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Isolation, cytotoxic evaluation, and simultaneous quantification of eight bioactive secondary metabolites from Cicer microphyllum by high‐performance thin‐layer chromatography
Author(s) -
Dar Alamgir A.,
Rath Santosh K.,
Qaudri Afnan,
Singh Buddh,
Tasduq Sheikh A.,
Kumar Anil,
Sangwan Payare L.
Publication year - 2015
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201500861
Subject(s) - chromatography , ethyl acetate , chemistry , luteolin , formic acid , thin layer chromatography , oleanolic acid , ammonium acetate , silica gel , high performance liquid chromatography , quercetin , organic chemistry , medicine , alternative medicine , pathology , antioxidant
Chemical investigation of Cicer microphyllum resulted in the isolation and characterization of eight natural products viz. Stigmasterol, Oleanolic acid‐3‐acetate, Oleanolic acid, Biochanin A, Genistein, Pratensein, Chrysoeriol, and Luteolin. Herein, we report a novel, accurate, and cost‐effective high‐performance thin‐layer chromatography method for the simultaneous quantification of the isolated natural products on silica‐gel 60F 254 plates using the solvent system n ‐hexane/ethyl acetate/formic acid (9.0:6.5:0.8, v/v/v). Natural products were quantified after postchromatographic derivatization with ceric ammonium sulfate. The method was validated as per the International Conference on Harmonization guidelines. All calibration curves showed a good linear relationship ( r > 0.9943) within the test range. Precision was assessed by intra‐ and interday tests with relative standard deviations <1.82%, accuracy validation recovery 98.38–99.57% with relative standard deviations <1.00%. On quantification, Pratensein was a major constituent (0.921%). The screening for cytotoxic activity using a 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assay resulted into identification of Luteolin as potent molecule with IC 50 3.5 and 25.6 μg/mL against murine melanoma and human epidermoid carcinoma cell lines, respectively.