An improved method for a rapid determination of phytase activity in animal feed.

Department of Animal Science, Cornell University, Ithaca, NY 14853, USA.
Journal of Animal Science (Impact Factor: 1.92). 05/2005; 83(5):1062-7.
Source: PubMed

ABSTRACT The current direct colorimetric assay for phytase activity in feeds has interference from high P background and other factors. Our objective was to develop a rapid and reliable spin column method to accurately determine phytase activity in feed ingredients or complete diets. After the feed sample was extracted by stirring in 0.2 M citrate buffer, pH 5.5, for 30 min at room temperature, the oily layer of the supernatant fraction was removed by passing through an acrodisc syringe filter (0.45-microm HT Tuffryn membrane, Gelman Laboratory, Ann Arbor, MI). The filtrate was then loaded onto a spin column (MW cutoff 30,000, Millipore, Bedford, MA) to remove free phosphate before the phytase activity assay. Compared with the direct assay, this new procedure improved both accuracy and reproducibility. When diets contained phytase at 0 to 1,500 U/kg (as fed), the CV for multiple assays of the same samples (n = 6) by the new method ranged from 1 to 6% compared with 28 to 39% by the direct method. A linear relationship was found between the added phytase activity in practical diets and the analyzed activity by the new method (r2 = 0.99; P < 0.01). In conclusion, the spin column method is an improved assay for phytase activity in animal feed, and may be used for quality control of phytase supplementation.

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    ABSTRACT: Phytase producing thirteen microbial strains were identified in poultry soil sample collected at Sathyamangalam, Erode district, Tamilnadu. Among thirteen isolates, two of them were the best of phytases enzyme producer and identified as Aspergillus fumigatus and Aspergillus terreus. Under solid substrate culture condition Aspergillus fumigatus was screened for phytase production on agricultural by-products. The results showed that wheat bran with sugarcane molasses was the best substrate for production of phytase. Phytase purified by non chromatography method, the purified enzyme has an apparent molecular weight 64kDa. Keywords: Phytase, Aspergillus fumigatus and Aspergillus terreus. INTRODUCTION Phytase is widespread in nature, occurring in plants, microorganisms, as well as in some animals. Microbial Phytase activity was most frequently detected in fungi. Phytic acid was discovered as early as in 1872 by Pfeffer and a first note on Phytase can be found in the literature in 1907. Phytase belongs to the group of phosphoric monoester hydrolases; it catalyzes the hydrolysis of myo-inositol hexakisphosphate to inorganic monophosphate and lower phosphoric esters of myo-inositol, or in some cases to free myo-inositol (Vats and Banerjee, 2004 and Howson and Davies, 2005). Shieh and Ware (1968) isolated 30 microorganisms from soil for extracellular Phytase activity. All Phytase producers were filamentous fungi, 28 of them belonged to the genus Aspergillus, one species belonging to Penicillium and on to Mucor. Most of the isolates produced only intracellular Phytase (Ghada et al., 2011). Similarly A. niger strains was best producers of extracellular Phytase while bacterial cultures produced only intracellular enzyme confirmed by (Volfov et al., 1994 and Sabu et al., 2005). Phytase production by yeasts was also studied. Saccharomyces cerevisiaewas found to produce Phytase (Nayini and Markakis, 2008) and further Candida tropicalis, Torulopsis candida, Debaryomycescastelii, Kluyveromycesfragilis and Schwanniomycescastellii were able to hydrolyze Phytate (Lambrechts et al., 1993). S. castellii, which showed the highest extracellular Phytase activity, was chosen for the further study (Lambrechts et al., 1993). There are two main areas where Phytase can be used. The first one is Phytate elimination in feed and food industries, and the second one a preparation of special myo-inositol phosphates as tools for biochemical investigation. The main reason for removal of Phytate from food and feed is its indigestibility for monogastric animals including man (Simell et al., 1989 and Traylor et al., 2001). Undigested Phytate together with part of additional
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