Purification Of Cholesterol From Wool Wax Alcohol

Cholesterol, which is a typical basic molecular in physiology, has been widely applied in various industry fields. The wool wax alcohol, which is one of the derivatives of lanolin, has plenty of cholesterol, so it is significant to isolate the cholesterol. This study was focus on the separation process of cholesterol, including the liquid-liquid extraction, column chromatography and crystallization.In wool wax alcohol, the components were complex. In order to get comprehensive analysis result, quantitative detection of several components was carried out on HPLC while the qualitative detection of all components was carried out on GC. So the analysis conditions of HPLC and GC were determined at first.The liquid-liquid extraction was used to remove the highly polar impurities in wool wax alcohol. A series of polar solvent was chosen to separate octadecanol from cholesterol in hexane and the mixture solvent of methanol/H2O was screened out as the proper extractant due to its high selectivity. Then the extraction of impurities in wool wax alcohol was studied. Several influencing factors were studied, such as the ratio of methanol to H2O, temperature and the concentration of wool wax alcohol. The optimized operation conditions were as follows:methanol/H2O (9:1,v/v) was used as extractant, the proper temperature was 35～40℃and the concentration of wool wax alcohol was 80-100mg/mL, then the value of selectivity coefficient was higher than 3. The optimum conditions above were used for countercurrent extraction. The acid value of wool wax alcohol was decreased from 3.16 to 0.91 after extraction and the purity of the cholesterol was evaluated by 5-6%.Then the extracted wool wax alcohol was purified by column chromatography. According to the resolution of cholesterol and lanosterol, silica was proved to be the proper adsorbent and the process conditions were optimized, including composition of eluent, temperature and the charge of column. The optimum conditions were as follows:the mixture solvent of acetone/hexane (4:96, v/v) was used as eluent, the proper temperature was 35℃and the proper charge of column was 2.4%. After separated by column chromatography under the conditions above, the purity of cholesterol was 88.63% and the recovery was 96.33%. The silica was regenerated by the mixture solvent of acetone/ hexane (50:50, v/v). It was formed that the extraction process ensured complete regeneration. The separation ability of silica gel remained almost unchanged after 7 circulations of usage and regeneration.The cholesterol was further purified by crystallization. The process conditions were optimized as follows:methanol was used as the crystallizing solvent, ratio of solid to liquid was 1:20, the temperature was 70℃at beginning and cooled to 30℃slowly in 3 hours, and then kept for 30 minutes. The purity of cholesterol was 98.7% at last while the recovery was 69.4%, and the melting range of which was between 147.1-147.4℃.