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Effects of microbial phytase, low calcium and phosphorus, and removing the dietary trace mineral premix on carcass traits, pork quality, plasma metabolites, and tissue mineral content in growing-finishing pigs^1,2

Department of Animal Sciences, Louisiana State University Agricultural Center, Baton Rouge 70803-4210

	Abstract

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An experiment was conducted to determine the effects of phytase addition, reduced Ca and available P (aP), and removing the trace mineral premix (TMP) on growth performance, plasma metabolites, carcass traits, pork quality, and tissue mineral content in growing-finishing swine. One hundred twenty crossbred pigs (initial and final BW of 22 and 109 kg, respectively) were allotted to five dietary treatments on the basis of weight within gender in a randomized complete block design. There were three replications of barrows and three replications of gilts, with four pigs per replicate pen. The dietary treatments were as follows: 1) corn-soybean meal (C-SBM), 2) C-SBM with reduced Ca and aP, 3) C-SBM with reduced Ca and aP plus 500 phytase units/kg of diet, 4) Diet 1 without the TMP, and 5) Diet 3 without the TMP. The Ca and aP were reduced by 0.10% in the low Ca and aP diets and the diets with added phytase. Daily gain, hot carcass weight, dressing percent, kilograms of carcass lean, bone ash percent, and bone strength were decreased (P = 0.10), but liver and kidney weight were increased (P = 0.10) in pigs fed diets with reduced Ca and aP; adding phytase reversed these responses (P = 0.10). The Commission Internationale de I’Eclairage L* was decreased (P = 0.09) in pigs fed the low Ca and aP diet plus phytase relative to those fed the control diet. Removing the TMP had no effect on overall growth performance, but it increased (P = 0.03) 10th-rib backfat thickness and fasting glucose and decreased (P = 0.03) carcass length and ham weight. Liver weight and liver weight as a percentage of final BW were not affected when phytase was added to the control diet, but removing the TMP increased liver weight and liver weight as a percentage of final BW; adding phytase reversed these responses (phytase x TMP, P = 0.06). Removing the TMP decreased (P = 0.08) Zn concentrations in the bone, muscle, and liver, and Cu and Fe concentrations in the bile but increased (P = 0.08) Mn concentrations in the bile and liver of pigs. The addition of phytase reversed the negative effects of the reduced Ca and aP diets. These data indicate that removing the TMP in diets for growing-finishing pigs has no negative effects on growth performance or pork quality, but it had negative effects on carcass traits and had variable effects on tissue mineral content.

Key Words: Carcass Composition • Carcass Quality • Phytase • Pigs • Trace Minerals

	Introduction

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Feed represents approximately 65% of the production costs of market pigs; thus, decreasing the cost of feed is important to the swine industry. Kim et al. (1997) and Mavromichalis et al. (1999) indicated that the trace mineral premix (TMP) could be deleted in diets for finishing pigs with no negative effects on growth or carcass traits or quality. There has been little research conducted on removing the TMP for the entire growing-finishing period, and most research deleting the TMP during the finishing phase also deleted the vitamin premix (Edmonds and Arentson, 2001; Shaw et al., 2002).

Phytate has the potential to form insoluble salts with Ca, Fe, Zn, Mn, and Cu (Vohra et al., 1965), which may decrease the availability of these minerals. Lei et al. (1993) and Adeola et al. (1995) reported that Zn bioavailability and retention were improved with the addition of phytase in the diet for weanling pigs. Moreover, Spears et al. (2001) reported that pigs fed phytase with no supplemental Zn performed as well as those fed supplemental Zn. Therefore, phytase addition to diets for pigs may decrease or eliminate the need for TMP supplementation.

If phytase is to be used in diets for growing-finishing pigs, effects on carcass traits and quality need to be determined. To date, there have been varying effects of phytase addition on carcass traits and quality. Rienstra et al. (2001) reported that pigs fed phytase had an increased LM area (LMA) and a decreased marbling and drip loss. Gebert et al. (1999) reported that phytase addition resulted in a paler LM and a decreased 45-min pH, and O’Quinn et al. (1997) reported a decreased dressing percent when phytase was added to diets for pigs.

The objectives of this experiment were to determine the effects of phytase addition, reduced Ca and available P (aP), and removing the TMP on growth performance, plasma metabolites, carcass traits and quality, and tissue mineral content in growing-finishing pigs.

	Materials and Methods

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All methods used in this experiment were approved by the Louisiana State University Agricultural Center Animal Care and Use Committee.

General.
One hundred twenty crossbred (Yorkshire x Landrace and Yorkshire x Duroc) gilts and barrows with an average initial and final BW of 22 and 109 kg, respectively, were used in this experiment. The pigs were allotted to five dietary treatments based on weight and gender with three replications of barrows and three replications of gilts, and with four pigs per replicate pen in a randomized complete block design. Ancestry was equalized within treatment as much as possible. The pigs were housed in total confinement in 1.5- x 3.0-m pens with concrete-slatted floors.

The dietary treatments were as follows: Diet 1) corn-soybean meal (C-SBM) positive control; Diet 2) C-SBM with reduced Ca and aP; Diet 3) C-SBM with reduced Ca and aP plus phytase (Natuphos 1200; BASF Corp., Mount Olive, NJ) to provide 500 phytase units per kilogram of diet; Diet 4) Diet 1 without the TMP supplementation (contained Ca and aP equal to that of the positive control diet); and Diet 5) Diet 3 without the TMP supplementation (Table 1). Actual analysis indicated that Diet 3 provided 877 phytase units per kilogram of diet, and Diet 5 provided 883 phytase units per kilogram of diet. To test the effect of reduced Ca and aP with and without phytase, Diets 1, 2, and 3 were used. To test the effects of phytase addition in diets with and without the TMP, the following diets were used: Diets 1, 3, 4, and 5.

Blood Metabolites.
On d 26 of the experiment, the feeders were removed from the pen at 1600. On d 27 at 0800, blood was collected via the anterior vena cava and placed into 7-mL tubes containing 17.5 mg sodium fluoride and 14.0 mg potassium oxalate (Monoject; Sherwood Medical, St. Louis, MO). The samples were then centrifuged at 1,500 x g and 4°C for 45 min. After centrifugation, the plasma from each sample was collected and frozen until analysis. Glucose concentrations were determined by a spectrophotometric procedure (Sigma, 1989). Blood samples were collected on d 27 because this is the approximate time at which Johnston et al. (2004) and Shelton et al. (2003b) reported changes in plasma glucose in pigs fed phytase. At slaughter, blood was collected during exsanguination for determination of hematocrit percent (Adams Autocrit Centrifuge, Papsippanny, NJ).

Carcass Evaluation.
On the day after the growth trial ended, three pigs per replicate pen were randomly selected and slaughtered by exsanguination after electrical stunning at the Louisiana State University Agricultural Center Meats Laboratory. Conventional carcass measurements and values from total body electrical conductivity (TOBEC, Model MQ1-27; Meat Quality Inc., Springfield, IL) were determined as described by Matthews et al. (2001a). The NPPC (1991) equation that assumes 5% estimation for intramuscular fat and compensates for unequal BW was used to evaluate the percentage of acceptable quality lean and kilograms of carcass lean. Leaf fat and the liver and kidneys were individually weighed at slaughter. After a 20-h chill at 2°C, the left front foot was removed and frozen.

Pork Quality.
Pork quality measurements were taken from the left side of the carcass after a 20-h chill at 2°C as described by Matthews et al. (2001a,c). Drip loss was determined by a suspension method (Shelton et al., 2003a). Cooking loss and shear force also were determined on a fresh chop taken from the 10th rib as outlined by Matthews et al. (2001c). Shear force was determined on three cores using an HD 250 Texture Machine (Texture Technologies Corp., Scarsdale, NY) fitted with a Warner-Bratzler head with a load cell capacity of 25 kg and a crosshead speed of 100 mm/min. One additional chop was taken from the ninth rib, deboned, external fat removed, and frozen for 90 d, and thaw and cook loss were determined.

Tissue Ash Determination and Mineral Content.
The third and fourth metacarpal bones from the left foot of each pig were removed and manually cleaned of adhering tissue. The fourth metacarpal bone was broken using an HD 250 Texture Machine fitted with a three-point bend rig with a load cell capacity of 250 kg and crosshead speed of 100 mm/min and a span over which the bone was set of 1.5 cm. Fat was removed from the third metacarpal bone by a 36-h Soxhlet extraction in ethyl alcohol followed by a 36-h extraction with diethyl ether, and then dried at 100°C.

A 20-g sample of kidney and liver was taken at slaughter and frozen for subsequent determination of ash percent. A 5-mL sample of bile was taken for determination of mineral content after drying at 100°C for 24 h. The liver sample also was analyzed for mineral content. A 1.27-cm chop from the eighth rib was taken, homogenized, and frozen for subsequent determination of ash percent and mineral content. Dry matter of the liver, kidney, and LM was determined by weighing a 5.0-g sample and drying at 100°C for 24 h. Percentage of ash was determined by placing the samples into a muffle furnace and ashing for 12 (liver, kidney, and LM) or 36 h (third metacarpal bone) at 550°C. The ash samples were dissolved in nitric acid, and mineral content was determined by inductively coupled plasma emission spectroscopy (Optima 3000; Perkin Elmer, Norwalk, CT).

Statistical Analyses.
Data were analyzed by analysis of variance procedures for a randomized complete block design using the GLM procedures of SAS (SAS Inst. Inc., Cary, NC). The statistical model included treatment, replication, and sex. The treatment x sex interaction was significant only for the bile mineral content data, and it was removed from the model for all other data. To test the effect of reduced Ca and aP with and without phytase, Diets 1, 2, and 3 were used. Treatment differences for pigs fed these three diets were separated by the PDIFF option of SAS. To test the effects of phytase addition in diets with and without the TMP, the following diets were used: Diets 1, 3, 4, and 5. These four diets were analyzed by contrast statements to evaluate phytase, TMP, and phytase x TMP effects as a 2 x 2 factorial arrangement of treatments. Treatment differences were considered significant at = 0.10. Final BW was used as a covariate for the carcass data. The pen of pigs was the experimental unit for all data.

	Results

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Results will be discussed in terms of the effects of Ca and aP with or without phytase and the effects of phytase x trace mineral supplementation.

Growth Performance
Low Ca and aP With or Without Phytase.
Daily gain was decreased (P = 0.02) in pigs fed the low Ca and aP diet, but adding phytase to the low Ca and aP diet increased (P = 0.02) ADG equal to that of pigs fed the control diet (Table 2). This response was noted during all phases of growth. The ADFI or G:F was not affected by Ca and aP with or without phytase.

Blood Metabolites
Low Ca and aP With or Without Phytase.
Hematocrit percent was increased (P = 0.10) in pigs fed the low Ca and aP diet plus phytase relative to those fed the low Ca and aP diet (Table 2). The dietary treatments did not affect the fasting plasma glucose concentrations of pigs.

Phytase x Trace Mineral.
Fasting plasma glucose concentration was increased (P = 0.09) in pigs fed the diets without the TMP. Hematocrit percent of pigs was not affected by these diets.

Carcass Evaluation
Diet did not affect LMA, average backfat thickness, lean gain per day, leaf fat, ham butt-face fat thickness, NPPC (National Pork Producers Council) percentage of acceptable quality lean, kilograms of carcass fat-free lean, percentage of carcass fat-free lean, kilograms of carcass fat, percentage of carcass fat, carcass lean:fat ratio, kilograms of ham fat-free lean, percentage of ham fat-free lean, kilograms of ham fat, or percentage of ham fat (Table 3).

Phytase x Trace Mineral.
Tenth-rib backfat thickness was increased (P = 0.03) but carcass length and ham weight were decreased (P = 0.03) in pigs fed the diets without the TMP. Liver weight and liver weight as a percentage of final BW were increased in pigs fed diets without the TMP, but this response was reversed by phytase (mineral, phytase, and phytase x mineral, P = 0.06).

Pork Quality
Diet did not affect color, marbling, Commission Internationale de I’Eclairage (CIE) a* value; CIE b* value; 45-min pH; 24-h temperature or pH; cook loss or shear force of a fresh chop; and thaw loss, cook loss, or total loss of a frozen (thawed) chop (Table 4).

Phytase x Trace Mineral.
Forty-five-minute temperature was increased (P = 0.07) in pigs fed the diets with added phytase.

Tissue Ash Determination and Mineral Content
Many tissue mineral concentrations were not affected by diet. Unaffected tissue mineral concentrations (mean ± pooled SEM, DM basis) were representative of 30 replications of three pigs per replicate pen. Bile: K, 0.37 ± 0.02%; Mg, 0.051 ± 0.002%; Na, 4.22 ± 0.22%. Muscle: ash 4.78 ± 0.10%; K, 1.20 ± 0.11%; Mg, 0.10 ± 0.01%; Na, 0.40 ± 0.05%. Liver: ash, 5.34 ± 0.04%; Mg, 0.058 ± 0.004%; Na, 0.50 ± 0.07%. Kidney: ash, 6.35 ± 0.09%.

Low Ca and aP With or Without Phytase.
Bone strength and ash percent were decreased in pigs fed the low Ca and aP diet, but adding phytase reversed the response (P = 0.10; Table 5). Copper and Mn concentrations in the bile and Na and Zn concentrations in the liver were increased in pigs fed the low Ca and aP diet (Table 6). Adding phytase reversed these responses in Cu and Mn concentrations in the bile (P = 0.10). Copper in the muscle and Zn in the liver were increased (P = 0.10), and Zn concentration in the bone was decreased (P = 0.10) in pigs fed the diet with phytase added to the low Ca and aP diet relative to those fed the control diet. Phosphorus in the liver was increased (P = 0.10) in pigs fed the diet with phytase added to the low Ca and aP diet relative to those fed the low Ca and aP diet. Potassium, Mg, and Mn concentrations in the bone were decreased (P = 0.10) in pigs fed the low Ca and aP diet relative to pigs fed the control diet.

	Discussion

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Low Ca and aP With or Without Phytase
Harper et al. (1997), O’Quinn et al. (1997), and Rienstra et al. (2001) reported that dietary phytase addition overcame the negative effects of reduced levels of dietary Ca and P on growth performance of pigs. Similarly, Kornegay and Qian (1996), Harper et al. (1997), and O’Quinn et al. (1997) reported a similar effect of phytase in reduced Ca and P diets on bone strength and bone ash percent. Our data agree with these previous reports.

The effect of phytase in reduced Ca and P diets on carcass traits has been inconsistent. O’Quinn et al. (1997) reported a decreased dressing percent when 300 phytase units per kilogram of diet were added to swine diets, but not when 0 or 500 phytase units per kilogram of diet were added (the Ca and P were reduced by 0.08% in diets with phytase). Rienstra et al. (2001) indicated that pigs fed phytase had an increased LMA, and Harper et al. (1997) indicated that adding phytase to swine diets had no effect on carcass length, backfat thickness, or LMA. Our data indicate that reduced Ca and P diets have negative effects on carcass traits of pigs, but the addition of phytase reversed these responses.

Gebert et al. (1999) reported a paler longissimus dorsi when pigs were fed phytase, which the authors attributed to an increase in the availability of trace minerals, mainly Fe and Cu. Also, Berg (2001) indicated that supranutritional levels of Cu may result in less desirable (paler) pork color. However, in our study, Cu concentration in the muscle was increased when phytase was added to the diet, but the chops were darker (CIE L* was decreased). Gebert et al. (1999) reported that adding phytase to pig diets resulted in a decreased 45-min pH, but we observed no effect of phytase on 45-min pH. Harper et al. (1997) indicated that adding phytase to pig diets had no effect on firmness or marbling, which agrees with our data; however, Rienstra et al. (2001) reported that pigs fed phytase had a decreased marbling and drip loss. In our study, there were numerical decreases in drip loss, thaw loss, and cook loss (fresh and frozen chops) in pigs fed diets with phytase and the diet with reduced Ca and P. The effect of phytase on moisture loss may be due to the reduction in monocalcium phosphate in the diet, which has been shown to decrease cooking loss (Mavromichalis et al., 1999).

Phytase x Trace Mineral
Previous research demonstrated that removing the TMP only during the finishing period does not affect growth performance of swine (Kim et al., 1997; Mavromichalis et al., 1999); however, weight ranges for these studies were 70 to 112 kg and 86 to 116 kg, respectively, whereas in our study the weight range was 22 to 109 kg. Other research has shown that removing the vitamin premix and TMP had no effect on growth performance in swine (Edmonds and Arentson, 2001; Shaw et al., 2002) or poultry (Skinner et al., 1992; Deyhim and Teeter, 1993; Christmas et al., 1995). Research could not be found in the literature on removing the TMP at the start of the growing period (22 to 109 kg). Nonetheless, Spears et al. (2001) indicated that removing the Zn and Cu for 22- to 91-kg pigs did not negatively affect growth performance. Although we observed no adverse effect of removing the TMP on growth performance, on commercial farms there are many stressors that can increase the requirement for TMP including temperature, stocking density, and degree of contamination (Cunha, 1977; Stahly et al., 1997).

Stahl et al. (1999) reported that phytase overcame the decreased hematocrit percent that resulted from Fe-deficient diets. In our study, hematocrit percent was not affected when pigs were fed diets with or without the TMP or with or without phytase. This lack of response could be due to the high Fe concentrations in the diets even with the removal of the TMP. Johnston et al. (2003) and Shelton et al. (2003b) reported an increase in fasting glucose concentrations when phytase was added to diets for growing pigs, but we observed no effect of phytase on fasting glucose. The increase in fasting glucose with the removal of the TMP from the diet that we observed may be due to an interaction of one or more of the trace minerals that were removed with another mineral such as Cr, which has been shown to affect glucose metabolism (Matthews et al., 2001b)

Kim et al. (1997) and Mavromichalis et al. (1999) reported no effect on carcass traits when the TMP was removed from diets for finishing pigs. In addition, Skinner et al. (1992) indicated that removing the TMP from poultry diets from 42 to 49 d had no effect on dressing or abdominal fat percents. Deyhim and Teeter (1993) indicated that removing the TMP from poultry diets from 28 to 49 d had no effect on dressing percent and breast and fat pad weight as a percentage of carcass weight. The negative effects (increased 10th-rib backfat and decreased carcass length and ham weight) noted in our study could be due to the length of time the TMP was removed from the diets, which was much longer than in previous research. Adding phytase did not reverse the negative effects in carcass traits observed with the removal of the TMP.

Shelton et al. (2003b) reported a decreased liver weight in growing pigs fed diets with phytase at 3.2 x maintenance level of growth, but a TMP was included in the diet in that study. Our results do not agree with Deyhim and Teeter (1993), who indicated that liver weight as a percentage of carcass weight was not affected when chicks were fed diets without the TMP from 28 to 49 d. In our study, reducing the Ca and P or removing the TMP resulted in an increase in liver weight of pigs, but phytase reversed the response. This response indicates that the increase in liver weight may result from a decrease in dietary mineral (macro or micro) levels.

Kim et al. (1997) and Mavromichalis et al. (1999) reported that removing the TMP in the diets of growing-finishing pigs had no effect on pork quality. Tian et al. (2001) reported no effect on pork quality when finishing pigs were fed diets with 50% of the requirement for Zn, Fe, Mn, Cu, I, and Se; these data agree with the results of our study. Although in our study there were no negative effects on pork quality when the TMP was removed, its effect on human nutrition needs to be determined because decreases in tissue trace minerals may have negative effects on the health of humans consuming pork as a source of these trace minerals.

Edmonds and Arentson (2001) and Shaw et al. (2002) indicated that removing the TMP had no effect on Cu, Fe, or Zn concentrations in the LM. In both studies, the vitamin premix was deleted, and, in the study by Shaw et al. (2002), two-thirds of the dicalcium phosphate was deleted. These data do not agree with our results, which indicated that removing the TMP had variable effects on tissue mineral concentration. The reason for this discrepancy may be due to the length of time the diets were fed. In the studies by Edmonds and Arentson (2001) and Shaw et al. (2002), the treatment diets were fed only during the finishing period, whereas in our study the treatment diets were fed during the growing and finishing periods.

Phytate has the potential to bind with some trace minerals, such as Zn, Cu, Mn, Mo, Co, Mg, and Fe, thereby decreasing their availability (Vohra et al., 1965; Erdman, 1979; Ravindran et al., 1995). Thus, it may be possible to decrease trace mineral levels in diets with phytase addition. Our data agree with those of Adeola et al. (1995), who indicated that Zn, Cu, and P absorption and retention were increased when phytase was added to the diet of pigs, but Mg absorption and retention were not affected by phytase supplementation. Also, Stahl et al. (1999) reported that phytase addition to the diet of young pigs increased Fe availability, but, in our study, Fe concentration in the bile was decreased and Fe concentrations in other tissues were not affected.

The effect of phytase supplementation in diets with and without the TMP on Mn indicate there may be an interaction between Zn and Mn because, when Zn concentration was decreased by removing the TMP, Mn concentration in the liver was increased; however, when Zn concentrations were increased by adding phytase to the diet, Mn concentration decreased. The same effect regarding Mn occurred in the bile, but there was only a numerical increase in Zn from the phytase addition (53.6 ppm Zn with the TMP removed and 57.2 ppm Zn with phytase added to that diet). Adeola et al. (1995) indicated that supplementing pig diets with Zn decreased Mg and Mn absorption and retention. Mohanna and Nys (1999) found no improvement in Mn retention by microbial phytase supplementation in broiler chicks, but Windisch and Kirchgessner (1996) indicated that phytase addition increased Mn retention by 3.0% in pigs.

The effects observed regarding the Cu concentrations may be explained by the interaction of Zn and Cu. Copper level in the liver was not affected, and, in bone, Cu concentration was increased when the TMP was removed from the diet, and adding phytase decreased the Cu concentrations in the bone and liver to levels below those of pigs fed the control diet. It has been documented that increasing the Zn concentration in the diet will decrease the availability of dietary Cu for pigs (Blakeborough and Salter, 1987) and rats (Frimpong and Magee, 1989), and the concentrations of Zn were increased in bone and liver when phytase was added to the diet.

Removing the TMP in the diets of pigs during the growing-finishing periods resulted in no effect on growth performance or pork quality components measured but resulted in some negative effects on carcass traits. Furthermore, removing the TMP will decrease the amount of trace minerals in pork. The effects of decreasing trace mineral concentrations in pork and its effect on human nutrition need to be further investigated.

	Implications

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The trace mineral premix can be removed from diets of growing-finishing pigs with no change in growth performance, but with minimal negative effects on carcass traits and pork quality. Stress that occurs in commercial settings may make the restrictions more severe. The addition of phytase improved trace mineral availability, and, if phytase is used to replace a portion of the Ca and P, the removal of the trace mineral premix will likely result in no negative effect. However, further research is needed to determine when the trace mineral premix can be removed and its potential effect with or without the addition of phytase on the nutrition of humans consuming pork.

	Footnotes

 
¹ Approved for publication by the director of the Louisiana Agric. Exp. Stn. as manuscript No. 03-18-1368.

² The authors thank M. Persica, J. Carothers, A. Guzik, R. Payne, B. Watson, and T. O’Connor-Dennie for assistance with data collection and laboratory analyses.

³ Correspondence—e-mail: lsouthern{at}agctr.lsu.edu.

Received for publication October 14, 2003. Accepted for publication May 18, 2004.

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