Submitted to: Communications in Soil Science and Plant Analysis
Publication Type: Peer reviewed journal
Publication Acceptance Date: 2/21/2008
Publication Date: 3/1/2009
Citation: He, Z., Waldrip, H., Erich, M., Honeycutt, C.W., Senwo, Z.N. 2009. Enzymatic Quantification of Phytate in Animal Manure. Communications in Soil Science and Plant Analysis. 40:566-575. Interpretive Summary: Phosphorus from soil and animal manures is a major contributor to water quality degradation. Before more effective phosphorus management practices can be developed, analytical techniques are needed for quantifying the specific phosphorus forms. In this work, we report our effort to develop a simple, rapid enzymatic method for routine analysis of phytate, a major organic phosphorus form in animal manure. Our data indicate that the procedure proposed in this work can be used to quantify this phosphorus form in dairy manure, poultry manure, and poultry litter. Additional research is being conducted to verify the effectiveness of this method for general use across a wider range of soils and manures.
Technical Abstract: Phytate (inositol hexaphosphate) has been identified as a major organic P form in soil, animal manure and other environmental samples. Although a number of methods are available for quantitative isolation and determination of phytate, they are time-consuming and not amenable to routine analysis. We developed a simple, rapid method for enzymatic determination of phytate in animal manure. Animal manure was extracted by H2O, 1 M HCl, 0.1 M sodium acetate (NaAc, pH 5.0) with or without 0.05 M EDTA, and 0.25 M or 0.5 M NaOH-0.05 M EDTA. Extracts were diluted (1/10-1/150) and adjusted to pH 5.0 in sodium acetate buffer. The diluted extracts were then incubated at 37 deg C for 1 h in the absence and presence of fungal 3-phytase (PHY) and potato acid phosphatase (PAP). Enzymatic hydrolyzable organic P was calculated as the difference in inorganic P (Pi) between the mixtures with and without enzymes. Our data indicated that enzymatic incubation of properly-diluted and pH-adjusted HCl or NaOH/EDTA extracts released phytate P. The complementary substrate specificity of the two enzymes is considered to enhance the effectiveness of enzymatic hydrolysis. Consequently, we recommend this method of combining PAP and PHY for quantifying phytate P. Additional research is being conducted to verify the effectiveness of this method for general use across a wider range of soils and manures.