Comparative determination of plasma phospholipids by automated gas--liquid chromatographic and manual colorimetric...

Plasma samples obtained during a prevalence study of hyperlipemia in a free-living urban population were analyzed for phosphatidylcholine, sphingomyelin and lysophosphatidylcholine content by automated high-temperature gas--liquid chromatographic (GLC) and manual colorimetric phosphorus (thin-layer chromatographic, TLC) methods. The GLC estimates were obtained from a quantitative analysis of the diacylglycerol, ceramide and monoacylglycerol moieties released from the parent phospholipids by digestion with phospholipase C, while the TLC estimates were derived by manual colorimetric phosphorus analyses of the individual phospholipid classes resolved by TLC. On samples analyzed over a two-year period the methods gave excellent correlation for the total phospholipids (r = 0.98), phosphatidylcholine (r = 0.98) and sphingomyelin (r = 0.90), but resulted in a poor agreement for lysophosphatidylcholine (r = 0.69). Comparable results were obtained for estimates of these phospholipids in plasma very low density, low density and high density lipoproteins. The between-method coefficient of variation ranged from 3 to 5% for phosphatidylcholine and from 5 to 10% for sphingomyelin. The relative error for the estimates of lysophosphatidylcholine ranged from 10 to 25%, and was due to the inclusion in the GLC estimates of a variable proportion of plasma free monoacylglycerols. Other differences between the two methods are due to various analytical errors and biases inherent in the two techniques. The within-day, within GLC, relative error averaged 1% for phosphatidylcholine, 3% for sphingomyelin and 5% for lysophosphatidylcholine. The apparent high precision and accuracy of the GLC method recommend it as an alternative to conventional direct methods of phospholipid analyses based on TLC isolation of lipid classes and colorimetric measurements of their phosphorus content. The GLC analyses of the plasma phospholipids are particularly convenient in conjunction with GLC measurements of plasma cholesterol and triacylglycerols, where a smaller throughput of samples is not a limitation and where both total amount and relative proportion of the lipids are of interest.