Research Article Received: 5 October 2014
Revised: 3 April 2015
Accepted article published: 15 April 2015
Published online in Wiley Online Library: 26 May 2015
(wileyonlinelibrary.com) DOI 10.1002/jsfa.7217
Effect of fermented medicinal plants on growth performance, nutrient digestibility, fecal noxious gas emissions, and diarrhea score in weanling pigs Pinyao Zhao,a Hanlin Li,a Yan Lei,a Tianshui Li,a Sunki Kimb and Inho Kima* Abstract BACKGROUND: Antibiotics used as growth promoters in livestock have been banned in the European Union since 2006. Alternatives of antibiotics have focused on phytogenic plants, such as herbs and medicinal plants. No studies have evaluated the use of fermented medicinal plants (FMP) made up of Gynura procumbens, Rehmannia glutinosa and Scutellaria baicalensis in weanling pigs. Therefore, an experiment was conducted to determine the effects of FMP on growth performance, nutrient digestibility, fecal noxious gas emissions and diarrhea score in weanling pigs. RESULTS: FMP supplementation increased (P < 0.05) average daily gain, average daily feed intake, gain:feed, apparent total tract digestibility of dry matter, nitrogen and gross energy compared with NC treatment, while a linear effect (P < 0.05) was observed on those criteria. Ammonia, total mercaptans and hydrogen sulfide concentrations were decreased (P < 0.05) by the supplementation of FMP compared with NC. Additionally, diarrhea score was lower (P < 0.05) by FMP addition compared with NC during days 0–7 and days 8–14. CONCLUSION: These results suggested that FMP could be used as an alternative to antibiotics by enhancing growth performance and nutrient digestibility, and decreasing fecal noxious gas emission and early diarrhea score of weanling pigs. © 2015 Society of Chemical Industry Keywords: diarrhea score; fecal noxious gas emission; fermented medicinal plants; growth performance; nutrient digestibility; weanling pig
INTRODUCTION
J Sci Food Agric 2016; 96: 1269–1274
herbs increase the intestinal absorption rate and bioavailability of the active compounds in medicinal herbs.17 Fermentation of medicinal herbs such as red ginseng and gumiganghwal-tang by microorganisms improved their pharmacological efficacy and neuroprotective activity by changing the contents of constituents of medicinal herbs.18,19 Ng et al.20 demonstrated that lactic acid bacterial fermentation could increase the antioxidant activity and reducing power of herbal plants. Min et al.21 reported that replacing soy protein concentrate with fermented soy protein appeared beneficial in growth performance, N and amino acid digestibility during the first 7 days after weaning. However, to the best of our knowledge, many studies have been conducted to evaluate the use of medicinal plants, but not enough studies have thus far concentrated on fermented medicinal plants (FMP) as an alternative to antibiotics in weanling pigs. The principal objective
∗
Correspondence to: Inho Kim, Department of Animal Resource and Science, Dankook University, Cheonan, Chungnam 330–714, South Korea. E-mail: [email protected]
a Department of Animal Resource and Science, Dankook University, Cheonan, Chungnam, 330-714, South Korea b Sunbio Ltd, Cheonan, Chungnam, 330-714, South Korea
www.soci.org
© 2015 Society of Chemical Industry
1269
Antibiotics used as feed additives for livestock have been completely banned in the European Union since 2006. Researchers have focused on developing antibiotic growth promoter alternatives to stabilize the health and growth performance in livestock. Phytogenic feed additives comprise a wide variety of herbs, spices and products derived therefrom, and are mainly essential oils. They have been widely used in recent decades as an alternative to antibiotics owing to their plant-derived properties and growth-promoting effects.1 – 4 Gynura procumbens is a well-known tropical Asian medicinal herbal plant, containing flavonoid sterol and steroid, as well as some proteins.5 Rehmannia glutinosa has been extensively used in traditional Chinese medicine to treat rheumatoid arthritis, asthma, urticaria and chronic nephritis.6 Some Scutellaria baicalensis is used to treat neurological disorders, cancer, inflammatory diseases, and viral and bacterial infections.7 Herbal extracts have been observed to change the intestinal microbiota,8 increase digestibility and nutrient absorption,9 enhance nitrogen absorption10 and improve the immune response11 in poultry and pigs. Moreover, previous studies indicated that essential oils may contribute to the improvement of pig performance,12,13 nutrient digestibility,14 immune status15 and the intestinal ecosystem.16 Fermented
www.soci.org of this study was to evaluate the effect of FMP on growth performance, nutrient digestibility, fecal noxious gas emissions and diarrhea score in weanling pigs.
MATERIALS AND METHODS The experimental protocol used in this study was approved by the Animal Care and Use Committee of Dankook University. Preparation of fermented medicinal plants The solid culture of the FMP product used in this study contained a mixture of lactic acid bacteria at 3.0 × 108 CFU g−1 (Lactobacillus plantarium species), yeasts at 7.5 × 107 CFU g−1 (Saccharomyces cerevisiae species), and fermenting bacteria at 8.0 × 108 CFU g−1 (Bacillus licheniformis species). The FMP were prepared as follows: the upper leaves of the Gynura procumbens plant were taken and thoroughly washed, and then cut to approximately 1 × 1 cm pieces. Processed leaves were mixed with L. plantarum inoculants, which were previously fermented with soybean meal (1:1, w/w). The fermentation condition of L. plantarum had estimated 40% moisture for 2 days at 30 ∘ C under solid-state fermentation (SSF). Rehmannia glutinosa roots were thoroughly washed and inoculated with 5% (v/w) S. cerevisiae which were fermented with soybean meal (1:1, w/w) for 2 months at low temperature, and then kept for 5 days at 25 ∘ C under SSF. Scutellaria baicalensis roots were thoroughly washed and inoculated with 5% (v/w) B. licheniformis, which were fermented with soybean meal (1:1, w/w) for 2 months at low temperature, and then kept for 5 days at 30 ∘ C under SSF.
P Zhao et al.
until analysis. Before chemical analysis, fecal samples were thawed at 57 ∘ C for 72 h, after which they were ground to pass through a 1 mm screen. All feed and fecal samples were analyzed for DM (method 930.15, AOAC, 2007)19 and crude protein (method 990.03, AOAC, 2007).24 Chromium was analyzed via UV absorption spectrophotometry (model UV-1201, Shimadzu, Kyoto, Japan). The GE was determined by measuring the heat of combustion in the samples using a Parr 6100 oxygen bomb calorimeter (Parr Instrument Co., Moline, IL, USA). The ATTD was calculated using the following formula: ) ( )]} { [( Digestibility = 1 − Nf × Cd ∕ Nd × Cf
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Experimental design, animals and housing A total of 150 weanling pigs [(Yorkshire × Landrace) × Duroc] with an average body weight (BW) of 6.08 ± 0.81 kg (21 days of age) were used in a 42-day experiment. Pigs were randomly allotted to five experimental diets according to their initial BW and sex (five replications; three gilts and three barrows per pen). Dietary treatment groups were: (i) negative control (NC), free antibiotics diet; (ii) positive control (PC), NC + 33 ppm apramycin; (iii) FMP05, NC + 0.5 g FMP kg−1 ; (iv) FMP10, NC + 1.0 g FMP kg−1 ; and (v) FMP20, NC + 2.0 g FMP kg−1 . The diets were formulated to meet or exceed the NRC (2012)22 nutrient requirements (Table 1). Treatment additives were included in the diet by replacing the same amount of corn. All the pigs were housed in an environmentally controlled room with a slatted plastic floor. Each pen was equipped with a one-sided self-feeder and a nipple waterer to allow the pig ad libitum access to feed and water throughout the experimental period. Temperature during week 1 was maintained at 32 ∘ C and lowered by 2.5 ∘ C each week thereafter.
where Nf = nutrient concentration in feces (g kg−1 DM), Nd = nutrient concentration in diet (g kg−1 DM), C d = chromium concentration in diet (g kg−1 DM), and C f = chromium concentration in feces (g kg−1 DM). Fresh feces and urine samples were collected randomly from at least two pigs in each pen on days 14 and 42 of the experiment. The urine was collected in a bucket via a funnel below the cage. Samples were kept in sealed containers and were immediately stored at −4 ∘ C for the duration of the period. After the collection period, feces and urine samples were pooled and each mixed well for each pen. As described by Zhao et al.,25 subsamples of slurry (150 g feces and 150 g urine were mixed well; 1:1 on a wet weight basis) were taken and stored in 2.6 L plastic boxes in duplicate. Each box had a small hole in the middle of one side wall, which was sealed with adhesive plaster. The samples were permitted to ferment for 7 days at room temperature (25 ∘ C). The concentrations of gas were determined at the end of the fermentation period. A gas sampling pump (model GV-100; Gastec Corp., Ayase, Japan) was utilized for gas detection (Gastec detector tube No. 3La for ammonia (NH3 ), No. 4LK for hydrogen sulfide (H2 S) and No. 70 for mercaptans; Gastec Corp.). Prior to the measurements, slurry samples were shaken manually for approximately 30 s to disrupt any crust formation on the surface of the slurry sample and to homogenize them. The adhesive plasters were punctured, and 100 mL of headspace air was sampled approximately 2.0 cm above the slurry surface. Two samples from each pen were measured and then the average was calculated. Subjective diarrhea scores were recorded daily from day 0 to 42 by the same person and were based on the following: 1 = hard, dry pellets in a small, hard mass; 2 = hard, formed stool that remains firm and soft; 3 = soft, formed, and moist stool that retains its shape; 4 = soft, unformed stool that assumes the shape of the container; 5 = watery, liquid stool that can be poured. Scores were recorded on a pen basis following observations of individual pig and signs of stool consistency in the pen. The score is reported as average daily diarrhea of individual pig score.
Sampling and measurements Individual pig BW was recorded at the beginning, on day 14, and at the end (day 42) of the experimental period, and feed consumption was recorded on a pen basis during the experiment to calculate average daily gain (ADG), average daily feed intake (ADFI) and gain:feed (G:F). From day 8 to 14 and from day 36 to 42, pigs were fed diets mixed with chromic oxide (2 g kg−1 ) as an indigestible marker for the determination of apparent total tract digestibility (ATTD) for dry matter (DM) and nitrogen (N).23 On days 14 and 42, fecal samples were collected from at least two pigs in each pen via rectal massage. All feed and fecal samples were stored at −20 ∘ C
Statistical analysis All experimental data were analyzed using the GLM procedure of SAS as a completely randomized block design (SAS Inst. Inc., Cary, NC, USA). The pen was used as the experimental unit. Orthogonal contrasts were used to test the overall effect of FMP supplementation (NC vs. FMP) and the effect of the difference between effect of PC and FMP (PC vs. FMP). Additionally, orthogonal comparison was conducted using polynomial regression to measure the linear and quadric effects of increasing concentration of FMP. Variability in the data was expressed as SEM, and a probability level of P < 0.05 was considered significant.
wileyonlinelibrary.com/jsfa
© 2015 Society of Chemical Industry
J Sci Food Agric 2016; 96: 1269–1274
Effect of fermented medicinal plants in weanling pigs
www.soci.org
Table 1. Compositions of basal weanling pig diets (as-fed basis)a
Items
Phase 1 Phase 2 (days 0–14) (days 15–42)
Ingredient (g kg−1 ) Extruded corn 368.0 Fermented oat 60.0 Soybean meal (440 g kg−1 crude protein) 267.2 Fish meal 50.0 Soybean oil 50.0 Lactose 60.0 Whey powder 100.0 Dicalcium phosphate 12.0 Sugar 20.0 L-Lys · HCl (780 g kg−1 ) 2.5 DL-Met (500 g kg−1 ) 1.5 L-Thr (890 g kg−1 ) 0.8 1.0 Vitamin premixb 2.0 Mineral premixc Limestone 3.0 Salt 2.0 Calculated composition (g kg−1 ) 3, 480 Metabolizable energy (kcal kg−1 ) Crude protein 217.8 Lysine 14.8 Methionine 5.2 Calcium 8.1 Total phosphorus 7.1 Analyzed composition (g kg−1 ) Crude protein 215.5 Lysine 14.4 Methionine 5.1 Calcium 7.8 Total phosphorus 7.3
492.8 60.0 289.2 20.0 38.0 – 73 15.0 – 1.5 1.5 – 1.0 2.0 4.0 2.0 3, 413 215.8 13.2 5.0 7.9 7.0 213.0 13.1 4.7 7.5 6.8
a Dietary treatments: NC, negative control, basal diet; PC, positive control, NC + 33 ppm apramycin; FMP05 = NC + 0.5 g FMP kg−1 ; FMP10 = NC + 1.0 g FMP kg−1 ; and FMP20 = NC + 2.0 g FMP kg−1 . b Provided per kilogram of complete diet: vitamin A, 11 025 IU; vitamin D3 , 1103 IU; vitamin E, 44 IU; vitamin K, 4.4 mg; riboflavin, 8.3 mg; niacin, 50 mg; thiamine, 4 mg; d-pantothenic, 29 mg; choline, 166 mg; and vitamin B12 , 33 μg. c Provided per kilogram of complete diet: Fe (as FeSO .7H O), 80 mg; 4 2 Cu (as CuSO4 .5H2 O), 12 mg; Zn (as ZnSO4 ), 85 mg; Mn (as MnO2 ), 8 mg; I (as KI), 0.28 mg; and Se (as Na2 SeO3 .5H2 O ), 0.15 mg.
RESULTS Growth performance During days 0–14, the supplementation of FMP improved ADG (P1 = 0.021 and linear, P = 0.019) and ADFI (P1 = 0.036 and linear, P = 0.031) (Table 2). During days 15–42, the supplementation of FMP improved ADG (P1 < 0.001, P2 = 0.012, and linear, P < 0.001), ADFI (P1 < 0.001, P2 = 0.030, and linear, P = 0.004, quadratic, P = 0.017), and G:F (P1 = 0.001, P2 = 0.032, and linear, P < 0.001). During the overall period, FMP supplementation improved ADG (P1 < 0.001, P2 = 0.020 and linear, P < 0.001), ADFI (P1 = 0.002 and linear, P = 0.003), and G:F (P1 = 0.03 and linear, P = 0.003).
J Sci Food Agric 2016; 96: 1269–1274
Noxious gas emission and diarrhea score FMP supplementation reduced ammonia (P1 = 0.011 for day 14, P1 < 0.001 and linear, P = 0.006 for day 42), hydrogen sulfide (P1 = 0.002, P2 = 0.043, and linear, P = 0.004 for day 42), and total mercaptans (P1 = 0.019 for day 14, P1 < 0.001, and linear, P = 0.004 for day 42) emissions (Table 4). Diarrhea score was decreased during days 0–7 (P1 < 0.001 and linear, P1 < 0.001) and days 8–14 (P1 = 0.025) (Table 5). No effect was observed on diarrhea score during days 22–28, 29–35 and 36–42 by the supplementation of FMP.
DISCUSSION Growth performance There is currently no information on the influence of FMP in diets fed to pigs, and thus no comparisons could be made with other studies. However, there have been some studies regarding phytogenic feed additives such as medicinal herbs and plant extracts. A previous study reported that both 1.0 and 3.0 g kg−1 Chinese medicinal herb supplementation composed of Panax ginseng, Dioscoreaceae batatas, Atractylodes macrocephala, Glycyrrhiza uralensis, Ziziphus jujuba and Platycodon grandiflorum improved ADG and decreased feed:gain during the first 2 weeks after weanling.26 Kwon et al.27 reported that with increasing addition of medicinal plant mixtures (0, 0.5, 1.0 and 1.5 g kg−1 Artemisia, Acanthopanax and garlic mixtures) a linear increased ADG, G:F and decreased ADFI were observed in weanling pigs from day 0 to 20. Oetting et al.9 observed that high dietary level of herbal extract showed better results on pig performance. Additionally, Yan et al.28 demonstrated that during the first week weanling pigs fed 0.25 g kg−1 herb extract mixture (buckwheat, thyme, curcuma, black pepper and ginger) supplemented diets led to a lower ADFI but greater G:F. Furthermore, dietary herb extract mixture supplementation increased ADG and G:F over 4–6 weeks. In our study, the supplementation of 0, 0.5, 1.0 and 2.0 g kg−1 fermented medicinal plants linearly increased ADG, ADFI and G:F during days 0–14, days 15–42 and the overall period. There was also a quadratic effect on ADFI during days 15–42. Those phytogenic feed additives mentioned above have beneficial effects on growth performance in weanling pigs at different dosages. Lin et al.29 and Czech et al.30 confirmed that herbs or herbal by-products could be used to stimulate growth performance of pigs, and beneficial effects of herbal extracts may be due to activation of feed intake and digestive secretion, immune stimulation, antibacterial, antiviral and antioxidant properties.31 – 35 Furthermore, herb additives could improve the flavor and palatability of feed, and subsequently increase the total feed intake, which would contribute to the increased growth performance.1,36 Nutrient digestibility In our study, the ATTD of DM, N, and GE increased in pigs fed FMP diets compared with those fed basal diet, and linear effects were observed as well. This indicated that FMP could improve nutrient digestibility in weanling pigs and higher digestibility could be achieved with higher concentration of FMP. Similar results were also reported in previous studies. Yan et al.28 noted that weanling
© 2015 Society of Chemical Industry
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Nutrient digestibility FMP supplementation improved ATTD of DM (day 14, P1 = 0.017 and linear, P = 0.012; day 42, P1 = 0.012 and linear, P = 0.012), N (day 14, P1 = 0.048 and linear, P = 0.049; day 42, P1 = 0.048 and
linear, P = 0.039), and GE (day 14, P1 = 0.046 and linear, P = 0.045; day 42, P1 = 0.027 and linear, P = 0.042) (Table 3). There was no effect on nutrient digestibility between PC and FMP addition treatments.
www.soci.org
P Zhao et al.
Table 2. Effect of fermented medicinal plants (FMP) supplementation on growth performance in weanling pigsa P-valueb Items Days 0–14 ADG (g) ADFI (g) G:F Days 15–42 ADG (g) ADFI (g) G:F Days 0–42 ADG (g) ADFI (g) G:F
PC
FMP05
FMP10
FMP20
SE
P1
P2
297 360 0.824
303 367 0.827
307 370 0.831
309 373 0.831
539 820 0.657
558 828 0.674
565 825 0.685
458 667 0.688
473 674 0.702
479 673 0.712
Linear
Quadratic
7 7 0.018
0.021 0.036 0.527
0.244 0.250 0.797
0.019 0.031 0.540
0.476 0.553 0.815
563 824 0.683
7 2 0.009
Revised: 3 April 2015
Accepted article published: 15 April 2015
Published online in Wiley Online Library: 26 May 2015
(wileyonlinelibrary.com) DOI 10.1002/jsfa.7217
Effect of fermented medicinal plants on growth performance, nutrient digestibility, fecal noxious gas emissions, and diarrhea score in weanling pigs Pinyao Zhao,a Hanlin Li,a Yan Lei,a Tianshui Li,a Sunki Kimb and Inho Kima* Abstract BACKGROUND: Antibiotics used as growth promoters in livestock have been banned in the European Union since 2006. Alternatives of antibiotics have focused on phytogenic plants, such as herbs and medicinal plants. No studies have evaluated the use of fermented medicinal plants (FMP) made up of Gynura procumbens, Rehmannia glutinosa and Scutellaria baicalensis in weanling pigs. Therefore, an experiment was conducted to determine the effects of FMP on growth performance, nutrient digestibility, fecal noxious gas emissions and diarrhea score in weanling pigs. RESULTS: FMP supplementation increased (P < 0.05) average daily gain, average daily feed intake, gain:feed, apparent total tract digestibility of dry matter, nitrogen and gross energy compared with NC treatment, while a linear effect (P < 0.05) was observed on those criteria. Ammonia, total mercaptans and hydrogen sulfide concentrations were decreased (P < 0.05) by the supplementation of FMP compared with NC. Additionally, diarrhea score was lower (P < 0.05) by FMP addition compared with NC during days 0–7 and days 8–14. CONCLUSION: These results suggested that FMP could be used as an alternative to antibiotics by enhancing growth performance and nutrient digestibility, and decreasing fecal noxious gas emission and early diarrhea score of weanling pigs. © 2015 Society of Chemical Industry Keywords: diarrhea score; fecal noxious gas emission; fermented medicinal plants; growth performance; nutrient digestibility; weanling pig
INTRODUCTION
J Sci Food Agric 2016; 96: 1269–1274
herbs increase the intestinal absorption rate and bioavailability of the active compounds in medicinal herbs.17 Fermentation of medicinal herbs such as red ginseng and gumiganghwal-tang by microorganisms improved their pharmacological efficacy and neuroprotective activity by changing the contents of constituents of medicinal herbs.18,19 Ng et al.20 demonstrated that lactic acid bacterial fermentation could increase the antioxidant activity and reducing power of herbal plants. Min et al.21 reported that replacing soy protein concentrate with fermented soy protein appeared beneficial in growth performance, N and amino acid digestibility during the first 7 days after weaning. However, to the best of our knowledge, many studies have been conducted to evaluate the use of medicinal plants, but not enough studies have thus far concentrated on fermented medicinal plants (FMP) as an alternative to antibiotics in weanling pigs. The principal objective
∗
Correspondence to: Inho Kim, Department of Animal Resource and Science, Dankook University, Cheonan, Chungnam 330–714, South Korea. E-mail: [email protected]
a Department of Animal Resource and Science, Dankook University, Cheonan, Chungnam, 330-714, South Korea b Sunbio Ltd, Cheonan, Chungnam, 330-714, South Korea
www.soci.org
© 2015 Society of Chemical Industry
1269
Antibiotics used as feed additives for livestock have been completely banned in the European Union since 2006. Researchers have focused on developing antibiotic growth promoter alternatives to stabilize the health and growth performance in livestock. Phytogenic feed additives comprise a wide variety of herbs, spices and products derived therefrom, and are mainly essential oils. They have been widely used in recent decades as an alternative to antibiotics owing to their plant-derived properties and growth-promoting effects.1 – 4 Gynura procumbens is a well-known tropical Asian medicinal herbal plant, containing flavonoid sterol and steroid, as well as some proteins.5 Rehmannia glutinosa has been extensively used in traditional Chinese medicine to treat rheumatoid arthritis, asthma, urticaria and chronic nephritis.6 Some Scutellaria baicalensis is used to treat neurological disorders, cancer, inflammatory diseases, and viral and bacterial infections.7 Herbal extracts have been observed to change the intestinal microbiota,8 increase digestibility and nutrient absorption,9 enhance nitrogen absorption10 and improve the immune response11 in poultry and pigs. Moreover, previous studies indicated that essential oils may contribute to the improvement of pig performance,12,13 nutrient digestibility,14 immune status15 and the intestinal ecosystem.16 Fermented
www.soci.org of this study was to evaluate the effect of FMP on growth performance, nutrient digestibility, fecal noxious gas emissions and diarrhea score in weanling pigs.
MATERIALS AND METHODS The experimental protocol used in this study was approved by the Animal Care and Use Committee of Dankook University. Preparation of fermented medicinal plants The solid culture of the FMP product used in this study contained a mixture of lactic acid bacteria at 3.0 × 108 CFU g−1 (Lactobacillus plantarium species), yeasts at 7.5 × 107 CFU g−1 (Saccharomyces cerevisiae species), and fermenting bacteria at 8.0 × 108 CFU g−1 (Bacillus licheniformis species). The FMP were prepared as follows: the upper leaves of the Gynura procumbens plant were taken and thoroughly washed, and then cut to approximately 1 × 1 cm pieces. Processed leaves were mixed with L. plantarum inoculants, which were previously fermented with soybean meal (1:1, w/w). The fermentation condition of L. plantarum had estimated 40% moisture for 2 days at 30 ∘ C under solid-state fermentation (SSF). Rehmannia glutinosa roots were thoroughly washed and inoculated with 5% (v/w) S. cerevisiae which were fermented with soybean meal (1:1, w/w) for 2 months at low temperature, and then kept for 5 days at 25 ∘ C under SSF. Scutellaria baicalensis roots were thoroughly washed and inoculated with 5% (v/w) B. licheniformis, which were fermented with soybean meal (1:1, w/w) for 2 months at low temperature, and then kept for 5 days at 30 ∘ C under SSF.
P Zhao et al.
until analysis. Before chemical analysis, fecal samples were thawed at 57 ∘ C for 72 h, after which they were ground to pass through a 1 mm screen. All feed and fecal samples were analyzed for DM (method 930.15, AOAC, 2007)19 and crude protein (method 990.03, AOAC, 2007).24 Chromium was analyzed via UV absorption spectrophotometry (model UV-1201, Shimadzu, Kyoto, Japan). The GE was determined by measuring the heat of combustion in the samples using a Parr 6100 oxygen bomb calorimeter (Parr Instrument Co., Moline, IL, USA). The ATTD was calculated using the following formula: ) ( )]} { [( Digestibility = 1 − Nf × Cd ∕ Nd × Cf
1270
Experimental design, animals and housing A total of 150 weanling pigs [(Yorkshire × Landrace) × Duroc] with an average body weight (BW) of 6.08 ± 0.81 kg (21 days of age) were used in a 42-day experiment. Pigs were randomly allotted to five experimental diets according to their initial BW and sex (five replications; three gilts and three barrows per pen). Dietary treatment groups were: (i) negative control (NC), free antibiotics diet; (ii) positive control (PC), NC + 33 ppm apramycin; (iii) FMP05, NC + 0.5 g FMP kg−1 ; (iv) FMP10, NC + 1.0 g FMP kg−1 ; and (v) FMP20, NC + 2.0 g FMP kg−1 . The diets were formulated to meet or exceed the NRC (2012)22 nutrient requirements (Table 1). Treatment additives were included in the diet by replacing the same amount of corn. All the pigs were housed in an environmentally controlled room with a slatted plastic floor. Each pen was equipped with a one-sided self-feeder and a nipple waterer to allow the pig ad libitum access to feed and water throughout the experimental period. Temperature during week 1 was maintained at 32 ∘ C and lowered by 2.5 ∘ C each week thereafter.
where Nf = nutrient concentration in feces (g kg−1 DM), Nd = nutrient concentration in diet (g kg−1 DM), C d = chromium concentration in diet (g kg−1 DM), and C f = chromium concentration in feces (g kg−1 DM). Fresh feces and urine samples were collected randomly from at least two pigs in each pen on days 14 and 42 of the experiment. The urine was collected in a bucket via a funnel below the cage. Samples were kept in sealed containers and were immediately stored at −4 ∘ C for the duration of the period. After the collection period, feces and urine samples were pooled and each mixed well for each pen. As described by Zhao et al.,25 subsamples of slurry (150 g feces and 150 g urine were mixed well; 1:1 on a wet weight basis) were taken and stored in 2.6 L plastic boxes in duplicate. Each box had a small hole in the middle of one side wall, which was sealed with adhesive plaster. The samples were permitted to ferment for 7 days at room temperature (25 ∘ C). The concentrations of gas were determined at the end of the fermentation period. A gas sampling pump (model GV-100; Gastec Corp., Ayase, Japan) was utilized for gas detection (Gastec detector tube No. 3La for ammonia (NH3 ), No. 4LK for hydrogen sulfide (H2 S) and No. 70 for mercaptans; Gastec Corp.). Prior to the measurements, slurry samples were shaken manually for approximately 30 s to disrupt any crust formation on the surface of the slurry sample and to homogenize them. The adhesive plasters were punctured, and 100 mL of headspace air was sampled approximately 2.0 cm above the slurry surface. Two samples from each pen were measured and then the average was calculated. Subjective diarrhea scores were recorded daily from day 0 to 42 by the same person and were based on the following: 1 = hard, dry pellets in a small, hard mass; 2 = hard, formed stool that remains firm and soft; 3 = soft, formed, and moist stool that retains its shape; 4 = soft, unformed stool that assumes the shape of the container; 5 = watery, liquid stool that can be poured. Scores were recorded on a pen basis following observations of individual pig and signs of stool consistency in the pen. The score is reported as average daily diarrhea of individual pig score.
Sampling and measurements Individual pig BW was recorded at the beginning, on day 14, and at the end (day 42) of the experimental period, and feed consumption was recorded on a pen basis during the experiment to calculate average daily gain (ADG), average daily feed intake (ADFI) and gain:feed (G:F). From day 8 to 14 and from day 36 to 42, pigs were fed diets mixed with chromic oxide (2 g kg−1 ) as an indigestible marker for the determination of apparent total tract digestibility (ATTD) for dry matter (DM) and nitrogen (N).23 On days 14 and 42, fecal samples were collected from at least two pigs in each pen via rectal massage. All feed and fecal samples were stored at −20 ∘ C
Statistical analysis All experimental data were analyzed using the GLM procedure of SAS as a completely randomized block design (SAS Inst. Inc., Cary, NC, USA). The pen was used as the experimental unit. Orthogonal contrasts were used to test the overall effect of FMP supplementation (NC vs. FMP) and the effect of the difference between effect of PC and FMP (PC vs. FMP). Additionally, orthogonal comparison was conducted using polynomial regression to measure the linear and quadric effects of increasing concentration of FMP. Variability in the data was expressed as SEM, and a probability level of P < 0.05 was considered significant.
wileyonlinelibrary.com/jsfa
© 2015 Society of Chemical Industry
J Sci Food Agric 2016; 96: 1269–1274
Effect of fermented medicinal plants in weanling pigs
www.soci.org
Table 1. Compositions of basal weanling pig diets (as-fed basis)a
Items
Phase 1 Phase 2 (days 0–14) (days 15–42)
Ingredient (g kg−1 ) Extruded corn 368.0 Fermented oat 60.0 Soybean meal (440 g kg−1 crude protein) 267.2 Fish meal 50.0 Soybean oil 50.0 Lactose 60.0 Whey powder 100.0 Dicalcium phosphate 12.0 Sugar 20.0 L-Lys · HCl (780 g kg−1 ) 2.5 DL-Met (500 g kg−1 ) 1.5 L-Thr (890 g kg−1 ) 0.8 1.0 Vitamin premixb 2.0 Mineral premixc Limestone 3.0 Salt 2.0 Calculated composition (g kg−1 ) 3, 480 Metabolizable energy (kcal kg−1 ) Crude protein 217.8 Lysine 14.8 Methionine 5.2 Calcium 8.1 Total phosphorus 7.1 Analyzed composition (g kg−1 ) Crude protein 215.5 Lysine 14.4 Methionine 5.1 Calcium 7.8 Total phosphorus 7.3
492.8 60.0 289.2 20.0 38.0 – 73 15.0 – 1.5 1.5 – 1.0 2.0 4.0 2.0 3, 413 215.8 13.2 5.0 7.9 7.0 213.0 13.1 4.7 7.5 6.8
a Dietary treatments: NC, negative control, basal diet; PC, positive control, NC + 33 ppm apramycin; FMP05 = NC + 0.5 g FMP kg−1 ; FMP10 = NC + 1.0 g FMP kg−1 ; and FMP20 = NC + 2.0 g FMP kg−1 . b Provided per kilogram of complete diet: vitamin A, 11 025 IU; vitamin D3 , 1103 IU; vitamin E, 44 IU; vitamin K, 4.4 mg; riboflavin, 8.3 mg; niacin, 50 mg; thiamine, 4 mg; d-pantothenic, 29 mg; choline, 166 mg; and vitamin B12 , 33 μg. c Provided per kilogram of complete diet: Fe (as FeSO .7H O), 80 mg; 4 2 Cu (as CuSO4 .5H2 O), 12 mg; Zn (as ZnSO4 ), 85 mg; Mn (as MnO2 ), 8 mg; I (as KI), 0.28 mg; and Se (as Na2 SeO3 .5H2 O ), 0.15 mg.
RESULTS Growth performance During days 0–14, the supplementation of FMP improved ADG (P1 = 0.021 and linear, P = 0.019) and ADFI (P1 = 0.036 and linear, P = 0.031) (Table 2). During days 15–42, the supplementation of FMP improved ADG (P1 < 0.001, P2 = 0.012, and linear, P < 0.001), ADFI (P1 < 0.001, P2 = 0.030, and linear, P = 0.004, quadratic, P = 0.017), and G:F (P1 = 0.001, P2 = 0.032, and linear, P < 0.001). During the overall period, FMP supplementation improved ADG (P1 < 0.001, P2 = 0.020 and linear, P < 0.001), ADFI (P1 = 0.002 and linear, P = 0.003), and G:F (P1 = 0.03 and linear, P = 0.003).
J Sci Food Agric 2016; 96: 1269–1274
Noxious gas emission and diarrhea score FMP supplementation reduced ammonia (P1 = 0.011 for day 14, P1 < 0.001 and linear, P = 0.006 for day 42), hydrogen sulfide (P1 = 0.002, P2 = 0.043, and linear, P = 0.004 for day 42), and total mercaptans (P1 = 0.019 for day 14, P1 < 0.001, and linear, P = 0.004 for day 42) emissions (Table 4). Diarrhea score was decreased during days 0–7 (P1 < 0.001 and linear, P1 < 0.001) and days 8–14 (P1 = 0.025) (Table 5). No effect was observed on diarrhea score during days 22–28, 29–35 and 36–42 by the supplementation of FMP.
DISCUSSION Growth performance There is currently no information on the influence of FMP in diets fed to pigs, and thus no comparisons could be made with other studies. However, there have been some studies regarding phytogenic feed additives such as medicinal herbs and plant extracts. A previous study reported that both 1.0 and 3.0 g kg−1 Chinese medicinal herb supplementation composed of Panax ginseng, Dioscoreaceae batatas, Atractylodes macrocephala, Glycyrrhiza uralensis, Ziziphus jujuba and Platycodon grandiflorum improved ADG and decreased feed:gain during the first 2 weeks after weanling.26 Kwon et al.27 reported that with increasing addition of medicinal plant mixtures (0, 0.5, 1.0 and 1.5 g kg−1 Artemisia, Acanthopanax and garlic mixtures) a linear increased ADG, G:F and decreased ADFI were observed in weanling pigs from day 0 to 20. Oetting et al.9 observed that high dietary level of herbal extract showed better results on pig performance. Additionally, Yan et al.28 demonstrated that during the first week weanling pigs fed 0.25 g kg−1 herb extract mixture (buckwheat, thyme, curcuma, black pepper and ginger) supplemented diets led to a lower ADFI but greater G:F. Furthermore, dietary herb extract mixture supplementation increased ADG and G:F over 4–6 weeks. In our study, the supplementation of 0, 0.5, 1.0 and 2.0 g kg−1 fermented medicinal plants linearly increased ADG, ADFI and G:F during days 0–14, days 15–42 and the overall period. There was also a quadratic effect on ADFI during days 15–42. Those phytogenic feed additives mentioned above have beneficial effects on growth performance in weanling pigs at different dosages. Lin et al.29 and Czech et al.30 confirmed that herbs or herbal by-products could be used to stimulate growth performance of pigs, and beneficial effects of herbal extracts may be due to activation of feed intake and digestive secretion, immune stimulation, antibacterial, antiviral and antioxidant properties.31 – 35 Furthermore, herb additives could improve the flavor and palatability of feed, and subsequently increase the total feed intake, which would contribute to the increased growth performance.1,36 Nutrient digestibility In our study, the ATTD of DM, N, and GE increased in pigs fed FMP diets compared with those fed basal diet, and linear effects were observed as well. This indicated that FMP could improve nutrient digestibility in weanling pigs and higher digestibility could be achieved with higher concentration of FMP. Similar results were also reported in previous studies. Yan et al.28 noted that weanling
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Nutrient digestibility FMP supplementation improved ATTD of DM (day 14, P1 = 0.017 and linear, P = 0.012; day 42, P1 = 0.012 and linear, P = 0.012), N (day 14, P1 = 0.048 and linear, P = 0.049; day 42, P1 = 0.048 and
linear, P = 0.039), and GE (day 14, P1 = 0.046 and linear, P = 0.045; day 42, P1 = 0.027 and linear, P = 0.042) (Table 3). There was no effect on nutrient digestibility between PC and FMP addition treatments.
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P Zhao et al.
Table 2. Effect of fermented medicinal plants (FMP) supplementation on growth performance in weanling pigsa P-valueb Items Days 0–14 ADG (g) ADFI (g) G:F Days 15–42 ADG (g) ADFI (g) G:F Days 0–42 ADG (g) ADFI (g) G:F
PC
FMP05
FMP10
FMP20
SE
P1
P2
297 360 0.824
303 367 0.827
307 370 0.831
309 373 0.831
539 820 0.657
558 828 0.674
565 825 0.685
458 667 0.688
473 674 0.702
479 673 0.712
Linear
Quadratic
7 7 0.018
0.021 0.036 0.527
0.244 0.250 0.797
0.019 0.031 0.540
0.476 0.553 0.815
563 824 0.683
7 2 0.009
