高效液相色谱测定血清高密度脂蛋白胆固醇分数和摩尔酯化速率

目的 建立一种高效液相色谱(HPLC)测定血清高密度脂蛋白胆固醇分数和摩尔酯化速率(FERHDL和MERHDDL)的方法.方法 用5,5'二硫代-2-硝基苯甲酸(DTNB)抑制全血中的卵磷脂胆固醇酰基转移酶(LCAT)活性,分离血清,制备HDL;HDL分别在含和不含2-巯基乙醇(ME)的条件下37℃温育1 h,HPLC法测定HDL游离胆固醇,计算FERHDL和MERHDL.结果 DTNB可以有效抑制全血中的LCAT活性,使游离胆固醇稳定在初始水平;ME可有效解除DTNB对LCAT的抑制作用;测定血清FERHDL和MERHDL的总变异系数分别为1.59%～3.74%和1.64%～2.88%;70名健康志愿者FERHDL.均值为18.7%/h(标准差7.2%/h),中位数16.1%/h;MERHDL均值42.7 μmol·L-1·h-1(标准差11.8 μmol·L-1·h-1),中位数41.6 μmol·L-1·h-1;FERHDL和MERHDL与多种心血管病危险因素显著相关.结论 本研究建立了血清FERHDL和MERHDL准确测定的方法,方法安全、简便、精密,将为进一步研究和应用FERHDL和MERHDL打下基础.

Fractional and molar esterification rate of high density lipoprotein cholesterol determined by high performance liquid chromatography

Objective To develop an HPLC method for the measurement of fractional and molaresterification rate of serum HDL(FER HDL and MERHDL).Methods Blood samples were mixed with 5,5'-dithiobis-(2-nitrobenzoic acid)(DTNB)and the sera were separated.Serum HDL fractions were prepared by precipitation with Dextran sulfate and magnesium and the fractions were incubated at 37℃for 1 h in the presence and absence of 2-mercaptoethanol(ME).Free cholesterol levels of the HDL fractions were analyzed by HPLC and FERHDL and MERHDL were calculated.Results Under the selected conditions,serum free cholesterol could be stabilized by inhibition of LCAT with DTNB and the inhibition be reversed by ME. The total CVs for FERHDL and MERHDL were 1.59%-3.74% and 1.64%-2.88%,respectively.The averages of FERHDL and MERHDL in 70 apparently healthy subjects were 18.7%/h and 42.7 μmol·L-1·h-1 with standard deviations of 7.2%/h and 11.8 μmol·L-1·h-1·respectively,and the medians were 16.1%/h and 11.8 μmol·L-1·h-1.Close correlations of FERHDL.and MERHDL with other cardiovascular disease risk factors were observed.Conclusion A new method for the measurements of FERHDL and MERHDL by HPLC has been estabished. The method is safe, precise and simple and applications in the assessment of cardiovascular diseases risks are expected.