Chemical composition and biological activities of essential oils from Alyssoides utriculata (L.) Medik

Ucuncu, O. (2024). "Chemical composition and biological activities of essential oils from Alyssoides utriculata (L.) Medik," BioResources 19(4), 8797–8811.

Abstract

The chemical compositions, antioxidant activities, and antimicrobial activities of the essential oils acquired from the separated parts of air-dried flowers, leaves, and stems of Alyssoides utriculata L. plant growing in Turkiye were determined. Three volatile oil components were acquired via hydrodistillation using a Clevenger apparatus and analyzed by the Gas Chromatography-Mass spectrometry/Flame Ionization Detection (GC-MS/FID) analysis. A total of 75, 67, and 76 compounds in the volatile oils of flower, leaves, and stem of A. utriculata were identified, respectively. The highest percentage of chemical compounds in the essential oils of A. utriculata were determined to be monoterpenes in flowers and leaves, (72.4% and 66.5%) and hydrocarbons (29.2%) in stems. While α-pinene (62.5% and 46.7%) was defined as the major compound in the flowers and leaves, nonane (21.2%) was determined to be so in the stem essential oil. The antioxidant activity of the obtained essential oils was determined according to free radical scavenging and total phenolic content (TPC), and antimicrobial activity against 12 bacteria and 5 fungi, using the agar dilution method. The amount of TPC and scavenging activity of the flower oil were found to be 440.61 ± 6.26 mg GAE/L and 46.00 ± 1.28%, respectively. Based on the antimicrobial activity results, all the essential oils of A. utriculata were determined to have antimicrobial activity against Escherichia coli and Bacillus subtilis.

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Chemical Composition and Biological Activities of Essential Oils from Alyssoides utriculata (L.) Medik

Osman Üçüncü *

DOI: 10.15376/biores.19.4.8797-8811

Keywords: Essential oil; Chemical composition; Antioxidant activity; Antimicrobial activity; α-Pinene; Alyssoides utriculata

Contact information: Department of Pharmacy Services, Macka Vocational School, Karadeniz Technical University, Trabzon, 61750 Turkiye; *Corresponding author: osmanucuncu@yahoo.com

INTRODUCTION

Since the beginning of humanity, human beings have been using plants to meet their basic needs such as nutrition, treatment, and warmth. In complementary medicine for treatment purposes, whole plants, including leaves, roots, and flowers, are used, as well as various extracts and essential oils obtained from them (Al Abboud et al. 2024; Alghonaim et al. 2023). The Brassicaceae (also known as Cruciferae) family has economic, agricultural, nutritional, and medicinal qualities (Preedy 2015). Foods, such as cabbage, broccoli, Bok-choy, and mustard, which have an important place in daily nutrition, are from the Brassicaceae family and contain glucosinolates, minerals, carotenoids, soluble sugars, polyphenols, vitamins, and antioxidant compounds (Preedy 2015; Luo et al. 2022). Members of Brassicaceae are widely used in traditional medicine and as veterinary medicines for livestock (Salehi et al. 2021).

The popularity and consumption of vegetable Brassicaceae family members are increasing due to their nutritional value and biological effects. Their phytochemical composition has been studied, as they contain valuable secondary metabolites, such as glucosinolates, phenolic compounds (hydroxycinnamic acids, flavonoids, anthocyanins, tocopherols, and carotenoids), terpenes, and fatty acids, which are directly linked to different biological activities (Favela-González et al. 2020). Brassicaceae metabolites are used in the treatment of chronic diseases such as obesity, type-2 diabetes, stroke, hypertension, and cancer (Favela-González et al. 2020). Previous studies have reported that the essential oils and extracts of Brassicaceae species are rich in glucosinolates and that they have biological activities such as anticancer, anti-inflammatory, antimicrobial, anti-obesity, cardioprotective, gastroprotective, and antioxidant activities (Favela-González et al. 2020; Salehi et al. 2021). They are also known to contain high amounts of carotenoids, tocopherol, and ascorbic acid, which have antioxidant effects (Singh et al. 2017).

The genus Alyssoides Mill., a member of the Brassicaceae family, is represented by two species (The Plant List 2013). Alyssoides is morphologically similar to the genus Physoptychis Boiss. according to Flora of Turkey (Cullen 1965) and distinguished from Physoptychis with less than 10 mm fruit diameter. According to phylogenetic-based studies, the genus Alyssoides is not monophyletic, and the members of the genus are grouped with members of Fibigia Medik. Alyssoides utriculata (L.) Medik is the only species of the genus found in Turkiye, and it is a yellow-flowered ornamental shrubby plant native to the country (Cullen 1965). This species is both an ornamental plant and used in some forms of treatment (rabies and hiccup) (Blazevic et al. 2013). Alyssoides utriculata var. utriculata is the only member of Alyssoides utriculata at the variety level in the Flora of Turkey (Cullen 1965; Mutlu 2012).

Essential oils are complex mixtures of low concentrations derived from different parts of plants and evaporate easily at room temperature (Fidan et al. 2022). The essential oils exhibit refreshing, pain-relieving, stress-relieving, insecticidal, antimicrobial, antifungal, and antioxidant activities and are used in the food preservation and cosmetic industries (Polatoğlu et al. 2013; Yılar et al. 2016; Cüce and Basançelebi 2021; Saruhan and Oz 2023). It has been reported that that the essential oils of Brassicaceae family members contain interesting natural phytochemicals such as allyl isothiocyanate (B. juncea, B, nigra), 1-butene-4-isothiocyanate (B. juncea, B. napus), benzyl isothiocyanate and 2-phenylethyl-isothiocyanate (Sinapis alba) as sulfur-containing compounds, hexahydrofarnesyl acetone (Arabis alpina, Eruca vesicaria), pulegone, isomenthone (B. campestris), phytol (Capsella bursa-pastoris), and β-elemene as terpene derivatives, and 2,6,10-trimethyldecane, nonacosane (Arabis alpina, Capsella bursa-pastoris) as hydrocarbons (Singh et al. 2015; Hichri et al. 2016; Salehi et al. 2021; Ucuncu 2021; Gumusok et al. 2023).

There is only one study in the literature on the essential oils and biological activities of A. utriculata. Blazevic et al. (2013) investigated the chemical composition of the essential oil obtained from A. utriculata and the acetyl cholinesterase activities of dichloromethane extracts. In the gas chromatography-mass spectrometry (GC/MS) analysis of essential oils of different parts of A. utriculata, chemical compounds belonging to the compound classes alcohols, carbonyls, alkanes, sulfur compounds, terpenes, fatty acids and esters, phenols, and phenylpropane derivatives were detected. According to this report, compounds such as but-3-enyl isothiocyanate, erucin, and sulforaphane, which are glucosinolates degradation products, are responsible for the acetylcholinesterase activity exhibited by the essential oil and extracts (Blazevic et al. 2013).

The chemical compositions and biological activities of A. urticulata with respect to individual parts of the plant, which may be important towards potential use, have not been explored. The goal of the present research is to determine the chemical compositions of essential oils in the air-dried parts (flower, leaf, and stem) of A. utriculata, which can be considered as a member of the Brassicaceae family, and has been subject to limited studies, and to investigate their antimicrobial and antioxidant capacities.

EXPERIMENTAL

Plant Materials

Alyssoides utriculata plant was collected from the roadsides between Torul and Kürtün, Gümüşhane: (40° 38′ 30″ N, 39° 11′ 39″ E at 800 m above sea level) in Turkiye (A7), a location with dry air and sandy soil, during June 2022. Flowers, leaves, and stems of A. utriculata were separated and air-dried at room temperature (20 to 22°C). The botanical identification of the plant was carried out by Prof. Kamil Coşkunçelebi in the Department of Biology, at Karadeniz Technical University (KTU), Trabzon, Turkiye. Voucher specimens were deposited with the number KTUB743 in the Herbarium of KTU.

Separation and Analysis of the Essential Oils

The volatile oils from air-dried plant parts (flower – 85 g, leaves – 54 g, and stems – 124 g) of A. utriculata were isolated using a modified Clevenger-type hydrodistillation apparatus (4 h, yields: 0.24%, 0.19%, and 0.08 % (w/w), respectively) (Ucuncu et al. 2019). Hydrodistillation for each sample was carried out three times, and the average value of the essential oil percentage (w/w) was used for the final evaluation and was detected on an air-dried weight basis. Essential oil yields were calculated with the following Eq. 1 (Fidan et al. 2022):

(1)

The essential oils obtained from the air-dried plants were taken by dissolving in 1.0 mL high-performance liquid chromatography (HPLC) grade n-hexane, dried over anhydrous sodium sulfate, and filtered (Ucuncu et al. 2019; Fidan et al. 2022).

A HP-5MS capillary chromatographic apolar column (film thickness 0.2 μm 30 m × 0.25 mm ID) was used for GC-FID (Agilent-7890A) and GC-MS (Agilent 5975C) analyses. These analyses were employed as described previously (Ucuncu et al. 2019; Fidan et al. 2022; Oz 2022).

The essential oils were analyzed twice. The GC peak areas of essential oil compounds were clarified by comparing the NIST and Willey libraries in the GC-MS device. The retention indices (RI) of components were determined through the retention times (RT) of homolog n-alkanes (C₆-C₃₂) and authentic compounds with linear interpolation. Identification of volatile compounds was determined by matching their RI values with NIST and Willey library data, comparing Kovats indices (KI) and literature value. Quantitative determination of components was performed with regard to peak area integration with GC-FID (Adams 2007; Ucuncu et al. 2019; Fidan et al. 2022; Oz 2022; Chemdata NIST 2023).

DPPH Assay and the Total Phenolic Content

The antioxidant activities of essential oils were determined by free radical scavenging capacity (DPPH assay), the most frequently used antioxidant/antiradical test, and by total phenolic contents (TPC) analysis, which shows not only total phenolic content but also total reducing capacity of the sample and is widely accepted as an antioxidant test (Sanchez-Moreno et al. 1998; Kasangana et al. 2015).

The free radical scavenging activities of volatile oils against stable 2,2-diphenyl-2-picrylhydrazyl hydrate (DPPH·) were spectrophotometrically determined. For this purpose, a DPPH solution prepared with 4 mL of 0.1 mM methanol was added to the volatile oils of A. utriculata. The change in color was measured at 517 nm on a UV-Vis spectrophotometer (Libra S60, Biochrom Ltd, Cambridge UK). The measurements were performed three times, and averaged. Trolox and ascorbic acid were used as standard antioxidants for comparison (Sağdıç et al. 2011; Ahmed et al. 2015).

The TPC amounts of volatile oils were determined by the Folin-Ciocalteu method. For this purpose, the absorbances of the samples were measured at 765 nm. TPC in essential oils were expressed as gallic acid equivalents (GAE) according to the method described previously (Ucuncu et al. 2019). The measurements were performed three times, and averaged (Kasangana et al. 2015).

Antimicrobial Activity Assessment

The antimicrobial activities of essential oils were determined using the agar-well diffusion method against 12 bacteria and 5 yeast samples. The antimicrobial tests were made in Gümüşhane University Food Engineering Laboratories. For this purpose, the samples of volatile oils were dissolved in HPLC-grade n-hexane to prepare stock solutions. Measurements were made according to previously described methods (Sağdıç and Özcan 2003; Matuschek et al. 2014). The results were expressed as inhibition zones (mm) of test microorganisms. The results of antimicrobial activity are given in Table 3.

Statistical Analysis

Statistical analyses were performed using Microsoft Excel software with XLSTAT (Addinsoft, Version 2024 New York, NY, USA). The consistency of measurements across the analysis was assessed using the relative standard deviation of repeatability (RSDr%) and the predicted relative standard deviation (PRSDr%).

RESULTS AND DISCUSSION

Chemical Composition of Essential Oils

The results of GC-MS and GC-FID analyses performed to determine the chemical compositions of essential oils of A. utriculata are given in Table 1, and their chemical class distributions are presented in Fig. 1. About 119 compounds were identified, constituting over 92.77%, 87.66%, and 86.26% total essential oil compositions of flowers, leaves, and stems of A. utriculata, respectively. The identified compounds are divided into 10 groups: alcohols, carbonyl compounds, fatty acids, hydrocarbons, monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons, oxygenated sesquiterpenes, diterpene, and ‘other’.

Table 1. Identified Components and Chemical Class Distribution in the Essential Oils of the Aerial Parts (Flower, Leaf, and Stem) of A. utriculata

^aPercentages obtained by FID peak-area normalization;

^bRetention index calculated from retention times relative to n-alkanes (C₆-C₃₂) on the non-polar HP-5MS column.

^cLiterature retention indices (RI) on HP-5MS column as seen in NIST, Willey, Kovats Index, and Adams (2007).

^dIncluded as authentic compound, NC: Numbers of compounds, and MS: Identification of mass spectrum.

For flower, leaves, and stem essential oils (Table 1), 75, 67, and 76 compounds were determined by GC-MS and GC-FID analyses, respectively Among them, three alcohols, 23 carbonyl compounds, six fatty acids, eight hydrocarbons, 13 monoterpene hydrocarbons, 25 oxygenated monoterpenes, 30 sesquiterpene hydrocarbons, nine oxygenated sesquiterpenes, one diterpene, and one other compound were identified (Fig. 1).

While monoterpene hydrocarbons were the main chemical class of flowers (74.4%), and leaf oils (66.5%), hydrocarbons were the abundant class of stem oils (29.21%). The main compounds were as follows: α-pinene (62.5%), o-cymene (3.4%), and limonene (2.3%) for flower oils; α-pinene (46.5%), o-cymene (6.4%), and limonene (5.5%) for leaf oils; nonane (21.2%), hexadecanoic acid (18.4%), and α-pinene (6.3%) for stem oils.

Approximately 36 compounds were common to all three essential oils. Nonane, α-pinene, o-cymene, limonene, borneol, α-amorphene, β-selinene, α-selinene, spathulenol, and hexahydrofarnesyl acetone were common components with relatively high quantities in all parts of A. utriculata. A study conducted by Blazevic et al. (2013) showed that 31 compounds were detected in the essential oils of the whole plant (A. utriculata), of which hexadecanoic acid (11.5%), nonacosane (10.0%), hexahydrofarnesyl acetone (5.9%), phytol (4.3%), and heptacosane (4.0%) made up the majority (Blazevic et al. 2013). When compared to the work of Blazevich et al. (2013) (E,E)-2,4-decadienal, tricosane, tetracosane, pentacosane, nonacosane, thymol, β-caryophyllene, α-cadinol, hexahydro-farnesyl acetone, tetradecanoic acid, pentadecanoic acid, and hexadecanoic acid were similarly detected in the current study. In another study, Saka et al. (2017) identified 34 and 39 compounds in the essential oils of Brassica rapa var. rapifera leaves and roots (Saka et al. 2017). In this study, methyl-5-hexenenitrile (52.6%), 2-phenylethanol (10.2%), menthol (5.3%), allyl isothiocyanate (4.6%), and hexahydrofarnesyl acetone (3.2%), were abundant compounds in leaf essential oil. When this study is compared with Saka et al. (2017), it is seen that terpene derivative compounds, such as geranyl acetone, hexahydrofarnesyl acetone, α-pinene, β-pinene, camphene, sabinene, α-terpinene, limonone, and α-terpineol, were similar.