Immobilization of alkaline phosphatase by sorption on Na-sepiolite

Alkaline phosphatase from bovine intestinal mucous membrane was immobilized by sorption on Na-sepiolite in order to obtain an enzyme-clay complex for soil application. Optimum conditions for immobilization were: phosphate buffer 0•1 at pH 7, 2 h of magnetic stirring at 4°C, 0•25 mg ml−1 for the concentration of enzyme and an enzyme/clay ratio of 10 ml g−1. The adsorption isotherm obeyed the Langmuir equation (r2 0•996 at P < 0•001), reaching a maximum amount of enzyme retention. Apparent Km values were 18•32 m for the immobilized enzyme and 2•36 m for the enzyme in aqueous solution; optimum pH (10•5) did not change as a consequence of immobilization; a first-order kinetic was obtained for incubation times of less than 1 h for both enzymes and the immobilized enzyme showed a lower incubation temperature optimum. The immobilized phosphatase showed less stability than the soluble enzyme in terms of storage at 30°C, thermal stability and resistance to proteolytic attack.