title: An enzymatic inosine 5'-monophosphate assay of increased specificity.
abstract: The enzymatic inosine 5'-monophosphate assay described by Grassl [in, Methods of Enzymatic Analysis (H. U. Bergman, ed.), pp. 2168-2171, Academic Press, New York (1974)] is highly nonspecific, as ITP, ATP, ADP, AMP, and adenosine react stoichiometrically. The reactivity with the adenine derivatives is due to the tri- and diphosphatase activity of alkaline phosphatase (AP), coupled with adenosine deaminase (and possibly AMP deaminase) contamination of commercially available preparations of AP, purine-nucleoside phosphorylase, and/or xanthine oxidase. The inclusion of coformycin (0.05 microgram/ml), a potent inhibitor of these deaminases, completely eliminated the cross-reactivity. ITP, however, still reacted stoichiometrically due to the tri- and diphosphatase activity of AP. Meyer and Terjung [Amer. J. Physiol. 237 C111-C118 (1979)] introduced a modification of Grassl's procedure, substituting 5'-nucleotidase for AP. It has been found that this disallows reactivity with ATP, ADP, and ITP but that AMP and adenosine still react completely. Coformycin prevents this cross-reactivity. It is therefore recommended that the assay be carried out with 5'-nucleotidase (instead of AP) and coformycin, in order to achieve a more specific assay, and one more suitable for use with whole tissue extracts.
authors.forename: H T
authors.initials: HT
authors.lastname: Kyriazi
authors.forename: R E
authors.initials: RE
authors.lastname: Basford
mesh_terms: D000244:Adenosine Diphosphate / Q000032:analysis; D000249:Adenosine Monophosphate / Q000032:analysis; D000255:Adenosine Triphosphate / Q000032:analysis; D003070:Coformycin; D007289:Cyclic IMP / Q000032:analysis*; D007292:Inosine Nucleotides / Q000032:analysis*; D013056:Spectrophotometry, Ultraviolet
chemical_list: D007292:Inosine Nucleotides; D003070:Coformycin; D007289:Cyclic IMP; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate