Blood AST, ALT and UREA/BUN Level Analysis

Abstract

AST (aspartate aminotransferase; GOT, glutamate oxalacetate transaminase) and ALT (alanine aminotransferase; GPT, glutamate pyruvate transaminase) are sensitive indicators to monitor the liver function under drugs treatment or with acute viral hepatitis. The elevated AST and ALT values in the blood sample indicate liver damage or injury. The determination of urea is the most widely used for the evaluation of kidney function. This protocol is for the quantitative determination of AST, ALT and UREA/BUN in serum and plasma on Roche automated clinical chemistry analyzers. The principle is shown below:For AST: α-ketoglutarate + L-aspartate L-glutamate + oxaloacetate (AST catalyzes this equilibrium reaction) oxaloacetate + NADH + H+ L-malate + NAD+ (malate dehydrogenase catalyzes this equilibrium reaction) The rate of the photometrically determined NADH decrease is directly proportional to the rate of formation of oxaloacetate and thus the AST activity. The above reactions were carried out at 37 °C and measured at a wavelength of 340 nm.For ALT: α-ketoglutarate + L-alanine L-glutamate + pyruvate (ALT catalyzes this equilibrium reaction) Pyruvate + NADH + H+ L-lactate + NAD+ (lactate dehydrogenase catalyzes this equilibrium reaction) The rate of the photometrically determined NADH decrease is directly proportional to the rate of formation of pyruvate and thus the ALT activity. The above reactions were carried out at 37 °C and measured at a wavelength of 340 nm.For UREA/BUN: Urea + H2O → 2 NH4+ + CO2 (urea is hydrolyzed by urease) α-ketoglutarate + NH4+ + NADH → L-glutamate + NAD+ + H2O (the presence of GLDH yields glutamate and NAD+) The decrease in absorbance due to consumption of NADH is measured kinetically. The above reactions were carried out at 37 °C and measured at a wavelength of 340 nm.