A meta-analysis of methane-mitigation potential of feed additives evaluated in vitro.

A systematic literature review of in vitro studies was performed to identify methane (CH4) mitigation interventions with a potential to reduce CH4 emission in vivo. Data from 277 peer-reviewed studies published between 1979 and 2018 were reviewed. Individual CH4 mitigation interventions were classified into 14 categories of feed additives based on their type, chemical composition, and mode of action. Response variables evaluated were absolute CH4 emission (number of treatment means comparisons = 1,325); total volatile fatty acids (n = 1,007), acetate (n = 783), propionate (n = 792), and butyrate (n = 776) concentrations; acetate to propionate ratio (n = 675); digestibility of dry matter (n = 489), organic matter (n = 277), and neutral detergent fiber (n = 177). Total gas production was used as an explanatory variable in the model for CH4 production. Relative mean difference between treatment and control means reported in the studies was calculated and used for statistical analysis. The robust variance estimation method was used to analyze the effects of CH4 mitigation interventions. In vitro CH4 production was decreased by antibodies (-38.9%), chemical inhibitors (-29.2%), electron sinks (-18.9%), essential oils (-18.2%), plant extracts (-14.5%), plant inclusion (-11.7%), saponins (-14.8%), and tannins (-14.5%). Overall effects of direct-fed microbials, enzymes, macroalgae, and organic acids supplementation did not affect CH4 production in the current meta-analysis. When considering the effects of individual mitigation interventions containing a minimum number of 4 degrees of freedom within feed additives categories, Enterococcus spp. (i.e., direct-fed microbial), nitrophenol (i.e., electron sink), and Leucaena spp. (i.e., tannins) decreased CH4 production by 20.3%, 27.1%, and 23.5%, respectively, without extensively, or only slightly, affecting ruminal fermentation and digestibility of nutrients. It should be noted, however, that although the total number of publications (n = 277) and treatment means comparisons (n = 1,325 for CH4 production) in the current analysis were high, data for most mitigation interventions were obtained from less than 5 observations (e.g., maximum number of observations was 4, 7, and 22 for nitrophenol, Enterococcus spp., and Leucaena spp., respectively), because of limited data available in the literature. These should be further evaluated in vitro and in vivo to determine their true potential to decrease enteric CH4 production, yield, and intensity. Some mitigation interventions (e.g., magnesium, Heracleum spp., nitroglycerin, ß-cyclodextrin, Leptospermum pattersoni, Fructulus Ligustri, Salix caprea, and Sesbania grandiflora) decreased in vitro CH4 production by over 50% but did not have enough observations in the database. These should be more extensively investigated in vitro, and the dose effect must be considered before adoption of mitigation interventions in vivo.

Lactational performance, enteric methane emission, and nutrient utilization of dairy cows supplemented with botanicals.

The objective of this study was to evaluate lactational performance, enteric gas emissions, ruminal fermentation, nutrient use efficiency, milk fatty acid profile, and energy and inflammatory markers in blood of peak-lactation dairy cows fed diets supplemented with Capsicum oleoresin or a combination of Capsicum oleoresin and clove oil. A 10-wk randomized complete block design experiment was conducted with 18 primiparous and 30 multiparous Holstein cows. Cows were blocked based on parity, days in milk, and milk yield (MY), and randomly assigned to 1 of 3 treatments (16 cows/treatment): (1) basal diet (CON); (2) basal diet supplemented with 300 mg/cow per day of Capsicum oleoresin (CAP); and (3) basal diet supplemented with 300 mg/cow per day of a combination of Capsicum oleoresin and clove oil (CAPCO). Premixes containing ground corn (CON), CAP, or CAPCO were mixed daily with the basal diet at 0.8% of dry matter intake (DMI). Supplementation of the diet with CAP or CAPCO did not affect DMI, MY, milk components, and feed efficiency of the cows. Body weight (BW) was increased during the last 2 wk of the experiment by CAP and CAPCO, compared with CON. The botanicals improved BW gain (0.85 and 0.66 kg/d for CAP and CAPCO, respectively, compared with -0.01 kg/d for CON) and CAP enhanced the efficiency of energy utilization, compared with CON (94.5% vs. 78.4%, respectively). Daily CH4 emission was not affected by treatments, but CH4 emission yield (per kg of DMI) and intensity (per kg of MY) were decreased by up to 11% by CAPCO supplementation, compared with CON and CAP. A treatment × parity interaction indicated that the CH4 mitigation effect was pronounced in primiparous but not in multiparous cows. Ruminal molar proportion of propionate was decreased by botanicals, compared with CON. Concentrations of trans-10 C18:1 and total trans fatty acids in milk fat were decreased by CAP and tended to be decreased by CAPCO, compared with CON. Total-tract apparent digestibility of nutrients was not affected by treatments, except for a tendency for decreased starch digestibility in cows supplemented with botanicals. Blood concentrations of ß-hydroxybutyrate, total fatty acids, and insulin were not affected by botanicals. Blood haptoglobin concentration was increased by CAP in multiparous but not in primiparous cows. Lactational performance of peak-lactation dairy cows was not affected by the botanicals in this study, but they appeared to improve efficiency of energy utilization and partitioned energy toward BW gain. In addition, CH4 yield and intensity were decreased in primiparous cows fed CAPCO, suggesting a potential positive environmental effect of the combination of Capsicum oleoresin and clove oil supplementation.

A meta-analysis of methane mitigation potential of feed additives evaluated in vitro.

A systematic literature review of in vitro studies was performed to identify methane (CH4) mitigation interventions with a potential to reduce CH4 emission in vivo. Data from 277 peer-reviewed studies published between 1979 and 2018 were reviewed. Individual CH4 mitigation interventions were classified into 14 categories of feed additives based on their type, chemical composition, and mode of action. Response variables evaluated were absolute CH4 emission (number of treatment means comparisons = 1,325); total volatile fatty acids (VFA; n = 1,007), acetate (n = 783), propionate (n = 792), and butyrate (n = 776) concentrations; acetate to propionate ratio (A:P; n = 675); digestibility of dry matter (DM; n = 489), organic matter (OM; n = 277), and neutral detergent fiber (NDF; n = 177). Total gas production was used as an explanatory variable in the model for CH4 production. Relative mean difference between treatment and control means reported in the studies were calculated and used for statistical analysis. Robust variance estimation method was used to analyze the effects of CH4 mitigation interventions. In vitro CH4 production was decreased by antibodies (-38.9%), chemical inhibitors (-29.2%), electron sinks (-18.9%), essential oils (-18.2%), plant extracts (-14.5%), plants inclusion (-11.7%), saponins (-14.8%), and tannins (-14.5%). Overall effects of direct fed microbials, enzymes, macroalgae, and organic acids supplementation did not affect CH4 production in the current meta-analysis. When considering the effects of individual mitigation interventions containing a minimum number of 4 degrees of freedom within feed additives categories, Enterococcus spp. (i.e., direct fed microbial), nitrophenol (i.e., electron sink), and Leucaena spp. (i.e., tannins) decreased CH4 production by 20.3, 27.1, and 23.5%, respectively, without extensively, or only slightly, affecting ruminal fermentation and digestibility of nutrients. It should be noted, however, that although the total number of publications (n = 277) and treatment means comparisons (n = 1,325 for CH4 production) in the current analysis were high, data for most mitigation interventions were obtained from less than 5 observations (e.g., maximum number of observations was 4, 7, and 22 for nitrophenol, Enterococcus spp., and Leucaena spp., respectively), because of limited data available in the literature. These should be further evaluated in vitro and in vivo to determine their true potential to decrease enteric CH4 production, yield, and intensity. Some mitigation interventions (e.g., magnesium, Heracleum spp., nitroglycerin, ß-cyclodextrin, Leptospermum pattersoni, Fructulus Ligustri, Salix caprea, and Sesbania grandiflora) decreased in vitro CH4 production by over 50% but did not have enough observations in the database. These should be more extensively investigated in vitro, and the dose effect must be considered before adoption of mitigation interventions in vivo.

Lactational performance effects of supplemental histidine in dairy cows: A meta-analysis.

The objective of this meta-analysis was to examine the effects of supplemental His on lactational performance, plasma His concentration and efficiency of utilization of digestible His (EffHis) in dairy cows. The meta-analysis was performed on data from 17 studies published in peer-reviewed journals between 1999 and 2022. Five publications reported data from 2 separate experiments, which were included in the analyses as separate studies, therefore resulting in a total of 22 studies. In 10 studies, His was supplemented as rumen-protected (RP) His; in 1 study, 2 basal diets with different dHis levels were fed; and in the remaining experiments, free His was infused into the abomasum (4 studies), the jugular vein (3 studies) or deleted from a mixture of postruminally infused AA (4 studies). The main forages in the diets were corn silage in 14 and grass silage in 8 studies. If not reported in the publications, the supplies of dietary CP, metabolizable protein (MP), net energy of lactation, and digestible His (dHis) were estimated using NRC (2001). An initial meta-analysis was performed to test the standard mean difference (SMD; raw mean difference of treatment and control means divided by the pooled standard deviation of the means), that is, effect size, and the corresponding 95% confidence interval (CI) in production parameters between His-supplemented groups versus control. Further, regression analyses were also conducted to examine and compare the relationships between several response variables and dHis supply. Across studies, His supplementation increased plasma His concentration (SMD = 1.39; 95% CI: 1.17-1.61), as well as DMI (SMD = 0.240; 95% CI: 0.051-0.429) and milk yield (MY; SMD = 0.667; 95% CI: 0.468-0.866), respectively. Further, milk true protein concentration (MTP; SMD = 0.236; 95% CI: 0.046-0.425) and milk true protein yield (MTPY; SMD = 0.581; 95% CI: 0.387-0.776) were increased by His supplementation. Notably, the increase in MTP concentration and MTPY were 3.9 and 1.3 times greater for studies with MP-deficient (according to NRC 2001) diets compared with studies with MP-adequate diets. The regression analyses revealed that production parameters (DMI, MY, and MTPY) responded in a nonlinear manner to increasing His supply. Further, we detected a difference in the magnitude of change in MTPY and plasma His concentration with the level of His supply and between His supplementation methods, being greater for infused His compared with RPHis. Lastly, a linear and negative relationship between EffHis and the ratio of total digestible His to net energy for lactation supply was observed, indicating an important interaction between dHis and energy supply and EffHis (i.e., utilization of dHis to support protein export). Overall, these analyses confirm His as an important AA in dairy cattle nutrition.

Effects of phytonutrients and yeast culture supplementation on lactational performance and nutrient use efficiency in dairy cows.

Yeast culture and phytonutrients are dietary supplements with distinct modes of action, and they may have additive effects on the performance of dairy cattle. The objective of this study was to investigate the effects of a preparation of phytonutrients and a yeast culture from Saccharomyces cerevisiae on lactational performance, total-tract digestibility of nutrients, urinary nitrogen losses, energy metabolism markers, and blood cells in dairy cows. Thirty-six mid-lactation Holstein cows (10 primiparous and 26 multiparous) were used in an 8-wk randomized complete block design experiment with a 2-wk covariate period, 2 wk for adaptation to the diets, and a 4-wk experimental period for data and samples collection. Following a 2-wk covariate period, cows were blocked by days in milk, parity, and milk yield and randomly assigned to 1 of 3 treatments (12 cows per treatment): basal diet supplemented with 14 g/cow per day yeast culture (YC; S. cerevisiae), basal diet supplemented with 1.0 g/cow per day phytonutrients (PN; 5.5% cinnamaldehyde, 9.5% eugenol, and 3.5% capsicum oleoresin), or basal diet supplemented with a combination of YC and PN (YCPN). Treatments were top-dressed once daily on the total mixed ration at time of feeding. Dry matter intake, milk yield, and feed efficiency were not affected by treatments. Milk composition and energy-corrected milk yield were also not affected by supplementation of YC, PN, and YCPN. There were no differences in intake or total-tract digestibility of dietary nutrients among treatments. Compared with YC, the PN and YCPN treatments tended to decrease the proportion of short-chain fatty acids in milk fat. There was an additive effect of YC and PN supplementation on urinary urea nitrogen (UUN) excretion relative to total nitrogen intake. Cows fed a diet supplemented with YCPN had lower UUN excretion than cows in YC and tended to have lower UUN excretion compared with PN. Blood monocytes count and percentage were decreased in cows fed PN and YCPN diets compared with YC. Treatments did not affect concentrations of blood ß-hydroxybutyrate and total fatty acids. Overall, lactational performance, digestibility of nutrients, energy metabolism markers, and blood cells were not affected by YC, PN, or YCPN supplementation. A combination of PN and YC had an additive effect on nitrogen excretion in dairy cows.

Symposium review: Effective nutritional strategies to mitigate enteric methane in dairy cattle.

Intensive research in the past decade has resulted in a better understanding of factors driving enteric methane (CH4) emissions in ruminants. Meta-analyses of large databases, developed through the GLOBAL NETWORK project, have identified successful strategies for mitigation of CH4 emissions. Methane inhibitors, alternative electron sinks, vegetable oils and oilseeds, and tanniferous forages are among the recommended strategies for mitigating CH4 emissions from dairy and beef cattle and small ruminants. These strategies were also effective in decreasing CH4 emissions yield and intensity. However, a higher inclusion rate of oils may negatively affect feed intake, rumen function, and animal performance, specifically milk components in dairy cows. In the case of nitrates (electron sinks), concerns with animal health may be impeding their adoption in practice, and potential emission trade-offs have to be considered. Tannins and tanniferous forages may have a negative effect on nutrient digestibility, and more research is needed to confirm their effects on overall animal performance in long-term experiments with high-producing animals. A meta-analysis of studies with dairy cows fed the CH4 inhibitor 3-nitrooxypropanol (3-NOP) at the Pennsylvania State University showed (1) a consistent 28 to 32% decrease in daily CH4 emissions or emissions yield and intensity; (2) no effect on dry matter intake, milk production, body weight, or body weight change, and a slight increase in milk fat concentration and yield (0.19 percentage units and 90 g/d, respectively); 3-NOP also appears to increase milk urea nitrogen concentration; (3) an exponential decrease in the mitigation effect of the inhibitor with increasing its dose (from 40 to 200 mg/kg of feed dry matter, corresponding to 3-NOP intake of 1 to 4.8 g/cow per day); and (4) a potential decrease in the efficacy of 3-NOP over time, which needs to be further investigated in long-term, full-lactation or multiple-lactation studies. The red macroalga Asparagopsis taxiformis has a strong CH4 mitigation effect, but studies are needed to determine its feasibility, long-term efficacy, and effects on animal production and health. We concluded that widespread adoption of mitigation strategies with proven effectiveness by the livestock industries will depend on cost, government policies and incentives, and willingness of consumers to pay a higher price for animal products with decreased carbon footprint.

Performance of dairy cows fed normal- or reduced-starch diets supplemented with an exogenous enzyme preparation.

The objective of this study was to investigate the effects of supplementation of an exogenous enzyme preparation (EEP) on performance, total-tract digestibility of nutrients, plasma AA profile, and milk fatty acids composition in lactating dairy cows fed a reduced-starch diet compared with a normal-starch diet (i.e., positive control). Forty-eight Holstein cows (28 primiparous and 20 multiparous) were enrolled in a 10-wk randomized complete block design experiment with 16 cows per treatment. Treatments were as follows: (1) normal-starch diet (control) containing (% dry matter basis) 24.8% starch and 33.0% neutral detergent fiber (NDF), (2) reduced-starch diet (RSD) containing 18.4% starch and 39.1% NDF, or (3) RSD supplemented with 10 g/cow per day of an EEP (ENZ). The EEP contained amylolytic and fibrolytic activities and was top-dressed on the total mixed ration at the time of feeding. Compared with normal-starch diet, dry matter intake and milk and energy-corrected milk (ECM) yields were lower (on average by 7.1, 9.5, and 7.2%, respectively) for cows on the RSD treatments. Concentrations, but not yields, of milk fat and total solids were increased by RSD. Energy-corrected milk feed efficiency did not differ among treatments. Total-tract digestibility of NDF tended to increase by RSD treatments. Plasma AA concentrations were not affected by treatment, except that of 3-methylhistidine was increased by ENZ, compared with RSD. Blood glucose concentration tended to be lower in cows on the RSD treatments, but ENZ increased glucose and tended to increase insulin concentrations at 4 h after feeding when compared with RSD. Cows on the RSD treatments had decreased concentrations of de novo fatty acids and tended to have increased concentrations of preformed fatty acids in milk. Overall, decreasing dietary starch concentration by 26% decreased dry matter intake, milk, and ECM yields, but ECM feed efficiency was not different among treatments. The negative effects of reducing dietary starch on production were not attenuated by the EEP.

Production effects and bioavailability of N-acetyl-l-methionine in lactating dairy cows.

Two experiments were conducted to investigate the production effects of N-acetyl-l-methionine (NALM; experiment 1) and to estimate its bioavailability (BA) and rumen escape (RE; experiment 2), respectively, in lactating dairy cows. In experiment 1, 18 multiparous Holstein cows were used in a replicated, 3 × 3 Latin square design experiment with three 28-d periods. Treatments were (1) basal diet estimated to supply 45 g/d digestible Met (dMet) or 1.47% of metabolizable protein (MP; control), (2) basal diet top-dressed with 32 g/d of NALM to achieve dMet supply of 2.2% of MP, and (3) basal diet top-dressed with 56 g/d of NALM to achieve dMet supply of 2.6% of MP. The NALM treatments supplied estimated 17 and 29 g/d dMet from NALM, respectively, based on manufacturer's specifications. In experiment 2, 4 rumen-cannulated lactating Holstein cows were used in a 4 × 4 Latin square design experiment with four 12-d periods. A 12-d period for baseline data collection and 4 d for determination of RE of NALM preceded the Latin square experiment. For determination of RE, 30 g of NALM were dosed into the rumen simultaneously with Cr-EDTA (used as a rumen fluid kinetics marker) and samples of ruminal contents were collected at 0 (before dosing), 1, 2, 4, 6, 8, 10, 14, 18, and 24 h after dosing. Rumen escape of NALM was calculated using the estimated passage rate based on the measured Cr rate of disappearance. Bioavailability of abomasally dosed NALM was determined using the area under the curve of plasma Met concentration technique. Two doses of l-Met (providing 7.5 and 15 g of dMet) and 2 doses of NALM (11.2 and 14.4 g of dMet) were separately pulse-dosed into the abomasum of the cows and blood was collected from the jugular vein for Met concentration analysis at 0 (before dosing), 1, 2, 4, 6, 8, 10, 12, 14, 18, and 24 h after dosing. Supplementation of NALM did not affect DMI, milk yield, feed efficiency, or milk protein and lactose concentrations and yields in experiment 1. Milk fat concentration and energy-corrected milk yield decreased linearly with NALM dose. Plasma Met concentration was not affected by NALM dose. The estimated relative BA of abomasally dosed NALM (experiment 2) was 50% when dosed at 14.4 g/cow (11.2 g/d dMet from NALM) and 24% when dosed at 28.8 g/cow (14.4 g/d dMet from NALM). The estimated RE of NALM was 19% based on the measured kp of Cr at 11%/h. The total availability of ingested NALM was estimated at 9.5% for the lower NALM dose when taking into account RE (19%) and bioavailability in the small intestine (50%). Overall, NALM supplementation to mid-lactation dairy cows fed a MP-adequate basal diet below NRC (2001) recommendations (45 g/d or 1.47% Met of MP) decreased milk fat and energy-corrected milk yields but did not affect milk or milk true protein yields. Further evaluation of BA of NALM at different doses is warranted. In addition, intestinal conversion of NALM to Met needs additional investigation to establish a possible saturation of the enzyme aminoacylase I at higher NALM doses.

Dietary supplementation with rumen-protected capsicum during the transition period improves the metabolic status of dairy cows.

In ruminants, it has been observed that capsicum oleoresin can alter insulin responses and that high-intensity artificial sweetener can increase glucose absorption from the small intestine. Because glucose metabolism and insulin responses are critical during early lactation, these supplements might have an effect on the metabolic status of dairy cows during the transition period. The objective of this experiment was to evaluate the effects of rumen-protected capsicum oleoresin fed alone or in combination with artificial sweetener during the transition period on lactational performance and susceptibility to subclinical ketosis in dairy cows. Fifteen primiparous and 30 multiparous Holstein cows (a total of 39 cows finished the study) were arranged in a randomized complete block design during d -21 to 60 relative to parturition. Cows within block were randomly assigned to one of the following treatments: no supplement (CON), supplementation with 100 mg of rumen-protected capsicum/cow per day (RPCap), or RPCap plus 2 g of high-intensity artificial sweetener/cow per day (RPCapS). For both the RPCap and RPCapS treatments, only rumen-protected capsicum was fed during the dry period. From d 8 to 11 of lactation, intake was limited to 70% of predicted dry matter intake to induce subclinical ketosis. Production variables were recorded daily, samples for milk composition were collected on wk 2, 4, 6, and 8, and blood samples were collected on wk -2, 1, 2, and 4 of the experiment for analysis of metabolic hormones and blood cell counts. Supplementation with rumen-protected capsicum increased serum insulin and decreased ß-hydroxybutyrate concentrations precalving, indicating a decrease in lipolysis. During the lactation period, RPCap was associated with a trend for increased milk production and feed efficiency following the ketosis challenge. Supplementation with RPCapS appeared to negate the response to rumen-protected capsicum. All cows developed subclinical ketosis during the challenge, and this was not affected by treatment. We conclude that treatments did not decrease susceptibility to subclinical ketosis; however, dietary supplementation with rumen-protected capsicum was effective at improving energy status precalving and tended to increased milk production and feed efficiency. The mechanism underlying these responses is unclear.

Lactational performance, enteric gas emissions, and plasma amino acid profile of dairy cows fed diets with soybean or canola meals included on an equal protein basis.

This study investigated the effects of feeding solvent-extracted canola meal (CM), extruded soybean meal (ESBM), or solvent-extracted soybean meal (SSBM) on an equivalent crude protein basis on performance, plasma AA profiles, enteric gas emissions, milk fatty acids, and nutrient digestibility in lactating dairy cows. Fifteen Holstein cows (95 ± 20 d in milk) were used in a replicated 3 × 3 Latin square design experiment with 3 periods of 28 d each. Treatments were 3 diets containing 17.1% CM, 14.2% ESBM, or 13.6% SSBM (dry matter basis). Vegetable oil was added (canola oil for CM or soybean oil for SSBM) to equalize the ether extract concentration of the diets. Rumen-protected Met was supplemented targeting digestible Met supply of 2.2% of metabolizable protein in all diets. Canola meal increased dry matter intake (DMI) by 5.9 and 8.9% in comparison with ESBM and SSBM, respectively. Milk urea nitrogen was lowest in CM, followed by SSBM, and was highest for ESBM. No differences were observed in feed efficiency, energy-corrected milk yield, and milk composition or component yields among treatments. Cows fed CM emitted less enteric CH4 per kg of DMI compared with both ESBM and SSBM, but CH4 emission intensity (CH4 per kg of energy-corrected milk) was similar among treatments. In summary, replacement of ESBM or SSBM with CM, on an equal crude protein basis, in the diet of lactating dairy cows enhanced DMI, but yields of energy-corrected milk and milk components and feed efficiency were similar among treatments.

Effects of the macroalga Asparagopsis taxiformis and oregano leaves on methane emission, rumen fermentation, and lactational performance of dairy cows.

Asparagopsis taxiformis (AT) is a source of multiple halogenated compounds and, in a limited number of studies, has been shown to decrease enteric CH4 emission in vitro and in vivo. Similarly, oregano has been suggested as a potential CH4 mitigating agent. This study consisted of 2 in vitro and 2 in vivo experiments. Experiment (Exp.) 1 was aimed at establishing the effect of AT on CH4 emission in vitro. Two experiments (Exp. 2 and 3) with lactating dairy cows were conducted to determine the antimethanogenic effect of AT and oregano (Exp. 3) in vivo. Another experiment (Exp. 4) was designed to investigate stability of bromoform (CHBr3) in AT over time. In Exp. 3, 20 Holstein cows were used in a replicated 4 × 4 Latin square design with four 28-d periods. Treatments were basal diet (control) or basal diet supplemented with (dry matter basis) 0.25% AT (LowAT), 0.50% AT (HighAT), or 1.77% oregano (Origanum vulgare L.) leaves. Enteric gas emissions were measured using the GreenFeed system (C-Lock Inc., Rapid City, SD), and rumen samples were collected for fermentation analysis using the ororuminal technique. In Exp.1 (in vitro), relative to the control, AT (at 1% dry matter basis, inclusion rate) decreased CH4 yield by 98%. In Exp. 3, HighAT decreased average daily CH4 emission and CH4 yield by 65% and 55%, respectively, in experimental periods 1 and 2, but had no effect in periods 3 and 4. The differential response to AT among experimental periods was likely a result of a decrease in CHBr3 concentration in AT over time, as observed in Exp. 4 (up to 84% decrease in 4 mo of storage). In Exp. 3, H2 emission was increased by AT and, as expected, the proportion of acetate in the total volatile fatty acids in the rumen was decreased and those of propionate and butyrate were increased by HighAT compared with the control. Compared with the control, HighAT decreased dry matter intake, milk yield, and energy-corrected milk yield in Exp. 3. Milk composition was not affected by treatment, except lactose percentage and yield were decreased by HighAT. Concentrations of iodine and bromide in milk were increased by HighAT compared with the control. Milk CHBr3 concentration and its organoleptic characteristics were not different between control and HighAT. Oregano had no effect on CH4 emission or lactational performance of the cows in Exp. 3. Overall, AT included at 0.50% in the ration of dairy cows can have a large mitigation effect on enteric CH4 emission, but dry matter intake and milk production may also decrease. There was a marked decrease in the CH4 mitigation potential of AT in the second half of Exp. 3, likely resulting from CHBr3 decay over time.

Short communication: Short-term effect of 3-nitrooxypropanol on feed dry matter intake in lactating dairy cows.

The objective of this study was to investigate the effect of 3-nitrooxypropanol (3-NOP), an enteric methane inhibitor under investigation, on short-term dry matter intake (DMI) in lactating dairy cows. Following a 1-wk adaptation period, 12 multiparous Holstein cows were fed a basal total mixed ration (TMR) containing increasing levels of 3-NOP during 5 consecutive, 6-d periods. The experiment was conducted in a tiestall barn. Feed bins were split in half by a solid divider, and cows simultaneously received the basal TMR supplemented with the following: (1) a placebo without 3-NOP or (2) 3-NOP included in the TMR at 30, 60, 90, or 120 mg/kg of feed dry matter (experimental periods 2, 3, 4, and 5, respectively). Cows received the control diet (basal TMR plus placebo premix) during experimental period 1. A premix containing ground corn grain, soybean oil, and dry molasses was used to incorporate 3-NOP in the ration. Cows were fed twice daily as follows: 60% of the daily feed allowance at 0800 h and 40% at 1800 h. Feed offered and refused was recorded at each feeding. During the morning feedings, each cow was offered either control or 3-NOP-treated TMR at 150% of her average intake during the previous 3 d. After collection of the evening refusals, cows received only the basal TMR without the premix until the next morning feeding. The test period for the short-term DMI data collection was defined from morning feeding to afternoon refusals collection during each day of each experimental period. Location (left or right) of the control and 3-NOP diets within a feed bin was switched every day during each period to avoid feed location bias. Dry matter intake of TMR during the test period was quadratically increased by 3-NOP compared with the control. Inclusion of 3-NOP at 120 mg/kg of feed dry matter resulted in decreased 10-h DMI compared with the lower 3-NOP doses, but was similar to the control. There was no effect of feed location (left or right) within feed bin on DMI. Data from this short-term study suggests that 3-NOP does not have a negative effect on DMI in lactating dairy cows.

Effects of lowering crude protein supply alone or in a combination with essential oils on productivity, rumen function and nutrient utilization in dairy cows.

Lowering dietary protein concentration is known to decrease urinary nitrogen (N) losses and increase milk N efficiency in dairy cows, but it may negatively affect animal productivity. Plant-derived essential oils (EO) may alleviate these negative effects by improving the efficiency of rumen fermentation in cows fed reduced feed protein diets. The experiment was conducted to investigate the effects of lowering crude protein (CP) supply alone or in a combination with an EO product on feed intake, milk production and composition, rumen fermentation, total tract digestibility and N utilization in dairy cows. Twenty-one Holstein cows were used in a replicated 3 × 3 Latin square design experiment. Each period consisted of 14 days for adaptation and 14 days for data collection and sampling. Cows were randomly assigned to one of three experimental diets: a 165 g/kg CP diet (control), a 155 g/kg CP diet (LCP) and LCP supplemented with 35 g/day per cow EO (LCPEO). The dry matter (DM) intake was decreased by LCP and LCPEO compared with the control; there was no effect of EO on DM intake. Milk yield and composition and feed efficiency were similar among treatments. Ruminal pH, lactate, ammonia and volatile fatty acids concentrations were not affected by treatment, except increased valerate concentration by LCPEO compared with LCP. The supplementation of EO tended to decrease protozoal counts. The LCP and LCPEO increased total tract digestibility of DM and organic matter and decreased CP digestibility compared with the control. Supplementation with EO did not affect total tract digestibility of dietary nutrients compared with the control or LCP. The LCP and LCPEO decreased urinary and fecal N excretions and increased milk N efficiency; nitrogen losses were not affected by EO. In this study, lowering dietary CP by 10 g/kg decreased urinary and fecal N excretion without affecting productivity. The supplementation of EO to LCP had only minor effects on rumen fermentation and did not affect productivity, digestibility and N excretion in lactating dairy cows.

Effects of phytonutrients alone or in combination with monensin on productivity in lactating dairy cows.

This experiment was conducted to investigate the effects of phytonutrients, compared with monensin as a positive control, on productivity, milk fatty acids, fat mobilization, and blood cells in lactating dairy cows. Thirty-six Holstein cows were used in a 9-wk randomized complete block design study. Following a 2-wk covariate period, cows were blocked by days in milk, parity, and milk yield and randomly assigned to 1 of 3 treatments (12 cows/treatment): 450 mg/cow per day of monensin (MO), 250 mg/cow per day of capsicum plus 450 mg/cow per day of MO (MOCAP), and 1,000 mg/cow per day of a mixture of cinnamaldehyde, eugenol, and capsicum (CEC). Dry matter intake and milk yield were not affected by treatment. Supplementation of CEC increased feed efficiency compared with MO, but did not affect feed efficiency on an energy-corrected milk basis. Milk composition (fat, protein, and lactose), milk fatty acid profile, and blood concentrations of nonesterified fatty acids and ß-hydroxybutyrate were also not affected by treatment. The expression of hormone-sensitive lipase in adipose tissues tended to increase for MOCAP compared with MO. Counts of total white blood cell, neutrophils, lymphocytes, eosinophils, and basophils were not affected by treatment, although monocytes count tended to be decreased by CEC. Treatments had no effect on red blood cells, hemoglobin, and platelets. Results indicate that dietary supplementation of CEC and capsicum had no production or other effects in dairy cows, compared with MO, except CEC increased feed efficiency and tended to decrease blood monocytes count.

The binding of orally dosed hydrophobic active pharmaceutical ingredients to casein micelles in milk.

Casein proteins (αS1-, αS2-, ß- and κ-casein) account for 80% of the total protein content in bovine milk and form casein micelles (average diameter = 130 nm, approximately 1015 micelles/mL). The affinity of native casein micelles with the 3 hydrophobic active pharmaceutical ingredients (API), meloxicam [351.4 g/mol; log P = 3.43; acid dissociation constant (pKa) = 4.08], flunixin (296.2 g/mol; log P = 4.1; pKa = 5.82), and thiabendazole (201.2 g/mol; log P = 2.92; pKa = 4.64), was evaluated in bovine milk collected from dosed Holstein cows. Native casein micelles were separated from raw bovine milk by mild techniques such as ultracentrifugation, diafiltration, isoelectric point precipitation (pH 4.6), and size exclusion chromatography. Acetonitrile extraction of hydrophobic API was then done, followed by quantification using HPLC-UV. For the API or metabolites meloxicam, 5-hyroxy flunixin and 5-hydroxy thiabendazole, 31 ± 3.90, 31 ± 1.3, and 28 ± 0.5% of the content in milk was associated with casein micelles, respectively. Less than ∼5.0% of the recovered hydrophobic API were found in the milk fat fraction, and the remaining ∼65% were associated with the whey/serum fraction. A separate in vitro study showed that 66 ± 6.4% of meloxicam, 29 ± 0.58% of flunixin, 34 ± 0.21% of the metabolite 5-hyroxy flunixin, 50 ± 4.5% of thiabendazole, and 33 ± 3.8% of metabolite 5-hydroxy thiabendazole was found partitioned into casein micelles. Our study supports the hypothesis that casein micelles are native carriers for hydrophobic compounds in bovine milk.

Histidine deficiency has a negative effect on lactational performance of dairy cows.

A 10-wk randomized complete block design experiment with 24 Holstein cows was conducted to investigate the long-term effects of feeding a His-deficient diet on lactational performance of dairy cows. Cows were blocked by days in milk, milk yield, and parity, and randomly assigned to 1 of the following 2 treatments: (1) His-adequate diet [HAD; providing +166 g/d over metabolizable protein (MP) requirements, according to the National Research Council (2001) and digestible His (dHis) supply of 68 g/d, or 2.5% of MP requirements] and (2) His-deficient diet (HDD; +37 g/d over MP requirements and dHis supply of 49 g/d, or 1.9% of MP requirements). Both HAD and HDD were supplemented with rumen-protected (RP) Met and Lys supplying digestible Met and digestible Lys at 2.4 and 2.4% and 7.2 and 7.1% of MP requirements, respectively. At the end of the 10-wk experiment, HDD was supplemented with RPHis (HDD+RPHis; total dHis supply of 61 g/d, or 2.4% of MP requirements) for an additional 9 d. Dry matter intake (DMI; 25.4 and 27.1 kg/d, standard error of the mean = 0.41), yields of milk (37.6 and 40.5 kg/d, standard error of the mean = 0.62), protein and lactose, energy-corrected milk, and milk and plasma urea-N were decreased by HDD compared with HAD. Feed and energy-corrected milk feed efficiencies, milk fat, protein and lactose concentrations, body weight, and body condition score of the cows were not affected by treatment. Apparent total-tract digestibility of dry and organic matter, crude protein, and neutral detergent fiber, and excretion of urinary N and urea-N were decreased by HDD compared with HAD. Concentration of plasma leptin tended to be decreased for HDD compared with HAD. Plasma concentrations of EAA (His, Leu, Lys, Val) and carnosine decreased and total EAA tended to be decreased in cows fed HDD compared with HAD. Muscle concentrations of free His, Leu, and Val decreased and Gly and ß-alanine tended to be increased by HDD compared with HAD. Cows fed HDD had a lower blood hemoglobin concentration than cows fed HAD. At the end of the 10-wk study, the 9-d supplementation of HDD with RPHis (i.e., HDD+RPHis) increased DMI and plasma His, and tended to increase energy-corrected milk yield and plasma carnosine, compared with HDD. In conclusion, feeding a diet deficient in dHis supplying adequate MP, digestible Met, and digestible Lys affected negatively lactational performance of dairy cows. These results confirm our previous findings that low dietary His supply can impair DMI, yields of milk and milk protein, and blood hemoglobin in dairy cows.

Effects of rumen-protected methionine, lysine, and histidine on lactation performance of dairy cows.

The objective of this study was to evaluate the effects of supplementing a metabolizable protein (MP)-deficient diet with rumen-protected (RP) Met, Lys, and His, individually or combined, on the performance of lactating dairy cows. The experiment was a 9-wk randomized complete block design with 72 Holstein cows. Following a 2-wk covariate period, cows were blocked by days in milk, milk yield, and parity, and randomly assigned to 1 of the following 6 treatments: (1) MP-adequate diet [MPA; +243g/d MP balance, according to the National Research Council (2001) requirements]; (2) MP-deficient diet (MPD; -54g/d MP balance); (3) MPD supplemented with RPMet (MPDM); (4) MPD supplemented with RPLys (MPDL); (5) MPD supplemented with RPHis (MPDH); and (6) MPD supplemented with RPMet, RPLys, and RPHis (MPDMLH). Dry matter intake (DMI), yields of milk and milk components (fat, protein, lactose) and energy-corrected milk (ECM), feed and ECM feed efficiencies, and milk and plasma urea N were decreased by MPD, compared with MPA. Supplementation of the MPD diet with RPLys increased milk protein content and plasma glucose concentration and tended to increase milk urea N. Addition of RPHis tended to increase DMI, increased milk protein concentration, and numerically increased yields of milk fat, protein, and ECM. In addition to the trends for increased DMI and milk fat content, and higher milk protein concentration, supplementation of the 3 RP AA also increased yields of milk fat, protein, and ECM and ECM feed efficiency. Relative to MPA, milk N efficiency tended to be increased by MPD. Concentrations of plasma essential AA (except Met and Thr) were decreased by MPD compared with MPA. Supplementation of RPMet, RPLys, and RPHis increased plasma Met (except for MPDM), Lys, and His concentrations, respectively. Cows fed MPD had lower blood hemoglobin concentration and numerically higher plasma ghrelin than cows fed MPA. Concentration of total saturated fatty acids in milk fat were or tended to be higher for MPD compared with MPA and MPDMLH, respectively. Concentration of total polyunsaturated and yield of milk odd- and branched-chain fatty acids were or tended to be decreased by MPD compared with MPA. Overall, the results of this study confirm our previous data and suggest that His stimulates DMI and the combination of the 3 RP AA (Met, Lys, and His) has the potential to improve milk and milk component yields in dairy cows fed MP-deficient diets.

Effects of dietary Capsicum oleoresin on productivity and immune responses in lactating dairy cows.

This study investigated the effect of Capsicum oleoresin in granular form (CAP) on nutrient digestibility, immune responses, oxidative stress markers, blood chemistry, rumen fermentation, rumen bacterial populations, and productivity of lactating dairy cows. Eight multiparous Holstein cows, including 3 ruminally cannulated, were used in a replicated 4×4 Latin square design experiment. Experimental periods were 25 d in duration, including a 14-d adaptation and an 11-d data collection and sampling period. Treatments included control (no CAP) and daily supplementation of 250, 500, or 1,000 mg of CAP/cow. Dry matter intake was not affected by CAP (average 27.0±0.64 kg/d), but milk yield tended to quadratically increase with CAP supplementation (50.3 to 51.9±0.86 kg/d). Capsicum oleoresin quadratically increased energy-corrected milk yield, but had no effect on milk fat concentration. Rumen fermentation variables, apparent total-tract digestibility of nutrients, and N excretion in feces and urine were not affected by CAP. Blood serum ß-hydroxybutyrate was quadratically increased by CAP, whereas the concentration of nonesterified fatty acids was similar among treatments. Rumen populations of Bacteroidales, Prevotella, and Roseburia decreased and Butyrivibrio increased quadratically with CAP supplementation. T cell phenotypes were not affected by treatment. Mean fluorescence intensity for phagocytic activity of neutrophils tended to be quadratically increased by CAP. Numbers of neutrophils and eosinophils and the ratio of neutrophils to lymphocytes in peripheral blood linearly increased with increasing CAP. Oxidative stress markers were not affected by CAP. Overall, in the conditions of this experiment, CAP did not affect feed intake, rumen fermentation, nutrient digestibility, T cell phenotypes, and oxidative stress markers. However, energy-corrected milk yield was quadratically increased by CAP, possibly as a result of enhanced mobilization of body fat reserves. In addition, CAP increased neutrophil activity and immune cells related to acute phase immune response.

Effects of slow-release urea and rumen-protected methionine and histidine on performance of dairy cows.

This experiment was conducted with the objective to investigate the effects of slow-release urea and rumen-protected (RP) Met and His supplementation of a metabolizable protein (MP)-deficient diet (according to NRC, 2001) on lactation performance of dairy cows. Sixty lactating Holstein cows were used in a 10-wk randomized complete block-design trial. Cows were fed a covariate diet for 2 wk and then assigned to one of the following treatments for an 8-wk experimental period: (1) MP-adequate diet [AMP; 107% of MP requirements, based on the National Research Council (NRC, 2001)]; (2) MP-deficient diet (DMP; 95% of MP requirements); (3) DMP supplemented with slow-release urea (DMPU); (4) DMPU supplemented with RPMet (DMPUM); and (5) DMPUM supplemented with RPHis (DMPUMH). Total-tract apparent digestibility of dry matter, organic matter, neutral detergent fiber, and crude protein, and urinary N and urea-N excretions were decreased by DMP, compared with AMP. Addition of slow-release urea to the DMP diet increased urinary urea-N excretion. Dry matter intake (DMI) and milk yield (on average 44.0±0.9kg/d) were not affected by treatments, except DMPUMH increased DMI and numerically increased milk yield, compared with DMPUM. Milk true protein concentration and yield were increased and milk fat concentration tended to be decreased by DMPUMH, compared with DMPUM. Cows gained less body weight on the DMP diet, compared with AMP. Plasma concentrations of His and Lys were not affected by treatments, whereas supplementation of RPMet increased plasma Met concentration. Plasma concentration of 3-methylhistidine was or tended to be higher for DMP compared with AMP and DMPU, respectively. Addition of RPHis to the DMPUM diet tended to increase plasma glucose and creatinine. In conclusion, feeding a 5% MP-deficient diet (according to NRC, 2001) did not decrease DMI and yields of milk and milk components, despite a reduction in nutrient digestibility. Supplementation of RPHis increased DMI and milk protein concentration and yield. These results are in line with our previous data and suggest that His may have a positive effect on voluntary feed intake and milk production and composition in high-yielding dairy cows fed MP-deficient diets.

Effect of technical cashew nut shell liquid on rumen methane emission and lactation performance of dairy cows.

Technical-grade cashew nut shell liquid (TCNSL) is a by-product of the cashew nut industry in tropical countries, and is known to exhibit a wide range of biological activities, including inhibitory effect against gram-positive bacteria. This study was conducted to investigate the effects of TCNSL (73.3% cardanol, 16.4% cardol, and 3.0% methylcardol) on rumen methane emission, nutrient digestibility, dry matter intake, and milk yield and composition in dairy cows. Eight multiparous Holstein cows were used in a crossover design trial with two 21-d experimental periods. The diet was based on corn silage and alfalfa haylage and was formulated to meet or exceed the energy and metabolizable protein requirements of the cows. Treatments were control (no TCNSL supplementation) or 30 g/cow per day of TCNSL. Rumen carbon dioxide emission was not affected by TCNSL. Treatment had no effect on methane emission (542 vs. 511±35.3 g/cow per day, respectively) and methane emission intensity (15.0 vs. 13.9±0.58 g/kg of energy-corrected milk, respectively) and tended to decrease methane emission per kilogram of dry matter intake (20.2 vs. 18.6±1.04 g/kg, respectively). Dry matter intake (average 26.9±1.00 kg/d), milk yield (40.0±1.73 kg/d), and milk composition were not different between treatments. The TCNSL had no effect on N losses in urine and feces and total-tract apparent digestibility of nutrients, except digestibility of neutral detergent fiber tended to be increased compared with the control. Plasma urea and glucose concentrations were not affected by TCNSL. Concentration of milk C18:0 tended to be decreased (17%) by TCNSL compared with the control. In this study, TCNSL did not alter absolute methane emission in the rumen, but tended to decrease it by 8% per kilogram of dry matter intake. The TCNSL had no effect on milk yield and composition in dairy cows.