Exploitation of dietary tannins to improve rumen metabolism and ruminant nutrition

Tannins (hydrolysable and condensed tannin) are polyphenolic polymers of relatively high molecular weight with the capacity to form complexes mainly with proteins due to the presence of a large number of phenolic hydroxyl groups. They are widely distributed in nutritionally important forage trees, shrubs and legumes, cereals and grains, which are considered as anti‐nutritional compounds due to their adverse effects on intake and animal performance. However, tannins have been recognised to modulate rumen fermentation favourably such as reducing protein degradation in the rumen, prevention of bloat, inhibition of methanogenesis and increasing conjugated linoleic acid concentrations in ruminant‐derived foods. The inclusion of tannins in diets has been shown to improve body weight and wool growth, milk yields and reproductive performance. However, the beneficial effects on rumen modulation and animal performance have not been consistently observed. This review discusses the effects of tannins on nitrogen metabolism in the rumen and intestine, and microbial populations (bacteria, protozoa, fungi and archaea), metabolism of tannins, microbial tolerance mechanisms to tannins, inhibition of methanogenesis, ruminal biohydrogenation processes and performance of animals. The discrepancies of responses of tannins among different studies are attributed to the different chemical structures (degree of polymerisation, procyanidins to propdelphinidins, stereochemistry and C? C bonding) and concentrations of tannins, and type of diets. An establishment of structure–activity relationship would be required to explain differences among studies and obtain consistent beneficial tannin effects. Copyright © 2010 Society of Chemical Industry     

Comparison of biological and chemical methods,and internal and external standards,for assaying tannins in Spanish shrub species

Tannin contents of stems, flowers and leaves of four shrub species (Erica arborea, Erica australis, Cytisus cantabricus and Genista occidentalis) were chemically assessed using two colorimetric assays (butanol–HCl and Folin–Ciocalteu) and several standards (tannic acid, purified quebracho condensed tannins (CT) and purified CT extracted from each species). A method based on the biological activity of tannins in the rumen was also used for assaying tannins. Depending on the species, the use of quebracho tannin as standard over‐ or underestimated CT contents, in comparison to the use of the respective internal standard. However, the high correlationship observed between in vitro gas production in the presence or absence of polyethylene glycol (PEG) and CT contents estimated from quebracho suggests that this standard could be used for a quicker estimate of the nutritive value of tannin‐containing plants. Tannin contents obtained by the Folin–Ciocalteu method, using tannic acid as standard, were also related to their activity on rumen fermentation. The use of the in vitro gas production technique, in the absence or presence of PEG, may provide more useful information than the chemical assays when the final aim of tannin analysis is to assess the nutritive value of feedstuffs for ruminants. Copyright © 2004 Society of Chemical Industry     

Interrelationships between the concentrations of total condensed tannin,free condensed tannin and lignin in Lotus sp. and their possible consequences in ruminant nutrition

Interrelationships between the concentrations of total condensed tannin (TCT), free condensed tannin (FCT) and lignin were studied to gain knowledge of how to manipulate nutritive value of fresh herbages containing condensed tannins fed to ruminants. FCT was defined as condensed tannin not bound by macerates of fresh plants, with both FCT and TCT being determined with vanillin HCl. Effects of spraying lotus with polyethylene glycol (mol. wt 3350; PEG) upon the relationship between FCT and TCT was also studied. Increasing soil nutrient and climatic stress caused large and similar increases in the concentrations of TCT and of lignin. Over the range 0–90 g kg^?1 DM, 10% of TCT in Lotus sp. was detected as FCT, with increments in TCT above 90 g kg^?1 DM being released almost entirely as FCT. PEG formed much stronger chemical bonds with condensed tannins than did plant proteins, and did not release FCT; consequently PEG application reduced the concentration of condensed tannin that was detectable with vanillin HCl. After disintegration of plant material, it is proposed that most condensed tannin is bound and co-precipitated as an insoluble complex with protein, that FCT is in equilibrium with this complex, and that bound and free tannin are indices of nutritionally beneficial and detrimental effects produced by condensed tannins in fresh forages eaten by ruminants. It was concluded that growing Lotus pedunculatus under conditions of stress leads to depressions in nutritive value through simultaneously increasing concentrations of lignin and FCT, both of which depress rumen carbohydrate digestion and voluntary intake, and that treatment with PEG offers a convenient method of separating effects due to condensed tannins from other factors influencing nutritive value.     

Complexes of the condensed tannins of sainfoin (Onobrychis viciifolia scop.) with fraction 1 leaf protein and with submaxillary mucoprotein,and their reversal by polyethylene glycol and pH

In vitro studies were made to determine the nature of complexes formed between the condensed tannins (i.e. pro-anthocyanidins or flavolans) of sainfoin (Onobrychis viciifolia) and either the major soluble dietary protein, Fraction 1 protein, of green leaves, or bovine salivary mucoprotein. Fraction 1 protein, uniformly labelled with ¹⁴C, formed only insoluble complexes with these tannins. Bovine salivary mucoprotein produced insoluble complexes only at temperatures below 25°C. Fraction 1 protein complexes were stable between pH 3.5 and pH 7.0. At pHs below 3.0 greater than 90% of the protein was solubilised in the presence of PEG but at pH 8.0 only 30% of the protein was released from the complex. In vivo experiments with sheep fed sainfoin, confirmed the stability of the tannin-Fraction 1 complexes in the rumen (pH 6.5) and break-up of the complexes in anterior duodenal samples (pH 2.5) as shown by the extractability of condensed tannin from the complex. The significance of these results with respect to (a) bloat in cattle, (b) nitrogen metabolism of ruminants, is discussed. Polyethylene-glycol, mol.wt. 4000 (PEG) exchanged with protein in the complex releasing protein into solution. The effectiveness of PEG was dependent on the amount of tannin in the complex and the age of the complex at the time of addition of PEG. The results explain the observation that PEG is unsuitable as a marker for rumen volume determination when animals are fed herbage that contains condensed tannins. A method for determining any deleterious effects of tannins on nitrogen metabolism by ruminants, is suggested.     

Degradation of condensed tannins by rumen microbes exposed to quebracho tannins (QT) in rumen simulation technique (RUSITEC) and effects of QT on fermentative processes in the RUSITEC

In an earlier study it was observed that the rumen microbes from cattle which had never consumed tannin-containing diets do not have enzymes for degrading condensed tannins. In this study, rumen microbes were exposed to small amounts of quebracho tannins for 8 days using a rumen simulation technique (RUSITEC). The levels maintained in the RUSITEC were 0.1, 0.2 and 0.4 mg of the spray-dried quebracho powder (SDQT) per ml of the medium. After 8 days of tannin exposure, the liquor containing ‘exposed/adapted’ microbes from RUSITEC was incubated for 40 h without and with purified tannins of quebracho and Dichostachys cinerea leaves in in vitro gas method. There was no degradation of condensed tannins. The enzymes for degradation of condensed tannins were not induced in rumen microbes by exposure to different concentrations of tannins for 8 days in the RUSITEC. In the RUSITEC, SDQT significantly reduced the number of total protozoa, entodiniomorphs and holotrichs; effect was higher on holotrichs. There was no significant change in the levels of short chain fatty acid but the molar proportion of propionate was significantly higher and of butyrate significantly lower at 0.4 mg SDQT ml^?1. Significantly lower levels of ammonia in the medium was also observed on injection of tannins. Microbial mass production, calculated using ¹⁵N incorporation, was similar at 0.1 and 0.2 mg SDQT ml^?1 but significantly lower (13%) at 0.4 mg SDQT ml^?1. The dry matter digestibility of the feed (80% hay and 20% barley) was not significantly affected by SDQT.     

Evaluation of tropical plants containing tannin on in vitro methanogenesis and fermentation parameters using rumen fluid

BACKGROUND: Methane (CH₄) produced during ruminal fermentation represents a loss of 10–11% of gross energy intake. The use of browse species containing tannin as feed supplement for ruminants tends to increase in order to reduce CH₄ production. The present study was conducted to evaluate some tropical plants containing tannin as feed supplement (200 g kg^?1) on in vitro CH₄ production and fermentation parameters. RESULTS: The crude protein (CP) content ranged from 87 to 390 g kg^?1 dry matter (DM) and was highest in Sesbania grandiflora (L.) Poiret. The neutral detergent fibre (NDF) concentration was highest in Pennisetum purpureum Schumach (725 g kg^?1 DM) and lowest in S. grandiflora (330 g kg^?1 DM). The ranking order of plants based on their total tannin content was Acacia mangium Willd. > Biophytum petersianum Klotzch > Jatropa curcas Linnaeus > Psidium guajava Linnaeus > Phaleria papuana > Persea americana Mill. > S. grandiflora. Methane gas production after 48 h of incubation was significantly (P < 0.05) lower with inclusion of A. mangium (PP + AM), B. petersianum (PP + BP), J. curcas (PP + JC) or P. guajava (PP + PG) as compared to control feed (PP). There was negative correlation between total tannin content and CH₄ production at 48 h of incubation (r = ? 0.76). Concentration NH₃‐N was significantly (P < 0.01) higher with inclusion of S. glandiflora. Inclusion of P. guajava significantly (P < 0.05) suppressed protozoa population by 49.7% relative to control feed (PP). CONCLUSION: It was concluded that A. mangium, B. petersianum, J. curcas and P. guajava have potential to be used as a feed supplement to reduce CH₄ production in ruminants. Copyright © 2009 Society of Chemical Industry     

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     

Difference in the nature of tannins on in vitro ruminal methane and volatile fatty acid production and on methanogenic archaea and protozoal populations

Six plant sources of hydrolyzable tannins (HT) or HT and condensed tannins (CT; designated as HT1, HT2, HT3, HT + CT1, HT + CT2, and HT + CT3) were evaluated to determine their effects in vitro on CH₄ production and on ruminal archaeal and protozoa populations, and to assess potential differences in biological activities between sources containing HT only or HT and CT. Samples HT1, HT2, and HT3 contained only HT, whereas samples HT + CT1, HT + CT2, and HT + CT3 contained HT and CT. In experiment 1, in vitro incubations with samples containing HT or HT + CT resulted in a decrease in CH₄ production of 0.6 and 5.5%, respectively, compared with that produced by incubations containing the added tannin binder polyethylene glycol-6000. Tannin also suppressed the population of methanogenic archaea in all incubations except those with HT2, with an average decrease of 11.6% in HT incubations (15.8, 7.09, and 12.0 in HT1, HT2, and HT3) and 28.6% in incubations containing HT + CT (35.0, 40.1, and 10.8 in HT + CT1, HT + CT2, and HT + CT3) when compared with incubations containing added polyethylene glycol-6000. The mean decrease in protozoal counts was 12.3% in HT and 36.2% in HT + CT incubations. Tannins increased in vitro pH, reduced total VFA concentrations, increased propionate concentrations, and decreased concentrations of iso-acids. In experiment 2, when a basal diet was incubated with graded levels of HT + CT1, HT + CT2, and HT + CT3, the total gas and CH₄ production and archaeal and protozoal populations decreased as the concentration of tannins increased. Our results confirm that tannins suppress methanogenesis by reducing methanogenic populations in the rumen either directly or by reducing the protozoal population, thereby reducing methanogens symbiotically associated with the protozoal population. In addition, tannin sources containing both HT and CT were more potent in suppressing methanogenesis than those containing only HT.     

Potential nutritive value and tannin bioassay of selected Acacia species from Kenya

Six Acacia forage species–A. brevispica, A. elatior, A. mellifera, A. nilotica, A. senegal and A. seyal—were analysed for their chemical composition, including phenolics and rumen fermentation characteristics. In vitro gas production technique was used to study the rumen fermentation characteristics and the effect of tannins present in the browse forages on in vitro fermentation by including polyethylene glycol (PEG‐6000) in the incubation. The forages had high crude protein content (145.7–270.1 g kg^?1 DM) and low to moderate content of neutral detergent fibre (220.2–442.8 g kg^?1 DM). The forages had variable content of total extractable phenolics ranging from 18.4 to 384.2 g kg^?1 DM and total extractable tannins ranging from 7.1 to 364.8 g kg^?1 DM (tannic acid equivalent). Fractionation of the condensed tannin flavonoids showed that the delphinidin/cyanidin ratio ranged from not detected:100 to 71:29. The gas production potential ranged from 28.4 to 40.8 mL gas 0.2 g^?1 DM. The rate of gas production was highest in A. elatior and lowest in A. nilotica. Addition of PEG‐6000 increased gas production significantly (P < 0.05) in all species except A. mellifera. PEG‐6000 addition did not have any significant effect on in vitro true dry matter and organic matter degradability but decreased the partitioning factor in all the species. Gas production and degradability parameters correlated positively with the CP content and negatively with the fibre and phenolics contents. Based on chemical composition, gas production and degradability, the forages have high potential nutritive value, especially as supplements to low‐quality feeds in the tropics and particularly during the dry season. However, the presence of tannins in terms of high levels and biological antinutritive activity may limit the nutritive potential of some of the forages, such as A. seyal and A. nilotica. Copyright © 2007 Society of Chemical Industry     

Comparison of the Precipitation of Alfalfa Leaf Protein and Bovine Serum Albumin by Tannins in the Radial Diffusion Method

The precipitation of protein by condensed and hydrolysable tannins was evaluated with the radial diffusion method of Hagerman (1987) using bovine serum albumin (BSA) and isolated leaf protein from fresh alfalfa (Medicago sativa). Alfalfa leaf protein (AALP) was included at two concentrations, 25 and 156 mg N litre^-1, at pH 6·8 and 39°C to simulate rumen conditions. The condensed tannins were purified from lyophilised samples of Arachis pintoi, Desmodium ovalifolium, Gliricidia sepium, Manihot esculenta and quebracho (Schinopsis balansae). Hydrolysable tannins from tannic acid (TA) were used as well. There was a significant interaction (P<0·001) between tannin and protein source, and protein level on protein precipitation. Most purified condensed tannins (CTs) precipitated more AALP than BSA when protein was included at the same level. Purified CT from quebracho and hydrolysable tannin from TA failed to precipitate AALP at both protein levels. In a second experiment, tannins from crude plant extracts were studied in the radial diffusion method using BSA and two levels of AALP. The crude plant extracts were obtained from lyophilised plant samples of A pintoi, Centrosema macrocarpum, Clitoria ternatea, D ovalifolium, Erythrina berteroana, E poepigiana, G sepium, M esculenta, Pueraria montana and P phaseoloides. The protein precipitated by soluble tannins in the plant samples was correlated to the total phenolic content and to the soluble CT estimated by the acid butanol assay or by the radial diffusion method. Tannins from different plant species precipitated different amounts of BSA and AALP. Therefore, the measures of the biological activity of tannins based on BSA precipitation may not reflect the ability of tannins to precipitate proteins of plant origin such as those commonly found in the diets of herbivores. The present study offers the possibility of using the radial diffusion method with plant proteins at precipitation conditions similar to those in the rumen. © 1997 SCI.     

Effect of adding an anaerobic fungal culture isolated from a wild blue bull (Boselophus tragocamelus) to rumen fluid from buffaloes on in vitro fibrolytic enzyme activity,fermentation and degradation of tannins and tannin‐containing Kachnar tree (Bauhinia variegata) leaves and wheat straw

The study investigated the effects of adding an anaerobic fungus (Piromyces sp FNG5; isolated from the faeces of a wild blue bull) to the rumen fluid of buffaloes consuming a basal diet of wheat straw and concentrates on in vitro enzyme activities, fermentation and degradation of tannins and tannin‐rich tree leaves and wheat straw. In experiment 1, strained rumen fluid was incubated for 24 and 48 h, in quadruplicate, with or without fungal culture using condensed tannin‐rich Bauhinia variegata leaves as substrates. In experiment 2, in vitro incubation medium containing wheat straw and different concentrations of added tannic acid (0–1.2 mg mL^?1) were incubated for 48 h, in quadruplicate, with strained buffalo rumen fluid with or without fungal culture. In experiment 3, tolerance of the fungal isolate to tannic acid was tested by estimating fungal growth in pure culture medium containing different concentrations (0–50 g L^?1) of tannic acid. In in vitro studies with Bauhinia variegata tree leaves, addition of the fungal isolate to buffalo strained rumen liquor resulted in significant (P < 0.01) increase in neutral detergent fibre (NDF) digestibility and activities of carboxymethyl cellulase (P < 0.05) and xylanase (P < 0.05) at 24 h fermentation. There was 12.35% increase (P < 0.01) in condensed tannin (CT) degradation on addition of the fungal isolate at 48 h fermentation. In in vitro studies with wheat straw, addition of the fungus caused an increase in apparent digestibility (P < 0.01), true digestibility (P < 0.05), NDF digestibility (P < 0.05), activities of carboxymethyl cellulase (P < 0.001), β‐glucosidase (P < 0.001), xylanase (P < 0.001), acetyl esterase (P < 0.001) and degradation of tannic acid (P < 0.05). Rumen liquor from buffaloes which had never been exposed to tannin‐containing diet had been found to have substantial inherent tannic acid‐degrading ability (degraded 55.3% of added tannic acid within 24 h of fermentation). The fungus could tolerate tannic acid concentration up to 20 g L^?1 in growth medium. The results of this study suggest that introduction of an anaerobic fungal isolate with superior lignocellulolytic activity isolated from the faeces of a wild herbivore may improve fibre digestion from tannin‐containing feeds and degradation of tannins in the rumen of buffaloes. Copyright © 2005 Society of Chemical Industry     

The condensed tannin content of vegetative Lotus pedunculatus,its regulation by fertiliser application,and effect upon protein solubility

The condensed tannin concentration in Lotus pedunculatus (cv. Grasslands ‘Maku’) was 8–11% DM when grown in acid soils without fertiliser application and 2–3% DM when grown in high fertility soils. Application of P & S fertiliser to the acid soils increased DM yield and reduced condensed tannin content to 4–5% DM, with over 88% of the variation in condensed tannin content being explained by variation in DM yield; it also increased plant total N concentration and halved the molar ratio of condensed tannin: protein (MR). Condensed tannins quantitatively precipitated soluble protein in lotus and also effectively precipitated protein in mixtures of lotus with white clover. The minimum concentration of condensed tannin necessary to precipitate protein was 2–4% DM, corresponding to MR values of 6–13. However, protein from such mixtures could be deaminated by fermentations with rumen fluid in vitro, but potentially soluble protein in pure lotus (MR 29) was protected from deamination by condensed tannins. A portion of lotus tannin could not be bound by plant constituents in finely ground fresh herbage and was designated ‘free’ tannin. This fraction was linearly related to total plant tannin content, was predicted to be zero at 1.8% DM total tannin, and increased at 0.15 units per unit increase in total tannin content above this figure. Maximum ‘free’ tannin comprised 17% of total condensed tannin. Effects of total and ‘free’ tannin content upon the intake and digestion of lotus by sheep are discussed.     

Meta‐analyses of effects of phytochemicals on digestibility and rumen fermentation characteristics associated with methanogenesis

BACKGROUND: A meta‐analysis study was conducted to investigate the changes in rumen fermentation characteristics when methane inhibition by phytochemicals is employed. The whole database containing 185 treatment means from 36 published studies was divided into four subsets according to the major phytochemicals used in the studies, i.e. saponins, tannins, essential oils (EO) and organosulfur compounds (OS). RESULTS: Changes in protozoal numbers showed linear relationships with changes in methane production by saponins (R² = 0.48), tannins (R² = 0.30) and EO (R² = 0.20) but not OS. Concentrations of total volatile fatty acids (VFA) and acetate did not show any relationship (P > 0.1) with changes in methane due to saponins. However, propionate production increased linearly with increasing inhibition of methane (R² = 0.31), which resulted in a linear (R² = 0.26) decrease in acetate/propionate ratio (A/P) with decreasing methane production. Concentrations of total VFA, acetate and propionate did not change with changes in methane production by tannins. However, A/P showed a significant linear relationship (R² = 0.27) with decreasing methane formation. Concentrations of total VFA (R² = 0.44) and propionate (R² = 0.15) changed linearly and positively with changes in methane production by EO. However, acetate production (R² = 0.22) and A/P (R² = 0.17) increased linearly with increasing inhibition of methane by EO. Changes in concentrations of total VFA (R² = 0.60) and acetate (R² = 0.35) decreased linearly while those of propionate increased linearly (R² = 0.23) with increasing inhibition of methane by OS. Consequently, A/P decreased linearly (R² = 0.30) with decreasing methane production by OS. Digestibilities of organic matter (OM) and neutral detergent fibre were not affected by inhibition of methane production by saponins, EO and OS, but digestibility of OM decreased with decreasing methane production by tannins. CONCLUSION: The inhibition of methane production by phytochemicals results in changes in rumen fermentation that differ depending on the types of phytochemicals. Copyright © 2010 Society of Chemical Industry     

Tea saponin reduced methanogenesis in vitro but increased methane yield in lactating dairy cows

The effect of tea saponin supplementation in the ruminant diet on methane emissions, rumen fermentation, and digestive processes is still under debate. The objective of this study was to assess the effect of this plant extract on methanogenesis, total-tract digestibility, and lactating performances of dairy cows. The work included 2 independent and successive experiments. First, the effect of 7 tea saponin doses (from 0 to 0.50 g/L) on methane emissions and protozoa concentrations was tested in 2 repeated in vitro batch culture incubations using bovine rumen contents as inoculum and a cereal mixture as substrate. After 18 h of incubation, total gas production and composition as well as rumen fermentation parameters and protozoa concentration were analyzed. Increasing dosage of the plant extract reduced methane production and protozoa concentration, with a maximum reduction of 29% for CH₄ (mL/g of substrate) and 51% for protozoa (10⁵/mL). Tea saponin did not affect volatile fatty acids concentration, but marginally decreased total gas production by 5% at the highest dose. Second, a 2-period crossover design experiment was carried out with 8 lactating dairy cows fed a basal diet (54% corn silage, 6% hay, and 40% pelleted concentrates on a dry matter basis) without (control) or with 0.52% tea saponin (TSP). Each experimental period lasted 5 wk. Animals were fed ad libitum during the first 3 wk of the period (wk 1, 2, and 3) and restricted (95% of ad libitum intake) during the last 2 wk (wk 4 and 5). Intake and milk production were recorded daily. Methane emissions were quantified using open chambers (2 d, wk 4). Total-tract digestibility and nitrogen balance were determined from total feces and urine collected separately (5 d, wk 5). Rumen fermentation parameters and protozoa concentration were analyzed from samples taken after morning feeding (1 d, wk 5). Milk production, dry matter intake, and feed efficiency were reduced with TSP (?18, ?12, and ?8%, respectively). As daily methane production (g/d) was not affected, methane emissions (g/kg of dry matter intake) increased by 14% with TSP. Total-tract digestibility and nitrogen balance were similar between diets, except for acid detergent fiber digestibility, which tended to be improved with TSP (+4 percentage units). Rumen fermentation parameters and protozoa concentration were relatively unchanged by diets. Under the conditions of this experiment, tea saponin is not efficient to reduce methane emissions from dairy cows.     

Invited review: Plant polyphenols and rumen microbiota responsible for fatty acid biohydrogenation,fiber digestion,and methane emission: Experimental evidence and methodological approaches

The interest of the scientific community in the effects of plant polyphenols on animal nutrition is increasing. These compounds, in fact, are ubiquitous in the plant kingdom, especially in some spontaneous plants exploited as feeding resources alternative to cultivated crops and in several agro-industry by-products. Polyphenols interact with rumen microbiota, affecting carbohydrate fermentation, protein degradation, and lipid metabolism. Some of these aspects have been largely reviewed, especially for tannins; however, less information is available about the direct effect of polyphenols on the composition of rumen microbiota. In the present paper, we review the most recent literature about the effect of plant polyphenols on rumen microbiota responsible for unsaturated fatty acid biohydrogenation, fiber digestion, and methane production, taking into consideration the advances in microbiota analysis achieved in the last 10 yr. Key aspects, such as sample collection, sample storage, DNA extraction, and the main phylogenetic markers used in the reconstruction of microbial community structure, are examined. Furthermore, a summary of the new high-throughput methods based on next generation sequencing is reviewed. Several effects can be associated with dietary polyphenols. Polyphenols are able to depress or modulate the biohydrogenation of unsaturated fatty acids by a perturbation of ruminal microbiota composition. In particular, condensed tannins have an inhibitory effect on biohydrogenation, whereas hydrolyzable tannins seem to have a modulatory effect on biohydrogenation. With regard to fiber digestion, data from literature are quite consistent about a general depressive effect of polyphenols on gram-positive fibrolytic bacteria and ciliate protozoa, resulting in a reduction of volatile fatty acid production (mostly acetate molar production). Methane production is also usually reduced when tannins are included in the diet of ruminants, probably as a consequence of the inhibition of fiber digestion. However, some evidence suggests that hydrolyzable tannins may reduce methane emission by directly interacting with rumen microbiota without affecting fiber digestion.     

Evaluation of edible flowers of agathi (Sesbania grandiflora L. Fabaceae) for in vivo anti-inflammatory and analgesic,and in vitro antioxidant potential

The methanol extract from flowers of agathi (Sesbania grandiflora L. Fabaceae) was evaluated for their in vitro antioxidant and cytotoxic activities, and in vivo anti-inflammatory and antinociceptive activities in several experimental models. The extract has sustainable concentrations of dietary polyphenolics, tannins, and flavonoids. The extract exhibited maximum radical scavenging activity on nitric oxide, superoxide, and hydroxyl radical and these values were significantly (p<0.05) higher over positive standards butylated hydroxyanisole and butylated hydroxytolune. The extract also exhibited potential cytotoxic activity against human cervical cancer cell line HeLa (IC₅₀ value of 0.13 mg/mL). Further, the methanol extracts showed significant inhibitory activity against inflammation (carrageenan and cotton pellet induced models) and on a pain model (hot plate test). The inhibitory values are comparable with positive standards. Owing to these properties, agathi flowers can be used as a potential source of natural nutraceutical food supplement.     

Effects of extracts of spices on rumen methanogenesis,enzyme activities and fermentation of feeds in vitro

BACKGROUND: An experiment was conducted to study the effects of boiling water, methanol and ethanol extracts (0, 0.25 and 0.50 mL) of seeds of Foeniculum vulgare (fennel), flower buds of Syzygium aromaticum (clove), bulbs of Allium sativum (garlic), bulbs of Allium cepa (onion) and roots of Zingiber officinalis (ginger) on rumen methanogenesis, fibrolytic enzyme activities and fermentation characteristics in vitro. RESULTS: Ethanol and methanol extracts of fennel, clove and garlic at 0.50 mL and clove at 0.25 mL inhibited (P < 0.05) methane production. Carboxymethylcellulase activity was reduced (P < 0.05) by ethanol and methanol extracts (0.50 mL) of fennel and clove (0.25 and 0.50 mL). The extracts of clove reduced (0.25 and 0.50 mL) xylanase and acetylesterase activities, and the fennel extract (0.50 mL) reduced (P < 0.05) xylanase activity. However, the extracts of garlic (0.50 mL) increased (P < 0.05) acetylesterase activity. Concentrations of volatile fatty acids were reduced (P < 0.05) by the extracts of garlic and onion. The extracts of garlic caused a decrease (P < 0.05) in acetate:propionate ratio (A:P) at 0.50 mL, whereas A:P was increased (P < 0.05) by the inclusion of 0.50 mL extracts of clove. Methanol and ethanol extracts of clove decreased (P < 0.05) in vitro organic matter degradability. Extracts (0.50 mL) of clove decreased (P < 0.05) the numbers of total protozoa, small entodiniomorphs and holotrichs, whereas extracts of onion, ginger and garlic enhanced (P < 0.05) protozoal numbers (both entodiniomorphs and holotrichs). CONCLUSION: Ethanol and methanol extracts of fennel and garlic have potential to inhibit rumen methanogenesis without adversely affecting rumen fermentation. Copyright © 2009 Society of Chemical Industry     

Condensed tannins in tropical fodder crops and their in vitro biological activity: Part 2

With the aim to evaluate the biological activity of purified condensed tannins of tropical forages we conducted two in vitro experiments. In the first, using a radial diffusion technique, the protein precipitation of free condensed tannins (FCT) from Manihot esculenta, leucaena leucocephala, Arachis pintoi, Guazuma ulmyfolia, Gliricidia sepium and of tannic acid on bovine serum albumin (ASB), papain, pepsin and trypsin at pH 5.0 and 6.8 was evaluated with a three‐way analysis of variance. Significant effects (P ≤ 0.05) for the tannin type, protein source, pH and their interactions were observed. Pepsin showed the highest protein precipitation (PP) at a pH of 5.0 (82.9 µg) with FCT of G ulmyfolia and the lowest (0 and 0.2 µg) of BSA with G sepium and A pintoi at pH 6.8. Experiments were then conducted using completely randomized designs in order to observe the effect of adding 0, 1.25 or 2.50 mg of FCT from M esculenta and L leucocephala to the rumen fluid‐buffer in an in vitro dry matter digestibility test (IVDMD) of Medicago sativa and Brachiaria decumbens, The IVDMD value of M sativa (757 g kg^?1) decreased with L leucocephala tannins, although with those of M esculenta it was increased (824 g kg^?1 and 871 g kg^?1, respectively) for 1.25 and 2.5 mg of FCT. The IVDMD value of B decumbens (774 g kg^?1 without tannins) diminished with any tannin and any dose (P < 0.05). We conclude that there are differences in the FCT contents of fodder crops and in their biological activity measured as the capacity to precipitate proteins, which is modified by the type of tannin, the protein and the pH. The result of an IVDMD is regulated by the type of tannin and its dose. Copyright © 2004 Society of Chemical Industry     

The ruminal degradability of fibre explains part of the low nutritional value and reduced methanogenesis in highly tanniniferous tropical legumes

BACKGROUND: Limited forage quality and low methanogenesis from fermentation of tropical shrub legumes is often attributed to high contents of condensed tannins (CTs), but characteristics of the fibre may be important, too. Using the rumen‐simulation technique an attempt was made to separate the influence of tannins and fibre on ruminal digestion and methanogenesis. Legumes with CTs (Calliandra calothyrsus, Flemingia macrophylla) and plants free of CTs (Vigna unguiculata, Brachiaria humidicola) were used. To separate effects, either CTs were inactivated using polyethylene glycol or purified fibres free of CTs were obtained. In experiment 1, legumes replaced one third of grass in the diet and urea was supplemented. In experiment 2, only purified fibres and casein were incubated. RESULTS: Purified fibres had a different degradability (low in legumes with CTs). Lignin prevented mainly hemicellulose from being degraded while the extent of degradation of hemicellulose seemed to have determined methanogenesis. In the mixed diets these effects were small when the supply of N was sufficient. When CTs were inactivated, fibre degradation was still lower with Calliandra than with Vigna. CONCLUSION: Properties of fibre, especially lignification, of CT‐rich plants help to explain the low feeding value of such forages, but CTs represent the over‐riding factor. Copyright © 2008 Society of Chemical Industry     

In vitro effects of and interactions between tannins and saponins and fate of tannins in the rumen

The rate of fermentation of hay was not affected by purified tannins from Ouercus incana and Dichostachys cinerea at 0.13 mg ml^?1 of the in vitro medium whereas a decrease of 4% in the rate was observed for Acioa barteri tannins at this level. At 0.23 mg and 0.47 mg of tannins per ml, the decrease in rate varied from 4 to 13% and 13 to 20%, respectively. The decrease in in vitro true digestibility of dry matter was 3, 6 and 7% for Q incana, D cinerea and A barteri, respectively, at a tannin concentration of 0.47 mg ml^?1, and 17, 21 and 27%, respectively, at 0.93 mg ml^?1. There was no change in the potential extent of digestion up to 0.47 mg ml^?1 for any of the tannins studied. These results suggested that the rate of digestion is affected to a greater extent than the potential extent of digestion or the in vitro true digestibility and that different tannins even at the same level have different degrees of effect. Tannin-saponin interactions were studied using tannic acid (TA), quebracho tannin (QT) and quillaja saponin (S) at different concentrations (0.2, 0.4 and 0.6 mg ml^?1) alone and in combination with each other. The decrease in the rate of digestion and true digestibility was additive when both the TA and the S were present. Similar results were obtained when TA was replaced by QT except that the effects with QT were more marked as compared to TA at the same concentration. Saponins had little effect on the potential extent of digestion and true digestibility. These tannins also decreased the production of short-chain fatty acid (SCFA) and their molar proportions (acetate decreased whereas propionate increased). Efficiency of microbial protein synthesis, expressed as the ratio of ¹⁵N incorporation per unit of SCFA production, was higher with both S and the tannins studied. A substantial amount (71–93%) of tannins soluble in aqueous acetone was released from leaves of some trees and shrubs on incubation in the in vitro medium for 48 h. The rumen liquor was not capable of degrading oligomeric condensed tannins.