Purification of Milk Process and Chemical Reactions Essay

Purification of Milk Process and Chemical Reactions Essay

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In this study, a three step purification of alkaline phosphatase from non-pasteurized milk was reported. It included cream extraction, n-butanol treatment and acetone precipitation. Different parameters like buffer concentration, temperature, pH, substrate concentration, acetone and n-butanol treatment were optimized to maximize the enzyme activity. The enzyme was fruitfully purified upto homogeneity from the milk, with percentage recovery and fold purification of 56.17 and 17.67 respectively. The kinetic parameter were determined to be 0.927 (Km) and 55.86 (Vmax), with specific activity of 11.31 U/mg. Other optimized parameters were estimated as buffer concentration of 0.5 M with pH 9.0, temperature optima at 37ºC, with n-butanol and acetone concentration of 20% (v/v) and 50% (v/v) respectively. This approach provides a simple and effective method for the purification of alkaline phosphatase from non-pasteurized milk.

INTRODUCTION
Alkaline phosphatase (orthophosphoric monoester phosphohydrolase, EC 3.1.3.1) is a non-specific monoesterases that catalyze the hydrolysis of various phosphate esters and anhydrides of phosphoric acid, under alkaline conditions (Junior et al. 2008). They are the non-specific phosphatases which are apparently ubiquitous in nature. They are widely used in recombinant DNA technology (Kopetzki et al. 1994) and DNA sequencing. It is also an important component of enzyme-linked immunosorbent assay (ELISA) based kits. So it’s purification on large scale is required for different commercial and research purposes. The enzymes have been isolated from different sources, which include bacteria, fungi, organs of mammals and invertebrates, but a little have been reported in plants. Commercially available alkaline ...


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...bronchiseptica. Can J Comp Med 48:175–8.
Morton RK (1953a) Alkaline phosphatase of milk. 1. Association of the enzyme with a particulate lipoprotein complex. Biochem J 55:786–95.
Morton RK (1953b) Alkaline phosphatase of milk. 2. Purification of the enzyme. Biochem J 55:795–800.
Neumann H, Wilson KJ, Hauck-Granoth R, Haran-Ghera N (1971) A Comparative Biochemical Study of Alkaline Phosphatases in Normal and Leukemic Mice. Cancer Res 31:1695–1701.
Njoku V, Chikezie P, Kaoje A (2013) Kinetic studies of alkaline phosphatase extracted from rabbit ( Lepus townsendii ) liver. African J Biotechnol 10:3157–3162. doi: 10.4314/ajb.v10i16.
Thompson LMM, Robert A (1974) Factors Affecting the Activity and Stability of Alkaline Phosphatase in a Marine Pseudomonad Factors Affecting the Activity and Stability of Alkaline Phosphatase in a Marine Pseudomonad. J Bacteriol 117:813–818.

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