Pelletier O, Wright L A, Breckenridge W C
Clin Chem. 1987 Aug;33(8):1403-11.
We describe a new gas-chromatographic/mass-spectrometric (GC/MS) isotope-dilution method for determination of serum cholesterol. The method has been fully optimized and documented to provide the high accuracy and precision expected for a Definitive Method. In the presence of [3,4-13C]cholesterol, cholesteryl esters in serum are hydrolyzed under optimum conditions and the entire cholesterol pool is extracted and derivatized to silyl ethers. The cholesterol derivatives are resolved from other sterols by gas-liquid chromatography on a fused silica column, and selected ions characteristic of cholesterol and the [3,4-13C]cholesterol are monitored with a GC/MS quandrupole system. We estimated the cholesterol content of samples by bracketing each sample with standards of comparable cholesterol concentration that also contained the [3,4-13C]cholesterol. The procedure was highly reproducible (CV less than 0.5%), better accuracy and precision being obtained with [3,4-13C]cholesterol than with heptadeuterated cholesterol. Mean values per gram of dry serum for one serum pool assayed by this method and that of the National Bureau of Standards differed by 0.5%. We conclude that the method satisfies the criteria for a Definitive Method.
我们描述了一种用于测定血清胆固醇的新的气相色谱/质谱（GC/MS）同位素稀释法。该方法已得到充分优化并记录在案，以提供确定性方法所期望的高精度和精密度。在[3,4-¹³C]胆固醇存在的情况下，血清中的胆固醇酯在最佳条件下水解，整个胆固醇库被提取并衍生为硅醚。胆固醇衍生物通过在熔融石英柱上的气液色谱法与其他甾醇分离，并用GC/MS四极杆系统监测胆固醇和[3,4-¹³C]胆固醇的特征选择离子。我们通过用含有[3,4-¹³C]胆固醇的胆固醇浓度相当的标准品对每个样品进行括值来估计样品中的胆固醇含量。该程序具有高度可重复性（变异系数小于0.5%），使用[3,4-¹³C]胆固醇比使用十七氘代胆固醇获得了更好的准确性和精密度。用该方法测定的一个血清池每克干血清的平均值与国家标准局的平均值相差0.5%。我们得出结论，该方法符合确定性方法的标准。
