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Presented at Cukurova University, Animal Science Dept.

Turkey , 30/11/2012

TITLE REVIEW : ENERGY EQUATION IN RUMINANT NURITION Muhamad Nasir Rofiq (Doctorate Student, Anmal Science Departement, Institute of Applied Science, Cukurova University)

Abstract This paper provides summative energy equation of ruminant nutrition and applies it to estimate the energy value of feed. Energy is the primary nutrient contributed by feed (TMR, forage and concentrate feed) . Some equation was developed using chemical compound (proximate analyzed or carbohydrate and protein fraction), digestibility experiment, rumen gas fermentation experiment and combined analyzed. Energy equation also was separated to classes such as forage, concentrate, animal source, fat feed and fatty acid. Specially for this paper is giving information about energy equation in ruminan nutrition by feed proximate fractionation, feed CNCPS fractionation, in vitro rumen gas fermentation, and in vitro rumen digestibility.

INTRODUCTION Feed is generally the major operating cost associated with livestock and ruminant operations. Over feeding situation is wasteful while underfeeding will decrease performance and profitability. Optimum feeding of animals is obtained by good

formulated feed which was contains nutrients enough and bio availabililty. To obtain good formulated feed, nutrient contain of feeding stuff should be meet to animal nutrient requirement. Nutrient in feeding stuff as chemical compound were analyzed at laboratory as procedure. But nutrient requirement f animal should be analyzed by bio research. Recently the result from chemical compound of feed and bio research could be use as mathematic formulation to estimate nutrient requirement and nutrient quality. Feeding stuffs of ruminant has high variety physical and chemical properties. It made high variety nutritive value for some of feeding stuffs (forages, by product, and some animal meals). The variety was caused by agronomic characteristics, sampling, geographic, and processing of feedstuffs. Their data were recorded well in nutrient list feed table. In feeding stuff tables, chemical and nutritive data are reported as well as
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

some brief information to characterize the feed (NRC, 1982, 2001; INRA, 1980, 1988, FAO, 2012). The fundamental characteristic of formulated rations for ruminant around which all other nutrients are structured, is its energy content. Its level energy in formulated ration is the sum of energies in its component feeds, but it cannot be analyzed as it represents the potential of feed, and its components, to do work as biological products, such as meat or milk or as heat. Recently accurate knowledge of the energy content of feeds is central to formulation of rations, which will maximize animal output of usable products, and minimize output of unused nutrients (i.e.waste , heat and gas). It could be estimated by bio experiment which its result is mathematic formulation as an equation to estimate their content. The purpose of this article is to review some energy equation in ruminant nutrition was resulted by some researches and applies to TMR, alfalfa hay and concentrate feed using data of chemical of feed, in vitro gas fermentation and in vitro true digestibility.

FEED CHEMICAL COMPOUND

The nutritive value of a feed is determined by the contribution it is able to make to nutritional requirements (energy, protein, and mineral) of the class of animal. Feed Analysis, most commercial laboratories offer standard feed test for forages, grain, or total mixed rations. Chemical nutrient Analyses in feed mostly for proximate analyze and van soest analyze to determine moisture, Crude protein, Extract ether, crude fiber, ADF and NDF is recommended when designing diets for ruminant. Furthermore we may wish to identify key mineral or minor nutrient interest. We should also know gross energy in feed analyzed by bomb calorimeter. Typically, results are reported on as fed and dry matter basis. After formulation with dry matter basis for nutrient balanced, values convert to an as fed basis. The result can estimate energy value of feed and digestibility. The original system for analyzing feeds, the Weende system of Proximate analysis was developed in the 1860s by Henneberg and Stohman at the Weende Experimental station in Germany. Their method was to separate feed into the nutrient components needed by the animal. That is water, crude protein, crude fat or ether extract,
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

ash or mineral matter, crude fiber, the indigestible fraction of the carbohydrates present and nitrogen free extract (NFE) the readily digestible carbohydrate fraction. The substantial amount of crude fiber (CF) where as much of the NFE in some feeds is not digested. This problem prompted van soest in 1963, to develop his system of detergent fiber analysis which determine lignin the totally indigestible fraction of ruminant feeds as separate entity (figure 1).

Figure 1. Schematic of detergent system of forage analysis (Van soest, 1996)

Inter fractionation of feed chemical nutrient between proximate system and detergent system (van soest) explain relation between their nutrient and substitute nutrient value each other (Figure 2). For further analysis, especially for energy source from carbohydrates estimated by carbohydrates fractionation analysis was supported by data analyzed.

Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

Figure 2. inter fractionation of chemical feed analysis between proximate and Van soest

Figure 3. Classification of forage fraction using van soest method

Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

Figure 4. Fractionation of plant carbohydrates

Recently, there is updated system for feed analysis which was used carbohydrate and protein data of feed. The system was the net carbohydrate and protein system (CNCPS) which was developed by Cornell University Nutrient managmen Planning System (UCD). The system was classified feed chemical into 8 components : 1. Dry matter (DM), 2. ash, 3. Estract ether (EE), 4. Neutral detergent fiber (NDF) and Acid detergent Lignin (ADL), 5. Total crude protein (CP), 6. Soluble protein (SCP), 7. Neutral detergent insoluble CP (NDICP) and Acid detergent insoluble CP (ADICP), 8. Non structural carbohydrate (NSC)

The CNCPS feed evaluation system data could be used for energy estimation using some equation which was developed combine with other data (in vitro digestibility or in vivo digestibility).
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

Figure 5. The CNCPS scheme for feed carbohydrate fractionation (Modified from Sniffen et al., 1992 in CNCPS ver 3. Manual and references, 2003)

Figure 6. The CNCPS scheme for Protein fractionation (Modified from Roe et al., 1990 in CNCPS ver 3. Manual and references, 2003 )

Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

ENERGY IN RUMINANT FEED

The energy in feed is a measure of that feeds ability to help the animal function and be productive. All feed have a gross energy value. Some of the gross energy (GE) is lost in feces. Some are absorbed by animal as digestibility energy (DE). From DE, some of them lost in urine, gas and heat to provide metabolisable energy (ME) to animal. The ME was used to maintenance, milk production, keep condition, activity, pregnancy and growth (Figure 7). The ME is energy available for ruminant as feed quality indicator.

Figure 7. The flow and partitioning of dietary energy through the ruminant (Moran, J. 2005) Energy can come from various part of the feed. Carbohydrates, fats, oils and even protein can provide energy.

Carbohydrat, plant tissue dry matter is about 75% carbohydrates. Carbohydrates are the main source of energy for livestock. Sugar molecules of various types are the building blocks of carbohydrates. The sugar are chemically bound together in different numbers and in variety of ways to form the three types of carbohydrate : soluble, storage and structural. Soluble carbohydrates are the simple or individual sugar found in cells of growing plants. They are digested and used almost instantly by the microbes in the rumen. Soluble
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carbohydrates are digested 100 times faster than storage carbohydrates. Storage carbohydrates are made up of sugar subunits which are chemically bound together and are found inside plant cells. Starch in example of a storage carbohydrates. Storages carbohydrates are digested 5 times faster than structural carbohydrates,. It is founded in grains, leaf, stem and in the bulbous roots of fodder crop. Structural carbohydrates are fibrous components of plant cell walls. They provide the structural support that plants need to grow upright. Pectin, hemicelluloses and cellulose are all structural carbohydrates. Lignin and silica are often associated with structural carbohydrates in plants. They give structural support toplants but are indigestible and are not actually carbohydrates. The can bind the structural carbohydrates and make them less digestibility. Fats and Oils, only 2% until 3% of forages are fat or oil. Fats and oils include vegetable oils and animal fat and processed fat. Fat can decrease the palatability of the diet and can coat fiber, interfering with its digestion by rumen microbes. Protein, the rumen microbes can use a surplus of protein in the rumen fro energy. This is, however and inefficient use of protein. (Semua sumber energy diatas dikasih contoh dan berapa jumalh energinya). Energy as feed quality indicator can be measured to three measurement, they are digestibility, metabolisable energy(ME) and total digestibility nutrient (TDN). Digestibility relates to the proportion of feed which is not excreted in the feces and so is available for use by ruminant. It is not direct measure of energy, but it does indicate overall feed quality. It is commonly measured as a percentage. The digestibility of various feed constituents can be determined with Organic Material Digestibility (OMD) as percentage of digestible organic matter per total dry weight. Metabolisable energy content of a feed also called its energy density is measured as megajoules of ME per kilogram of dry matter (MJ /Kg DM), or Megacalories per kilogram of dry matter (Mcal/Kg DM). the intake of ME is expressed in MJ/day or Mcal/day. Total Digestible Energy (TDN) is an older energy system used in USA and some countries. It is not used formally in Australia and England. It is a less accurate measurement of energy than ME. It does not take into account energy loss via gas (methane,) and urine. TDN expressed as an percentage, with total digestible nutrients intake expressed in Kg/day.
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

THE PROBLEM FOR ENERGY ESTIMATION

Prediction of energy availability from laboratory analyses usually requires specific equations for each type of feed. The accuracy of energy estimation is a function of the accuracy of laboratory analyses and the accuracy of the bio experimentation used to develop estimation equation. Digestibility and energy value can be measured under a variety of conditions that influence the resulted values. Cattle, sheep and dairy cows will obtain different digestibility for the same feed. In addition,the level of feed intake has a significant effect on digestibility and the utilization of its energy. Major variable affecting the measurement of digestibility is the amount of selection allowed by the animal. They have difference feed preferences. However, most scientist measure ad libitum intake and digestibility in the same trial by offering the animals 5 15 % more than they consume. Others problem is that the botanical description of feedstuffs and time of year that it was harvested for forage. They have difference nutrient value, so that they have difference equation for estimation energy availability. Type of feed processing was obtained from industrial waste or agricultural industrial also effect on feed nutrient value and digestibility.

ESTIMATION ENERGY IN RUMINANT NUTRITION

Basic Principle for estimation of available energy and digestibility is estimated from chemical composition and bio experiment. There were two factors that determine the energy value of feedstuff for ruminant are its content of fat, due to its high energy value, and the digestibility of its structural fibre (i.e. NDF), due to its generally high content in forage. Most energy values are predicted from fiber analyses because fiber is negatively related to the animals ability to digest and use nutrient in the feed.

The traditional and still most common, approach to estimating the energy value of feedstuff has been to calculate its total digestible nutrient (TDN) level using a factorial equation based upon analyzable component of feedstuffs. Many equation calculate TDN
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

as the sum of digestible crude protein (CP), digestible fat (multipled by 2.25), digestible neutral detergent fiber (NDF) and digestible non-structural carbohydrate (NSC) all corrected for a metabolic cost of digestion by the animal. The estimated of TDN can be used to estimate the digestible energy (DE), metabolisable energy (ME) and/or NEl values of individual feedstuffs.

TDN% = Digestible CP + Dig.CF + Dig. Nitrogen free extract (NFE) + Dig.fat x 2.2.5

Some Estimation Energy from Chemical compound of Feed

Some energy estimation were approached using chemical compound (Table ). The some of variables used in the equation was indicate the correlation between the variable and energy balue (ME, TDN , NEl, etc).

Table 1. Some energy estimation equation for feed evaluation using chemical components of feed 1 D.R.Martens Legumes %TDNm= 86.2 - ( 0.513 x NDF ) NEL(Mcal/lb)= 1.054 - ( 0.0098 x NDF ) %TDNm= 84.2 - ( 0.598 x ADF ) NEL(Mcal/lb)= 1.011 - ( 0.0113 x ADF ) Grasses %TDNm= 105.2 - ( 0.667 x NDF ) NEL(Mcal/lb)= 1.297 - ( 0.119 x NDF ) %TDNm= 97.6 - ( 0.974 x ADF ) NEL(Mcal/lb)= 1.12 - ( 0.0159 x ADF ) 2 Moran, Jhon TDN(%)=5.31 + 0.412 CP% + 0.249 CF% + 1.444 EE% + 0.937 NFE% 2005 TDN (%)= 5.4 ME + 10.2 ME (Mcal.kg DM) = 0.185 TDN 1.89 ME (MJ/kg DM) = 11.78 + 0.00654CP + (0.000665EE) 2 Alderman, CF(0.00414EE) 0.0118A (UI eq. 25. Page 29) 3 1985 Variables are in %DM ME (Kcal/kg OM) = 3260+0.455CP + 3.517EE - 4.037CF 4. TSE(9610) Variables are in g/Kg OM 5. MAFF (84) ME (Mcal/kg DM) = 3227 - 35.85%ADF + 33.46%EE - 35.85%Ash
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Energy estimation (eq. (25 UI)) resulted from Alderman (1985) was selected for legislation for estimate ME values. Equation (25) (UI) has a square term for EE, which improves the fit with high energy (high fat) feeds. This agrees with what is known about the metabolism of fat, but implies non-linearity of the ME of fat when added to diets. Other energy estimation using chemical such as MAFF (1984), TSE(9621) and Moran J (2005) using different chemical compound of feed.

NRC (2001) Energy Estimation

Energy estimation approached by NRC (2001) equation was mostly used for feed evaluation. The dietary energy requirements in NRC (2001) consider feedstuff digestion dynamics as well as the energy requirements for maintenance, growth, lactation, reproductive status and activity of the animal (linn, J. 2003). The approach used in the NRC (2001) is to calculate the enrgy value of feeds and diest directly from their nutrient composition. the chemical component requirements and in vitro assay : dNDF48 : Ash, fat, crude protein, ADICP, NDF, 48h NDF, PAF Lignin : Ash, fat, crude protein, ADICP, NDF, lignin, PAF.

The NRC-2001 energy equation system was evaluated by VandeHaar (2002) which was reported by Linn J (2003) that found some legitimate concerns with the energy system when it is applied to formulation of diets. These were : Energy value of protein feed may be over evaluated. The effect of the over is the energy concentration in diets can be increased by feeding higher protein diets. The NDF digestibility equation does not consider feed type. Variable effect of feed type due to affect of lignin contain and in vitro digestibility of NDF after 48 hour does not substitute directly for the calculated NDF digestibility value (Wisconsin research), and the in vitro dNDF48 h was founded had higher value than in sacco NDF digestion (Robinson, P.H. et al., 1999) For high energy feed or diet, the digestibility discount may be too aggressive. Fiber length of feed or particle size is not accounted for in the model ad this will impact DMI.

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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

The NRC-2001 equation can be found on page 13 to 17 of nutrient requirements of dairy cattle in NRC (2001). The equation for calculating could be started from true digestible (td) of feed fraction then estimated DE1x or TDN1x, here the equation for most feed from NRC (2001) :
tdCPf (%) = [(CP*exp(-0.012*ADICP/CP))] tdCPc (% ) = [(1- (0.04*ADICP/CP)]*CP tdNFC(%) = 0.98*(100-[(NDF NDICP) + CP + EE + Ash])*PAF tdFA (%) = FA, note : if EE < 0 Fa = 0, if EE>1 FA = EE 1 tdNDF (%) = [0.75 * ((NDF-NDICP) Lignin)* (1-(lignin/(NDF-NDICP))0.667 or = tdNDF 48 hour or = tdNDF30 h * 1.16 DE1x (MCal/kg DM) = (tdNDF/100)*4.2 + (tdNDF/100)*4.2 + (tdCP/100)*5.6 + (FA/100)*9.4 0.3 TDN1x (%) = DE/0.04409 If TDN = < 60%, the discount for diets was set to 1.0 (no discount) If TDN > 60%, DE1x should be corrected with discount for productive level DEp Discount = [(TDN1x [(0.18*TDN1x0-10.3)] * intake)]/TDN1x Intake = 2, (3 -1) for cow 3x maintanenace for above maintenance DEp (MCal/kg) = DE1x*Discount MEp (Mcal/Kg) = [1.01*(DEp) 0.45] + 0.0046*(EE-3) , if EE > 3 MEp(Mcal/Kg) = 1.01*DEp 0.45, if EE < 3 NELp (Mcal/kg) = [0.703**MEp)] 0.19 NEm (Mcal/kg) = 1.37ME 0.138 ME2 + 0.0105ME3 1.12 NEg (Mcal/kg) = 1.42ME 0.174 ME2 + 0.0122ME3 1.65 NRC (20010)

UCD Energy Estimation UCD energy estimation was tried to correction the weakness of NRC (2001) energy system with different time NDF digestibility (dNDF 30 h) and considered the protein soluble contain of feeds. UCD energy estimation used chemical component of feed and in vitro gas production test. The chemical components requirements were : dNDF30 : Ash, fat, crude protein, soluble CP, ADICP, NDF and 30 h NDF Gas : 24 h gas production in vitro.

The estimation from 24 hour gas production was a modification of the method of Menke and Steingass (1988) as well as estimates of the feeds level of CP and Fat.
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TDN1x (%) = ((CP-ADICP)*(FT/5)*0.98)+((CPADICP)*(1-(FT/5))*0.80) + ((EE-1)*0.98*2.25) + (NDF*dNDF) + (0.98*(100-ASH-EE-NDF-CP))) DE1x (Mcal/Kg) = (0.04409 x TDN1x) ME1x (Mcal/Kg) = ((DE1x)*1.01) 0.45 NEL 1x (MCal/Kg) = (TDN1x*0.0266) 0.12 Discount = ((0.033 + (0.132*NDF(%DM))) (0.033*NEL1x,Mcal/Kg))) + (NSC(%DM)*0.05) NEL3x (MCal/KgDM) = NEL1x ((NEL1x)*Discount energy/100)*intake)) Intake = 2 , from (3x maintenance 1) NEm(MCal/KgDM)= ((1.37*(DE1x*0.82))- ((0.138*(DE1x*0.82)^2)) + ((0.0105*(DE1x)*0.82)^3)) 1.12 NEG(MCal/KgDM)= ((1.42*(DE1x)*0.82))- ((0.174*(DE1x)*0.82)^2)) + ((0.0122*(DE1x*0.82)^3)) 1.65
Where : CP = crude protein (% of DM) ADICP = acid detergent insoluble CP (% of DM) FT = feed type (1=silage,2 =wet by-product, 3 = others) EE = ether extract (% of DM) NDF = ash-free NDF assayed with sodium sulphite and amylase (% of DM) dNDF = in vitro NDF digestibility at 30 hours (% of NDF) Ash = ash (% DM) NE1 = energy value at 1xM intake NSC = non-fibre carbohydrate calculated as 100-ASH-EE-NDF-CP DE1x= Digestible energy at maintenance level NEL1x = Net energy lactation at maintenance level NEL3x = Net energy lactation at production level NEm = Net energy maintenance at production level NEG = Net energy gain at production level

Robinson P.H. (2001)

TDN1x is the TDN value of a feed or diet at maintenance intake (NRC, 2001). Because NEL1x at maintenance is not representative of the energy value of feed or diet at production level, a discount factor was developed to correct for decrease net energy level at production (NEl, 3x). In UCD factorial approach to estimate feed energy levels. A discount energy factor was formulated based on NDF and NSC content of feedstuff and NEl(1xM) values as % per unit of energy intake (M). Net energy values for lactation at level production (NEl 3xM). The energy

content of a feedstuff is not constant value. As its intake by the animal increase, its energy content tends to decline, since it passes through the intestine faster allowing rumen microorganism and intestinal enzymes less time to digest available nutrient. To
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

extent of the change, refer to as the energy discount or simply discount, quantity the extent of this change. The discount is a reflection of the NDF and NSC content of the feedstuff, and it can be calculated as % per unit of energy intake .

ME (MJ/kg DM) = 1.25 + (0.0292*gas24) +(2.46*fat) + (0.0143 c (CP-ADICP))


Where : gas24 is gas production in 24 hour (ml/g DM) and others variable are in g/kg DM

The UCD equation evaluation result in low value of precision of prediction value (r2) for UCD gas (0.50) , while it had higher value for UCD30 (0.83) than NRC48 or NRClig.

Energy Equation in TMR (Boguhn J. et. al 2003)

Total mixed ration, complete diet for ruminant was less predicted by equation and have so far not beed developed, and TMR can only be evaluated if the composition and the energy value of the ingredients ae known. Boguhn J. et al. (2003) reported prediction of ME in TMR using chemical feed data, gas production and enzymatically degraded organic matter (EDOM).
ME (MJ/kg DM) = 6.0756 + 0.19123EE + 0.02459CP 0.000038CF.CF 0.002139 EE.EE 0.000060 CP.CP ( r = 0.83) ME (MJ/kg DM) = 8.9695 + 0.04095GP - 0.01267CF + 0.04108EE + 0.00387CP + 0.00508Ash ( r = 0.83) ME (MJ/kg DM) = 13.903 0.0147 Ash 0.01114 EUDOM + 0.00234CP (r = 0.82)
Where : GP = gas production in ml/200 mg DM, others variable in g/kg DM EUDOM = enzumatic undegraded organic matter

The equation was determined from 30 TMR which was analyzed for nutrients, gas production and enzymatic degraded or undegraded organic matter. The equation no.3 and 8 from table 4 pages 263 was recommended (Boguhn J. et al., 2003).
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Presented at Cukurova University, Animal Science Dept. Turkey , 30/11/2012

RESUME

Estimation energy value of feed by some equation raised overestimated or underestimated value from truly energy value. It should be considered four using selected equation depend on available data. Using chemical component analyze only was considered for some type of feed which clearly information known. Best estimation of energy value of feed is using combination between chemical component of feed and digestibility data (in vitro or in vivo). NRC-2001 system for energy estimation which was commonly used and recorded in NRC table still not enough precision for some feed. However the UCD30 was resulted in some correction for it.

Acknowledgements

The authors thanks Professor Murat Gorgulu and Professor Hasan Rustu Kutlu for literature sources and attention.

REFFERENCES

Alderman, G. 1985. Prediction of the energy value of compound feeds : In Haresign W and D.J.A Cole. Recent Advances in Animal Nutrition 1985 . Univ of Nottingham. UK. Boguhn J., H. Kluth, O. Stenhofel, M. Peterhansel and M. Rodehutscord. 2003. Nutrient digestibility and prediction metabolizable energy in total mixed ration for ruminants. Arch. Anim. Nutr., 57(4) : 253-266. Fox. D. G., T.P. tylutki, L.O. Tedeschi, M.E. van Amburgh, L.E. Chase, A.N. Pell, T.R. Overton and J.B. Russell. 2003. The net carbohydrate and protein system for evaluating herd nutrition and nutrient excretion. Anim. Sci. Mimeo Model Doc. Cornell University. Gorgulu, M. 2010-2011. RUMNANT YEMLERNDE SE, ME, TDN, NEM, NEG, NEL DEERLERNN HAM BESIN MADDELERNDEN HESAPLANMASI. Article. http://muratgorgulu.com.tr/altekran.asp?id=97

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Linn, J. 2003. EEnergy in the 2001 Dairy NRC : understanding the system. Proceeding of the Mnnesota Dairy Health Conference. Univ. Minesota.

MAFF (Ministry of Agriculture, Fisheries and Food). 1984. Energy allowances and feeding systems for ruminants. Reference Book 433. HMSO (Her Majesty's Stationery Office), London, UK. 85 pp. Martens. D.R. 1987. Predicting intake and digestibility using mathematical models of ruminal function. J. Anim. Sci. 64, 1548-1558

Moran, J. 2005. Tropical dairy farming : feeding management for small holder dairy farmer in the humid tropics. Landlink press. P.312 National Research Council. 1989. Nutrient Requirements of Dairy Cattle. 6th rev. Ed. National Academy of Sciences, Washington DC National Research Council. 2001. Nutrient Requirements of Dairy Cattle. 7th rev. Ed. National Academy of Sciences, Washington DC. Robinson, P.H., M. C. Mathews, J.G. Fadel. 1999. Influence of storage time and temperature on in vitro digestion of neutral detergent fibre at 48 h, and comparison to 48 h in sacco neutral detergent fiber digestion. Anim. Feed and Tech. 80 : 257-266 Robinson, P.H. 2001. Estimating the energy value of corn silage and other forages. Proceeding 31st California Alfalfa and Forage symposium. UCD. Robinsonp.H., D.I. Givens, G. Getachew. 2004. Evaluation of NRC, UC Davis and ADA approaches to estimates the metabolizable energy values of feeds at maintenance energy intake from equation utilizing chemical assays and in vitro determinations. Anim. Feed Sci. and Tech. 114: 75-90 VandeHarr, M.J. 2002. Energy and protein in the 2001 dairy NRC : challenges for a ration formulation program. Tri-State (IN, MI, OH) dairy nutrition Conf. Fort Wayne, IN, pages 81-98

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