Total and pancreatic amylase measured with 2-chloro-4-nitrophenyl-4-O-beta-D-galactopyranosylmaltoside.
BACKGROUND
: Many different methods have been used to assay amylase activity, using nitrophenylated oligosaccharides as substrate; however, the hydrolysis steps in these methods are complex.


METHODS
: 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.


RESULTS
: 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.


CONCLUSIONS
: This novel assay for T-AMY and P-AMY measures both activities stoichiometrically, directly, and easily, and may be suitable for routine procedures.
Morishita
Y
Y
Department of Clinical Laboratory, Nagoya University Hospital, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8560, Japan.
Iinuma
Y
Y
Nakashima
N
N
Majima
K
K
Mizuguchi
K
K
Kawamura
Y
Y
D000681:Amylases / Q000032:analysis* / Q000097:blood / Q000737:chemistry / Q000652:urine; D001826:Body Fluids / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007202:Indicators and Reagents; D007527:Isoenzymes / Q000032:analysis / Q000097:blood / Q000737:chemistry / Q000652:urine; D009844:Oligosaccharides / Q000737:chemistry*; D010179:Pancreas / Q000737:chemistry*; D012680:Sensitivity and Specificity
C411063:2-chloro-4-nitrophenyl-4-O-galactopyranosylmaltoside; D007202:Indicators and Reagents; D007527:Isoenzymes; D009844:Oligosaccharides; D000681:Amylases