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Article

Antioxidative and Photoprotective In Vitro Potential of Lavandula Angustifolium

by
Magdalena Stelmach
1,
Adam Klimowicz
2,*,
Agnieszka Wróblewska
3,*,
Daria Oshetkova
4,
Sylwia Gajewska
3 and
Joanna Siemak
3
1
Independent Researcher, PL-70236 Szczecin, Poland
2
Department of Cosmetic and Pharmaceutical Chemistry, Pomeranian Medical University in Szczecin, Powstańców Wlkp. 72, PL-70111 Szczecin, Poland
3
Faculty of Chemical Technology and Engineering, Department of Catalytic and Sorbent Materials Engineering, West Pomeranian University of Technology in Szczecin, Piastów Ave. 42, PL-71065 Szczecin, Poland
4
Student’s Scientific Club at the Department of Cosmetic and Pharmaceutical Chemistry, Pomeranian Medical University in Szczecin, Powstańców Wlkp. 72, PL-70111 Szczecin, Poland
*
Authors to whom correspondence should be addressed.
Appl. Sci. 2025, 15(11), 6004; https://doi.org/10.3390/app15116004
Submission received: 28 April 2025 / Revised: 22 May 2025 / Accepted: 24 May 2025 / Published: 27 May 2025
(This article belongs to the Special Issue Extraction of Functional Ingredients and Their Application)
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Versions Notes

Abstract

:
People are paying more and more attention to their physical appearance. One way is the use of cosmetics containing antioxidants that slow down the skin ageing process. The application of photoprotective agents is another factor that protects the skin against ageing. Preparations based on natural raw materials are considered to be more safe. The evaluation of both antioxidant and photoprotective potential seems to be of interest for formulating new cosmetics. The aim of this study was to evaluate the antioxidant and in vitro photoprotective potential of Lavandula angustifolia alcoholic extracts. Two methods, i.e., DPPH and ABTS, based on spectrophotometric analysis were applied to determine antioxidant activity. Additionally, the in vitro sun protection factor (SPF) of these extracts was determined and a correlation between this parameter and the antioxidant potential of the extracts was also evaluated. The extracts of dry flowers and herbs of lavender were prepared using ultrasound-assisted extraction. As extractants, four short-chain alcohols, i.e., methanol, ethanol, n-propanol, and isopropanol, in three concentrations were applied to obtain the extracts. To evaluate the stability of the extracts, the determination of antioxidant activity by the DPPH and ABTS methods as well as the SPF value in vitro were performed twice: immediately after the preparation of the extracts and twelve months later. Moreover, the GC-MS analysis of certain extracts was also performed. In extracts made in diluted alcohols, a higher antioxidant potential was observed. A similar observation was made for the in vitro SPF determination. A significant correlation was seen between the antioxidant activity determined by the ABTS method and SPF (for herbs analysed immediately after extract preparation and twelve months later, r = 0.713 and 0.936, respectively, and for flower extracts, r = 0.640 and 0.801, respectively). For the DPPH method, a significant correlation was found only for herb extracts (r = 0.520 and 0.623, respectively). In general, slightly higher antioxidant or photoprotective in vitro potential were observed in later-analysed extracts. However, no significant differences were noted between the antioxidant activity or the photoprotection factor of the extracts determined immediately after their preparation and twelve months later, except for the flower extracts evaluated using the DPPH method (p < 0.0001). A very high correlation was found between the SPF values for both herb and flower extracts evaluated immediately and twelve months later, r = 0.953 and 0.899, respectively. Based on the obtained results, the extracts of Lavandula angustifolia Hidcote Blue variety could be considered as a possible component of anti-ageing cosmetics.

1. Introduction

In contemporary medicine and cosmetology, there is an increasing recognition of the interconnection between skin condition and mental well-being [1]. Skin condition may, among other things, influence self-esteem and social life. In this context, maintaining a nice skin appearance may also contribute to improved psychosocial well-being [2,3]. Therefore, it is important to take a holistic approach and verify the various factors that can adversely affect the human body. The contemporary human condition, particularly with regard to exposure to environmental stress in the form of various types of industrial and municipal pollution, has been demonstrated to promote changes in the metabolic processes of the body, thereby intensifying the formation of excessive amounts of reactive oxygen species [4,5].
Oxidative stress can result in the damage of important macromolecules present in our body, thereby contributing to the development of various diseases, including neurodegenerative diseases (Alzheimer’s, Parkinson’s), cardiovascular diseases, and cancers [6,7]. This raises interest in the role of externally supplied antioxidants, e.g., provided as components of our diet, especially those of plant origin (fruits, vegetables and herbs), which are considered an important part of preventive health care [8,9]. The vast majority of plants contain significant amounts of natural antioxidants. Antioxidants, due to their high bioactivity and low toxicity, are currently considered to be compounds necessary for maintaining human health and vitality [10,11].
The key exogenous factors leading to the production of free radicals include UV radiation, environmental pollution, and tobacco smoke. Some researchers suggest that up to 80% of degenerative changes in unprotected skin can be caused by exposure to UV radiation [12,13,14]. Therefore, a fundamental role in protecting the skin from this radiation is played by photoprotective preparations containing physical filters that reflect and scatter UV radiation and chemical filters that absorb the radiation [15]. Plants contain a number of substances that can reflect and absorb harmful UV radiation, so there is an increased interest in their use in anti-ageing formulations [16,17]. In addition, it should be noted that plant-based UV filters contain substances with antioxidant activity, which play an important role in neutralizing harmful free radicals, thus supporting the anti-ageing effect [18,19,20].
In this context, a particularly noteworthy plant is the Lavandula angustifolia ‘Hidcote’. It is one of 39 species from the Lavandula genus. Lavender has been used in traditional medicine for the management of various ailments due to its calming, mood-enhancing, muscle-relaxing, anti-inflammatory, antibacterial, and antioxidant effects [21,22]. Since the early twentieth century, this plant has been cultivated on an industrial scale primarily for the purpose of obtaining its essential oils and for the production of absolutes through solvent extraction methods [23,24]. This plant, beyond its application in herbal medicine, is extensively utilized in food production, in aromatherapy, in the cosmetics industry, and in perfumery [25,26]. Lavender, originally a Mediterranean species, is indigenous to countries such as France, Spain, Andorra, and Italy. However, it is also cultivated in other regions, including Poland. Its adaptability to various climates has contributed to its widespread cultivation beyond its native area of occurrence [27].
Lavandula angustifolia, often referred to as true lavender or English lavender, is the most widely cultivated species that is used for essential oil production [28]. However, not only the essential oil fraction obtained by the hydrodistillation is rich in beneficial compounds for the human body. It has been shown, among other things, that ethanol and water extracts of L. angustifolia are rich in polyphenols and flavonoids and exhibit strong antioxidant activity [23,29,30]. In addition, it has been studied that lavender essential oil shows a UV protection factor of about SPF 6. This information is of significant value, as the application of skincare products enriched with diverse bioactive compounds targeting multiple biological pathways may contribute to the mitigation of the photoageing process. When used in conjunction with consistent sunscreen application, this approach can effectively reduce UV-B-induced oxidative stress mediated by reactive oxygen species [31].
Plant-derived compounds, due to their proven safety and wide range of beneficial properties, hold strong potential for use as accessible and low-cost ingredients in the formulation of sunscreen products and skincare products [16,32]. However, in order to fully utilize the beneficial properties of plants, it is essential to select appropriate extraction methods that ensure the optimal isolation of active constituents. The initial phase in the investigation of medicinal plants involves the extraction process, which holds a pivotal influence on the overall effectiveness and reliability of the study’s outcomes. The most classic methods of extraction include the following methods: Soxhlet extraction, maceration and hydrodistillation. However, these methods have certain limitations, which has led to growing interest in more modern and advanced extraction techniques. Among the most advanced and effective methods are ultrasound-assisted extraction, enzyme-assisted extraction, pulsed electric field-assisted extraction, microwave-assisted extraction, supercritical fluid extraction, and pressurized liquid extraction [33,34]. The first method is based on the generation of ultrasonic waves that enhance the interaction between the plant material and the surrounding solvent [35]. In addition, this method allows the reduction in the time of the extraction process and it is also possible to apply a lower temperature in this process [36]. The use of ultrasound not only increases the extraction yield of the main product compared to other methods but also significantly speeds up the manufacturing process [37]. This method is considered environmentally friendly due to its minimal ecological footprint. Key variables to take into account include the choice of solvent, the type of plant material, the duration of extraction, and the temperature conditions applied [38].
The presented results of multicentre studies suggest that lavender is a source of many valuable ingredients that have found practical application. Therefore, it was decided to investigate the antioxidant and in vitro photoprotective potential of alcohol extracts from Lavandula angustifolia.
The aim of the study was to evaluate the in vitro antioxidant and photoprotective potential of Lavandula angustifolia extracts prepared in four short-chain alcohols applied in three concentrations using ultrasound-assisted extraction. To evaluate antioxidant activity, two frequently applied spectrophotometric methods, i.e., DPPH and ABTS, were used. Moreover, the in vitro sun protection factor (SPF) was determined. All of these assays were performed in extracts evaluated immediately after preparation and in the same extracts after twelve months. Additionally, in extracts obtained after a one-hour extraction, the qualitative identification of ingredients was performed.

2. Materials and Methods

2.1. Reagents

ABTS (2,2′-azino-bis(3-ehylbenzothiazline-6-sulphonic acid) diammonium salt, DPPH (2,2-diphenyl-1-picrylhydrazyl), and Trolox (6-Hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid) were purchased from Sigma-Aldrich (St. Louis, MO, USA), and methanol, ethanol, n-propanol, and isopropanol were purchased from Chempur, Piekary Śląskie (Poland). All of the reagents were of analytical grade.

2.2. Raw Material

The plant material consisted of the aerial parts (herb and flower) of Lavandula angustifolia (Hidcote Blue variety), collected in September 2023. Lavandula material was from Szczecin (West Pomeranian Voivodeship; 53°24′29″ N, 14°41′05″ E) (Poland). The raw material was dried in a ventilated place in the dark at ambient temperature. Samples of raw material were marked as LH-01/2024 and LF-01/2024 and were deposited in the Department of Cosmetic and Pharmaceutical Chemistry of the Pomeranian Medical University in Szczecin.

2.3. Preparation of the Extracts

The extracts were made in methyl, ethyl, n-propyl, and isopropyl alcohols according to a previously described methodology [39,40,41]. All the alcohols were used at 40% (v/v), 70% (v/v), and undiluted. To prepare the extracts, 0.5 g of ground raw material was placed in a glass-stoppered tube and 10 mL of one of the solvents was added, followed by ultrasound-assisted extraction at a frequency of 40 kHz (80 W; Sonic 0.5, Polsonic, Poland) for either 15, 30 or 60 min. After extraction, the plant material was separated using filtration. The obtained extracts were stored in tightly closed plastic tubes at room temperature in a dark place pending further analyses which included the determination of antioxidant potential and photoprotection parameters. Assays of all extracts were performed twice: immediately after the preparation of the extracts (marked as 2024) and after twelve months (marked as 2025).

2.4. Determination of Antioxidant Potential of the Lavender Extracts

The antioxidant activity of the extracts was determined by applying two frequently applied techniques. They are based on the reduction of DPPH and ABTS radicals. These methods were described in detail in our previous papers [39,42,43]. Briefly, to determine the antioxidant potential using the DPPH technique, stock DPPH solution in ethanol was prepared as follows: an amount of 0.012 g of DPPH was dissolved in 100 mL of 96% (v/v) ethanol using a magnetic stirrer. A working solution with an absorbance of 1.00 ± 0.02 measured at 517 nm (in 1 cm cuvettes) was prepared from the stock solution by dilution with ethanol. The determination of the antioxidant activity included the addition of 132 µL of the extract to 2500 µL of the working solution of DPPH and thorough mixing. After 10 min of incubation in the dark at ambient temperature, the absorbance of the samples at 517 nm was measured (Hitachi U-5100 spectrophotometer) [41,42]. Three determinations of each extract were performed [20,40]. The antioxidant activity of the extract was presented as Trolox equivalent antioxidant capacity (TEAC), i.e., in mg Trolox/g raw material (RM) [41]. To calculate the results, a calibration curve of absorbance vs. Trolox concentration (y = −0.9704x + 0.9093, R2 = 0.9915) was applied.
Evaluation of the antioxidant activity with the ABTS method included the addition of 20 µL of the extract to 2500 µL of 7mM ABTS solution and thorough mixing. The samples were incubated for 6 min at room temperature. Measurements of absorbance were conducted at 734 nm. The 7 mM ABTS solution was prepared by dissolving 0.038 g of ABTS in 10 mL of 2.45 mM aqueous potassium persulphate solution. After 24 h, a period necessary to form the ABTS cation radical, the obtained solution was diluted with 50% (v/v) methanol in such a way that the absorbance of the ABTS solution in 1 cm cuvettes was 1.00 ± 0.02 at 734 nm. Three samples of each extract were prepared. The antioxidant potential of the extracts was expressed as TEAC, i.e., in mg Trolox/g RM [41]. For this purpose, the absorbance vs. Trolox concentration calibration curve was prepared (y = −0.1803x + 1.0185, R2 = 0.9942). The results are presented as arithmetical mean ± standard deviation (SD).
The in vitro sun protection factor (SPF) was determined using the slightly modified method applied in our previous study [41] according to a methodology described by other authors [44,45,46,47]. To determine the absorbance spectrum, 1 cm quartz cuvettes were used. To obtain samples for SPF determination, 70 µL of extract was added to 2030 µL of the solvent applied for extraction and mixed. UV spectra from 290 nm to 320 nm were measured using Hitachi U-5100 (Hitachi High-Tech Corporation, Tokyo, Japan) and DR6000 Hach (Hach Lange GmbH, Düsseldorf, Germany) spectrophotometers. SPF was calculated using the Mansur equation presented below:
S P F = C F · ∑ 290 320 E E ( λ )   ·   I ( λ )   ·   A ( λ )
where CF—correction factor (=10); EE(λ)—erythmogenic effect of radiation with wavelength λ; and A(λ)—absorbance at wavelength λ. The absorbance was determined at 290–320 nm every 5 nm. The values EE(λ) · I(λ) are constants and were determined by Sayre et al. [46]. Table 1 presents the values used in this study to evaluate the SPF value of the studied extracts.

2.5. GC-MS Analyses of Selected Extracts

Antioxidant capacity studies of the extracts obtained showed that the most active extracts were the alcoholic extracts obtained after the 60 min extraction. For the qualitative identification of the ingredients (organic compounds) in those 24 extracts obtained from the flowers and herbs of Lavandula angustifolia in solvents such as ethanol, methanol, n-propanol, and isopropanol in three concentrations, the GC-MS method was applied.
The extracts were analysed using a GC/MS system, consisting of a Nexis GC-2030 gas chromatograph coupled with a GCMS-QP2020 NX mass spectrometer. The chromatographic column which was used for the studies was column SH-I-5MS (30 mm × 0.25 mm × 0.25 mm). Extract samples of 1 µL were applied to the chromatographic column. The injector temperature was 250 °C, and the carrier gas (helium 6.0) was used with a flow rate of 0.94 mL/min. The temperature program included an initial isothermal step at 40 °C for 2 min, followed by a temperature increase of 10 °C/min to 300 °C, and the final isothermal step was established as 300 °C for 2 min. The interface temperature between the GC and MS was set to 250 °C, with the ion source temperature at 200 °C. The mass spectrometer operated in the mass range of 35–500 u. Data analysis was performed using LabSolutions software version 4.53SP1 with NIST20 libraries.

2.6. Statistical Analysis of the Results

The results of the determinations are presented as arithmetical means ± standard deviations (SD). Most groups of results differed significantly from the normal distribution as checked by the Lilliefors test. Therefore, the Kruskal–Wallis and Mann–Whitney, i.e., non-parametric, tests were used to estimate the differences between the evaluated groups of results. Additionally, linear regression parameters and correlation coefficients between the studied groups were also calculated. For this purpose, the ProStat v. 5.5 and Excel programs were used.

3. Results

Figure 1 and Figure 2 present the antioxidant activity of Lavandula angustifolia herb and flower extracts in four short-chain alcohols determined with the DPPH method immediately after preparation and after twelve months of storage at room temperature. As concerns the herbal extracts, the lowest activity was observed for extracts prepared in undiluted alcohols except for those in methanol. Significant differences were found between flower extracts assayed immediately and after twelve months (p < 0.0001) and between herb and flower extracts assayed twelve months after preparation (p = 0.0001) (Table 2).
Figure 3 and Figure 4 present the antioxidant activity of Lavandula angustifolia herb and flower extracts in four short-chain alcohols determined with the ABTS method immediately after preparation and after twelve months of storage at room temperature. The higher activities were found for extracts prepared in shorter-chain alcohols. Significant differences were found between herb and flower extracts assayed after twelve months (p = 0.0009) (Table 2).
Figure 5 and Figure 6 present the in vitro sun protection factor (SPF) of Lavandula angustifolia herb and flower extracts in four short-chain alcohols determined immediately after preparation and after twelve months of storage at room temperature. Higher values were found for extracts prepared in shorter-chain alcohols, particularly for herbal extracts. The lowest values of this parameter were found for undiluted alcohols. Significant differences were found between herb and flower extracts assayed both immediately after preparation (p = 0.0028) and after twelve months (p = 0.0038) (Table 2).
In Figure 7, correlations between the evaluated antioxidant activity of Lavandula angustifolia extracts determined using DPPH and ABTS methods after the preparation of extracts and twelve months later are presented. Statistically significant correlations were found for the activity of both herb and flower extracts determined either by DPPH or ABTS methods. A significant linear relationship between the antioxidant activity of Lavandula angustifolia alcoholic extracts of herbs and flowers was observed both after the application of the DPPH and ABTS methods (p < 0.001)
Figure 8 presents correlations between the in vitro SPF value of the studied extracts determined after the preparation of Lavandula angustifolia alcoholic extracts and twelve months later. A highly significant linear relationship between the SPF value of Lavandula angustifolia alcoholic extracts of herb and flower was observed (correlation coefficient r = 0.9526 and r = 0.8989, respectively, p < 0.0001).
Figure 9 presents the correlation between the in vitro SPF values and antioxidant activity determined using the DPPH and ABTS methods. A significant linear relationship was found between antioxidant activity determined with the ABTS method both for herb and flower extracts, whereas this was found only for the herb extracts for the DPPH method.

GC-MS Analysis of the Obtained Alcoholic Extracts by the Ultrasound-Assisted Extraction

The antioxidant capacity studies of the obtained extracts showed that the most active were the alcoholic extracts obtained during the 60 min extraction. For the qualitative identification of ingredients (organic compounds) in 24 extracts prepared from the flowers and herbs of Lavandula angustifolia in solvents such as ethanol, methanol, n-propanol, and isopropanol, the GC-MS method was used.
Table 3 presents the list of area percentages (area %) of the organic compounds identified by the GC-MS method in the alcoholic extracts for which antioxidant activity has been described in scientific literature. These compounds include: linalool (LIN), 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-(4-HEX), α-terpineol (α-TER), geraniol (GER), coumarin (COU), 2H-1-benzopyran-2-one, 7-methoxy-(2H-1), linalyl acetate (LIA), 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-, acetate (4-HEA), γ-sitosterol (ɣ-SIT), trans-β-ocimene (t-β-OCI), 4H-pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl-(4H-PYR), 3,7-octadiene-2,6-diol, 2,6-dimethyl-(3,7-OCT), D-allose (D-ALL), τ-cadinol (τ-CAD), 2-methoxy-4-vinylphenol (2-MET), and phytol (PHY). The structural formulas of these compounds are presented in Figure 10 and Figure 11.

4. Discussion

Nowadays, people are paying more and more attention to their physical appearance. It affects not only their appearance but also their well-being. Therefore, opportunities are being sought to ensure a nice, youthful appearance [3,48]. One way is to use cosmetics containing antioxidants that slow down the skin ageing process, which affects a favourable appearance [49,50]. Since one of the factors responsible for the formation of free radicals is ultraviolet radiation, it seems beneficial to combine photoprotective properties with antioxidant potential. Formulations with such properties would be helpful for reducing the risk of various skin diseases caused by the adverse effects of UV radiation, including skin cancer [51]. People are looking for agents based on natural raw materials, which are considered to be more safe. Hence, a great interest in cosmetics containing natural ingredients is observed [52]. One of the raw materials of interest is Lavandula angustifolia, particularly its Hidcote Blue variety. Studies on this plant have been carried out by many researchers, using different extraction techniques classified as both classical [53,54] and modern, more ecological (green) techniques [22,55]. Few studies have included a comparison of different solvents used for the preparation of extracts, so we decided to investigate the in vitro antioxidant and photoprotective activity of extracts from the herbs and flowers of this plant prepared in four low-molecular-weight alcohols, i.e., methanol, ethanol, n-propanol, and isopropanol, using an ultrasound-assisted extraction method categorised as a so-called green extraction technique [56,57,58]. To prepare the extracts, 40% (v/v), 70% (v/v), and undiluted alcohols were used. These parameters were also determined in the same extracts stored at room temperature away from light after 12 months. Additionally, the composition of the selected extracts was also examined by the GC-MS method to identify the compounds for which antioxidant activity has been previously described in scientific reports.
In the presented study, the highest antioxidant potential assayed by the DPPH method and expressed in mg Trolox/g RM in extracts of the Lavandula angustifolia herb determined immediately after their preparation was found in extracts prepared in 40% and 70% isopropanol after a 30 min extraction (3.57 ± 0.01 and 3.62 ± 0.04 TEAC, respectively) and in 40% isopropanol and 96% ethanol after a 60 min UAE (3.62 ± 0.03 and 3.57 ± 0.01 TEAC, respectively).
For flower extracts evaluated after their preparation, the highest antioxidant potential was found for extracts prepared in concentrated methanol and 70% ethanol following a 15 min UAE (3.78 ± 0.05 and 3.71 ± 0.09 TEAC, respectively) and in undiluted ethanol and undiluted isopropanol after a 30 min extraction (3.74 ± 0.18 and 3.73 ± 0.09 TEAC, respectively).
The antioxidant potential of Lavandula angustifolia was also evaluated by others. Rather et al. evaluated groups of phytochemicals from the leaf extracts of Lavandula angustifolia [59]. Moreover, the antioxidant potential of methanolic extracts was also evaluated by the DPPH method. For this purpose, 100 g of plant powder was added to 1 L of methanol and mixed by continuous agitation for 24 h. The prepared methanolic extract at the concentration of 250 µg/mL scavenged 63.5% radicals in a DPPH assay [59]. However, the observed differences can be related to the use of a slightly different procedure as well as a different source of raw material.
Blažeković et al. determined the antioxidant potential of Lavandula x intermedia ‘Budrovka’ and Lavandula angustifolia [43]. To compare these two plants, several methods frequently used to evaluate antioxidant potential, including the DPPH assay, were applied. The extracts were obtained by ultrasound-assisted extraction using 30 g of powdered plant material in 500 mL of 80% ethanol. The IC50 values for DPPH scavenging activity were 11.4 ± 0.7 and 10.62 ± 0.02 µg/mL for flower and leaf extracts, respectively [55].
Dobros et al. evaluated the extracts obtained from the inflorescences of different cultivars of Lavandula angustifolia [22]. The authors used three different techniques to obtain the extracts, including ultrasound-assisted extraction. Plant material consisted, among others, of the Lavandula angustifolia Hidcote variety. To obtain the extracts, 20 mL of 50% ethanol was added to 1.0 g of raw material and subjected to sonication for 30 min, with 10 mL added in the second and third stages, lasting 15 min. The antioxidant activity determined using the DPPH method was 156.8 µM Trolox/g d.w., found for extracts obtained using ultrasound-assisted extraction. This result was higher as compared with the extracts obtained using maceration or decoction [22].
Betley et al. estimated the antioxidant potential of flower and leaf extracts of Lavandula angustifolia cultivated in southern Poland [48]. The extracts were prepared by shaking 10 g of lavender (either flower or leaf) in 60% ethanol for 72 h. The mean DPPH radical scavenging activity was 97.5 µmol Trolox/g for flower extracts and 198.2 µmol Trolox/g for leaf extracts [60].
In the presented study, the same extracts were also evaluated twelve months later. They showed slightly higher activity determined by the DPPH method. For herb extracts, the highest activity was found for extracts in 70% methanol after 30 and 60 min extractions and in 70% ethanol after a 60 min extraction (4.11 ± 0.01, 4.05 ± 0.09 and 4.20 ± 0.03 TEAC). The differences between the antioxidant activities of lavender herb extracts determined at an interval of 12 months were statistically insignificant. This result proved that the prepared extracts were, in general, stable during the observation period. In the case of flower extracts evaluated after twelve months, the highest antioxidant potential was observed for extracts prepared in undiluted isopropanol after 15 and 30 min of ultrasound-assisted extraction (4.48 ± 0.09 and 4.51 ± 0.04 TEAC, respectively). The differences between the antioxidant activities of lavender flower extracts determined at an interval of 12 months were statistically significant (p < 0.001, Table 2). Moreover, the differences in the antioxidant activity in herb and flower extracts determined immediately after their preparation were statistically insignificant, but for determinations carried out twelve months later, the differences were statistically significant (Table 2).
The second method applied in our study to evaluate the antioxidant potential of the obtained extracts was the ABTS technique. In the presented study, the highest antioxidant potential assayed by the ABTS method (expressed similarly to the DPPH method as TEAC in mg Trolox/g of raw material) in extracts of the Lavandula angustifolia herb determined immediately after their preparation was found in extracts prepared in 40% n-propanol and in 70% ethanol after a 15 min extraction and in 40% ethanol after a 60 min extraction (24.6 ± 0.8, 23.6 ± 1.4 and 23.2 ± 0.6 TEAC, respectively).
For flower extracts, after their preparation, evaluated by the ABTS method, the highest antioxidant activity was observed for extracts prepared after a 60 min extraction in 70% methanol, 70% ethanol, and 40% n-propanol (26.9 ± 2.7, 31.8 ± 3.6, and 35.4 ± 2.01 TEAC, respectively). No significant differences were found between antioxidant activity in herb and flower extracts evaluated immediately after their preparation (Table 2).
The same extracts evaluated twelve months later showed slightly higher activity determined by the ABTS method, similar to the DPPH method. For herb extracts, the highest activity was found for extracts in 40% methanol and in 40% ethanol after a 60 min extraction (23.8 ± 0.6 and 23.3 ± 0.2 TEAC, respectively). The differences between the antioxidant activities of lavender herb extracts determined at an interval of 12 months were statistically insignificant. In the case of flower extracts evaluated after twelve months, the highest antioxidant capacity was found for extracts in 40% n-propanol and 40% isopropanol after a 60 min ultrasound-assisted extraction (30.8 ± 6.0 and 24.3 ± 0.4 TEAC, respectively). The differences between the antioxidant activities of lavender flower extracts determined at an interval of 12 months were statistically insignificant (Table 2). Moreover, the differences in antioxidant activity in herb and flower extracts determined immediately after their preparation were statistically insignificant, but for determinations carried out twelve months later, the differences were statistically significant (p = 0.0009, Table 2).
Marovska et al. evaluated the antioxidant potential of Lavandula angustifolia extracts in 70% ethanol [29]. The studied material included extracts of dried untreated lavender and three extracts of lavender by-products. To prepare the extracts, 300 g of raw material was heated with 2 L 70% (v/v) ethanol at 60–65 °C for 1 h and macerated overnight at room temperature. After filtration, 1000 mL of the same solvent was added to the remaining raw material and the procedure was repeated. Both filtrates were combined. The antioxidant activity determined using the ABTS method was 125.0 ± 18.9 mM TE (Trolox equivalent)/g and was lower than the activities of by-product extracts. Similar observations can be made for the DPPH method (the activity of the extract of untreated lavender was 185.0 ± 6.7 mM TE/g) [29].
Dvorackova et al. evaluated the antioxidant potential and phenolic content in 17 plants, including spices [60]. Antioxidants were extracted using different techniques. The extracts were prepared using 2 g of raw material and 20 mL of water. The highest antioxidant activity determined by the ABTS method was observed for the extracts obtained by Randall extraction, while the lowest was obtained after sonication (20.19 and 4.54 mg/100 g DW) [60].
Betley et al. determined the antioxidant potential of the flower and leaf extracts of Lavandula angustifolia cultivated in southern Poland also using the ABTS method. As previously mentioned, the extracts were prepared by shaking 10 g lavender (either flowers or leaves) in 60% ethanol for 72 h. The mean antioxidant activity determined using the ABTS method was 168.9 ± 2.9 µmol Trolox/g for lavender flower extracts and 274.8 ± 0.7 µmol Trolox/g for leaf extracts [61].
Another parameter evaluated in the presented study was the in vitro sun protection factor (SPF). This parameter is important for the initial evaluation of the photoprotective ability of the studied extracts. In most cases, higher SPF values were observed for the herb extracts evaluated twelve months after their preparation as compared to the extracts evaluated immediately after preparation. Similar results were obtained for flower extracts. It should be noted that no statistically significant differences were observed between herb extracts assayed immediately after their preparation and after twelve months later. Similar observations can be made for flower extracts of Lavandula angustifolium. However, it should be noted that statistically significant differences were found between the SPF of herb and flower extracts evaluated both immediately and twelve months after their preparation (Table 2). Moreover, a very high correlation was observed between the SPF values obtained immediately and after twelve months, both for herb and flower extracts (r = 0.9526 and 0.8989, respectively). Studies on the in vitro determination of the SPF in plant extracts are rarely conducted. The results of the assessment of this parameter in lavender extracts obtained in our study are probably the first determinations of this parameter, taking into account the number of prepared and analysed extracts.
Studies on the qualitative evaluation of Lavandula angustifolia extracts have been performed by others. However, some of these studies have been focused on the analysis of essential oils isolated from this plant [30,62,63,64]. The studies presented in this paper included GC-MS analysis of 24 extracts of both lavender flowers and herbs obtained in four solvents with three different concentrations. Based on the results obtained during the studies by the GC-MS method, it can be said that compounds present in the highest area percentages in alcoholic extracts of Lavandula angustifolia include linalool, coumarin, 2H-1-benzopyran-2-one, 7-methoxy-, and linalyl acetate. The highest content of individual organic compounds mentioned above was observed in the following samples: flower 40% (v/v) nProOH–LIN 28.76 Area%, flower 40% (v/v) isoProOH–4-HEX 7.84 Area%, flower 40% (v/v) nProOH–α-TER 6.48 Area%, flower 40% (v/v) MeOH–GER 2.22 Area%, herb 70% (v/v) EtOH–COU 16.25 Area%, herb 40% (v/v) EtOH–2H-1 30.43 Area%, flower 96% (v/v) EtOH–LIA 32.42 Area%, flower 99% (v/v) MeOH–4-HEA 10.05 Area%, herb 40% (v/v) isoProOH–ɣ-SIT 4.72 Area%, flower 99% (v/v) isoProOH–t-β-OCI 2.41 Area%, herb 40% (v/v) MeOH–4H-PYR 2.46 Area%, flower 99% (v/v) isoProOH–3,7-OCT 2.76 Area%, herb 99% (v/v) MeOH–D-ALL 2.88 Area%, herb 99% (v/v) nProOH–τ-CAD 3.69 Area%, herb 40% (v/v) EtOH–2-MET 2.72 Area%, and herb 40% (v/v) nProOH–PHY 3.64 Area%.
Moreover, a detailed analysis of Table 3 shows that linalool is the main component only in extracts obtained from Lavandula angustifolia flowers. The lowest content of this compound (17.85%) was determined for the flower extract obtained using 99% (v/v) n-ProOH, and the highest (28.76%) was for the same solvent but at a concentration of 40% (v/v). It can be further assumed that in the extracts obtained from flowers using MeOH, EtOH, n-ProOH, and isoProOH, linalool plays a key role as an antioxidant compound. In addition, Table 3 shows that its concentration in extracts decreases with increasing concentrations of the solvent used. The linalool molecule contains a hydroxyl group (Figure 10), which is the main structural element responsible for the antioxidant activity of this compound, although this antioxidant activity is weaker than in phenols (e.g., flavonoids), due to the lack of the aromatic system and the lower stability of the radical after hydrogen donation. The hydroxyl group (-OH) allows hydrogen donation (H•) in reaction with free radicals, which allows their neutralization. Thanks to this, linalool can act as a hydrogen donor and interrupt radical chain reactions. The free radical neutralization reactions involving linalool can be represented as follows:
ROO• + R−OH→ROOH + R−O•
The resulting alkoxy radical (R-O•) from linalool is relatively stable, which stops the oxidation reaction [65].
The extracts mentioned above also contain significant amounts of linalyl acetate (from 9 to 32%). In the case of this ester compound, unlike linalool, the content of linalyl acetate increases with the concentration of the solvent used for extraction. It can be assumed that linalyl acetate acts here in synergy with linalool, which has an -OH group and can directly scavenge radicals. Both compounds can jointly stabilize lipid free radicals and protect lipids from oxidative damage, thus affecting the stabilization of cell membranes. A similar synergy of these two compounds has been observed earlier in essential oils [66].
The extracts obtained from flowers of Lavandula angustifolia using MeOH, EtOH, n-ProOH, and isoProOH also contain coumarin and 2H-1-benzopyran-2-one,7-methoxy- (7-methoxycoumarin). The content of the first compound is a maximum of 4.9%, and the second compound is 5.9%. These compounds also show antioxidant activity, previously described in the scientific literature—with the methoxy derivative of coumarin being greater. Studies have shown that 7-methoxycoumarin has the ability to neutralize free radicals and has a hepatoprotective effect [67]. However, in the studies presented in this work, attention is drawn to the very high content of these two compounds in the extracts obtained from the herb of Lavandula angustifolia using MeOH, EtOH, n-ProOH, and isoProOH and at the same time the very low content of linalool and linalyl acetate in them. It can therefore be assumed that coumarin and its methoxy derivative are responsible for the antioxidant activity of extracts from the herb of Lavandula angustifolia obtained in this work. It is also worth noting that for these extracts, the content of these two compounds decreases with the increase in the concentration of the solvent used for extraction. It is therefore beneficial to use the solvents tested as extractants while maintaining their lowest concentration of 40% (v/v).

5. Conclusions

Taking into account the results of our research, it has been established that the investigated alcoholic extracts of Lavandula angustifolia Hidcote Blue variety have antioxidant and photoprotective potential. Better results were obtained if diluted alcohols were applied as solvents to obtain both the herb and the flower extracts using UAE. GC-MS analyses of Lavandula angustifolia extracts showed the presence of 16 compounds with antioxidant activity. However, taking into account the content of these compounds in the extracts, the compounds mainly responsible for the antioxidant activity of the extracts from flowers should be assumed to be linalool and linalyl acetate, while the compounds responsible for the antioxidant activity of the extracts from herbs should be assumed to be 7-methoxycoumarin and coumarin.
A satisfactory correlation was observed between the antioxidant activity determined immediately after the preparation of extracts and twelve months later, which may indicate the stability of the prepared extracts. Similar results were found for the evaluation of the in vitro sun protection factor, which could confirm the above-mentioned conclusion. The obtained results could suggest the possible use of Lavandula angustifolia extracts in the development of cosmetics. In the next stage, further studies of antioxidant activity using additional methods for the evaluation of the antioxidant potential of the lavender extracts, as well as their application to formulate cosmetic preparations, should be performed. Moreover, additional evaluation is required taking into account, among others, toxicological, antimicrobial, and allergy tests to assess the safety of such products.

Author Contributions

Conceptualization, M.S., A.K. and A.W.; methodology, M.S., A.K., A.W., D.O. and S.G.; software, A.K. and A.W.; validation A.K. and A.W.; formal analysis, M.S., A.K., D.O. and S.G.; investigation, M.S., A.K., D.O., S.G. and J.S.; resources, M.S. and A.K.; data curation, A.K. and A.W.; writing—original draft preparation, A.K., A.W. and J.S.; writing—review and editing, A.K. and A.W.; visualization, M.S., A.K., A.W. and S.G.; supervision, A.K. and A.W.; project administration, A.K.; funding acquisition, A.K. 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.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available upon request from the corresponding author.

Conflicts of Interest

The authors declare no conflict of interest.

Abbreviations

The following abbreviations are used in this manuscript:
ABTS(2,2′-azino-bis(3-ehylbenzothiazline-6-sulphonic acid) diammonium salt
DPPH2,2-diphenyl-1-picrylhydrazyl
SDStandard deviation
SPFSun protection factor
TEACTrolox equivalent antioxidant capacity

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Applsci 15 06004 g001
Figure 1. Mean antioxidant activity of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation using the DPPH method. The results are presented as TEAC (mg Trolox/g RW). Vertical lines denote standard deviations (SD). No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
Figure 1. Mean antioxidant activity of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation using the DPPH method. The results are presented as TEAC (mg Trolox/g RW). Vertical lines denote standard deviations (SD). No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
Applsci 15 06004 g001
Applsci 15 06004 g002
Figure 2. Mean antioxidant activity of Lavandula angustifolia flower alcoholic extracts evaluated immediately and twelve months after preparation using the DPPH method. The results are presented in mg Trolox/g raw material (TEAC). Vertical lines denote standard deviations (SD). The statistically significant differences were found between flower extracts assayed immediately and after twelve months (p < 0.0001).
Figure 2. Mean antioxidant activity of Lavandula angustifolia flower alcoholic extracts evaluated immediately and twelve months after preparation using the DPPH method. The results are presented in mg Trolox/g raw material (TEAC). Vertical lines denote standard deviations (SD). The statistically significant differences were found between flower extracts assayed immediately and after twelve months (p < 0.0001).
Applsci 15 06004 g002
Applsci 15 06004 g003
Figure 3. Mean antioxidant potential of Lavandula angustifolia herb alcoholic extracts (in mg Trolox/g raw material (TEAC)) evaluated immediately and twelve months after preparation using the ABTS method. Vertical lines correspond to standard deviations (SD). No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
Figure 3. Mean antioxidant potential of Lavandula angustifolia herb alcoholic extracts (in mg Trolox/g raw material (TEAC)) evaluated immediately and twelve months after preparation using the ABTS method. Vertical lines correspond to standard deviations (SD). No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
Applsci 15 06004 g003
Applsci 15 06004 g004
Figure 4. Mean antioxidant potential of Lavandula angustifolia flower alcoholic extracts (in mg Trolox/g raw material (TEAC)) evaluated immediately and twelve months after preparation using the ABTS method. Vertical lines correspond to standard deviations (SD). No statistically significant differences were found between flower extracts assayed immediately and after twelve months.
Figure 4. Mean antioxidant potential of Lavandula angustifolia flower alcoholic extracts (in mg Trolox/g raw material (TEAC)) evaluated immediately and twelve months after preparation using the ABTS method. Vertical lines correspond to standard deviations (SD). No statistically significant differences were found between flower extracts assayed immediately and after twelve months.
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Applsci 15 06004 g005
Figure 5. In vitro sun protection factor of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation. No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
Figure 5. In vitro sun protection factor of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation. No statistically significant differences were found between herb extracts assayed immediately and after twelve months.
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Applsci 15 06004 g006
Figure 6. In vitro sun protection factor of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation. No statistically significant differences were found between flower extracts assayed immediately and after twelve months.
Figure 6. In vitro sun protection factor of Lavandula angustifolia herb alcoholic extracts evaluated immediately and twelve months after preparation. No statistically significant differences were found between flower extracts assayed immediately and after twelve months.
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Applsci 15 06004 g007
Figure 7. Correlation between the evaluated antioxidant activity of Lavandula angustifolia alcoholic extracts determined using DPPH and ABTS methods after preparation of extracts and twelve months later r—correlation coefficient, p—probability.
Figure 7. Correlation between the evaluated antioxidant activity of Lavandula angustifolia alcoholic extracts determined using DPPH and ABTS methods after preparation of extracts and twelve months later r—correlation coefficient, p—probability.
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Applsci 15 06004 g008
Figure 8. Correlations between the in vitro SPF values of the Lavandula angustifolia herb and flower extracts evaluated after their preparation and twelve months later r—correlation coefficient, p—probability.
Figure 8. Correlations between the in vitro SPF values of the Lavandula angustifolia herb and flower extracts evaluated after their preparation and twelve months later r—correlation coefficient, p—probability.
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Applsci 15 06004 g009
Figure 9. Correlations between in vitro SPF values and antioxidant activity of the Lavandula angustifolia herb and flower extracts evaluated after their preparation and twelve months later r—correlation coefficient, p—probability.
Figure 9. Correlations between in vitro SPF values and antioxidant activity of the Lavandula angustifolia herb and flower extracts evaluated after their preparation and twelve months later r—correlation coefficient, p—probability.
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Applsci 15 06004 g010
Figure 10. The structural formulas of compounds present in studied extracts for which antioxidant activity has been described in the scientific literature: linalool, linalyl acetate, α-terpineol, geraniol, coumarin, 2H-1-benzopyran-2-one, 7-methoxy-, 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-, 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-, acetate.
Figure 10. The structural formulas of compounds present in studied extracts for which antioxidant activity has been described in the scientific literature: linalool, linalyl acetate, α-terpineol, geraniol, coumarin, 2H-1-benzopyran-2-one, 7-methoxy-, 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-, 4-hexen-1-ol, 5-methyl-2-(1-methylethenyl)-, acetate.
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Applsci 15 06004 g011
Figure 11. The structural formulas of compounds present in the studied extracts for which antioxidant activity has been described in scientific literature: γ-sitosterol, trans-β-ocimene, 4H-pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl-, 3,7-octadiene-2,6-diol, 2,6-dimethyl- -D-allose, τ-cadinol, 2-methoxy-4-vinylphenol, phytol.
Figure 11. The structural formulas of compounds present in the studied extracts for which antioxidant activity has been described in scientific literature: γ-sitosterol, trans-β-ocimene, 4H-pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl-, 3,7-octadiene-2,6-diol, 2,6-dimethyl- -D-allose, τ-cadinol, 2-methoxy-4-vinylphenol, phytol.
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Table 1. Values of EE(λ) · I(λ) [45,46].
Table 1. Values of EE(λ) · I(λ) [45,46].
Wavelength [nm]EE(λ) · I(λ)
2900.0150
2950.0817
3000.2874
3050.3278
3100.1864
3150.0839
3200.0180
Table 2. Statistical analysis of the examined parameters of Lavandula angustifolia extracts.
Table 2. Statistical analysis of the examined parameters of Lavandula angustifolia extracts.
Compared ParametersZ-Valuep
Antioxidant activity evaluated with DPPH method
herb assayed in 2024herb assayed in 2025−1.79090.0733 (NS)
flower assayed in 2024flower assayed in 2025−5.7612<0.00001
herb assayed in 2024flower assayed in 2024−1.35170.1765 (NS)
herb assayed in 2025flower assayed in 2025−3.86930.0001
Antioxidant activity evaluated with ABTS method
herb assayed in 2024herb assayed in 2025−1.67240.0944 (NS)
flower assayed in 2024flower assayed in 2025−0.00560.9955 (NS)
herb assayed in 2024flower assayed in 2024−1.42470.1542 (NS)
herb assayed in 2025flower assayed in 2025−3.32300.0009
Sun protection factor
herb assayed in 2024herb assayed in 2025−1.07550.2821 (NS)
flower assayed in 2024flower assayed in 2025−0.01690.9865 (NS)
herb assayed in 2024flower assayed in 2024−2.99010.0028
herb assayed in 2025flower assayed in 2025−2.89350.0038
NS—not statistically significant.
Table 3. The area percentage of chemical compounds identified by the GC-MS method in the obtained extracts from the flowers and herbs of Lavandula angustifolia after 60 min of extraction. Only the compounds with known antioxidant properties based on the data described in the scientific literature are presented.
Table 3. The area percentage of chemical compounds identified by the GC-MS method in the obtained extracts from the flowers and herbs of Lavandula angustifolia after 60 min of extraction. Only the compounds with known antioxidant properties based on the data described in the scientific literature are presented.
EXTRACTS of Lavandula
angustifolia (60 min)		AREA [%]
LIN4-HEXα-TERGERCOU2H-1LIA4-HEAɣ-SITt-β-OCI4H-PYR3,7-OCTD-ALLτ-CAD2-METPHY
flower 40% (v/v) MeOH25.325.963.522.224.95.9----1.311.17-0.74--
flower 70% (v/v) MeOH28.61.53.25-3.974.8897.6---0.99-1.76--
flower 99% (v/v) MeOH20.880.66--2.533.331.6510.05-1.19-1.32-1.34-0.57
flower 40% (v/v) EtOH24.863.193.94-3.244.5413.295.65---0.86-1.21--
flower 70% (v/v) EtOH24.475.192.38-3.895.118.293.63---1.92-1.77--
flower 96% (v/v) EtOH19.50.49--1.090.9232.428.481.182.38-0.62-1.24--
flower 40% (v/v) n-ProOH28.767.746.48-3.464.12--2.76--1.27-1.43--
flower 70% (v/v) n-ProOH20.691.790.64-2.864.0226.956.64-1.31-0.86-1.1--
flower 99% (v/v) n-ProOH17.85---1.021.430.528.68-2.19-0.91-1.21--
flower 40% (v/v) isoProOH25.547.844.44-4.294.8913.230.91---0.85-1.4--
flower 70% (v/v) isoProOH20.595.280.92-3.113.8726.943.63---1.68-1.4-0.68
flower 99% (v/v) isoProOH19.130.5----32.178.99-2.41-2.76-1.28--
herb 40% (v/v) MeOH----15.1529.72----2.46---1.83-
herb 70% (v/v) MeOH0.61---13.7428.79----0.75-2.422.851.721.25
herb 99% (v/v) MeOH----10.7517.040.84-----2.883.442.21.4
herb 40% (v/v) EtOH0.67---16.2530.43--1.92-1.62--1.692.72-
herb 70% (v/v) EtOH0.57---13.1928.220.45---0.48-1.783.411.662.32
herb 96% (v/v) EtOH----9.4316.380.95-1.75---2.072.761.552.35
herb 40% (v/v) n-ProOH0.54---10.220.330.34------2.9313.64
herb 70% (v/v) n-ProOH----8.3918.220.48-1.58---1.252.871.222.11
herb 99% (v/v) n-ProOH0.75---4.447.280.95-1.66----3.690.661.82
herb 40% (v/v) isoProOH----5.7411.12--4.72---1.422.220.831.79
herb 70% (v/v) isoProOH0.78---5.488.130.74-1.86----2.73-1.4
herb 99% (v/v) isoProOH1.06---1.462.972.11-2.16----3.62-0.89
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Stelmach, M.; Klimowicz, A.; Wróblewska, A.; Oshetkova, D.; Gajewska, S.; Siemak, J. Antioxidative and Photoprotective In Vitro Potential of Lavandula Angustifolium. Appl. Sci. 2025, 15, 6004. https://doi.org/10.3390/app15116004

AMA Style

Stelmach M, Klimowicz A, Wróblewska A, Oshetkova D, Gajewska S, Siemak J. Antioxidative and Photoprotective In Vitro Potential of Lavandula Angustifolium. Applied Sciences. 2025; 15(11):6004. https://doi.org/10.3390/app15116004

Chicago/Turabian Style

Stelmach, Magdalena, Adam Klimowicz, Agnieszka Wróblewska, Daria Oshetkova, Sylwia Gajewska, and Joanna Siemak. 2025. "Antioxidative and Photoprotective In Vitro Potential of Lavandula Angustifolium" Applied Sciences 15, no. 11: 6004. https://doi.org/10.3390/app15116004

APA Style

Stelmach, M., Klimowicz, A., Wróblewska, A., Oshetkova, D., Gajewska, S., & Siemak, J. (2025). Antioxidative and Photoprotective In Vitro Potential of Lavandula Angustifolium. Applied Sciences, 15(11), 6004. https://doi.org/10.3390/app15116004

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