Rastegar A, Pelletier A, Duportail G, Freysz L, Leray C
Centre de Neurochimie du CNRS, Strasbourg, France.
J Chromatogr. 1990 Sep 28;518(1):157-65. doi: 10.1016/s0021-9673(01)93171-1.
A sensitive high-performance liquid chromatographic (HPLC) method for the separation and determination of diacylglycerophospholipid and diacylglycerol (DAG) molecular species has been developed. Phospholipids are hydrolysed with phospholipase C and the resulting DAGs are reacted with naproxen chloride in the presence of 4-dimethylaminopyridine. The naproxen-DAGs were purified by thin-layer chromatography on silica gel G plates. Molecular species were separated using reversed-phase HPLC with isocratic elution and determined by measuring the absorbance at 230 nm or fluorescence at 352 nm (excitation at 332 nm). The method was applied to the determination of diacylglycerophosphoethanolamine in rat cerebrum and cerebellum. The molar absorption coefficient of the naproxen derivatives was 53,000 lmol-1 cm-1 at 230 nm, permitting the generation of linear concentration-dependent determinations down to less than 10 pmol. A ten-fold increase in sensitivity was obtained with a fluorescence detection system owing to the fluorescent properties of the proposed adduct.
已开发出一种灵敏的高效液相色谱（HPLC）方法，用于分离和测定二酰基甘油磷脂和二酰基甘油（DAG）分子种类。磷脂用磷脂酶C水解，所得的DAG在4-二甲基氨基吡啶存在下与氯萘普生反应。萘普生-DAG通过硅胶G板上的薄层色谱法纯化。分子种类使用反相HPLC通过等度洗脱进行分离，并通过测量230nm处的吸光度或352nm处的荧光（332nm激发）进行测定。该方法应用于大鼠大脑和小脑中二酰基甘油磷酸乙醇胺的测定。萘普生衍生物在230nm处的摩尔吸收系数为53,000 lmol-1 cm-1，可进行低至小于10 pmol的线性浓度依赖性测定。由于所提出加合物的荧光特性，使用荧光检测系统可使灵敏度提高十倍。
