DEVELOPMENT OF STUDENTS' RESEARCH SKILLS IN THE PROCESS OF DETERMINING THE BIOLOGICAL ACTIVITY OF THYMUS ASIATICUS SERG

ABSTRACT

This study aims to determine the antiradical, antioxidant, and cytotoxic activity of the essential oil and extracts obtained from Thymus asiaticus Serg. using various solvents (hexane, chloroform, ethyl acetate, ethanol). The results showed that the ethanol extract exhibited the highest antiradical (87.70%) and antioxidant activity (1.8995), while the essential oil demonstrated the highest cytotoxic activity (96% mortality). The study results contribute to a deeper understanding of the biological activity of  Th. asiaticus and expand its potential applications in the pharmaceutical field. These studies are based on the laboratory topics of the educational-methodological complex: "Determination of antiradical and antioxidant activity in medicinal plants" and "Study of cytotoxic activity in medicinal plants."

Keywords: Thymus asiaticus Serg., antiradical activity, antioxidant activity, cytotoxic effect, educational process.

Introduction:

Currently, comprehensive research on medicinal plants and their integration into the educational process is one of the pressing issues in education and medicine. Despite the abundance of medicinal plants growing in Kazakhstan, knowledge about their healing properties and applications remains insufficient. The natural compounds extracted from plants are of particular interest due to their biological activity, availability, and minimal side effects. Among such plants, Thymus asiaticus Serg., a representative of the Lamiaceae family, is known for its anti-inflammatory, antimicrobial, and antioxidant properties, widely used in traditional medicine [1].

Figure 1. Th. Asiaticus Serg. Plant

Th. asiaticus is a medicinal plant belonging to the Lamiaceae family. It is widespread in Central Asia, particularly in arid and semi-arid regions. The plant contains essential oils (thymol, carvacrol), flavonoids, and phenolic compounds, which provide antioxidant, antimicrobial, and anti-inflammatory effects. In folk medicine, it is used to treat respiratory and digestive system disorders. Scientific studies confirm its high biological activity [2].

The properties of Th. asiaticus Serg. and its essential oils have been studied by scientists from many countries, including Central Asia, Europe, the USA, China, and the Middle East:

1. Central Asia – Scientists from Kazakhstan, Uzbekistan, and Kyrgyzstan have investigated Th. asiaticus species, identifying their distribution in arid and semi-arid regions and their biological activity. In this region, thyme has been traditionally used in folk medicine for centuries [3].
2. Europe – Researchers from France, Spain, Greece, Bulgaria, and Poland have studied the antioxidant, antimicrobial, and insecticidal properties of thyme essential oils. Particularly, Spanish and Greek scientists have focused on identifying chemotypes based on the plant’s chemical composition [4].
3. USA – American researchers have examined thyme essential oils as food additives and natural preservatives, studying their antibacterial properties. In addition, the immunomodulatory and anti-inflammatory effects of thyme have been extensively explored [4].
4. China – Chinese scientists have studied the effects of thyme on glucose metabolism and its ability to inhibit lactic acid production, suggesting its potential antidiabetic properties [5].
5. Middle East (Iran, Turkey, Saudi Arabia) – Researchers in this region have focused on the antiviral, antifungal, and pharmaceutical applications of thyme essential oils [5].

Thus, thyme is a highly valuable medicinal and technological plant with a broad range of applications [6].

This study aimed to determine the antiradical, antioxidant, and cytotoxic activity of Th. asiaticus Serg. essential oil and extracts. The effectiveness of these compounds was evaluated by comparing their antiradical and antioxidant activity using gallic acid and their cytotoxic effect using brine shrimp lethality tests. The study sought to assess their potential as sources of biologically active compounds. The results contribute to a deeper understanding of the plant's pharmacological potential and its biochemical properties. This research was conducted as part of the educational-methodological complex for laboratory studies on “Determination of antiradical and antioxidant activity in medicinal plants” and “Study of cytotoxic activity in medicinal plants” [7]. Importance in Education

The significance of this study in the field of education is substantial, as teaching the chemical composition, pharmacological properties, and potential applications of medicinal plants helps students develop scientific research skills. The findings can enhance the educational process in chemistry and pharmaceuticals. From a pedagogical perspective, the research can be integrated into the educational process through the following methodological approaches:

Laboratory experiments – Organizing practical lessons where students extract essential oils and plant extracts, analyze their chemical composition, and investigate their biological activity. Project-based learning – Encouraging students to conduct scientific projects on medicinal plant properties and present their findings. Problem-based learning – Assigning students research problems related to medicinal plants, allowing them to conduct independent scientific studies.

Research Methodology

Plant Material: The Th. asiaticus plant was collected in the Akmola region, in the Ormandybulaq area near Karsak village (coordinates: 52.8973201N, 68.8130265E) during the third decade of July 2023. The collected material was dried in the shade at room temperature and then used for further studies. Essential oil (Th.As.0) and extracts with different solvents, including hexane (Th.As.1), chloroform (Th.As.2), ethyl acetate (Th.As.3), and ethanol (Th.As.4), were obtained from the aerial parts of the Th. asiaticus plant. Each sample was tested three times, and experiments were conducted at 20°C under natural light conditions.

Antiradical activity was determined using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical inhibition method. Optical density was measured using a spectrophotometer at a wavelength of 520 nm [8].

Antioxidant activity was evaluated using the ferric-reducing antioxidant power (FRAP) assay. Samples dissolved at specific concentrations were incubated with phosphate buffer, potassium ferricyanide, and FeCl₃ at 50°C. The reaction was terminated with trichloroacetic acid, and the mixture was centrifuged. Antioxidant activity was measured at a wavelength of 700 nm [9].

Cytotoxic activity was assessed by incubating brine shrimp eggs in artificial seawater for three days. A test tube was partially covered with foil to create light and dark regions. The larvae that migrated toward the light were collected using a Pasteur pipette [10].

Research Results

The study focused on evaluating the antiradical, antioxidant, and cytotoxic activity of essential oil and extracts obtained from the aerial parts of Th. asiaticus. The results are presented in Figures 2 and 3, and Table 1.

Figure 2. Dynamics of Antiradical Activity

The study results showed that the ethanol extract (Th.As.4) exhibited the highest antiradical activity, reaching 87.70% at a concentration of 1.0 mg/ml. This value was significantly higher compared to other extracts. Hexane (Th.As.1) and ethyl acetate (Th.As.3) extracts showed moderate antiradical activity, whereas essential oil (Th.As.0) and chloroform extract (Th.As.2) demonstrated the lowest values, indicating weaker antioxidant properties.

The FRAP method is based on the ability of antioxidants to reduce Fe³⁺ ions to Fe²⁺. The process involves the reduction of K₃[Fe(CN)₆] to K₄[Fe(CN)₆], forming a yellow complex. The oxidation inhibition capacity of antioxidants was measured using gallic acid as a standard. The study was conducted at concentrations of 0.25, 0.5, 0.75, and 1 mg/ml.

Figure 3. Effect of Substance Concentration on Antioxidant Activity

According to the analysis, the antioxidant activity of different Th. asiaticus extracts varied depending on concentration. The essential oil (Th.As.0) exhibited low to moderate activity. Hexane (Th.As.1) and ethyl acetate (Th.As.3) extracts showed moderate to higher activity. The chloroform extract (Th.As.2) was generally at a moderate level. The highest antioxidant activity was observed in the ethanol extract (Th.As.4), particularly at a concentration of 1 mg/ml.

Table 1.

 Average values of the experiment based on cytotoxic activity

Based on the results of the conducted experiment, the cytotoxic activity of various extracts and essential oils obtained from the aerial parts of Th. asiaticus plant was investigated. Specifically, the essential oil exhibited pronounced cytotoxic properties at all concentrations (96% mortality), while hexane, chloroform, and ethanol extracts did not show significant cytotoxic activity (4-30% mortality). The ethyl acetate extract showed cytotoxic effects at a concentration of 10 mg/ml (60% mortality), but its activity was lower at lower concentrations.

Conclusion

The conducted study provided a comprehensive evaluation of the biological activity of the essential oil and extracts obtained from the aerial parts of Th. asiaticus plant. The ethanol extract (Th.As.4) exhibited the highest antioxidant activity, with a level of 87.70% at a concentration of 1.0 mg/ml. In terms of antioxidant activity, the ethanol extract showed the highest results, while the essential oil demonstrated a pronounced effect in terms of cytotoxic activity. Other extracts showed moderate activity, and the ethyl acetate extract showed cytotoxic effects at higher concentrations. These results may expand the potential scientific application of the medicinal properties of Th. asiaticus and increase its potential in the production of pharmaceuticals and food supplements. Additionally, the study contributes to the educational process by teaching the chemical and pharmacological properties of medicinal plants. The laboratory work on determining the antioxidant, anti-radical, and cytotoxic activities of medicinal plants, as part of the educational and methodological complex “Determining Anti-Radical and Antioxidant Activities in Medicinal Plants” and “Investigating the Cytotoxic Activity of Medicinal Plants,” was conducted.

References:

1. Aliyeva G.A., Ahmetova A.B. Biologically Active Compounds of Plants of the Genus Thymus // Bulletin of KazNU. Biological Series. – 2020. – №3 (83). – P. 45–53.
2. Ivanov P.V. Antioxidant Properties of Thyme Essential Oils // Pharmaceutical Journal. – 2018. – №2. – P. 112–118.
3. Kuanova Zh.K., Sarsenbayeva G.T. Chemical Composition of Medicinal Plants in the Flora of Kazakhstan // Bulletin of Kazakh National University. – 2019. – No. 1 (71). – P. 37–42.
4. Smith J., Brown K. Chemical Composition and Pharmacological Effects of Thymus Essential Oils // Journal of Medicinal Plants Research. – 2017. – Vol. 11, No. 4. – P. 56–63.
5. Zhang L., Wang Y. Antimicrobial and Antioxidant Activity of Thymus Species // Chinese Journal of Natural Medicines. – 2021. – Vol. 19, No. 2. – P. 87–94.
6. Hosseini M., Farahani A. Antiviral and Antifungal Properties of Thymus Essential Oils // Iranian Journal of Pharmacology. – 2022. – Vol. 24, No. 3. – P. 101–110.
7. Perdekhanova A.A., Iskakova Zh.B. Development of Students' Research Competence in the Study of Medicinal Plants // L.N. Gumilyov Eurasian National University, "GYLYM JANE BILIM – 2025" 20th International Scientific Conference. – 2025, April 11.
8. Tleubayeva, M.I., Abdullabekova, R.M., Datkhayev, U., Serikbayeva, E.A., Flisyuk, E.V. Investigation of CO₂ Extract of Portulaca oleracea for Antioxidant Activity from Raw Material Cultivated in Kazakhstan. International Journal of Biomaterials, 2022, 2022, 6478977.
9. Sisengalieva, G.G., Adilkhanova, A.A., Gorovoi, P.G., Iskakova, Zh.B. Composition and Biological Activity of Essential Oil from Artemisia keiskeana. Chemistry of Natural Compounds, 2019, 55(1), 154–156.
10. Iskakova, Zh.B., Dudkin, R.V., Ross, S.A., Martins, C.H.G. Composition and Biological Activity of Essential Oils from East-Asian Species Angelica viridiflora, A. cincta, and Coelopleurum gmelinii. Chemistry of Natural Compounds, 2014, 50(6), 1136–1139.