Enantiomer resolution of glycerolipid components of fish oils by chromatography

以含(R)-(+)-1-(1-萘基)乙胺的手性色谱柱(25 cm长)为固定相，建立了多不饱和n-1，2-和n-2，3-二酰基甘油对映体的高效液相色谱快速拆分方法。用正己烷-1，2-二氯乙烷-乙醇(体积比40：30：1)等温洗脱法，在15min内完成了亚油酸、亚麻酸、花生四烯酸、二十碳五烯酸和二十二碳六烯酸的单酸sn-1，2-和sn-2，3-二酰基甘油对映体的拆分，并用于某些海洋生物中三酰甘油和磷脂酰胆碱的对映体拆分。金枪鱼眼眶、太平洋鳗、沙丁鱼、鲱鱼、鲑鱼卵和扇贝肝胰腺的三酰甘油在格氏降解过程中产生的sn-1，2(2，3)-二酰基甘油被清晰地拆分为两组，分别代表sn-1，2-和sn-2，3-二酰基甘油对映体。这种清晰的拆分使得手性柱和收集的组分的极性毛细管气相色谱能够制备分离双酯甘油，这将得到详细的对映体组成。从刀鱼和海胆的三酰甘油中分离得到的sn-1，2(2，3)-二酰基甘油得到了令人满意的拆分，但在含有多不饱和酸的sn-1，2-对映体和含有长链单不饱和酸的sn-2，3-对映体之间有一些峰重叠。用磷脂酶C毒法从秋刀鱼、太平洋鳗和海胆的磷脂酰胆碱中分离得到的甘油二酯仅为sn-1，2-对映体。该方法还用于猪胰腺、扇贝肝胰腺和一些微生物脂肪酶的立体特异性测定。

A rapid ans sensitive high-performance liquid chloromatographic method for the enantiomer resolution of polyunsaturated sn-1,2- and sn-2,3- diacylglycerols as their 3,5-dinitrophenylurethanes was developed with a chiral column (25 cm long) containing (R)-(+)-1-(1-naphthyl)ethylamine. By an isocratic elution with n-hexane-1,2-dichloroethane-ethanol (40 : 30 : 1 by volume), complete enantiomer resolution was obtained within 15 min for monoacid sn-1,2- and sn-2,3-diacylglycerols of linoleic, linolenic, arachidonic, eicosapentaenoic and docosahexaenoic acids.The methos was applied to the enantiomer resolution of the sn-1,2(2,3)-diacylglycerol moieties of triacylglycerols and phosphatidylcholines from some marine organisms. The sn-1,2(2,3)-diacylglycerols generated by Grignard degradation from the triacylglycerols of tuna fish orbital, pacific pomfret flesh, sardine flesh, mackerel flesh, salmon egg and scallop hepatopancreas were clearly resolved into two groups, which represented sn-1,2- and sn-2,3-diacylglycerol enantiomers. This clear resolution permits the preparative separation of the enatimeric diacylglycerols on the chiral column and the polar capillary GLC of the collected fractions, which would give the detailed enantiomer composition. Satisfactory enantiomer resolution was obtained for the sn-1,2(2,3)-diacylglycerols generated from the triacylglycerols of saury flesh and sea urchin gonard, although some peak overlapings between sn-1,2-enantiomers containing polyunsaturated acids and sn-2,3-enantiomers containing long-chain monounsaturated acids were observed. The diacylglycerols derived from phosphatidylcholines of saury, pacific pomfret and sea urchin by venom phospholipase C were only the sn-1,2-enantiomers. The method presented was also applied to the determination of the stereospecificity of lipases from porcine pancreas, scallop hepatopancreas and some microorganisms.