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The apparent digestibility of phytate phosphorus and the influence of supplemental phytase in horses^1,2

Department of Nutrition, Faculty of Veterinary Medicine, Utrecht University, 3584 CM Utrecht, The Netherlands

Abstract

Availability of phytate-bound P as influenced by supplemental phytase was studied in eight horses consuming four diets in a 4 x 4 Latin square design experiment. The treatments were a control (containing a low P level, 18.4 g/d) and three high-P diets. These diets contained P as monocalcium phosphate (MCP; 43.7 g/d), myoinositol hexakisphosphate in the form of wheat and rice bran (MIHP; 41.8 g/d), or MIHP with microbial phytase (MIHPP; 42.5 g/d). The proportions of phytate-bound P were 3, 1, 55, and 56% for the control, MCP, MIHP, and MIHPP, respectively. The MIHPP diet was supplemented with 300 phytase units (FTU)/kg (as-fed basis). Feces and urine were collected quantitatively and analyzed for P, Ca, and Mg. Urinary P excretion was lower (P < 0.05) with the control diet (0 g of P/d) than with the MCP diet (1.0 g of P/d). The low urinary P excretion (0.3 g of P/d) for the MIHP diet suggested low P availability compared with the MCP diet, but apparent digestibility of P expressed as a percentage of intake did not differ (P = 0.065) between these diets. Apparent Ca digestibility was lower (P < 0.05) for the MIHP diet than for the MCP diet (26.4 vs. 42.4%). This difference may have been caused by the origin of the Ca in these diets. Phytase supplementation increased apparent Ca digestibility from 26.4 to 31.5% (P < 0.05). Magnesium was not influenced by the level of phytate in the diet. Our data indicate that phytase supplementation had more influence on Ca digestibility than on P digestibility and suggest that phytase supplementation may be beneficial for improving Ca digestibility for horses receiving a phytate-rich diet.

Key Words: Calcium • Horses • Magnesium • Phytase • Phytate • Phosphorus

Introduction

Phytate, the salt of phytic acid (myoinositol hexakisphosphate or IP6), has very little P availability for monogastric animals. In contrast to ruminants, monogastric animals only have a negligible activity of the digestive enzyme phytase before the site of P absorption (Pallauf and Rimbach, 1997). The ruminal microorganisms produce phytase so that P is released from dietary phytate and absorbed in the small intestine (Kemme, 1998). Moreover, phytate decreases the bioavailability of minerals and trace elements in nonruminants due to its molecular structure. Negative charges in phytate bind positively charged cations into a stable complex (Pallauf and Rimbach, 1997). Dietary proteins and AA can also form complexes with phytate (Pallauf and Rimbach, 1997; Kemme, 1998).

Microbial phytase supplemention to the diet of nonruminant animals increases P availability from phytate and enhances the absorption of cations, proteins, and AA. There are only a few studies concerning phytate-P digestibility and the effect of phytase in horses. In two studies (Schryver et al., 1971b; Matsui et al., 1999), it was concluded that P from sodium phytate is available to horses. However, Matsui et al. (1999) used a marker to estimate digestibility in several parts of the digestive tract and observed a negative total-tract digestibility of P. Moreover, there is evidence that P from sodium phytate is more efficiently absorbed than is P from phytate in natural feed ingredients such as wheat bran (Hintz et al., 1973). The objective of our study was to measure the digestibility of phytate-P and the influence of supplemental phytase on apparent P digestibility in horses. Knowledge about the digestibility of phytate-P in natural feed ingredients and the effect of phytase supplementation may markedly affect the selection of feedstuffs to formulate concentrates, and it may decrease the excretion of P into the environment.

Materials and Methods

Animals and Diets

Eight mature trotters (BW = 476 ± 62 kg, geldings, age 6 ± 1 yr) were assigned to four treatments in a 4 x 4 Latin square design. Therefore, two horses were subjected to each sequence of the treatments at the same time. The horses were housed in individual tie stalls. They were checked for normal urinary function before the start of the experiment.

In the present study, we fed diets (Table 1) that had the same level of total P, but which contained either monocalcium phosphate (MCP), phytate without phytase (MIHP), or phytate with phytase (MIHPP). The MCP diet vs. the diet low in P (control) can be considered to be the positive control because the P from monocalcium phosphate is readily available and its ingestion should raise urinary P excretion. In addition to urinary P excretion, the apparent digestibility of P was measured for the four treatments.

Table 2 shows the analyzed macronutrient composition of the concentrates and hay. The analyzed P, phytate, Ca, and Mg content (g/kg), and phytase activity (FTU) of the semipurified concentrates and hay are shown in Table 3. The true phytase activity of the MIHPP diet seemed to be lower than intended.

Each experimental period consisted of 21 d for adaptation and 7 d for sample collection. During the first 3 d of each adaptation period, the horses were gradually transferred from the current to the next concentrate diet. The horses were bedded on straw during the first two weeks of each feeding period and on rubber mats during the last 2 wk of the experimental period. Tap water was available during adaptation periods, but the horses were supplied manually with water during the collection period to measure daily water intake. During the experiment, the horses were given a daily 1-h walk per day on a tredmill, but during the collection period, they were walked manually for 10 min.

Before the experiment, the hay and alfalfa meal, grass meal, wheat bran, and rice bran were analyzed for DM, P, Ca, and CP contents. Contents of other nutrients in the raw materials used to formulate the experimental diets were calculated on the basis of the Dutch Feed Tables (Centraal Veevoederbureau, 1999). The differences in CP and N-free Extract of the control and MCP vs. the MIHP and MIHPP concentrates were kept as small as possible, taking into account the choice of raw materials.

The experimental protocol was approved by the animal experiments committee of the Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.

Urine and Feces Collection

During the collection periods urine (5 d) and feces (7 d) were collected quantitatively. Urine was collected by the use of urine quivers (Genap B.V., s’-Heerenberg, The Netherlands), designed according to Tasker et al. (1965). Before the experiment, all horses were accustomed to the urine quivers. At 0900, urine samples were taken from the total urine collected. Daily collected feces were frozen until further analysis. By using this method, fecal or urine samples could be easily excluded for those days on which the accuracy of collection was not adequate. One horse showed signs of colic on d 6 of the third period, and another horse on d 7. The feces collected on these days were excluded from the week sample. In case of improper urine collection, the samples were replaced by those taken on d 6 or 7 of the collection period.

Chemical Analyses

Macronutrient contents of the concentrates and hay were determined according to methods described earlier by Schonewille et al. (1999). The NDF and ADF were determined according to the procedures of Goering and Van Soest (1970). Before analysis, feces were pooled per horse, per collection period; defrosted; and mixed in a concrete mixer. Two homogenous 1-kg samples were obtained per horse per collection period, which were analyzed separately. The samples were dried at 60°C for 72 h, ground and stored until analysis. Before analysis, samples were dried overnight at 100°C, and subsequently ashed at 500°C for 6 h. Unfortunately, one DM value was not determined. The sample concerned was given the mean value of the DM values determined for the other collection periods for the corresponding horse. Diet samples were ground, dried overnight at 100°C, and ashed at 500°C for 6 h. Phosphorus in feed, feces, and water was analyzed by stannous chloride-hydrazine reduction of phosphomolybdic acid (vanadate yellow method; Quinlan and DeSesa, 1955). Phosphorus in urine was analyzed by the molybdenum blue method (Quinlan and DeSesa, 1955). Calcium and Mg in feed, feces, urine, and water were measured using atomic absorption spectroscopy (Perkin Elmer 3110 PC, Norwalk, CT). Urine samples were acidified directly after urine sampling with 6 M HCl to prevent the complex forming of Mg and P, which has also been described previously (Quinlan and DeSesa, 1955). The acidification of the urine was such that the samples contained 90% urine and 10% acid (6 M HCl). Diet samples were further analyzed for phytic acid (Bos et al., 1991) and for phytase activity (Engelen et al., 1994). The amount of P bound in phytic acid was expressed as P from phytate (phytate-P) and used the fact that phytic acid contains 28.2% P (Kemme, 1998).

Calculations and Statistical Methods

Average BW, urine production, DMI, water intake, and fecal excretion were calculated. Apparent digestibility (%) of P, Ca, Mg, and DM were calculated as follows:

Urine P excretion was used as index for P absorption. Urinary mineral excretion was expressed as percentage of mineral intake. The percentage of phytate-P in the total diet was calculated as follows:

Genstat Release 4.2 for PC (Numerical Algorithms Group, Ltd., Wilkinson House, Oxford, U.K.) was used for all statistical analyses. Besides ANOVA, the R-pair procedure was used to detect differences between treatments. The level of statistical significance was preset at P < 0.05.

Results

Body Weight, Feed and Water Intake, and Dry Matter Digestibility

There were no differences in BW, water intake and urine production between treatments (Table 4). The apparent DM digestibility differed between the control and MCP diet vs. the MIHP and MIHPP diet.

Table 4 shows small differences in P intake between the MCP, MIHP, and MIHPP diets. Compared with the control diet the MCP diet resulted in a higher urinary P excretion (expressed in g/d and as percentage of intake) and P retention, indicating that urinary P excretion and P retention are related to P intake. The apparent P digestibility (%) was lower for the control diet, but there were no differences (P < 0.05) between the MCP, MIHP, and MIHPP diets. The percentage of urinary P excreted ranged from 0.2% (control) to 2.3% (MCP). The percentage of urinary P excreted on the MCP diet was higher than the percentage measured on the MIHP diet, suggesting a low digestibility of P from phytate. The apparent P digestibility (%) on the MIHP diet tended (P = 0.065) to be lower than that on the MCP diet. The P retention on the MIHP diet was not lower than that measured on the MCP diet. No differences were observed as to fecal P excretion and urinary P excretion on the MIHP and MIHPP diets, but urinary P excretion on the MIHPP diet was intermediate when compared with the MCP and MIHP diets.

Calcium Digestibility and Urinary Excretion

Table 4 shows that despite a somewhat lower Ca intake on the MCP diet, there was a higher fecal Ca excretion, and thus a lower apparent Ca digestibility, compared with the control diet. Compared with the control diet, urinary Ca excretion was also lower on the MCP diet (32.0 vs. 25.5 g), indicating a depressing effect of high P intake on Ca digestibility. Calcium balances for the control and MCP diet were positive.

Compared with the MCP diet, fecal Ca excretion on the MIHP diet (39.2 vs. 51.0 g) was much higher, resulting in a lower apparent Ca digestibility. Urinary Ca excretion on the MIHP diet was also lower than urinary Ca excretion on the MCP diet. The Ca balance was higher on the MCP diet compared with the MIHP diet (3.3 vs. –0.2 g).

Phytase supplementation decreased fecal Ca excretion and increased apparent Ca digestibility. Further, urinary Ca excretion for the MIHPP vs. MIHP diet was slightly increased and the horses had a positive Ca balance on the MIHPP diet.

Magnesium Digestibility and Urinary Excretion

There were differences (Table 4) in Mg intake between the control and MCP diet, which resulted in differences in fecal excretion for these diets; however, when compared with the MCP diet, the apparent digestibility of Mg was higher on the control diet. The apparent Mg digestibility pointed at a lower amount of Mg absorbed when the horses were fed on the MCP diet compared with the control diet, even though the MCP diet had a higher Mg content. Compared with the control diet, urinary Mg excretion was lower (P < 0.05) with the MCP diet. The horses retained less Mg when fed on the control diet compared with the other diets.

Magnesium intake on the MIHP and MIHPP diets was higher than on the MCP diet, which resulted in a higher (P < 0.05) fecal excretion of Mg and lower (P < 0.05) apparent Mg absorption, but not in differences in urinary Mg excretion. There were no differences between Mg variables for the MIHP and MIHPP diet.

Discussion

The DM digestibility of the rations was low because they contained a high percentage of hay (60% of total DMI). To realize a control ration with a low P level, a low-quality hay was used so that a low DM digestibility could be expected. The differences in apparent DM digestibility between diets with and without phytate can be explained by the differences in ingredients. Water intake and urine production did not differ among dietary treatments.

The MCP diet resulted in a higher urinary P excretion when compared with the control diet, which is consistent with earlier reports (Schryver et al., 1971a,b; Caple et al., 1982; Meyer, 1990; Buchholz-Bryant et al., 2001). Most of these reports measured urinary P excretion quantitatively except for Caple et al. (1982), who expressed the urine concentration of P as the ratio of the concentration of total urine solutes (µmole/mosmole) and as creatinine clearance ratios (P:%Cr). In this study, the MIHP diet resulted in a lower urinary P excretion (g/d) than the MCP diet, but urinary P excretion was not different from that for the control or MIHPP diet. The apparent digestibility for the MIHPP diet was not different from between the MCP and MIHP diets. The low urinary P excretion, combined with a lower apparent digestibility seen for the MIHP diet, may point at a lower absorption of P from phytate than from monocalcium phosphate. Hintz et al. (1973) found an apparent P digestibility (expressed as a percentage of intake) of 18.2 (±5.0) for diets in which wheat bran supplied a significant amount of dietary P. They concluded that the P in wheat bran was about half as available as that from NaH₂PO₄ (39.5 ± 5.0). Thus, our results did not agree with the observations of Hintz et al. (1973).

As the phytase most likely acts in the stomach (due to its low pH optimum), the liberated P would be available for absorption in the small intestine and the dorsal large colon, which are the major sites of P absorption (Schryver et al., 1972a). Our results may indicate that dietary phytase improved phytate degradation, but it remains unclear to what extent the liberated P was absorbed. Digestibility and urinary variables for the MIHPP diet indicated a higher digestibility of P for the MIHPP than for the MIHP diet, but the differences lacked statistical significance. Our results do not confirm those of Patterson et al. (2002), who observed no effect of supplementing 300, 600, or 900 FTU/kg of concentrate on P availability. These researchers suggested that the lack of phytase effect was due to the high P levels in the diets used (28 to 33 g/d); however, the relatively low phytate content of the total diet may also explain the lack of effect of phytase supplementation in that study. Because the analyzed phytase activity of the MIHPP diet was lower than the intended concentration, it remains unclear whether a higher concentration would have resulted in more contrast between the MIHP and MIHPP diet. Additionally, the efficiency of supplemental phytase may have been somewhat decreased in our diets due to the high levels of Ca as described for growing pigs (Lei et al., 1994; Lantzsch et al., 1995).

Urinary P excretion (mg/kg BW) in our study was 0.07, 2.15, 0.61, and 1.18 for the control, MCP, MIHP, and MIHPP diets, respectively. Schryver et al. (1971b) calculated a linear relationship (y = 0.240x – 7.103) between P intake in mg of P/kg of BW (x) and P excretion in urine in mg of P/kg of BW (y) for young ponies. Phosphorus intake ranged between 30 to 116 mg/kg BW. The predicted values (mg/kg BW) based on the regression of Schryver et al. (1971b) were 2.12, 15.65, 13.81, and 15.07 for the control, MCP, MIHP, and MIHPP diets, respectively. Meyer (1990) derived a relationship from literature data, and his formula (y = 0.11x – 2.35) resulted in values of 1.88, 8.08, 7.23 and 7.81 for the control, MCP, MIHP, and MIHPP diets, respectively. The predicted urinary excretions (mg/kg BW) were much higher than values measured in the present study. This may be explained by differences in age (Schryver et al., 1971b), diet composition (Meyer, 1990), and roughage:concentrate ratios (Schryver et al., 1971b; Meyer, 1990). Despite the low values in our study, urinary P excretion did follow a pattern similar to that of apparent P digestibility. This confirms that, on average, urinary P excretion reflects differences in apparent P digestibility and thus can be used to interpret treatment effects. This is supported by the observations of Schryver et al. (1970a, 1971b), Whitlock et al. (1970), and Caple et al. (1982).

Phosphorus retention increased with P intake, which is consistent with studies in both young (Schryver et al., 1971a,b) and mature horses (Ott et al., 1975; Buchholz-Bryant et al., 2001). The P retention cannot be explained by P losses to sweat (Schryver et al., 1972a) or storage in hair (Wells et al., 1990) or hoof wall (Ley et al., 1998). Enteroliths might be a cause of high P retentions as enteroliths consist primarily of magnesium ammonium phosphate (Hassel et al., 2001) and have been related to dietary management (Bray, 1995). However, it seems unlikely from earlier reported dietary considerations that the rations fed in our experiment led to enterolith formation as not all conditions for formation were fulfilled (Bray, 1995). Schryver et al. (1971a) suggested that excess phosphate is retained in the hydroxyapatite crystal of the bone at the expense of carbonate. Such storage has been shown to occur in rats (Sobel et al. 1945a,b). Buchholz-Bryant et al. (2001) studied the effect of Ca and P supplementation in young (2 to 3 yr old), mature (7 to 11 yr old), and aged (15 to 21 yr old) horses with different exercise regimens. They reported higher retentions for all age groups when the horses were at rest and were fed either a normal or a high-P diet. However, during periods with intensive exercise, the mean apparent P balances were lower. This suggests that mature horses receiving exercise are better able to maintain a P balance close to zero, although higher P intake still resulted in higher P balances compared with the control diet. However, Nielsen et al. (1998) did not observe differences in P retention in young horses receiving various amounts of Ca and P and different exercise regimens. The effects of both age and exercise on P metabolism remain to be investigated.

We found that fecal and urinary Ca excretion were different for the control and MCP diets (Table 4), which may be explained by the depressing effect of high-P diets on Ca digestibility, as reported by Schryver et al. (1971a). We found that feeding of the MIHP diet increased fecal Ca and decreased urinary Ca excretion. On the control and MCP diets, the apparent Ca digestibility was 47.7 and 42.4% and was much higher than when the horses were fed the MIHP diet (26.4%). Compared with the MCP diet, Ca retention for the MIHP diet was lower. This difference may be caused by the origin of the Ca in these diets. Phytase supplementation improved apparent Ca digestibility despite the high dietary Ca levels used in this experiment. Phytase supplementation has been reported to increase Ca availability in other monogastric animals (Lei et al., 1994; Lantzsch et al., 1995; Pallauf et al., 1997). However, apparent Ca digestibility was higher (P < 0.05) for the control and MCP diets than for the MIHPP diet. If phytate P is partly available for horses, the depressing effect of P on Ca may also be an explanation for the lower apparent Ca digestibility seen for the MIHP and MIHPP diets. Our results are consistent with those of Hintz et al. (1973), who also observed a lower apparent Ca digestibility when wheat bran (67% of the total P was phytate P) was fed to ponies.

It has been reported that the linear regression equation of urinary Ca excretion (y) on Ca intake (x) is y = 0.33x – 2.13 (Meyer, 1990). Based on the average Ca intake in the present study, the predicted Ca excretion was 46 mg Ca/kg BW. We found an average Ca excretion of 50.2 mg Ca/kg BW. Despite the lower urinary Ca excretion on the MIHP and MIHPP diets, a somewhat higher Ca excretion was seen in our experiment than predicted.

Although Mg intake was lower on the control diet, the apparent MG digestibility and urinary Mg excretion were higher than those for the MCP, MIHP, and MIHPP diets. Different Mg levels may not affect apparent Mg digestibilities in horses (Hintz et al., 1972). Differences in apparent Mg digestibilities and urinary Mg excretion between the control vs. the MCP diet may therefore be explained by the inhibitory effect of high levels of P intake on Mg digestibility. Furthermore, sodium phytate or wheat bran did not affect Mg metabolism in ponies (Hintz and Schryver, 1972), but studies in other species have indicated that apparent Mg digestibility is affected by phytate in the diet (Palauf and Rimbach, 1997). Because phytase supplementation did not improve Mg availability in our study, we suggest that the low availability of Mg seen for the MCP, MIHP, and MIHPP diets may also be explained by the inhibitory effect of high P intake on Mg digestibility.

It has been shown, on the basis of 72 trials, that the linear regression equation of urinary Mg excretion (y) on Mg intake (x) is y = 0.188x + 2.80 (30). The calculated urinary Mg excretion for the average Mg intake (32.1 mg Mg/kg BW) resulted in a value of 8.8 mg Mg/kg BW. We found an average urinary Mg excretion of 8.5 which is in line with the reported regression equation (Hintz and Schryver, 1972). It seems that the predicted urinary excretion of both Ca and Mg are closer to the values observed than the predicted urinary P excretion. This implies that urinary P excretion is not an accurate indicator of P intake, which has also been reported by Meyer et al. (1990). However, urinary P, Ca, and Mg excretion reflected the corresponding apparent digestibilities. This suggests that urinary mineral excretion can be used to assess treatment effects on mineral provided that the concentrate:roughage of the rations is kept constant (Meyer, 1990).

Urinary P excretion was lower (P < 0.05) and apparent P digestibility tended (P = 0.065) to be lower for the MIHP diet compared with the MCP diet, suggesting a lower apparent digestibility of P from phytate than P from monocalcium phosphate. However, the observed digestibility of P for the MIHP diet indicated that a considerable amount of P was still absorbed on the MIHP diet compared with the MCP diet. We observed a small effect of phytase supplementation on P digestibility and no significant improvement in apparent P digestibility. This can be explained by the uncertainty about the fate of the P liberated from phytate and the relative small contribution of the increase in urinary P excretion (250 mg). Phytase improved apparent Ca digestibility. Our data show that feeding P as phytate has more influence on Ca digestibility than on P digestibility, which suggests that Ca forms stable complexes with phytate (Pallauf and Rimbach, 1997). Because the phytase added to the diet is mainly active in the gastric digesta (as a result of its low pH optimum), Ca becomes available for absorption in the small intestine which is the main site of Ca absorption in the horse (Schryver et al., 1970b). Phytase supplementation may be beneficial for improving apparent Ca digestibility in phytate-rich diets for horses.

Implications

For monogastrics, phytate-phosphorus represents a poorly available source of phosphorus, especially in the case of diets low in intrinsic phytase activity. According to published reports, the phosphorus in phytate from natural phosphorus sources is less available than that in pure sodium phytate. Our study showed that phytate phosphorus in feedstuffs is partly available for horses. Phytase supplementation with feedstuffs seemed to have a greater influence on apparent calcium digestibility than on phosphorus digestibility. This implies that phytase supplementation may be beneficial when phytate-rich diets are fed to horses.

Footnotes

¹ This study was funded by the Product Board Animal Feed (Produktschap Diervoeder), The Hague, The Netherlands.

² The assistance of B. Barlo, H. Bosch, D. van Velsen, L. de Jong, J. van der Kuilen, and the animal care keepers and veterinarians of the Dept. of Equine Sci., Faculty of Vet. Med., Utrecht Univ., The Netherlands, is gratefully acknowledged.

³ Correspondence: Yalelaan 16, P.O. Box 80153, 3508 TD Utrecht (phone: +31-(30)-253-4002/1234; fax: +31-(30)-253-1817; e-mail: a.c.beynen{at}vet.uu.nl).

Received for publication July 18, 2003. Accepted for publication February 26, 2004.

Literature Cited

Bos, K. D., C. Verbeek, C. H. P. van Eeden, P. Slump, and M. G. E. Wolters. 1991. Improved determination of phytate by ion-exchange chromatography. J. Agric. Food. Chem. 39:1770–1773.

Bray, R. E. 1995. Enteroliths: Feeding and management recommendations. J. Equine Vet. Sci. 15:474–477.

Buchholz-Bryant, M. A., L. A. Baker, J. L. Pipkin, B. J. Mansell, J. C. Haliburton, and R. C. Bachman. 2001. The effect of calcium and phosphorus supplementation, inactivity and subsequent aerobic training on the mineral balance in young, mature and aged horses. J. Equine Vet. Sci. 21:71–77.

Caple, I. W., P. A. Doake, and P. G. Ellis. 1982. Assessment of the calcium and phosphorus nutrition in horses by analysis of urine. Austr. Vet. J. 58:125–131.

Centraal Veevoederbureau. 1996. Het definitieve VEP- en VREp systeem, CVB documentatierapport No. 15. Centraal Veevoederbureau, Lelystad, The Netherlands.

Centraal Veevoederbureau. 1999. Veevoedertabel 1999. Lelystad, The Netherlands.

Engelen, A. J., F. C. van der Heeft, P. H. Randsdorp, and E. L. Smit. 1994. Simple and rapid determination of phytase activity. J. Assoc. Off. Anal. Chem. 77:760–764.

Goering, H. K. and P. J. van Soest. 1970. Forage fiber analyses. Agriculture Handbook No. 379. ARS, USDA, Washington, DC.

Hassel, D. M., P. S. Schiffman, and J. R. Snyder. 2001. Petrographic and geochemic evaluation of equine enteroliths. Am. J. Vet. Res. 62:350–358.[Medline]

Hintz, H. F., and H. F. Schryver. 1972. Magnesium metabolism in the horse. J. Anim. Sci. 35:755–759.

Hintz, H. F., A. J. Wiliams, J. Rogoff, and H. F. Schryver. 1973. Availability of phosphorus in wheat bran when fed to ponies. J. Anim. Sci. 36:522–525.[Abstract/Free Full Text]

Kemme, P. A. 1998. Phytate and phytases in pig nutrition. Ph.D. Diss., Utrecht Univ., Utrecht, The Netherlands.

Lantzsch, H. J., S. Wjst, and W. Drochner. 1995. The effect of dietary phytase on the efficacy of microbial phytase in rations for growing pigs. J. Anim. Physiol. A. Anim. Nutr. 73:19–26.

Lei, X. G., P. K. Ku, E. R. Miller, M. T. Yokoyama, and D. E. Ullrey. 1994. Calcium level affects the efficacy of supplemental microbial phytase in corn-soybean meal diets of weanling pigs. J. Anim. Sci. 72:139–143.[Abstract]

Lewis, L. D. 1995. Feeding and Care of the Horse. Lippincott, Williams & Wilkins, Philadelphia, PA.

Ley, W. B., R. Scott Pleasant, and E. A. Dunnington. 1998. Effects of season and diet on tensile strength and mineral content of the equine hoofwall. Equine Vet. J. 26(Suppl.):46–50.

Matsui, T., H. Murakami, H. Yano, H. Fujikawa, T. Ossawa, and Y. Asai, 1999. Phytate and phosphorus movements in the digestive tract of horses. Equine Vet. J. 30(Suppl.):505–507.

Meyer, H. 1990. Assessing of the mineral supply of horses by urine analysis. Pages 86–97 in Contributions to Water and Mineral Metabolism of the Horse. H. Meyer, ed. Stadermann, Institut fur Tierernahrung, Tierarzliche Hochschule, Hannover, Germany.

NRC. 1989. Nutrient Requirements of Horses. Natl. Acad. Press, Washington, DC.

Nielsen, B. D., G. D. Potter, L. W. Greene, E. L. Morris, M. Murray-Gerzik, W. B. Smith, and M. T. Martin. 1998. Response of young horses in training to varying concentrations of dietary calcium and phosphorus. J. Equine Vet. Sci. 18:397–404.

Ott, E. A., J. P. Fealter, and J. P. Panco. 1975. Effect of calcium and phosphorus levels on availability of trace minerals. Pages 61–64 in Proc. 4th Equine Nutr. Physiol. Symp., Equine Nutr. and Physiol. Soc., California Polytechnic State Univ., Pomona.

Pallauf, J., and G. Rimbach. 1997. Nutritional significance of phytic acid and phytase. Arch. Anim. Nutr. 50:301–319.

Patterson, D. P., S. R. Cooper, D. W. Freeman, and R. G. Teeter. 2002. Effects of varying levels of phytase supplementation on dry matter and phosphorus digestibility in horses fed a common textured ration. J. Equine Vet. Sci. 22:456–459.

Schonewille, J. Th., A. C. Beynen, A. Th. van’t Klooster, H. Wouterse, and L. Ram. 1999. Dietary potassium bicarbonate and potassium citrate have a greater inhibitory effect than does potassium chloride on magnesium absorption in wethers. J. Nutr. 129:2043–2047.[Abstract/Free Full Text]

Schryver, H. F., P. H. Craig, and H. F. Hintz. 1970a. Calcium metabolism in ponies fed varying levels of calcium. J. Nutr. 100:955–964.

Schryver, H. F., P. H. Craig, and H. F. Hintz. 1970b. The site of calcium absorption in the horse. J. Nutr. 100:1127–1132.

Schryver, H. F., and H. F. Hintz. 1972a. Calcium and phosphorus requirements of the horse: a review. Feedstuffs 44(28):35–36.

Schryver, H. F., H. F. Hintz, and P. H. Craig. 1971a. Calcium metabolism in ponies fed a high phosphorus diet. J. Nutr. 101:259–264.

Schryver, H. F., H. F. Hintz, and P. H. Craig. 1971b. Phosphorus metabolism in ponies fed varying levels of phosphorus. J. Nutr. 101:1257–1263.

Schryver, H. F., H. F. Hintz, P. H. Craig, D. E. Hogue, and J. E. Lowe. 1972b. Site of phosphorus absorption from the intestine of the horse. J. Nutr. 102:143–148.

Sobel A. E., M. Rockenmacher, and B. Kramer. 1945a. Carbonate content of bone in relation to the composition of blood and diet. J. Biol. Chem. 158:475–489.[Free Full Text]

Sobel A. E., M. Rockenmacher, and B. Kramer. 1945b. Composition of bone in relation to blood and diet. J. Biol. Chem. 159:159–171.[Free Full Text]

Tasker, J. B. 1965. Fluid and electrolyte studies in the horse. II. An apparatus for the collection of total daily urine and feces from horses. Cornell Vet. 57:77–84.

Quinlan, K. P., and M. A. DeSesa. 1955. Spectrophotometric determination of phosphorus as molybdovanadophosphoric acid. Anal. Chem. 27:1626–1629.

Wells, L. A., R. LeRoy, and S. L. Ralston. 1990. Mineral intake and hair analysis of horses in Arizona. J. Equine Vet. Sci. 10:412–416.

Whitlock, R. H., H. F. Schryver, L. Krook, H. F. Hintz, and P. H. Craig. 1970. The effects of high dietary calcium in horses. Page 127–134 in Proc. 16th Am. Assoc. Equine Pract. Lexington, KY.

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