Nutrition Research 22 (2002) 623 632

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Phytase and acetic acid supplementation in the diet of early weaned piglets: effect on performance and apparent nutrient digestibility
Zully Valencia, M.Sc., Eduardo R. Chavez, Ph.D.
Department of Animal Science, McGill University, 21,111 Lakeshore Rd. Ste. Anne de Bellevue, Quebec H9X 3V9 Received 13 July 2001; received in revised form 8 February 2002; accepted 10 February 2002

Abstract A study with Landrace x Yorkshire piglets weaned at 21 days of age was conducted for four weeks to assess the growth promoting effects on nutrient utilization of either phytase, acetic acid or a mix of the two in the starter diet. The treatments (trt) were corn-soybean meal based as follows: 1.control diet, 0.35% of available P; 2. low P, 0.15% Pi; 3. as 2 plus 1000 phytase units/kg diet; 4. as 3 plus 1% acetic acid; and 5. as 1 plus 1% acetic acid. Supplementing phytase to the low P diet, signicantly improved the ADG. There was no difference in growth rate between normal P level and low P plus phytase. A signicant additional effect was obtained when acetic acid was incorporated in diet 3. Apparent digestibility of P and calcium (Ca) were signicantly increased by both phytase and acetic acid, and by phytase-acetate supplementation. These results showed an additional effect of acetic acid combined with phytase in promoting better mineral digestibility. 2002 Elsevier Science Inc. All rights reserved.
Keywords: Piglets; Phosphorus; Phytase; Acetic acid; Apparent digestibility

1. Introduction Theoretically, the phosphorus (P) content in corn and soybean meal should be enough to meet the requirement of piglets. However, about two thirds of the P in these feed ingredients are not available to the animal because the element comes in the chemical form of phytate

* Corresponding author. Tel.: 1-514-398-7795; fax: 1-514-398-7964. E-mail address: chavez@macdonald.mcgill.ca (E.R. Chavez).
0271-5317/02/$ see front matter 2002 Elsevier Science Inc. All rights reserved. PII: S 0 2 7 1 - 5 3 1 7 ( 0 2 ) 0 0 3 7 1 - 8

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[1]. The very limited ability of monogastrics to utilize P from feedstuffs of plant origin is well documented. Non-ruminant animals lack intestinal phytase to hydrolyse P from the phytic acid molecule. The endogenous phytase activity in the intestinal mucosa of pigs is negligible [2]. In the early-weaned pig this inefciency is aggravated because its capacity for digesting feeds containing plant materials is still very limited [3]. Attempts have been made to eliminate the weaning stress and to improve weaning performance of pigs. Several feed additives such as synthetic enzymes [4], organic acids and antibiotics have been developed. Microbial phytase is an enzyme that is being added to the diets of poultry [4 6], and pigs [711] to improve the utilization of phytate-P and at the same time to reduce the amount of P excrete in faeces which represents a hazard to the integrity of the environment. Organic acids have been added to the diets of pigs [1213] to improve the feed utilization and live weight gain during the post weaning lag period. An acidication of the diet of the earlyweaned pig may help to reduce the gastric pH and consequently, stimulate the activity of pepsin and pancreatic proteases [3]. Protein digestibility as well as the digestibility of other nutrients, was reported to be improved by the addition of organic acids [14]. More recently, Jongbloed et al [15] reported a synergistic effect of organic acids with microbial phytase for ash, P and Mg apparent digestibility. Most published studies of supplementation of the diets of weaned pigs have included only one feed additive at the time, phytase or organic acid, as a mechanism to improve growth rate, and most have been done with heavier pigs. The objective of the experiment reported herein was to evaluate the growth promoting effects and the effects in the bioavailbility of minerals and protein of supplemental phytase, acetic acid or both on early-weaned pigs.

2. Materials and Methods 2.1. Animals and housing One hundred and thirty Landrace x Yorkshire piglets weaned at 21 days of age in three different batches a few weeks apart, weighing on average 6.54 kg were randomly allotted to ve different treatments balanced for litter, weight and sex. Animal care was according to the guidelines of the Canadian Council on Animal Care [16]. Piglets had access to creep feed during the suckling period. A three-day adaptation period after weaning (litter-mate pigs remained together) was allowed for pigs to adjust to the feeding regimen (a commercial starter diet was offered during this period). Thereafter, pigs were distributed in groups and assigned one experimental diet. The animals were housed by groups of four in the rst two blocks and groups of 5 in block 3 in an environmentally controlled nursery. The weaner-deck (0.9m x 2.4m) was equipped with a heat lamp, a ten-space standard nursery feeder and low pressure water-biting nipple suspended on the pen sidewall. 2.2. Experimental diets Piglets were fed one of ve corn-soybean meal diets. They differed only in P, phytase or acetic acid supplementation. Experimental diet 1, was the control diet. It had 0.35%, the

Z. Valencia, E.R. Chavez / Nutrition Research 22 (2002) 623 632 Table 1 Composition of the basal mix and the experimental diets (%) Ingredients Experimental diets (%) Control (1) Corn Soybean meal Tallow Di-Ca phosphate Calcium carbonate Min Vita Choline-choride L-lysine Mecadow Chronic oxide Phytaseb Acetic acidc Calculated chemical composition (%) Crude protein Calcium Phosphorus Analysed chemical composition (%) Crude protein Calcium Phosphorus
a

625

Low-P (2) 54.48 32.00 10.00 0.82 1.65 0.40 0.20 0.10 0.10 0.25

As 2 (3) 54.48 32.00 10.00 0.82 1.65 0.40 0.20 0.10 0.10 0.25

phytase

As 3 acid (4) 53.48 32.00 10.00 0.82 1.65 0.40 0.20 0.10 0.10 0.25 1.00

acetic

As 1 acid (5) 53.16 32.00 10.00 1.92 0.87 0.40 0.20 0.10 0.10 0.25 1.00 20.0 0.80 0.70 20.6 0.93 0.66

acetic

54.16 32.00 10.00 1.92 0.87 0.40 0.20 0.10 0.10 0.25

20.0 0.80 0.70 20.7 0.97 0.69

20.0 0.80 0.55 20.9 0.92 0.47

20.0 0.80 0.55 21.0 0.94 0.48

20.0 0.80 0.55 20.4 0.90 0.47

Amount provided (g kg 1 diet: Fe 100; Zn 100; Cu 6; Mn 4; Se 0.3; I 0.14; NaCl 0.25%. (mg kg 1 diet) vit K 0.5; biotin 0.1; folic acid 0.3; niacin 20; pantothenic acid 18; riboavin 6; thiamine 2; BHT antioxidant 50; vit B-12 30 g; (IU kg 1) vit A 4,000; vit D3 500; vit E 25. b Natuphos. Fisher Scientic, Montreal, Canada. Added at 1,000 PTU/kg diet. c Acetic acid Lab Grade, Caledon Laboratories Ltd., Georgetown, Ont., Canada. Concentraiton 99% diluted to 1:1 with water.

NRC [17] recommendation of the available P provided from an inorganic P (Pi) source (di-Ca- phosphate). Treatments (trt) 2,3, and 4 were low (0.15%) in available P. Microbial phytase (Natuphos, Fisher Scientic, Montreal, Canada) at a rate of 1000 PTU/kg diet was added to trt 3. Experimental diet of trt 4 was as diet 3 supplemented with 1% acetic acid added at the expense of corn. In trt 5, the diet with acetic acid supplementation was offered to the piglets for only the rst two weeks of the 4-week trial. Then, they were fed the control diet (Table 1). The acetic acid (99% Lab Grade, Caledon Laboratories Ltd) was diluted (1:1) with water before being added. Pigs were allowed ad libitum access to water and feed. Chromic oxide was added (0.25%) to the experimental diets and fed during the fourth week to determine the apparent digestibility values of minerals. 2.3. Measurements Experimental diets were offered to piglets in group-pens for four consecutive weeks. Individual pigs and pen feed consumption were weighed weekly. Individual average daily

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gain (ADG), average-pen daily feed intake (ADFI) and the pen feed/gain or feed conversion ratio (FCR) values were calculated. Random samples of faeces were collected from each pen during the fourth week of the trial for four consecutive days. Samples of faeces were taken from each pig by anal stimulation and pooled daily by pen in aluminium trays. A representative faecal sample was freeze-dried, ground and stored at room temperature until it was analysed. 2.4. Chemical analysis Each diet was analysed for dry matter, ether extract, and ash according to AOAC methods [18]. Crude protein was determined in a LECO nitrogen analysis system (LECO Corporation, model FP-428). Crude protein was calculated using a factor of 6.25. Gross energy determination was made in an adiabatic bomb calorimetry (Parr Instrument). Diets and faeces were analysed for Ca, Mn, Zn, Fe, Cu, and Cr with an atomic absorption spectro-photometer (Perkin Elmer, model 2380) after wet ashing with concentrated nitric and perchloric acids [19]. Phosphorus content was determined using a UV spectrophotometer at 400 nm (Bekhman, model DU-20). 2.5. Calculations and statistical analysis If not otherwise specied, the formula units are in percent. Chromic oxide was added to the diets as an indigestible marker. Apparent mineral digestibility was calculated by pengroup as follows: AD %
MIN

100

100

min

FECES

Cr

DIET

/ min

DIET

Cr

FECES)

(1)

Table 2 Effect of dietary treatment on pig growth performance Treatment 1 2 3 4 5 Control Low-P As 2 phytase As 3 acetic acid As 1 acetic acid SEM p-value Daily body weight gain (g) 289 229 296 350 313 14.9 0.0002 ADG 2 1 4 1 0.0169** 0.7719 0.0500* 0.3599 Daily feed intake (g) 479 501 508 530 502 23.3 0.6635 Feed efciency (feed:gain) 1.70 2.29 1.72 1.52 1.62 0.07 0.0001 FCR 0.0001*** 0.8488 0.0778 0.4491

Contrast trt trt trt trt 3 3 3 5 vs vs vs vs trt trt trt trt

*/**/*** Different levels of signicant differences.

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Where in formulae 1, ADMIN are the apparent digestibility of individual minerals in percent, [min]FECES is the mineral concentration in the faeces, [min]DIET is the mineral concentration in the diet, [Cr]DIET is the chromic oxide concentration in the diet and [Cr]FECES is the chromic oxide concentration in the faeces. The animal performance and apparent mineral digestibility data were analysed in the Statistical Analysis System [20] using the General Linear Models procedure. The performance data were analysed for effect of treatment, sex, pen, block and treatment sex. The digestibility data were analysed for effect of treatment, sex, block, pen, day of collection and the treatment x sex interaction. The contrasts, established a priori, among treatment means using t-test were: trt 1 vs trt 3, trt 1 vs trt 5, trt 2 vs trt 3, and trt 3 vs trt 4. The observed probability value (p) was taken as signicant at the 0.05 levels.

3. Results Proximate analysis of the experimental diets is showed in Table 1. The low P diet (trt 2) was estimated to contain 0.15% available P coming from di-Ca phosphate. Thus, assuming that a corn-soybean meal diet has 0.40% P, the total estimated P in the diet was 0.55%. However, laboratory analysis of feed revealed a lower actual value for total P (0.47%). 3.1. Performance parameters No signs of diarrhea or poor health due to trt effects were observed on the piglets throughout the study. The trt effects on ADG, ADFI and FCR are presented in Table 2. Average daily gain (g/d) was signicantly different among trts (p 0.0003) with mean values of 289, 229, 296, 350, and 313, for trts 1 to 5, respectively. Piglets fed the low-P diet supplemented with phytase (trt. 3) responded with a greater growth (p 0.01) and better feed conversion (p 0.0001) than pigs fed the low-P diet (trt 2). There was no signicant difference (p 0.77) in daily gain between pigs fed the control diet (trt 1) and pigs fed the low-P diet supplemented with phytase (trt 3). An increased ADG (p 0.05) was obtained by piglets fed the low-P diet supplemented with phytase plus acetic acid (trt 4) when compared to those animals fed low P plus phytase alone. Overall, piglets in trt 4 obtained the best growth performance among trts. Although some improvement was recorded, there was no signicant difference (p 0.3) in growth rate between animals that were offered either the control diet (trt.1) or the control diet plus acetic acid (trt.5) for the rst two weeks after weaning. Average daily feed intake was not signicantly affected (p 0.6) by Pi level or by supplementation with either phytase or acetic acid. Feed conversion ratio followed the trend of the ADG. The mean FCR values were 1.70, 2.29, 1.72, 1.52, and 1.62 (p 0.0001) for trts 1 to 5, respectively. A signicant improvement (p 0.0001) in the FCR was observed when the low-P diet was supplemented with phytase and plus acetic acid. Even though piglets fed the low-P diet (trt 2) had an ADFI (501g/d) similar to the animals in trts 3 and 5 (508 g/d and 502 g/d respectively), their ADG and FCR were much lower. Animals fed the low-P diet supplemented with phytase plus acetic acid showed the best FCR (Table 2).

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Table 3 Effect of dietary treatment on apparent digestibility (%) of dry matter, protein and minerals Description Dietary Treatment 1 Number of observations Dry matter Protein Phosphorus Calcium Iron Zinc Cooper Magnanese Contrasts Item Protein Dry matter Phosphorus Calcium Iron Zinc Copper Manganese Trt 1 vs Trt 3 0.0008*** 0.0017*** 0.0001*** 0.0287* 0.9828 0.0032*** 0.0746 0.0012*** Trt 2 vs Trt 3 0.7986 0.6600 0.0001*** 0.0199** 0.1832 0.1294 0.6287 0.4644 Trt 3 vs Trt 4 0.3102 0.1590 0.0130*** 0.0203* 0.0517* 0.1462 0.0448* 0.1095 Trt 1 vs Trt 5 0.0047*** 0.0067*** 0.0002*** 0.2875 0.0390* 0.0015*** 0.0023*** 0.0053*** 26 83.42 79.42 48.01 55.93 20.21 0.69 3.73 11.40 2 26 85.55 83.44 41.48 55.46 26.55 5.55 3.37 0.37 3 26 85.84 83.72 60.18 62.32 20.31 11.09 1.91 1.55 4 26 86.80 84.93 66.21 69.15 29.70 16.35 8.33 2.77 5 26 85.43 82.91 58.33 58.88 30.24 12.29 6.86 3.77 SEM 0.48 0.79 1.59 1.95 3.27 2.52 2.16 1.87

*/**/*** different levels of signicant differences.

3.2. Apparent digestibility of minerals The dietary trt effects on the apparent digestibility of the dry matter, protein, P, Ca, Fe, Zn, Cu, and Mn are presented in Table 3. The apparent digestibility of dry matter ranged from 83.4% to 86.8%. It was signicantly improved when comparing trts 1 vs 3 and 1 vs 5. The apparent protein digestibility values were: 79.4%, 83.4%, 83.7%, 84.9%, and 82.9% for trts 1 to 5, respectively. It was signicantly (p 0.005) improved in trts 3 and 5 when the comparisons against the control diet were made. The results shown in the table indicate that the apparent digestibility of P ranged from a low 41.5% of trt 2 to a high 66.2% of trt 4 supplemented with phytase plus acetic acid. The apparent digestibility of P increased (p 0.0001) when phytase was added to the low P diet. By adding acetic acid (trt 3) the P digestibility was further increased (p 0.01). The addition of acetic acid alone to the control diet, also signicantly improved (p 0.0002) the apparent digestibility of P from 48% to 58%. The apparent digestibility of Ca ranged from a low 55% in trt 2 to a high 69% in trt 4 and followed the trend of P digestibility. Calcium digestibility was not improved by adding acetic acid to the control diet. Consequently, the addition of phytase or phytase plus acetic acid resulted in a relative reduction of both, P and Ca faecal excretion. Phytase reduced faecal excretion of P by 44% and Ca by 12%, while acetic acid supplementation reduced this faecal excretion further up to 51% and 28% for P and Ca, respectively (Table 4). This signicant reduction in mineral faecal excretion promoted by acetic acid supplementation in

Z. Valencia, E.R. Chavez / Nutrition Research 22 (2002) 623 632 Table 4 Effect of dietary treatment on apparent P and Ca absorption and fecal excretion Description Units Dietary Treatment 1 Feed Consumption Phosphorus in diets Phosphorus intake Apparent P absorption P excretion Relative excretion Calcium in diet Calcium intake Apparent Ca absorption Ca excretion Relative excretion (g/d) (%) (g/d) (g/d) (g/d) (%) (%) (g/d) (g/d) (g/d) (%) 479 0.69 3.31 1.59 1.72 1.00 0.97 4.65 2.60 2.05 100 2 501 0.47 2.35 0.98 1.38 80.2 0.92 4.61 2.56 2.05 100 3 508 0.48 2.44 1.47 0.97 56.5 0.94 4.78 2.98 1.80 87.9 4 530 0.47 2.49 1.65 0.84 49.0 0.90 4.77 3.30 1.47 71.9 5

629

502 0.66 3.31 1.93 1.38 80.3 0.93 4.67 2.75 1.92 93.8

the presence of phytase was also observed when the control diet was supplemented only with acetic acid. In this trial phytase supplementation to the low-P diet (trt3), did not affect (p 0.1) the apparent digestibility of trace minerals considered, namely Fe, Zn, Cu, and Mn. However, when acetic acid was added to the low-P phyatse supplemented diet (trt4), the apparent digestibility of P, Ca, Fe and Cu were signicantly (p 0.05) increased. Moreover, acetic acid supplemented to the control diet for the rst two weeks after weaning promoted a highly signicant improvement in the digestibility of most trace minerals considered on this study (Table 3). Calcium was the only exception. The results of the P and Ca balance were consistent with the signicant improvement on the apparent digestibility pf these two macro-elements (Table 4). Phytase increased the retention of dietary P from 41% to 60% and the addition of acetic acid increased it to 66%. Thus, in comparative terms the piglets fed the low-P diet supplemented with phytase plus acetic acid grew faster and excreted 51% less P and 28% less Ca into the manure than the group fed with the control diet. The addition of only acetic acid to the control diet also reduced the excretion of dietary P by 20% and Ca excretion by 6% (Table 4).

4. Discussion There are several physiological restrictions on early-weaned piglets when switching from the sows milk to the new feeding regimen. Immediately after weaning, the digestive system of the pig has to adapt to solid feed with respect to pH, enzyme secretion and gut motility [21]. One of the most important factors in causing digestive disturbance is the low activity of proteolytic enzymes [22]. It is related to the development of the gastric pH of the weaned pig which is low in acidity for the rst 40 days [3]. Consequently, proteins of plant origin are digested to a lesser extent than milk proteins, resulting in poor performance when fed to newly-weaned piglets [2324]. Improvements in post-weaning growth rate and efciency of feed utilization have been achieved by supplementing starter diets with fumaric or citric acids [12,25].

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Average daily gain was signicantly improved by supplementing either phytase (trt 3) or phytase plus acetic acid (trt 4) to the low-P diet (trt 2). This was an indication of increased availability of dietary P. Average DFI was not different among treatments. This suggests that the improved gain occurred as a result of improved efciency at similar intakes (Table 2). The improved performance observed in pigs receiving the low P diet supplemented with phytase plus acetic acid would seem to support the hypothesis that the young weanling pig may be unable to adequately digest certain nutrients. Due to the lack of adequate levels of hydrochloric acid in the stomach [26] it may limit the activation of pepsinogen, and hence limit protein utilization. It seems also, that the lower gastric pH enhanced phytase activity (which is pH dependent) since there was not only a further increase in P and Ca digestibility but also a signicant increase on the apparent digestibility of Cu and Fe (trt 4). It appears that by increasing the acidity of the digesta with acetic acid, a weak acid, it does supplement the low level of hydrochloric acid being secreted or perhaps stimulates its secretion. Consequently, the pig seems to be more capable of making the necessary digestive adjustments at weaning to avoid the growth lag normally observed at this time. Phytase supplementation signicantly improved the ADG and FCR of a low P diet (trt 2), and there was no signicant difference in ADG and FCR between a low-P diet supplemented with phytase and the control diet that had the NRC [[16] recommended level of available P. There were no differences in ADFI but there was a signicant increase in the FCR when comparing the low P diet (trt 2) and the same diet supplemented with phytase (trt 3). Similar results were found by Jongbloed et al [26] and Yi et al [27]. Supplementing the control diet with acetic acid and offering it to piglets for two weeks after weaning had no effect on the ADG or FCR (Table 2). The apparent digestibility of dry matter, protein and minerals (except Ca) were signicantly increased by the addition of acetic acid to the control diet (Table 3), but this was not reected in an increased growth rate. Radecki et al [13], obtained similar results when supplementing a corn-soybean meal diet with citric acid and offered it to 4-week old pigs. Kirchgessner and Roth [14] reported an improvement in protein digestibility when the diet was acidied with fumaric acid. In a review on the effects of organic acid supplementation in weanling pigs, Gabert and Sauer [28] reported that the ADG may be increased by up to 26.9% or decreased by up to 15%. In this study, an additional 18% increase in growth rate was observed when both acetic acid and phytase (trt 4) were added to the low-P plus phytase diet. The increase in apparent digestibility and absorption of P, Ca, Fe, Zn, Cu, and Mn could be the primary reason of a better growth performance observed when both, phytase and acetic acid were supplemented. Previously, the addition of microbial phytase to plant based diets has been shown to improve the utilization of Ca, Zn and Cu in pig diets [29]. Kirchgessner and Roth [14] reported that fumaric acid supplementation increased the apparent faecal digestibility of Ca and P, as well, as Ca and P retention. This could be explained by the growth-promoting effects of phytase and acetic acid on speeding up the digestion in the small intestine. This means, that more nutrients are made available for absorption in the upper parts of the small intestine and less material is thus available for microbial fermentation in the hindgut, which leads to less energy losses [30]. It was interesting to observe that acetic acid supplementation did not alter the apparent digestibility

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of Ca in the control diet but did improve it on the low-P diet supplemented with phytase that may suggest that Ca in the control diet was also tight up by phytate. In conclusion, dietary phytase supplementation enhanced the utilization of P from plant sources, also improving growth rate and nutrient utilization of feeds. In addition, it reduced the need for expensive P supplementation in the feed, reduced P excretion and thus, reduced P pollution of the environment. The combined action of phytase and acetic acid in the ration gave further boosts to growth and feed efciency by improving the digestibility and absorption of most nutrients measured, resulting in further reduction of faecal mineral excretion and thus allowing a further reduction of environmental pollution. Phosphorus excretion can be reduced with estimates ranging from 25 to 50%, by the addition of phytase to the diet [11]. The present study found a 44% reduction of P excretion with phytase supplementation and it was further reduced to 51% when acetic acid was also supplemented.

Acknowledgments Z. Valencia was the principal researcher of this study as part of her M.Sc. thesis requirements for the degree under the direction and supervision of Dr. E.R. Chavez. We greatly appreciate the assistant of Mr Luc Normand for his help in handling the animals during the trial.

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