Evan C. Titgemeyer

Fermentation of various dietary fiber sources by human fecal bacteria

Abstract Eleven fiber-rich substrates were subjected to in vitro incubation with fecal bacteria from each of three adult male volunteers to assess substrate organic matter disappearance (OMD), short-chain fatty acid (SCFA) production, and the potential water holding capacity (PWHC) of fermented residues. Substrate OMD ranged from 83.3% (citrus pectin) to 5.8% (oat fiber). Total SCFA production (mmol/g substrate OM) was highly correlated to OMD and ranged from 9.64 (gum arabic) to 0.48 (oat fiber). Molar ratios of acetate:propionate produced during fermentation varied considerably among substrates, ranging from 1.5 for guar gum to 12.6 for citrus pectin. PWHC, an estimate of potential fecal bulking effects, was negatively related to OMD (r = −0.88) and ranged from 4.47 (oat fiber) to 0.71 (citrus pectin) g H 2 O/g substrate dry matter. In vitro fermentation techniques can be potentially useful predictors of in vivo responses to dietary fiber sources.

Effect of rumen-degradable intake protein supplementation on urea kinetics and microbial use of recycled urea in steers consuming low-quality forage

We evaluated the effect of increasing amounts of rumen-degradable intake protein (DIP) on urea kinetics in steers consuming prairie hay. Ruminally and duodenally fistulated steers (278 kg of BW) were used in a 4 x 4 Latin square and provided ad libitum access to low-quality prairie hay (4.9% CP). The DIP was provided as casein dosed ruminally once daily in amounts of 0, 59, 118, and 177 mg of N/kg of BW daily. Periods were 13 d long, with 7 d for adaptation and 6 d for collection. Steers were in metabolism crates for total collection of urine and feces. Jugular infusion of (15)N(15)N-urea, followed by determination of urinary enrichment of (15)N(15)N-urea and (14)N(15)N-urea was used to determine urea kinetics. Forage and N intake increased (linear, P < 0.001) with increasing DIP. Retention of N was negative (-2.7 g/d) for steers receiving no DIP and increased linearly (P < 0.001; 11.7, 23.0, and 35.2 g/d for 59, 118, and 177 mg of N/kg of BW daily) with DIP. Urea synthesis was 19.9, 24.8, 42.9, and 50.9 g of urea-N/d for 0, 59, 118, and 177 mg of N/kg of BW daily (linear, P = 0.004). Entry of urea into the gut was 98.9, 98.8, 98.6, and 95.9% of production for 0, 59, 118, and 177 mg of N/kg of BW daily, respectively (quadratic, P = 0.003). The amount of urea-N entering the gastrointestinal tract was greatest for 177 mg of N/kg of BW daily (48.6 g of urea-N/d) and decreased (linear, P = 0.005) to 42.4, 24.5, and 19.8 g of urea-N/d for 118, 59, and 0 mg of N/kg of BW daily. Microbial incorporation of recycled urea-N increased linearly (P = 0.02) from 12.3 g of N/d for 0 mg of N/kg of BW daily to 28.9 g of N/d for 177 mg of N/kg of BW daily. Provision of DIP produced the desired and previously observed increase in forage intake while also increasing N retention. The large percentage of urea synthesis that was recycled to the gut (95.9% even when steers received the greatest amount of DIP) points to the remarkable ability of cattle to conserve N when fed a low-protein diet.

Effects of encapsulated niacin on metabolism and production of periparturient dairy cows

Nicotinic acid (niacin) can suppress lipolysis, but responses to dietary niacin have been inconsistent in cattle. Our aim was to determine if 24 g/d of encapsulated niacin (EN; providing 9.6g/d of bioavailable nicotinic acid) alters lipid metabolism and productivity of transition cows. Beginning 21 d before expected calving, primiparous (n = 9) and multiparous (n = 13) cows (body condition score of 3.63 ± 0.08) were sequentially assigned within parity to EN (12 g provided with ration twice daily) or control through 21 d postpartum. Liver biopsies were collected on d -21, -4, 1, 7, and 21 relative to parturition. Blood samples were collected on d -21, -14, -7, -4, 1, 4, 7, 14, and 21 relative to parturition. On d 7 postpartum, a caffeine clearance test was performed to assess liver function, and on d 21 to 23 postpartum, blood samples were collected every 8h to monitor posttreatment nonesterified fatty acid (NEFA) responses. Data were analyzed using mixed models with repeated measures over time. A treatment × time × parity effect was observed on prepartum dry matter intake (DMI), which was caused by a 4 kg/d decrease in DMI of EN-treated multiparous cows compared with control multiparous cows during the final 4 d prepartum. A significant increase in plasma nicotinamide concentration occurred in EN-treated cows on d -7 and 21 relative to parturition. Prepartum glucose concentration decreased in treated animals, with no difference in plasma insulin concentration. Treatment × time × parity effects were detected for NEFA and β-hydroxybutyrate concentrations during the postpartum period. Plasma NEFA peaked at 1,467 ± 160 μM for control animals compared with 835 ± 154 μM for EN-treated animals. After treatments ended on d 21, no evidence was found for a plasma NEFA rebound in either parity group. A treatment × parity × time interaction was detected for liver triglyceride content, indicating a tendency for less liver triglyceride in EN-treated primiparous cows, but caffeine clearance rates were not affected by treatment. No treatment effects were observed for body condition score, body weight, energy balance, or milk or milk component production. A high dose of EN can decrease postpartum plasma NEFA concentration, but may also decrease prepartum DMI.

Effect of a wide range in the ratio of supplemental rumen degradable protein to starch on utilization of low-quality, grass hay by beef steers

Hereford×Angus steers were used in a 14-treatment, 2-period, crossover design experiment to examine effects of a wide range in the ratio of supplemental starch to rumen degradable protein (RDP) on low-quality forage utilization and ruminal characteristics. Steers were given ad libitum access to grass hay (4.9% CP, 42.4% ruminally degradable) and supplemented in a 2×7 factorial arrangement of treatments. All supplements were administered directly into the rumen and delivered one of two levels of ruminally degradable starch (cornstarch grits; 0 and 0.3% of initial BW) and one of seven levels of RDP (sodium caseinate; 0, 0.015, 0.051, 0.087, 0.123, 0.159, and 0.195% of initial BW). Supplementation with RDP increased consumption of forage OM, total OM, NDF, and digestible OM in a quadratic (P<0.01) fashion (intake increased and then declined). Starch supplementation depressed (P<0.01) forage OM and NDF intakes. In general, RDP supplementation elicited a positive quadratic response on NDF digestion (P=0.02). However, an interaction between supplemental starch and RDP level was observed (P<0.01) for NDF digestion. At the four lowest levels of supplemental RDP, starch supplementation substantially reduced NDF digestion, although for steers receiving the three highest levels of supplemental RDP, starch supplementation had little effect on NDF digestion. In contrast, neither the starch×RDP interaction nor the starch main effect was significant for OM digestion, which increased linearly (P<0.01) with supplemental RDP. Supplementation with RDP altered passage rate of acid detergent insoluble ash in a quadratic (P=0.05) manner that paralleled the intake response. However, liquid passage rate was not affected significantly. A decline in ruminal pH was associated (P=0.02) with increasing supplemental RDP and tended (P=0.07) to be associated with increasing starch, but it was not requisite for starch-induced depressions in NDF digestion. Ruminal NH3 concentration increased in response to increasing RDP, although the increase when starch was supplemented was less than that observed without starch (P=0.03). Supplemental starch generally elicited negative effects on low-quality forage intake and fiber digestion, but the effects on fiber digestion were overridden by adequate supplemental RDP. Supplemental RDP exerted a highly positive effect on consumption and digestion of this low-quality forage.

Fermentation of Dietary Fibre by Human Colonic Bacteria: Disappearance of, Short-Chain Fatty Acid Production from, and Potential Water-Holding Capacity of, Various Substrates

Several dietary fibre-rich substrates were fermented in vitro with human colonic bacteria obtained from each of three adult male subjects to assess the extent of substrate fermentation short-chain fatty acid (SCFA) production, and the potential effect of fermented residues on faecal bulk. Substrates tested were two varieties of oat hull fibre, gum arabic, carboxymethylcellulose (CMC), soy fibre, psyllium, and six blends containing oat fibre, gum arabic, and CMC in various proportions. All substrates contained greater than 900 g/kg of total dietary fibre except for CMC (816 g) and soy fibre (778 g). In vitro organic matter disappearance during fermentation was greatest for gum arabic (69.5%), intermediate for soy fibre (56.4%), and less than 20% for the two oat fibres, CMC, and psyllium. Averaged across substrates, acetate, propionate, and butyrate were produced in the molar proportion of 64:24:12. Potential water-holding capacity (PWHC) of substrates, a measure of faecal bulking potential, was greatest for CMC (13.5 g H2O/g substrate) and lowest for gum arabic (1.92 g) and soy fibre (1.71 g). Organic matter disappearance and SCFA production of blends were directly proportional to their gum arabic content. Blend PWHC was proportional to CMC content. In vitro procedures are useful in predicting the actions of fibre blends formulated to produce desirable effects in vivo.

Effect of frequency and amount of rumen-degradable intake protein supplementation on urea kinetics and microbial use of recycled urea in steers consuming low-quality forage.

We evaluated the effect of frequency and amount of rumen-degradable intake protein (DIP) on urea kinetics in steers consuming prairie hay. Five ruminally and duodenally fistulated steers (366 kg of BW) were used in a 5 x 5 Latin square and provided ad libitum access to low-quality prairie hay (4.7% CP). Casein was provided daily in amounts of 61 and 183 mg of N/kg of BW (61/d and 183/d) and every third day in amounts of 61, 183, and 549 mg of N/kg of BW per supplementation event (61/3d, 183/3d, and 549/3d). Periods were 18-d long with 9 d for adaptation and 9 d for collection. Steers were in metabolism crates for total collection of urine and feces. Jugular infusion of (15)N(15)N-urea followed by determination of urinary enrichment of (15)N(15)N-urea and (14)N(15)N-urea was used to determine urea kinetics. Treatment means were separated to evaluate the effects of increasing DIP supplementation and the effects of frequency at the low (61/d vs. 183/3d) and at the high (183/d vs. 549/3d) amounts of DIP provision. Forage OM and total digestible OM intakes were linearly (P < or = 0.05) increased by increasing DIP provision but were not affected by frequency of supplementation at either the low or high amounts. Production and gut entry of urea linearly (P < or = 0.006) increased with DIP provision and tended to be greater (P < or = 0.07) for 549/3d than 183/d but were not different between 61/d and 183/3d. Microbial N flow to the duodenum was linearly (P < 0.001) increased by increasing DIP provision. Additionally, 183/d resulted in greater (P = 0.05) microbial N flow than 549/3d. Incorporation of recycled urea-N into microbial N linearly (P = 0.04) increased with increasing DIP. Microbial incorporation of recycled urea-N was greater for 549/3d than 183/d, with 42 and 23% of microbial N coming from recycled urea-N, respectively. In contrast, there was no difference due to frequency in the incorporation of recycled urea-N by ruminal microbes at the low level of supplementation (i.e., 61/d vs. 183/3d). This study demonstrates that urea recycling plays a substantial role in the N supply to the rumen and to the animal, particularly in steers supplemented infrequently with high levels of protein.

Dietary molasses increases ruminal pH and enhances ruminal biohydrogenation during milk fat depression

Feeding high-concentrate diets has the potential to cause milk fat depression, but several studies have suggested that dietary sugar can increase milk fat yield. Two experiments were conducted to evaluate the ability of dietary molasses to prevent milk fat depression in the presence of a 65% concentrate diet. In trial 1, molasses replaced corn grain at 0, 2.5, or 5% of diet dry matter in diets fed to 12 second-lactation Holstein cows (134±37 d in milk) in a 3×3 Latin square design. Trial 1 demonstrated that replacing up to 5% of dietary dry matter from corn with molasses had positive effects on de novo fatty acid synthesis, increasing the yield of short- and medium-chain fatty acids during diet-induced milk fat depression. Increasing inclusion rate of molasses increased milk fat concentration, but decreased milk yield and milk protein yield. Trial 2 used 7 ruminally cannulated, multiparous, late-lactation Holstein cows (220±18 d in milk) to evaluate effects of dietary molasses on ruminal parameters and milk composition, and also to assess whether increased metabolizable protein supply would alter these responses. Cows were randomly assigned to a dietary treatment sequence in a crossover split plot design with 0 and 5% molasses diets. Dietary treatments were fed for 28 d, with 16 d for diet adaptation, and the final 12 d for 2 abomasal infusion periods in a crossover arrangement. Abomasal infusions of water or AA (5 g of l-Met/d+15 g of l-Lys-HCl/d+5 g of l-His-HCl-H(2)O/d) were administered 3 times daily for 5 d, with 2 d between infusion periods. Administration of AA had no effect on concentration or yield of any milk components. Addition of molasses increased milk fat concentration (2.71 vs. 2.94±0.21%), but had no effect on yields of milk fat or protein. Dietary molasses decreased total volatile fatty acid concentration (141 vs. 133±4.6mM), decreased the molar proportion of propionate, and increased the molar proportion of butyrate in ruminal fluid. Molasses also increased ruminal pH (5.73 vs. 5.87±0.06), decreased the yield of trans-10 C18:1, and increased the yield of trans-11 C18:1 in milk fat. These data provide evidence that molasses may promote mammary de novo fatty acid synthesis in cows fed high-energy rations by moderating ruminal pH and altering ruminal fatty acid biohydrogenation pathways.

Effect of undegradable intake protein supplementation on urea kinetics and microbial use of recycled urea in steers consuming low-quality forage

We evaluated the effect of undegradable intake protein (UIP) on urea kinetics and microbial incorporation of urea-N in ruminally and duodenally fistulated steers (n 4; 319 kg) provided ad libitum access to grass hay in a 4 x 4 Latin square. Casein was continuously infused abomasally in amounts of 0, 62, 124 and 186 mg N/kg body weight per d to simulate provision of UIP. Periods were 13 d long with 7 d for adaptation and 6 d for collection. Jugular infusion of [15N15N]urea followed by determination of urinary enrichment of [15N15N]urea and [14N15N]urea was used to measure urea kinetics. Forage and N intake increased (quadratic, P<0.02) with increasing UIP. Urea synthesis was 27.1, 49.9, 82.2 and 85.8 g urea-N/d for 0, 62, 124 and 186 diets, respectively (linear, P<0.01). The proportion of urea synthesis that entered the gastrointestinal tract was 0.96 for steers receiving no UIP and decreased linearly (P=0.05) to a low of 0.89 for steers receiving 186. The amount of urea entering the gastrointestinal tract was least for 0 (26.3) and increased (linear, P<0.01) to 48.7, 77.2 and 76.6 g urea-N/d for 62, 124 and 186 diets, respectively. Microbial incorporation of recycled urea-N increased quadratically (P=0.04) from 13.9 for 0 to 47.7 g N/d for 124. The proportion of microbial N derived from recycled urea increased (quadratic, P=0.05) from 0.31 to 0.58 between 0 and 124 and dropped to 0.44 for 186 mg N/kg body weight per d. UIP increased intake of hay and provided a N source for ruminal microbes via urea recycling.

An unusual distribution of the niacin receptor in cattle.

Responses to pharmacological doses of niacin, an agonist for GPR109A (niacin receptor), were different in cattle than in humans and rodents. Thus, the tissue distribution of GPR109A was investigated in cattle. Samples of tail head fat, back fat, perirenal fat, longissimus muscle, and liver were analyzed for abundance of GPR109A mRNA by quantitative real-time reverse transcription-PCR and for abundance of GPR109A protein by Western blotting. Niacin receptor transcript and protein were detected in all tissues analyzed. The mRNA for GPR109A was more abundant in liver than in the other tissues sampled (GPR109A:RPS9 mRNA abundance = 0.56 in liver compared with 0.06 in longissimus muscle, 0.15 in kidney fat, 0.11 in back fat, 0.23 in tail head fat; standard error of the mean = 0.028). Additionally, mRNA for GPR109A was found (GPR109A:RPS9 mRNA abundance ≥ 0.004) in each of the 5 regions of bovine brain that were analyzed: cerebral cortex, cerebellum, thalamus, hypothalamus, and brain stem. Evaluation of liver tissue by immunofluorescence suggested that GPR109A was expressed in parenchymal cells and not localized exclusively to immune-system cells. Finally, analysis of the putative bovine GPR109A sequence verified that AA residues required for binding niacin in human GPR109A are conserved, suggesting that the bovine sequence identified encodes a functional niacin receptor. The identification of GPR109A in bovine liver, muscle, and brain is a novel finding.

Effect of nitrogen supplementation on urea kinetics and microbial use of recycled urea in steers consuming corn-based diets.

We studied the effects of supplementing N as distillers dried grains with solubles (DDGS) or urea to steers consuming corn-based diets. Six ruminally and duodenally cannulated steers (244 kg) were used in 2 concurrent 3 x 3 Latin squares and fed 1 of 3 corn-based diets: control (10.2% CP), urea (13.3% CP), or DDGS (14.9% CP). Periods were 14 d, with 9 d for adaptation and 5 d for collection of urine and feces. Urinary (15)N(15)N-urea enrichments, resulting from venous infusions of (15)N(15)N-urea, were used to measure urea kinetics. Dry matter intake (6.0 kg/d) was not affected by treatment, but N intake differed (99, 151, and 123 g/d for the control, DDGS, and urea treatments, respectively). Urea-N synthesis tended to be greater (P = 0.09) for DDGS (118 g/d) than for the control treatment (52 g/d), with the urea treatment (86 g/d) being intermediate. Urea-N excreted in the urine was greater (P < 0.03) for the DDGS (35 g/d) and urea treatments (29 g/d) than for the control treatment (13 g/d). Gastrointestinal entry of urea-N was not statistically different among treatments (P = 0.25), but was numerically greatest for DDGS (83 g/d), intermediate for urea (57 g/d), and least for the control (39 g/d). The amount of urea-N returned to the ornithine cycle tended to be greater (P = 0.09) for the DDGS treatment (47 g/d) than for the urea (27 g/d) or control treatment (16 g/d). The fraction of recycled urea-N that was apparently used for anabolism tended (P = 0.14) to be greater for the control treatment (0.56) than for the DDGS treatment (0.31), with the urea treatment (0.45) being intermediate, but no differences were observed among treatments in the amount of urea-N used for anabolism (P = 0.66). Urea kinetics in cattle fed grain-based diets were largely related to the amount of N consumed. The percentage of urea production that was captured by ruminal bacteria was greater (P < 0.03) for the control treatment (42%) than for the DDGS (25%) or urea treatment (22%), but the percentage of duodenal microbial N flow that was derived from recycled urea-N tended (P = 0.10) to be greater for the DDGS treatment (35%) than for the urea (22%) or control treatment (17%). Thus, ruminal microbes were more dependent on N recycling when the protein supplement was largely resistant to ruminal degradation.