Pharmacokinetics and bioavailability after intramuscular injection of the 5‐HT1A serotonin agonist R‐8‐hydroxy‐2‐(di‐n‐propylamino) tetralin (8‐OH‐DPAT) in domestic goats (Capra aegagrus hircus)

To determine the bioavailability and pharmacokinetic properties of the serotonin 5‐HT_1A receptor agonist R‐8‐OH‐DPAT in goats, and 0.1 mg kg^?1 R‐8‐OH‐DPAT hydrobromide was administered intramuscularly (i.m.) and intravenously (i.v.) to six goats in a two‐phase cross‐over design experiment. Venous blood samples were collected from the jugular vein 2, 5, 10, 15, 20, 30, 40 and 60 min following treatment and analysed by liquid chromatography tandem mass spectrometry. Bioavailability and pharmacokinetic parameters were determined by a one‐compartment analysis. Mean bioavailability of R‐8‐OH‐DPAT when injected i.m. was 66%. The mean volume of distribution in the central compartment was 1.47 L kg^?1. The mean plasma body clearance was 0.056 L kg^?1 min^?1. All goats injected i.v. and two of six goats injected i.m. showed signs of serotonin toxicity. In conclusion, R‐8‐OH‐DPAT is well absorbed following i.m. injection and the observed pharmacokinetics suggest that administration via dart is feasible. Administration of R‐8‐OH‐DPAT hydrobromide, at a dosage of 0.1 mg kg^?1, resulted in the observation of clinical signs of serotonin toxicity in the goats. It is suggested that dosages for the clinical use of the compound should be lower in order to achieve the desired clinical effect without causing serotonin toxicity.     

Effects of maternal treatment with β‐hydroxy‐β‐metylbutyrate and 2‐oxoglutaric acid on femur development in offspring of minks of the standard dark brown type

The aim of the study was to evaluate the effect of the diet, mother type and sex of the offspring on the mechanical and geometric parameters of long bones as well as bone tissue density in minks. Primiparous and multiparous dams were supplemented with β‐hydroxy β‐methylbutyrate (a metabolite of leucine, at the daily dosage of 0.02 g/kg of body weight) and/or 2‐oxoglutaric acid (a precursor of glutamine, at the daily dosage of 0.4 g/kg of body weight) during gestation. The diet did not influence bone tissue density and the length of the humerus. An increase in the length of the femur was noted in male offspring delivered by multiparous dams. The diet resulted in an increase in the weight of the humerus in males from multiparous dams and a decrease in offspring from primiparous dams. Heavier femora were noted in male offspring delivered by both types of dams. The maximum elastic strength of the humerus was higher in the offspring delivered by multiparous than primiparous dams, irrespective of the offspring sex. The diet resulted in reduction in the ultimate strength of the femur in the male offspring delivered by primiparous dams. Only females born by multiparous dams, irrespective of the diet, showed a significant increase in the cross‐sectional area of the humerus, while a significant decline was noted in males delivered by multiparous dams and in all the offspring delivered by primiparous dams. An increase in the cross‐sectional area of the femur was noted in the offspring delivered by multiparous dams, while reduction was observed in the offspring delivered by primiparous dams. These results have shown for the first time that the presence of β‐hydroxy‐β‐methylbutyrate or 2‐oxoglutaric acid in the diet of pregnant primiparous or multiparous dams unambiguously affects the geometry and mechanical properties of offspring's long bones.     

Effects of β‐carotene‐enriched dry carrots on β‐carotene status and colostral immunoglobulin in β‐carotene‐deficient Japanese Black cows

Data from 18 β‐carotene‐deficient Japanese Black cows were collected to clarify the effects of feeding β‐carotene‐enriched dry carrots on β‐carotene status and colostral immunoglobulin (Ig) in cows. Cows were assigned to control or carrot groups from 3 weeks before the expected calving date to parturition, and supplemental β‐carotene from dry carrots was 138 mg/day in the carrot group. Plasma β‐carotene concentrations in the control and carrot groups at parturition were 95 and 120 μg/dL, and feeding dry carrots slightly improved plasma β‐carotene at parturition. Feeding dry carrots increased colostral IgA concentrations in cows and tended to increase colostral IgG₁, but colostral IgM, IgG₂, β‐carotene and vitamin A were not affected by the treatment. Feeding dry carrots had no effects on plasma IgG₁, IgA and IgM concentrations in cows, but plasma IgG₁ concentrations decreased rapidly from 3 weeks before the expected calving date to parturition. These results indicate that feeding β‐carotene‐enriched dry carrots is effective to enhance colostral IgA and IgG₁ concentrations in β‐carotene‐deficient cows.     

Non‐phytate phosphorus requirements and efficacy of a genetically engineered yeast phytase in male lingnan yellow broilers from 1 to 21 days of age

This experiment was conducted to investigate the requirement of non‐phytate phosphorus (nPP) and efficacy of a genetically engineered yeast phytase in performance and tibia characteristics by male Lingnan Yellow broilers from 1 to 21 days of age. A total of 2640 1‐day‐old male chicks were randomly allotted to one of 11 dietary treatments, which consisted of six replicate floor pens with 40 birds per pen. All treatments had the same levels of all nutrients except for phosphorus and phytase. The control group (treatment 1) was fed the basal diet without dicalcium phosphate or phytase supplementation. Dietary concentrations of nPP were 0.11%, 0.19%, 0.27%, 0.35%, 0.43%, 0.51% and 0.59% respectively for treatments 1, 2, 3, 4, 5, 6 and 7, through addition of dicalcium phosphate (chemistry grade) to the basal diet. Diets 8–11 were supplemented with a genetically engineered yeast phytase 250, 500, 750 U/kg and a commercial phytase product 500 U/kg in basal diet respectively. The results showed that 0.46% and 0.51% nPP were required for maximum body‐weight gain and optimum tibia development indicators respectively. However, 0.59% nPP had a negative effect on bird growth. The equivalency value of the genetically engineered yeast phytase was estimated to be 874 U/kg to liberate 0.1% nPP.     

Effects of supplementation with β‐carotene on the growth performance and intestinal mucosal barriers in layer‐type cockerels

β‐carotene is a robust modulator of mucosal barriers, and it can amplify the immunoglobulin A (IgA) response via the retinoic acid (RA)–mediated pathway. We investigated the influence of β‐carotene on intestinal barriers in layer‐type cockerels. In this study, β‐carotene has a positive influence on growth performance and intestinal morphology. β‐carotene remarkably enhanced serum secretory immunoglobulin A (sIgA) levels, jejunal mucosal sIgA, and IgA concentrations. β‐Carotene significantly enhanced mRNA expression levels of IgA, CC chemokine receptor‐9 (CCR9), polymeric immunoglobulin receptor (pIgR), and retinoic acid receptor α (RARα) in the ileal tissues and pIgR in the jejunal tissues. β‐Carotene improves mRNA expression of intestinal barrier‐related proteins including: mucin‐2 (MUC‐2), zonula occludens‐2 (ZO‐2), occludins (OCLN), and zonula occludens‐1 (ZO‐1) in the ileal tissues. Moreover, β‐carotene decreased the levels of Escherichia coli and elevates the levels of Lactobacillus. The results indicate that β‐carotene can promote growth performance and contribute to the gradual development of intestinal barriers in Hyline Brown chicks. This study enriches our knowledge about the effects of β‐carotene on intestinal barrier and highlights a theoretical basis of β‐carotene application in the poultry industry.     

Effects of supplemental β‐carotene on colostral immunoglobulin and plasma β‐carotene and immunoglobulin in Japanese Black cows

Data from 26 Japanese Black cows were collected to clarify the effects of supplemental β‐carotene on colostral immunoglobulin (Ig) and plasma β‐carotene and Ig in the cows. Cows were assigned to control or β‐carotene groups from 21 days before the expected calving date to 60 days after parturition. Supplemental β‐carotene was provided at 500 mg/day in the β‐carotene group. Supplemental β‐carotene drastically increased plasma β‐carotene concentrations in the cows from parturition to 60 days after parturition, and plasma β‐carotene concentrations in the control and β‐carotene groups at parturition were 202 and 452 μg/dl, respectively. Supplemental β‐carotene had no effects on plasma IgG₁, IgA or IgM concentrations at parturition. Supplemental β‐carotene increased colostral IgG₁ concentrations in the cows, but colostral β‐carotene, IgA and IgM concentrations were not affected by supplemental β‐carotene. These results indicate that supplemental β‐carotene is effective to enhance colostral IgG₁ concentrations and plasma β‐carotene concentrations in Japanese Black cows.     

Effects of β‐hydroxybutyricacid on the synthesis and assembly of very low‐density lipoprotein in bovine hepatocytes in vitro

β‐Hydroxybutyricacid (BHBA) is an important metabolite that involved in the development of ketosis and fatty liver in dairy cows. Dairy cows with fatty liver displayed high blood concentration of BHBA and very low‐density lipoprotein (VLDL) assembly. The effects of BHBA on VLDL synthesis and assembly in hepatocytes of cows were unclear. In this study, bovine hepatocytes were cultured and treated with different concentrations of BHBA. We found that BHBA treatment upregulated the mRNA and protein levels of apolipoprotein B100 (ApoB 100), apolipoprotein E (ApoE) and microsomal triglyceride transfer protein (MTTP) and showed in a firstly increased and then decreased trend. Meanwhile, the mRNA and protein levels of LDLR showed in a reverse trend. Consequently, VLDL content was significantly increased in medium‐dose BHBA treatment group, while decreased in high‐dose group. These results indicate that the effects of BHBA on the VLDL synthesis showed in a dose‐dependent manner that low levels of BHBA increase VLDL synthesis and high levels of BHBA decrease VLDL synthesis.     

Effects of (−)‐hydroxycitric acid on lipid droplet accumulation in chicken embryos

This study was conducted to determine the impact of (?)‐hydroxycitric acid ((?)‐HCA) on biochemical indices and lipid metabolism parameters in chicken embryos. Two hundred and forty fertilized eggs were divided into six groups and injected with (?)‐HCA at concentrations of 0, 0.1, 0.5, 1.0, 10.0 and 50 mg/kg (n = 40). After 19 days of incubation, serum and liver were collected for analysis of biochemical indices and lipid metabolism parameters. Results showed no significant differences on serum biochemical indices: 1–50 mg/kg (?)‐HCA significantly increased serum glucose and hepatic glycogen contents (P < 0.05). Oil Red O staining analysis showed total area, counts of lipid droplets and hepatic triglyceride content were significantly decreased (P < 0.01), meanwhile hepatic lipase and lipoprotein lipase activity were significantly increased (P < 0.05). ACLY, ME1, SREBP‐1c messenger RNA (mRNA) levels in 0.5–10 mg/kg groups and FAS mRNA level in 1–10 mg/kg groups were significantly decreased (P < 0.05), while PPARα mRNA level, serum adiponectin content and AdipoR1 mRNA level were significantly increased in 0.5–50 mg/kg groups (P < 0.05). These results indicated (?)‐HCA treatment inhibited triglyceride synthesis via decreasing lipogenesis‐related factors, mRNA expression level and accelerated lipolysis by enhancing lipoprotein lipase and hepatic lipase activity, which finally reduced lipid droplet accumulation, and this action may be associated with activating the adiponectin signaling pathway.     

Effects of a novel missense polymorphism within the SIGLEC5 gene on fertility traits in Holstein‐Friesian cattle

The aim of the study was to find functional polymorphism within two exons of the SIGLEC5 (sialic acid‐binding Ig‐like lectin‐5) gene and to examine its effects on the production and fertility traits of cows and bulls. Two hundred seventytwo Holstein‐Friesian cows and 574 bulls were included in the study. Novel missense polymorphism (A > G) within exon 3 causing substitution of amino acid arginine by glutamate in position 260 of SIGLEC5 protein (R260Q) was identified by sequencing and digestion by restriction enzyme Msp I. Basic production and fertility traits of cows and estimated breeding values (EBV) of bulls were analysed. The study demonstrated a significant association of SIGLEC5 R260Q polymorphism with days open and calving interval in cows as well as with breeding value for calving interval in bulls. An opposite effect of SIGLEC5 alleles for production and fertility traits was observed: the allele G increased the breeding value for the protein yield, while the allele A increased the breeding value for the calving interval. The current study suggests the involvement of SIGLEC5 R260Q polymorphism in biological processes related to fertility traits. This finding can be applied as a biomarker for a genetic improvement programme in Holstein‐Friesian cattle.     

Effects of concentrate‐to‐forage ratios and 2‐methylbutyrate supplementation on ruminal fermentation,bacteria abundance and urinary excretion of purine derivatives in Chinese Simmental steers

This study evaluated the effects of dietary concentrate levels and 2‐methylbutyrate (2MB ) supplementation on performance, ruminal fermentation, bacteria abundance, microbial enzyme activity and urinary excretion of purine derivatives (PD ) in steers. Eight ruminally cannulated Simmental steers (12 months of age; 389 ± 3.7 kg of body weight) were used in a replicated 4 × 4 Latin square design with a 2 × 2 factorial arrangement. Moderate‐concentrate (400 g/kg diet [MC ]) or high‐concentrate (600 g/kg diet [HC ]) diets were fed with or without 2MB (0 g/day [2MB ?] or 15.0 g/day [2MB +]). Dry matter intake and average daily gain increased, but feed conversion ratio decreased with the HC diet or 2MB supplementation. Ruminal pH decreased, but total volatile fatty acid increased with the HC diet or 2MB supplementation. Molar proportion of acetate and acetate‐to‐propionate ratio decreased with the HC diet, but increased with 2MB supplementation. Propionate molar proportion and ruminal NH ₃‐N content increased with the HC diet, but decreased with 2MB supplementation. Neutral detergent fibre degradability decreased with the HC diet, but increased with 2MB supplementation. Crude protein degradability increased with the HC diet or 2MB supplementation. Abundance of Ruminococcus albus , Ruminococcus flavefaciens , Fibrobacter succinogenes and Bufyrivibrio fibrisolvens as well as activities of carboxymethyl cellulase, cellobiase, xylanase and pectinase decreased with the HC diet, but increased with 2MB supplementation. However, abundance of Prevotella ruminicola and Ruminobacter amylophilus as well as activities of α‐amylase and protease increased with the HC diet or 2MB supplementation. Total PD excretion also increased with the HC diet or 2MB supplementation. The results suggested that growth performance, ruminal fermentation, CP degradability and total PD excretion increased with increasing dietary concentrate level from 40% to 60% or 2MB supplementation. The observed diet × 2MB interaction indicated that supplementation of 2MB was more efficacious for improving growth performance, ruminal fermentation and total PD excretion with promoted ruminal bacteria abundance and enzyme activity in the MC diet than in the HC diet.     

Effects of transforming growth factor‐β1 treatment on muscle regeneration and adipogenesis in glycerol‐injured muscle

Transforming growth factor (TGF)‐β1 is associated with fibrosis in many organs. Recent studies demonstrated that delivery of TGF‐β1 into chemically injured muscle enhances fibrosis. In this study, we investigated the effects of exogenous TGF‐β1 on muscle regeneration and adipogenesis in glycerol‐injured muscle of normal mice. Tibialis anterior (TA) muscles were injured by glycerol injection. TGF‐β1 was either co‐injected with glycerol, as an ‘early treatment’ group, or injected at day 4 after glycerol, as a ‘late treatment’ group and the TA muscles were collected at day 7 after initial injury. Myotube density was significantly lower in the early treatment group than in the glycerol‐injured group (without TGF‐β1 treatment). Moreover, the Oil red O‐positive area was significantly smaller in the early treatment group than in the late treatment group and glycerol‐injured group. Furthermore, TGF‐β1 treatment increased endomysial fibrosis and induced immunostaining of α‐smooth muscle actin. The greater inhibitory effects of early TGF‐β1 treatment than that of late TGF‐β1 treatment during regeneration in glycerol‐injured muscle suggest a more potent effect of TGF‐β1 on the initial stage of muscle regeneration and adipogenesis. Combination of TGF‐β1 with glycerol might be an alternative to enhance muscle fibrosis for future studies.     

Effects of exposure to artificial long days on milk yield,maternal insulin‐like growth factor 1 levels and kid growth rate in subtropical goats

This study was designed to determine whether any relationship exists between exposure to artificial long days, milk yield, maternal plasma insulin‐like growth factor 1 (IGF‐1) levels, and kid growth rate in goats. One group of lactating goats was maintained under naturally decreasing day length (control group; n = 19), while in another one, they were kept under artificial long days (LD group; n = 19). Milk yield was higher in goats from the LD group than that in the control group (P < 0.05). Maternal IGF‐1 levels at day 57 of lactation were higher (P < 0.05) in goats from the LD group than the levels in the control group and were positively correlated with the total milk yields per goat at days 43 and 57 of lactation (r = 0.77 and r = 0.84, respectively; P < 0.01). Daily weight gain at week 4 was higher (P < 0.01) in kids from the LD group than that in kids from the control group and was correlated with total and average IGF‐1 maternal levels (r = 0.60 and r = 0.60, P < 0.05). It was concluded that submitting lactating goats to artificial long days increases milk yield, plasma IGF‐1 maternal levels and the growth rate of the kids.     

Effects of 25‐hydroxycholecalciferol with two D3 vitamin levels on production and immunity parameters in broiler chickens

This study was performed in Ross 308 chickens aged 1–21 days and aimed to evaluate whether the addition of 25‐hydroxycholecalciferol (25(OH)D₃) to broiler chicken diets affects their growth performance and immunity. A completely random 2 × 2 factorial arrangement was used with two levels of vitamin D₃ and the absence or presence of 25(OH)D₃, corresponding to four treatments based on sorghum + soya bean diets: (i) 200 IU of vitamin D₃/kg of feed (Diet 1) (NRC, 1994 ), (ii) Diet 1 + 69 μg of 25(OH)D₃/kg of feed (Diet 2), (iii) 5,000 IU of vitamin D₃/kg of feed (Diet 3) and (iv) Diet 3 + 69 μg of 25(OH)D₃/kg of feed (Diet 4). Each treatment was conducted with six replicates of 10 chickens each. Water and feed was supplied ad libitum. The results showed significantly increased growth and tibia ash (p < .05) in the birds fed 5,000, IU of vitamin D₃/kg + 25(OH)D₃. Additionally, the cellular immune response increased significantly (p < .05) in both treatments with added 25(OH)D_3. Based on the results obtained under the current test conditions, the addition of 25(OH)D₃ at a rate of 69 μg/kg to diets containing vitamin D₃ improved the cellular immune response and mineral deposition in the bones of broilers aged 1–21 days. Because these parameters are very important in modern poultry farming, these results indicate that supplementation with 25(OH)D₃ should improve broiler production.     

Effects of Wnt10b on dermal papilla cells via the canonical Wnt/β‐catenin signalling pathway in the Angora rabbit

Wnt10b is a member of Wnt family that plays a variety of roles in biological functions, including those in the development of hair follicles. To investigate the effect of Wnt10b on hair growth in the Angora rabbit and to determine the underlying molecular mechanism, we cultured dermal papilla (DP) cells with exogenous Wnt10b in vitro. We observed the expressions of downstream critical gene β‐catenin and lymphoid enhancer‐binding factor 1 (LEF1) in Wnt/β‐catenin pathway. The levels of β‐catenin mRNA and protein were higher in the Wnt10b group of DP cells than in the Control group, and the mRNA level of LEF1 in the Wnt10b group was higher than in the Control group. Moreover, translocation of β‐catenin from cytoplasm to nucleus was activated in the Wnt10b group. Furthermore, the mRNA levels of the hair follicle‐regulatory genes, insulin‐like growth factor‐1 (IGF‐1) and alkaline phosphatase (ALP), and the protein activity of ALP was also upregulated in the Wnt10b group compared to their corresponding levels in the Control group. These data suggest that Wnt10b could activate the canonical Wnt/β‐catenin signalling pathway to induce DP cells in the Angora rabbit. In addition, the proliferation of DP cells was significantly promoted when cultured with Wnt10b for 48 and 72 hr, suggesting that Wnt10b plays a pivotal role in the proliferation and maintenance of DP cells in vitro. In conclusion, this study demonstrates that Wnt10b may promote hair follicle growth in Angora rabbit through the canonical Wnt/β‐catenin signalling pathway that promotes the proliferation of DP cells.     

Effects of 25‐hydroxycholecalciferol supplementation in maternal diets on milk quality and serum bone status markers of sows and bone quality of piglets

Twenty primiparous sows were allocated to two treatments to evaluate the effects of maternal 25‐hydroxycholecalciferol (25OHD₃) supplementation during gestation and lactation on milk quality and serum bone status markers of sows and bone quality of piglets. Immediately after mating, sows were randomly allotted to one of two diets supplemented with 50 µg/kg 25OHD₃ or basal diets without 25OHD₃. Blood and milk samples were obtained. At birth and weaning, 10 piglets from each treatment were killed for bone quality analysis. 25OHD₃‐fed sows provided one more piglet at farrowing and 1.17 more piglets at weaning than sows fed basal diets. The contents of solids not‐fat, protein, fat or lactose were increased in milk from days 7 and 14 of lactation in 25OHD₃‐supplemented sows and 25OHD₃ concentrations in milk were increased by dietary 25OHD₃ supplementation. Dietary 25OHD₃ supplementation increased serum alkaline phosphatase activity but had no effect on serum tartrate‐resistant acid phosphatase activity of sows. Maternal 25OHD₃ supplementation improved bone strength, density and ash content of newborn piglets rather than those of weaning piglets. In conclusion, 25OHD₃ supplementation in maternal diets improved reproductive performance, milk quality and bone status of sows as well as bone quality of newborn piglets.     

Effects of calcium soap of rice bran oil fatty acids supplementation alone and with DL‐α‐tocopherol acetate in lamb diets on performance,digestibility, ruminal parameters and meat quality

Thirty‐six Malpura lambs (28 day old and 6.7 ± 0.25 kg BW) were distributed equally in three groups having six males and six female. They were ad libitum fed individually three different experimental diets containing calcium soap of fatty acids (CA‐FA) at 0 (T₁) and 40 (T₂ and T₃) g/kg concentrate up to six months of age. Animals in T₃ were supplemented additionally with 40 mg DL‐α‐tocopherol acetate/kg of concentrate. The roughage moiety included ad libitum dry Prosopis cineraria and fresh Azadirachata indica leaves. All the lambs were allowed to suckle from their dam up to weaning (90 day of age). Supplementation of Ca‐FA improved weight gain and feed conversion ratio during both pre‐ (28–90 days) and post‐weaning (91–180 days) phases; however, no effect of DL‐α‐tocopherol was observed. Metabolic parameters during post‐weaning phase revealed non‐significant effect on digestibility but improved nitrogen balance in the test groups. The effect on biochemical attributes did not show any significant alteration in ruminal parameters, blood biochemicals and urinary purine derivatives. Carcass traits revealed higher (p < 0.05) dressing yield and loin eye area with Ca‐FA supplementation. The value of thiobarbituric reactive substances for nuggets prepared from frozen carcasses revealed significant (p < 0.05) reduction in T₃ compared to the other dietary groups. Fatty acid profile of adipose tissue revealed higher (p < 0.001) 9‐octadecanoic, 9‐12‐octadecadienoic, polyunsaturated fatty acids (PUFA), higher ratio of PUFA/saturated fatty acids (SFA), ω‐6/ω‐3 and lower SFA in Ca‐FA‐supplemented groups. It is concluded that supplementation of 40 g/kg calcium soap prepared from industrial grade rice bran oil in lamb ration provided additional energy intake, improved N utilization, gain and feed conversion ratio besides improving dressing yield and meat quality with CLA enriched fatty acid profile. DL‐α‐tocopherol acetate when supplemented at 40 mg/kg feed reduced lipid oxidation of meat products thus improving its keeping quality.     

Effects of dietary protein levels and 2‐methylbutyrate on ruminal fermentation,nutrient degradability,bacterial populations and urinary purine derivatives in Simmental steers

The objective of this study was to evaluate the effects of dietary crude protein (CP ) levels and 2‐methylbutyrate (MB ) supplementation on ruminal fermentation, bacterial populations, microbial enzyme activity and urinary excretion of purine derivatives (PD ) in Simmental steers. Eight ruminally cannulated Simmental steers, averaging 18 months of age and 465 ± 8.6 kg of body weight (BW ), were used in a replicated 4 × 4 Latin square design by a 2 × 2 factorial arrangement. Low protein (98.5 g CP /kg dry matter [LP ] or high protein (128.7 g CP /kg dry matter [HP ]) diets were fed with MB supplementation (0 g [MB ?] or 16.8 g steer^?1 day^?1 [MB +]). Steers were fed a total mixed ration with dietary corn straw to concentrate ratio of 50:50 (dry matter [DM ] basis). The CP  × MB interaction was observed for ruminal total VFA , molar proportions of acetate and propionate, acetate to propionate ratio, ammonia‐N, effective degradability of neutral detergent fibre (NDF ) and CP , microbial enzyme activity, bacterial populations and total PD excretion (p  < .05). Ruminal pH decreased (p  < .05), but ruminal total VFA concentration increased (p  < .05) with increasing dietary CP level or MB supplementation. Acetate molar proportion increased (p  = .043) with MB supplementation, but was not affected by dietary CP level. Propionate molar proportion decreased (p  < .05) with increasing dietary CP level or MB supplementation. Consequently, acetate‐to‐propionate ratio increased (p  = .001) with MB supplementation, but was not affected by dietary CP level. Ruminal ammonia‐N content increased (p  = .034) with increasing dietary CP level, but decreased (p  = .012) with MB supplementation. The effective degradability of NDF and CP increased (p  < .05) with increasing dietary CP level or MB supplementation. Microbial enzyme activity, bacterial populations and total PD excretion also increased (p  < .05) with increasing dietary CP level or MB supplementation. The results indicated that ruminal fermentation, nutrient degradability, microbial enzyme activity, ruminal bacterial populations and microbial protein synthesis improved with increasing dietary CP level or MB supplementation in steers.