Optimization of adenosine deaminase assay by response surface methodology.
The effect of four variables (adenosine, glutamate dehydrogenase, phosphate buffer, and pH) on the measured catalytic concentration of adenosine deaminase (EC 3.5.4.4) was studied by Response Surface Methodology (RSM). This multivariate methodology offers an empirical approach to the study of enzyme assays and allows to detect the interaction between different variables of the system. Response-surface data showed maximum adenosine deaminase catalytic concentration at pH 7.2, adenosine 20 mmol/l, phosphate buffer 200 mmol/l and glutamate dehydrogenase 850 mu kat/l when pleural fluids were used as samples. Optimum conditions for a material containing purified adenosine deaminase from human erythrocytes differed only slightly from that obtained for the pleural fluid.
Bota
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Departament de Bioquímica i de Biologia Molecular, Universitat Autònoma de Barcelona, Bellaterra, Spain.
Gella
F J
FJ
Canalias
F
F
D000241:Adenosine / Q000378:metabolism; D000243:Adenosine Deaminase / Q000032:analysis* / Q000592:standards; D002021:Buffers; D004912:Erythrocytes / Q000201:enzymology; D005969:Glutamate Dehydrogenase / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007656:Ketoglutaric Acids / Q000378:metabolism; D010994:Pleura / Q000201:enzymology*
D002021:Buffers; D007656:Ketoglutaric Acids; D005969:Glutamate Dehydrogenase; D000243:Adenosine Deaminase; D000241:Adenosine