Condensed tannins,attine ants,and the performance of a symbiotic fungus

Field experiments indicate that the foliar concentration of condensed tannin affects the selection of leaf material ofInga oerstediana Benth., a tropical legume tree, by leaf cutter ants. In one study an increase in tannin concentration was correlated with a decrease in the acceptability of leaves to leaf-cutter ants, except at low tannin concentrations. Protein concentration was not correlated with acceptability nor was the ratio of protein to tannin. Results from a second study suggest that when the concentration of tannin was low the ants appear to select leaves on the basis of nutrient availability. Laboratory assays with the ants indicated that quebracho tannin, a commercially available condensed tannin, inhibits foraging ants. Again, at lower concentrations, quebracho tannin appeared to have little affect on the ants. The fungus the ants cultivate is a wood-rotting Basidiomycete that produces enzymes, such as polyphenol oxidase (PPO), that are capable of inactivating tannins. The activity of these PPOs may explain why leaf-cutter ants are undeterred by low concentrations of condensed tannins. I hypothesized that PPO activity would be absent from fungal cultures without tannin and that only high concentrations of tannin would inhibit the fungus. Cultures with and without tannin showed similar PPO activity. Thus PPO activity is constitutive. In fact, as fungal biomass increased, so did PPO activity. As hypothesized, only high concentrations of quebracho tannin inhibited PPO activity and fungal growth. However, it is not clear whether the ants can discriminate between concentrations that do and do not inhibit the fungus.     

Plant phenolics as chemical defenses: Effects of natural phenolics on survival and growth of prairie voles (Microtus ochrogaster)

Very few studies have shown experimentally that plant chemical defenses actually reduce the performance of individual mammalian herbivores, much less the density of mammalian populations. We investigated the effects of representatives of three classes of plant phenoiics on the survival and growth of prairie voles by incorporating the compounds into artificial diets and feeding them to weanlings for three weeks. At low levels of protein, both quercetin (a flavonoid) and tannic acid (a hydrolyzable tannin) caused reduced growth rates; no effect occurred at high levels of protein. Quebracho (a condensed tannin) inhibited feeding and thus was lethal at all levels of protein. These results indicate that plant phenolics are likely to influence the performance and dynamics of natural populations of microtine rodents by reducing the quality of available forage. The hypothesis that the primary mode of action of the phenoiics is the reduction of digestibility of protein was not supported. The reduced growth caused by both quercetin and tannic acid could be attributed primarily to their toxicity. The effect of quebracho resulted from reduced intake (unpalatability).     

Chemical and Biological Assays to Evaluate Bacterial Inhibition by Tannins

The interactions among purified tannins from three different plants with five strains of ruminal bacteria were investigated. Tannins were purified from crude quebracho (Schinopsis balansae spp.), desmodium (Desmodium ovalifolium), and myrtle (Mirtus communis). The ruminal bacteria studied were Streptococcus bovis JB1, Ruminococcus albus 8, Fibrobacter succinogenes S85, Prevotella ruminicola H₁4, and a recently isolated tannin-tolerant bacterium. Bacterial binding to tannins, rate of tannin binding, and the inhibitory concentrations for each bacterium to each tannin were evaluated. The effects of tannins on glucose utilization and fermentation product formation also were studied. Tannin binding was rapid with little additional binding occurring 10 min after tannin addition. There was variation among bacteria in the amount of tannins bound and in the amount of tannin required to inhibit growth. Condensed tannins from myrtle and desmodium had the highest binding capacity and were most inhibitory to microbes. Absorbance readings from the total phenolics assay were highest for myrtle and lowest for quebracho. Similarly, readings for condensed tannins were highest for desmodium, which also had the largest polymer size. Myrtle and quebracho gave the largest precipitate rings in the radial diffusion assay, a biological assay that measures precipitation of bovine serum albumin by tannins. The relative absorbance values obtained from the total phenolics assay were the most useful predictor of tannin inhibition of microbial growth. The acid–butanol assay and the radial diffusion assay using bovine serum albumin as the precipitable protein were much less useful predictors of tannin biological activity against the bacteria studied.     

Tolerance of acridids to ingested condensed tannin

Four species of Acridoidea were fed on wheat leaves with and without the condensed tannin, quebracho. In no case was it deleterious to survival and growth at levels below about 10% dry weight on the food. Similarly, consumption, digestibility, and utilization of food were unaffected at up to 10% dry weight. At higher concentrations, however, the consumption and the efficiency of conversion of digestion were reduced, although digestibility was little affected. The possible mechanisms for such tolerance are discussed, and contrast made with insects which are very sensitive to ingested condensed tannin.     

Effects of large mammalian herbivores and ant symbionts on condensed tannins of Acacia drepanolobium in Kenya

Condensed tannins have been considered to be important inducible defenses against mammalian herbivory. We tested for differences in condensed tannin defenses in Acacia drepanolobium in Kenya over two years among different large mammalian herbivore treatments [total exclusion, antelope only, and megaherbivore (elephants and giraffes) + antelope] and with four different ant symbiont species on the trees. We predicted that (1) condensed tannin concentrations would be lowest in the mammal treatment with the lowest level of herbivory (total exclusion), (2) trees occupied by mutualist ants that protect the trees most aggressively would have lower levels of tannins, and (3) if chemical defense production is costly, there would be a trade-off between tannin concentrations, growth, and mechanical defenses. Mean tannin concentrations increased from total exclusion treatments to wildlife-only treatments to megaherbivore + antelope treatments. In 1997, condensed tannin concentrations were significantly lower in trees occupied by the ant Crematogaster nigriceps, the only ant species that actively removed axillary buds. Contrary to our prediction, trees occupied by ant species that protect the trees more aggressively against mammalian herbivores did not have lower overall levels of condensed tannins. There was no consistent evidence of a trade-off between tannin concentrations and growth rate, but there was a positive correlation between mean thorn length and mean tannin concentrations across species of ant inhabitants and across herbivore treatments in 1997. Contrary to our expectation, trees had higher tannin concentrations in the upper parts of the canopy where there is little herbivory by mammals.     

Tannin sensitivity in larvae ofMalacosoma disstria (Lepidoptera): Roles of the peritrophic envelope and midgut oxidation

Final-instarMalacosoma disstria fed artificial diets containing tannic acid develop lethal pupal deformities. We examined some of the factors potentially underlying tannin sensitivity in this species, including the permeability of the peritrophic envelope to tannic acid and the chemical fate of tannic acid in the gut. Tannic acid does not penetrate the peritrophic envelope ofM. disstria, demonstrating that the containment of tannic acid within the endoperitrophic space is not sufficient to protect an insect herbivore from the adverse effects of ingested tannins. Ingested tannic acid undergoes extensive chemical modification in the midgut. Only 19–21 % of the high molecular weight components of the tannic acid ingested was recovered in the frass. Of two possible chemical fates of ingested tannic acid, oxidation is the predominant chemical transformation, whereas little hydrolysis occurs. Measurements of gut redox parameters showed that conditions in the midgut favor the oxidation of phenols. However, similar conditions occur in the midguts ofOrgyia leucostigma, in which no oxidation occurs. Therefore, oxidizing gut redox conditions do not necessarily lead to polyphenol oxidation in lepidopteran larvae. We conclude that the sensitivity ofM. disstria to ingested tannins is a consequence of their oxidation in the midgut.     

Reassessment of interaction between gut detergents and tannins in lepidoptera and significance for gypsy moth larvae

Surface-active lysophospholipids are thought to interfere with tannin-protein interactions and may comprise an adaptation in Lepidoptera to tannin-rich diets. We found several lysophospholipids and phospholipids at about eight times the critical micelle concentration (CMC) in the midguts of gypsy moth larvae, which exhibited appropriately reduced surface tensions. We confirmed the interfering activity of lysophosphatidylcholine (lysolecithin) in a model astringency assay using tannic acid and hemoglobin (hemanalysis), but discovered that tannic acid and red oak tannin extracts precipitate the lipid from simulated midgut solutions quantitatively, even at pH 10. Leaf tannin extracts from trees on which gypsy moths grew and reproduced poorly precipitated lysolecithin more effectively than did extracts from trees on which gypsy moths performed well. Adding tannic acid to midgut fluid elevated surface tension, and about 25% of larvae feeding on oak leaves exhibited elevated midgut surface tension, suggesting a loss of surfactants. Larvae appear able to replace lost surfactants to a limited degree. An important effect of leaf tannins, and perhaps other phenolics, may be to reduce concentrations of surface-active phospholipids in the midgut and produce lipid or other dietary deficiencies in insects.     

Loss of Tannins and Other Phenolics from Willow Leaf Litter

A litter bag experiment was conducted to determine the fate of condensed tannins during the initial phase of willow leaf decomposition. Chemical methods and HPLC were used to identify and quantify the phenolics in aqueous acetone extracts of the leaves during the first eight weeks after leaf fall. Extractable phenolics and tannins were lost rapidly from the leaves and had half lives of about 2.4 weeks. Lower molecular weight tannins were lost more rapidly than higher molecular weight tannins, suggesting that the primary route for loss of tannins is leaching. There was no evidence for decomposition of the tannin during the leaf decay process. After leaching, the tannin apparently sorbs tightly to the soil.     

Tannin Composition Affects the Oxidative Activities of Tree Leaves

We examined whether tannin composition plays an important role in explaining the oxidative activities of tree leaves of Acer saccharum (sugar maple) and Quercus rubra (red oak). Sugar maple leaves contained substantial amounts of ellagitannins, condensed tannins, and galloyl glucoses, whereas red oak leaves contained almost exclusively condensed tannins. Oxidative activities of the crude phenolics from both species, and the phenolic fractions from sugar maple, were measured with electron paramagnetic resonance (EPR) spectrometry and UV-visible spectrophotometry. The two assays produced similar results: (1) sugar maple phenolics produced larger semiquinone radical concentrations,and higher semiquinone decay rates and browning rates than did red oak phenolics;(2) ellagitannin levels were positively associated with the three measures of oxidative activity; and (3) condensed tannin and galloyl glucose levels were negatively associated with these measures. The negative relationship between condensed tannin levels and oxidative activity resulted from the antioxidant effects of condensed tannins on hydrolyzable tannins; several purified condensed tannins significantly decreased the concentrations of semiquinone radicals and browning rates of pedunculagin (an ellagitannin) and pentagalloyl glucose. As expected, whole-leaf extracts from sugar maple produced elevated levels of semiquinone radicals, but none were observed in red oak extracts when the two species were compared with an EPR time-course assay. We conclude that the oxidative activities of tree leaves may be affected by tannin composition, and that the prooxidant activity of ellagitannins may be decreased by co-occurring condensed tannins.     

Inhibition of Cyanogenesis by tannins

During isolation of two biosynthetic types of cyanogenic glycosides fromCarica papaya, weak cyanide tests were obtained from initial fractions. Upon final purification, strongly positive cyanide tests were obtained. Pretreatment of extracts to remove polyphenolics alleviated inhibition of cyanogenesis, which led us to suspect that tannins were inhibitory agents. Qualitative and quantitative measures of inhibition were made using standard cyanogenic glycosides and polyphenolics. Cyanogenesis was inhibited quantitatively when condensed tannins (quebracho, wattle, and chestnut), or hydrolyzable tannin (tannic acid) were added. When tannins were precipitated from the reaction mixture, cyanide tests proceeded optimally. These results stress the need to interpret negative cyanide tests with care and indicate possible ecological synergisms between plant defensive chemicals.     

Chestnut bark tannin assays and growth of chestnut blight fungus on extracted tannin

Tannins extracted from the green bark of each of two Chinese, Japanese, and American chestnut trees were assayed in a protein-binding test. Four levels of tannins were added to a buffered, minimal growth medium, and a standard, virulent strain of the chestnut blight fungus was grown. There were only slight differences in protein binding between the extracts from different species. Fungal growth was better with tannin than without, but there was no difference between species extracts in their ability to improve fungal growth rate. There was also no inhibition of blight fungus growth by any of the tree tannins, so tannin toxicity is not the reason for Asian chestnut tree resistance.     

Factors affecting levels of some phenolic compounds,digestibility, and nitrogen content of the mature leaves ofBarteria fistulosa (Passifloraceae)

Levels of total phenolics, condensed tannins, acid detergent fiber, pepsin/cellulase digestibility, and nitrogen in mature leaves of 26 individuals of the ant-plant,Barteria fistulosa, have been determined. Analysis of the results in terms of the presence or absence of ants and the position of the branch from which the leaves were collected showed no relationship with concentrations of phenolics or fiber and only a weak relationship with digestibility and nitrogen. By contrast, light intensity strongly influenced levels of phenolics, notably condensed tannins, so that mature leaves of individuals growing in direct sunlight were less digestible and appeared to be of lower quality as food for herbivores than did mature leaves of individuals in shaded positions. Possible reasons for the variation in condensed tannin levels are discussed.     

Effects of addition of condensed tannin on the structure and properties of silk fibroin film

Condensed tannin is a polymeric polyphenol compound, which has considerable protein‐binding ability and a variety of bioactivities, such as anti‐inflammatory, antithrombotic and antimicrobial activities. In this study, silk fibroin film was modified by adding condensed tannin. It was found that strong intermolecular interactions occurred between silk fibroin and tannin, and incorporation of 1–10% (w/w) tannin in the blend films resulted in more compact, stiffer but less elastic films. Swelling and dissolution trials showed that the addition of tannin significantly reduced the swelling and solubility of the films. The mechanical properties, swelling and solubility of the blend films varied as a function of tannin concentration, showing optimum values when containing 10% (w/w) tannin. The silk fibroin–tannin films all showed significant antioxidant activity and antibacterial activity against Gram‐positive bacteria, and the activities were positively related to the concentration of tannin. The results emphasized that the addition of condensed tannin could simultaneously improve the structural stability of silk fibroin film and impart functional properties. © 2016 Society of Chemical Industry     

Effects of Acacia condensed tannins on urinary parameters,body mass,and diet choice of an Acacia specialist rodent,Thallomys nigricauda

The aim of this study was to investigate the dietary and physiological effects of condensed tannin ingestion on foregut fermenters, using Thallomys nigricauda, a folivorous rodent, as a model. We initially investigated the variability in physiological parameters, such as daily body mass (DM_b), daily feed intake, daily fecal energy loss (FE), daily energy intake (DEI), daily urine pH, and daily urinary ammonia and urea concentrations, in response to different diets with low condensed tannin levels. This experiment was conducted to identify which physiological variables showed the least variation in the absence of tannin. In a second experiment, we investigated the response of the same dietary and physiological parameters to the effects of high dietary condensed tannin ingestion in T. nigricauda. We hypothesized that DM_b, daily feed intake, FE, and DEI of T. nigricauda would be adversely affected by high dietary tannin content. We predicted that detoxification activity by T. nigricauda would increase at higher tannin levels. Ingestion of tannins affected the nutritional status of T. nigricauda, as shown by a decrease in body mass at high tannin levels. We also found that fewer ammonium ions were excreted in the urine by T. nigricauda, as would be expected if this were a means of regulating metabolic acidosis. The urine produced was more alkaline. This result indicates that T. nigricauda is not metabolizing these allelochemicals. Urea production was initially reduced, indicating conservation of bicarbonate ions that will neutralize blood acidity if there is detoxification. A diet choice experiment showed that tree rats avoid high tannin diets, even to the extent that they lose body mass on an alternative diet. This last-mentioned result is noteworthy because previous studies of the effects of tannins on herbivorous mammals have shown that there is physiological control rather than behavioral avoidance of the negative effects of tannin ingestion.     

Effect of Nitrogen Availability on Expression of Constitutive and Inducible Chemical Defenses in Tomato, Lycopersicon esculentum

Young tomato plants (Lycopersicon esculentum) grown in sand in a greenhouse and subjected to different fertilization regimes were used to test the effects of nitrogen availability on constitutive levels of phenolics and on constitutive and inducible activities of polyphenol oxidase and proteinase inhibitors. Theories that emphasize physiological constraints on the expression of phytochemicals predict an increase in levels of carbon-based allelochemicals under moderate nitrogen stress but predict, under the same conditions, an attenuation of chemical responses involving nitrogen-containing compounds such as proteinase inhibitors and polyphenol oxidase. We found that nitrogen availability had a strong effect on constitutive levels of phenolics; weaker effects on constitutive polyphenol oxidase activity, constitutive proteinase inhibitor activity, and inducible polyphenol oxidase activity; and no effect on inducible proteinase inhibitor activity. These results point to a need for the integration of theories that emphasize physiological influences on secondary metabolism with those that emphasize ecological influences on secondary metabolism and suggest that current theories of plant defense do not adequately account for enzymatic and proteinaceous defenses against arthropods.     

Reassessment of the roles of the peritrophic envelope and hydrolysis in protecting polyphagous grasshoppers from ingested hydrolyzable tannins

We examined several of the mechanisms that have been reported to enable polyphagous grasshoppers (Orthoptera: Acrididae) to tolerate ingested hydrolyzable tannins: hydrolysis, adsorption on the peritrophic envelope, and peritrophic envelope impermeability. None of these mechanisms explain the tolerance ofMelanoplus sanguinipes to ingested tannic acid. In this species, tannin hydrolysis was 12–47% complete, adsorption accounted for less than 1% of the tannic acid contained in the midgut, and the peritrophic envelope was permeated by several gallotannins. The foregut is the main site for the chemical transformation of tannic acid in this species. InPhoetaliotes nebrascensis, hydrolysis was more extensive (82% complete), but the peritrophic envelope was readily permeated by two gallotannins. Oxidizing redox conditions were found in the guts of both species, and ingested tannins were oxidized inM. sanguinipes. We hypothesize that the tolerance of some polyphagous grasshoppers to ingested hydrolyzable tannins may be the consequence of their ability to tolerate the reactive oxygen species generated by polyphenol oxidation, whereas others may rely on rapid and extensive hydrolysis.     

Effects of Purified Persimmon Tannin and Tannic Acid on Survival and Reproduction of Bean Bug, Riptortus clavatus

We evaluated the effects of tannic acid and purified perrsimmon tannin on survival and reproduction of bean bugs, Riptortus clavatus.Feeding behavior of R. clavatus was also examined on sweet (cv. Fuyu) and astringent (cv. Chongdosi) persimmon fruits. Soluble tannin in sweet persimmon fruits decreased from 3% in early June to 0.5% in late September, but it increased from 2 to 8% during the same period in astringent persimmon fruits. More bugs visited sweet than astringent persimmon. Numbers of piercing/sucking spots were higher on sweet than on astringent persimmon. When fed 1 and 3% solutions of persimmon tannin, adult bugs ingested only 64.1 and 9.5% of the amount of water ingested by those offered the control (distilled water). Amounts of persimmon tannin ingested by the adult bugs were 6.5 and 2.8 times higher at 1 and 3% tannin solutions compared to a 0.1% solution. Persimmon tannin exerted negative effects on survival and reproduction of R. clavatus at higher concentrations (1 and 3% solutions). Feeding of R. clavatus adults decreased with increasing tannin concentrations. When results from both sexes were pooled, 50% mortality was achieved at 11 and 4 days after treatment with the 1 and 3% tannin solutions, respectively. Reproduction decreased with 1% tannin, and no eggs were produced with 3% tannin solution. Tannic acid was similar in its effects on R. clavatus. All nymphs died 14, 12, and 7 days after feeding on 0.1, 1, and 3% tannic acid, respectively. Adults were less sensitive than nymphs, and their survival was not affected by 0.1% tannic acid. However. 1 and 3% tannic acid solutions were fatal. Survivorship decreased to 50% at 11 and 6 days after supplying tannin solutions of 1 and 3% concentrations. Higher concentrations (1 and 3%) resulted in reduced reproduction, as was seen with persimmon tannin. Our data may explain why R. clavatus does not invade sweet persimmon orchards until late July, when concentrations of soluble tannin are low enough to allow them to feed.     

Effect of tannic acid concentration on development of the western treehole mosquito,Aedes sierrensis (Diptera: Culicidae)

Populations ofAedes sierrensis (Ludlow) completed development in tannic acid solutions over a wide concentration range (i.e., 0–1.0 g/liter) in artificial microcosms exposed to field conditions. The most notable effects of high tannic acid concentration were to slow larval developmental rates and to reduce numbers of adults produced; adult size and sex ratio were minimally affected. Vector potential of the western treehole mosquito is discussed in terms of tannin concentration.     

Synthesis,characterization, and thermal behaviors of tannin stearates prepared from quebracho and pine bark extracts

Reported is the preparation of various condensed tannin stearates and the associated chemical characterization and thermal properties of these products. Stearate esters of condensed tannins from both quebracho and pine bark extracts were prepared in generally high, isolated yields from reaction with stearic acid chloride. Tannin esterification was confirmed by both Fourier transform infrared and nuclear magnetic resonance (NMR), and the average degree of stearate substitution was calculated from ¹H‐NMR analysis. Product degree of substitution (DS) was observed to proportionately increase with higher stearic acid chloride ratio with maximum DS values of 4.0 and 5.1 achieved for quebracho and pine tannins, respectively. Thermal analysis revealed that tannin stearate products have increased thermal stability with a degradation onset at significantly higher temperature for those samples possessing relatively greater DS. Analysis using differential scanning calorimetry revealed isolated products to consist of multiple components which exhibit interesting melt behaviors, a likely result from their work up on isolation. However, multiple melt features of the individual components in products were lost on heating to give single, broad melt endotherms due to coalescence of sample components. Furthermore, tannin stearate samples with high DS show relatively greater endothermic melting at higher temperature than those samples with lower DS. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010     

高纯度食品级鞣酸的制备

采用溶剂萃取法，以五倍子为原料，水为溶媒进行浸提，再用乙酸乙酯为溶剂进行萃取，通过脱色制备高纯度食品鞣酸。结果表明，高纯度食品鞣酸的收率达60％，单宁酸含量达97．5％以上。