrossRef] [PubMed]
  41. Ferguson, L.R. Role of Plant Polyphenols in Genomic Stability. Mutat. Res. Mol. Mech. Mutagen. 2001, 475, 89–111. [Google Scholar] [CrossRef] [PubMed]
  42. Rajput, V.D.; Harish; Singh, R.K.; Verma, K.K.; Sharma, L.; Quiroz-Figueroa, F.R.; Meena, M.; Gour, V.S.; Minkina, T.; Sushkova, S.; et al. Recent Developments in Enzymatic Antioxidant Defence Mechanism in Plants with Special Reference to Abiotic Stress. Biology 2021, 10, 267. [Google Scholar] [CrossRef] [PubMed]
  43. Mehdizadeh, L.; Moghaddam, M.; Ganjeali, A.; Rahimmalek, M. Phenolic Compounds, Enzymatic and Non-Enzymatic Antioxidant Activities of Mentha piperita Modified by Zinc and Methyl Jasmonate Concentrations. Sci. Hortic. 2024, 329, 112980. [Google Scholar] [CrossRef]
  44. Fujita, M.; Hasanuzzaman, M. Approaches to Enhancing Antioxidant Defense in Plants. Antioxidants 2022, 11, 925. [Google Scholar] [CrossRef] [PubMed]
  45. Stahl, W.; Sies, H. Antioxidant Activity of Carotenoids. Mol. Asp. Med. 2003, 24, 345–351. [Google Scholar] [CrossRef] [PubMed]
  46. Landrum, J.T.; Bone, R.A. Lutein, Zeaxanthin, and the Macular Pigment. Arch. Biochem. Biophys. 2001, 385, 28–40. [Google Scholar] [CrossRef] [PubMed]
  47. Maslova, T.G.; Markovskaya, E.F.; Slemnev, N.N. Functions of Carotenoids in Leaves of Higher Plants (Review). Biol. Bull. Rev. 2021, 11, 476–487. [Google Scholar] [CrossRef]
  48. Larsen, E.; Christensen, L.P. Simple Saponification Method for the Quantitative Determination of Carotenoids in Green Vegetables. J. Agric. Food Chem. 2005, 53, 6598–6602. [Google Scholar] [CrossRef] [PubMed]
  49. Žnidarčič, D.; Ban, D.; Šircelj, H. Carotenoid and Chlorophyll Composition of Commonly Consumed Leafy Vegetables in Mediterranean Countries. Food Chem. 2011, 129, 1164–1168. [Google Scholar] [CrossRef] [PubMed]
  50. Xu, C.-J.; Fraser, P.D.; Wang, W.-J.; Bramley, P.M. Differences in the Carotenoid Content of Ordinary Citrus and Lycopene-Accumulating Mutants. J. Agric. Food Chem. 2006, 54, 5474–5481. [Google Scholar] [CrossRef] [PubMed]
  51. Lanfer-Marquez, U.M.; Barros, R.M.C.; Sinnecker, P. Antioxidant Activity of Chlorophylls and Their Derivatives. Food Res. Int. 2005, 38, 885–891. [Google Scholar] [CrossRef]
  52. Mehdipoor Damiri, G.R.; Motamedzadegan, A.; Safari, R.; Shahidi, S.A.; Ghorbani, A. Evaluation of Stability, Physicochemical and Antioxidant Properties of Extracted Chlorophyll from Persian Clover (Trifolium resupinatum L.). J. Food Meas. Charact. 2021, 15, 327–340. [Google Scholar] [CrossRef]
  53. Martins, T.; Barros, A.N.; Rosa, E.; Antunes, L. Enhancing Health Benefits through Chlorophylls and Chlorophyll-Rich Agro-Food: A Comprehensive Review. Molecules 2023, 28, 5344. [Google Scholar] [CrossRef] [PubMed]
  54. Sarker, U.; Oba, S. Nutrients, Minerals, Pigments, Phytochemicals, and Radical Scavenging Activity in Amaranthus Blitum Leafy Vegetables. Sci. Rep. 2020, 10, 3868. [Google Scholar] [CrossRef] [PubMed]
  55. Buhăianu, S.; Jităreanu, D. The Relationship between Chlorophyll Content and Antioxidant Activity of Abies Alba and Nepeta Pannonica Extracts According to Phenophase and Harvesting Area. Cercet. Agron. Mold. 2019, 52, 158–165. [Google Scholar] [CrossRef]
  56. Mishra, N.; Jiang, C.; Chen, L.; Paul, A.; Chatterjee, A.; Shen, G. Achieving Abiotic Stress Tolerance in Plants through Antioxidative Defense Mechanisms. Front. Plant Sci. 2023, 14, 1110622. [Google Scholar] [CrossRef] [PubMed]
  57. Hosseini, R.; Heidari, M. Impact of Drought Stress on Biochemical and Molecular Responses in Lavender (Lavandula angustifolia Mill.): Effects on Essential Oil Composition and Antibacterial Activity. Front. Plant Sci. 2025, 16, 1506660. [Google Scholar] [CrossRef] [PubMed]
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Figure 1. (A) Total carotenoid, (B) xanthophyll, and (C) carotenoid content in µg of fresh weight (FW). Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Figure 1. (A) Total carotenoid, (B) xanthophyll, and (C) carotenoid content in µg of fresh weight (FW). Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Figure 2. (A) Xanthophylls and (B) carotenes in the total pool of carotenoids. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Figure 2. (A) Xanthophylls and (B) carotenes in the total pool of carotenoids. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Agriculture 15 01586 g003
Figure 3. (A) Lutein content in µg of fresh weight (FW) and (B) lutein in the total pool of xanthophylls. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Figure 3. (A) Lutein content in µg of fresh weight (FW) and (B) lutein in the total pool of xanthophylls. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Figure 4. Content of (A) α- and (B) β-carotenes and (C) β-carotene in the total pool of carotenoids. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Figure 4. Content of (A) α- and (B) β-carotenes and (C) β-carotene in the total pool of carotenoids. Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Figure 5. (A) Total chlorophyll, (B) chlorophyll a, and (C) b content in µg of fresh weight (FW). Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Figure 5. (A) Total chlorophyll, (B) chlorophyll a, and (C) b content in µg of fresh weight (FW). Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Figure 6. The heatmap shows scaled values of the measured traits, normalised from 0 (minimum) to 100 (maximum) within each trait. The colour intensity reflects the relative magnitude of each parameter.
Figure 6. The heatmap shows scaled values of the measured traits, normalised from 0 (minimum) to 100 (maximum) within each trait. The colour intensity reflects the relative magnitude of each parameter.
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Table 1. Plant species used in research with their families.
Table 1. Plant species used in research with their families.
Latin NameCommon NameFamily
1Anethum graveolens L.dillApiaceae
2Levisticum officinale W.D.J. Kochlovage
3Petroselinum Hortense (Mill.) A. W. Hillparsley
4Achillea millefolium L.yarrowAsteraceae
5Artemisia dracunculus L.tarragon
6Helichrysum italicum (Roth) G. DonItalian strawflower
7Lamium album L.white dead-nettleLamiaceae
8Mentha x citrata Ehrh.bergamot-mint
9Mentha spicata var. crispa L.curly mint
10Mentha arvensis L.wild mint or corn mint
11Ocimum basilicum L.great basil
12Origanum vulgare L.oregano
13Salvia rosmarinus Spenn.rosemary
14Salvia officinalis L.common sage
15Thymus vulgaris L.common thyme
Table 2. Total polyphenol content expressed as the gallic acid equivalent (GAE) and ABTS radical scavenging capacity described as the Trolox equivalent (TE) in selected herbs representing diversified families.
Table 2. Total polyphenol content expressed as the gallic acid equivalent (GAE) and ABTS radical scavenging capacity described as the Trolox equivalent (TE) in selected herbs representing diversified families.
FamilySpeciesPolyphenols
[mg GAE g−1 FW]		Antioxidant Capacity
[µmol TE g−1 FW]
ApiaceaeAnethum graveolens0.35 ± 0.018 b *44 ± 3.3 e
Levisticum officinale0.67 ± 0.058 e14 ± 1.1 b
Petroselinum hortense0.64 ± 0.030 e70 ± 4.7 g
AsteraceaeAchillea millefolium0.41 ± 0.014 bc5 ± 0.5 a
Artemisia dracunculus0.7 ± 0.039 e25 ± 6.0 c
Helichrysum italicum0.99 ± 0.019 i57 ± 4.2 f
LamiaceaeLamium album0.41 ± 0.026 c29 ± 4.9 cd
Mentha x citrata0.77 ± 0.021 f32 ± 2.5 d
Mentha spicata0.84 ± 0.067 g66 ± 6.4 g
Mentha arvensis0.79 ± 0.024 fg41 ± 1.6 e
Ocimum basilicum0.24 ± 0.015 a15 ± 2.0 b
Origanum vulgare0.8 ± 0.023 fg29 ± 3.5 cd
Salvia rosmarinus0.55 ± 0.032 d78 ± 9.1 h
Salvia officinalis0.92 ± 0.048 h30 ± 2.7 cd
Thymus vulgaris0.78 ± 0.052 fg44 ± 3.1 e
* Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
Table 3. The activity of antioxidant enzymes—catalase (CAT) and superoxide dismutase (SOD) normalised to protein content in the tested species belonging to different families.
Table 3. The activity of antioxidant enzymes—catalase (CAT) and superoxide dismutase (SOD) normalised to protein content in the tested species belonging to different families.
FamilySpeciesCAT
[µmol H2O2 min−1 mg−1 Protein]		SOD
[U mg−1 Protein]
ApiaceaeAnethum graveolens35 ± 5.6 fg *54 ± 6.6 b
Levisticum officinale59 ± 6.2 h96 ± 7.7 gh
Petroselinum hortense26 ± 3.7 cd38 ± 4.2 a
AsteraceaeAchillea millefolium27 ± 4.3 cde132 ± 4.2 i
Artemisia dracunculus42 ± 5.9 g107 ± 3.0 h
Helichrysum italicum36 ± 5.0 fg81 ± 3.9 ef
LamiaceaeLamium album9 ± 0.9 a58 ± 8.2 bc
Mentha x citrata55 ± 4.0 h67 ± 3.7 cd
Mentha spicata42 ± 6.7 g53 ± 4.7 b
Mentha arvensis24 ± 4.2 bcd54 ± 8.4 b
Ocimum basilicum8 ± 1.5 a127 ± 17.5 i
Origanum vulgare21 ± 3.1 bc80 ± 4.5 ef
Salvia rosmarinus17 ± 2.9 b88 ± 10.2 fg
Salvia officinalis31 ± 4.1 def103 ± 4.1 h
Thymus vulgaris34 ± 5.0 ef74 ± 4.2 de
* Different letters indicate means ± SDs that are significantly different according to one-way ANOVA and the post hoc Tukey honest significant difference (HSD) test at p < 0.05.
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Rusaczonek, A.; Sankiewicz, P.; Duszyn, M.; Górecka, M.; Chwedorzewska, K.; Muszyńska, E. Interspecific Variation in the Antioxidant Potential of Culinary and Medicinal Herbs. Agriculture 2025, 15, 1586. https://doi.org/10.3390/agriculture15151586

AMA Style

Rusaczonek A, Sankiewicz P, Duszyn M, Górecka M, Chwedorzewska K, Muszyńska E. Interspecific Variation in the Antioxidant Potential of Culinary and Medicinal Herbs. Agriculture. 2025; 15(15):1586. https://doi.org/10.3390/agriculture15151586

Chicago/Turabian Style

Rusaczonek, Anna, Patryk Sankiewicz, Maria Duszyn, Mirosława Górecka, Katarzyna Chwedorzewska, and Ewa Muszyńska. 2025. "Interspecific Variation in the Antioxidant Potential of Culinary and Medicinal Herbs" Agriculture 15, no. 15: 1586. https://doi.org/10.3390/agriculture15151586

APA Style

Rusaczonek, A., Sankiewicz, P., Duszyn, M., Górecka, M., Chwedorzewska, K., & Muszyńska, E. (2025). Interspecific Variation in the Antioxidant Potential of Culinary and Medicinal Herbs. Agriculture, 15(15), 1586. https://doi.org/10.3390/agriculture15151586

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