Exogenous Fibrolytic Enzymes Addition in Concentrate Ration of Lactating Nili Ravi Buffaloes: Effects on Milk Production and Diet Digestibility

Burhan Azam1, Muhammad Naeem Tahir2,*, Faisal Shahzad2, Abdul Ghaffar3, Ghulam Abbas4, Madiha Gohar5 and Saima1

1Department of Animal Nutrition, Faculty of Animal Production and Technology, University of Veterinary and Animal Sciences, Lahore 54000, Pakistan

2University College of Veterinary and Animal Sciences, The Islamia University of Bahawalpur, Bahawlpur 63100, Pakistan

3Department of Life Sciences, The Islamia University of Bahawalpur, Bahawlpur 63100, Pakistan

4Centre of Excellence in Marine Biology, University of Karachi, Karachi 75270, Pakistan

5Alltech International, Industrial Area, Sector I-9/3, Islamabad, Pakistan

ABSTRACT

Poor availability of good quality feedstuffs for the feeding of livestock has been identified as the major hurdles in the development of livestock sector in Pakistan. To improve the poor quality roughages, exogenous fibrolytic enzyme may play important role by enhancing digestibility of fibre. This study was carried out to evaluate the effects of varying level of fibrolytic enzyme in concentrate ration on dry matter (DM) intake, milk production and composition, and diet digestibility in Nili Ravi buffaloes. Four lactating Nili-Ravi buffaloes of 3rd and 4th parity and milk production of 8±2.3 kg/day were selected and offered a basal diet based on sorghum forage and concentrate ration supplemented with fibrolytic enzyme (Fibrozyme™, Alltech Inc. Company, USA) at four levels (T1 = 0, T2 = 10, T3 = 15 and T4 = 20 g/animal/day) in a 4 × 4 Latin Square change-over design with four experimental periods. Each period consisted of a 14-days enzyme supplementation period following a 3-day transition period without enzyme supplementation between consecutive experimental periods. The results showed a significant linear (P<0.001) increase in the intakes of DM, CP and NDF by the animals with increasing level of enzyme in the diet. The milk yield increased until an enzyme supplementation of 15 g/day (T3) and then dropped. The yield of milk protein, fat, solids not fat and lactose were not affected (P>0.05). Milk fat, solids not fat and lactose concentrations differed (P<0.001) among the treatments but did not show (P>0.05) any linear or quadratic trends with increasing level of enzyme in the diet. Milk protein concentration linearly (P<0.001) increased with increasing level of enzyme in the diets. It is concluded that fibrolytic enzyme addition in the diets of lactating Nili Ravi buffaloes improves the dry matter intake and diet digestibility which results in improved milk production.

Article Information

Received 11 July 2016

Revised 10 February 2017

Accepted 03 April 2017

Available online 12 July 2017

Authors’ Contribution

BA and S conceived the idea and conducted the experiments. MNT and FS analysed the data and wrote the manuscript. AG, GA and MG helped in writing and revising the manuscript.

Key words

Concentrate ration, Feed intake, Fibrozyme™, Lactating buffaloes, Plant cell wall.

DOI: http://dx.doi.org/10.17582/journal.pjz/2017.49.4.1359.1364

* Corresponding author: [email protected]

0030-9923/2017/0004-1359 $ 9.00/0

Copyright 2017 Zoological Society of Pakistan

Introduction

 

Fibre degradation in ruminants depends upon the type of feed and number of microbes present in rumen. The fibre from the tropical plants is more resistant to rumen degradation by microbes present in forestomach which produce enzymes to digest cellulose, hemicelluloses and lignin (Van Soest et al., 1978). Exogenous fibrolytic enzymes, in the form of feed additives are included in the diets of ruminants consuming tropical region plants worldwide and they are reported to have exerted variable effects on the digestibility of fibre and animal performance (Beauchemin et al., 2000; Dhiman et al., 2002). Fibrolytic enzymes used during ensiling improve fermentation by hydrolyzing the cell wall and digestibility of corn- (Colombatto et al., 2003) and Bermuda grass-silage (Dean et al., 2005).

Fibrozyme™ which is a fibrolytic enzyme have cellulases and xylanase like activities (Alltech Inc. Company, USA) and breaks beta 1-4 glucosidic linkages present in fibre, provides readily fermentable substrate to ruminal microbes (McAllister et al., 2001; Bowman et al., 2002) which results in increased microbial colonization by increasing their number (Nsereko et al., 2000), thus increasing the fibre degradation in the rumen and improving feed efficiency (Yang et al., 1999; Meale et al., 2014). In addition, exogenous polysaccharidases stay in small intestine for long time and maintain the digesting activity in that environment too (Morgavi et al., 2002). The purpose of the present research was to evaluate the effect of dietary addition of fibrolytic enzyme on the performance of dairy Nili Ravi buffaloes.

 

Table I.- Chemical composition of forage and concentrate ration.

Ingredients	Concentrate ration	Sorghum
Sorghum	---	100.0
Maize	8.0	---
Wheat bran	32.0	---
Maize gluten	20.0	---
Cotton Seed cake	22.0	---
Rape seed cake	3.0	---
Molasses	14.0	---
Mineral mixture	1.0	---
Chemical composition
Dry matter	88.0	36.1
Crude Protein	18.0	8.3
Ether Extract	2.9	5.6
Neutral detergent fibre	32.1	51.8
Ash	1.7	5.6

All values are presented as percentage of dry matter unless otherwise notified.

 

Materials and Methods

Animals, experimental design and diets

This research was conducted at Buffalo Research Institute (BRI), Bhunikey, Kasur, Pakistan. Four Nili-Ravi lactating buffaloes of the 3rd & 4th parity and 8±2.3 kg/day milk production were randomly selected from the herd maintained at BRI and designated to four experimental dietary treatments in a 4 × 4 Latin square change over design with four experimental periods. Each period consisted of a 14-days enzyme supplementation period following a 3-day transition period without enzyme supplementation between consecutive experimental periods. All animals were tied up and fed individually. Animals were de-wormed using Nilzan® Plus at the rate of 2.5 ml/10 kg body weight and vaccinated using Niab® HS™ oil based at the rate of 3-5 ml/animal before the initiation of the experiment. The live weight of the animals was measured fortnightly early in the morning using a digital weighing scale.

The basal diets consisted of sorghum forage which was provided ad libitum and the concentrate ration at a rate of 4 kg/animal/day. Fibrozyme™ (Alltech Inc., Nicholasville, KY, USA) was supplemented at a rate of 0 (T1), 10 (T2), 15 (T3) and 20 (T4) g/animal/day by mixing with the concentrate ration (Table I). According to the manufacturers, Fibrozyme is a fibrolytic enzyme powder preparation containing xylanase and cellulose activities (from Aspergillus niger and Trichoderma viride fermentation extract) with a cellulose and xylanase activity of 31.0 and 43.4 IU, respectively. Fresh, clean water was available all the time.

Sampling and chemical analyses

Feed, experimental diets and orts were sampled weekly, composited for each experimental treatment and analyzed for dry matter (DM), crude protein (CP), ether extract and ash according to AOAC (1999) and for neutral detergent fibre (NDF) according to Van Soest et al. (1991) at Nutrition Laboratory of BRI.

Forage intake was measured weekly as feed offered minus refusal collected. Milk yield was recorded twice a day at 0545 h and 1745 h. Representative milk samples were collected weekly from each animal, stored at -20oC, composted against each animal for morning and evening milkings and evaluated for milk composition (fat, protein, solids not fat and lactose) using LactiCheck™-01 RapiRead (Page and Pedesen International Ltd. USA).

Determination of digestibility of dry matter, crude protein and neutral detergent fibre

During last three days of each experimental period, total faeces were collected using polythene bags during 24 h and composited for each animal. Bags were covered from top to avoid any loss of moisture from faeces. Representative faecal samples were analyzed for DM, CP and NDF and digestibility for these nutrients, which is hereby denoted by X was calculated according to the following equation:

 

Statistical analyses

Data from the feeding trial were analyzed according to Latin Square Design using GLM procedure of MINITAB® (version 16.1.1.0). Each animal on a specific diet was considered as the experimental unit. The linear and quadratic effects of increasing level of fibrolytic enzyme in the concentrate ration were examined by replacing the qualitative variable diet in the model with the quantitative variable proportion of fibrolytic enzyme using Fitted Line Polynomial Regression Analysis in MINITAB® (version 16.1.1.0). Data are presented as mean±standard error of mean. The level of significance was set at P<0.05.

Following mathematical model was applied:

Yijkl = µ+Pi+Aj+Tk+Eijk

Where, Yijkl is dependent variable, µ is overall mean, Pi is effect of period i, Aj is effect of animal j, Tk is effect of treatment k and Eijk is residual error.

 

Results and Discussion

Effects on feed intake

The results showed a significant linear (P<0.001) increase in the intakes of DM, CP and NDF by the animals with increasing level of enzyme in the diet (Table II). The results of our study for increase in DM intake were similar to those of Atrian and Shahryar (2012) who observed higher DM intakes when the enzyme in liquid form was supplemented to beef cattle and also with those of Gaafar et al. (2010) in lactating buffaloes and Bowman et al. (2003) in lactating cows. However, the findings of current study are not similar with those of Dhiman et al. (2002) who found no effects of fibrolytic enzyme supplementation on DM intake in Holstein cows. A linear increase in DM intake of the buffaloes with the enzyme supplementation is attributed to the corresponding increases in the digestibility of all nutrients resulting in more digestible energy supply at rumen level.

Effects on milk production and composition

The milk yield (Table III) increased with enzyme supplementation of 15 g/day (T3) and then dropped. The yield of milk protein, fat, solids not fat and lactose were not affected (P>0.05). The results of current study are similar to the findings of Kung et al. (2002) who found increased milk production of cows fed on total mixed ration supplemented with fibrolytic enzymes and also with those of Shekhar et al. (2010) in Murrah buffaloes.

 

Table II.- Effects of increasing level of fibrolytic enzyme in concentrate rations on intake parameters in lactating Nili-Ravi buffaloes.

Item	Experimental diets1				SEM	P value (diet)	L	Q
Intake (kg/day)	T1	T2	T3	T4
Dry matter	19.22d	19.86c	20.28b	20.79a	0.283	<0.001	0.002	0.853
Crude protein	1.89d	1.94c	1.97b	2.01a	0.023	<0.001	<0.001	0.780
Neutral detergent fibre	9.25d	9.58c	9.79b	10.06a	0.145	<0.001	0.028	0.901

1T1, T2, T3 and T4 represent the diets containing 0, 10, 15 and 20 g/animal/day supplementation of exogenous fibrolytic enzymes in the concentrate ration. SEM, standard error of mean; L, linear effect of increasing level of fibrolytic enzyme addition in experimental diets; Q, quadratic effect of increasing level of fibrolytic enzyme addition in experimental diets.

 

Table III.- Effects of increasing level of fibrolytic enzyme in concentrate rations on milk yield and composition in lactating Nili-Ravi buffaloes.

Item	Experimental diets1				SEM	P value (diet)	L	Q
	T1	T2	T3	T4
Yield (kg/day)	8.49c	9.53b	9.98a	9.40b	0.968	<0.001	<0.001	<0.001
Milk protein	0.33c	0.37b	0.39a	0.37b	0.005	0.215	0.636	0.301
Milk fat	0.48d	0.54b	0.58a	0.50c	0.007	0.246	0.576	0.370
Solids not fat	0.83c	0.96b	1.01a	0.95b	0.012	0.117	0.616	0.228
Lactose	0.44c	0.51ab	0.53a	0.50b	0.006	0.149	0.663	0.237
Composition (%)
Milk protein	3.85a	3.91b	3.92b	3.94b	0.035	0.001	0.040	0.442
Milk fat	5.68ab	5.70ab	5.88a	5.41b	0.293	0.032	0.354	0.111
Solids not fat	9.82b	10.04a	10.07a	10.15a	0.113	<0.001	0.441	0.441
Lactose	5.23b	5.34a	5.31a	5.34a	0.048	<0.001	0.074	0.282

1T1, T2, T3 and T4 represent the diets containing 0, 10, 15 and 20 g/animal/day supplementation of exogenous fibrolytic enzymes in the concentrate ration. For abbreviations, see Table II.

 

The findings of current study are also similar to those of Dean et al. (2007) who fed low and high energetic diets with or without fibrolytic enzyme supplementation and determined that feeding high energetic diets along with fibrolytic enzyme increased milk production. However, our results of milk production also suggest that the animals’ production performance was improved to a certain level of enzyme supplementation i.e. T3 diet (15 g/kg diet DM) as depicted in Figure 1. Although dry matter intake kept on increasing with level of enzyme; this increased energy intake was not reflected by the milk production. It might be true that the surplus energy had been utilized to build up the body reserves.

 

 

Milk fat, solids not fat and lactose concentrations differed (P<0.001) among the treatments but did not show (P>0.05) any linear or quadratic trends with increasing level of enzyme in the diet. Milk protein concentration linearly (P<0.001) increased with increasing level of enzyme in the diets. The results of current trial agree with the findings of Dean et al. (2007) who examined the effects of fibrolytic enzyme on milk composition by mixing it in total mixed ration, silage, concentrate and fresh fodder. Animals fed on concentrate + enzyme produced more protein in milk while the animals fed on TMR + enzyme produced more fat and protein in milk. Also our results are in accordance with those of Miller et al. (2008) who determined the effects of adding different levels of liquid fibrolytic enzyme in lactating cows’ diets and observed that enzyme supplementation linearly increased milk protein and quadratically increased milk fat concentrations. The lactose concentration of milk remained unaltered across many studies (e.g. Mohamed et al., 2013; Shadmanesh, 2014) and did the same in our study.

Effects on diet digestibility

Table IV shows that the increasing level of enzyme in the diets had a positive linear (P<0.001) effect on the digestibility of DM (2%), CP (3%) and NDF (3%) at the final supplementation level of enzyme. Performance of dairy animals is determined by amount of digestible nutrient that it consumes each day; which is the product of intake and digestibility. The digestibility of forage is related to the content and digestibility of NDF (Mertens, 2009).

The results of current study for digestibility agree with the findings of Yang et al. (1999) and El-Kady et al. (2006) who observed positive effects of fibrolytic enzyme on digestibility in the diets of cows and buffaloes, respectively. Our results are also similar to those of Arriola et al. (2011) where the diets of lactating cows were supplemented with fibrolytic enzyme at low and high concentrate proportions and it was observed that supplemented high concentrate groups had higher digestibility compared to non-supplemented groups. Titi and Tabbaa (2004) studied the effect of cellulase on growth performance and diet digestibility and observed that digestibility of DM and NDF significantly increased with the fibrolytic enzyme supplementation. However, our results are not in accordance with those of Sutton et al. (2003) and Muwalla et al. (2007) who reported no effects of fibrolytic enzyme supplementation in the diets on productive performance of Holstein cows and growth performance of Awassi lambs, respectively.

 

Table IV.- Effects of increasing level of fibrolytic enzyme in concentrate rations on diet digestibility in lactating Nili-Ravi buffaloes.

Item	Experimental diets1				SEM	P value (diet)	L	Q
Digestibility (%)	T1	T2	T3	T4
Dry matter	71.31	71.43	72.98	74.23	1.145	<0.001	<0.001	0.008
Crude protein	52.91	53.03	55.96	57.67	2.232	<0.001	<0.001	0.038
Neutral detergent fibre	51.39	51.51	53.49	55.05	1.580	<0.001	<0.001	0.014

1T1, T2, T3 and T4 represent the diets containing 0, 10, 15 and 20 g/animal/day supplementation of exogenous fibrolytic enzymes in the concentrate ration. For abbreviations, see Table II.

 

Conclusions

 

Based on the results presented, it is concluded that increasing level of enzyme supplementation in the diets of lactating Nili Ravi buffaloes increases the amount of digestible nutrients intake; which is attributable to improved digestibility.

 

Acknowledgements

 

The authors are grateful to the Higher Education Commission of Pakistan who provided the full grant money for this research project.

 

Statement of conflict of interest

Authors have declared no conflict of interest.

 

References

 

AOAC, 1999. Official methods of analysis, 18th ed. Association of Official Analytical Chemists, Washington, DC, USA, pp. 38-39.

Arriola, K.G., Kim, S.C., Staples, C.R. and Adesogan, A.T., 2011. Effect of fibrolytic enzyme application to low- and high-concentrate diets on the performance of lactating dairy cattle. J. Dairy Sci., 94: 832-839. https://doi.org/10.3168/jds.2010-3424

Atrian, P. and Shahryar, H.A., 2012. Effect of Fibrolytic enzyme treated Alfalfa on performance of Holstein beef cattle. Eur. J. exp. Biol., 2: 270-273.

Beauchemin, K.A., Rode, L.M., Maekawa, M.M., Morgavi, D.P. and Kampen, R., 2000. Evaluation of a non starch polysaccharidase feed enzyme in dairy cow diets. J. Dairy Sci., 83: 543–553. https://doi.org/10.3168/jds.S0022-0302(00)74914-9

Bowman, G.R., Beaucheman, K.A. and Shelford, J.A., 2003. Fibrolytic enzyme and parity effects on feeding behavior, salivation and ruminal pH of lactating dairy cows. J. Dairy Sci., 86: 565-575. https://doi.org/10.3168/jds.S0022-0302(03)73635-2

Bowman, G.R., Beauchemin, K.A. and Shelford, J.A., 2002. The proportion of the diet to which firbolytic enzyme are added affects nutrient digestion by lactating dairy cows. J. Dairy Sci., 85:3420–3429. https://doi.org/10.3168/jds.S0022-0302(02)74430-5

Colombatto, D., Morgavi, D.P., Furtado, A.F. and Beauchemin K.A., 2003. Screening of exogenous enzymes for ruminants diets: Relationship between biochemical characteristics and in vitro ruminal degradation. J. Anim. Sci., 81: 2628-2638. https://doi.org/10.2527/2003.81102628x

Dean, D.B., Adesogan, A.T., Krueger, N. and Littell, R.C., 2005. Effect of fibrolytic enzymes on the fermentation characteristics, aerobic stability, and digestibility of dairy cows. J. Dairy Sci., 88: 994-1003. https://doi.org/10.3168/jds.S0022-0302(05)72767-3

Dean, D.B., Adesogan, A.T., Staples, C.R., Littell, R.C. and Kim, S.C., 2007. Effect of adding the fibrolytic enzyme to different components of a dairy cow diet on feed intake, digestibility, milk production, ruminal fermentation, and blood metabolites. J. Dairy Sci., 18: 92-110.

Dhiman, T.R., Zaman, M.S., Gimenez, R.R., Walters, J.L. and Treacher, R., 2002. Performance of dairy cows fed forage treated with fibrolytic enzymes prior to feeding. Anim. Feed Sci. Technol., 101: 115–125. https://doi.org/10.1016/S0377-8401(02)00177-3

El-Kady, R.I., Mawadalla, I., Mohamed, M.I., Fadel, M. and El-Ahman, H.H., 2006. Effect of exogenous enzymes on the growth performance and digestibility of growing buffalo calves. Int. J. agric. Biol., 8: 354-359.

Gaafar, H.M.A., Abdel, E.M. and El-Reidy, K.F.A., 2010. Effect of fibrolytic enzyme supplementation and fiber content of total mixed ration on productive performance of lactating buffaloes. Slovak. J. Anim. Sci., 43: 147–153.

Kung, L., Cohen, M.A., Rode, L.M. and Treacher, R.J., 2002. The effect of fibrolytic enzymes sprayed onto forages and fed in a total mixed ration to lactating dairy cows. J. Dairy Sci., 85: 396-402. https://doi.org/10.3168/jds.S0022-0302(02)74321-X

Mcallister, T.A., Hristov, A.N., Beauchemin, K.A., Rode, L.M. and Cheng, K.J., 2001. Enzymes in ruminant diets. In: Enzymes in farm animal nutrition (eds. M. Bedford and G. Partridge), CAB International Publishing, Oxon, UK, pp. 273–298.

Meale, S.J., Beauchemin, K.A., Hristov, A.N., Chaves, A.V., McAllister, T.A., 2014. Opportunities and challenges in using exogenous enzymes to improve ruminant production. J. Anim. Sci., 92: 427–442. https://doi.org/10.2527/jas.2013-6869

Mertens, D.R., 2009. Impact of NDF and digestibility on dairy cow performance. Adv. Dairy Technol., 21: 191-201.

Miller, D.R., Granzin, B.C., Elliott, R. and Norton, B.W., 2008. Effects of an exogenous enzyme, Roxazyme® G2 Liquid, on milk production in pasture fed dairy cows. Anim. Feed Sci. Technol., 145: 194–208. https://doi.org/10.1016/j.anifeedsci.2007.05.049

Morgavi, D.P., Nsereko, V.L., Rode, L.M., Beauchemin, K.A., Mcallister, T.A. and Wang Y., 2002. Effect of trichoderma feed enzyme on growth and substrate degradation by Fibrobacter succinogens. Reprod. Nutr. Dev., 40: 219–228.

Mohamed, D.E.A., Borhami, B.E., El-Shazly, K.A. and Sallam, S.M.A., 2013. Effect of dietary supplementation with fibrolytic enzymes on the productive performance of early lactating dairy cows. J. agric. Sci., 5: 146-155. https://doi.org/10.5539/jas.v5n6p146

Muwalla, M.M., Haddad, S.G. and Hijazeen, M.A., 2007. Effect of fibrolytic enzyme inclusion in high concentrate fattening diets on nutrient digestibility and growth performance of Awassi lambs. Livest. Sci., 111: 255–258. https://doi.org/10.1016/j.livsci.2007.03.003

Nsereko, V.L., Morgavi, D.P., Rode, L.M., Beauchemin, K.A. and Mcalliater, T.A., 2000. Effects of fungal enzyme preparations and hydrolysis and subsequent degradation of alfalfa hay fiber by mixed rumen micro-organisms in vitro. Anim. Feed Sci. Technol., 88: 153–170. https://doi.org/10.1016/S0377-8401(00)00225-X

Shadmanesh, A., 2014. Effect of dietary suppplement with fibrolytic enzymes on the productive performance of early lactating dairy cows. Indian J. Fund. appl. Life Sci., 4: 396-401.

Shekhar, C., Thakur, S.S. and Shelke, S.K., 2010. Effect of exogenous fibrolytic enzymes supplementation on milk production and nutrient utilization in Murrah buffaloes. Trop. Anim. Hlth. Prod., 42: 1465-1470. https://doi.org/10.1007/s11250-010-9578-2

Sarwar, M., Khan, M.A. and Iqbal, Z., 2002. Feed resources for livestock in Pakistan. Int. J. agric. Biol., 4: 186–192.

Sutton, J.D., Fipps, R.H., Beever, D.E., Hampries, D.H., Hartnell, G.H., Vicini, J.L. and Hard, D.L., 2003. Effect of method of application of a fibrolytic enzyme product on digestive processes and milk production in Holstein-Friesian cows. J. Dairy Sci., 86: 546-556. https://doi.org/10.3168/jds.S0022-0302(03)73633-9

Titi, H.H. and Tabbaa, M.J., 2004. Efficacy of exogenous cellulase on digestibility and growth of dairy calves. Livest. Prod. Sci., 87: 207-214. https://doi.org/10.1016/j.livprodsci.2003.07.012

Van Soest, P.J., Mertens, D.R. and Deinum B., 1978. Pre harvest factors influencing quality of conserved forage. J. Anim. Sci., 47: 712-720. https://doi.org/10.2527/jas1978.473712x

Van Soest, P.J., Robertson, J.B. and Lewis, B.A., 1991. Methods for dietary fiber, neutral detergent fiber, and nonstarch polysaccharides in relation to animal nutrition. J. Dairy Sci., 74: 3583-3597. https://doi.org/10.3168/jds.S0022-0302(91)78551-2

Yang, W.Z., Beauchemin, K.A. and Rode L.M., 1999. Effects of an enzyme feed additive on extent of digestion and milk production of lactating dairy cows. J. Dairy Sci., 82: 391-403. https://doi.org/10.3168/jds.S0022-0302(99)75245-8