Development of Peroxyoxalate Chemiluminescence Assay Method for Platelet Activationg Factor

对流动注射分析法测定血小板活化因子（PAF）和含胆碱磷脂进行了初步研究。1)采用戊二醛法将磷脂酶D和胆碱氧化酶固定在烷基胺玻璃微珠上，装入不锈钢小柱中，制成柱反应器。2)流动注射分析条件为：载体溶液为10 mM咪唑缓冲液（pH 8），含10 mM Co（NO_3）_2和0. 1% Triton-X，化学发光试剂为0.2mM TCPO和0.25mM TMP的乙腈溶液，流速为1.0ml/min。[1994.4-1995.3]考察了PAF与磷脂的HPLC分离。1)蒸发对应于每种含胆碱磷脂的洗脱液，并将其溶解在0. 1% Triton-X中，将所得物的等分试样施加到FIA系统中。校准曲线线性高达180 pmol的柱上与较低的pmol水平的检测限。2)正常人血清中的总胆碱浓度为102-168 mg/dL，与市售试剂盒获得的浓度相当。4)正常人血清中磷脂酰胆碱、鞘磷脂和溶血磷脂酰胆碱的比例为80：15：5。该方法简便、灵敏，可用于PAF及其相关物质的测定。

[1993.4-1994.3] A preliminary study on a flow-injection analysis (FIA) has been performed for the determination of platelet activating factor (PAF) and choline-containing phospholipids. 1) Phospholipase D and sholine oxidase were immobilized to alkylamine glass beads by a glutaraldehyde method, packed into a small stainless steel column to prepare column reactor. 2) FIA conditions are as follows : carrier solution was 10 mM imidazole buffer (pH 8) containing 10 mM Co (NO_3)_2 and 0.1% Triton-X ; chemiluminescence reagent was a mixture of 0.2 mM TCPO and 0.25mM TMP in acetonitrile ; flow rate was 1.0ml/min. 3) Calibration curves were linear up to 200 pmol with the detection limits of lower pmol levels.[1994.4-1995.3] HPLC separation of PAF and phospholipids were examined. 1) The eluent corresponding to each choline-containing phospholipid was evaporated and dissolved in 0.1% Triton-X,and an aliquot of the resultant was applied to the FIA system. Calibration curves were linear up to 180 pmol on column with the detection limits of lower pmol levels. 2) Total choline concentrations in normal human sera were 102-168 mg/dL,which were well corresponded to those obtained by the commercially available kits. 4) The ratio of phosphatidylcholine, sphingomyeline, and lyso-phosphatidylcholine in normal serum was 80 : 15 : 5.As the proposed method is convenient and sensitive, it should be useful for the determination of PAF and related compounds in biomedical and clinical studies.