Phytochemical Profiling, Antioxidant Activity, and Protective Effect against H2O2-Induced Oxidative Stress of Carlina vulgaris Extract

Abstract

Carlina vulgaris is a little-understood plant with unexplored biological potential, and the papers regarding its chemical composition are scarce. In our study, for the first time, the phytochemical profile of the plant, focusing on polar metabolites, was established using modern chromatographic techniques including LC-HRMS-QTOF-CAD, UHPLC-PDA-MS. Phytochemical analysis revealed that the species is a rich source of polyphenolic components, with the most abundant being chlorogenic acid and C-glycosides of luteolin, including carlinoside, orientin, isoorientin, and C-glycosides of apigenin, schaftoside, isoschaftoside, and vitexin. Furthermore, we assessed the impact of the polyphenolic-rich fraction of C. vulgaris extracts on human skin fibroblasts using the MTT and NR assays. It was found that the extract was non-toxic and exhibited potent antioxidant activity in the cells subjected to induced oxidative stress. Additionally, it effectively protected the cells against H₂O₂-induced cytotoxicity. Our study contributes to the general trend of searching for new phytotherapeutics with potential applications in pharmacy and medicine. The results indicate that further exploration of C. vulgaris species is worthwhile, as they can serve as valuable plant material for cosmetic use.

Keywords: H2O2-induced stress; antioxidant; flavonoid C-glycosides; human skin fibroblast; polyphenols.

Figure 1

An example of C. vulgaris growing in its natural habitat.

Figure 2

Chromatograms of extracts from C. vulgaris obtained using liquid chromatography with a quadrupole-time-of-flight high-resolution mass spectrometer (LC-HRMS-QTOF) and a charged aerosol detector (CAD). (A)—MS chromatogram (electrospray ionization—ESI); (B)—CAD chromatogram.

Figure 3

The chromatograms obtained from ultra-performance liquid chromatography with mass spectrometry and electrospray ionization UHPLC-ESI-MS(−) chromatograms (a) and the UHPLC with photodiode detector—PDA (254 nm) (b). Fractions obtained through liquid–liquid extraction from the extract of C. vulgaris. ECV—methanol extract, HCV—hexane fraction, EaCV—acetate fraction, BCV—butanol fraction, H₂OCV—water fraction.

Figure 4

Effect of the different concentrations of ethyl acetate fraction (EaCV), obtained from methanol/water extract of Carlina vulgaris on cell viability determined by the NR (a) and MTT (b) assay, expressed as a % of control (0.5% of DMSO in medium). The data are means (n = 3) ± SD. One-way ANOVA followed by Dunnett’s post hoc test; the differences were considered significant at p < 0.05. * indicates statistically significant difference.

Figure 5

Effect of different concentrations of ethyl acetate fraction (EaCV), obtained from the methanol/water extract of Carlina vulgaris, on H₂O₂-treated cells evaluated in terms of (a) cell viability (NR) and (b) cellular metabolism (MTT). Cells were pretreated with extract at different concentrations prior to the H₂O₂ exposure. The results are expressed as a percentage of the control (0.5% DMSO). The data are means ± SD (n = 3). * indicates a statistically significant difference (p < 0.05) versus H₂O₂-treated cells assessed using one-way ANOVA followed by Dunnett’s multiple comparison post hoc test.

Figure 6

Relative fluorescence of 2′,7′-dichlorodihydrofluorescein (DCF) in human skin fibroblast cells calculated as a percentage in comparison with untreated control cells. (a)—the cells were treated with H₂O₂ or different concentrations of ethyl acetate fraction (EaCV), obtained from the methanol/water extract of Carlina vulgaris. * indicates a statistically significant difference (p < 0.05) versus untreated controls. (b)—the cells were pretreated with EaCV prior to the H₂O₂ exposure. * indicates a statistically significant difference (p < 0.05) versus the H₂O₂-treated cells. The data are means ± SD (n = 3). One-way ANOVA followed by Dunnett’s multiple comparison post hoc test.