Morishita Y, Iinuma Y, Nakashima N, Majima K, Mizuguchi K, Kawamura Y
Department of Clinical Laboratory, Nagoya University Hospital, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8560, Japan.
Clin Chem. 2000 Jul;46(7):928-33.
: Many different methods have been used to assay amylase activity, using nitrophenylated oligosaccharides as substrate; however, the hydrolysis steps in these methods are complex.
: We developed a new continuously monitoring assay for amylase activity in biological fluids, using 2-chloro-4-nitrophenyl-4-O-beta-D-galactopyranosylmaltoside (GalG2CNP) as the substrate; this assay was used with anti-human salivary amylase monoclonal antibodies for specific determination of the pancreatic isoenzyme. Amylase converted GalG2CNP into beta-D-galactopyranosylmaltose and 2-chloro-4-nitrophenol, which was measured at 405 nm.
: GalG2CNP was cleaved between 2-chloro-4-nitrophenol and beta-D-galactopyranosylmaltose and did not undergo transfer reactions. The within-assay CVs (n = 20) for total amylase (T-AMY) and pancreatic amylase (P-AMY) were 0.6-1.6% and 0.5-2.5%, respectively; and day-to-day CVs (n = 10) for T-AMY and P-AMY were 0.8-3.7% and 0.6-4.1%, respectively. T-AMY and P-AMY activities in serum or urine obtained by the proposed method correlated well with those determined by the 2-chloro-4-nitrophenyl 4-O-beta-D-galactopyranosyl-beta-maltotetraoside method or the modified IFCC method.
: This novel assay for T-AMY and P-AMY measures both activities stoichiometrically, directly, and easily, and may be suitable for routine procedures.
已使用多种不同方法以硝基苯基化寡糖为底物来测定淀粉酶活性；然而，这些方法中的水解步骤很复杂。
我们开发了一种用于生物体液中淀粉酶活性的新型连续监测测定法，使用2-氯-4-硝基苯基-4-O-β-D-吡喃半乳糖基麦芽糖苷（GalG2CNP）作为底物；该测定法与抗人唾液淀粉酶单克隆抗体一起用于胰腺同工酶的特异性测定。淀粉酶将GalG2CNP转化为β-D-吡喃半乳糖基麦芽糖和2-氯-4-硝基苯酚，在405nm处对其进行测定。
GalG2CNP在2-氯-4-硝基苯酚和β-D-吡喃半乳糖基麦芽糖之间裂解，且未发生转移反应。总淀粉酶（T-AMY）和胰腺淀粉酶（P-AMY）的批内变异系数（n = 20）分别为0.6 - 1.6%和0.5 - 2.5%；T-AMY和P-AMY的日间变异系数（n = 10）分别为0.8 - 3.7%和0.6 - 4.1%。通过所提出的方法获得的血清或尿液中的T-AMY和P-AMY活性与通过2-氯-4-硝基苯基4-O-β-D-吡喃半乳糖基-β-麦芽四糖苷法或改良的国际临床化学和检验医学联合会（IFCC）法测定的活性具有良好的相关性。
这种用于T-AMY和P-AMY的新型测定法能以化学计量的方式直接且轻松地测量两种活性，可能适用于常规检测程序。
