This study examined the effect of 3-nitrooxypropanol (3-NOP), a substance under investigation, on enteric methane (CH 4 ) emission, rumen fermentation, lactational performance, sensory properties of milk, and the resumption of ovarian cyclicity in early-lactation dairy cows. Fifty-six multi-and primiparous Holstein cows, including 8 that were rumen cannulated, were used in a 15-wk randomized complete block design experiment. Cows were blocked based on parity and previous lactation milk yield (MY) or predicted MY, and within each block were randomly assigned to one of 2 treatments: (1) control (CON), administered no 3-NOP, or (2) 3-NOP applied at 60 mg/kg of feed dry matter (3-NOP). Enteric CH 4 emission was measured during experimental wk 2, 6, 9, and 15, using the GreenFeed system. Dry matter intake (DMI) and MY data were collected daily throughout the experiment, and milk composition samples were collected 7 times during the experiment. Milk samples were collected from 14 to 60 (±2) d after calving, 3 d per week, and assayed for progesterone concentration to determine resumption of ovarian activity. Compared with CON, 3-NOP decreased daily CH 4 emission by 26%, CH 4 yield (CH 4 per kg of DMI) by 21%, and CH 4 emission intensity [CH 4 per kg of MY or energy-corrected milk (ECM)] by 25%. Enteric emission of carbon dioxide was decreased by 5%, and hydrogen emission was increased 48-fold by 3-NOP. Inclusion of 3-NOP decreased concentration of total volatile fatty acids (by 9.3%) and acetate but increased butyrate molar proportion, ethanol, and formate concentrations in ruminal fluid. Dry matter intake was lower for 3-NOP compared with CON, but DMI expressed as a percentage of body weight was not different between treatments. Treatment had no effect on milk and ECM, body weight change, or body condition score. Milk composition and milk fat and protein yields were not affected by treatment, except that concentrations of short-chain fatty acids in milk were increased by 3-NOP. Nutrient digestibility and blood metabolites and hormones were not affected by 3-NOP, except that insulin was decreased by 3-NOP. There was no effect of 3-NOP on postpartum resumption of ovarian activity, including days to first and second luteal phases, length of first and second luteal phases, and interval from first to second luteal phase. Sensory properties of milk from cows fed 3-NOP and cheese made from that milk were not affected by treatment. In this experiment, 3-NOP decreased daily enteric CH 4 emission, emission yield, and emission intensity, improved feed efficiency, and did not affect lactational performance or onset of ovarian activity in early-lactation dairy cows.

This study was conducted to investigate the effect of dextrose, starch, NDF, and a carbohydrate (CHO) mix on utilization of ruminal ammonia in dairy cows. Four ruminally and duodenally cannulated Holstein cows (BW = 788 +/- 31 kg; 217 +/- 35 d in milk) were allocated to four treatments in a 4 x 4 Latin square design trial. Cows were fed an all alfalfa diet at 12-h intervals (DMI = 22.2 +/- 0.25 kg/d). Treatments were control, white oat fiber (NDF); corn dextrose (GLU); cornstarch (STA); and a CHO mix (25% of each): apple pectin, GLU, STA, and NDF (MIX). Carbohydrates were introduced intraruminally during feeding at 20% of dietary DMI. Ruminal ammonia was labeled with (15)N. Ruminal pH was the highest for NDF followed by STA and MIX and GLU (P < 0.001). Ruminal ammonia concentration and pool size were decreased by GLU and STA compared with NDF (P < 0.001 and P = 0.03, respectively). Acetate, isobutyrate, isovalerate, and total VFA concentration in the rumen were decreased (P = 0.009 to 0.001), and butyrate was increased (P < 0.001) by GLU compared with the other CHO. Microbial N flow to the duodenum was decreased (P < 0.05) by NDF compared with the other CHO, and the flow of microbial N formed from ammonia was greater for STA compared with GLU and NDF (P = 0.04 and 0.03, respectively). Urinary N loss was decreased (P = 0.05) by GLU and STA, but overall (feces plus urine) N losses were not affected (P = 0.73) by treatment. Milk urea concentration was lowered by GLU and STA compared with NDF and MIX (P = 0.002). The proportion of bacterial N synthesized from ammonia in the rumen was greater with STA than with NDF and MIX and was least for GLU (P = 0.02). Irreversible ammonia loss and flux were lower (P = 0.09 and 0.02, respectively) for GLU than for STA and NDF. As a percentage of the dose given, cumulative secretion of (15)N ammonia in milk protein was greater for STA than for GLU or NDF (P = 0.01 and 0.001, respectively). This experiment demonstrated that provision of readily fermentable energy can decrease ammonia concentrations in the rumen through decreased ammonia production (GLU), or through enhanced uptake of ammonia for microbial protein synthesis (STA). Rapidly fermentable energy in the rumen decreased ammonia production and flux, but the overall efficiency of ammonia utilization for milk protein synthesis was only increased by enhancing ruminal microbial ammonia uptake.

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This experiment was designed to test the effect of inclusion rate of 3-nitrooxypropanol (3-NOP), a methane inhibitor, on enteric methane emissions in dairy cows. The study was conducted with 49 multiparous Holstein cows in a randomized complete block design in 2 phases; phase 1 was with 28 cows, and phase 2 with 21 cows. Cows were fed a basal total mixed ration ad libitum and were blocked based on days in milk, milk yield, and enteric methane emissions during a 14-d covariate period. Treatments were control (no 3-NOP) and 40, 60, 80, 100, 150, and 200 mg of 3-NOP/kg of feed dry matter. Following a 14-d adaptation period, enteric gaseous emissions (methane, carbon dioxide, and hydrogen) were measured using the GreenFeed system (C-Lock Inc., Rapid City, SD) over a 3-d period. Compared with the control, inclusion rate of 3-NOP quadratically decreased daily enteric methane emissions from 22 to 40%. Maximum mitigation effect was achieved with the 3 highest 3-NOP doses (with no statistical difference among 100, 150, and 200 mg/ kg). The decrease in methane emission yield and emission intensity ranged from 16 to 36% and from 25 to 45%, respectively. Emissions of hydrogen quadratically increased 6-to 10-fold, compared with the control; the maximum increase was with 150 mg/kg 3-NOP. Treatment did not affect daily emissions of carbon dioxide, but a linear increase in carbon dioxide emission yield was observed with increasing 3-NOP doses. Dry matter intake and milk yield of the cows was not affected by 3-NOP. Milk fat concentration and yield were increased by 3-NOP due to increased concentration of de novo synthetized short-chain fatty acids in milk. Inclusion of 3-NOP also tended to increase milk urea nitrogen but had no other effects on milk components. In this shortterm experiment, 3-NOP decreased enteric methane emissions without affecting dry matter intake or milk yield and increased milk fat in dairy cows. Maximum mitigation effect was achieved at 100 to 200 mg/kg of feed dry matter.

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The effect of type of grain (corn vs. barley) and amylopectin content of barley grain (normal vs. waxy) on ruminal fermentation, digestibility, and utilization of ruminal ammonia nitrogen for milk protein synthesis was studied in a replicated 3 x 3 Latin square design trial with 6 lactating dairy cows. The experimental treatments were (proportion of dietary dry matter): CORN, 40% corn grain, NBAR, 30% normal Baronesse barley:10% corn grain, and WBAR, 30% high-amylopectin (waxy) Baronesse barley:10% corn grain. All grains were steam-rolled and fed as part of a total mixed ration. The NBAR and WBAR diets resulted in increased ruminal ammonia concentrations compared with CORN (8.2, 7.4, and 5.6 mM, respectively), but other ruminal fermentation parameters were not affected. Ruminal digestibility of dietary nutrients and microbial protein synthesis in the rumen were also not affected by diet. Corn grain had greater in situ effective ruminal dry matter degradability (62.8%) than the barley grains (58.2 and 50.7%, respectively), and degradability of the normal barley starch was greater than that of the waxy barley (69.3 and 58.9%, respectively). A greater percentage of relative starch crystallinity was observed for the waxy compared with the normal barley grain. Total tract apparent digestibility of dry matter and organic matter were decreased by WBAR compared with CORN and NBAR. Total tract starch digestibility was greater and milk urea nitrogen content was lower for CORN compared with the 2 barley diets. In this study, the extent of processing of the grain component of the diet was most likely the factor that determined the diet responses. Minimal processing of barley grain (processing indexes of 79.2 to 87.9%) reduced its total tract digestibility of starch compared with steam-rolled corn (processing index of 58.8%). As a result of the increased ammonia concentration and reduced degradability of barley dry matter in the rumen, the utilization of ruminal ammonia nitrogen for microbial protein synthesis was decreased with the barley diets compared with the corn-based diet. In this study, waxy Baronesse barley was less degradable in the rumen and the total digestive tract than its normal counterpart. The most likely reasons for these effects were the differences in starch characteristics and chemical composition, and perhaps the different response to processing between the 2 barleys.

The objective of this experiment was to compare ruminal fluid samples collected through rumen cannula (RC) or using an oral stomach tube (ST) for measurement of ruminal fermentation and microbiota variables. Six ruminally cannulated lactating Holstein cows fed a standard diet were used in the study. Rumen samples were collected at 0, 2, 4, 6, 8, and 12 h after the morning feeding on two consecutive days using both RC and ST techniques. Samples were filtered through two layers of cheesecloth and the filtered ruminal fluid was used for further analysis. Compared with RC, ST samples had 7% greater pH; however, the pattern in pH change after feeding was similar between sampling methods. Total volatile fatty acids (VFA), acetate and propionate concentrations in ruminal fluid were on average 23% lower for ST compared with RC. There were no differences between RC and ST in VFA molar proportions (except for isobutyrate), ammonia and dissolved hydrogen (dH2) concentrations, or total protozoa counts, and there were no interactions between sampling technique and time of sampling. Bacterial ASV richness was higher in ST compared with RC samples; however, no differences were observed for Shannon diversity. Based on Permanova analysis, bacterial community composition was influenced by sampling method and there was an interaction between sampling method and time of sampling. A core microbiota comprised of Prevotella, S24-7, unclassified Bacteroidales and unclassified Clostridiales, Butyrivibrio, unclassified Lachnospiraceae, unclassified Ruminococcaceae, Ruminococcus, and Sharpea was present in both ST and RC samples, although their relative abundance varied and was influenced by an interaction between sampling time and sampling method. Overall, our results suggest that ruminal fluid samples collected using ST (at 180 to 200 cm depth) are not representative of rumen pH, absolute values of VFA concentrations, or bacterial communities &gt;2 h post-feeding when compared to samples of ruminal fluid collected using RC. However, ST can be a feasible sampling technique if the purpose is to study molar proportions of VFA, protozoa counts, dH2, and ammonia concentrations.