Enzymatic Characterization of Organic Phosphorus in Animal Manure

Abstract

Information on the forms of P present in animal manure may improve our ability to manage manure P. In most investigations of manure P composition, only inorganic and total P are determined, and the difference between them is assigned as organic P. In this study, we explored the possibility of identifying and quantifying more specific organic P forms in animal manure with orthophosphate-releasing enzymes. Pig (Sus scrofa) manure and cattle (Bos taurus) manure were first sequentially fractionated into water-soluble P, NaHCO3-soluble P, NaOH-soluble P, HCl-soluble P, and residual P. The fractions were separately incubated with wheat phytase, alkaline phosphatase, nuclease P1, nucleotide pyrophosphatase, or their combinations. The released orthophosphate was determined by a molybdate blue method. Part of the organic P in those fractions could be identified by the enzymatic treatments as phytate (i.e., 39% for pig manure and 17% for cattle manure in water-soluble organic P), simple phosphomonoesters (i.e., 43% for pig manure and 15% for cattle manure in NaOH-soluble organic P), nucleotide-like phosphodiesters (2-12%), and nucleotide pyrophosphate (0-4%). Our data indicate that the enzymatic treatment is an effective approach to identify and quantify the organic P forms present in animal manures.

Phosphorus Compounds in Sequential Extracts of Animal Manures: Chemical Speciation and a Novel Fractionation Procedure

“…38, NO. 22,2004 is immobile in soils, because it sorbs strongly to clays and reacts with metals to form insoluble precipitates (7,8). It is also difficult for organisms to access phytic acid once it is stabilized in soil (7).…”

“…In addition, there is little information on phosphorus compounds in sequential extracts, so it is impossible to assign environmental significance to the extracted fractions. He and Honeycutt (22) used phosphatase enzymes to determine functional classes of hydrolyzable phosphorus in sequential fractions of swine and dairy manures, including simple phosphate monoesters, phytatelike phosphorus, and nucleotide-like phosphorus. However, a large proportion of the organic phosphorus was not identified, and no information was obtained on organic phosphorus in HC1 extracts.…”

Soil phosphorus fractions in sandy soils amended with cattle manure for long periods

“…Data on the effects of animal residue application on soil P forms come mainly from countries where meat production is based on confined animals (e.g. poultry and pig production, ruminant feedlots) and where significant amounts of this type of residue are applied to crops in liquid (Hountin et al, 2000), semisolid (He & Honeycutt, 2001) and solid form (Hao et al, 2008).…”

A Modified Molybdenum Blue Method for Orthophosphate Determination Suitable for Investigating Enzymatic Hydrolysis of Organic Phosphates

“…Phosphatase hydrolysis has emerged as a useful tool to characterize soluble P o from various sources (He and Honeycutt 2001;He, Honeycutt, and Griffin 2003;Pant and Warman 2000;Shand and Smith 1997;Turner, McKelvie, and Haygarth 2002). In these studies, enzymatically hydrolyzable P o in a given sample is calculated as the difference between P i contents determined in the presence and absence of enzyme(s).…”

“…Thus, reducing chemical hydrolysis of labile P o that occurs during P i determination will make a significant contribution to accuracy of enzymatic characterization. Another concern is interference by protein precipitates under the acidic conditions for P i determination (He and Honeycutt 2001;Shand and Smith 1997;Pant, Vaughan, and Edwards 1994). One way to reduce protein interference is to keep the enzyme concentrations low (He and Honeycutt 2001;Turner, McKelvie, and Haygarth 2002).…”