Identification of secondary metabolites in polar organic extract of the fungus Cladobotryum virescens (Hypocreaceae, Ascomycota) by UHPLC-ESI-MS/MS: Cytotoxic and mutagenic activity

The genus Cladobotryum is known for causing cobweb disease in edible mushroom crops. Its parasitic nature has prompted interest in its biocontrol potential and ability to produce cytotoxic compounds. The aim of this study was to identify secondary metabolites in a metahnolic extract of Cladobotryum virescens G.R.W. Arnold, and to evaluate its cytotoxicity against four cancer cell lines and its mutagenic effect in vitro. The chemical identification of metabolites was conducted using ultra-high performance liquid chromatography electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) in negative ionization mode. Cytotoxicity was assessed via MTT assay on MCF-7, MDA-MB-231 (both breast cancer), H460 (lung cancer), and L1210 (murine lymphocytic leukemia line) cell lines. A bacterial reversion test using Salmonella typhimurium strains TA98 and TA100 (Ames' test) was carried out to evaluate mutagenicity. UHPLC-ESI-MS/MS analysis allowed us to tentatively identify a total of 13 compounds in the extract, including polyketides, anthraquinones, and polyphenols mainly glycosylated flavonoids, among others. The methanolic extract of C. virescens showed significant dose-dependent cytotoxic against various cancer cell lines (IC₅₀ = 5.02, 50.15, 12.05 μg/mL for MCF-7, MDA-MB-231, and L1210, respectively) and remarkable primary mutagenic potential. These findings highlight the bioactive potential of C. virescens as a promising source of secondary metabolites for cancer treatment. Particularly, antioxidants may play a role in their cytotoxic effect and support further exploration of Cladobotryum species as a source of biologically active compounds.