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Isolation and identification of the multiple forms of soybean phytases

Hamada, J.S.
Journal of the American Oil Chemists' Society 1996 v.73 no.9 pp. 1143
Glycine max, soybeans, seed germination, phytases, phosphoric monoester hydrolases, isozymes, enzyme activity, enzyme substrates, phytic acid, enzymatic hydrolysis, feed processing, food processing, polyethylene glycol, high performance liquid chromatography, anion exchange resins
Preparative-scale purification of acid phosphatases from soybeans was undertaken. The purified enzymes were subsequently characterized to assess their utility for phytate reduction in processed plant foods. Sequential precipitation with polyethylene glycol (PEG) and KCl followed by preparative anion exchange HPLC was used to isolate 6 acid phosphatases (APases) from germinating soybean seeds. KCl inhibited the precipitation of APases by PEG, thereby enabling individual APases to be isolated directly from crude extracts by controlling PEG and KCl concn. 4 of the APases isolated (numbers 3-6) were phytases. APases 3 and 5 exhibited optimal activity at pH 4.5 and 3.0, respectively, and APases 4 and 6 had a dual optimum pH of 3.0 and 5.5, and 3.5 and 5.8, respectively. Phytase activity was at its peak at 40°C for APase 4 and at 60°C for APases 3, 5 and 6. It is concluded that this isolation method can be used to identify endogenous and/or exogenous phytases that are suitable for phytic acid hydrolysis in foods, especially during processing of oilseeds and cereal-based ingredients.