title: Evaluation of a new alpha-amylase assay using 4.6-ethylidene-(G7)-1-4-nitrophenyl-(G1)-alpha-D-maltoheptaoside as substrate.
abstract: The determination of alpha-amylase activity using an ethylidene-blocked 4-nitrophenyl-maltoheptaoside (EPS) has been evaluated in five laboratories on eight different analysers at 25 degrees C, 30 degrees C and 37 degrees C. The protecting ethylidene group inhibits hydrolysis at the non-reducing end of the substrate molecule by the auxiliary enzyme, alpha-glucosidase. The combined reagent is therefore stable for at least 10 days at 2-8 degrees C. HEPES is used, because the molar absorbance of 4-nitrophenol is independent of temperature in the presence of this buffer. Compared with the method using unprotected substrate 4-nitrophenyl-alpha-D-maltoheptaoside (4NP-G7), the present method is equal or better with respect to the imprecision, linearity and interlaboratory transferability of results in human and control sera. Since the protected and unprotected substrates differ in their turnover rate, the new assay yields activities which differ from those of the 4-nitrophenyl-alpha-D-maltoheptaoside method. Based on the homogeneous results obtained in method comparisons between EPS and 4-nitrophenyl-alpha-D-maltoheptaoside, and in order to maintain the 4-nitrophenyl-alpha-D-maltoheptaoside reference values, a conversion factor was derived to eliminate the above differences: activityEPS x 2.50 = activity4NP-G7. The temperature and instrument independence of this relationship was demonstrated in a total of 720 human sera and plasmas.
authors: Institut für Klinische Chemie und Laboratoriumsmedizin, Katharinenhospital, Stuttgart, FRG.
authors: J D
authors: JD
authors: Kruse-Jarres
authors: C
authors: C
authors: Kaiser
authors: J C
authors: JC
authors: Hafkenscheid
authors: W
authors: W
authors: Hohenwallner
authors: W
authors: W
authors: Stein
authors: J
authors: J
authors: Bohner
authors: G
authors: G
authors: Klein
authors: W
authors: W
authors: Poppe
authors: E
authors: E
authors: Rauscher
mesh_terms: D005960:Glucosides; D006027:Glycosides; D006801:Humans; D007202:Indicators and Reagents; D013379:Substrate Specificity; D000516:alpha-Amylases / Q000097:blood*
chemical_list: D005960:Glucosides; D006027:Glycosides; D007202:Indicators and Reagents; C058250:4,6-ethylidene-4-nitrophenyl maltoheptaoside; D000516:alpha-Amylases