Effects of intramuscular injections of vitamin B12 on lactation performance of dairy cows fed dietary supplements of folic acid and rumen-protected methionine

The experiment was undertaken to determine the effects of i.m. injections of vitamin B(12) on lactational performance of primiparous dairy cows fed dietary supplements of folic acid and rumen-protected methionine from 4 to 18 wk of lactation. Fourteen primiparous Holstein cows were assigned to 7 blocks of 2 cows each, according to milk production during the third week of lactation. All cows were fed a basal diet supplemented daily with rumen-protected methionine (18 g of supplement, to bring the estimated supply of methionine to 2.2% of metabolizable protein) plus folic acid (4 mg per kg of BW). Within each block, the cows received a weekly i.m. injection (2 mL) of saline or 10 mg of vitamin B(12). Milk production was recorded daily. Milk and blood were sampled every 2 wk. Supplementary vitamin B(12) increased energy-corrected milk from 25.8 to 29.0 (SE 1.6) kg/d, as well as milk yields of solids [3.52 to 3.90 (SE 0.22) kg/d], fat [0.87 to 1.01 (SE 0.06) kg/d], and lactose [1.48 to 1.64 (SE 0.11) kg/d]. Supplementation also increased concentrations and amounts of vitamin B(12) secreted in milk but had no significant effect on dry matter intake and concentrations and amounts of folates in milk. Packed cell volume, blood hemoglobin, and serum vitamin B(12) were increased by supplementary vitamin B(12), whereas serum methylmalonic acid was decreased. Serum concentrations of sulfur amino acids were unchanged by treatment. These findings support the hypothesis that, in early lactation, supply of vitamin B(12) was not optimal and limited the lactation performance of the cows.     

Influence of methionine supply on the response of lactational performance of dairy cows to supplementary folic acid and vitamin B12

The present experiment was undertaken to determine if the effects of supplementary folic acid on lactational performance were caused by improved methylneogenesis and if the supply in vitamin B₁₂ could affect this metabolic pathway. In this eventuality, supplementary Met, a major source of preformed methyl groups, should reduce the requirements for these vitamins. Sixty multiparous Holstein cows were assigned to 10 blocks of 6 cows each according to their previous milk production. Within each block, 3 cows were fed a diet estimated to supply Met as 1.83% metabolizable protein and 3 cows were fed the same diet supplemented with 18 g of rumen-protected methionine (RPM) to supply Met as 2.23% of metabolizable protein. Within each level of Met, cows received no vitamin supplement or weekly intramuscular injections of 160 mg of folic acid alone or combined with 10 mg of vitamin B₁₂ from 3 wk before to 16 wk after calving. There was no treatment effect on dry matter intake during pre- and postcalving periods: 13.4 ± 0.4 and 21.8 ± 0.4 kg/d, respectively. Milk production was not affected by RPM supplementation. Folic acid and vitamin B₁₂ given together tended to increase milk production during the 16 wk of lactation. This effect was more pronounced during the first 4 wk of lactation: 37.5, 37.7, and 40.3 ± 0.9 kg/d for cows receiving no vitamin supplement, folic acid alone, or folic acid combined with vitamin B₁₂, respectively. Milk fat yield was not affected by treatments. Lactose, crude protein, and total solid yields were greater, in early lactation, in cows injected with folic acid and vitamin B₁₂ together but this effect diminished as lactation progressed. Intramuscular injections of folic acid alone or combined with vitamin B₁₂ tended to decrease plasma concentrations of homocysteine from 5.51 μM with no vitamin supplement to 4.54 and 4.77 ± 0.37 μM, respectively. Results of the present experiment suggest that the effects of the combined supplement of folic acid and vitamin B₁₂ on lactational performance of dairy cows were not due to an improvement in methyl groups supply, because RPM supplement, a source of preformed methyl groups, did not alter the cow responsiveness to vitamin supplements.     

Effect of different forms of methionine on lactational performance of dairy cows

Methionine is one of the first limiting AA in dairy cows. The use of rumen-protected Met to correct deficient diets is limited by the lack of a product that could be incorporated into a pelleted concentrate. The main objective of this trial was to test, at practical doses (approximately 10 g of absorbable Met), the efficacy of 2 forms of pelletable Met hydroxy analogs, d,l-2-hydroxy-4-(methylthio)-butanoic acid (HMB) and the isopropyl ester of HMB (HMBi), to provide Met to cows, especially for milk protein synthesis, compared with a negative control and to Smartamine M (SmM). These treatments were tested according to a 4 × 4 Latin square in 16 Holstein cows. Plasma Met concentrations were increased by 110 and 65% that of the control value after HMBi and SmM treatments, respectively. Milk protein yield increased by 32 and 41 g/d for HMBi and SmM, respectively. d,l-2-Hydroxy-4-(methylthio)-butanoic acid supplementation did not improve Met availability to the cows for milk protein synthesis. The HMBi treatment induced an increase in 15:0 in milk at the expense of a general reduction in even-numbered short-and medium-chain fatty acids. Moreover, HMBi and SmM supplements led to an increase in the saturation level of C₁₈ fatty acids consistent with the improvement of Met supply. It was concluded that HMBi is a new “rumen-protected” form of Met that can be supplied to cows integrated into pellets.     

Effects of supplementary folic acid and vitamin B12 on hepatic metabolism of dairy cows according to methionine supply

The present experiment was undertaken to study the interactions between dietary supplements of rumen-protected methionine (RPM) and intramuscular injections of folic acid and vitamin B₁₂, given from 3 wk before calving to 16 wk of lactation, on hepatic metabolism of lactating dairy cows. Sixty multiparous Holstein cows were assigned to 10 blocks of 6 cows each according to their previous milk production. Within each block, 3 cows were fed a diet calculated to supply Met as 1.83% of metabolizable protein, whereas the 3 other cows were fed the same diet supplemented with 18 g of RPM calculated to provide Met as 2.23% of metabolizable protein. Within each level of Met, the cows received no vitamin supplement or weekly intramuscular injections of 160 mg of folic acid alone or combined with 10 mg of vitamin B₁₂. Liver biopsies were taken at 2, 4, 8, and 16 wk of lactation. Liver concentrations of folates and vitamin B₁₂ were increased by their respective supplements but this response to vitamin supplements was altered by methionine supply. Concentrations of total lipids and triglycerides increased in livers of cows fed RPM, whereas concentrations of cholesterol ester, cholesterol, diglycerides, phosphatidylethanolamine, and phosphatidylcholine were not affected. Folic acid, alone or combined with vitamin B₁₂, tended to increase the ratio of phosphatidylcholine to phosphatidylethanolamine. Gene expression of 5,10-methylene-tetrahydrofolate reductase, microsomal transfer protein, and phosphatidylethanolamine methyltransferase were higher in liver of cows fed RPM supplements. The relative mRNA abundance of 5,10-methylene-tetrahydrofolate reductase and methylmalonyl-CoA mutase were increased by the combined injections of folic acid and vitamin B₁₂, whereas those of methionine synthase and methionine synthase reductase were not affected by treatments. These results suggest that increasing supply of methyl groups, as preformed labile methyl groups or through methylneogenesis, affected the methylation cycle but had a limited effect on dairy cow performance. The observed effects of the combined supplement of folic acid and vitamin B₁₂ on lactational performance of dairy cows probably result from an improvement of energy metabolism during early lactation.     

Effects of supplements of folic acid, vitamin B12, and rumen-protected methionine on whole body metabolism of methionine and glucose in lactating dairy cows

The present experiment was undertaken to determine the effects of dietary supplements of rumen-protected methionine and intramuscular injections of folic acid and vitamin B₁₂, given 3 wk before to 16 wk after calving, on glucose and methionine metabolism of lactating dairy cows. Twenty-four multiparous Holstein cows were assigned to 6 blocks of 4 cows each according to their previous milk production. Within each block, 2 cows were fed a diet estimated to supply methionine as 1.83% metabolizable protein, equivalent to 76% of methionine requirement, whereas the 2 other cows were fed the same diet supplemented daily with 18 g of rumen-protected methionine. Within each diet, the cows were administrated either no vitamin supplement or weekly intramuscular injections of 160 mg of folic acid plus 10 mg of vitamin B_12. To investigate metabolic changes at 12 wk of lactation, glucose and methionine kinetics were measured by isotope dilution using infusions of 3[U-¹³C]glucose, [¹³C]NaHCO₃ and 3[1-¹³C,²H₃] methionine. Milk and plasma concentrations of folic acid and vitamin B₁₂ increased with vitamin injections. Supplementary B-vitamins increased milk production from 34.7 to 38.9 ± 1.0 kg/d and increased milk lactose, protein, and total solids yields. Whole-body glucose flux tended to increase with vitamin supplementation with a similar quantitative magnitude as the milk lactose yield increase. Vitamin supplementation increased methionine utilization for protein synthesis through increased protein turnover when methionine was deficient and through decreased methionine oxidation when rumen-protected methionine was fed. Vitamin supplementation decreased plasma concentrations of homocysteine independently of rumen-protected methionine feeding, although no effect of vitamin supplementation was measured on methionine remethylation, but this could be due to the limitation of the technique used. Therefore, the effects of these B-vitamins on lactation performance were not mainly explained by methionine economy because of a more efficient methylneogenesis but were rather related to increased glucose availability and changes in methionine metabolism.     

Effects of dietary supplements of folic acid and vitamin B12 on metabolism of dairy cows in early lactation

The present experiment was undertaken to determine the effects of dietary supplements of folic acid and vitamin B₁₂ given from 3 wk before to 8 wk after calving on lactational performance and metabolism of 24 multiparous Holstein cows assigned to 6 blocks of 4 cows each according to their previous milk production. Supplementary folic acid at 0 or 2.6 g/d and vitamin B₁₂ at 0 or 0.5 g/d were used in a 2 × 2 factorial arrangement. Supplementary folic acid increased milk production from 38.0 ± 0.9 to 41.4 ± 1.0 kg/d and milk crude protein yield from 1.17 ± 0.02 to 1.25 ± 0.03 kg/d. It also increased plasma Gly, Ser, Thr, and total sulfur AA, decreased Asp, and tended to increase plasma Met. Supplementary B₁₂ decreased milk urea N, plasma Ile, and Leu and tended to decrease Val but increased homocysteine, Cys, and total sulfur AA. Liver concentration of phospholipids was higher in cows fed supplementary B₁₂. Plasma and liver concentrations of folates and B₁₂ were increased by their respective supplements, but the increase in plasma folates and plasma and liver B₁₂ was smaller for cows fed the 2 vitamins together. In cows fed folic acid supplements, supplementary B₁₂ increased plasma glucose and alanine, tended to decrease plasma biotin, and decreased K_m of the methylmalonyl-coenzyme A mutase in hepatic tissues following addition of deoxyadenosylcobalamin, whereas it had no effect when cows were not fed folic acid supplements. There was no treatment effect on plasma nonesterified fatty acids as well as specific activity and gene expression of Met synthase and methylmalonyl-coenzyme A mutase in the liver. Ingestion of folic acid supplements by cows fed no supplementary B₁₂ increased total lipid and triacylglycerols in liver, whereas these supplements had no effect in cows supplemented with B₁₂. The increases in milk and milk protein yields due to folic acid supplements did not seem to be dependent on the vitamin B₁₂ supply. However, when vitamin B₁₂ was given in combination with folic acid, utilization of the 2 vitamins seems to be increased, probably more so in extrahepatic tissues. Metabolic efficiency seems also to be improved as suggested by similar lactational performance and dry matter intake for cows fed supplementary folic acid but increased plasma glucose and decreased hepatic lipids in cows fed folic acid and vitamin B₁₂ together.     

Effects of providing two forms of supplemental methionine to periparturient Holstein dairy cows on feed intake and lactational performance

Eighteen primiparous and 42 multiparous Holstein cows were blocked according to parity and expected calving date and assigned randomly to 1 of 3 dietary treatments: 1) a basal diet (negative control), 2) the basal diet plus 2-hydroxy-4-methylthio butanoic acid isopropyl ester (MetaSmart, Adisseo Inc., Antony, France), or 3) the basal diet plus rumen-protected Met (Smartamine M, Adisseo Inc., Alpharetta, GA). Treatments were initiated 21 d before expected calving and continued through 140 d postpartum. Diets were similar in ingredient and chemical composition, except for the content of Met in metabolizable protein. MetaSmart [0.35% prepartum and 0.54% postpartum in diet dry matter (DM)] and Smartamine M (0.06% prepartum and 0.10% postpartum in diet DM) were added to the basal diet in amounts needed to achieve a 3.0:1 ratio of Lys to Met in metabolizable protein. Prepartum DM intake (DMI; 13.5 kg/d), body weight (687 kg), body condition score (3.81), postpartum milk yield (42.0 kg/d), milk fat yield (1,549 g/d), milk fat content (3.66%), milk true protein yield (1,192 g/d), and milk urea N content (12.9 mg/dL) were not different among treatments. Postpartum DMI and body condition score were greater and the ratios of milk:DMI and milk N:feed N were less for cows fed the MetaSmart diet than for cows fed the control and Smartamine M diets. Milk protein content was greater for the Smartamine M (2.87%) and MetaSmart (2.81%) treatments than for the control treatment (2.72%). Concentrations of Met and Met + Cys in total plasma AA were different among treatments, with values for the Smartamine M treatment being the highest, followed by the MetaSmart and control treatments. The results indicated that both MetaSmart and Smartamine M are effective in providing metabolizable Met, but clarification of their relative contributions to metabolizable Met is still needed.     

Whole-body propionate and glucose metabolism of multiparous dairy cows receiving folic acid and vitamin B12 supplements

This study was undertaken to evaluate the effect of supplementation of folic acid and vitamin B₁₂ on glucose and propionate metabolism. Twenty-four multiparous cows were assigned according to a complete block design in a 2 × 2 factorial arrangement to one of the following treatments: (1) saline 0.9% NaCl, (2) 320 mg of folic acid, (3) 10 mg of vitamin B₁₂, or (4) 320 mg of folic acid and 10 mg of vitamin B₁₂. Intramuscular injections were given weekly from 3 wk before the expected calving date until 9 wk postpartum. At 63 d in milk, d-[6,6-²H₂]-glucose (16.5 mmol/h; jugular vein) and [1-¹³C]-sodium propionate (13.9 mmol/h; ruminal vein) were simultaneously infused for 4 h; blood samples were collected from 2 to 4 h of the infusion period. Liver biopsies were carried out the following day. Supplements of folic acid and vitamin B₁₂ respectively increased folate and vitamin B₁₂ concentrations, both in milk and liver. Although dry matter intake was unaffected by treatments, milk and milk lactose yields tended to be lower by 5.0 and by 0.25 kg/d, respectively, for cows receiving the folic acid supplement. Plasma β-hydroxybutyrate concentration with the folic acid supplement followed the same tendency. Hepatic gene expression of methylmalonyl-CoA mutase and S-adenosylhomocysteine hydrolase was higher for cows receiving the combined folic acid and vitamin B₁₂ supplement compared with cows receiving only the supplement of folic acid, whereas no treatment effect was noted for cows not receiving the folic acid supplement. Whole-body glucose rate of appearance and the proportion of whole-body glucose rate of appearance secreted in milk lactose decreased by 229 g/d and 5%, respectively, for animals receiving the folic acid supplement, concomitant with the lower milk lactose synthesis in these cows, indicating that supplementary folic acid may alter energy partitioning in cows. The absence of treatment effect on plasma concentrations of methylmalonic acid as well as on the proportion of glucose synthesized from propionate, averaging 60%, supports the fact that vitamin B₁₂ supply was sufficient in control cows in the current study. Our results suggest that the folic acid supplement reduced glucose-derived lactose synthesis by redirecting glucose for other metabolic activity in the mammary gland or in other tissues.     

Improving intestinal amino acid supply of pre- and postpartum dairy cows with rumen-protected methionine and lysine

Eighty-four Holstein cows were assigned to a randomized block experiment to determine effects of supplementing pre- and postpartum diets containing highLys protein supplements with rumen-protected Met and Lys. Before parturition, cows received a basal diet with 1) no rumen-protected amino acids (AA), 2) 10.5 g/d of Met from rumen-protected Met, or 3) 10.2 g/d of Met and 16.0 g/d of Lys from rumen-protected Met plus Lys. After parturition, cows continued to receive AA treatments but switched to diets balanced for 16.0 or 18.5% crude protein (CP). Diets were corn-based; supplemental protein was provided by soybean products and blood meal. Cows received treatments through d 105 of lactation. Compared with basal and Met-supplemented diets, Met + Lys supplementation increased yield of energy-corrected milk, fat, and protein, and tended to increase production of 3.5% fat-corrected milk. Significant CP x AA interactions were observed only for milk protein and fat content. Supplementation of the 16% CP diet with Met and Met + Lys had no effect on milk true protein and fat content. However, Met and Met + Lys supplementation of the 18.5% CP diet increased milk protein content by 0.21 and 0.14 percentage units, respectively, and Met supplementation increased fat content by 0.26 percentage units. Results of this study indicate that early-lactation cows fed corn-based diets are responsive to increased intestinal supplies of Lys and Met and that the responses depend on dietary CP concentration, supply of metabolizable protein, and intestinal digestibility of the rumen-undegradable fraction of supplemental proteins.     

Milk performance and glucose metabolism in dairy cows fed rumen-protected fat during mid lactation

Feeding rumen-protected fat (RPF) can improve energy supply for dairy cows but it affects glucose metabolism. Glucose availability is a precondition for high milk production in dairy cows. Therefore, this study investigated endocrine regulation of glucose homeostasis and hepatic gene expression related to glucose production because of RPF feeding in lactating cows. Eighteen Holstein dairy cows during second lactation were fed either a diet containing RPF (mainly C16:0 and C18:1; FD; n = 9) or a control diet based on corn starch (SD; n = 9) for 4 wk starting at 98 d in milk (DIM). Feed intake and milk yield were measured daily and milk composition once a week. Blood samples were taken weekly for analyses of plasma triglyceride, nonesterified fatty acids (NEFA), β-hydroxybutyrate, bilirubin, urea, lactate, glucose, insulin, and glucagon. At 124 DIM, an intravenous glucose tolerance test (GTT; 1 g/kg of BW^0.75) was performed after a 12-h period without food. Blood samples were taken before and 7, 14, 21, and 28 min after glucose administration, and plasma concentrations of glucose, insulin, and glucagon were measured. Glucose half-life as well as areas under the concentration curve for glucose, insulin, and glucagon were calculated. After slaughter at d 28 of treatment, liver samples were taken to measure mRNA abundance of pyruvate carboxylase, cytosolic phosphoenolpyruvate carboxykinase, glucose 6-phosphatase (G6Pase), and facilitative glucose transporter 2. Dry matter intake, but not energy and protein intake, was lower in FD than in SD. Milk yield during lactation decreased more in SD than in FD, and milk protein was lower in FD than in SD. Plasma concentrations of triglycerides and NEFA were higher in FD than in SD. Plasma insulin concentrations were lower and the glucagon:insulin ratios were higher in FD than in SD. Fasting glucose concentration before GTT was lower, and fasting glucagon concentrations tended to be higher in FD than in SD. In liver, fat content tended to be higher and G6Pase mRNA abundance was lower in FD than in SD. Lower hepatic G6Pase mRNA abundance was associated with reduced fasting plasma glucose concentrations, but the glucose-induced insulin response was not affected by RPF feeding. Hepatic G6Pase gene expression might be affected by DMI and might be involved in the regulation of glucose homeostasis in dairy cows, resulting in a lower hepatic glucose output after RPF feeding.     

Effects of intramuscular injections of folic acid,vitamin B12, or both,on lactational performance and energy status of multiparous dairy cows

The purpose of this experiment was to gain understanding on changes in energy partitioning when folic acid and vitamin B₁₂ supplements, alone or combined, were given by weekly intramuscular injections from 3 wk before the expected calving date until 7 wk postpartum. Twenty-four multiparous cows were assigned to 6 blocks of 4 cows each according to previous 305-d lactation yield to either 0 or 320 mg of folic acid and 0 or 10 mg of vitamin B₁₂ in a 2 × 2 factorial arrangement. Plasma concentration of folates was increased by folic acid supplement, and this increase was greater with the combined supplement. Vitamin B₁₂ supplement increased plasma concentration of vitamin B₁₂. Even though postpartum energy balance was similar among treatments, postpartum body condition score was higher for cows receiving folic acid supplement compared with cows that did not. Milk yield of cows receiving folic acid supplement reached a plateau earlier than for cows that did not. Fat and protein, as well as total solid concentrations and yields, were unaffected by treatments. Postpartum plasma concentrations of glucose and insulin were higher and postpartum plasma concentration of nonesterified fatty acids was lower for cows that received weekly folic acid supplement compared with cows that did not. Plasma concentration of methylmalonic acid was low and unaffected by treatments, suggesting that vitamin B₁₂ supply was not limiting, even for unsupplemented cows. Postpartum plasma concentrations of Cys, His, Phe, and Tyr were increased, whereas plasma concentration of Gly was decreased, by folic acid supplement. In the present study, supplementary folic acid altered energy partitioning in early lactation as suggested by similar milk total solid yield and postpartum energy balance, lower plasma nonesterified fatty acid concentration and body condition score losses, and higher plasma glucose and insulin concentrations for cows receiving folic acid supplement compared with cows that did not.     

Effects of supplementing rumen-protected lysine and methionine during prepartum and postpartum periods on performance of dairy cows

An experiment was conducted to examine effects of prepartum, postpartum, or continuous prepartum and postpartum supply of rumen-protected lysine (RPLys) and rumen-protected methionine (RPMet) on performance and blood metabolites of transition cows. The experiment consisted of a prepartum (3 wk), postpartum (3 wk), and carryover (10 wk) period. Eighty-eight prepartum cows (36 primiparous and 52 multiparous cows) were blocked by parity and expected calving date and assigned to 1 of 4 treatments arranged factorially. Treatments were a prepartum diet (12% crude protein on a dry matter basis) without (Pre?) or with supplemental RPLys (10 g of digestible Lys/cow per day) and RPMet (4 g of digestible Met/cow per day; Pre+) followed by postpartum diets (16% crude protein on a dry matter basis) without (Post?) or with supplemental RPLys (26 g of digestible Lys/cow per day) and RPMet (11 g of digestible Met/cow per day; Post+). Prepartum, only 2 treatments were applied, but postpartum cows received treatments of Pre?Post?, Pre?Post+, Pre+Post?, or Pre+Post+. During the prepartum period, treatment did not affect dry matter intake and body weight. During the postpartum period, milk protein content was greater (3.23 vs. 3.11%) for Post+ compared with Post? independent of prepartum treatment. However, dry matter intake, body weight, milk yield, and yields of milk components were not affected by Post+ versus Post?. No effects of prepartum treatment or interactions between pre- and postpartum treatments were observed on postpartum performance of cows. No effects of pre- and postpartum supplementation of RPLys and RPMet on performance during the carryover period were found except prepartum supplementation of RPLys and RPMet decreased somatic cell count (4.60 vs. 4.83; log₁₀ transformed) compared with Pre? in the postpartum period and this effect continued during the carryover period [i.e., 4.42 and 4.55 (log₁₀ transformed) for Pre+ and Pre?, respectively]. Prepartum supplementation of RPLys and RPMet increased or tended to increase plasma concentration of Lys, Met, and branched-chain AA compared with Pre? in prepartum cows. Cows on Post+ tended to have greater plasma Lys concentration compared with Post?, but plasma Met concentration was not affected. Health events of postpartum cows were not affected by treatments. In conclusion, we did not observe positive effects of supplementing with RPLys and RPMet on performance of prepartum and postpartum cows. However, prepartum supply of RPLys and RPMet may have potential to improve udder health and immune status of fresh cows.     

Evaluation of rumen-protected methionine sources and period length on performance of lactating dairy cows within Latin squares

Multiparous Holstein cows (n = 16) were used in a replicated (n = 4) Latin square design with 2-wk periods to determine whether length of the experimental period chosen within Latin square designs would influence experimental outcomes for performance-related variables. Cows were fed a basal TMR formulated to supply Lys in excess of the predicted requirements and either no rumen-protected Met (RP-Met; control), or 6 (M6) or 12 (M12) g/d of RP-Met (Mepron), or 12 (S12) g/d of RP-Met (Smartamine M). Performance outcomes were evaluated separately using data collected at the end of each week of each period, and as overall means for each period. Milk yield was not affected by treatment in any period. Supplementation of RP-Met from all sources tended to increase milk fat percentage when evaluated using wk-1 data, but responses evaluated using wk-2 data only were not significant. Supplementation of M12 and S12 tended to increase milk fat percentage over the entire experimental period. Supplementation of M6 and M12 tended to increase milk true protein percentage when evaluated using wk-1 data only; however, responses to treatments during wk 2 were not significant. All supplemental RP-Met treatments either tended to increase (M6 and S12) or increased (M12) milk true protein content over the entire experimental period relative to the control treatment. Yields of fat and true protein were not affected by treatment. Carryover effects of treatment on production variables were largely not significant. Predictions of nutrient supply with 3 models used in dairy ration formulation and evaluation (CPM-Dairy, AminoCow, and 2001 NRC Dairy) indicated that energy, protein, and Met all were supplied in excess of requirements for all treatments. Overall, results suggest that interpretation of performance outcomes can vary depending on selection of the experimental period in Latin squares, and responses to increased Met supply based on ratio-based formulation may not be independent of the grams of Met supplied.     

Effects of rumen-protected methionine and other essential amino acid supplementation on milk and milk component yields in lactating Holstein cows

Objectives of this study were to investigate the effects of supplementing rumen-protected methionine (RP-Met), threonine (RP-Thr), isoleucine (RP-Ile), and leucine (RP-Leu) individually or jointly to a low-protein diet, on the performance of lactating dairy cows, as well as to determine the effects of these amino acids (AA) on the mammalian target of rapamycin (mTOR) in vivo. Ten lactating Holstein cows were randomly allocated to a repeated 5 × 5 Latin square experiment with five 19-d periods. Treatments were high-protein diet (16% crude protein, positive control; HP), low-protein diet (12% crude protein, negative control; LP), LP plus RP-Met (LPM), LP plus RP-Met and RP-Thr (LPMT), and LP plus RP-Met, RP-Thr, RP-Ile, and RP-Leu (LPMTIL). The dry matter intakes (DMI) of the LP, LPM, and LPMT diets were lower than that of the HP diet, whereas the DMI of the LPMTIL diet was intermediate between the HP diet and the other LP diets. Supplementing RP-Met to the LP diet increased the yields of milk and milk protein, increased the content of milk urea N, and tended to increase milk N efficiency. Co-supplementation of RP-Thr with RP-Met resulted in no further milk production increase. Co-supplementation of all 4 rumen-protected amino acids (RP-AA) increased milk and lactose yields to the level of the HP diet and tended to increase milk protein yield compared with the LPMT diet. We found no significant differences in the contents and yields of milk components between the LPMTIL and HP diets except for a lower milk urea N content in the LPMTIL diet. Venous concentrations of the measured AA were similar across the LP and LP diets supplemented with RP-AA. Relative to levels of the HP diet, LP diets had higher venous concentrations of Met and Gly and tended to have higher Phe concentration and lower concentrations of Val and BCAA. The LPMTIL diet had higher venous concentrations of Arg, Lys, Met, Phe, and Glu, and a lower Val concentration. Phosphorylation status of the measured mTOR components in LPM and LPMT treatments were similar to those in the LP treatment but phosphorylation status of mTOR and eIF4E-binding protein 1 (4eBP1) in LPMTIL treatment were higher. The phosphorylation rates of eukaryotic elongation factor 2 (eEF2) in the 4 LP and LP plus RP-AA diets were higher than that of the HP diet. Overall, results of the present study supported the concept that under the relatively short time of this experiment, supplementing RP-AA, which are believed to stimulate the mTOR signal pathway, can lead to increased milk protein yield. This increase appears to be due to increased DMI, greater mTOR signaling, and greater eEF2 activity.     

Effects of feeding rumen-protected methionine pre- and postpartum in multiparous Holstein cows: Lactation performance and plasma amino acid concentrations

Objectives were to evaluate the effect of feeding rumen-protected methionine (RPM) in pre- and postpartum total mix ration (TMR) on lactation performance and plasma AA concentrations in dairy cows. A total of 470 multiparous Holstein cows [235 cows at University of Wisconsin (UW) and 235 cows at Cornell University (CU)] were enrolled approximately 4 wk before parturition, housed in close-up dry cow and replicated lactation pens. Pens were randomly assigned to treatment diets (pre- and postpartum, respectively): UW control (CON) diet = 2.30 and 2.09% of Met as percentage of metabolizable protein (MP) and RPM diet = 2.83 and 2.58% of Met as MP; CU CON = 2.22 and 2.19% of Met as percentage of MP, and CU RPM = 2.85 and 2.65% of Met as percentage of MP. Treatments were evaluated until 112 ± 3 d in milk (DIM). Milk yield was recorded daily. Milk samples were collected at wk 1 and 2 of lactation, and then every other week, and analyzed for milk composition. For lactation pens, dry matter intake (DMI) was recorded daily. Body weight and body condition score were determined from 4 ± 3 DIM and parturition until 39 ± 3 and 49 DIM, respectively. Plasma AA concentrations were evaluated within 3 h after feeding during the periparturient period [d ?7 (±4), 0, 7 (±1), 14 (±1), and 21 (±1); n = 225]. In addition, plasma AA concentrations were evaluated (every 3 h for 24 h) after feeding in cows at 76 ± 8 DIM (n = 16) and within 3 h after feeding in cows at 80 ± 3 DIM (n = 72). The RPM treatment had no effect on DMI (27.9 vs. 28.0 kg/d) or milk yield (48.7 vs. 49.2 kg/d) for RPM and CON, respectively. Cows fed the RPM treatment had increased milk protein concentration (3.07 vs. 2.95%) and yield (1.48 vs. 1.43 kg/d), and milk fat concentration (3.87 vs. 3.77%), although milk fat yield did not differ. Plasma Met concentrations tended to be greater for cows fed RPM at 7 d before parturition (25.9 vs. 22.9 µM), did not differ at parturition (22.0 vs. 20.4 µM), and were increased on d 7 (31.0 vs. 21.2 µM) and remained greater with consistent concentrations until d 21 postpartum (d 14: 30.5 vs. 19.0 µM; d 21: 31.0 vs. 17.8 µM). However, feeding RPM decreased Leu, Val, Asn, and Ser (d 7, 14, and 21) and Tyr (d 14). At a later stage in lactation, plasma Met was increased for RPM cows (34.4 vs. 16.7 µM) consistently throughout the day, with no changes in other AA. Substantial variation was detected for plasma Met concentration (range: RPM = 8.9–63.3 µM; CON = 7.8–28.8 µM) among cows [coefficient of variation (CV) > 28%] and within cow during the day (CV: 10.5–27.1%). In conclusion, feeding RPM increased plasma Met concentration and improved lactation performance via increased milk protein production.     

Effect of dietary protein concentration and degradability on response to rumen-protected methionine in lactating dairy cows

An incomplete 8 × 8 Latin square trial (4-wk periods; 12 wk total) using 32 multiparous and 16 primiparous Holstein cows was conducted to assess the production response to crude protein (CP), digestible rumen-undegraded protein (RUP), and rumen-protected Met (RPM; fed as Mepron; Degussa Corp., Kennesaw, GA). Diets contained [dry matter (DM) basis] 21% alfalfa silage, 34% corn silage, 22 to 26% high-moisture corn, 10 to 14% soybean meal, 4% soyhulls, 2% added fat, 1.3% minerals and vitamins, and 27 to 28% neutral detergent fiber. Treatments were a 2 × 2 × 2 factorial arrangement of the following main effects: 15.8 or 17.1% dietary CP, with or without supplemental rumen-undegraded protein (RUP) from expeller soybean meal, and 0 or 9 g of RPM/d. None of the 2- or 3-way interactions was significant. Higher dietary CP increased DM intake 1.1 kg/d and yield of milk 1.7 kg/d, 3.5% fat-corrected milk (FCM) 2.2 kg/d, fat 0.10 kg/d, and true protein 0.05 kg/d, and improved apparent N balance and DM and fiber digestibility. However, milk urea N and estimated urinary excretion of urea-N and total-N also increased, and apparent N efficiency (milk-N/N-intake) fell from 33 to 30% when cows consumed higher dietary CP. Positive effects of feeding more RUP were increased feed efficiency and milk fat content plus 1.8 kg/d greater FCM and 0.08 kg/d greater fat, but milk protein content was lower and milk urea N and urinary urea excretion were elevated. Supplementation with RPM increased DM intake 0.7 kg/d and FCM and fat yield by 1.4 and 0.06 kg/d, and tended to increase milk fat content and yield of milk and protein.     

Glucose and insulin responses to an intravenous glucose tolerance test administered to feed-restricted dairy cows receiving folic acid and vitamin B12 supplements

The present experiment was conducted to determine whether, during periods of negative energy balance, the increase in glucose availability, despite similar DMI and greater milk production, induced by a combined supplement of folic acid and vitamin B₁₂ was related to effects of insulin on metabolism. Sixteen multiparous Holstein cows averaging 45 days in milk (standard deviation: 3) were assigned to 8 blocks of 2 animals each according to their milk production (45 kg/d; standard deviation: 6) during the week preceding the beginning of the experiment. Within each block, they received weekly intramuscular injections of either saline (CON) or folic acid and vitamin B₁₂ (VIT) during 5 consecutive weeks. During the last week, the cows were fed 75% of their ad libitum intake during 4 d. Blood samples were taken the morning before starting the feed restriction and on the third day of feed restriction. On the fourth day of feed restriction, the daily meal was not served and an intravenous glucose tolerance test was performed. During the 4 wk preceding the feed restriction, milk production and DMI were not affected by treatments. During the feed restriction, the vitamin supplement tended to decrease milk fat concentration and increase milk concentration of lactose. Plasma concentrations of homocysteine, Ile, Leu, Val, and branched-chain AA increased in VIT cows during the restriction but not in CON cows. During the glucose tolerance test, insulin peak height was lower and insulin incremental positive area under the curve tended to be lower for VIT than for CON [83 (95% confidence interval, CI: 64–108) vs. 123 (95% CI: 84–180) µg·180 min/L, respectively]. Free fatty acid nadir was reached earlier for VIT than for CON [34 (95% CI: 26–43) vs. 46 (95% CI: 31–57) min, respectively]. Glucose area under the curve, clearance rate and peak height, insulin time to reach the peak and clearance rate, and free fatty acid nadir did not differ between VIT and CON. The reduction in insulin release during a glucose tolerance test without changes in glucose clearance rate or area under the curve suggests that the vitamin supplement improved insulin sensitivity in feed-restricted lactating dairy cows.     

Effect of ruminally protected methionine on splanchnic metabolism of amino acids in lactating dairy cows

The effect of ruminally protected Met (RPM) on splanchnic metabolism was measured in 3 primiparous and 3 multiparous Holstein cows. Doses of RPM (0, 36, and 72 g/d) were tested in a replicated 3 × 3 Latin square design, over 3 consecutive 14-d experimental periods. A mixed ration was fed in 12 equal meals per d (average dry matter intake: 17.5 ± 0.08 kg/d). Indwelling catheters were surgically implanted in the mesenteric artery and the portal and hepatic veins for blood collection, as well as in 2 distal branches of the mesenteric vein for infusion of p-aminohippurate to determine blood flow. On d 14 of each period, a temporary catheter was inserted into a mammary vein and 6 hourly blood samples were collected to determine plasma concentrations of metabolites, hormones, and their respective fluxes across the splanchnic bed and mammary glands. Yields of milk (32.8, 32.0, and 32.9 ± 0.92 kg/d) and protein (1,028, 1,053, and 1,075 ± 28.7 g/d) were unaffected by level of RPM. However, the true protein content in milk from primiparous cows increased linearly (2.92, 3.09, and 3.34 ± 0.077%). The addition of RPM linearly increased the net flux of Met across the portal-drained viscera, which resulted in increased arterial Met concentrations (25, 29, and 40 ± 1.1 μM). Although it had no significant effect on net portal and hepatic fluxes of other essential amino acids, RPM resulted in a linear increase in the total splanchnic output of Ile, Leu, Phe, and Thr. These results suggest that feeding RPM triggered a homeostatic response resulting in less utilization of certain essential amino acids through the gastrointestinal tract and liver. Net mammary uptake of Met did not change with the addition of RPM. However, mammary extraction of Met decreased in a linear fashion in response to increased arterial inflow.     

Effects of dietary supplementation of rumen-protected conjugated linoleic acid in dairy cows during established lactation

Short-term studies (< 5 d) involving abomasal infusion of a mixture of CLA isomers or pure trans-10, cis-12 CLA have demonstrated that supplements of conjugated linoleic acids (CLA) reduce milk fat synthesis during established lactation in dairy cows. Our objective was to assess longer term effects of supplementation during established lactation using a dietary supplement of rumen-protected CLA. Thirty Holstein cows were blocked by parity and received a dietary fat supplement of either Ca-salts of palm oil fatty acids (control) or a mixture of Ca-salts of palm oil fatty acids plus Ca-salts of CLA (CLA treatment). Supplements provided about 90 g/d of fatty acids and were topdressed on the TMR. The CLA supplement provided 30.4 g/d of CLA in which the predominant isomers were: trans-8, cis-10 (9.2%), cis-9, trans-11 (25.1%), trans-10, cis-12 (28.9%), and cis-11, trans-13 (16.1%). All cows were pregnant; treatments were initiated on d 79 of pregnancy (approximately 200 d prepartum) and continued for 140 d until dry off. Twenty-three cows completed the study; those receiving CLA supplement had a lower milk fat test (2.90 versus 3.80%) and a 23% reduction in milk fat yield (927 versus 1201 g/d). Intake of DM, milk yield, and the yield and content of true protein and lactose in milk were unaffected by treatment. Milk fat analysis indicated that the CLA supplement reduced the secretion of fatty acids of all chain lengths. However, effects were proportionally greater on short and medium chain fatty acids, thereby causing a shift in the milk fatty acid composition to a greater content of longer-chain fatty acids. Changes in body weight gain, body condition score, and net energy balance were not significant and imply no differences in cows fed the CLA supplement in replenishment of body reserves in late lactation. Likewise, maintenance of pregnancy, gestation length, and calf birth weight were unaffected by treatment. Overall, feeding a dietary supplement of rumen-protected CLA to pregnant cows over the last 140 d of the lactation cycle resulted in a marked reduction in milk fat content and yield, and a shift in milk fatty acid composition, but other milk components, DMI, maintenance of pregnancy, and cow well-being were unaffected.