Antioxidant and anticancer potentiality of F13 fraction of Pleurotus sajar caju against cancer cell lines and in silico analysis

Novel drugs, isolated from natural compounds, are currently being tested and sought out, and here, Pleurotus sajar caju was considered as a source for potentially novel drugs. The current study focuses on identifying the bioactive compounds of partially purified fractions (F1-F20) from PSME (Pleurotus sajar caju methanolic extract) in order to demonstrate the antioxidant activity and anticancer activity of F13 on cervical, lung, and breast cancer cell lines with mechanisms and also in silico study of selected compounds. The results of the qualitative analysis of PSME exhibited that phenol, flavonoid, carbohydrate, and alkaloid contents were in high (+++) amounts. FT-IR analysis of the extract showed several functional groups, like O-H stretching for carboxylic acids, C-H stretching for alkanes and alkyl groups, etc. The PSME underwent column chromatography for fractionation and partial purification, obtaining 20 fractions (F1-F20). Fractions were tested for antioxidant content and activity, and network analysis of the co-occurrence pattern within the antioxidant content and activity of different fractions of PSME was done by R software, and F13 was judged as the best fraction. The F13 had the highest radical scavenging activity with EC₅₀ values of 21.65 ± 0.81 µg.mL^-1 by DPPH (1,1 diphenyl-2-picrylhydrazyl), and it also had maximum scavenging activity among other fractions by the FRAP (ferric reducing antioxidant power) method. Total phenolic content, flavonoid content, and ascorbic acid content of F13 were maximum among other fractions. TLC analysis of the F13 fraction revealed six distinct spots with Rf values (0.36, 0.52, 0.65, 0.74, 0.36, and 0.87), but three spots matched perfectly with the Rf values of the standards of quinine (0.37), quercetin (0.74), and p-coumaric acid (0.87). The cytotoxicity effect of F13 from PSME at the highest concentration of 1500 µg.mL^-1 on the HeLa cell line was 90.66 ± 3.05% cell growth inhibition, compared to other cell lines at 24 h. The F13 induced apoptosis and LDH leakage. It upregulated gene expression of Caspase 3 & 9 and P53 and downregulated BcL2 genes of all cell lines, as proved by our Western blotting experiment. In silico analysis exhibited that quercetin and p-coumaric acid strongly bound with receptors BcL2 and Caspase 3, inhibiting BcL2 function and enhancing Caspase 3 function. In conclusion, the present study has demonstrated that F13, having six bioactive compounds, was a highly potent antioxidant and anti-breast, anti-lung, and anti-cervical cancer agent. After in vivo and clinical trials in humans, it would be an important product for next-generation drug preparation for cancer management.