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The 5'-NT (EC3水解酶)assay is based on the enzymatic hydrolysis of 5'-monophosphate (5'-IMP) to form inosine which is converted to hypoxanthine by purine nucleoside phosphorylase (PNP). Hypoxanthine is then converted to uric acid and hydrogen peroxide (H₂O₂) by xanthine oxidase (XOD). H₂O₂ is further reacted with N-Ethyl- N-(2-hydroxy-3-sulfopropyl)-3-methylaniline (EHSPT) and 4- aminoantipyrine (4-AA) in the presence of peroxidase (POD) to generate quinone dye which is monitored in a kinetic manner.

reactionenzyme 5'-monophosphate (5'-IMP)to form inosine
The 5'-NT 
 inosine---> hypoxan...
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The 5'-NT (EC3水解酶)assay is based on the enzymatic hydrolysis of 5'-monophosphate (5'-IMP) to form inosine which is converted to hypoxanthine by purine nucleoside phosphorylase (PNP). Hypoxanthine is then converted to uric acid and hydrogen peroxide (H₂O₂) by xanthine oxidase (XOD). H₂O₂ is further reacted with N-Ethyl- N-(2-hydroxy-3-sulfopropyl)-3-methylaniline (EHSPT) and 4- aminoantipyrine (4-AA) in the presence of peroxidase (POD) to generate quinone dye which is monitored in a kinetic manner.

reaction	enzyme
5'-monophosphate (5'-IMP)to form inosine	The 5'-NT
inosine---> hypoxanthine	purine nucleoside phosphorylase (PNP)
hypoxanthine--->uric acid and hydrogen peroxide (H2O2)	xanthine oxidase (XOD)
H2O2     +       Ethyl- N-(2-hydroxy-3-sulfopropyl)-3-methylaniline (EHSPT) and 4- aminoantipyrine (4-AA) 呈色	peroxidase (POD)
PH:  7.4~7.5 (weak basic)    inhibitor :Ni+     activator: Mn2+