Chromatographic purification of gamma oryzanol from by-products of rice bran oil refinery

Abstract

The objective of this study is to separate and purify γ-oryzanol from the extract of saponified soapstock or the extract of hydrolyzed acid oil, by-products from rice bran oil processing, using normal phase chromatography. To remove the main impurity which is glycerides, soapstock was saponified with NaOH in methanol, and acid oil was hydrolyzed with aqueous solution of NaOH, prior to extraction with ethyl acetate. Upon saponification of soapstock and hydrolysis of acid oil, glycerides contents are much reduced, however loss of γ-oryzanol was also observed, particularly in the case of soapstock in which as high as 41% loss of γ-oryzanol (from 6.73% to 4.07%) was resulted. Thus the extract of hydrolyzed acid oil which has higher γ-oryzanol content of 5.38 %wt was selected as a suitable raw material for γ-oryzanol recovery. Chromatography was employed as a purification process and it began with determining the suitable composition of mobile phase by thin layer chromatography (TLC), in which hexane and ethyl acetate mixture at 75:25 v/v was found to be most suitable. This solvent system was employed in a semi-preparative chromatography operated with isocratic mode, in which the effects of the amounts (5 g, 10 g, and 15 g) and particle sizes of silica gel (15-25 μm, 25-40 μm and 40-63 μm) on the column performance were determined. The result suggested that the column packed with 10 g silica gel of 25-40 μm particle size gave the highest yield (83.64%) of high purity γ-oryzanol (>95%). The yield and purity of the resulted product could be further improved to 90.15% and 100%, respectively, by applying gradient elution mode, in which 85:15 v/v of hexane and ethyl acetate mixture was used as a mobile phase in the first hour before it was switch to 75:25 v/v of hexane and ethyl acetate.