Bioscience Division, Tosoh Corporation, Kanagawa, Japan.
J Lipid Res. 2010 May;51(5):1237-43. doi: 10.1194/jlr.D003624. Epub 2009 Oct 30.
We previously established an analysis method for determining the cholesterol levels of five major lipoprotein classes [HDL, LDL, intermediate density lipoprotein (IDL), VLDL, and chylomicrons] in serum by an anion-exchange (AEX)-HPLC method, but lipoprotein(a) [Lp(a)], a well-known risk factor for atherosclerotic diseases, was not determinable. Therefore, we established new AEX-HPLC separation conditions for analyzing the cholesterol levels of six lipoprotein classes, including Lp(a). Serum lipoproteins were separated by HPLC with a diethylaminoethyl-ligand nonporous polymer-based column by elution with a stepwise gradient of the sodium perchlorate concentration. In this improved method, HDL, LDL, IDL, VLDL, chylomicrons, and Lp(a) were each eluted from the column. The cholesterol levels of the eluted lipoproteins were measured enzymatically by a postcolumn reaction. The within-day assay and between-day assay coefficients of variation for the lipoprotein cholesterol levels were in the ranges of 0.29-11.86% and 0.57-11.99%, respectively. The Lp(a) cholesterol levels determined by AEX-HPLC were significantly correlated with the amounts of Lp(a) protein measured by an immunoturbidimetry method available commercially (r = 0.9503, P < 0.0001). Taken together, this AEX-HPLC method may be effectively applied to the analysis of serum lipoproteins with high levels of Lp(a).
我们之前建立了一种通过阴离子交换（AEX）-HPLC 方法测定血清中五种主要脂蛋白类（HDL、LDL、中间密度脂蛋白（IDL）、VLDL 和乳糜微粒）胆固醇水平的分析方法，但脂蛋白（a）[Lp(a)]，一种众所周知的动脉粥样硬化疾病的危险因素，无法确定。因此，我们建立了新的 AEX-HPLC 分离条件，用于分析包括 Lp(a)在内的六种脂蛋白类的胆固醇水平。血清脂蛋白通过 HPLC 用基于二乙基氨基乙基配体的多孔聚合物柱分离，通过逐步梯度增加高氯酸钠浓度进行洗脱。在这种改进的方法中，HDL、LDL、IDL、VLDL、乳糜微粒和 Lp(a)从柱上依次洗脱出来。用柱后反应通过酶法测定洗脱脂蛋白的胆固醇水平。脂蛋白胆固醇水平的日内和日间测定变异系数分别在 0.29-11.86%和 0.57-11.99%的范围内。通过 AEX-HPLC 测定的 Lp(a)胆固醇水平与市售免疫比浊法测定的 Lp(a)蛋白量呈显著相关（r=0.9503，P<0.0001）。综上所述，这种 AEX-HPLC 方法可能有效地应用于分析高 Lp(a)水平的血清脂蛋白。
