Samet J M, Friedman M, Henke D C
Department of Medicine and Curriculum in Toxicology, University of North Carolina, Chapel Hill 27599-7020.
Anal Biochem. 1989 Oct;182(1):32-6. doi: 10.1016/0003-2697(89)90713-6.
An HPLC method for the separation and analysis of arachidonic acid and eight phospholipid classes is described: phosphatidylglycerol, phosphatidylinositol, cardiolipin, phosphatidylserine, phosphatidylethanolamine, phosphatidylcholine, sphingomyelin, and 2-lysophosphatidylcholine. The separation is carried out at 60 degrees C on 2 cyanopropyl columns using a gradient of acetonitrile and 5 mM sodium acetate (pH 5.0). Cyanopropyl columns require a lower proportion of water in the mobile phase to elute the more polar phospholipids than other types of columns and are thus less prone to equilibration problems. The method is highly reproducible (average coefficient of variation for each retention time less than or equal to 3.5%) and permits analysis of peaks by phosphorus content. Data obtained by analyzing lipid extracts from rat alveolar macrophages prelabeled with [G-3H]-arachidonic acid were analyzed by this HPLC method and compared to standard analysis by TLC. There was a significant correlation between the radioactivity profiles obtained with the two chromatographic methods (HPLC versus TLC) by linear regression analysis [HPLC = 0.83 (TLC) + 3.58, n = 25, r = 0.95, P less than 0.001].
磷脂酰甘油、磷脂酰肌醇、心磷脂、磷脂酰丝氨酸、磷脂酰乙醇胺、磷脂酰胆碱、鞘磷脂和2-溶血磷脂酰胆碱。分离在60℃下于两根氰丙基柱上进行，使用乙腈和5 mM醋酸钠（pH 5.0）的梯度洗脱。与其他类型的柱子相比，氰丙基柱在流动相中需要较低比例的水来洗脱极性更强的磷脂，因此不太容易出现平衡问题。该方法具有高度的重现性（每个保留时间的平均变异系数小于或等于3.5%），并允许通过磷含量分析峰。用此高效液相色谱法分析了用[G-3H] -花生四烯酸预标记的大鼠肺泡巨噬细胞脂质提取物获得的数据，并与薄层色谱法的标准分析结果进行了比较。通过线性回归分析，两种色谱方法（高效液相色谱法与薄层色谱法）获得的放射性图谱之间存在显著相关性[高效液相色谱法 = 0.83（薄层色谱法）+ 3.58，n = 25，r = 0.95，P < 0.001]。
