Optimization of cell suspension cultures of endemic plant Salvia pseudeuphratica Rech.f. and effects of some AgNP applications on total phenolic, flavonoid, antioxidant activity, and some terpenoid compound levels

The species belonging to the Salvia genus are important medicinal sources containing many therapeutic properties used in traditional medicine. This thesis focuses on the establishment of cell suspension cultures of Salvia pseudeuphratica Rech.f., especially an endemic and endangered species, using biotechnological methods. In this context, the aim is to contribute to the sustainable production of Salvia pseudeuphratica Rech.f. through the evaluation of antioxidant activity, enzyme inhibition effects, and the effects on terpenoid compounds of plant materials obtained via cell suspension cultures. Additionally, the effects of AgNP (silver nanoparticle) applications on the terpenoid content, total phenolic and flavonoid amounts, and antioxidant capacity values of Salvia pseudeuphratica Rech.f. are evaluated. Through the conducted studies within the scope of this thesis, callus and cell suspension cultures of Salvia pseudeuphratica Rech.f., a critically endangered and locally endemic species, have been established for the first time using biotechnological techniques. Furthermore, the effects of some AgNP applications (100, 200, 400, and 800 µg/ml) on the amounts of total phenolics, flavonoids, antioxidant activity, enzyme inhibition, and some terpenoid compounds in cell suspension cultures have been investigated for the first time. The findings indicate significant effects of silver nanoparticles (AgNPs) applied to cell suspension cultures of Salvia pseudeuphratica Rech.f. on antioxidant activities. Particularly, a significant increase in antioxidant activities has been observed with AgNPs applied at concentrations of 400 and 800 µg/ml for 24 and 48 hours. Additionally, antioxidant activity studies conducted with DPPH, ABTS, and CUPRAC methods revealed that the application of 400 µg/ml AgNP for 12 hours in the DPPH method (IC50: 100.00±1.98 µg/mL), 100 µg/ml AgNP for 24 hours in the ABTS method (IC50: 56.82±0.59 µg/mL), and 400 µg/ml AgNP for 24 hours in the CUPRAC method (A0.5: 79.591±0.59) exhibited the highest antioxidant activity. In addition to antioxidant effects, significant effects of AgNPs on enzyme activities in plant cells have been observed. Particularly, a significant increase in BChE enzyme activity has been detected within 24 hours of applying 400 µg/ml AgNP. This demonstrates that AgNP applications regulating the biological activities of Salvia pseudeuphratica Rech.f., especially their effects on BChE, may vary depending on time and concentration. Moreover, the results have determined that the application of 400 µg/ml AgNP for 24 hours yielded good results compared to other parameters in terms of both total phenolic (22.43±0.50 μg PEs/mg extract) and total flavonoid (11.02±0.26 μg QEs/mg extract) contents. Terpenoid analysis results indicate that AgNP applications significantly affect the terpenoid profile in plant cultures and particularly lead to significant increases in oleanolic acid levels. While oleanolic acid was not found in the control group samples without AgNP application, it was observed that all AgNP applications stimulated its synthesis. The highest amount of oleanolic acid produced in suspension cultures was determined to be 3995.80 µg analyte/g extract in the 24-hour 400 µg/ml AgNP application. This demonstrates the potential of AgNPs to influence plant metabolism and increase certain terpenoids. In conclusion, this study comprehensively examines the effects of AgNP applications on antioxidant activities, enzyme activities, and terpenoid profiles in cell suspension cultures of Salvia pseudeuphratica Rech.f. Keywords: AgNP applications, biological activity, cell suspension cultures, terpenoid production, Salvia pseudeuphratica Rech.f.