芳香酯酶测定底物探讨及临床应用评价

目的该科合成芳香酯酶(ArylesterhydrolaseEC3.1.1.2,ArE)底物4-已酰氧基-3-甲氧基-ω-硝基乙烯;建立手工的和自动生化仪检测方法;评价在常见肝病中的应用。方法底物合成:香草醛加冰乙酸、乙酸铵。在乙醇溶液中用硝基四烷进行硝基化,滤取结晶。用乙酸酐进行酯化,滤取结晶,重结晶。方法建立。程序:取pH8.6Tris-HCL缓冲液2.5ml,加血清30ul,再加3mmol/l底物液0.4ml,37℃10min,加5ml/dl正丁醇3ml。混匀,在520nm,读取吸光度。摩尔消光系数(20345±280)A。计算:ArEU/L=971.3×A。结果反应条件:底物溶剂以甲醇13.65%为最佳。产物光吸收峰498nm,选用520nm,吸光度达93.20%。择取pH8.6缓冲液,加0.51mmol/LCa2 和68.3mmol/LNa 可提酶活力14.00%。批间CV为3.21%~4.52%。溶血、黄疸和脂血标本对ArE结果无干扰。临床应用:90例健康人ArE(470.6±49.7)U/L,参考值:<421U/L。肝硬化、肝癌、肝炎3组ArE结果x±S,U/L分别是(266±77),(35±84)和(366±111);ROC曲线下面积分别是0.979,0.910和0.807。面积愈大诊断价值愈高,ArE愈低,肝功能愈差。尤其对肝硬化、肝癌。肝硬化组<421U/L阳性率为92.30%;肝癌ArE(Y)与脂碱脂酶CHE相关系数为0.909,Y=0.045X 190.53。结论ArE简便可靠。在肝硬化、肝癌、肝炎中阳性率性高,特异性好。有望     

Characterization of the esterases of guinea pig liver and kidney

The esterases of cavian liver and kidney were separated electrophoretically into 12 and 14 distinct, tissue-specific bands, respectively. Based on substrate specificity and subcellular fractionation studies, the tissue esterases were identified as carboxylesterases (aliesterases) of microsomal origin. There was no evidence of arylesterase or cholinesterase activity in either tissue. Inhibition studies revealed the presence of two types of carboxylesterases in each tissue, one type being sensitive to organophosphorus esters, the second being resistant. Gel filtration and molecular weight estimation on a Sephadex G-200 column yielded two distinct molecular forms of hepatic enzymes having mol. wt. of 240,000 and 56,000, respectively. The higher molecular weight form could be dissociated into units of 56,000 mol. wt by a combination of acidic pH and high salt concentration. One molecular form was observed for renal esterases, the molecular weight being of the order of 56,000. The presence of electrophoretically different multiple forms of activity in the fractions isolated by gel filtration suggested the existence of charge isomers as well as readily formed aggregates.     

慢性肾功能衰竭血清芳香酯酶活性改变的研究

目的研究慢性肾功能衰竭 (CRF)患者血清芳香脂酶 (ArE)活性改变的临床意义。方法 :通过测定CRF早期、晚期与终末期患者血清ArE活性、胆固醇、高密度脂蛋白胆固醇 (HDLc)、氧修饰低密度脂蛋白 (ox -LDL)与脂蛋白 (a) [Lp(a) ]及内生肌酐清除率 (Ccr)等的水平 ,进行分析。结果 :CRF患者血清ArE活性与HDLc水平低于对照组 ,血清ox -LDL、Tc与LP(a)水平高于对照组。三期肾功能衰竭患者血清ArE活性与HDLc水平降低程度自小至大依次为早期、中期与晚期 ,血清ox -LDL、Tc与LP(a)升高程度在三组间有显著性差别 ,其中以终末期最高。血清ArE活性与HDLc、白蛋白、Ccr呈正相关 ,与血清肌酐呈负相关 ,和Lp(a)无相关性。结论 :测定CRF患者血清ArE活性不仅了解其血脂与脂蛋白的代谢异常 ,还可为了解肾功能损害程度提供参考依据。