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Herbivores learn to select compounds that attenuate the aversive effects of plant secondary metabolites (PSM), but can they increase intake of PSM they typically avoid when these PSM provide medicinal effects? We hypothesized that herbivores learn to increase intake of PSM-containing feeds when experiencing a gastrointestinal parasitic infection. Ten lambs with natural gastrointestinal parasitic burdens (PB) and 10 nonparasitized lambs (NP) were offered a choice of alfalfa (Medicago sativa) and alfalfa mixed with 10% quebracho tannin (Schinopsis quebrachocolorado; alfalfa:tannins) before and after they were conditioned with the postingestive effects of tannins. Preference for alfalfa:tannins did not differ between groups before experiencing the postingestive effects of tannins (P = 0.85) or when parasite loads were terminated due to the administration of ivermectin (P = 0.18). In contrast, when tested with a parasite burden, lambs in PB consumed more alfalfa:tannins (P = 0.08), showed greater preference for alfalfa:tannins (P = 0.07), and consumed less alfalfa than lambs in NP (P = 0.06). Ingestion of tannins by lambs in PB was followed by reduced fecal egg counts (FEC; P = 0.006), and there was a direct proportional relationship between preference for alfalfa:tannins and FEC (P = 0.07). In summary, parasitized lambs increased their intake of alfalfa:tannins when they experienced a parasite burden, which suggests they self-medicated with tannins against parasites. Self-selection of PSM has implications for the quest for alternatives to chemoprophylaxis in the treatment and well-being of parasitized wild and domestic animals grazing in pasturelands and in confinement. Key words: diet selection, helminth, Ovis aries, parasitism, tannin doi: 10.2527/jas.2009-2272

Tanniferous temperate legumes are assumed to possess anthelmintic properties, but it is unclear whether this is the direct result of condensed tannins (CT) or is mediated indirectly via an improved metabolic protein supply. A metabolism experiment was conducted to differentiate between these factors by feeding the CT plant sainfoin (19.7% CP in DM) to lambs infected with the abomasal blood-sucking nematode Hemonchus contortus. A total of 18 infected lambs were fed sainfoin either untreated or treated with polyethylene glycol, a CT-inactivating agent, or a grass-clover mixture (13.2% CP in DM) over 3 wk (n = 6). Six uninfected lambs received the grass-clover mixture as a control. In addition to indicators of infection (fecal egg count, packed-cell volume, abomasal worm burden, and serum protein), nutrient digestibility, the balance of N and selected minerals, ruminal fluid characteristics, and plasma AA levels were determined mostly in the final experimental week. The specific effects of the sainfoin CT, the extra CP with sainfoin, and the infection were statistically evaluated by contrast analysis. The sainfoin CT exerted no beneficial effects on resilience to nematode infection and exerted only minor effects on ruminal ammonia or blood urea concentrations and the excretory pattern of N. Plasma alanine, aspartate, and proline concentrations tended to be greater (P [less than or equal to] 0.09) because of the sainfoin CT, whereas the other AA remained unaffected. Intake of the mineral supplement was lower (P < 0.001) for lambs fed sainfoin compared with those fed sainfoin treated with polyethylene glycol. Feeding the high-protein sainfoin instead of the grass-clover mixture increased (P < 0.001) N retention and apparent OM digestibility, whereas digestibility of NDF and ADF were decreased (P < 0.001). Feeding sainfoin also decreased (P [less than or equal to] 0.04) plasma alanine, glycine, isoleucine, and total nonessential AA compared with the grass-clover mixture. Although fecal egg count, worm burden, and packed cell volume were not affected by the greater CP supply associated with sainfoin feeding, serum albumin level was increased (P = 0.008). The lack of effects of sainfoin on resilience to nematode infection might have been the result of the unexpectedly low CT content (3.6% in DM) of the material used. It cannot be excluded that longer term feeding of this batch of sainfoin might have been effective. Key words: Haemonchus contortus, nitrogen, polyethylene glycol, ruminant, sainfoin, sheep

An extracellular tannase was produced from solid-state cultures of Aspergillus roger. The enzyme was purified to homogeneity from the cell-free culture broth by preparative isoelectric focusing and by FPLC using anion-exchange and gel-filtration chromatography. SDS-PAGE analysis as well as gel localization studies of purified tannase indicated the presence of two enzyme forms, with molecular masses of 90 kDa and 180 kDa. The tannase had an isoelectric paint of 3.8, a temperature optimum of 60-70[degrees]C and a pH optimum of 6.0. The substrata specificity of the tannase was determined by HPLC analysis of tannin substrates and products. The enzyme was able to remove gallic acid from both condensed and hydrolysable tannins. Internal sequences were obtained from each of the gel-purified and trypsin-digested tannese forms. The peptide sequences obtained from both forms were identical to sequences within a [beta]-glucosidase from Aspergillus kawachii. The purified tannase was tested for [beta]-glucosidase activity and was shown to hydrolyse cellobiose efficiently, However, no [beta]-glucosidase activity was detected when the enzyme was assayed in the presence of tannic acid.

We determined if supplemental macronutrients or polyethylene glycol (PEG) influenced intake of a tannin diet. Sheep (lambs 5 mo age, 36 kg) and goats (kids 7 mo age, 32 kg) were fed supplements high in either energy or protein or offered a choice between the two supplements before and after receiving a meal containing 15% quebracho tannin. The effect of PEG, a compound that attenuates the negative effects of tannins, was assessed by offering PEG while animals consumed the tannin diet for 4 h/d. Intake of the tannin diet was influenced by both macronutrients and PEG. Animals that chose their own supplements or that received the high-protein supplement, consumed more of the tannin diet than animals fed the high-energy supplement: 34 and 36 vs 31 g/[kg.sup.0.75] (lambs) and 41 and 39 vs 34 g/[kg.sup.0.75] (kids), respectively (P < 0.05). Animals supplemented with PEG ate much more of the tannin diet than unsupplemented animals: 70 vs 39 g/ [kg.sup.0.75] (lambs) and 63 vs 34 g/[kg.sup.0.75] (kids), respectively (P < 0.001). Sheep and goats consumed more tannin food when given PEG than when supplemented with macronutrients (51 and 38 g/[kg.sup.0.75], P < 0.001). Sheep and goats offered a choice between supplements consumed more CP than animals fed the high-energy supplement and more ME than animals fed the high-protein supplement (P < 0.05). In so doing, they selected a combination of foods that yielded a more balanced intake of macronutrients, while achieving high levels of intake of the tannin food. Sheep and goats can be used as an environmentally safe and economically sound means to reduce the abundance of tannin-rich vegetation. Macronutrients and PEG enhance use of tannin-containing plants, which may increase production of alternate forages and create a more diverse mix of species in a plant community. Key Words: Food Preference, Goats, Polyethylene Glycol, Sheep, Supplements, Tannins

Research has been conducted on the tannin-like p-tert-butylcalix[4]arene 1,3-digallate. The synthesis of this compound has been carried out and its conformational property has been investigated via the dynamic (super)1 H NMR and X-ray crystallography.

We conducted an experiment to evaluate the effects that a diet containing condensed tannins with and without polyethylene glycol (PEG) has on lamb growth and meat quality. Twenty-three male Comisana lambs were introduced to the three experimental diets between 45 and 50 d of age. Eight were given a diet containing 56% of Ceratonia siliqua (carob) pulp (tannin group), another eight received the same diet with a supplement of 40 g of PEG for each kilogram of diet (PEG group), and the remaining seven lambs were given a conventional maize-based diet (maize group). Voluntary feed intake and live weights were measured until slaughter at 105 d of age, and digestibility measurements were undertaken toward the end of the feeding trial. Carcass yield, meat quality characteristics, and a taste panel evaluation were conducted. The tannin-based diet contained 2.5% condensed tannins (DM basis), and lambs given this diet had lower growth rates and poorer feed efficiencies (P < .01) compared with the other treatment groups. Daily gain was similar between the maize and PEG lambs, although the efficiency of feed conversion was highest in the maize group. The digestibility of DM, N, and fiber was reduced (P < .05) by the condensed tannins. Lambs fed the tannin diet had a lower carcass yield (P < .05) and had less fat (P < .05), and the meat had a higher ultimate pH (P < .01) than those given the PEG or maize diets. Condensed tannins affected meat color, which was lighter (L*) than meat from lambs given the PEG-containing diet (P < .01). Sensory evaluation showed that panelists preferred meat from lambs receiving PEG and maize treatments compared with those receiving the tannin diet, and this could be related to differences in meat ultimate pH and carcass fatness. These results show that condensed tannins from carob pulp are very detrimental to feed digestibility and lamb performance. Inclusion of 40 g of PEG/kg diet eliminated the effects of condensed tannins so that lamb performance and meat quality were similar to lambs given a maize-based diet. Key Words: Carobs, Meat Quality, Polyethylene Glycol, Sheep, Tannins

Effect of tannic acid on sodium ([Na.sup.+]) balance of captive meadow voles (Microtus pennsylvanicus) was studied by measuring size of the zona glomerulosa of the adrenal gland, a reflection of [Na.sup.+] status, in voles provided diets containing three levels of [Na.sup.+] and either 0, 1.5, or 5% tannic acid at each level of [Na.sup.+]. [Na.sup.+], but not tannic acid, had a significant effect on size of the zona glomerulosa. The zona glomerulosa hypertrophied significantly in voles on diets containing the two lowest levels of [Na.sup.+]. Both tannic acid and [Na.sup.+] had significant effects on relative body mass, indicating a nutritional constraint. However, results of this study do not support the widely cited hypothesis that tannins, such as tannic acid, disrupt [Na.sup.+] balance in herbivores. Key words: Microtus pennsylvanicus, meadow voles, sodium, mineral balance, tannins, tannic acid, adrenal glands

Tannic acid was investigated to find out whether it elicits neurogenic plasma exudation from the oral mucosa in vivo in hamsters. Using intravital microscopy, it was observed that suffusion of tannic acid elicits substantial concentration-dependent leaky site formation and increase in clearance of fluorescein isothiocyanate-dextran from the in situ hamster cheek pouch. Results indicated that tannic acid elicits L-arginine-NO biosynthetic pathway-dependent neurogenic plasma exudation from the in situ hamster cheek pouch.

Japanese researchers continue to study the health benefits of drinking green tea. Green tea was imported from China in the 12th Century and is now very common. Researchers found that one of the principal tannins in green tea is a compound called (-)-epigallocatechin gallate (EGCG). There is much evidence that EGCG can prevent carcinogenesis. It inhibits carcinogenesis in skin, stomach, colon, liver, pancreas, lung and breast. Green tea consumption also prolongs the survival of cancer patients.

Dietary tannin in six captive food hoarding granivores, Western Scrub-Jays, suppresses protein, and the Jays are unable to physiologically counteract these inhibiting effects completely. When exposed to dietary lipids and proteins in artificial diets varying in tannin activity, the Jays show loss of mass with low-protein diet, and loss of most mass with a low-protein high tannin diet. These birds fail to show any significant changes in weight with lipid treatment.

We investigated the ability of the meadow vole, Microtus pennsylvanicus, to produce tannin-binding salivary proteins when fed quebracho tannin-containing diets or when injected with the drug isoproterenol. None of the treatments induced production of new salivary proteins. None of the salivary proteins from voles bound tannin in vitro, and none of the proteins in vole saliva reacted with antibodies to tannin-binding proteins from saliva of rats. Diets containing 2.9% crude quebracho tannin had no effect on growth or survival of voles. Voles consuming diets containing 6% crude quebracho tannin had increased rates of loss of body mass and mortality and decreased rates of food consumption early in the trial, but began to accommodate to the diet by the end of the 10-day trial. Voles fed diets containing 6% tannin prepared by moistening and heating a mixture of lab chow and quebracho tannin had significantly higher rates of mortality than voles fed 6% tannin added directly to dry chow. Nitrogen, dry matter, and caloric digestibilities did not differ significantly among treatments. Key words: Microtus pennsylvanicus, meadow voles, condensed tannins, plant phenolics, secondary chemicals, salivary proteins

Secondary compounds in fruits are thought to be a defense against invertebrates or pathogens which might otherwise damage or destroy the fruits or seeds without contributing to seed dispersal. We incorporated ground, nonripe green or ripe red fruits of American holly Ilex opaca into standard diets of two species of polyphagous caterpillars to test the hypothesis that nonripe fruits are better defended than ripe fruits. Fall armyworm Spodoptera frugiperda larvae grew more slowly on diet augmented with 10% (dry wt) ripe fruits than on control diet with 10% cellulose powder, and slower still on diet with nonripe fruits. However, survival, pupal weight and length of the pupal period were not affected by either treatment. Growth rates and survival of fall webworms Hyphantria cunea were reduced to a similar degree by both nonripe and ripe fruits. Methanolic extracts of nonripe fruits brushed on leaf disks of sassafras, a preferred host, deterred feeding by Japanese beetles Popillia japonica Newman, a highly polyphagous species. Extracts of ripe fruits were not repellent. Saponin content was highest in nonripe green fruits, while tannin levels were highest in ripening fruits. In choice tests, red holly fruits were removed from bird feeders at a faster rate than nonripe green fruits. Levels of secondary compounds in fruits of I. opaca may be a compromise between the continued need to protect the seeds from predispersal damaging agents without deterring potential vertebrate dispersers.

Condensed tannins (CT) in birdsfoot trefoil (Lotus corniculatus L.) are associated with reduced degradation of protein and dry matter digestibility by ruminant animals. The objectives of this study were to isolate the effect of CT concentration on in vitro degradable crude protein (IVDCP) and in vitro digestible dry matter (IVDDM) in birdsfoot trefoil herbage containing diverse CT concentrations, to determine the relative influence of IVDCP compared with IVDDM, and to determine minimum CT concentrations effecting reductions in IVDCP and IVDDM. Twelve birdsfoot trefoil clones, genetically diverse for CT concentration, were harvested at St. Paul and Rosemount, MN. Herbage samples with uniform fiber concentrations were arrayed in five tannin groups (n = 3) at regular intervals from minimum to maximum CT concentrations at both locations. Herbage samples were compared for differences between polyethylene glycol (PEG) and control treatments for IVDCP and IVDDM analyses. The protease ficin was used to determine IVDCP. As CT concentration increased, the difference between PEG and control treatments increased for IVDCP and IVDDM ([R.sub.2] = 0.91 and 0.69, respectively). For tannin groups with mean CT concentrations [is greater than or equal to] 27.0 g catechin equivalent (CE) [kg.sup.-1] DM, the PEG and control treatments differed for IVDCP compared with tannin groups containing minimum CT concentrations at both locations. For the tannin group with the maximum CT concentration at Rosemount (85.3 g CE [kg.sup.-1] DM), the PEG and control treatments differed for IVDDM compared with the tannin group containing the minimum CT concentration. It was concluded that paired-subsample comparisons between PEG and control treatments was a convenient and consistent method for separating the effect of CT concentration from other sources of variation. These data imply that CT concentrations in the range of 25 to 85 g CE [kg.sup.-1] DM may reduce ruminal protein degradation, with little or no corresponding reduction in dry matter digestibility.

The effects of condensed tannins on the digestion of cellulose by the rumen bacteria Fibrobacter succinogenes S85 were determined. The results showed that condensed tannins induced a transient increase in the activity of cell-associated endoglucanases, accompanied by a decrease in the activity of extracellular endoglucanases. Howver, as the concentration of condensed tannins approached 400 micrograms per milliliter, all endoglucanases were inhibited. These results confirm earlier reports on the inhibitory effects of condensed tannins on cellulose digestion.

Condensed tannins reportedly act as feeding deterrents to bollworm (Helicoverpa zea (Boddie)), tobacco budworm (Heliothis virescens (F.)), and twospotted spider mite (Tetranychus urticae (Koch)) in cotton (Gossypium hirsutum L.). This study was conducted to determine the effects of planting date, plant density, growth stage, and leaf age on the condensed tannin concentration of six cotton genotypes planted at College Station, TX, in 1988 and 1989. Planting commenced on 25 April in both years and consisted of three plantings at approximately 2-wk intervals. The topmost unfolded leaf from the terminal and the fourth mainstem leaf from the terminal were sampled at five growth stages from plants spaced 8 and 30 cm, within rows spaced at 102 cm. Genotypes included two commercial checks and four high-tannin breeding lines developed in a spider mite resistance program. Condensed tannin concentration, expressed as a percentage of fresh weight, was determined spectrophotometrically following extraction with HCl-butanol. The younger leaf tissue had significantly higher levels of condensed tannins than the more mature leaf tissue. Condensed tannin concentration increased with plant age through the first-bloom growth stage. The condensed tannin concentration of plants sown on Planting Dates 1, 2, and 3 did not differ in 1988, but in 1989, plants from Planting Date 2 had higher condensed tannin concentrations than those from Planting Dates 1 or 3. Plant density did not significantly affect condensed tannin concentration. Many interactions of other factors with genotypes suggest that breeding for high tannin levels would require close attention to leaf age, growth stage, and environment.

Bacteria capable of degrading tannin-protein complexes (T-CP) were isolated from the feces and cecal wall of koalas. The predominant strains were shown to belong to the Enterobacteriaceae based on morphological features and biochemical reactions. A selective medium, based on Wilkins-Chalgren anaerobe agar supplemented with arginine, tannin and vancomycin, was developed for these isolated strains. The medium supported the growth of the enterobacteria while excluding Streptococcus bovis, another common digestive tract T-CP degrader.

Mule deer (Odocoileus hemionus), domestic sheep (Ovis aries), and American black bears (Ursus americanus) were fed quebracho (Schinopsis sp.) tannin to determine the contribution of salivary proteins to nitrogen-and fiber-digestive efficiencies and tannin metabolism. These values were compared to previously published values for laboratory rats (Rattus rattus) and prairie voles (Microtus ochrogaster). Mule deer, black bears, and laboratory rats consuming this condensed tannin produced tannin-binding salivary proteins that reduced fecal-nitrogen losses per unit of ingested and reduced tannin metabolism relative to domestic sheep and prairie voles. Digestibility of the plant fiber was reduced significantly by tannins in domestic sheep, but not in mule deer. Although virtually all ingested tannin (98.3 [+ or -] 5.0%) was recovered in feces of mule deer and black bears, ca. 25% was not recovered in feces of domestic sheep and presumably was metabolized. The defensive role of tannins as digestion inhibitors or toxins is dependent upon the molecular characteristics of the tannin interacting with the physiological capability of the animal. Results from one plant-mammal interaction cannot be used to interpret others without an understanding of the characteristics of the tannins and the physiology, ecology, and evolution of the animal.

A study was conducted to assess the effects of tannins on ruminal biohydrogenation (BH) due to shifts in the ruminal microbial environment in sheep. The obtained results indicated that quebracho tannins altered BH by changing ruminal microbial populations.

Research has been conducted on tannins, polyphenols with antimicrobial effects. The study of the tannin toxicity mechanism toward Escherichia coli has been carried out via the use of Acacia mearnsii extract as a condensed tannin source, and the details are reported.

Tannins occur in many plant species, and they often suppress intake by reducing nutrient availability or by causing malaise. Polyethylene glycol (PEG) binds to tannins and may thereby increase the availability of macronutrients and decrease malaise. Supplemental PEG increases intake of tannin-containing plants by sheep, goats, and cattle. Given the strong response to supplemental PEG, we speculated that animals might self-regulate their intake of PEG when offered foods high in tannins. The objective of the first experiment was to determine if the amount of supplemental PEG (0, 25, 50, 75, or 100 g; molecular weight, 3,350) affected intake by lambs of a food (milotannin mix) containing 20% quebracho tannin. There was a linear relationship (Y = 272 + 1.2X; [R.sup.2] = .86; P = .023) between the amount of supplemental PEG ingested and the subsequent intake of milo-tannin food by lambs. The objective of the second experiment was to determine whether lambs self-regulated intake of PEG when fed a ration that contained 0, 5, 10, 15, or 20% quebracho tannin and whether they adjusted their intake of PEG when tannin was removed from the diet. There was a positive relationship between the amount of PEG ingested and intake of food and tannin (P = .0001). Lambs fed high-tannin diets ate more PEG than controls (P = .03). Lambs fed the 20% tannin diet ate the most PEG, and controls ate the least PEG. Tannin limited intake of the diets, but PEG attenuated the response to a great degree (P = .065). Immediately after tannin was removed from the ration, lambs that formerly had been fed the 20% tannin ration ate more PEG than lambs fed the other rations (P = .0075). Ten of the lambs (5 from the 20% tannin group, 1 from the 15% tannin, and 2 each from the 10 and 5% groups) continued to eat PEG for 7 d after tannin was removed from their ration. When they were tested again 6 wk after the trial and offered tannin-free diets, their intake of PEG had decreased. Key Words: Intake, Polyethylene Glycol, Sheep, Tannins

THE EFFECTS OF TANNINS ON FREE RADICALS. Twaskia S. Johnson and William N. Setzer, University of Alabama in Huntsville, Department of Chemistry, Huntsville, AL 35899. The use and search for [...]

Tannins are polyphenolic compounds in plants that readily bind protein and can reduce palatability, digestibility and cause toxicity. Many mammals that come in contact with tannins in their diets have developed the ability to secrete praline-rich proteins (PRPs) in their saliva that preferentially bind to tannins. It is thought that the PRPs bind to tannins forming indigestible complexes that escape both enzymatic and microbial degradation and are excreted in the feces; thus protecting the animal from digestibility reduction and metabolic costs associated with the detoxification of absorbed tannin components. However, no one has shown quantitatively that PRPs increase fecal tannin excretion while decreasing detoxification products in urine. Thus we tested whether the presence of salivary PRPs would decrease the metabolic cost of pentagaloyl glucose (PGG - a hydrolysable tannin) consumption in laboratory rats. We used radiolabeled pentagaloyl [C14] glucose to determine if rats secreting PRPs excreted more radiolabeled tannin in their feces than rats not secreting PRPs. We also collected urine from the rats to determine if the rats secreting PRPs excreted less glucuronic acid (a measure of detoxification) in their urine than rats not secreting PRPs. We found that rats which were secreting PRPs excreted four times more of the ingested radiolabel in their feces than rats that were not secreting PRPs (20.5+/-5.1% vs. 5.0+/-3.3%). We also found that rats which were secreting PRPs excreted less than half as much glucuronic acid in their urine as rats that were not secreting PRPs (11.9+/-5.5 vs. 28.4+/-19.0 mg glucuronic acid/g PGG consumed). Thus we conclude that the presence of salivary PRPs does decrease the metabolic costs of PGG consumption in rats. Supported by USDA (Hatch) WISO4322 NSF IBN-9318675 IBN-9723793