Phytochemical Profile, Antioxidant Capacity and Anticancer Potential of Water Extracts from In Vitro Cultivated Salvia aethiopis

Abstract

Salvia aethiopis L. (Mediterranean sage) is a medicinal plant known for its rich phenolic content and different therapeutic properties. This study evaluated the phytochemical composition, antioxidant capacity and anticancer potential of water extracts from in vitro cultivated S. aethiopis. The extract exhibited a high total polyphenol (110.03 ± 0.7 mg GAE/g) and flavonoid (7.88 ± 0.25 mg QE/g) content, along with a strong oxygen radical absorbance capacity (an ORAC value of 3677.9 ± 24.8 µmol TE/g). LC-HRMS analysis identified 21 bioactive compounds, including salvianic acid C, rosmarinic acid, salvianolic acid K and various organic acids. A cytotoxicity evaluation using the Neutral Red Uptake assay showed that the extract had a low toxicity to non-cancerous BALB/3T3 cells. An antiproliferative activity assessment via the MTT assay revealed selective cytotoxicity against Hep G2 hepatocellular carcinoma cells (IC₅₀ = 353.8 ± 21.8 µg/mL) and lung (A549) and prostate (PC-3) carcinoma cell lines. Migration assays and cytopathological evaluations confirmed the significant inhibition of cancer cell proliferation, the suppression of migration and G2/M cell cycle arrest. Flow cytometry revealed considerable increases in apoptotic and necrotic cell populations following treatment with S. aethiopis extract. These findings showed the potential of S. aethiopis as a promising source of bioactive compounds with antioxidant and anticancer properties, supporting its further exploration for therapeutic applications.

Keywords: LC-HRMS; Salvia aethiopis; antioxidant activity; apoptosis; cytotoxicity; hepatocellular carcinoma; phytochemicals.

Figure 1

Cytotoxicity of S. aethiopis extract on BALB/3T3 cells assessed by the Neutral Red Uptake (NRU) test. Data are expressed as mean ± SD. Statistical significance: *** p < 0.001.

Figure 2

Antiproliferative effect of S. aethiopis extract on the cell viability of human cell lines assessed by the MTT test after 72 h of exposure. Data are presented as means ± SD. Statistical significance: * p < 0.05, ** p < 0.01 and *** p < 0.001.

Figure 3

Effect of S. aethiopis extract on the migration capacity of hepatocellular carcinoma (Hep G2) cells. Statistical significance: *** p < 0.001.

Figure 4

Cytopathological alterations induced by S. aethiopis extract in hepatocellular carcinoma (Hep G2) cells. (a,d) Negative control (untreated cells); (b,e) cells treated with S. aethiopis extract (300 µg/mL); (c,f) positive control (cells treated with 6 µg/mL doxorubicin). Upper panel: AO/EB staining. Lower panel: DAPI staining. Fluorescence microscopy, 40× objective.

Figure 5

Effect of S. aethiopis extract on the cell cycle progression of Hep G2 hepatocellular carcinoma cells. Cells were analyzed for distribution in the individual phases of the cell cycle by examining FL2 peak heights (FL2-H, Fluorescence Channel 2—Height) (left and middle panel). Statistical significance: ** p < 0.01 and *** p < 0.001.

Figure 6

Apoptotic response of Hep G2 hepatocellular carcinoma cells following treatment with S. aethiopis extract as assessed by flow cytometry. Statistical significance: *** p < 0.001.

Figure 7

Cultivated S. aethiopis in the experimental field.