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Post-ruminal phytate degradation in sheep
Animal Feed Science and Technology 101 (2002) 55–60
Post-ruminal phytate degradation in sheep W.-Y. Park, T. Matsui∗ , H. Yano Division of Applied Biosciences, Graduated School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan Received 7 June 2001; received in revised form 3 June 2002; accepted 3 June 2002
Abstract This experiment was conducted to study phytate (inositol hexaphosphate) degradation in the post-ruminal digestive tract of sheep. Three sheep were fed a diet containing 20% rapeseed meal at intervals of 2 h for 5 days and then digesta were collected from the abomasum, small intestine, upper large intestine and lower large intestine. Levels of phytate and its hydrolysis products (i.e. inositol tri-, tetra- and pentaphosphates) were measured in digesta samples. Approximately 35% of dietary phosphorus in the form of inositol phosphates (IPP) reached the abomasum. Passage of IPP did not differ markedly between the abomasum and the small intestine. The flow of IPP into the lower large intestine was less (P < 0.05) than that into the small intestine but did not differ between the upper and the lower large intestine. Consequently, 12% of dietary IPP was recovered from the lower large intestine. Results suggest that phytate is partly degraded in the large intestine of sheep but the major site of phytate degradation is the forestomach. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Phytate degradation; Digestive tract; Sheep
1. Introduction Oilseed meals contain high levels of phosphorus and more than half exists in the form of phytate (Weremko et al., 1997). Morse et al. (1992) reported that ruminants efficiently utilized phytate phosphorus because phytate was easily degraded in the rumen and phytate was almost completely degraded in the digestive tract of steers (Nelson et al., 1976) and dairy cows (Clark et al., 1986). However, our previous studies suggested that more than 30% of phytate in rapeseed meal was not degraded in the rumen of sheep (Konishi et al., Abbreviations: CP, crude protein; DM, dry matter; IP, inositol phosphate; IPP, phosphorus in the form of inositol phosphates; IP3, inositol triphosphates; IP4, inositol tetraphosphates; IP5, inositol pentaphosphates; IP6, inositol hexaphosphates; LW, live weight ∗ Corresponding author. Tel.: +81-75-753-6056; fax: +81-75-753-6344. E-mail address: [email protected] (T. Matsui). 0377-8401/02/$ – see front matter © 2002 Elsevier Science B.V. All rights reserved. PII: S 0 3 7 7 - 8 4 0 1 ( 0 2 ) 0 0 1 8 1 - 5
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1999; Park et al., 1999) and that 22% of phytate in rapeseed meal reached the duodenum of sheep (Park et al., 2000). The availability of phytate phosphorus in rapeseed meal was 19% in pigs (Weremko et al., 1997), which suggested that phytate was partly degraded in their digestive tract. Some researchers reported that small-intestinal mucosa contained phytase activity and this enzyme effectively hydrolyzed phytate in rats (Lopez et al., 2000) and pigs (Hu et al., 1996). Therefore, phytate escaping ruminal degradation may be hydrolyzed in the small intestine of sheep, and the released inorganic phosphorus may be utilized. The present study was conducted to examine phytate degradation in the post-ruminal digestive tract of sheep. 2. Materials and methods 2.1. Animals and diet Three Corriedale × Suffolk crossbred ewes (55 ± 10.0 kg live weight (LW)) were individually housed in cages with automatic feeders and cared for according to the Guide for the Care and Use of Laboratory Animals (Kyoto University Animal Care Committee). The experimental diet consisted of 50% alfalfa hay and 50% pelleted concentrate (56% corn starch, 4% sucrose, 39% rapeseed meal and 1% Yb-labeled rapeseed meal) on a dry matter (DM) basis. The animals were fed the diet at a level of 1.6% of LW per day in 12 equal meals daily and had free access to water and a mineral block that contained (%): NaCl 97.1; Fe3 O3 ·H2 O 0.174; (FeO2 , FeO3 ) 0.02; CuSO4 0.038; CoSO4 0.007; ZnSO4 0.124; MnCO3 0.105; Ca(IO3 )2 0.008; NaSeO3 0.003; Vitamin E powder 2.421 (2000 IU). 2.2. Sampling procedure The sheep were exsanguinated under pentobarbital anesthesia after a 5 days adjustment period. Digesta were immediately collected from the abomasum, small intestine (i.e. between 3 and 5 m cranial to the ileocecal valve; jejunum), upper large intestine (i.e. the middle of the ascending colon) and lower large intestine (i.e. rectum). Samples were freeze–dried and used for analyses of phytate, its hydrolyzed products and Yb. Alfalfa hay, and pelleted concentrate were also collected for determination of phytate, its hydrolyzed products, phosphorus, calcium, crude protein (CP) and DM. 2.3. Chemical analyses The amounts of inositol phosphates (IPs; i.e. hexa-, penta-, tetra-, and triphosphates, IP6, IP5, IP4 and IP3) were determined in pelleted concentrate, alfalfa hay and digesta samples by ion-pair high performance liquid chromatography (Park et al., 2000). Dietary phosphorus and calcium and Yb concentrations in digesta were measured using an inductively coupled plasma emission spectrometer (ICPS-1000, Shimadzu, Kyoto, Japan) after digestion with concentrated nitric acid and perchloric acid (5:1, v/v). Alfalfa hay and
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pelleted concentrate were analyzed for DM and CP (N × 6.25) using the methods of AOAC (1990): 930.15 and 984.13, respectively. 2.4. Estimation of the flow of IPs into each segment of the digestive tract The daily passage of IPs in each segment of the digestive tract was calculated by the following equation daily flow of IP into each segment of the digestive tract (mol per day) IP concentration in digesta = × daily intake of Yb Yb concentration in digesta Total phosphorus in the form of IPs (IPP) was calculated as the sum of individual IPs multiplied by the number of phosphorus atoms that each IP contained. 2.5. Statistical analysis Statistical analysis was performed using SAS (1990). Data were analyzed as a randomized block design and the model for analysis was Y = µ + αi + βj + εij where Y is the response, µ the overall mean, α i the effect of animal (block), β j the effect of sampling site and εij the error. The difference of IP inflow between each part of the digestive tract was determined by Scheffe’s test. Differences were considered statistically significant if P < 0.05.
3. Results 3.1. Chemical composition of feed The diet contained 16.5% CP, 0.79% calcium and 0.36% phosphorus on a DM basis. Animals consumed all the feed offered and the daily intakes of CP, calcium and phosphorus were sufficient for the requirements of maintenance ewes weighing 50 kg (NRC, 1985). Inositol phosphates were not found in alfalfa hay. The pelleted concentrate contained 55.74 mol/g IPP on a DM basis, and the concentrations of IP6, IP5, IP4 and IP3 were 7.51, 1.60, 0.45 and 0.30 mol/g, respectively. 3.2. Daily flow of IPs into each segment of the digestive tract Inositol hexaphosphate was the major IP in the digesta of each segment (Table 1). Recoveries of dietary IP6 and IPP from the abomasum were 23.8 and 35.3%, respectively. The flow of IP5 into the abomasum was approximately half of the dietary IP5. However, the inflow of IP3 was four-fold more than its intake.
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Table 1 Daily flow of phosphorus in the form of inositol phosphates in the digestive tract of sheep (mol/kg live weight per day)a
Abomasum Small intestine Upper large intestine Lower large intestine S.E.M. Significance value (P)
Hexaphosphateb Pentaphosphateb Tetraphosphateb Triphosphateb (360.3) (63.8) (14.5) (7.2)
Total phosphatec (445.8)
85.9 d 77.5 d 31.1 e 25.2 e 5.7 0.006
157.3 d 140.7 d, e 58.4 e 53.7 e 12.2 0.021
31.0 28.1 12.8 12.7 3.8 0.161
12.1 17.7 8.0 8.7 2.4 0.375
28.3 17.5 6.5 7.1 3.0 0.056
Values in parenthesis are the intake values. a Values with different letters (d, e) in the same column were significantly different, P < 0.05. Values are means for three sheep. b Hexaphosphate, pentaphosphate, tetraphosphate and triphosphate denote phosphorus in the form of inositol hexa-, penta-, tetra- and triphosphate, respectively. c Total phosphorus in the form of inositol phosphates.
The passages of IP6, IP5, IP4 and IPP were not markedly different between the abomasum and the small intestine, although the flow of IP3 tended to decrease in the small intestine. The passage of IP6 in the upper large intestine was less (P < 0.05) than that in the small intestine. The similar tendency was observed in the inflow of IPP and other IPs. As a result, 13.1% of dietary IPP reached the upper large intestine. The inflow of each IP and IPP did not differ between the upper and lower segments of the large intestine.
4. Discussion Rapeseed meal was considered to be the major source of IPs in the diet because IPs were not found in alfalfa hay, which was consistent with Eeckhout and De Paepe (1994). Approximately 35% of dietary IPP reached the abomasum of sheep. This result supported our previous report (Park et al., 1999) showing that 32% of phytate in rapeseed meal escaped ruminal degradation. Inositol hexaphosphate and IP5 flows were decreased in the abomasum but the flow of IP3 was increased, which was supported by our previous results showing decreased passage of IP6 and IP5 until digesta reached the duodenum, while that of IP3 was increased in sheep given rapeseed meal (Park et al., 2000). These results suggested that the production of IP3 from highly phosphorylated IPs exceeded the degradation of IP3 in the forestomach. Phytase produced by ruminal microbes may preferentially degrade highly phosphorylated inositols. The passages of IPP, IP6 and IP5 did not differ markedly between the abomasum and small intestine (jejunum), which suggested that highly phosphorylated inositols were not degraded between the abomasum and the jejunum. On the other hand, the inflow of IP3 tended to be decreased in the small intestine. A recent study indicated that duodenal mucosa showed high phytase activity in rats and this enzyme effectively degraded phytate (Lopez et al., 2000). Hu et al. (1996) observed that phytase isolated from the intestinal mucosa of
W.-Y. Park et al. / Animal Feed Science and Technology 101 (2002) 55–60
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pigs had lower activity toward highly phosphorylated inositols than IP3. Therefore, mucosal phytase may degrade IP3 in the small intestine of sheep. As IP3 was a minor constituent of IP in the gut, the degradation of IP3 did not markedly affect the inflow of IPP. The upper large intestine showed less daily passage of IPP than the small intestine, which indicated that IPs were degraded in the upper large intestine. As a result, 13% of dietary IPP reached the upper large intestine. Sandberg et al. (1993) showed that the amount of phytate in feces was lower than that in the ileal digesta of pigs given rapeseed meal and suggested that most phytate degradation took place in the large intestine. A study in horses also suggested that a large amount of phytate was degraded in the large intestine (Matsui et al., 1999). Wise and Gilburt (1982) reported that phytate was degraded by microbial phytase in the large intestine of rats. Phytate is considered to be partly degraded by the action of microbial phytase in the upper large intestine of sheep. However, the reduction of IPP flow into the abomasum is more marked than that into the large intestine, which suggests that phytate is mainly degraded in the forestomach. The flow of each IP was decreased similarly between the small intestine and the upper large intestine, which suggests that the microbial phytase in the upper large intestine does not selectively degrade IPs. Therefore, the substrate preference of phytase may be different between ruminal and large intestinal microbes. As phosphorus is mainly absorbed in the small intestine of ruminants (Grace et al., 1974; Pfeffer et al., 1970), phytate degradation in the large intestine is unlikely to have any nutritional significance for sheep. Alternatively, Höller et al. (1988) reported that inorganic phosphate was absorbed in the large intestine of sheep when inorganic phosphate was perfused into the large intestine. This report suggested that phosphorus was potentially absorbed from the large intestine of sheep. Degradation of phytate possibly increases the amount of absorbable phosphorus in the large intestine. However, further studies are needed to investigate the absorption of inorganic phosphorus released from phytate in the large intestine. 5. Conclusions Phytate in rapeseed meal was only partly degraded in the large intestine of sheep. Although sheep possibly utilized the released phosphorus from phytate in the large intestine, more than 10% of dietary phosphorus in the form of inositol phosphates was excreted in the feces. This suggests that rapeseed meal contains unavailable phosphorus for sheep and that degradation of phytate in the forestomach principally affects phosphorus availability of rapeseed meal. Acknowledgements This work was partly supported by Livestock Industry’s Environmental Improvement Organization of Japan. References AOAC, 1990. Official Methods of Analysis, 15th ed. Association of Official Analytical Chemists, Arlington, VA. Clark Jr., W.D., Wohlt, J.E., Gilbreath, R.L., Zajac, P.K., 1986. Phytate phosphorus intake and disappearance in the gastrointestinal tract of high producing daily cows. J. Dairy Sci. 69, 3151–3155.
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Eeckhout, W., De Paepe, M., 1994. Total phosphorus, phytate-phosphorus and phytate activity in plant feedstuffs. Anim. Feed Sci. Technol. 47, 19–29. Grace, N.D., Ulyatt, M.J., Macrae, J.C., 1974. Quantitative digestion of fresh herbage by sheep. Part III. The movement of Mg, Ca, P, K and Na in the digestive tract. J. Agric. Sci. 82, 321–330. Höller, H., Figge, A., Richter, F., Breves, G., 1988. Calcium and inorganic phosphate net absorption from the sheep colon and rectum perfused in vivo. J. Anim. Physiol. Anim. Nutr. 59, 9–15. Hu, H.L., Wise, A., Henderson, C., 1996. Hydrolysis of phytate and inositol tri-, tetra-, and pentaphosphates by the intestinal mucosa of the pig. Nutr. Res. 16, 781–787. Konishi, C., Matsui, T., Park, W.Y., Yano, H., Yano, F., 1999. Heat treatment of soybean meal and rapeseed meal suppresses rumen degradation of phytate phosphorus in sheep. Anim. Feed Sci. Technol. 80, 112–115. Lopez, H.W., Vallery, F., Levrat, V.M.A., Coudray, C., Demigne, C., Remesy, C., 2000. Dietary phytic acid and wheat bran enhance mucosal phytase activity in rat small intestine. J. Nutr. 130, 2020–2025. Matsui, T., Murakami, Y., Yano, H., Fujikawa, H., Osawa, T., Asai, Y., 1999. Phytate and phosphorus movements in the digestive tract of horses. Equine Vet. J. 30 (Suppl.), 505–507. Morse, D., Head, H.H., Wilcox, C.J., 1992. Disappearance of phosphorus in phytate from concentrates in vitro and from rations fed to lactating dairy cows. J. Dairy Sci. 75, 1975–1986. National Research Council, 1985. Nutrient Requirements of Sheep, 6th ed. (revised). National Academy Press, Washington, DC. Nelson, T.S., Daniels, L.B., Hall, J.R., Shields, L.G., 1976. Hydrolysis of natural phytate phosphorus in the digestive tract of calves. J. Anim. Sci. 42, 1509–1512. Park, W.Y., Matsui, T., Konishi, C., Kim, S.W., Yano, F., Yano, H., 1999. Formaldehyde treatment suppresses ruminal degradation of phytate in soybean meal and rapeseed meal. Br. J. Nutr. 81, 467–471. Park, W.Y., Matsui, T., Yano, F., Yano, H., 2000. Heat treatment of rapeseed meal increases phytate flow into the duodenum of sheep. Anim. Feed Sci. Technol. 88, 31–37. Pfeffer, E., Thompson, A., Armstrong, D.G., 1970. Studies on intestinal digestion in the sheep. Part 3. Net movement of certain inorganic elements in the digestive tract on rations containing different proportions of hay and rolled barley. Br. J. Nutr. 24, 197–204. Sandberg, A.S., Larsen, T., Sandstrom, B., 1993. High dietary calcium level decreases colonic phytate degradation in pigs fed a rapeseed diet. J. Nutr. 123, 559–566. Statistical Analysis Systems, 1990. SAS User’s Guide. SAS Institute Inc., Cary, NC. Weremko, D., Fandrejewski, H., Zebrowska, T., Han, I.K., Kim, J.H., Cho, W.T., 1997. Bioavailability of phosphorus in feeds of plant origin for pigs (review). Asian Aust. J. Anim. Sci. 10, 551–556. Wise, A., Gilburt, D.J., 1982. Phytate hydrolysis by germ-free and conventional rats. Appl. Environ. Microb. 43, 753–756.
Post-ruminal phytate degradation in sheep W.-Y. Park, T. Matsui∗ , H. Yano Division of Applied Biosciences, Graduated School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan Received 7 June 2001; received in revised form 3 June 2002; accepted 3 June 2002
Abstract This experiment was conducted to study phytate (inositol hexaphosphate) degradation in the post-ruminal digestive tract of sheep. Three sheep were fed a diet containing 20% rapeseed meal at intervals of 2 h for 5 days and then digesta were collected from the abomasum, small intestine, upper large intestine and lower large intestine. Levels of phytate and its hydrolysis products (i.e. inositol tri-, tetra- and pentaphosphates) were measured in digesta samples. Approximately 35% of dietary phosphorus in the form of inositol phosphates (IPP) reached the abomasum. Passage of IPP did not differ markedly between the abomasum and the small intestine. The flow of IPP into the lower large intestine was less (P < 0.05) than that into the small intestine but did not differ between the upper and the lower large intestine. Consequently, 12% of dietary IPP was recovered from the lower large intestine. Results suggest that phytate is partly degraded in the large intestine of sheep but the major site of phytate degradation is the forestomach. © 2002 Elsevier Science B.V. All rights reserved. Keywords: Phytate degradation; Digestive tract; Sheep
1. Introduction Oilseed meals contain high levels of phosphorus and more than half exists in the form of phytate (Weremko et al., 1997). Morse et al. (1992) reported that ruminants efficiently utilized phytate phosphorus because phytate was easily degraded in the rumen and phytate was almost completely degraded in the digestive tract of steers (Nelson et al., 1976) and dairy cows (Clark et al., 1986). However, our previous studies suggested that more than 30% of phytate in rapeseed meal was not degraded in the rumen of sheep (Konishi et al., Abbreviations: CP, crude protein; DM, dry matter; IP, inositol phosphate; IPP, phosphorus in the form of inositol phosphates; IP3, inositol triphosphates; IP4, inositol tetraphosphates; IP5, inositol pentaphosphates; IP6, inositol hexaphosphates; LW, live weight ∗ Corresponding author. Tel.: +81-75-753-6056; fax: +81-75-753-6344. E-mail address: [email protected] (T. Matsui). 0377-8401/02/$ – see front matter © 2002 Elsevier Science B.V. All rights reserved. PII: S 0 3 7 7 - 8 4 0 1 ( 0 2 ) 0 0 1 8 1 - 5
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1999; Park et al., 1999) and that 22% of phytate in rapeseed meal reached the duodenum of sheep (Park et al., 2000). The availability of phytate phosphorus in rapeseed meal was 19% in pigs (Weremko et al., 1997), which suggested that phytate was partly degraded in their digestive tract. Some researchers reported that small-intestinal mucosa contained phytase activity and this enzyme effectively hydrolyzed phytate in rats (Lopez et al., 2000) and pigs (Hu et al., 1996). Therefore, phytate escaping ruminal degradation may be hydrolyzed in the small intestine of sheep, and the released inorganic phosphorus may be utilized. The present study was conducted to examine phytate degradation in the post-ruminal digestive tract of sheep. 2. Materials and methods 2.1. Animals and diet Three Corriedale × Suffolk crossbred ewes (55 ± 10.0 kg live weight (LW)) were individually housed in cages with automatic feeders and cared for according to the Guide for the Care and Use of Laboratory Animals (Kyoto University Animal Care Committee). The experimental diet consisted of 50% alfalfa hay and 50% pelleted concentrate (56% corn starch, 4% sucrose, 39% rapeseed meal and 1% Yb-labeled rapeseed meal) on a dry matter (DM) basis. The animals were fed the diet at a level of 1.6% of LW per day in 12 equal meals daily and had free access to water and a mineral block that contained (%): NaCl 97.1; Fe3 O3 ·H2 O 0.174; (FeO2 , FeO3 ) 0.02; CuSO4 0.038; CoSO4 0.007; ZnSO4 0.124; MnCO3 0.105; Ca(IO3 )2 0.008; NaSeO3 0.003; Vitamin E powder 2.421 (2000 IU). 2.2. Sampling procedure The sheep were exsanguinated under pentobarbital anesthesia after a 5 days adjustment period. Digesta were immediately collected from the abomasum, small intestine (i.e. between 3 and 5 m cranial to the ileocecal valve; jejunum), upper large intestine (i.e. the middle of the ascending colon) and lower large intestine (i.e. rectum). Samples were freeze–dried and used for analyses of phytate, its hydrolyzed products and Yb. Alfalfa hay, and pelleted concentrate were also collected for determination of phytate, its hydrolyzed products, phosphorus, calcium, crude protein (CP) and DM. 2.3. Chemical analyses The amounts of inositol phosphates (IPs; i.e. hexa-, penta-, tetra-, and triphosphates, IP6, IP5, IP4 and IP3) were determined in pelleted concentrate, alfalfa hay and digesta samples by ion-pair high performance liquid chromatography (Park et al., 2000). Dietary phosphorus and calcium and Yb concentrations in digesta were measured using an inductively coupled plasma emission spectrometer (ICPS-1000, Shimadzu, Kyoto, Japan) after digestion with concentrated nitric acid and perchloric acid (5:1, v/v). Alfalfa hay and
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pelleted concentrate were analyzed for DM and CP (N × 6.25) using the methods of AOAC (1990): 930.15 and 984.13, respectively. 2.4. Estimation of the flow of IPs into each segment of the digestive tract The daily passage of IPs in each segment of the digestive tract was calculated by the following equation daily flow of IP into each segment of the digestive tract (mol per day) IP concentration in digesta = × daily intake of Yb Yb concentration in digesta Total phosphorus in the form of IPs (IPP) was calculated as the sum of individual IPs multiplied by the number of phosphorus atoms that each IP contained. 2.5. Statistical analysis Statistical analysis was performed using SAS (1990). Data were analyzed as a randomized block design and the model for analysis was Y = µ + αi + βj + εij where Y is the response, µ the overall mean, α i the effect of animal (block), β j the effect of sampling site and εij the error. The difference of IP inflow between each part of the digestive tract was determined by Scheffe’s test. Differences were considered statistically significant if P < 0.05.
3. Results 3.1. Chemical composition of feed The diet contained 16.5% CP, 0.79% calcium and 0.36% phosphorus on a DM basis. Animals consumed all the feed offered and the daily intakes of CP, calcium and phosphorus were sufficient for the requirements of maintenance ewes weighing 50 kg (NRC, 1985). Inositol phosphates were not found in alfalfa hay. The pelleted concentrate contained 55.74 mol/g IPP on a DM basis, and the concentrations of IP6, IP5, IP4 and IP3 were 7.51, 1.60, 0.45 and 0.30 mol/g, respectively. 3.2. Daily flow of IPs into each segment of the digestive tract Inositol hexaphosphate was the major IP in the digesta of each segment (Table 1). Recoveries of dietary IP6 and IPP from the abomasum were 23.8 and 35.3%, respectively. The flow of IP5 into the abomasum was approximately half of the dietary IP5. However, the inflow of IP3 was four-fold more than its intake.
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Table 1 Daily flow of phosphorus in the form of inositol phosphates in the digestive tract of sheep (mol/kg live weight per day)a
Abomasum Small intestine Upper large intestine Lower large intestine S.E.M. Significance value (P)
Hexaphosphateb Pentaphosphateb Tetraphosphateb Triphosphateb (360.3) (63.8) (14.5) (7.2)
Total phosphatec (445.8)
85.9 d 77.5 d 31.1 e 25.2 e 5.7 0.006
157.3 d 140.7 d, e 58.4 e 53.7 e 12.2 0.021
31.0 28.1 12.8 12.7 3.8 0.161
12.1 17.7 8.0 8.7 2.4 0.375
28.3 17.5 6.5 7.1 3.0 0.056
Values in parenthesis are the intake values. a Values with different letters (d, e) in the same column were significantly different, P < 0.05. Values are means for three sheep. b Hexaphosphate, pentaphosphate, tetraphosphate and triphosphate denote phosphorus in the form of inositol hexa-, penta-, tetra- and triphosphate, respectively. c Total phosphorus in the form of inositol phosphates.
The passages of IP6, IP5, IP4 and IPP were not markedly different between the abomasum and the small intestine, although the flow of IP3 tended to decrease in the small intestine. The passage of IP6 in the upper large intestine was less (P < 0.05) than that in the small intestine. The similar tendency was observed in the inflow of IPP and other IPs. As a result, 13.1% of dietary IPP reached the upper large intestine. The inflow of each IP and IPP did not differ between the upper and lower segments of the large intestine.
4. Discussion Rapeseed meal was considered to be the major source of IPs in the diet because IPs were not found in alfalfa hay, which was consistent with Eeckhout and De Paepe (1994). Approximately 35% of dietary IPP reached the abomasum of sheep. This result supported our previous report (Park et al., 1999) showing that 32% of phytate in rapeseed meal escaped ruminal degradation. Inositol hexaphosphate and IP5 flows were decreased in the abomasum but the flow of IP3 was increased, which was supported by our previous results showing decreased passage of IP6 and IP5 until digesta reached the duodenum, while that of IP3 was increased in sheep given rapeseed meal (Park et al., 2000). These results suggested that the production of IP3 from highly phosphorylated IPs exceeded the degradation of IP3 in the forestomach. Phytase produced by ruminal microbes may preferentially degrade highly phosphorylated inositols. The passages of IPP, IP6 and IP5 did not differ markedly between the abomasum and small intestine (jejunum), which suggested that highly phosphorylated inositols were not degraded between the abomasum and the jejunum. On the other hand, the inflow of IP3 tended to be decreased in the small intestine. A recent study indicated that duodenal mucosa showed high phytase activity in rats and this enzyme effectively degraded phytate (Lopez et al., 2000). Hu et al. (1996) observed that phytase isolated from the intestinal mucosa of
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pigs had lower activity toward highly phosphorylated inositols than IP3. Therefore, mucosal phytase may degrade IP3 in the small intestine of sheep. As IP3 was a minor constituent of IP in the gut, the degradation of IP3 did not markedly affect the inflow of IPP. The upper large intestine showed less daily passage of IPP than the small intestine, which indicated that IPs were degraded in the upper large intestine. As a result, 13% of dietary IPP reached the upper large intestine. Sandberg et al. (1993) showed that the amount of phytate in feces was lower than that in the ileal digesta of pigs given rapeseed meal and suggested that most phytate degradation took place in the large intestine. A study in horses also suggested that a large amount of phytate was degraded in the large intestine (Matsui et al., 1999). Wise and Gilburt (1982) reported that phytate was degraded by microbial phytase in the large intestine of rats. Phytate is considered to be partly degraded by the action of microbial phytase in the upper large intestine of sheep. However, the reduction of IPP flow into the abomasum is more marked than that into the large intestine, which suggests that phytate is mainly degraded in the forestomach. The flow of each IP was decreased similarly between the small intestine and the upper large intestine, which suggests that the microbial phytase in the upper large intestine does not selectively degrade IPs. Therefore, the substrate preference of phytase may be different between ruminal and large intestinal microbes. As phosphorus is mainly absorbed in the small intestine of ruminants (Grace et al., 1974; Pfeffer et al., 1970), phytate degradation in the large intestine is unlikely to have any nutritional significance for sheep. Alternatively, Höller et al. (1988) reported that inorganic phosphate was absorbed in the large intestine of sheep when inorganic phosphate was perfused into the large intestine. This report suggested that phosphorus was potentially absorbed from the large intestine of sheep. Degradation of phytate possibly increases the amount of absorbable phosphorus in the large intestine. However, further studies are needed to investigate the absorption of inorganic phosphorus released from phytate in the large intestine. 5. Conclusions Phytate in rapeseed meal was only partly degraded in the large intestine of sheep. Although sheep possibly utilized the released phosphorus from phytate in the large intestine, more than 10% of dietary phosphorus in the form of inositol phosphates was excreted in the feces. This suggests that rapeseed meal contains unavailable phosphorus for sheep and that degradation of phytate in the forestomach principally affects phosphorus availability of rapeseed meal. Acknowledgements This work was partly supported by Livestock Industry’s Environmental Improvement Organization of Japan. References AOAC, 1990. Official Methods of Analysis, 15th ed. Association of Official Analytical Chemists, Arlington, VA. Clark Jr., W.D., Wohlt, J.E., Gilbreath, R.L., Zajac, P.K., 1986. Phytate phosphorus intake and disappearance in the gastrointestinal tract of high producing daily cows. J. Dairy Sci. 69, 3151–3155.
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