Purification, characterization, and cellular antioxidant activity of 4,4‐dimethylsterols and 4‐desmethylsterols

A mixture of 4,4-dimethylsterols (TA) and 4-desmethylsterols (PS) was produced as valuable byproducts during the large-scale production of ferulic acid by gamma-oryzanol. However, the utilization of both byproducts is impeded by this mixed form. This study has developed a novel method for separating TA and PS through a one-step separation procedure using flash chromatography. The optimal conditions were as follows: mobile phase of hexane-ethyl acetate (9:1, vol/vol), the flow rate of 40 ml/min, loading volume of 20 ml, run time of 20 min. Under optimized conditions, TA with a yield of 41.8% +/- 2.8% with a purity of 93.4% +/- 0.4% was obtained, and 36.8% +/- 1.7% and 98.2% +/- 0.8% for that of PS. Their chemical structures were further confirmed by ultraviolet spectroscopy (UV), Fourier-transform infrared spectroscopy (FT-IR), and gas chromatography-mass spectrometry (GC-MS). The time-fluorescence value curve indicated that TA and PS had inhibitory effects on peroxyl radical-induced 2 ',7 '-dichlorofluorescin (DCFH) oxidation. The results showed that TA had higher cellular antioxidant activity (CAA) than PS. TA was more suitable as a potential natural antioxidant. The present study provides a convenient method for high-purity TA and PS production, which will improve the value-added byproducts from the large-scale production of ferulic acid.