Published online 5 January 2006
Published in J Environ Qual 35:390-393 (2006)
DOI: 10.2134/jeq2005.0036
© 2006 American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America
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SHORT COMMUNICATIONS
Phosphorus Fractions in Manure from Growing Pigs Receiving Diets Containing Micronized Peas and Supplemental Enzymes
D. V. Igea,
O. O. Akinremia,*,
C. M. Nyachotib and
W. Guenterb
a Department of Soil Science
b Department of Animal Science, University of Manitoba, Winnipeg, MB, Canada R3T 2N2
* Corresponding author (akinremi{at}ms.umanitoba.ca)
Received for publication February 1, 2005.
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ABSTRACT
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Different livestock feeds manipulations have been reported to reduce the total P concentration in manure. Information on the influence of these dietary manipulation strategies on the forms of P in manure is, however, limited. This study was, therefore, conducted to investigate the effect of diet manipulation through feed micronization and enzyme supplementation on the forms of P in swine manure. Eight growing pigs were fed four diets: barley–raw pea (BRP), barley–micronized pea (BMP), barley–raw pea with enzyme (BRPE), and barley–micronized pea with enzyme (BMPE) in a 4 x 4 Latin square design. Because we are interested in the effect of enzyme cocktail and pea micronization on manure P, we did not reduce the non-phytate P with enzyme addition in this study. The fecal material and urine were collected and analyzed for total P. Fecal material was fractionated to determine the total P in H2O-, NaHCO3–, NaOH-, and HCl-extractable fractions. The total P in the residual fractions was also determined. About 98% of the total P excreted by the pigs was found in the fecal material. Inclusion of micronized pea in pig diet did not have any significant effect (p > 0.1) on either the total P or the different P fractions in the manure. The labile P (the sum of H2O-P and NaHCO3–P) was significantly reduced (p < 0.05) by the addition of enzyme to swine diets. Pigs fed the BRPE and BMPE had 14 and 18% lower labile P, respectively, compared with pigs fed the BRP. Enzyme addition to pig diets reduced not only the total P in manure, but also the labile P fraction, which is of great environmental concern. Thus, the potential of P loss to runoff and the subsequent eutrophication can be reduced by enzyme addition to pig diets.
Abbreviations: BMP, barley–micronized pea • BMPE, barley–micronized pea with enzyme • BRP, barley–raw pea • BRPE, barley–raw pea with enzyme
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INTRODUCTION
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FEEDING DIETS supplemented with phytase and/or containing low phytic acid grains to swine has been reported to decrease the P concentration in manure (Veum et al., 2002; Thacker et al., 2003). Yi et al. (1996) reported a 25 to 50% reduction in fecal P excretion by swine following diet modification with phytase and reduction of supplementary P addition to the diet. A reduction in P content of manure is good for the environment as this will reduce P loading when manure is applied to the soil.
Another dietary manipulation that has been reported to increase P utilization efficiency is micronization. Micronization disrupts the cell wall component of grains, thus increasing the dry matter and nutrient digestibility in swine (Lawrence, 1973) and poultry (Igbasan and Guenter, 1997). Zhang et al. (2003) reported a 17% reduction in the total P content of manure from swine fed micronized pea diets.
Sequential fractionation schemes have been used to separate manure P into different fractions based on their reactivity and solubility (Dou et al., 2000; Ajiboye et al., 2004). This has been used to compare different organic amendments regarding the lability of their phosphorus. For example, Ajiboye et al. (2004) reported that 60% of hog manure P was labile while only 24% of biosolids P was in the labile fraction. There is a general consensus that the reactivity and the risk of manure P to the environment can be assessed by the forms of phosphorus in manure (Ajiboye et al., 2004; Maguire et al., 2004; Wienhold and Miller, 2004). Efforts have been made in recent times to characterize the forms of P extracted by the different extracting agents used in sequential extraction of manure P. Turner and Leytem (2004) reported that while water and NaHCO3 extracted, primarily, inorganic phosphate (orthophosphate) with a small amount of soluble organic P compounds from swine manure, NaOH and HCl extracted appreciable amounts of less soluble organic P. A similar observation was made by Maguire et al. (2004) and Toor et al. (2005).
While diet manipulation clearly has the potential of reducing the total P in manure, thus producing the environmental benefit of reducing soil P load, the question as to how this will affect the solubility and reactivity of manure P in soil has not been fully addressed. Furthermore, there is no consensus on the effect of diet manipulation on the composition of P in manure. Gilley et al. (2001) and Applegate et al. (2003) reported that enzyme addition to animal diets did not have any significant effect on manure P fractions. Maguire et al. (2003), on the other hand, reported a significant decrease in the water-soluble P fraction following phytase addition to and reduction of non-phytate P in turkey diets. The reduction of P content of manure following dietary manipulation such as the use of enzymes and micronization is good for the environment, but this benefit may be lost if the manure P resulting from these diets is more labile and more susceptible to loss in the environment. There is, therefore, a need for a better understanding of the effect of diet manipulation, such as micronization and the use of enzymes, on the solubility and lability of manure P. The objective of this study was, therefore, to evaluate the effect of diet manipulation, through feed micronization and enzyme supplementation, on the forms of P in swine manure.
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MATERIALS AND METHODS
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Eight growing Cotswold barrows were used to investigate the effect of including micronized peas with or without enzyme supplementation in growing pigs' diets on manure P composition. Pigs were obtained from the University of Manitoba's Glenlea Swine Research Farm (Winnipeg, MB, Canada) and housed in individual metabolism crates with smooth transparent walls and tenderfoot flooring. The experimental diets were: barley–raw pea based diet (BRP; control), barley–micronized pea based diet (BMP), barley–raw pea with enzyme blend (BRPE), and barley–micronized pea with enzyme blend (BMPE). The enzyme blend used was a multi-enzyme blend that supplied 500 U of glucanase and 300 U phytase kg–1 of diet (supplied by Canadian Biosystem, Calgary, AB) plus a broad spectrum of other enzymes such as protease, amylase, cellulase, and pectinase. Micronized peas were prepared as described by Zhang et al. (2002) by tempering peas at 25% moisture level using a Monarch Mixer (Machinery Co. Ltd., Winnipeg).
The experiment was designed and conducted according to a 4 x 4 Latin square design in which each of the four diets was randomly assigned to four pigs each time in two blocks until all the pigs in a block had each of the diets, giving eight observations for each experimental diet. The total (4.3 g kg–1), phytate (2.4 g kg–1), and non-phytate (1.9 g kg–1) P were the same for all diets and were set to meet or exceed the recommendation of the National Research Council (1998). As we are interested in the effect of enzyme cocktail and pea micronization on manure P, we did not reduce the non-phytate P with enzyme addition.
The pigs were allowed to adjust to their respective experimental diets for 5 d before feces and urine collection over a 3-d period. Urine was collected into plastic pails containing 50 mL of 5% H2SO4. After measuring the daily volume, 100-mL aliquots were taken for each 24-h period and kept frozen until required for analysis. Feces were collected frequently and stored in plastic zip-lock bags. After the 24-h collection, the feces were weighed and stored at –20°C until required for analysis.
All analyses were performed in duplicate. Fecal samples were freeze-dried and, along with diet samples, ground to pass through a 1-mm screen, then thoroughly mixed before taking samples for analysis. Total phosphorus content in feces and diets were determined according to the method of AOAC International (1990). A 4.4-mL portion of sulfuric acid–hydrogen peroxide digestion mixture was added to 0.4 g of manure or diet in a Kjeldahl digestion tube and the mixture was digested in a digestion block for 3 h at 350°C. Phosphorus was determined in the digest using the molybdate blue method (Murphy and Riley, 1962). Phytate P in the diet was determined by the method described by Haug and Lantzsch (1983). Ten milliliters of 0.2 M HCl was added to 100 mg of diet and the mixture shaken for 3 h at room temperature and then filtered. Then, 0.5 mL of distilled water and 2 mL ferric solution were added to 0.5 mL filtrate and the mixture was boiled for 30 min. The resulting solution was centrifuged at 2400 x g, and 1.5 mL 2 2' bipyridine solution was added to a 1-mL aliquot of the supernatant. The absorbance of the mixture was read against distilled water at 519 nm using a Pharmacia Ultrospec 2000 spectrophotometer. The non-phytate P was determined by the difference between the total and phytate P.
The sequential extraction scheme described by Ajiboye et al. (2004) was employed for the fractionation of P into H2O-, NaHCO3–, NaOH-, and HCl-extractable P, and residual P. A 0.3-g portion of dried manure was sequentially extracted with 30 mL of deionized H2O, 0.5 M NaHCO3, 0.1 M NaOH, and 1M HCl solutions. The total P in the extract was determined by the method of Akinremi et al. (2003) by adding 1.1 mL of sulfuric acid–hydrogen peroxide acid digestion mixture to an aliquot of the extract and digesting the mixture in a digestion block at 350°C for 1 h. The P in the digest was determined using the molybdate blue method (Murphy and Riley, 1962). The residue left after all extractions was also digested using the wet oxidation method of Akinremi et al. (2003) and the P determined by the molybdate blue method (Murphy and Riley, 1962). Urine phosphorus content was determined at the Veterinary Services Branch of Manitoba Agriculture and Food using inductively coupled plasma–atomic emission spectroscopy (ICP–AES). Data were analyzed using the General Linear Models procedure (SAS Institute, 2000).
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RESULTS AND DISCUSSION
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The total P in the fecal material of pigs fed the enzyme-supplemented diet, BRPE (18 g kg–1) or BMPE (17.3 g kg–1), was significantly lower (p 
0.05) than that from pigs fed without enzyme supplementation, that is, either the BRP (20.2 g kg–1) or BMP (20.5 g kg–1), showing that enzyme supplementation reduced the total P in swine feces (Table 1). This was similar to the results reported by Baxter et al. (2003). While enzyme supplementation reduced fecal P content, micronization of peas did not influence the P content of the feces in this study as the fecal P excreted by pigs fed the BRPE and BMPE were not statistically different. This result differs from that of Zhang et al. (2003) who reported that pigs fed diets similar to ours, the BRP and the BMP diets, had fecal P contents that were significantly different.
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Table 1. Phosphorus concentrations in feces and urine from growing pigs fed a barley-based diet containing raw or micronized peas with or without enzyme supplementation.
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While the fecal P was reduced by enzyme supplementation, the urine P was increased by enzyme supplementation with urine from pigs fed the enzyme-supplemented diets (either BRPE or BMPE) having significantly higher urine P than those fed diets without enzyme supplementation (BRP or BMP; Table 1). This result is consistent with that of Zhang et al. (2003). The percent P in urine was, however, much lower than that of the feces. The mean urine P for all the pigs was 0.3 g kg–1 while the mean fecal P was 19 g kg–1. MacLean et al. (1983) reported that about 90% of P excreted by swine was in the feces. Inclusion of micronized peas in the pig diet did not influence the urine P content. The lower fecal P and higher urine P observed in pigs receiving enzyme-supplemented diets suggested that enzyme addition to pig diets aided P digestion and hydrolysis of phytate P in pigs, which resulted in greater absorption of P in the small intestine. However, excess P absorbed was eliminated through the kidney in the urine (Zhang et al., 2003). This justifies the recommendation of Maguire et al. (2004) that phytase addition to poultry diets should be accompanied by adequate reduction in non-phytate P to avoid excessive P absorption.
A higher N to P ratio was observed in manure from pigs fed the enzyme-supplemented diet (N data not shown). This is desirable due to the reduced P load on soil when the manure is applied to soil, since manure is often applied to meet the N requirement of the crops. Thus, when manure from pigs receiving modified diets is applied on an N basis, a higher N to P ratio would result in a 10 to 14% reduction in applied P.
While the addition of enzyme resulted in a 14% reduction in manure P from pigs fed the micronized pea diet, only a 10% reduction was observed from pigs fed the enzyme-supplemented raw pea diet. Although micronization did not have a significant effect on the total P in feces or urine (Table 1), there appears to be a synergistic effect of enzyme supplementation and pea micronization on the reduction of P content of manure.
Forms of Phosphorus in Swine Feces
No significant difference (p 
0.10) was observed in the amount of water-extractable P in the fecal material of the pigs fed the various experimental diets (Fig. 1
). Enzyme supplementation did not have a significant effect on water-extractable P, although a 6% decrease in P concentration of this fraction was observed in the feces of pigs fed enzyme-supplemented diets.
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Fig. 1. Concentrations of phosphorus extracted by deionized water (H2O-P), 0.5 M NaHCO3 (pH 8.5, NaHCO3–P), 0.1 M NaOH (NaOH-P), and 1 M HCl (HCl-P) and residual P fraction in pig manure as influenced by diet manipulation. Columns indicated with the same letter in the same group were not significantly different from one another (p > 0.10). BMP, barley–micronized pea; BMPE, barley–micronized pea with enzyme; BRP, barley–raw pea; BRPE, barley–raw pea with enzyme.
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Enzyme supplementation reduced the NaHCO3–P fraction in fecal material as pigs fed the enzyme-supplemented diets (BRPE and BMPE) had significantly lower (p 0.05) NaHCO3–P than pigs fed the diets with no enzyme supplement (BRP and BMP). The H2O- and NaHCO3–extractable P fractions can be considered as the labile fraction of P in manure, as such, any treatment that reduces these fractions is desirable, since the labile fraction is vulnerable to loss by runoff and leaching (Sharpley and Moyer, 2000). Maguire et al. (2004) and Turner and Leytem (2004) reported that H2O and NaHCO3 extracted mainly the orthophosphate P in manure. The labile P fraction accounted for 84% of the total P in the manure with a mean of 18 143 mg kg–1 P in manure from BRP and BMP diets and a mean value of 15 233 mg kg–1 P in manure from enzyme-supplemented diets. Qian and Schoenau (2000) reported that 70% of the total P in hog manure was present in the labile form. Ajiboye et al. (2004) also reported that P in hog manure was mainly in labile forms. The labile P in manure was significantly reduced by diet manipulation. Enzyme addition resulted in a 14 and 18% reduction in labile P from pigs fed the raw and micronized peas, respectively. Micronization did not influence the amount of labile P in the manure as no significant difference existed between labile P fraction in manure from pigs fed the raw or micronized diet with and without enzyme supplementation. The lower labile P in manure from pigs fed the BRPE or BMPE further affirms the fact that enzyme addition to diets enhanced P retention in growing pigs.
The difference observed in the residual- and NaOH-P in manure from pigs fed the four experimental diets was not significant (p 0.10), which implied that diet manipulation did not influence these fractions of manure P. However, these fractions may not be of great agronomic or environmental significance based on their relative size (14% of total P) and their recalcitrant nature (Ajiboye et al., 2004).
When expressed as a percentage of total P, the distribution of P in the various extracts was in the order H2O-P > NaHCO3–P > NaOH-P > HCl-P > residual P (Fig. 2
). Wienhold and Miller (2004) reported P distribution in swine manure in the order H2O-P > HCl-P > NaHCO3–P > NaOH-P > residual P, while Ajiboye et al. (2004) reported P distribution in the order H2O-P > NaHCO3–P > HCl-P > NaOH-P > residual P. The differences in the P distribution as reported by different authors may be due to pig diets, especially the level of mineral supplementation. While Wienhold and Miller (2004) used a corn–soybean-based diet with a total P of 2.9 to 3.1 g kg–1, a barley–soybean–pea-based diet was employed in the present study with a total P of 4.3 g kg–1. The phytate P was, however, the same in both studies, although while Wienhold and Miller (2004) used phytase enzyme, we used a cocktail of enzymes.
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Fig. 2. Percentage of the total phosphorus extracted by deionized water (H2O-P), 0.5 M NaHCO3 (pH 8.5, NaHCO3–P), 0.1 M NaOH (NaOH-P), and 1 M HCl (HCl-P) and the residual P fraction in pig manure as influenced by diet manipulation. BMP, barley–micronized pea; BMPE, barley–micronized pea with enzyme; BRP, barley–raw pea; BRPE, barley–raw pea with enzyme.
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Including micronized peas in a growing pig diet did not have any significant effect on the manure P content. Supplementation of pig diets with enzyme reduced the total and labile P (which is the fraction of greatest concern) in manure. Thus, the potential of P loss to runoff and the subsequent eutrophication can be reduced by diet manipulation in pigs.
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ABSTRACT
INTRODUCTION
