Evaluation of a simple colorimetric assay for serum angiotensin-converting enzyme: comparison with a new ion-pair liquid chromatography-assisted assay.
The development and validation of two different assays for serum angiotensin-converting enzyme are reported. The first step in both analytical systems is based on enzymatic cleavage of the synthetic substrate, hippuryl-histidyl-leucine, under conditions advocated by Cushman and Cheung. The hippuric acid released is then assayed either by colorimetry following an azlactone condensation reaction with an aromatic aldehyde, or by reversed-phase ion-pair high-performance liquid chromatography (HPLC). Both procedures reveal good linearity in the range 0-80 nmol/ml per min, with an inter-assay coefficient of variation of 6.2% for the colorimetric assay and 4.5% for the HPLC-assisted assay. Recovery values measured for the colorimetric method ranged from 97% to 102% and for the HPLC-assisted method from 98% to 101%. The colorimetric method is suitable for performance in small hospital laboratories since it can be carried out with simple analytical instrumentation. It is obvious nevertheless, that the HPLC-assisted assay reveals better validation criteria. The method is also simple and rapid and it has successfully been used in our laboratory for the last two years.
D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003124:Colorimetry / Q000379:methods*; D006626:Hippurates / Q000032:analysis; D006801:Humans; D009842:Oligopeptides / Q000378:metabolism*; D007703:Peptidyl-Dipeptidase A / Q000097:blood* / Q000378:metabolism; D012015:Reference Standards
D006626:Hippurates; D009842:Oligopeptides; C010980:hippuryl-histidyl-leucine; D007703:Peptidyl-Dipeptidase A; C030514:hippuric acid