Nutrient content,in vitro ruminal fermentation characteristics and methane reduction potential of tropical tannin‐containing leaves

BACKGROUND: Plant tannins as rumen modifiers are better than chemicals or antibiotic‐based modifiers since these compounds are natural products which are environmentally friendly and therefore have a better acceptance with regard to feed safety issues. Tropical plants containing phenols such as tannins were found to suppress or eliminate protozoa from the rumen and reduce methane and ammonia production. The screening of these plants is an important step in the identification of new compounds and feed additives which might contribute to mitigate rumen methanogenesis. The present study was carried out to determine the efficacy of tannins from tropical tree leaves for their methane reduction properties. RESULTS: Activity of tannins, as represented by the increase in gas volume with the addition of polyethylene glycol (PEG)‐6000 as a tannin binder (tannin bioassay) was highest in Ficus bengalensis (555%), followed by Azardirachta indica (78.5%). PEG addition did not alter (P > 0.05) methane percentage in Ficus racemosa, Glyricidia maculata, Leucena leucocephala, Morus alba and Semaroba glauca, confirming that tannins in these samples did not affect methanogenesis. The increase (P < 0.05) in protozoa population with PEG was maximal in Ficus religiosa (50), followed by Moringa oleifera (31.2), Azardirachta indica (29.9) and Semaroba glauca (27.5). There was no change (P > 0.05) in the protozoa population in Autocarpus integrifolia, Ficus bengalensis, Jatropha curcus, Morus alba and Sesbania grandiflora, demonstrating that methane reduction observed in these samples per se was not due to defaunation effect of the tannin. The increase in total volatile fatty acid concentration in samples with PEG ranged from 0.6% to > 70%. The highest increase (%) in NH₃‐N was recorded in Azardirachta indica (67.4), followed by Ficus mysoriensis (35.7) and Semaroba glauca (32.6) leaves, reflecting strong protein binding properties of tannin. CONCLUSION: The results of our study established that in vitro methanogenesis was not essentially related to the density of protozoa population. Tropical tree leaves containing tannins such as Autocarpus integrifolia, Jatropha curcus and Sesbania grandiflora have the potential to suppress methanogenesis. Therefore tannins contained in these plants could be of interest in the development of new additives in ruminant nutrition. Copyright © 2012 Society of Chemical Industry     

Effect of sarsaponin on ruminal fermentation with particular reference to methane production in vitro

This experiment was designed to investigate the effects of different concentrations (0, 1.2, 1.8, 2.4, and 3.2 g/L) of sarsaponin on ruminal microbial methane production using the substrates soluble potato starch, cornstarch, or hay plus concentrate (1.5:1). Ruminal fluid was collected from a dairy cow, mixed with phosphate buffer (1:2) and incubated (30 ml) anaerobically at 38 degrees C for 6 and 24 h with or without sarsaponin. Excluding the lower level of sarsaponin, pH of the medium was slightly decreased. Ammonia-N concentration and numbers of protozoa were decreased in a dose-dependent manner. Total volatile fatty acids and total gas production were increased. Molar proportion of acetate was decreased and propionate was increased with a corresponding decrease in acetate:propionate ratio. Hydrogen production was decreased. As the concentration of sarsaponin increased from 1.2 to 3.2 g/L, fermentation of soluble potato starch, cornstarch, or hay plus concentrate decreased methane production from 20 to 60% (6 h) and 17 to 50% (24 h), 21 to 58% (6 h) and 18 to 52% (24 h), and 23 to 53% (6 h) and 15 to 44% (24 h), respectively. Excluding the lower dose concentration (1.2 g/L) of sarsaponin, in vitro disappearance of dry matter of hay plus concentrate was decreased after 24 h. In conclusion, these results show that sarsaponin stimulated the mixed ruminal microorganism fermentation as well as to inhibit methane production in vitro.     

Effects of acetate,propionate, and pH on volatile fatty acid thermodynamics in continuous cultures of ruminal contents

To investigate the effects of acetate, propionate, and pH on thermodynamics of volatile fatty acids (VFA) in the rumen, a dual-flow continuous culture study was conducted to quantify production of major VFA, interconversions among the VFA, and H₂ and CH₄ emissions in a 4 × 4 Latin square design. The 4 treatments were (1) control: pH buffered to an average of 6.75; (2) control plus 20 mmol/d of infused acetate (InfAc); (3) control plus 7 mmol/d of infused propionate (InfPr); and (4) a 0.5-unit decline in pH elicited by adjustment of the buffer (LowpH). All fermentors were fed 40 g of a pelleted diet containing whole alfalfa pellets and concentrate mix pellets (50:50) once daily. After 7 d of treatment, sequential, continuous infusions of [2-¹³C] sodium acetate (3.5 mmol/d), [U-¹³C] sodium propionate (2.9 mmol/d), and [1-¹³C] sodium butyrate (0.22 mmol/d) were carried out from 12 h before feeding for 36 h. Filtered liquid effluent (4 mL) was sampled at 0, 2, 4, 6, 8, 12, 16, and 22 h after feeding, and assessed for VFA concentrations, with another filtered sample (20 mL) used to quantify aqueous concentrations of CH₄ and H₂. Headspace CH₄ and H₂ gases were monitored continuously. Ruminal microbes were isolated from the mixed effluent samples, and the microbial community structure was analyzed using the 16S rRNA amplicon sequencing technique. The digestibility of neutral detergent fiber, acid detergent fiber, and starch and microbial C sequestrated from VFA were not affected by treatments. The LowpH treatment increased net propionate production and decreased H₂ and CH₄ headspace emissions, primarily due to shifts in metabolic pathways of VFA formation, likely due to the observed changes in bacterial community structure. Significant interconversions occurred between acetate and butyrate, whereas interconversions of other VFA with propionate were relatively small. The InfAc and InfPr treatments increased net acetate and propionate production, respectively; however, interconversions among VFA were not affected by pH, acetate, or propionate treatments, suggesting that thermodynamics might not be a primary influencer of metabolic pathways used for VFA formation.     

Insect growth regulators and in vitro volatile fatty acid production.

Three insect growth regulators were tested for their effects on in vitro volatile fatty acid production. Each material was tested at 100 and 200 ppm of feed in a triplicated trial. At the concentrations used, there was a trend toward lowered acetate:propionate ratios with inconsistent effects on total volatile fatty acid production for all materials. Of the individual acids, only the relative production of valeric was affected significantly.     

Inoculant effects on alfalfa silage: in vitro gas and volatile fatty acid production

Alfalfa silages from 2 similar trials were analyzed for in vitro ruminal gas production. In both trials, there were 15 treatments: alfalfa treated at ensiling with 1 of 14 lactic acid bacterial inoculants or untreated alfalfa. First-cut (477 g of dry matter/kg) and second-cut (393 g of dry matter/kg) alfalfa were ensiled in glass jars for a minimum of 35 d at room temperature (∼22°C). At opening, a portion of each silage was wet-ground with a mixer. Each silage was then assessed for in vitro ruminal gas production in 3 replicate runs with the wet-ground silage, 1 on the fresh silage and 2 on frozen and thawed silage. In vitro gas production was measured in 160-mL sealed serum vials incubated at 39°C. One gram of silage was incubated with 17.1 mL of nutrient solution, 0.9 mL of reducing solution, and 12 mL of ruminal inoculum (1:2 vol.vol mixture of rumen fluid and buffer). Gas production was measured manually by using a pressure gauge at 3, 6, 9, 24, 48, and 96 h. At 96 h, the rumen fluid was analyzed for pH and volatile fatty acids. In the 2 trials, the untreated control silage produced either numerically the highest or one of the highest levels of gas production per unit of dry matter incubated. In first-cut silage, 9 of the inoculant treatments at 9 h and 4 treatments at 96 h had reduced gas production compared with the control. In second-cut silage, 10 inoculant treatments at both 9 and 96 h had reduced gas production compared with the control. Furthermore, in first-cut silage, the fraction of total gas production at 3, 6, and 9 h was numerically the highest for the control, and only 4 treatments were not significantly lower than the control at 9 h. In second-cut silage, 2 of 14 inoculated treatments produced faster fractional rates of gas production than the control, but most inoculated treatments had numerically slower fractional rates (4 significant) in the first 9 h. The in vitro fermented wet-ground control silages had one of the highest acetate:propionate ratios in both trials, significantly higher than 12 and 8 of the inoculated treatments in first- and second-cut silage, respectively. The response in acetate:propionate ratio in both cuts was similar, even though the control silage was highest in lactic acid in one trial and lowest in the other. Overall, inoculation of crops at ensiling appears to affect in vitro ruminal fermentation of wet-ground silages, even in the absence of large effects during silage fermentation.     

Assessment of the effect of condensed (acacia and quebracho) and hydrolysable (chestnut and valonea) tannins on rumen fermentation and methane production in vitro

BACKGROUND: Tannins added to animal diets may have a positive effect on energy and protein utilisation in the rumen. The objective of this study was to examine the impact of different sources and concentrations (20, 50, 100, 150 and 200 g kg^?1 dry matter (DM)) of condensed (acacia and quebracho) and hydrolysable (chestnut and valonea) tannins on rumen microbial fermentation in vitro. The experiment also included a negative control with no tannins (control) and a positive control with monensin (10 mg L^?1). RESULTS: In vitro gas production and total volatile fatty acid (VFA) concentration decreased as tannin concentration increased. Addition of acacia, chestnut or valonea tannins at ≥ 50 g kg^?1 or quebracho tannins at ≥ 100 g kg^?1 resulted in a decrease (up to 40%) in methane (CH₄) production compared with the control. Valonea tannins were the only tannin source that reduced (?11%) CH₄ production at 50 g kg^?1 without affecting VFA concentration. Tannin treatments reduced ammonia (NH₃) and branched‐chain VFA concentrations, indicating a reduction in ruminal protein degradation. Monensin reduced CH₄ production (?37%) and NH₃ concentration (?20%) without affecting total VFA concentration. CONCLUSION: Supplying acacia, chestnut or valonea tannins at 50 g kg^?1 has the potential to reduce CH₄ production and ruminal protein degradation with minimum detrimental effects on efficiency of ruminal fermentation. Copyright © 2012 Crown in the right of Canada. Published by John Wiley & Sons, Ltd     

Effects of bacterial direct-fed microbials on ruminal characteristics,methane emission,and milk fatty acid composition in cows fed high- or low-starch diets

This study investigated the effects of bacterial direct-fed microbials (DFM) on ruminal fermentation and microbial characteristics, methane (CH₄) emission, diet digestibility, and milk fatty acid (FA) composition in dairy cows fed diets formulated to induce different ruminal volatile fatty acid (VFA) profiles. Eight ruminally cannulated dairy cows were divided into 2 groups based on parity, days in milk, milk production, and body weight. Cows in each group were fed either a high-starch (38%, HS) or a low-starch (2%, LS) diet in a 55:45 forage-to-concentrate ratio on a dry matter (DM) basis. For each diet, cows were randomly assigned to 1 of 4 treatments in a Latin square design of (1) control (CON); (2) Propionibacterium P63 (P63); (3) P63 plus Lactobacillus plantarum 115 (P63+Lp); (4) P63 plus Lactobacillus rhamnosus 32 (P63+Lr). Strains of DFM were administered at 10¹⁰ cfu/d. Methane emission (using the sulfur hexafluoride tracer technique), total-tract digestibility, dry matter intake, and milk production and composition were quantified in wk 3. Ruminal fermentation and microbial characteristics were measured in wk 4. Data were analyzed using the mixed procedure of SAS (SAS Institute Inc., Cary, NC). The 2 diets induced different ruminal VFA profiles, with a greater proportion of propionate at the expense of acetate and butyrate for the HS diet. Greater concentrations of total bacteria and selected bacterial species of methanogenic Archaea were reported for the HS diet, whereas the protozoa concentration in HS decreased. For both diets, bacterial DFM supplementation raised ruminal pH (+0.18 pH units, on average) compared with CON. Irrespective of diet, P63+Lp and P63+Lr increased ruminal cellulase activity (3.8-fold, on average) compared with CON, but this effect was not associated with variations in ruminal microbial numbers. Irrespective of diet, no effect of bacterial DFM on ruminal VFA was observed. For the LS diet, supplementing cows with P63+Lr tended to decrease CH₄ emission (26.5%, on average, when expressed per kilogram of milk or 4% fat-corrected milk). Only P63 supplementation to cows fed the HS diet affected the concentration of some milk FA, such as cis isomers of 18:1 and intermediates of ruminal biohydrogenation of polyunsaturated FA. Overall, bacterial DFM could be useful to stabilize ruminal pH. Their effects on CH₄ production mitigation and milk FA profile depended on DFM strain and diet and should be confirmed under a greater variation of dietary conditions.     

Effect of the addition of fumarate on methane production by ruminal microorganisms in vitro

The effect of fumarate used as a feed additive on the reduction of methanogenesis in the rumen was evaluated by in vitro experiments. The addition of fumarate to the culture of mixed ruminal microorganisms that were fermenting hay powder and concentrate reduced methane production. Most fumarate was metabolized to propionate, and a slight increase was noted in other volatile fatty acids. Fumarate was utilized by mixed bacteria but not by mixed protozoa. Fibrobacter succinogenes, Selenomonas ruminantium ssp. ruminantium, Selenomonas ruminantium ssp. lactilytica, Veillonella parvula, and Wollinella succinogenes oxidized H2 by using fumarate as a final electron acceptor, suggesting that these bacteria compete with methanogens for H2, which is the main substrate for methanogenesis in the rumen. However, the affinity of these bacteria to H2 was lower than their affinity to methanogens. These fumarate-utilizing bacteria metabolized malate to products that were similar to those from fumarate, suggesting the possession of fumarate dehydratase. Fibrobacter succinogenes, V. parvula, and W. succinogenes utilized formate, another substrate for methanogenesis, as an electron donor for fumarate reduction. The affinity of these bacteria to formate was higher than the affinity methanogenes have for formate. When methanogens were cocultured with an equal cellular amount of each of the fumarate-utilizing bacteria, methane production was markedly decreased, not only from formate, but also from H2. These results suggest that the addition of fumarate to ruminant feed reduces methanogenesis and enhances propionate production in the rumen.     

Effects of tannic acid and quebracho tannins on in vitro ruminal fermentation of wheat and corn grain

Current understanding of the effect of moderate amounts of tannins on the utilization of protein contained in forages indicates beneficial effects when fed to ruminants. Nevertheless, there is little knowledge about the effects of tannins on the ruminal hydrolysis of starch‐rich concentrates. Modulation of the hydrolysis of starch may reduce the occurrence of metabolic disorders, such as acidosis, in ruminants consuming high‐grain diets. The effects of commercial tannic acid (hydrolysable tannins) and quebracho tannins (condensed tannins) (50 g kg^?1 DM) on the in vitro fermentation of ground wheat and corn grains by mixed ruminal bacteria was examined. The architecture and chemical composition of wheat and corn endosperms might explain differences in fermentation rates, as well as in response to similar concentrations of tannins. Regardless of the source of tannin, microbial fermentation was inhibited in both grains, as demonstrated by a decline in gas production, DM disappearance, volatile fatty acids and ammonia production. However, these effects were more pronounced for wheat than corn grain, mostly during the initial stages of the incubation. Scanning electron microscopy revealed that both sources of tannins inhibited the microbial hydrolysis of the endosperm protein matrix. Tannins did not prevent bacterial attachment to starch granules, but starch hydrolysis was slowed indirectly as a result of a tannin‐mediated reduction in the degradation of the surrounding protein matrix. Tannins are likely to be more effective at modulating the rate of starch digestion in grains that possess a readily degradable protein matrix. Copyright © 2006 Society of Chemical Industry     

Effects of chestnut tannins and coconut oil on growth performance, methane emission, ruminal fermentation, and microbial populations in sheep

This study was conducted to evaluate the effects of chestnut tannins (CT) and coconut oil (CO) on growth performance, methane (CH₄) emission, ruminal fermentation, and microbial populations in sheep. A total of 48 Rideau Arcott sheep (average body weight 31.5 ± 1.97 kg, 16 wk old) were randomly assigned into 6 treatment groups in a 3 × 2 factorial design, with CT and CO as the main effects (8 sheep per group). The treatments were control diet (CTR), 10 or 30 g of CT/kg of diet (CT10 and CT30), 25 g of CO/kg of concentrate (CO25), and 10 or 30 g of CT/kg of diet + 25 g of CO/kg of concentrate (CT10CO25 and CT30CO25). After the feeding trial (60 d), all sheep were moved to respiratory chambers to measure CH₄ emission. After CH₄ emission measurements, all sheep were slaughtered to obtain rumen fluid samples. Results showed that the addition of CT, CO, and CT + CO had no significant effects on growth performance of sheep but reduced CH₄ emission. Addition of CT reduced the NH₃-N concentration in rumen fluid in CT30. Addition of CO decreased the concentration of total volatile fatty acids in rumen fluid. No significant differences were observed in pH and molar proportion of volatile fatty acids among treatments. Addition of CT, CO, and CT + CO significantly decreased methanogen and protozoa populations. Moreover, CO decreased counts of Fibrobacter succinogenes. No significant differences were observed in populations of fungi, Ruminococcus flavefaciens, or Ruminococcus albus among treatments. In conclusion, supplementation of CT and CO seemed to be a feasible means of decreasing emissions of CH₄ from sheep by reduction of methanogen and protozoa populations with no negative effect on growth performance.     

The effect of oak tannin (Quercus robur) and hops (Humulus lupulus) on dietary nitrogen efficiency,methane emission,and milk fatty acid composition of dairy cows fed a low-protein diet including linseed

The objective of this study was to test the effects of inclusion of hop pellets (HP) and oak tannin extracts (OT) alone or in combination on N efficiency, methane (CH₄) emission, and milk production and composition in 2 experiments with dairy cows fed low-N rations supplemented with linseed. In both experiments, 6 lactating Holstein cows were assigned to 3 dietary treatments in a 3 × 3 duplicated Latin square design (21-d periods). Cows were fed a total mixed ration at a restricted level to meet their nutrient requirements. In experiment 1, 169 g dry matter (DM) of OT or 56 g DM of HP was included separately in the control diet (C1). In experiment 2, the additives were included together (OT-HP) in the control diet (C2) similar to C1. Diet C2 was compared with a control without linseed (C0). In experiment 1, the supplementation of the control diet with OT decreased urinary N excretion by 12%. In experiment 2, the combination of OT and HP decreased urinary N by 7%. Oak tannin extracts and HP alone or in combination did not influence the daily enteric CH₄ production of cows. Cows fed diet C0 produced 17% more enteric CH₄ daily than those fed diet C2. Intake of diet C2, which contained 6.7% extruded linseed on a DM basis (experiment 2), decreased the sum of 6:0 to 14:0 fatty acids (?16%) and palmitic acid (?26%) and increased the stearic acid (+50%), oleic acid (+36%), vaccenic acid (trans-11 18:1; +285%), rumenic acid (cis-9,trans-11 18:2; +235%), and α-linolenic acid (+100%) in milk fat. The supplementation of diet C2 with the OT-HP mixture further improved the milk's fatty acid composition. Intake of the OT alone increased α-linolenic acid by 17.7% (experiment 1). The results of this study show that at the economically acceptable dose we tested, hops had no effect on urinary N excretion, CH₄ emission, milk production, and milk composition. By contrast, supplementation of diets with oak tannin extract can be considered for reducing urinary N excretion. The combination of oak tannin and hops had no more effect than oak tannin alone except on the milk fatty acid profile, which was favorably influenced from a nutritional point of view.     

Nutrient and energy content, in vitro ruminal fermentation characteristics and methanogenic potential of alpine forage plant species during early summer

BACKGROUND: Plants growing on alpine meadows are reported to be rich in phenols. Such compounds may affect ruminal fermentation and reduce the plants' methanogenic potential, making alpine grazing advantageous in this respect. The objective of this study was to quantify nutrients and phenols in Alpine forage grasses, herbs and trees collected over 2 years and, in a 24 h in vitro incubation, their effects on ruminal fermentation parameters. RESULTS: The highest in vitro gas production, resulting in metabolisable energy values around 10 MJ kg^?1, were found with Alchemilla xanthochlora and Crepis aurea (herbaceous species) and with Sambucus nigra leaves and flowers (tree species). Related to the amount of total gas production, methane formation was highest with Nardus stricta, and lowest with S. nigra and A. xanthochlora. In addition, Castanea sativa leaves led to an exceptional low methane production, but this was accompanied by severely impaired ruminal fermentation. When the data were analysed by principal component analysis, phenol concentrations were negatively related with methane proportion in total gas. CONCLUSION: Variation in methane production potential across the investigated forages was small. The two goals of limited methane production potential and high nutritive value for ruminants were met best by A. xanthochlora and S. nigra. Copyright © 2011 Society of Chemical Industry