Small Ruminant Research 41 (2001) 229±233
Effect of supplementation with a by-product of molasses fermentation or a non-protein nitrogen/mineral mix on feed intake and microbial protein supply in sheep consuming chopped oat (Avena sativa) hay Z.J. Liu1, N.P. McMeniman* School of Veterinary Science, The University of Queensland, St. Lucia, Qld 4072, Australia Accepted 5 April 2001
Abstract Crossbred ewes, weighing 30±40 kg, were assigned to three groups of six animals. One group of sheep was fed chopped oat hay (control), the second group was fed the control diet plus 30 g per head per day spray dried residue from the fermentation of molasses and the third group was fed the control diet plus 30 g per head per day of a non-protein nitrogen/mineral mix. Voluntary feed intake, digestibility of DM, OM and nitrogen, nitrogen balance and microbial nitrogen ¯ow to the intestines were signi®cantly increased by supplementation but ef®ciency of microbial protein production was not affected. # 2001 Elsevier Science B.V. All rights reserved. Keywords: Sheep; Supplement; Intake; Microbial protein production
1. Introduction The availability of energy and nitrogen are the major determinants of the amount of microbial protein synthesised in the rumen. However, moderate mineral de®ciencies or improper proportions of minerals in the diet often depress food intake and this depression may be at least partly due to impaired activity of rumen microorganisms (Durand and Komisarczuk, 1987).
* Corresponding author. Tel.: 61-7-33651108; fax: 61-7-33651288. E-mail address:
[email protected] (N.P. McMeniman). 1 Present address: Chinese Medical Material College, Jilin Agricultural University, Changchun 130118, PR China.
Also, it has been shown that some rumen bacteria have a requirement for some vitamins (Hungate, 1966) and it may be possible for a vitamin de®ciency to develop in rumen microbes and suppress their activity. Identi®cation of nutrients that stimulate microbial activity in sheep fed low quality roughages could lead to development of more effective methods of supplementing sheep. In this experiment, two supplements, Rumevite-EC,2 which is the spray dried residue left after molasses has been fermented to alcohol and a non-protein nitrogen/mineral mix, Pro-phos,2 were fed to sheep with a low quality chopped oat hay diet to investigate the effect of the supplements on feed intake and microbial nitrogen supply. 2 Ridley Agriproducts, Pty, Ltd., 1325 Progress Road, 4077 Wacol.
0921-4488/01/$ ± see front matter # 2001 Elsevier Science B.V. All rights reserved. PII: S 0 9 2 1 - 4 4 8 8 ( 0 1 ) 0 0 2 0 9 - 7
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2. Materials and methods 2.1. Animals Eighteen crossbred (12-month-old) ewes, weighing 30±40 kg, were used in this experiment. The ewes were weighed before feeding at the beginning and end of the experiment and housed in individual metabolism cages that allowed for the separate collection of faeces and urine by gravity. 2.2. Experimental design and feeding
chopped oat hay (about 100 g in excess of each ewe's previous day's voluntary feed intake) was given to the ewes with supplements placed on top of the chopped hay. There was a 2-week preliminary feeding period followed by a 5-day experimental period. Feed residues were weighed each morning, bulked for 5 days, mixed well and a sample was taken for analysis. The animals had free access to fresh water. 2.3. Sample collection
The ewes were randomly allocated into three groups of six with each group being given one of the following diets: chopped oat hay only (OC); OC supplemented with 30 g of spray dried residue left after molasses had been fermented to alcohol (EC); and OC supplemented with 30 g of a non-protein nitrogen/mineral mix (PP). The chemical composition of the basal diet and the supplements is given in Table 1. The ewes were individually fed once daily at 9 a.m. Weighed
During each day of the 5-day experimental period, all urine and faeces produced by each ewe during the previous 24 h were collected. Urine was collected into containers to which had been added 30 ml of 10% sulphuric acid to keep the pH below 3. Urine was weighed and a 1% sample taken and bulked for 5 days for each sheep. Bulked urine samples were diluted 1:20 with 0.1 M (NH4)2HPO4 and stored in a freezer ( 208C) until analysis. Faeces were also weighed each morning and a 10% sample taken and dried in an oven at 1058C for 24 h. Dried faeces samples were bulked for 5 days for individual sheep, mixed well and a sub-sample was taken for further analysis.
Table 1 Chemical composition of basal diet (OC), and the two supplements (EC and PP)
2.4. Chemical analyses
Basal diet
a
Dry matter (g/kg) Organic matter (g/kg DM)a Nitrogen (g/kg DM)a True protein nitrogen (%) Calcium (g/kg DM) Phosphorus (g/kg DM) Sulphur (g/kg DM) Magnesium (g/kg DM) Sodium (g/kg DM) Potassium (g/kg DM) Manganese (mg/kg DM) Iron (mg/kg DM) Copper (mg/kg DM) Cobalt (mg/kg DM) Zinc (mg/kg DM) Iodine (mg/kg DM) Selenium (mg/kg DM)
Supplements b
OC
EC
PP
933 932 10.1
950 570 20.3 100 80.0 5.0 22.0 13.0 5.0 103.0 150 1000 90 1.3 45 ± ±
967 276 45.5 13.3 118.0 51.0 16.0 1.0 117.0 7.0 9.6 75 306.3 30.1 508.3 30 2.54
1.0a 2.0a 1.0a 1.0a 2.0a 9.0a 89.0a 81.0a 1.5a 0.0a 17.0a ± 0.0a
a Analysed value, other data were provided by the manufacturer; DM: dry matter. b The PP supplement contained 10% EC (Stansby, personal communication).
Dry matter (DM) in feed, supplements, faeces and refusal samples was determined by drying at 1008C for 24 h. The ash from samples, dried in an electric muf¯e furnace at 6008C for 4 h, were used for organic matter (OM) determination. The total N content of samples was determined with a Leco automatic N analyser (Leco Corporation). Purine derivatives (PD), including allantoin, uric acid, hypoxanthine and xanthine were estimated by HPLC analysis (Balcells et al., 1992). 2.5. Calculations Microbial nitrogen supply was calculated from urine PD excretion (Chen and Gomes, 1992). Organic matter apparently digested in the rumen for each ewe for each diet was calculated given that the proportion of digestible organic matter apparently digested in the rumen was 0.602, which had been determined in a separate experiment with sheep cannulated at the abomasum and fed a chopped oat hay diet (Liu, 2000).
Z.J. Liu, N.P. McMeniman / Small Ruminant Research 41 (2001) 229±233
2.6. Statistical analyses All data, both directly measured and derived, were analysed using a linear model that included ®xed effect due to supplement and residual error. Differences between means were tested by LSD only when the P-value was signi®cant. Relationships between microbial nitrogen supply and OM and digestible OM intake were determined with linear regression analysis. 3. Results 3.1. Intake and digestion of DM and OM Voluntary feed intakes and digestibility values for the experimental diets are presented in Table 2. Supplementation with EC and PP increased the DM and OM intake of the whole diet and of the chopped oat hay (P < 0:01), but DM and OM intakes for the two supplemented diets did not differ (P 0:10 and 0.17, respectively). The digestibility of DM and OM in the entire tract was 45.4 and 46.4% for the control diet with these values being increased with EC and PP supplementation (P < 0:01). The digestible organic
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matter intakes of EC and PP supplemented diets were also higher than for the control diet (P < 0:01) due to their higher voluntary intake and DM digestibility (Table 2). 3.2. Nitrogen intake, digestion and balance Total nitrogen intake and nitrogen intake from the OC were different between diets (P < 0:01), the lowest being for the control diet and the highest for the PP supplemented diet (Table 3). Faecal nitrogen excretion was not affected by treatments (P > 0:05), while urine nitrogen excretion was higher for the PP supplemented diet than for control and EC diets (P < 0:01). The apparent digestibility of nitrogen in the entire tract was improved by supplementation (P < 0:01) as was the nitrogen retained by the experimental animals (P < 0:05, Table 3). 3.3. PD excretion and microbial nitrogen supply As shown in Table 4, PD excretion increased with supplementation (P < 0:01) as did microbial nitrogen supply, being 5.43, 10.06 and 9.51 g per day for OC, EC and PP diets, respectively. Microbial nitrogen supply (MNS, g per day) was related to OM intake
Table 2 Intake and digestion of a chopped oat hay diet (OC), an OC diet supplemented with the dried residue left after molasses fermentation (EC) and an OC diet supplemented with a non-protein nitrogen/mineral mix (PP)a Diets
S.E.M.
P-value
OC
EC
PP
Total DM intake (g per day) g/kg LWc From OC From supplement
582.2 b 17.9 b 582.2 b 0.0
767.8 a 24.6 a 739.3 a 28.5
863.6 a 26.4 a 834.6 a 29.0
38.66 0.10 38.66
OMd intake (g per day) From OC From supplement
542.8 b 542.8 b 0.0
706.4 a 690.2 a 16.2
786.5 a 778.5 a 8.0
36.05 36.10
Digestion (%) DM OM DOMIe (g per day)
45.4 b 46.4 b 255.0 b
56.3 a 58.4 a 415.2 a
57.2 a 58.9 a 466.8 a
2.43 2.45 32.86
b
a
Means with different letters in the same row are signi®cantly different ( P < 0:01). DM: dry matter. c LW: live weight. d OM: organic matter. e DOMI: digestible organic matter intake. b
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Table 3 Daily nitrogen (N) intake, digestion and retention by ewes fed with chopped oat hay (OC), OC supplemented with the dried residue left after molasses fermentation (EC) and an OC diet supplemented with a non-protein nitrogen/mineral mix (PP)a Diets OC Total N intake (g per day) From OC From supplement Faecal N (g per day) Urine N (g per day) Digestibility of N (%) N balance (g per day) a
5.9 5.9 0.0 3.6 1.5 37.7 0.7
EC c c
8.0 7.4 0.5 3.9 2.0 50.0 2.1
b b b
S.E.M.
P-value
0.39 0.39
0.21 0.20 2.51 0.39
n.s.
PP b b
9.6 a 8.4 a 1.2 4.2 3.2 a 56.31 a 2.1 a
b a a
Means with different letters in the same row are signi®cantly different (P < 0.05,
P < 0.01; n.s.: not signi®cant).
Table 4 Purine derivatives (PD) excretion, microbial nitrogen supply (MN) and ef®ciency of microbial nitrogen supply (EMNS) in ewes fed with chopped oat hay (OC), OC supplemented with the dried residue left after molasses fermentation (EC) and an OC diet supplemented with a non-protein nitrogen/mineral mix (PP)a Diets OC PD (mmol per day) MN (g per day) EMNS g/kg DOMIb g/kg OMADRc
6.59 b 5.43 b 20.0 33.5
S.E.M.
P-value
EC
PP
11.70 a 10.06 a
11.10 a 9.51 a
0.806 0.716
23.3 38.7
21.2 35.2
1.97 3.81
n.s. n.s.
a
Means with different letters in the same row are signi®cantly different (P < 0.01; n.s.: not signi®cant). DOMI: digestible organic matter intake. c OMADR: organic matter apparently digested in the rumen. b
(OMI, g per day) and digestible OM intake (DOMI, g per day) MNS 3:9459 0:0173
0:0047OMI; r 0:68; P < 0:01 MNS 1:6418 0:0171
0:0045DOMI; r 0:69; P < 0:01 The ef®ciency of microbial nitrogen supply was not affected by treatments (P > 0:05). 4. Discussion It is known that feed intake of ruminants is in¯uenced by the properties of food consumed by them. Feed intake usually increases when the digestibility of
the feed increases. As a signi®cant proportion of food is digested in the rumen with the aid of rumen microorganisms, the digestibility of food gives an indication of the activity or growth of rumen microbes. When the control diet was supplemented with EC, digestibility of the diet DM, OM and N were signi®cantly increased as was the feed intake and microbial nitrogen supply. The proportionate increase in DM intake was similar to that which would be predicted from the relationship between digestibility and intake of low quality pastures (Minson, 1990). Therefore, the response to the supplement appears to have been entirely associated with stimulation of rumen microbial activity and for such stimulation to occur the supplement must have provided a nutrient or nutrients that were limiting microbial growth. The control diet was theoretically marginally de®cient in rumen
Z.J. Liu, N.P. McMeniman / Small Ruminant Research 41 (2001) 229±233
digestible nitrogen (RDN). If it assumed that the nitrogen in the oat hay had a rumen degradability of 0.8 (ARC, 1980) then the mean RDN supply from the hay was 5.8 and 7.7 g per day when the control (OC) and UC diets were fed. The theoretical requirements can be calculated to have been 5.1 and 8.5 g per day (SCA, 1990), respectively. It is suggested that recycling of urea nitrogen to the rumen would have made up these small de®cits in RDN. In addition, 30 g of the EC supplement would only provide 0.3 g RDN, an amount that would not be likely to satisfy a de®ciency if one did exist. It is possible that the EC supplement supplied a mineral or minerals that were limiting microbial growth. For sheep, the control diet was de®cient in calcium, copper, cobalt, zinc and selenium (SCA, 1990). However, 30 g of the EC supplement would not have provided signi®cant quantities of any of these minerals except calcium and there is no record of calcium de®cient diets adversely affecting rumen microbial growth. Another explanation for the response to the EC supplement is that it contained other compounds that stimulated the rumen microbes. The EC supplement is a by-product of the process whereby molasses is fermented to alcohol and its organic component contains, amongst other constituents, high concentrations of B group vitamins. It is possible that these vitamins or other organic constituents were responsible for the observed responses to the supplement. It has been reported (Nangia and Sharma, 1994) that microbial protein synthesis in buffaloes was signi®cantly improved by supplementation with niacin. The PP supplement gave similar responses to the EC supplement. As mentioned previously, it is possible that the control diet was marginally de®cient in rumen degradable nitrogen and that the PP supplement satis®ed this requirement. Another possibility is that there was suf®cient EC component in the PP supplement (10%) to result in the response that was obtained. More work would be required to isolate the factor(s) responsible for the responses to both the EC and PP supplements. Although daily microbial nitrogen ¯ow to the small intestine increased signi®cantly with supplementation of EC and PP, the ef®ciency of microbial protein production was not signi®cantly affected. As microbial nitrogen production is dependent on ME intake (SCA, 1990), the increase in production associated with supplementation can be explained by the
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increased intake of DOM when the supplements were fed. Nitrogen balance was also signi®cantly improved by supplementation with no signi®cant difference between the two supplemented diets, although the nitrogen intake for the PP supplemented diet was higher than for EC supplemented diet. While microbial nitrogen production with the two supplemented diets was similar, urine nitrogen excretion was signi®cantly higher for PP diet. This indicates that the increased intake of nitrogen in the form of urea in the PP supplemented diet was degraded and excreted in urine and not used for microbial synthesis. This is a further indication that the control diet was not de®cient in RDN and the increase in nitrogen balance of the sheep fed the supplemented diets was solely due to the increased microbial nitrogen supply. In conclusion, improved intake, dry and organic matter digestibility, microbial nitrogen production and nitrogen balance may be achieved when low quality forage diets are supplemented with the spray dried residue that is left after molasses is fermented to alcohol.
References ARC, 1980. The Nutrient Requirements of Ruminant Livestock. Agricultural Research Council. Commonwealth Agricultural Bureaux, Farnham Royal, UK. Balcells, J., Guada, J.A., Peiro, J.M., Parker, D.S., 1992. Short communication: simultaneous determination of allantoin and oxypurines in biological ¯uids by high-performance liquid chromatography. J. Chromatogr. 575, 133±157. Chen, X.B., Gomes, M.J., 1992. Estimation of microbial protein supply to sheep and cattle based on urinary excretion of purine derivatives Ð an overview of the technical details. Occasional Publication of the International Feed Unit, Rowett Research Institute, Buckburn, Aberdeen AB2 9SB, UK. Durand, M., Komisarczuk, S., 1987. In¯uence of major minerals on rumen microbiota. J. Nutr. 118, 249±260. Hungate, R.E., 1966. The Rumen and its Microbes. Academic Press, New York. Liu, Z., 2000. Microbial N ¯ow in the duodenum of sheep estimated using purine derivative excretion in the urine. Ph.D. Thesis, University of Queensland, Australia. Minson, D.J., 1990. Forage in Ruminant Nutrition. Academic Press, San Diego, USA. Nangia, O.P., Sharma, R., 1994. In¯uence of niacin supplementation on rumen fermentation and microbial protein synthesis in buffaloes. Indian Vet. J. 71, 978±982. SCA, 1990. Feeding Standards for Australian Livestock. Standing Committee on Agriculture, CSIRO, Melbourne, Australia.