Partitioning of amino acids flowing to the abomasum into feed, bacterial, protozoal, and endogenous fractions

We partitioned the flow of amino acids (AA) to the abomasum among rumen undegradable protein (RUP) and bacterial, protozoal, and endogenous fractions using four Holstein cows in midlactation that were equipped with ruminal and abomasal cannulas. A 2 x 2 factorial design with four diets, combinations of high or low ruminally degradable organic matter, and rumen degradable protein, was employed. Crude protein (CP) and AA contents of ruminal bacteria and protozoa and abomasal digesta were determined. Equations for the source compositions and in vivo flows of CP and 16 AA were then solved simultaneously with a linear program to estimate the contribution of RUP, bacterial, protozoal, and endogenous CP to AA flows. The flows of RUP and bacterial AA were not affected by diet. Low dietary RDP increased the flow of protozoal AA to the abomasum, but the ruminally degradable organic matter content of the diet did not affect protozoal AA flow. Across diets, RUP, bacterial, protozoal, and endogenous fractions provided 55, 33, 11, and <1% of the CP, and 62, 26, 12, and <1% of the AA that reached the abomasum. The linear program was a useful tool for partitioning AA that flows to the abomasum. The technique may also allow dietary effects on ruminal microbes and the AA profile of protein flowing to the duodenum to be better understood and perhaps manipulated.     

Evaluation of the National Research Council (2001) dairy model and derivation of new prediction equations. 2. Rumen degradable and undegradable protein

This work evaluated the National Research Council (NRC) dairy model (2001) predictions of rumen undegradable (RUP) and degradable (RDP) protein compared with measured postruminal non-ammonia, nonmicrobial (NANMN) and microbial N flows. Models were evaluated using the root mean squared prediction error (RMSPE) as a percent of the observed mean, mean and slope biases as percentages of mean squared prediction error (MSPE), and concordance correlation coefficient (CCC). The NRC (2001) over-estimated NANMN by 18% and under-estimated microbial N by 14%. Both responses had large mean biases (19% and 20% of MSPE, respectively), and NANMN had a slope bias (22% of MSPE). The NRC NANMN estimate had high RMSPE (46% of observed mean) and low CCC (0.37); updating feed library A, B, and C protein fractions and degradation rate (K_d) estimates with newer literature only marginally improved fit. The re-fit NRC models for NANMN and microbial N had CCC of 0.89 and 0.94, respectively. When compared with a prediction of NANMN as a static mean fraction of N intake, the re-derived NRC approach did not have improved fit. A protein system of intermediate complexity was derived in an attempt to estimate NANMN with improved fit compared with the static mean NANMN model. In this system, postruminal appearance of A, B, and C protein fractions were predicted in a feed-type specific manner rather than from estimated passage and degradation rates. In a comparison to independent data achieved through cross-validation, the new protein system improved RMSPE (34 vs. 36% of observed mean) and CCC (0.42 vs. 0.30) compared with the static mean NANMN model. When the NRC microbial N equation was re-derived, the RDP term dropped from the model. Consequently, 2 new microbial protein equations were formulated, both used a saturating (increasing at a decreasing rate) form: one saturated with respect to TDN and the other saturated over increasing intakes of rumen degraded starch and NDF. Both equations expressed maximal microbial N production as a linear function of RDP intake. The function relating microbial N to intake of rumen degradable carbohydrate improved RMSPE (24 vs. 28% of the observed mean) and CCC (0.63 vs 0.30) compared with the re-derived NRC model. The newly derived equations showed modest improvements in model fit and improved capacity to account for known biological effects; however, substantial variability in NANMN and microbial N estimates remained unexplained.     

Improved Feed Protein Fractionation Schemes for Formulating Rations with the Cornell Net Carbohydrate and Protein System

Adequate predictions of rumen-degradable protein (RDP) and rumen-undegradable protein (RUP) supplies are necessary to optimize performance while minimizing losses of excess nitrogen (N). The objectives of this study were to evaluate the original Cornell Net Carbohydrate Protein System (CNCPS) protein fractionation scheme and to develop and evaluate alternatives designed to improve its adequacy in predicting RDP and RUP. The CNCPS version 5 fractionates CP into 5 fractions based on solubility in protein precipitant agents, buffers, and detergent solutions: A represents the soluble nonprotein N, B1 is the soluble true protein, B2 represents protein with intermediate rates of degradation, B3 is the CP insoluble in neutral detergent solution but soluble in acid detergent solution, and C is the unavailable N. Model predictions were evaluated with studies that measured N flow data at the omasum. The N fractionation scheme in version 5 of the CNCPS explained 78% of the variation in RDP with a root mean square prediction error (RMSPE) of 275 g/d, and 51% of the RUP variation with RMSPE of 248 g/d. Neutral detergent insoluble CP flows were overpredicted with a mean bias of 128 g/d (40% of the observed mean). The greatest improvements in the accuracy of RDP and RUP predictions were obtained with the following 2 alternative schemes. Alternative 1 used the inhibitory in vitro system to measure the fractional rate of degradation for the insoluble protein fraction in which A = nonprotein N, B1 = true soluble protein, B2 = insoluble protein, C = unavailable protein (RDP: R² = 0.84 and RMSPE = 167 g/d; RUP: R² = 0.61 and RMSPE = 209 g/d), whereas alternative 2 redefined A and B1 fractions as the non-amino-N and amino-N in the soluble fraction respectively (RDP: R² = 0.79 with RMSPE = 195 g/d and RUP: R² = 0.54 with RMSPE = 225 g/d). We concluded that implementing alternative 1 or 2 will improve the accuracy of predicting RDP and RUP within the CNCPS framework.     

Animal and dietary factors affecting feed intake during the prefresh transition period in Holsteins

Parity, body condition score (BCS), and dry matter intake (DMI) data of 699 Holsteins fed 49 different diets during the final 3 wk of gestation (prefresh transition period) were compiled from 16 experiments conducted at eight universities. The objectives of this study were to determine the effects of animal and dietary factors on DMI and to elucidate interactions between animal and dietary factors and among dietary factors on DMI during the prefresh transition period. Animal factors examined were parity and BCS, whereas dietary factors examined were rumen undegradable protein (RUP), rumen degradable protein (RDP), neutral detergent fiber (NDF), and ether extract (EE). DMI decreased 32% during the final 3 wk of gestation, and 89% of that decline occurred during the final week of gestation. Day of gestation, animal factors, and dietary factors accounted for 56.1, 19.7, and 24.2% of explained variation in DMI, respectively, and R2 of this linear multivariable model was 0.18. Cows had higher DMI than heifers. DMI decreased linearly as BCS, RUP, and NDF increased, decreased quadratically as EE increased, and increased quadratically as RDP increased. Moreover, the magnitude of DMI depression as animals approached parturition was affected by characteristics of animals and dietary nutrient composition. There were significant parity x EE, BCS x NDF, RUP x NDF, RDP x NDF, NDF x EE, and RUP x EE interactions on DMI. In conclusion, parity, BCS, and concentrations of organic macronutrients in diets affected DMI during the prefresh transition period, and the magnitude of DMI depression as animals approached parturition.     

Potential protein degradation balance and total metabolizable protein supply to dairy cows from heat‐treated faba beans

The effects of pressure toasting (100, 118 and 136 °C for 3, 7, 15 and 30 min) on potential protein nutritional value of faba beans were evaluated with the NRC 2001 dairy model, by determining undegraded (RUP) and degraded rumen protein (RDP), undegraded (RUST) and degraded rumen starch (RDST), truly absorbed undegraded protein (ARUP), microbial protein (MCP_RDP) synthesized in the rumen from rumen‐available protein, truly absorbed rumen synthesized microbial protein (AMCP), truly absorbed rumen endogenous protein (AECP), total metabolizable protein (MP) in the small intestine, and the protein degradation balance (PDB). The treatments increased RUP, RUST, ARUP and MP (p < 0.001), and decreased RDP, RDST, MCP_RDP and PDB (p < 0.001), the effects increasing with increasing temperature and time. The treatments increased (p < 0.001) ARUP without affecting AECP and AMCP, so that the net absorbable total MP in the small intestine was increased. The PDB was reduced (p < 0.001) but never became negative. These results indicated that potential microbial protein synthesis would not be impaired due to sufficient nitrogen in the rumen, but the high positive PDB values with most treatments, except 136 °C for 15 min (PDB 2.0 g kg^?1 DM) indicated that there were large potential losses of nitrogen in the rumen, particularly for the control with a value of 88.9 g kg^?1 dry matter. It is concluded that predicted potential protein degradation balance and total metabolizable protein supply from faba beans were improved by the treatments. Copyright © 2005 Society of Chemical Industry     

Lowering rumen-degradable and rumen-undegradable protein improved amino acid metabolism and energy utilization in lactating dairy cows exposed to heat stress

The objective of this study was to evaluate the effects of reducing dietary rumen-degradable protein (RDP) and rumen-undegradable protein (RUP) on protein and energy metabolism in heat-stressed dairy cows. Eighteen primiparous and 30 multiparous mid-lactation Holstein cows were used in a completely randomized design arranged in a 2 × 2 factorial (n = 12/treatment). Cows were randomly assigned to 1 of 4 dietary treatments that included 2 levels of RDP (10 and 8%; D) and 2 levels of RUP (8 and 6%; U) of dry matter for 21 d as (1) 10D:8U, (2) 8D:8U, (3) 10D:6U, and (4) 8D:6U. Diets were isoenergetic and contained 50% forage and 50% concentrate (dry matter basis). Cows were housed in a freestall barn. Three weeks before start of treatments, all animals were fed the 10D:8U diet and received supplemental cooling to prevent heat stress. During the treatment period, cows experienced a daily increment in temperature-humidity index from 74 to 82 for 1000 to 2000 h. Blood samples were collected on d ?1 and 21 of the treatment period to determine plasma concentrations of AA, glucose, insulin, fatty acids, and β-hydroxybutyrate. For primiparous cows, reducing from 10 to 8% RDP decreased insulin concentrations. For multiparous cows, we found significant RDP by RUP interactions for insulin, β-hydroxybutyrate, fatty acids, total essential AA, and 3-methylhistidine concentrations. Reducing from 10 to 8% RDP decreased insulin concentrations at 6% RUP, but concentrations did not change when reducing RDP at 8% RUP. Reducing from 10 to 8% RDP decreased β-hydroxybutyrate concentrations at 8% RUP, but concentrations did not change when reducing RDP at 6% RUP. Reducing from 10 to 8% RDP increased nonesterified fatty acid and total essential AA concentrations at 8% RUP, but concentrations did not change when reducing RDP at 6% RUP. Reducing from 8 to 6% RUP decreased 3-methylhistidine concentration at 8% RDP, but not at 10% RDP. Reducing from 8 to 6% RUP increased milk protein yield efficiency in primiparous and multiparous cows. These results indicate that reducing RDP and RUP lowers circulating insulin, which was associated with mobilization and utilization of fatty acids. Reduced RDP and RUP increases the use of AA to maintain milk protein synthesis and limit AA catabolism in cows exposed to warm climates.     

Predictions of ruminal outflow of essential amino acids in dairy cattle

The objective of this work was to update and evaluate predictions of essential AA (EAA) outflows from the rumen. The model was constructed based on previously derived equations for rumen-undegradable (RUP), microbial (MiCP), and endogenous (EndCP) protein outflows from the rumen, and revised estimates of ingredient composition and EAA composition of the protein fractions. Corrections were adopted to account for incomplete recovery of EAA during 24-h acid hydrolysis. The predicted ruminal protein and EAA outflows were evaluated against a data set of observed values from the literature. Initial evaluations indicated a minor mean bias for non-ammonia, non-microbial nitrogen flow ([RUP + EndCP]/6.25) of 16 g of N per day. Root mean squared errors (RMSE) of EAA predictions ranged from 26.8 to 40.6% of observed mean values. Concordance correlation coefficients (CCC) of EAA predictions ranged from 0.34 to 0.55. Except for Leu, all ruminal EAA outflows were overpredicted by 3.0 to 32 g/d. In addition, small but significant slope biases were present for Arg [2.2% mean squared error (MSE)] and Lys (3.2% MSE). The overpredictions may suggest that the mean recovery of AA from acid hydrolysis across laboratories was less than estimates encompassed in the recovery factors. To test this hypothesis, several regression approaches were undertaken to identify potential causes of the bias. These included regressions of (1) residual errors for predicted EAA flows on each of the 3 protein-driven EA flows, (2) observed EAA flows on each protein-driven EAA flow, including an intercept, (3) observed EAA flows on the protein-driven EAA flows, excluding an intercept term, and (4) observed EAA flows on RUP and MiCP. However, these equations were deemed unsatisfactory for bias adjustment, as they generated biologically unfeasible predictions for some entities. Future work should focus on identifying the cause of the observed prediction bias.     

Diets deficient in rumen undegraded protein did not depress milk production

The objective of this study was to evaluate the National Research Council's recommendations for feeding levels of rumen undegraded protein (RUP) for cows fed a one-group total mixed ration. Sixty Holstein cows were paired by parity (1 to 6) and DIM (23 to 315) and were randomly assigned to one of two treatment sequences. Diets contained alfalfa silage (30% diet DM) and corn silage (26% diet DM), and were isonitrogenous (16% CP) and isocaloric (1.71 Mcal/kg). Soybean meal, protected soybean meal (Soy Best), and urea were used to make ration protein fractions that were predicted to be 35 or 29% RUP. The 35% RUP diet was formulated to provide 98 and 105% of the average requirement for RUP and rumen degraded protein (RDP), respectively. The ration containing 29% RUP provided 79 and 117% of average required RUP and RDP, respectively. All cows were group-fed the high RUP diet during a 2-wk pretreatment period, and then were fed one ration for 4 wk followed by the other for 4 wk according to their assigned treatment sequence. Data were collected in the last wk of each period. Mean milk production, milk fat, and milk protein were 32.6 kg/d, 4.35%, and 3.36%, respectively, with no treatment differences. Treatment response was not affected by degree of predicted RUP deficiency. National Research Council requirements for RUP may be too high for cows fed diets similar in energy to a one-group total mixed ration. Alternatively, estimates of RUP content of feedstuffs may be low.     

Effect of nitrate supplementation,dietary protein supply,and genetic yield index on performance,methane emission,and nitrogen efficiency in dairy cows

The objective was to investigate the effect of nonprotein nitrogen source, dietary protein supply, and genetic yield index on methane emission, N metabolism, and ruminal fermentation in dairy cows. Forty-eight Danish Holstein dairy cows (24 primiparous cows and 24 multiparous cows) were used in a 6 × 4 incomplete Latin square design with 4 periods of 21-d duration. Cows were fed ad libitum with the following 6 experimental diets: diets with low, medium, or high rumen degradable protein (RDP):rumen undegradable protein (RUP) ratio (manipulated by changing the proportion of corn meal, corn gluten meal, and corn gluten feed) combined with either urea or nitrate (10 g NO₃⁻/kg of dry matter) as nonprotein nitrogen source. Samples of ruminal fluid and feces were collected from multiparous cows, and total-tract nutrient digestibility was estimated using TiO₂ as flow marker. Milk samples were collected from all 48 cows. Gas emission (CH₄, CO₂, and H₂) was measured by 4 GreenFeed units. We observed no significant interaction between dietary RDP:RUP ratio and nitrate supplementation, and between nitrate supplementation and genetic yield index on CH₄ emission (production, yield, intensity). As dietary RDP:RUP ratio increased, intake of crude protein, RDP, and neutral detergent fiber and total-tract digestibility of crude protein linearly increased, and RUP intake linearly decreased. Yield of milk, energy-corrected milk, and milk protein and lactose linearly decreased, whereas milk fat and milk urea nitrogen concentrations linearly increased as dietary RDP:RUP ratio increased. The increase in dietary RDP:RUP ratio resulted in a linear increase in the excretion of total purine derivatives and N in urine, but a linear decrease in N efficiency (milk N in % of N intake). Nitrate supplementation reduced dry matter intake (DMI) and increased total-tract organic matter digestibility compared with urea supplementation. Nitrate supplementation resulted in a greater reduction in DMI and daily CH₄ production and a greater increase in daily H₂ production in multiparous cows compared with primiparous cows. Nitrate supplementation also showed a greater reduction in milk protein and lactose yield in multiparous cows than in primiparous cows. Milk protein and lactose concentrations were lower for cows receiving nitrate diets compared with cows receiving urea diets. Nitrate supplementation reduced urinary purine derivatives excretion from the rumen, whereas N efficiency tended to increase. Nitrate supplementation reduced proportion of acetate and propionate in ruminal volatile fatty acids. In conclusion, no interaction was observed between dietary RDP:RUP ratio and nitrate supplementation, and no interaction between nitrate supplementation and genetic yield index on CH₄ emission (production, yield, intensity) was noted. Nitrate supplementation resulted in a greater reduction in DMI and CH₄ production, and a greater increase in H₂ production in multiparous cows than in primiparous cows. As the dietary RDP:RUP ratio increased, CH₄ emission was unaffected and RDP intake increased, but RUP intake and milk yield decreased. Genetic yield index did not affect CH₄ production, yield, or intensity.     

Stage of lactation and corresponding diets affect in situ protein degradation by dairy cows

The influence of stage of lactation and corresponding diets on rates of protein degradation (k_d) is largely unstudied. Study objectives were to measure and compare in situ ruminal k_d of crude protein (CP) and estimate rumen CP escape (rumen-undegradable protein; RUP) of selected feeds by cows at 3 stages of lactation fed corresponding diets, and to determine the incubation times needed in an enzymatic in vitro procedure, using 0.2 units of Streptomyces griseus protease per percent of true CP, that predicted in situ RUP. Residue CP was measured after in situ fermentation for 4, 8, 12, 24, 36, 48, and 72 h of 5 protein sources and 3 total mixed rations, which were fed to the in situ cows. Two nonlactating (dry) cows and 2 cows each at 190 (mid) and 90 (peak) days of lactation were used. Each pair of cows was offered free-choice diets that differed in composition to meet their corresponding nutrient requirements. Diets had decreasing proportions of forages and contained (dry matter basis) 11.9, 15.1 and 16.4% CP and 54.3, 40.3 and 35.3% neutral detergent fiber, for dry, mid, and peak TMR (TMR1, TMR2, and TMR3), respectively. Intakes were 10.3, 21.4, and 23.8 kg of dry matter/d, respectively. Kinetic CP fractions (extractable, potentially degradable, undegradable, or slowly degradable) were unaffected by treatment. Lag time and k_d varied among feeds. The k_d was faster for all feeds (0.136/h) when incubated in dry-TMR1 cows compared with mid-TMR2 (0.097/h) or peak-TMR3 (0.098/h) cows, and no differences in lag time were detected. Calculated RUP, using estimated passage rates for each cow based on intake, differed between dry-TMR1 (0.382) and mid-TMR2 (0.559) or peak-TMR3 (0.626) cows, with a tendency for mid-TMR2 to be different from peak-TMR3. Using the average k_d and lag time obtained from dry-TMR1 to calculate RUP for mid-TMR2 and peak-TMR3 cows using their passage rates reduced RUP values by 6.3 and 9.5 percentage units, respectively. Except for that of herring meal, in vitro residue CP at 6, 12, and 48 h of enzymatic hydrolysis was correlated (r = 0.90) with in situ RUP of peak-TMR3, mid-TMR2, and dry-TMR1, respectively. Although confounded within treatments, stage of lactation, diet, and intake appeared to affect CP degradation parameters and RUP. Using k_d from nonlactating cows, or the RUP calculated from them, may bias diet evaluation or ration formulation for lactating cows. In addition, enzymatic in vitro predictions of RUP should be measured using incubation times that are appropriate for lactating cows.     

Lowering rumen-degradable protein maintained energy-corrected milk yield and improved nitrogen-use efficiency in multiparous lactating dairy cows exposed to heat stress

The objective of this study was to examine the effect of reducing rumen-degradable protein (RDP) and rumen-undegradable protein (RUP) proportions on feed intake, milk production, and N-use efficiency in primiparous and multiparous cows exposed to warm climates. Eighteen primiparous and 30 multiparous mid-lactation Holstein cows were used in a completely randomized design with a 2 × 2 factorial arrangement of treatments. Cows were randomly assigned to 1 of 4 dietary treatments formulated to contain 2 proportions of RDP (10 and 8%) and 2 proportions RUP (8 and 6%) of dry matter (DM) indicated as follows: (1) 10% RDP, 8% RUP; (2) 8% RDP, 8% RUP; (3) 10% RDP, 6% RUP; and (4) 8% RDP, 6% RUP. Protein sources were manipulated to obtain desired RDP and RUP proportions. Diets were isoenergetic and contained 50% forage and 50% concentrate (DM basis). Cows were individually fed the 10% RDP, 8% RUP diet 3 wk before treatment allocation. Cows were exposed to the prevailing Tennessee July and August temperature and humidity in a freestall barn with no supplemental cooling. Main effects and their interaction were tested using the Mixed procedure of SAS (least squares means ± standard error of the mean; SAS Institute Inc., Cary, NC). Observed values of nutrient intake and milk production were used to obtain NRC (2001) model predictions. Cows showed signs of heat stress throughout the study. Reducing from 10 to 8% RDP decreased dry matter intake (DMI; 0.9 kg/d) at 8% RUP, but increased DMI (2.6 kg/d) at 6% RUP in primiparous cows. Reducing from 10 to 8% RDP decreased milk yield (10%) at 8% RUP, but increased yield (14%) at 6% RUP. Treatments did not affect yield of energy-corrected milk. For multiparous cows, treatments did not affect DMI. Reducing from 10 to 8% RDP decreased yield of energy-corrected milk (3.4%) at 8% RUP, but increased yield (8.8%) at 6% RUP. Reducing from 10 to 8% RDP and 8 to 6% RUP both increased N-use efficiency for primiparous and multiparous cows. The NRC model underestimated metabolizable protein and RUP supply, and overestimated RUP requirements, resulting in predictive losses of milk yield 1.4 to 5.8 times greater than observed values. In summary, the reduction of RDP and RUP proportions did not affect DMI, whereas the RUP reduction at 10% RDP had a small negative effect on energy-corrected milk yield. However, reduction of RDP and RUP consistently improved N-use efficiency of heat-stressed multiparous cows. The reduction of RDP and RUP proportions reduced DMI and milk yield but did not affect energy-corrected milk yield in primiparous cows, indicating a limited supply of nutrients.     

Lactation Performance of Mid-Lactation Dairy Cows Fed Ruminally Degradable Protein at Concentrations Lower Than National Research Council Recommendations

The aim of this study was to test whether feeding of diets containing lower proportions of ruminally degradable protein (RDP) but with a constant proportion of ruminally undegradable protein (RUP) alters feed intake, milk production and yield, and the apparent efficiency of N utilization by mid-lactation dairy cows. During the covariate period (d 1 to 28), 40 mid-lactation cows (36 Holstein and 4 Jersey × Holstein cross-breds) were fed a common diet formulated to contain 11.3% of diet dry matter (DM) as RDP. During the treatment period (d 29 to 47), cows were randomly assigned to 1 of 4 diets formulated to contain 11.3, 10.1, 8.8, or 7.6% RDP, whereas ruminally undegradable protein remained constant at 7.1% of DM. All diets contained 47.5% forage and 52.5% concentrate on a DM basis. Dry matter intake was significantly reduced for the 7.6% RDP diet. The lowest RDP content was associated with a trend for reduced milk yield. Dietary RDP had no effect on body weight or milk fat, protein, and lactose contents. Milk protein yield was not affected by RDP level; however, milk fat yield decreased linearly as dietary RDP was reduced. Concentrations of plasma essential amino acids were unaffected, whereas milk urea-N concentrations decreased linearly as dietary RDP content was reduced. The apparent efficiency of N utilization for milk N production increased from 27.7% on the 11.3% RDP diet to 38.6% on the 7.6% RDP diet. The dietary RDP requirement of cows in this study was apparently met between 15.9 and 14.7% dietary crude protein. Milk production was not significantly affected by the 8.8% RDP (15.9% crude protein) diet even though the NRC (2001) model predicted that RDP supply was 87% of that required, suggesting the current NRC recommendations for RDP may be overestimated for mid-lactation dairy cows in this study.     

Comparison of four markers for quantifying microbial protein flow from the rumen of lactating dairy cows

Eight ruminally cannulated lactating cows from a study on the effects of dietary rumen degraded protein (RDP) on production and N metabolism were used to compare ¹⁵N, total purines, amino acid (AA) profiles, and urinary excretion of purine derivatives (PD) as microbial markers for quantifying the flow of microbial protein at the omasal canal. Dietary RDP was gradually decreased by replacing solvent soybean meal and urea with lignosulfonate-treated soybean meal. The purine metabolites xanthine and hypoxanthine were present in digesta and microbial samples and were assumed to be of microbial origin. The sum of the purines and their metabolites (adenine, guanine, xanthine, and hypoxanthine) were defined as total purines (TP) and used as a microbial marker. Decreasing dietary RDP from 13.2 to 10.6% of dry matter (DM) reduced microbial nonammonia N (NAN) flows estimated using TP (from 415 to 369 g/d), ¹⁵N (from 470 to 384 g/d), AA profiles (from 392 to 311 g/d), and PD (from 436 to 271 g/d). Averaged across diets, microbial NAN flows were highest when estimated using TP and ¹⁵N (398 and 429 g/d), lowest when using PD (305 g/d), and intermediate when using AA profiles (360 g/d) as microbial markers. Correlation coefficients between ¹⁵N and TP for fluid-associated bacteria, particle-associated bacteria, and total microbial NAN flows were 0.38, 0.85, and 0.69, respectively. When TP was used as the microbial marker, ruminal escape of dietary NAN was not affected by replacing solvent soybean meal with lignosulfonate-treated soybean meal in the diets. The direction and extent of response of dietary and microbial NAN flow to dietary treatments were similar when estimated using ¹⁵N, AA profiles, and PD, and were in agreement with previously published data and National Research Council predictions. Microbial and dietary NAN flows from the rumen estimated using ¹⁵N appeared to be more accurate and precise than the other markers. Caution is required when interpreting results obtained using TP as the microbial marker.     

Ratio of dietary rumen degradable protein to rumen undegradable protein affects nitrogen partitioning but does not affect the bovine milk proteome produced by mid-lactation Holstein dairy cows

Little is known about the bovine milk proteome or whether it can be affected by diet. The objective of this study was to determine if the dietary rumen degradable protein (RDP):rumen undegradable protein (RUP) ratio could alter the bovine milk proteome. Six Holstein cows (parity: 2.5 ± 0.8) in mid lactation were blocked by days in milk (80 ± 43 d in milk) and milk yield (57.5 ± 6.0 kg) and randomly assigned to treatment groups. The experiment was conducted as a double-crossover design consisting of three 21-d periods. Within each period, treatment groups received diets with either (1) a high RDP:RUP ratio (RDP treatment: 62.4:37.6% of crude protein) or (2) a low RDP:RUP ratio (RUP treatment: 51.3:48.7% of crude protein). Both diets were isonitrogenous and isoenergetic (crude protein: 18.5%, net energy for lactation: 1.8 Mcal/kg of dry matter). To confirm N and energy status of cows, dry matter intake was determined daily, rumen fluid samples were collected for volatile fatty acid analysis, blood samples were collected for plasma glucose, β-hydroxybutyrate, urea nitrogen, and fatty acid analysis, and total 24-h urine and fecal samples were collected for N analysis. Milk samples were collected to determine the general milk composition and the protein profile. Milk samples collected for high-abundance protein analysis were subjected to HPLC analysis to determine the content of α-casein, β-casein, and κ-casein, as well as α-lactalbumin and β-lactoglobulin. Samples collected for low-abundance protein analysis were fractionated, enriched using ProteoMiner treatment, and separated using sodium dodecyl sulfate-PAGE. After excision and digestion, the peptides were analyzed using liquid chromatography (LC) tandem mass spectrometry (MS/MS). The LC-MS/MS data were analyzed using PROC GLIMMIX of SAS (version 9.4, SAS Institute Inc., Cary, NC) and adjusted using the MULTTEST procedure. All other parameters were analyzed using PROC MIXED of SAS. No treatment differences were observed in dry matter intake, milk yield, general milk composition, plasma parameters, or rumen volatile fatty acid concentrations, indicating no shift in total energy or protein available. Milk urea N and plasma urea N concentrations were higher in the RDP group, indicating some shift in N partitioning due to diet. A total of 595 milk proteins were identified, with 83% of these proteins known to be involved in cellular processes. Although none of the low-abundance proteins identified by LC-MS/MS were affected by diet, feeding a diet high in RUP decreased β-casein, κ-casein, and total milk casein concentration. Further investigations of the interactions between diet and the milk protein profile are needed to manipulate the milk proteome using diet.     

Quantifying ruminal nitrogen metabolism using the omasal sampling technique in cattle—A meta-analysis

Mixed model analysis of data from 32 studies (122 diets) was used to evaluate the precision and accuracy of the omasal sampling technique for quantifying ruminal-N metabolism and to assess the relationships between nonammonia-N flow at the omasal canal and milk protein yield. Data were derived from experiments in cattle fed North American diets (n = 36) based on alfalfa silage, corn silage, and corn grain and Northern European diets (n = 86) composed of grass silage and barley-based concentrates. In all studies, digesta flow was quantified using a triple-marker approach. Linear regressions were used to predict microbial-N flow to the omasum from intake of dry matter (DM), organic matter (OM), or total digestible nutrients. Efficiency of microbial-N synthesis increased with DM intake and there were trends for increased efficiency with elevated dietary concentrations of crude protein (CP) and rumen-degraded protein (RDP) but these effects were small. Regression of omasal rumen-undegraded protein (RUP) flow on CP intake indicated that an average 32% of dietary CP escaped and 68% was degraded in the rumen. The slope from regression of observed omasal flows of RUP on flows predicted by the National Research Council (2001) model indicated that NRC predicted greater RUP supply. Measured microbial-N flow was, on average, 26% greater than that predicted by the NRC model. Zero ruminal N-balance (omasal CP flow = CP intake) was obtained at dietary CP and RDP concentrations of 147 and 106 g/kg of DM, corresponding to ruminal ammonia-N and milk urea N concentrations of 7.1 and 8.3 mg/100 mL, respectively. Milk protein yield was positively related to the efficiency of microbial-N synthesis and measured RUP concentration. Improved efficiency of microbial-N synthesis and reduced ruminal CP degradability were positively associated with efficiency of capture of dietary N as milk N. In conclusion, the results of this study indicate that the omasal sampling technique yields valuable estimates of RDP, RUP, and ruminal microbial protein supply in cattle.     

Evaluation of the passage rate equations in the 2001 Dairy NRC model

Dairy ration formulation to meet protein and amino acid requirements with the National Research Council Nutrient Requirements of Dairy Cattle (NRC, 2001) model depends on accuracy of predicting feed passage rates out of the rumen. The NRC (2001) passage rate (Kp) equations were evaluated for validity and sensitivity to input variables in predicting supplies of rumen degraded protein, rumen undegraded protein, and metabolizable protein. The database used in the development of the 3 Kp equations (for dry forage, wet forage, and concentrate) was used to independently derive the 3 equations using a meta-analysis technique. To extract quantitative relationships between statistically significant input variables and rate of passage, a random coefficients model that used each study effect as a random variable was used. The database was comprised of studies that only used rare earth markers. Outliers were identified by acceptance criteria defined a priori or the difference in fit statistic (DFFITS) value; 319, 63, and 139 treatment means were used to develop the Kp equations for dry forage, wet forage, and concentrate, respectively. We found that the sign of the regression coefficient for concentrate content in diet dry matter in the equation for Kp dry forage was inverted; it should be positive. A sensitivity analysis was conducted with a spreadsheet version of the NRC (2001) model developed for this study, using the Monte Carlo technique. The sensitivity analysis indicated that all Kp predictions were the most sensitive to variation in DM intake, and thus accurate measurement of DM intake is the most important factor in predicting Kp. Predictions for protein supply (rumen degraded protein, rumen undegraded protein, and metabolizable protein) were sensitive to variability in amount of feed crude protein (CP, %DM), digestion rate (Kd) of the B fraction of feed CP (%/h), and the Kp for concentrate (%/h), due to the high proportion of dietary CP in lactating dairy rations coming from concentrates. The sensitivity analysis indicated that accurate determinations of DMI, the Kd of the B fraction of feed CP, and feed CP are the most important variables needed to predict MP supply in lactating dairy cows with the NRC (2001) model. We conclude that the empirical Kp equations in the model are suitable for predicting passage rate in lactating dairy cows. More accurate predictions of Kp will require the development of a more mechanistic model that accounts for more of the biologically important variables (e.g., physical property of particles, liquid flow, and timely variation of intake) affecting passage rate.     

Effect of feeding protein supplements of differing degradability on omasal flow of microbial and undegraded protein

Ten ruminally cannulated lactating Holstein cows that were part of a larger trial studying the effects of feeding different proteins on milk production were used in a replicated 5 x 5 Latin square to quantify flows of microbial and rumen-undegradable protein (RUP) in omasal digesta. Cows were fed total mixed rations containing (dry matter basis) 44% corn silage, 22% alfalfa silage, 2% urea, and 31% concentrate. The basal diet contained 31% high-moisture corn; equal N from one of four protein supplements was added to the other diets at the expense of corn: 9% solvent soybean meal (SSBM), 10% expeller soybean meal (ESBM), 5.5% blood meal (BM), and 7% corn gluten meal (CGM). Omasal sampling was used to quantify total AA N (TAAN) and nonammonia N (NAN) flows from the rumen. Estimates of RUP were made from differences between total and microbial N flows, including a correction for RUP in the basal diet. Modifying a spectrophotometric assay improved total purine recovery from isolated bacteria and omasal samples and gave estimates of microbial TAAN and NAN flows that were similar to a standard HPLC method. Linear programming, based on AA patterns of the diet and isolated omasal bacteria and ruminal protozoa, appeared to overestimate microbial TAAN and NAN flows compared to the purine assays. Yields of microbial TAAN and NAN determined using any method was not affected by diet and averaged 32 to 35 g NAN per kilogram of organic matter truly digested in the rumen. On average, National Research Council (NRC) equations underpredicted microbial N flows by 152 g/d (vs. HPLC), 168 g/d (vs. spectrophotometry), and 244 g/d (vs. linear programming). Estimates of RUP (means from the HPLC and spectrophotometric methods) were: SSBM, 27%, ESBM, 45%, BM, 60%, and CGM, 73%. Except for CGM, RUP values averaged about 20 percentage units lower than those reported by the NRC.     

Intestinal digestibility of amino acids in rumen undegradable protein estimated using a precision-fed cecectomized rooster bioassay: I. Soybean meal and SoyPlus

The objectives of this experiment were to measure intestinal digestibility of AA in rumen undegradable protein (RUP-AA) in soybean meal (SBM) and expeller SBM (SoyPlus, West Central, Ralston, IA; SP) and to determine if these feeds contain a constant protein fraction that is undegradable in the rumen and indigestible in the small intestine, as assumed in the French Institut National de la Recherche Agronomique (Paris, France) and Scandinavian AAT-PBV (AAT = AA absorbed from small intestine; PBV = protein balance in the rumen) models. Three samples of SBM and 3 samples of SP were obtained from the Feed Analysis Consortium Inc. (Savoy, IL). To obtain the RUP fraction, samples were ruminally incubated in situ for 16 h in 4 lactating cows, and the collected rumen undegraded residues (RUR) were pooled by sample. Subsamples of the intact feeds and RUR were crop intubated to 4 cecectomized roosters, and total excreta were collected for 48 h. Intact feeds, RUR, and excreta were analyzed for AA. Basal endogenous AA loss estimates were obtained from fasted birds and were used to calculate standardized digestibility of AA in the intact feeds and RUP-AA. Indigestibility coefficients of the intact feeds were calculated as (100 – % standardized AA digestibility), and indigestibility of the RUR was calculated as (100 – % ruminal degradation of AA) × [(100 – % standardized RUP-AA digestibility)]/100. Results indicated that standardized digestibility of feed-AA was similar to standardized digestibility of RUP-AA for SBM and SP samples and that standardized digestibility of individual AA differed within samples. Standardized feed-AA and RUP-AA digestibility values were lowest for Lys and Cys and highest for Trp and Met. Results also indicated that SBM and SP did not contain a constant protein fraction that was both undegradable in the rumen and indigestible in the small intestine. Indigestibility values of RUR were lower than in intact feeds, suggesting that SBM and SP contain a protein fraction that is indigestible in the intestine but partly degradable in the rumen, digestible in the intestine after ruminal incubation, or both.     

Ruminal degradation and intestinal digestibility of protein and amino acids in high-protein feedstuffs commonly used in dairy diets

A study was conducted to determine the rumen degradation and intestinal digestibility of crude protein (CP) and AA, and AA composition of the rumen-undegradable protein (RUP) from 3 sources of blood meal (BM1, BM2, and BM3), canola meal (CM), low-fat distillers dried grains with solubles (LFDG), soybean meal (SBM), and expeller soybean meal (ESBM). Two Holstein cows fitted with ruminal and proximal duodenal cannulas were used for in situ incubation of 16 h and for the mobile bag technique. To correct for bacterial contamination of the RUP, 2 methods were used: purines and DNA as bacterial markers. Ruminal degradations of CP were 85.3, 29.8, 40.7, 75.7, 76.9, 68.8, and 37.0 ± 3.93% for BM1, BM2, BM3, CM, LFDG, SBM, and ESBM, respectively. Ruminal degradation of both total essential AA and nonessential AA followed a similar pattern to that of CP across feedstuffs. Based on the ratio of AA concentration in the RUP to AA concentration in the original feedstuff, ruminal incubation decreased (ratio <1) the concentrations of His, Lys, and Trp, and increased (ratio >1) the concentrations of Ile and Met across feedstuffs. Compared with purines, the use of DNA as bacterial marker resulted in a higher estimate of bacterial CP contamination for CM and lower estimates for LFDG and ESBM. Intestinal digestibility of RUP could not be estimated for BM1, BM3, and SBM due to insufficient recovery of residue. For the remaining feedstuffs, intestinal digestibility of RUP was highest for ESBM, followed by BM2 and LFDG, and lowest for CM: 98.8, 87.9, 89.7, and 72.4 ± 1.40%, respectively. Intestinal absorbable dietary protein was higher for BM2 compared with CM and LFDG, at 61.7, 17.9, and 20.7 ± 2.73% CP, respectively. As prices fluctuate, intestinal absorbable protein or AA may be used as a tool to aid in the selection among feedstuffs with different protein quality.     

Ruminal and intestinal degradability of distillers grains plus solubles varies by source

Currently in the dairy industry, there is a concern about the variability in the nutrient content among sources of distillers grains plus solubles (DG), but little research has evaluated the variability in metabolizable AA among sources. The ruminal degradability of crude protein (CP) in soybean meal (SBM), dried DG from 5 sources (DG1, DG2, DG3, DG4, and DG5), and 1 source of wet DG (WDG) were determined using 2 lactating ruminally cannulated Holstein cows. Feeds were incubated in the rumen for 3, 6, 12, 18, 24, and 36 h. Intestinal CP digestibility via pepsin and pancreatin and AA profiles were measured on residue from 12-h ruminal incubation of feeds. Ruminal undegradable protein (RUP) was less for SBM (46.4%) than for DG. The WDG (53.6%) had less RUP than dried DG. The RUP concentrations of DG3 (59.1%) and DG5 (60.3%) were lower than DG1 (71.7%) and DG4 (67.5%), with DG1 having more than DG2 (63.7%) and DG4. Intestinal digestibility of RUP was greater for SBM (86.7%) than DG. The DG2 (76.8%) and DG3 (74.2%) had greater intestinal digestibility compared with DG1 (59.2%), DG4 (63.0%), and DG5 (68.1%). The intestinal digestibility in WDG (65.8%) was similar to all other DG except for DG1, which was lower. Total digestibility of CP was greater in SBM (93.9%) compared with DG. Among the DG sources, the CP in DG2 (85.3%) and DG3 (84.9%) was more digestible compared with DG1 (70.7%), DG4 (74.9%), and DG5 (80.8%) but not WDG (81.9%). Based on the milk protein score (MPS), which is an estimate of the proportion of milk protein that a protein source can sustain until the first limiting AA is depleted, Met was the first limiting AA in SBM and Lys in DG. The concentrations of essential AA in the RUP were not different among DG sources, but the greater MPS in WDG (0.306) compared with the dried DG (0.240) sources indicated that WDG may have been the more ideal RUP source; but, the MPS of the metabolizable protein indicated that the protein quality of WDG was similar to that in DG2, DG3, and DG5. In conclusion, protein degradability and digestibility differed greatly among DG sources, but these differences were actually not as prominent in the concentrations of metabolizable AA and MPS among these sources.