- Email: [email protected]
Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration
G Model
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
Small Ruminant Research xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Small Ruminant Research journal homepage: www.elsevier.com/locate/smallrumres
Short communication
Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration S.U. Mahfuz a,∗ , M.R. Chowdhury c , M.M.H. Khan c , M.A. Baset b a
Department of Animal Nutrition, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh Department of Livestock Production and Management, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh c Department of Biochemistry and Chemistry, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh b
a r t i c l e
i n f o
Article history: Received 5 December 2013 Received in revised form 2 April 2014 Accepted 3 April 2014 Available online xxx
Keywords: Triple super phosphate Rice straw In vitro In vivo Degradability Digestibility
a b s t r a c t Two way analysis of variance in duplicate were used to assess the degradability of rice straw with triple super phosphate (TSP: Ca (H2 PO4 )2 ·CaSO4 ·2H2 O) as a source of phosphorous (P) at six levels (0, 2, 4, 6, 8 and 12 g/kg) for each of the five different incubation times (0, 6, 12, 24 and 48 h). The effects of P (TSP) levels and time were significant for in vitro dry matter and organic matter degradability. For NH3 -N concentration in rumen fluid the effect of P and time were significant. For in vivo digestibility eight bucks average live weight 7.63 ± 0.19 kg were selected. The DM, ADF and OM digestibility were significantly higher in P group than control group. The digestibility of CP was higher in P group than control group. It can be concluded that P has an important role in ruminant digestion and metabolism. As low quality forages are deficit in P, supplementation of P can improve the low quality forages. Thus ruminants can proper utilize this low quality forages. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Rice straw which is commonly used as basal feed for ruminants in the developing countries has low nutritive values. The high level of lignification and silicification are the main constrain for rice straw digestibility in ruminants (Van Soest, 2006). Rice straw is an agricultural waste and its availability is abundant, but up to now its utilization is still limited (Syamsu et al., 2007). Feeding ruminants with rice straw only will cause live weight loss and possibly health problem (Nguyen Van et al., 2003). Supplements increase the utilization of low quality forages, but the requirement
∗ Corresponding author. Tel.: +880 1724671408. E-mail address: [email protected] (S.U. Mahfuz).
for these supplements is more than their availability, especially in developing countries (Devendra and Sevilla, 2002). Therefore, it is necessary to look for alternative natural or artificial feed supplements that can improve the utilization of rice straw. Phosphorus (P) is very important for normal rumen metabolism, skeletal growth, production and reproduction. Rumen microbes have specific P requirements to degrade the cell walls of feedstuffs. Also, rumen microbes need P to maintain metabolism and growth (Komisarczuk et al., 1988); and total P content of rumen microorganisms ranges from 2 to 6% of the dry matter (Valk et al., 2000). Bacterial proliferation is strongly dependent on a sufficient supply of P. Phosphorus deficiency occurs in young calves and dry cows due to lack of supplementary feeding (McDonald et al., 2002). Most of the normal forages consumed by
http://dx.doi.org/10.1016/j.smallrumres.2014.04.003 0921-4488/© 2014 Elsevier B.V. All rights reserved.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model RUMIN-4714; No. of Pages 5
ARTICLE IN PRESS S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
2
ruminants are low in P content than the requirement (Khan and Chaudhry, 2012). In herds reared at pasture, Phosphorus can be considered, worldwide, the major mineral deficiency causing economic impact (Underwood and Suttle, 1999). Therefore, increasing use of poor quality roughages and by-products, generally deficient in P, animals need to be supplemented with this mineral to meet their nutritional requirements. So farmers from developing countries are looking for alternative source of P that is cheap and easily available. TSP can be used as an alternative source which contains 48% phosphorus penta oxide (P2 O5 ) and 21.12% P. It was not only the sources of P but also provides calcium and sulphur. Therefore the present study was carried out to examine the effect of triple super phosphate supplements to enhance the utilization of low quality forages like rice straw by ruminants. 2. Materials and methods 2.1. Experimental work plan This study was evaluated the suitability of different levels of TSP as a source of P on the in vitro rumen degradability and fermentation profiles of rice straw at different incubation times. Proximate analyses of feed samples were performed following the methods of AOAC (2004) and in vivo digestibility trial was conducted with bucks. 2.2. Characterization of feed sample 2.2.1. Processing of feed sample Rice straw was dried in the sun. After drying the sample was coarse ground to approximate size (1 mm) using a grinder (blender machine) and measured by analytical balance (Setra, M-EL-410s, USA).
2.6. In vivo trial 2.6.1. Live weight of bucks and dietary treatment Eight young bucks (8–9 month) of Black Bengal goat were kept into individual pens for control and P group where each group contains four bucks and the average live weight was 7.63 ± 0.19 kg. In the control group 500 g napier grass and 250 g concentrate mixture (keshari bran 35%, soyabean meal 10%, wheat bran 10%, crushed gram 25%, crushed maize 20%) were provided per buck per day. For P group, 500 g napier grass and 250 g concentrate mixture along with 5 g TSP was provided per buck per day.
2.6.2. Measurement of feed intake Feed intake and refusals were recorded daily. Bucks consumed all the concentrate but sometimes refused napier stem. For measuring feed intake napier grass was weighed every day before supplying to the buck; next morning left over of napier grass stem was weighed by top loading balance.
2.6.3. Collection and chemical analysis of faeces sample Digestibility was determined by using total collection method. During collection period (10 days), complete collection of faeces from each group of bucks were taken in a polythene sheet daily in the morning, weighed, mixed thoroughly and 5% of it was sampled and stored at −20 ◦ C. At the end of collection period faecal samples were composited by group and 10% of the composited samples were taken for analysis following the methods of AOAC (2004).
2.7. Statistical analyses The in vitro data were analyzed by using ANOVA in General Linear model of Minitab to compare degradability and NH3 -N concentration of forages with different levels of P supplementation and the in vivo data were analyzed by using ANOVA following the principles of CRD using computer package GENSTAT (Lawes Agricultural Trust, 1997). Significant differences between means of different groups were compared by using the Tukey’s test at p > 0.05.
2.3. Preparation of buffer solution
3. Result The buffer solution was prepared as described by McDougall (1948) according to the formula for synthetic saliva with some modification (Khan and Chaudhry, 2010). 2.4. Collection of rumen fluid and preparation of buffer inoculum The rumen fluid was collected from immediately after slaughtered a mature cow. The fluid was then transferred into a flask via filtering with a filter cloth. The filtered rumen fluid was transported to the laboratory by a flask (37 ◦ C) for in vitro trial. The rumen fluid was pooled through two layers of the muslin cloth into pre warmed flask. The rumen fluid was mixed with pre warmed buffer at 1:3 (rumen fluid: buffer) ratio in a bottle which was covered with black polythene. 2.5. In vitro incubation The incubations of rice straw were conducted in 50-ml centrifuge tubes each containing about 0.4 g of ground (1 mm) sample. TSP was added at six different levels (0, 2, 4, 6, 8 and 12 g/kg). Then 40 ml of buffered rumen fluid were added to each tube. The tubes were sealed with rubber stoppers fitted with pressure release narrow glass rod. Incubation was conducted at 38 ◦ C in a water bath. After 0, 6, 12, 24 and 48 h the tubes were collected from water bath and submerged in an ice box to stop further fermentation. The liquid and residue were separated by filtering with filter cloth. The supernatant of the buffered rumen fluid was collected to determine ammonia concentration in rumen fluid and 20 ml of supernatant were acidified with 10 ml of 1 N HCl and kept in a tube. Residues were washed with distilled water and used to determine DM degradability. Acidified sample was distillated with 40 ml 40% NaOH solution into kjeldahl flask. Afterward, 20 ml 2% boric acid solution was placed into distillation set. After the distillation the sample was titrated with 0.1 N HCl.
3.1. Chemical composition of roughages and concentrate mixture The chemical composition of rice straw, napier grass and concentrate mixture used during the experimental period was shown in Table 1. DM and ADF were higher in rice straw than napier grass. On the other hand, CP and Ash were lower in rice straw than napier grass. The concentrate mixture contains higher CP and lower ADF and ash than rice straw and napier grass. 3.2. Effects of phosphorus supplementation (triple super phosphate, TSP: Ca (H2 PO4 )2 ·CaSO4 ·2H2 O) on rice straw degradability and fermentation profile 3.2.1. In vitro DM degradability of rice straw The effects of P levels and time were significant for in vitro DM degradability. The DM degradability increased with higher level of P and longer incubation time (Table 2). 3.2.2. In vitro OM degradability of rice straw The effects of P levels and time were significant for in vitro OM degradability. The OM degradability was increased with higher level of P and longer incubation time like dry matter degradability (Table 3).
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
3
Table 1 Chemical compositions (g/100 g DM) of forages and concentrate mixture (mean ± std). Feed items
DM (% of fresh weight)
Rice straw Napier grass Concentrate mixture
92.80 ± 0.23 19.32 ± 0.20 87.58 ± 2.08
% of DM OM
CP
ADF
EE
NFE
Ash
92.32 ± 0.19 89.2 ± 0.32 92.69 ± 1.58
3.54 ± 0.22 8.67 ± 0.56 15.60 ± 1.42
35.37 ± 0.61 29.18 ± 0.78 11.93 ± 1.01
1.16 ± 0.03 2.14 ± 0.22 9.27 ± 0.58
52.24 ± 0.63 49.21 ± 1.27 56.36 ± 1.23
7.68 ± 0.18 10.80 ± 0.09 6.83 ± 0.84
Table 2 In vitro DM degradability of rice straw (g/kg) at different incubation times. Level of phosphorus (g/kg)
Time
0 2.0 4.0 6.0 8.0 12.0
SEM
0h
6h
12 h
24 h
48 h
97.50 108.8 136.3 132.5 166.3 222.5
150.0 152.5 201.3 265.0 306.2 361.3
167.5 292.5 316.2 355.0 343.7 405.0
175.0 301.3 325.0 366.3 347.5 453.8
287.5 326.3 351.2 428.8 458.7 562.5
20.9 29.5 27.8 34.6 33.2 37.3
350
350
300
300
Ammonia N (mg/L)
Ammonia N (mg/L)
Level of phosphorus; p < 0.01, time; p < 0.01.
250 200 150 100
250 200 150 100 50
50
0
0 0
2
4
6
8
0
12
10
20
30
40
50
Different incubation time (h)
Level of phosphorus (g/kg)
Fig. 1. NH3 -N concentration (mg/L) in rumen fluid with rice straw at different levels of phosphorus.
3.2.3. Effects of phosphorus supplementation on NH3 -N concentration (mg/L) in rumen fluid with rice straw The effect of P and time were significant for NH3 -N concentration in rumen fluid. The NH3 -N concentration increased with increased level of P and longer incubation time (Figs. 1 and 2). 3.3. In vivo digestibility 3.3.1. Chemical composition of goat faeces The chemical compositions of goat faeces for control and P group are shown in Table 4. The result showed the DM,
Fig. 2. NH3 -N concentration (mg/L) in rumen fluid with rice straw at different incubation times.
CP, ADF and Ash were lower in P group than control group. On the other hand, OM and NFE were higher in P group than control group.
3.3.2. Effect of phosphorus supplementation (TSP) on in vivo digestibility The in vivo digestibility of DM, ADF and OM were significantly higher in P group than control group and there was a tendency to increase digestibility with the presence of P. On the other hand, the digestibility of CP was higher in P group (Table 5).
Table 3 In vitro OM degradability of rice straw (g/kg) at different incubation times. Level of phosphorus (g/kg)
0 2.0 4.0 6.0 8.0 12.0
Time
SEM
0h
6h
12 h
24 h
48 h
120.6 130.1 150.4 166.7 187.0 196.5
219.5 227.6 243.9 256.1 307.6 378.0
256.1 307.6 322.5 334.7 353.7 394.3
319.8 325.2 337.4 369.9 380.8 458.0
357.7 375.3 417.3 448.5 486.4 592.1
27.5 28.7 30.4 32.3 32.7 42.8
Level of phosphorus; p < 0.01, time; p < 0.01.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
4 Table 4 Chemical composition of goat faeces. Group
DM (% of fresh weight)
Control Phosphorus
69.24 67.03
% of DM CP% 8.18 7.32
ADF%
Ash%
EE%
OM%
NFE%
17.62 14.26
10.384 8.70
1.91 1.92
89.616 91.304
61.914 67.80
Table 5 Effect of phosphorus supplementation (TSP) on in vivo digestibility. Parameter
DM digestibility (g/kg) CP digestibility (g/kg) ADF digestibility (g/kg) OM digestibility (g/kg)
Group
SED and level of significant
Control
Phosphorus
802.1 879.3 795.3 806.48
808.7 895.8 841.3 810.68
3.10** 12.53NS 13.10** 1.886**
NS: not significant. ** Significant at the 0.01 level.
4. Discussion 4.1. Chemical composition of roughages It was expected that napier grass should have higher CP content than rice straw and rice straw should have higher ADF content than napier grass. In the present experiment the ash value of rice straw was unexpectedly lower than napier grass and published value (Khan and Chaudhry, 2010). It was may be due to different in variety, soil quality and any other environmental effect. 4.2. Effects of phosphorus supplementation on DM and OM degradability of rice straw The DM and OM degradability increased due to the presence of P that helps in increased microbial activity during the process of digestion. The higher degradability of DM causes higher rumen metabolism. The DM and OM degradability of rice straw without any treatment was very low due to various ligno-cellulose bonds. TSP as a source of P played an important role in synthesis of protein and was crucial for the production of ATP. Due to increased microbial activity and higher protein synthesis OM and DM degradability were increased with higher level of TSP. Phosphorus is necessary for normal growth and function of rumen micro-organisms especially for cellulose digestion that leads to increased DM and OM degradability (Harris et al., 2003). Insoluble carbohydrate became soluble and available by various enzymatic changes with the presence of P supplementation. Supplementation of P and sulphur were effective in stimulating dry matter, organic matter, protein and fibrous fraction digestibility in cassava leaves (Zain et al., 2010). 4.3. Effects of phosphorus supplementation on NH3 -N concentration (mg/L) in rumen fluid with rice straw Due to higher degradability of DM and OM enhanced higher metabolism that helped to increase ammonia nitrogen concentration. The activity of bacterial fibrolytic enzymes is strongly dependent on the supply of available
P. Kampa and Wanapat (2006) stated that NH3 -N, CO2 and CH4 in the rumen are used to synthesize microbial cells. The ammonia concentration in rumen fluid is the key intermediate in the microbial degradation and synthesis of protein. The optimum concentration of ammonia in rumen fluid varies widely, from 85 to over 300 mg/L. In the present study, rumen ammonia of cow was average 300 mg/L which was within the acceptable range to maintain rumen function. Increased NH3 -N concentration indicated increased DM and OM degradability (Sommart et al., 2000). 4.4. Effect of phosphorus supplementation on in vivo digestibility The nutrient digestibility was higher in P group where 5 g TSP was added with control feeding. Phosphorus supplementation acts as a mediator of higher digestion and metabolism of nutrient. The present study indicated that improvement in fibre degradation by P supplementation occurred through its specific stimulation on growth of rumen cellulolytic bacteria and anaerobic rumen fungi (Khan and Chaudhry, 2012). Since P takes part in the formation of DNA and RNA, the number of rumen microbes may increased at the presence of TSP and ultimately the digestibility increased. 5. Conclusion A diet of rice straw alone is not adequate even to maintain animals, which will lose weight if no supplementary protein or mineral is given. This is true of all straws and roughages containing less than 4 percent protein, since a level of approximately 6 percent in the diet are needed to prevent weight loss. This study has shown that P supplementation improved DM, CP, ADF and OM degradability and digestibility of low quality forage like rice straw. TSP may be a source of supplementation to improve the low quality forages. It can be used at higher doses (20 g/kg) without any side effect. Finally it may be concluded that TSP supplementation can improve the digestibility of low quality forage.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model RUMIN-4714; No. of Pages 5
ARTICLE IN PRESS S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
Conflict of interest The author feels immerse proud to express his heartfelt respect and deepest sense of gratitude to his research Cosupervisor Dr. Mohammad Mehedi Hasan Khan, Chairman, Department of Biochemistry and Chemistry, Faculty of Veterinary and Animal Science, Sylhet Agricultural University (SAU), Sylhet for his scholastic guidance, valuable suggestions and untiring assistance in all phases of the research work and in the preparation of this thesis. References AOAC, 2004. Official Methods for Analysis, 13th ed. Association of Official Analytical Chemists, Washington, DC. Devendra, C., Sevilla, C.C., 2002. Availability and use of feed resources in crop-animal systems in Asia. Agric. Syst. 71, 59–73. Harris, B., Adams, A.L., Van Horn, H.H., 2003. Mineral Needs of Dairy Cattle. Animal Science Department, Institute of Food and Agricultural Sciences, University of Florida, Florida, USA. Kampa, S., Wanapat, M., 2006. Influenceces of energy sources and level on ruminal fermentation and microbial protein synthesis in dairy stress. J Nutr. Vet. Sci. 5, 294–300. Khan, M.M.H., Chaudhry, A.S., 2010. Chemical composition of selected forages and spices and the effect of these spices on in vitro rumen degradability of some forages. Asian-Australas. J. Anim. Sci. 23, 889–900. Khan, M.M.H., Chaudhry, A.S., 2012. Spice supplementation of forage based ruminant diets (in vitro study). LAP Lambert Academic Publishing Gmblt & Co, KG Saarbrucken, Germany.
5
Komisarczuk, S., Gaudet, G., Hannequart, G., Fonty, G., Durand, M., 1988. Effects of a sub-deficiency in phosphorus on some aspects of cellulolytic activity of Bacteroides succinogenes. Reprod. Nutr. 28, 79–80. Lawes Agricultural Trust, 1997. A General Statistical Programme. Genstat 5, 5th ed. (Beta). Rothamsted Exptl. Stat., Harpenden, Hertfordshire, UK. McDonald, P., Edwards, R.A., Greenhalgh, J.F., Morgan, C.A., 2002. Animal Nutrition. Longman Group Limited, Harlow. McDougall E.I., Studies on ruminant saliva. The composition and output of sheep’s saliva. Institute of animal pathology, University of Cambridge, 1948, 99-100. Nguyen Van, T., Udén, P., 2003. Feces as an alternative to rum fluid for in vitro digestibility measurement. Buffalo J. Thail. 19, 4. Sommart, K., Parker, D.S., Linson, P., 2000. Fermentation characteristics and microbial protein synthesis in an in vitro system using cassava rice straw and dried ruzi grass as substrate. Asian Australas J. Anim. Sci. 13, 1084–1093. Syamsu, J.A., Sofyan, L.A., Mudikdjo, K.G., Said, E.G., 2007. Power support agricultural waste as ruminant feed resources. Wartazona 24 (3), 43–45. Underwood, E.J., Suttle, N.F., 1999. Phosphorus. In: The Mineral Nutrition of Livestock, 3rd ed. CABI, London, pp. 105–148. Valk, H., Metcalf, J.A., Withers, P.J.A., 2000. Prospects for minimizing phosphorus excretion in ruminants by dietary manipulation. J. Environ. 29, 28–36. Van Soest, P.J., 2006. Rice straw, the role of silica and treatments to improve quality. Anim. Feed Sci. Technol. 130, 137–171. Zain, M., Ninggrat, R., Jamarun, N.A.T., 2010. Effect of phosphorus supplementation of ammoniated rice straw on rumen fermentability, synthesised microbial protein and degradability in vitro. In: Advances in Animal Bioscience: Proceedings of the British Society of Animal Science Annual Conference, Belfast, Northern Ireland, UK, p. 210.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
Small Ruminant Research xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Small Ruminant Research journal homepage: www.elsevier.com/locate/smallrumres
Short communication
Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration S.U. Mahfuz a,∗ , M.R. Chowdhury c , M.M.H. Khan c , M.A. Baset b a
Department of Animal Nutrition, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh Department of Livestock Production and Management, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh c Department of Biochemistry and Chemistry, Faculty of Veterinary and Animal Science, Sylhet Agricultural University, Sylhet 3100, Bangladesh b
a r t i c l e
i n f o
Article history: Received 5 December 2013 Received in revised form 2 April 2014 Accepted 3 April 2014 Available online xxx
Keywords: Triple super phosphate Rice straw In vitro In vivo Degradability Digestibility
a b s t r a c t Two way analysis of variance in duplicate were used to assess the degradability of rice straw with triple super phosphate (TSP: Ca (H2 PO4 )2 ·CaSO4 ·2H2 O) as a source of phosphorous (P) at six levels (0, 2, 4, 6, 8 and 12 g/kg) for each of the five different incubation times (0, 6, 12, 24 and 48 h). The effects of P (TSP) levels and time were significant for in vitro dry matter and organic matter degradability. For NH3 -N concentration in rumen fluid the effect of P and time were significant. For in vivo digestibility eight bucks average live weight 7.63 ± 0.19 kg were selected. The DM, ADF and OM digestibility were significantly higher in P group than control group. The digestibility of CP was higher in P group than control group. It can be concluded that P has an important role in ruminant digestion and metabolism. As low quality forages are deficit in P, supplementation of P can improve the low quality forages. Thus ruminants can proper utilize this low quality forages. © 2014 Elsevier B.V. All rights reserved.
1. Introduction Rice straw which is commonly used as basal feed for ruminants in the developing countries has low nutritive values. The high level of lignification and silicification are the main constrain for rice straw digestibility in ruminants (Van Soest, 2006). Rice straw is an agricultural waste and its availability is abundant, but up to now its utilization is still limited (Syamsu et al., 2007). Feeding ruminants with rice straw only will cause live weight loss and possibly health problem (Nguyen Van et al., 2003). Supplements increase the utilization of low quality forages, but the requirement
∗ Corresponding author. Tel.: +880 1724671408. E-mail address: [email protected] (S.U. Mahfuz).
for these supplements is more than their availability, especially in developing countries (Devendra and Sevilla, 2002). Therefore, it is necessary to look for alternative natural or artificial feed supplements that can improve the utilization of rice straw. Phosphorus (P) is very important for normal rumen metabolism, skeletal growth, production and reproduction. Rumen microbes have specific P requirements to degrade the cell walls of feedstuffs. Also, rumen microbes need P to maintain metabolism and growth (Komisarczuk et al., 1988); and total P content of rumen microorganisms ranges from 2 to 6% of the dry matter (Valk et al., 2000). Bacterial proliferation is strongly dependent on a sufficient supply of P. Phosphorus deficiency occurs in young calves and dry cows due to lack of supplementary feeding (McDonald et al., 2002). Most of the normal forages consumed by
http://dx.doi.org/10.1016/j.smallrumres.2014.04.003 0921-4488/© 2014 Elsevier B.V. All rights reserved.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model RUMIN-4714; No. of Pages 5
ARTICLE IN PRESS S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
2
ruminants are low in P content than the requirement (Khan and Chaudhry, 2012). In herds reared at pasture, Phosphorus can be considered, worldwide, the major mineral deficiency causing economic impact (Underwood and Suttle, 1999). Therefore, increasing use of poor quality roughages and by-products, generally deficient in P, animals need to be supplemented with this mineral to meet their nutritional requirements. So farmers from developing countries are looking for alternative source of P that is cheap and easily available. TSP can be used as an alternative source which contains 48% phosphorus penta oxide (P2 O5 ) and 21.12% P. It was not only the sources of P but also provides calcium and sulphur. Therefore the present study was carried out to examine the effect of triple super phosphate supplements to enhance the utilization of low quality forages like rice straw by ruminants. 2. Materials and methods 2.1. Experimental work plan This study was evaluated the suitability of different levels of TSP as a source of P on the in vitro rumen degradability and fermentation profiles of rice straw at different incubation times. Proximate analyses of feed samples were performed following the methods of AOAC (2004) and in vivo digestibility trial was conducted with bucks. 2.2. Characterization of feed sample 2.2.1. Processing of feed sample Rice straw was dried in the sun. After drying the sample was coarse ground to approximate size (1 mm) using a grinder (blender machine) and measured by analytical balance (Setra, M-EL-410s, USA).
2.6. In vivo trial 2.6.1. Live weight of bucks and dietary treatment Eight young bucks (8–9 month) of Black Bengal goat were kept into individual pens for control and P group where each group contains four bucks and the average live weight was 7.63 ± 0.19 kg. In the control group 500 g napier grass and 250 g concentrate mixture (keshari bran 35%, soyabean meal 10%, wheat bran 10%, crushed gram 25%, crushed maize 20%) were provided per buck per day. For P group, 500 g napier grass and 250 g concentrate mixture along with 5 g TSP was provided per buck per day.
2.6.2. Measurement of feed intake Feed intake and refusals were recorded daily. Bucks consumed all the concentrate but sometimes refused napier stem. For measuring feed intake napier grass was weighed every day before supplying to the buck; next morning left over of napier grass stem was weighed by top loading balance.
2.6.3. Collection and chemical analysis of faeces sample Digestibility was determined by using total collection method. During collection period (10 days), complete collection of faeces from each group of bucks were taken in a polythene sheet daily in the morning, weighed, mixed thoroughly and 5% of it was sampled and stored at −20 ◦ C. At the end of collection period faecal samples were composited by group and 10% of the composited samples were taken for analysis following the methods of AOAC (2004).
2.7. Statistical analyses The in vitro data were analyzed by using ANOVA in General Linear model of Minitab to compare degradability and NH3 -N concentration of forages with different levels of P supplementation and the in vivo data were analyzed by using ANOVA following the principles of CRD using computer package GENSTAT (Lawes Agricultural Trust, 1997). Significant differences between means of different groups were compared by using the Tukey’s test at p > 0.05.
2.3. Preparation of buffer solution
3. Result The buffer solution was prepared as described by McDougall (1948) according to the formula for synthetic saliva with some modification (Khan and Chaudhry, 2010). 2.4. Collection of rumen fluid and preparation of buffer inoculum The rumen fluid was collected from immediately after slaughtered a mature cow. The fluid was then transferred into a flask via filtering with a filter cloth. The filtered rumen fluid was transported to the laboratory by a flask (37 ◦ C) for in vitro trial. The rumen fluid was pooled through two layers of the muslin cloth into pre warmed flask. The rumen fluid was mixed with pre warmed buffer at 1:3 (rumen fluid: buffer) ratio in a bottle which was covered with black polythene. 2.5. In vitro incubation The incubations of rice straw were conducted in 50-ml centrifuge tubes each containing about 0.4 g of ground (1 mm) sample. TSP was added at six different levels (0, 2, 4, 6, 8 and 12 g/kg). Then 40 ml of buffered rumen fluid were added to each tube. The tubes were sealed with rubber stoppers fitted with pressure release narrow glass rod. Incubation was conducted at 38 ◦ C in a water bath. After 0, 6, 12, 24 and 48 h the tubes were collected from water bath and submerged in an ice box to stop further fermentation. The liquid and residue were separated by filtering with filter cloth. The supernatant of the buffered rumen fluid was collected to determine ammonia concentration in rumen fluid and 20 ml of supernatant were acidified with 10 ml of 1 N HCl and kept in a tube. Residues were washed with distilled water and used to determine DM degradability. Acidified sample was distillated with 40 ml 40% NaOH solution into kjeldahl flask. Afterward, 20 ml 2% boric acid solution was placed into distillation set. After the distillation the sample was titrated with 0.1 N HCl.
3.1. Chemical composition of roughages and concentrate mixture The chemical composition of rice straw, napier grass and concentrate mixture used during the experimental period was shown in Table 1. DM and ADF were higher in rice straw than napier grass. On the other hand, CP and Ash were lower in rice straw than napier grass. The concentrate mixture contains higher CP and lower ADF and ash than rice straw and napier grass. 3.2. Effects of phosphorus supplementation (triple super phosphate, TSP: Ca (H2 PO4 )2 ·CaSO4 ·2H2 O) on rice straw degradability and fermentation profile 3.2.1. In vitro DM degradability of rice straw The effects of P levels and time were significant for in vitro DM degradability. The DM degradability increased with higher level of P and longer incubation time (Table 2). 3.2.2. In vitro OM degradability of rice straw The effects of P levels and time were significant for in vitro OM degradability. The OM degradability was increased with higher level of P and longer incubation time like dry matter degradability (Table 3).
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
3
Table 1 Chemical compositions (g/100 g DM) of forages and concentrate mixture (mean ± std). Feed items
DM (% of fresh weight)
Rice straw Napier grass Concentrate mixture
92.80 ± 0.23 19.32 ± 0.20 87.58 ± 2.08
% of DM OM
CP
ADF
EE
NFE
Ash
92.32 ± 0.19 89.2 ± 0.32 92.69 ± 1.58
3.54 ± 0.22 8.67 ± 0.56 15.60 ± 1.42
35.37 ± 0.61 29.18 ± 0.78 11.93 ± 1.01
1.16 ± 0.03 2.14 ± 0.22 9.27 ± 0.58
52.24 ± 0.63 49.21 ± 1.27 56.36 ± 1.23
7.68 ± 0.18 10.80 ± 0.09 6.83 ± 0.84
Table 2 In vitro DM degradability of rice straw (g/kg) at different incubation times. Level of phosphorus (g/kg)
Time
0 2.0 4.0 6.0 8.0 12.0
SEM
0h
6h
12 h
24 h
48 h
97.50 108.8 136.3 132.5 166.3 222.5
150.0 152.5 201.3 265.0 306.2 361.3
167.5 292.5 316.2 355.0 343.7 405.0
175.0 301.3 325.0 366.3 347.5 453.8
287.5 326.3 351.2 428.8 458.7 562.5
20.9 29.5 27.8 34.6 33.2 37.3
350
350
300
300
Ammonia N (mg/L)
Ammonia N (mg/L)
Level of phosphorus; p < 0.01, time; p < 0.01.
250 200 150 100
250 200 150 100 50
50
0
0 0
2
4
6
8
0
12
10
20
30
40
50
Different incubation time (h)
Level of phosphorus (g/kg)
Fig. 1. NH3 -N concentration (mg/L) in rumen fluid with rice straw at different levels of phosphorus.
3.2.3. Effects of phosphorus supplementation on NH3 -N concentration (mg/L) in rumen fluid with rice straw The effect of P and time were significant for NH3 -N concentration in rumen fluid. The NH3 -N concentration increased with increased level of P and longer incubation time (Figs. 1 and 2). 3.3. In vivo digestibility 3.3.1. Chemical composition of goat faeces The chemical compositions of goat faeces for control and P group are shown in Table 4. The result showed the DM,
Fig. 2. NH3 -N concentration (mg/L) in rumen fluid with rice straw at different incubation times.
CP, ADF and Ash were lower in P group than control group. On the other hand, OM and NFE were higher in P group than control group.
3.3.2. Effect of phosphorus supplementation (TSP) on in vivo digestibility The in vivo digestibility of DM, ADF and OM were significantly higher in P group than control group and there was a tendency to increase digestibility with the presence of P. On the other hand, the digestibility of CP was higher in P group (Table 5).
Table 3 In vitro OM degradability of rice straw (g/kg) at different incubation times. Level of phosphorus (g/kg)
0 2.0 4.0 6.0 8.0 12.0
Time
SEM
0h
6h
12 h
24 h
48 h
120.6 130.1 150.4 166.7 187.0 196.5
219.5 227.6 243.9 256.1 307.6 378.0
256.1 307.6 322.5 334.7 353.7 394.3
319.8 325.2 337.4 369.9 380.8 458.0
357.7 375.3 417.3 448.5 486.4 592.1
27.5 28.7 30.4 32.3 32.7 42.8
Level of phosphorus; p < 0.01, time; p < 0.01.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model
ARTICLE IN PRESS
RUMIN-4714; No. of Pages 5
S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
4 Table 4 Chemical composition of goat faeces. Group
DM (% of fresh weight)
Control Phosphorus
69.24 67.03
% of DM CP% 8.18 7.32
ADF%
Ash%
EE%
OM%
NFE%
17.62 14.26
10.384 8.70
1.91 1.92
89.616 91.304
61.914 67.80
Table 5 Effect of phosphorus supplementation (TSP) on in vivo digestibility. Parameter
DM digestibility (g/kg) CP digestibility (g/kg) ADF digestibility (g/kg) OM digestibility (g/kg)
Group
SED and level of significant
Control
Phosphorus
802.1 879.3 795.3 806.48
808.7 895.8 841.3 810.68
3.10** 12.53NS 13.10** 1.886**
NS: not significant. ** Significant at the 0.01 level.
4. Discussion 4.1. Chemical composition of roughages It was expected that napier grass should have higher CP content than rice straw and rice straw should have higher ADF content than napier grass. In the present experiment the ash value of rice straw was unexpectedly lower than napier grass and published value (Khan and Chaudhry, 2010). It was may be due to different in variety, soil quality and any other environmental effect. 4.2. Effects of phosphorus supplementation on DM and OM degradability of rice straw The DM and OM degradability increased due to the presence of P that helps in increased microbial activity during the process of digestion. The higher degradability of DM causes higher rumen metabolism. The DM and OM degradability of rice straw without any treatment was very low due to various ligno-cellulose bonds. TSP as a source of P played an important role in synthesis of protein and was crucial for the production of ATP. Due to increased microbial activity and higher protein synthesis OM and DM degradability were increased with higher level of TSP. Phosphorus is necessary for normal growth and function of rumen micro-organisms especially for cellulose digestion that leads to increased DM and OM degradability (Harris et al., 2003). Insoluble carbohydrate became soluble and available by various enzymatic changes with the presence of P supplementation. Supplementation of P and sulphur were effective in stimulating dry matter, organic matter, protein and fibrous fraction digestibility in cassava leaves (Zain et al., 2010). 4.3. Effects of phosphorus supplementation on NH3 -N concentration (mg/L) in rumen fluid with rice straw Due to higher degradability of DM and OM enhanced higher metabolism that helped to increase ammonia nitrogen concentration. The activity of bacterial fibrolytic enzymes is strongly dependent on the supply of available
P. Kampa and Wanapat (2006) stated that NH3 -N, CO2 and CH4 in the rumen are used to synthesize microbial cells. The ammonia concentration in rumen fluid is the key intermediate in the microbial degradation and synthesis of protein. The optimum concentration of ammonia in rumen fluid varies widely, from 85 to over 300 mg/L. In the present study, rumen ammonia of cow was average 300 mg/L which was within the acceptable range to maintain rumen function. Increased NH3 -N concentration indicated increased DM and OM degradability (Sommart et al., 2000). 4.4. Effect of phosphorus supplementation on in vivo digestibility The nutrient digestibility was higher in P group where 5 g TSP was added with control feeding. Phosphorus supplementation acts as a mediator of higher digestion and metabolism of nutrient. The present study indicated that improvement in fibre degradation by P supplementation occurred through its specific stimulation on growth of rumen cellulolytic bacteria and anaerobic rumen fungi (Khan and Chaudhry, 2012). Since P takes part in the formation of DNA and RNA, the number of rumen microbes may increased at the presence of TSP and ultimately the digestibility increased. 5. Conclusion A diet of rice straw alone is not adequate even to maintain animals, which will lose weight if no supplementary protein or mineral is given. This is true of all straws and roughages containing less than 4 percent protein, since a level of approximately 6 percent in the diet are needed to prevent weight loss. This study has shown that P supplementation improved DM, CP, ADF and OM degradability and digestibility of low quality forage like rice straw. TSP may be a source of supplementation to improve the low quality forages. It can be used at higher doses (20 g/kg) without any side effect. Finally it may be concluded that TSP supplementation can improve the digestibility of low quality forage.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003
G Model RUMIN-4714; No. of Pages 5
ARTICLE IN PRESS S.U. Mahfuz et al. / Small Ruminant Research xxx (2014) xxx–xxx
Conflict of interest The author feels immerse proud to express his heartfelt respect and deepest sense of gratitude to his research Cosupervisor Dr. Mohammad Mehedi Hasan Khan, Chairman, Department of Biochemistry and Chemistry, Faculty of Veterinary and Animal Science, Sylhet Agricultural University (SAU), Sylhet for his scholastic guidance, valuable suggestions and untiring assistance in all phases of the research work and in the preparation of this thesis. References AOAC, 2004. Official Methods for Analysis, 13th ed. Association of Official Analytical Chemists, Washington, DC. Devendra, C., Sevilla, C.C., 2002. Availability and use of feed resources in crop-animal systems in Asia. Agric. Syst. 71, 59–73. Harris, B., Adams, A.L., Van Horn, H.H., 2003. Mineral Needs of Dairy Cattle. Animal Science Department, Institute of Food and Agricultural Sciences, University of Florida, Florida, USA. Kampa, S., Wanapat, M., 2006. Influenceces of energy sources and level on ruminal fermentation and microbial protein synthesis in dairy stress. J Nutr. Vet. Sci. 5, 294–300. Khan, M.M.H., Chaudhry, A.S., 2010. Chemical composition of selected forages and spices and the effect of these spices on in vitro rumen degradability of some forages. Asian-Australas. J. Anim. Sci. 23, 889–900. Khan, M.M.H., Chaudhry, A.S., 2012. Spice supplementation of forage based ruminant diets (in vitro study). LAP Lambert Academic Publishing Gmblt & Co, KG Saarbrucken, Germany.
5
Komisarczuk, S., Gaudet, G., Hannequart, G., Fonty, G., Durand, M., 1988. Effects of a sub-deficiency in phosphorus on some aspects of cellulolytic activity of Bacteroides succinogenes. Reprod. Nutr. 28, 79–80. Lawes Agricultural Trust, 1997. A General Statistical Programme. Genstat 5, 5th ed. (Beta). Rothamsted Exptl. Stat., Harpenden, Hertfordshire, UK. McDonald, P., Edwards, R.A., Greenhalgh, J.F., Morgan, C.A., 2002. Animal Nutrition. Longman Group Limited, Harlow. McDougall E.I., Studies on ruminant saliva. The composition and output of sheep’s saliva. Institute of animal pathology, University of Cambridge, 1948, 99-100. Nguyen Van, T., Udén, P., 2003. Feces as an alternative to rum fluid for in vitro digestibility measurement. Buffalo J. Thail. 19, 4. Sommart, K., Parker, D.S., Linson, P., 2000. Fermentation characteristics and microbial protein synthesis in an in vitro system using cassava rice straw and dried ruzi grass as substrate. Asian Australas J. Anim. Sci. 13, 1084–1093. Syamsu, J.A., Sofyan, L.A., Mudikdjo, K.G., Said, E.G., 2007. Power support agricultural waste as ruminant feed resources. Wartazona 24 (3), 43–45. Underwood, E.J., Suttle, N.F., 1999. Phosphorus. In: The Mineral Nutrition of Livestock, 3rd ed. CABI, London, pp. 105–148. Valk, H., Metcalf, J.A., Withers, P.J.A., 2000. Prospects for minimizing phosphorus excretion in ruminants by dietary manipulation. J. Environ. 29, 28–36. Van Soest, P.J., 2006. Rice straw, the role of silica and treatments to improve quality. Anim. Feed Sci. Technol. 130, 137–171. Zain, M., Ninggrat, R., Jamarun, N.A.T., 2010. Effect of phosphorus supplementation of ammoniated rice straw on rumen fermentability, synthesised microbial protein and degradability in vitro. In: Advances in Animal Bioscience: Proceedings of the British Society of Animal Science Annual Conference, Belfast, Northern Ireland, UK, p. 210.
Please cite this article in press as: Mahfuz, S.U., et al., Effect of triple super phosphate supplementation on degradability of rice straw and ammonia nitrogen concentration. Small Ruminant Res. (2014), http://dx.doi.org/10.1016/j.smallrumres.2014.04.003