Purfication Of Polyprenole From Ginkgo Biloba L. Leaves And Synthesis Of S-dolichol Thereof

Polyprenols from Ginkgo biloba L. leaves generally compose of 15 to 21 unsaturated isoprene units and are terminated by an unsaturated isoprene unit bearing a primary hydroxyl group. Polyprenols are natural products. They are not only non-toxic, non-mutagenic, non-teratogenic, and non-carcinogenic in humans but also have distinct physiological and pharmacological functions. Polyprenols mainly exist in the form of acetates and a few are in the form of free alcohols. These account for 0.5 %² % according to the dry G. biloba L. leaves. S-dolichol is a linear polyisoprene compound with a terminal unit of saturated isopropane bearing a primary hydroxyl group, which is found in the lipid fraction of animal and plant tissues. S-dolichol is the key carrier in the biosynthesis of glycoprotein, with biological functions such as free radical scavenging, antioxidation, hematopoiesis etc. Polyprenols are ideal as initial building blocks for the semisynthesis of S-dolichol.The purity of polyprenols was determined by High Performance Liquid Chromatography (HPLC) using external standard method. The regression equation obtained from the standard curve was y=1.5751x+1.1546 (R²=0.9991), and y is the peak area of the 5 main peaks, x is the amount of polyprenols. The purity of polyprenols was calculated according to the standard curve. The linear relation between the amount of polyprenols (4.6²3.3μg) and peak area was good.Decoloration technology was applied to the purification of polyprenols, and the traditional purification process for polyprenols was optimized. L₉(3⁴) orthogonal experiment design was employed to optimize the conditions for decoloration. Decoloration of polyprenols could be achieved effectively by stirring crude polyprenols in petroleum ether (1:10, w/v) and adding mixed decolorant (activated carbon/attapulgite ratio of 1:5, w/w) with a polyprenols/decolorant ratio of 1:1.2 (w/w) for 20 min at 70℃. Decoloration applied under the optimal condition could improve the purity of polyprenols from 38.5 % to 49.6 % without loss of polyprenols. The purity could be further improved to over 90 % through only one cycle of silica gel column chromatography. This developed purification method is convenient, rapid, economical, high-yield, and suitable for industrial production, among others.Due to the lack of commercially available optically pure compounds with a similar structure, C5 synthon ((R)-4-benzyloxy-1-bromo-2-methylbutane) was synthesized from the readily available 3-methyl-3-buten-1-ol using a slightly more complex procedure involving an enzymatic enantio-resolution step. At first, the hydroxyl function of 3-methyl-3-buten-1-ol was protected with a benzyl group carried out under phase-transfer conditions. The unsaturated benzyl ether was converted to the racemic primary alcohol by a hydroboration–oxidation reaction, followed by the stereoselective acetylation and bromination.Polyprenols with high purity were conventionally acetylated with acetic anhydride and pyridine. S-dolichyl benzyl ether was synthesized using Grignard coupling reaction between polyprenyl acetates and the Grignard reagent of C5, which was carried out in THF catalyzed by Li₂CuCl₄. The S-dolichyl benzyl ether could be deprotected by reduction with alkali metal in ammonia solution at -78℃and S-dolichol can be obtained.