Fluorimetric microplate assay for the determination of extracellular alkaline phosphatase kinetics and inhibition kinetics in activated sludge

  • The microbial enzyme alkaline phosphatase contributes to the removal of organic phosphorus compounds from wastewaters. To cope with regulatory threshold values for permitted maximum phosphor concentrations in treated wastewaters, a high activity of this enzyme in the biological treatment stage, e.g., the activated sludge process, is required. To investigate the reaction dynamics of this enzyme, to analyze substrate selectivities, and to identify potential inhibitors, the determination of enzyme kinetics is necessary. A method based on the application of the synthetic fluorogenic substrate 4-methylumbelliferyl phosphate is proven for soils, but not for activated sludges. Here, we adapt this procedure to the latter. The adapted method offers the additional benefit to determine inhibition kinetics. In contrast to conventional photometric assays, no particle removal, e.g., of sludge pellets, is required enabling the analysis of the whole sludge suspension as well as of specific sludge fractions. The high sensitivity of fluorescence detection allows the selection of a wide substrate concentration range for sound modeling of kinetic functions. - Fluorescence array technique for fast and sensitive analysis of high sample numbers - No need for particle separation – analysis of the whole (diluted) sludge suspension - Simultaneous determination of standard and inhibition kinetics

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Author:Klaus Fischer, Marion Wacht
Parent Title (English):MethodsX
Place of publication:Amsterdam
Document Type:Article
Date of completion:2023/06/20
Date of publication:2023/06/20
Publishing institution:Universität Trier
Contributing corporation:The publication was funded by the Open Access Fund of Universität Trier and the German Research Foundation (DFG)
Release Date:2023/12/12
Tag:Biological wastewater treatment; Enzymatic reactions; Enzyme inhibition; Enzyme kinetics; Organic phosphorus compounds; Phosphor elimination
GND Keyword:Abwasserreinigung; Belebtschlamm; Enzymimmunassay; Enzymkinetik; Phosphatelimination; Phosphororganische Verbindungen
Volume (for the year ...):2023
Issue / no.:Band 11
Number of pages:11
Institutes:Fachbereich 6 / Raum- und Umweltwissenschaften
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 572 Biochemie
Licence (German):License LogoCC BY: Creative-Commons-Lizenz 4.0 International

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