No Major Role for Binding by Salivary Proteins as a Defense Against Dietary Tannins in Mediterranean Goats

We investigated whether Mediterranean goats use salivary tannin-binding proteins to cope with tannin-rich forages by determining the affinity of salivary or parotid gland proteins for tannic acid or quebracho tannin. Mixed saliva, sampled from the oral cavity, or parotid gland contents were compared to the intermediate affinity protein bovine serum albumin with a competitive binding assay. Goats that consume tannin-rich browse (Damascus) and goats that tend to avoid tannins (Mamber) were sequentially fed high (Pistacia lentiscus L.), low (vetch hay), or zero (wheat hay) tannin forages. Affinity of salivary proteins for tannins did not differ between goat breeds and did not respond to presence or absence of tannins in the diet. Proteins in mixed saliva had slightly higher affinity for tannins than those in parotid saliva, but neither source contained proteins with higher affinity for tannins than bovine serum albumin. Similarly, 3 months of browsing in a tannin-rich environment had little effect on the affinity of salivary proteins for tannin in adult goats of either breed. We sampled mixed saliva from young kids before they consumed forage and after 3 months of foraging in a tannin-rich environment. Before foraging, the saliva of Mamber kids had higher affinity for tannic acid (but not quebracho tannin) than the saliva of Damascus kids, but there was no difference after 3 months of exposure to tannin-rich browse, and the affinity of the proteins was always similar to the affinity of bovine serum albumin. Our results suggest there is not a major role for salivary tannin-binding proteins in goats. Different tendencies of goat breeds to consume tannin-rich browse does not appear be related to differences in salivary tannin-binding proteins.     

Interspecific Differences in Tannin Intakes of Forest-Dwelling Rodents in the Wild Revealed by a new Method Using Fecal Proline Content

Mammalian herbivores adopt various countermeasures against dietary tannins, which are among the most widespread plant secondary metabolites. The large Japanese wood mouse Apodemus speciosus produces proline-rich salivary tannin-binding proteins in response to tannins. Proline-rich proteins (PRPs) react with tannins to form stable complexes that are excreted in the feces. Here, we developed a new method for estimating the tannin intake of free-living small rodents, by measuring fecal proline content, and applied the method to a field investigation. A feeding experiment with artificial diets containing various levels of tannic acid revealed that fecal proline content was clearly related to dietary tannin content in three species (A. speciosus, Apodemus argenteus, and Myodes rufocanus). We then used fecal proline content to estimate the tannin intakes of these three forest-dwelling species in a forest in Hokkaido. In the autumn, estimated tannin intakes increased significantly in the Apodemus species, but not in M. rufocanus. We speculated that an increase in tannin intake during autumn may result from consumption of tannin-rich acorns. This hypothesis was consistent with population fluctuation patterns of the three species, which were well-synchronized with acorn abundance for the Apodemus species but not for M. rufocanus.     

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.     

Salivary Proteins as a Defense Against Dietary Tannins

Tannins, a diverse group of water-soluble phenolics with high affinity to proteins, are widely distributed in various parts of plants, and have negative effects in herbivores after ingestion. Some mammalian species are thought to counteract tannins by secreting tannin-binding salivary proteins (TBSPs). Several types of TBSPs are found in the saliva of laboratory animals, livestock, and wildlife. Among them, proline-rich proteins (PRPs) and histatins are effective precipitators of tannins. It is widely accepted that, at the least, PRPs act as a first line of defense against tannins. Many observations support this idea: in vitro affinity of PRPs to tannins is far higher than that of other proteins such as bovine serum albumin; complexes formed between PRPs and tannins are stable even under the conditions in the stomach and intestine; and PRP production is induced by ingesting tannins. It is believed that species that usually ingest tannins as part of their natural diets produce high levels of PRPs, whereas species not exposed to tannins produce little or no PRPs. This hypothesis is generally supported, although studies on TBSPs in wildlife are limited. This work stresses the importance of gathering basic information on such items as the characteristics of unidentified TBSPs, and seasonal and geographical variations in PRP production.     

Interaction of proteins with sorghum tannin: Mechanism,specificity and significance

The grain of some varieties of sorghum contains 2% or more condensed tannin; many other varieties contain no tannin at all. Agronomic advantages, e.g., resistance to bird depredation, are associated with high-tannin sorghums, which have relatively low nutritional value for nonruminants. The biological effects of tannin are a result of its propensity for binding proteins; both hydrogen bonding and hydrophobic interactions are involved. Sorghum tannins can bind dietary proteins and reduce their digestibility. Purified digestive enzymes are inhibited by tannin, but significant inhibition in vivo is unlikely. Proteins differ greatly in their affinity for tannin. Those with highest affinity are large, have an open structure, contain no bound carbohydrate and are rich in proline. Sorghum proteins of the alcohol-soluble prolamine fraction associate strongly with tannin, are difficult to remove during tannin purification and are found combined with tannin in the indigestible residue after in vitro digestion with pepsin. On germination, the seed may sacrifice a portion of these proteins to bind the tannin that might otherwise interfere with metabolism by inhibiting seed enzymes. During seed development, tannin molecules are relatively short and do not effectively precipitate proteins; as the seed dries, tannins undergo polymerization to an average of ca. 6 flavan-3-ol units/molecule. The antinutritional effects of sorghum tannins can be eliminated by soaking the grain in dilute aqueous alkali, but not by cooking. When rats are put on high-tannin sorghum diets, their parotid glands undergo hypertrophy and produce a group of unique salivary proteins with extremely high affinity for tannin. These proteins contain over 40% proline and are devoid of sulfur-containing and aromatic amino acids. This metabolic adaption may protect rats against tannin by binding and inactivating it immediately when it enters the digestive tract. Presented at the AOCS Meeting, May 11, 1983.     

Comparison of Electrophoretic Protein Profiles from Sheep and Goat Parotid Saliva

Saliva provides a medium for short-term adaptation to changes in diet composition, namely, the presence of plant secondary metabolites. Salivary proteins have biological functions that have particular influence on oral homeostasis, taste, and digestive function. Some salivary proteins, such as proline-rich proteins, are present in browsers but absent in grazers. Despite the significance of salivary proteins, their expression patterns in many herbivores are unknown. We investigated the sodium dodecyl sulfate-polyacrylamide gel electrophoresis profile of parotid salivary proteins from two domesticated species, one a grazer, the sheep, Ovis aries, and the other a mixed feeder, the goat, Capra hircus, both fed on the same conventional diet. With 12.5% polyacrylamide linear gels, we observed uniform patterns of salivary proteins within the two species. In the goat profile, 21 major bands were observed, and 19 in the sheep profile. Each band was subjected to peptide mass fingerprinting for purposes of identification, allowing for 16 successful protein identifications. Marked differences were observed between the species in the region of 25–35 kDa molecular weights: one band was present in significantly different intensities; three bands were present only in goats; and one band was present only in sheep. This is the first report of a comparison of the protein salivary composition of sheep and goats and suggests that future research should be conducted to reveal a physiological function for salivary proteins related to the differences in feeding behavior of these species.     

Tannin-binding proteins in saliva of deer and their absence in saliva of sheep and cattle

A method has been developed for detecting tannin-binding proteins in the saliva of herbivores. The method is simple and requires only small quantities of crude saliva. The saliva of deer, a browsing ruminant, has been compared to that of domestic sheep and cow, which are grazing ruminants. The browser, which normally ingests dietary tannin, produces tannin-binding proteins, while the grazers do not produce such proteins. The tannin-binding protein from deer saliva is a small glycoprotein containing large amounts of proline, glycine, and glutamate/glutamine. The protein is not closely related to the proline-rich salivary proteins found in rats and other nonruminant mammals.     

Dry bean tannins: A review of nutritional implications

Tannins are one of several antinutritional factors present in dry beans and are located mainly in the seed coat or testa. The tannin content of dry beans ranges from 0.0 to 2.0% depending on the bean species and color of the seed coat. Many high tannin bean varieties are of lower nutritional quality than low tannin varieties of beans. Naturally occurring food legume tannins are reported to interact with proteins (both enzyme and nonenzyme proteins) to form tannin-protein complexes resulting in inactivation of digestive enzymes and protein insolubility. Both in vitro and in vivo studies indicate that bean tannins decrease protein digestibility, either by inactivating digestive enzymes or by reducing the susceptibility of the substrate proteins after forming complexes with tannins and absorbed ionizable iron. Other deleterious effects of tannins include a lowered feed efficiency and growth depression in experimental animals. The antinutritional activity of bean tannins can be reduced by processing (1 or a combination of 2 or more methods), for example dehulling, soaking, cooking and germination. Genetic selection also may help in breeding varieties low in tannins. Potential chemical treatments such as use of tannin complexing agents are discussed. Presented at the AOCS Meeting, Dallas, Texas, April 1984.     

Do Salivary Proline-Rich Proteins Counteract Dietary Hydrolyzable Tannin in Laboratory Rats?

We hypothesized that dietary hydrolyzable tannins would not act as digestibility reducing substances but would be excreted in the feces if the tannin were ingested by rats producing salivary proline-rich proteins (PRPs). To test that hypothesis we used two groups of Sprague–Dawley rats: tannin-naïve rats that were secreting basal levels of salivary PRPs and tannin-habituated rats that were secreting elevated levels of PRPs. The animals were fed for 10–18 d on diets containing 3% (w/w) purified hydrolyzable tannin [pentagalloyl glucose (PGG)] that was periodically spiked with chemically synthesized, radiolabeled 1,2,3,4,6-penta-O-galloyl-[U-¹⁴ C]-d-glucopyranose (1 μCi per gram diet). The PGG-habituated rats excreted three times more of the consumed ¹⁴ C in their feces than did the PGG-naïve rats (11.4% for PGG-habituated rats vs. 3.5% for PGG-naïve rats, P < 0.05). The addition of 3% PGG to the diet of the PGG-naïve rats had no significant effect on apparent dry matter or nitrogen digestibility (P > 0.05). However, dry matter digestibility and nitrogen digestibility were significantly decreased by PGG in the diets of the PGG-habituated rats (7 and 25%, P < 0.001, respectively). Production of PRPs increased the amount of PGG excreted intact in the feces but at the cost of diminishing apparent dry matter and nitrogen digestibility.     

Growth, development and dental caries in rats fed two experimental diets

In order to determine the nutritional adequacy of diets MIT 200 and NIH 2000 on the growth and development of experimental animals, these two diets were fed to two groups of animals during three periods of development: 1) pregnancy; 2) lactation, and 3) post-weaning. These diets were compared with a diet that satisfied the requirements of the National Research Council for growth and maintenance. It was found that for the two experimental diets, growth was compromised due to a caloric deficit in both diets. In addition, NIH 2000 was deficient in other nutrients such as iron. Analysis of other parameters such as hemoglobin, hematocrit, salivary protein, saliva flow and weight of vital organs upon autopsy revealed a picture of iron deficiency in the NIH 2000 group. Caries formation can be precipitated by a mechanism influenced by a nutrient deficit.     

The Influence of Condensed Tannin Structure on Rate of Microbial Mineralization and Reactivity to Chemical Assays

We examined how tannin structure influences reactivity in tannin assays and carbon and nitrogen mineralization. Condensed tannins from the foliage of ten tree and shrub species and from pecan shells (Carya illinoensis) had different proportions of: (a) epicatechin (cis) and catechin (trans) isomers, (b) procyanidin (PC) and prodelphinidin (PD) monomers, and (c) different chain lengths. The response of each tannin to several widely used tannin assays was determined. Although there was some variation in response to proanthocyanidin (butanol/HCl) and Folin Ciocalteu assays, we did not deduce any predictable relationship between tannin structure and response to either assay. There was little variation in protein precipitation among the different tannins. To assess biological activity, six of the tannins were incubated with forest humus for 22 days. We determined that, while PC-based tannins remained at least partly extractable for the duration of the incubation, tannins with a high proportion of PD subunits rapidly became unextractable from soil. There was a positive correlation between net nitrogen mineralization and cis chemical structure. Carbon mineralization was enhanced initially by the addition of tannins to humus, but after 22 days, a negative correlation between the proportion of cis subunits and respiration was determined. Overall, we were not able to demonstrate consistent effects of structure on either microbial mineralization or reactivity to chemical assays; such relationships remain elusive.     

Salivary proline-rich proteins in mammals: Roles in oral homeostasis and counteracting dietary tannin

We review information on the structure of proline-rich proteins (PRPs), their various functions related to oral homeostasis and dietary tannin, and the structural basis of these functions. Consideration of the multifunctional nature of these salivary proteins helps explain both the subtle and large variations found in structure and secretion rates both within individuals and between species. We propose that the ancestral function of PRPs is in maintaining oral homeostasis and that counteracting dietary tannins by binding with them is a derived function. PRPs are effective in oral homeostasis at low secretion levels, whereas counteracting tannin depends on high secretion levels. In the dietary habits ranging from carnivores through omnivores to exclusively planteaters, the dietary nitrogen level is progressively reduced, and plant allelochemical intake, including tannins, increases. We suggest that during this evolution from meat-eater to plant-eater, there was some point in omnivory at which selective pressure from nitrogen limitations, arising from a low nitrogen/high tannin diet, became sufficiently great for the evolution of increased secretion level and diversification of PRPs for dealing with tannin. If this hypothesis is correct, carnivorous mammals should secrete low levels of PRPs for oral homeostasis, but should never secrete high levels, unless they are secondarily carnivorous. Omnivores consuming a diet of very little animal tissue but higher levels of tannin-containing foliage or fruit should generally have the capacity to produce high levels of salivary PRPs. Browsers and frugivores should also produce high levels of PRPs, but grazers may have reduced secretion rates depending on the antiquity of the dietary habit. This hypothesis is consistent with the limited information available on the abundance, type, and distribution of PRPs in mammals. Studies are suggested which would test the functional and evolutionary arguments presented.     

Patterns and sources of leaf tannin variation in yellow birch (Betula allegheniensis) and sugar maple (Acer saccharum)

Leaves from forest-grown sugar maple (Acer saccharum Marsh) and yellow birch (Betula allegheniensis Britt.) trees were analyzed for four tannin measures (hydrolyzable and condensed tannins, total phenolics, and protein binding) at three times during the growing season. Fifteen-year-old half-sib sugar maples from four provenances, representing the geographical extremes of the sugar maple range and growing in a common garden, were examined for the same traits. We found no significant geographic or seed source component to variation in three of the four tannin measures. We found significant seasonal changes in both birch and maple leaf tannins. Withincanopy leaf tannin variation tended to obscure differences between trees in maple, but in birches between-tree differences in leaf tannin content were more readily found. We also found a significant negative correlation between leaf protein binding capacity and leaf wet weight.     

Phlorotannin-protein interactions

Tannins are one of the most broadly distributed types of plant secondary compounds, and have been the focal point for many studies of plant/herbivore interactions. Tannins interact strongly with proteins, so that the fate and effects of ingested tannins are in part dependent on the mode of interaction of the tannin with dietary and endogenous proteins in an herbivore's gut. We investigated the factors affecting the precipitation of proteins by phlorotannins from three species of marine brown algae:Carpophyllum maschalocarpum, Ecklonia radiata, andLobophora variegata. Phlorotannins were precipitated by proteins in a pH-dependent and concentration-dependent fashion. Precipitation also varied as a function of the presence of reducing agent, the type of phlorotannin or protein used, and the presence of organic solvents such as hydrogen bond inhibitors. Of particular significance was the ability of some phlorotannins to oxidize and form covalent bonds with some proteins. In contrast, under similar experimental conditions three types of terrestrial tannins (procyanidins, profisetinidins, and gallotannins) apparently did not form covalent complexes with proteins. Our results suggest several ways in which the biological activity of phlorotannins may vary as a function of the properties of the gut environment of marine herbivores. Moreover, we identify specific structural characteristics of phlorotannins which affect their tendency to oxidize, and thus, their potential effects on marine herbivores.     

Resistance and Tolerance of Terminalia sericea Trees to Simulated Herbivore Damage Under Different Soil Nutrient and Moisture Conditions

Resource availability, degree of herbivore damage, genetic variability, and their interactions influence the allocation of investment by plants to resistance and tolerance traits. We evaluated the independent and interactive effects of soil nutrients and moisture, and simulated the effects of herbivore damage on condensed tannins (resistance) and growth/regrowth (tolerance) traits of Terminalia sericea, a deciduous tree in the Kalahari desert that constitutes a major component of livestock diet. We used a completely crossed randomized-block design experiment to examine the effects of nutrients, water availability, and herbivore damage on regrowth and resistance traits of T. sericea seedlings. Plant height, number of branches, internode length, leaf area, leaf mass for each seedling, combined weight of stems and twigs, and root mass were recorded. Condensed tannin concentrations were 22.5 and 21.5% higher under low nutrients and low soil moisture than under high nutrient and high water treatment levels. Tannin concentrations did not differ significantly between control and experimental seedlings 2 mo after simulated herbivore damage. Tannin concentrations correlated more strongly with growth traits under low- than under high-nutrient conditions. No trade-offs were detected among individual growth traits, nor between growth traits and condensed tannins. T. sericea appeared to invest more in both resistance and regrowth traits when grown under low-nutrient conditions. Investment in the resistance trait (condensed tannin) under high-nutrient conditions was minimal and, to a lesser degree, correlated with plant growth. These results suggest that T. sericea displays both resistance and tolerance strategies, and that the degree to which each is expressed is resource-dependent.     

Soy-based,tannin-modified plywood adhesives

In this research, two different types of commercial tannins, namely a hydrolysable tannin (chestnut) and a condensed flavonoid tannin (mimosa), were used to prepare two types of soy-based (soy flour (SF) and soy protein isolate) adhesives for making plywood. Thermogravimetric properties (TGA) and its derivative as function of temperature (DTG) of different soy-based adhesive were measured in the range 40°C–300°C. Thermomechanical analysis (TMA) from 25°C to 250°C was done for the different resin formulations. Duplicate three-ply laboratory plywood panels were prepared by adding 300 g/m² of the adhesives’ total resin solid content composed of SF or isolated soy protein (ISP), urea, chestnut, and mimosa tannin extracts with hexamine as hardener. Based on the results obtained, tannins can improve SF adhesion properties. The TMA showed that chestnut tannin extract appeared to react well with SF, while mimosa tannin extract appeared to react well with ISP. Matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry also showed that among other reactions, the soy protein amino acids reacted with the tannins. Furthermore, delamination and shear strength test results showed the good water resistance of plywood bonded with soy-based tannin modified adhesive.     

Experience early in life affects voluntary intake of blackbrush by goats

Low nutritional quality and high levels of condensed tannins adversely affect voluntary intake of blackbrush (Coleogyne ramosissima Torr.) by goats. We studied: (1) how consumption of blackbrush or alfalfa pellets by young goats affected their consumption of those foods later in life, and (2) whether previous ingestion of blackbrush or alfalfa pellets affected the excretion of condensed tannins and total phenols from blackbrush in urine and feces, production of proline-rich proteins in saliva, excretion of nitrogen in feces, and mass of the liver, kidneys, parotid glands, and reticuloramen in goats. From 6 to 26 weeks of age, experienced goats were exposed to blackbrush, while inexperienced goats ate alfalfa pellets. Following exposure, both groups were offered older-growth blackbrush twigs (OG) or a choice between OG and current season's blackbrush twigs (CSG). A similar feeding trial was repeated nine months after exposure, and, in addition, both groups were offered a choice between OG ad libitum and alfalfa pellets at six levels of availability. Immediately after exposure, experienced goats ingested 95% more (P < 0.01) OG per unit of body weight than inexperienced goats, but both groups rejected CSG. Nine months after exposure, experienced goats ingested 27% more (P < 0.01) OG than inexperienced goats. Experienced goats ingested 30% more OG than inexperienced goats at every level of alfalfa pellet availability. The fate of condensed tannins and total phenols was similar for both groups, but experienced goats excreted 63% more (P < 0.05) uronic acids per unit of body weight. Neither experienced nor inexperienced goats produced proline-rich proteins in saliva. Experienced goats excreted 32% more (P < 0.01) nitrogen in feces per unit of nitrogen ingested than did inexperienced goats. The mass of the reticulorumen was 30% greater (P < 0.05) for experienced than for inexperienced goats one month after exposure, but did not differ 10 months after exposure. The mass of the liver, kidneys, and parotid glands did not differ between treatments. The results show that experience early in life can have profound and persistent effects on consumption of diets high in chemical defenses and low nutrional quality. The results also suggest that several physiological and morphological factors are involved.     

Do defoliation and subsequent phytochemical responses reduce future herbivory on oak trees?

Perennial plants are thought to respond to partial or complete defoliation by producing new foliage that is less susceptible to herbivores because of induction of allelochemicals. Here, I tested this hypothesis by manually removing primary foliage from branches ofQuercus emoryi (Fagaceae) at two different times in the season and monitoring changes in protein and tannin levels and the amount of herbivory relative to control branches. New, secondary leaves had 2.5 × greater hydrolyzable tannin content than mature foliage of control branches. Condensed tannins, which constitute a relatively low fraction of leaf mass, were lower, while protein content was temporarily greater, in new secondary leaves relative to mature leaves. Despite large increases in hydrolyzable tannins, herbivory levels were greater on refoliated branches than on control branches. New foliage is susceptible to herbivory regardless of when it is produced in the season, possibly because lower toughness and higher water content override any induced or developmentally related changes in allelochemistry. My results do not support the hypothesis that postherbivore changes in phytochemistry protect perennial plants from future herbivory, at least within a growing season.     

In‐vitro study on the efficacy of tannin fractions of edible nuts as antioxidants

Tannin fractions were isolated from crude acetonic extracts of defatted walnut, hazelnut and almond kernels using Sephadex LH‐20 column chromatography. The obtained material was characterized by content of total phenolics and electrophoretic separations using capillary zone electrophoresis (CZE). The antioxidant activities of the tannin fractions were analyzed by several methods: DPPH and ABTS assays, photochemiluminescence (PCL) method, as well as in two lipid model systems: emulsion with β‐carotene‐linoleic acid and L ‐α‐lecithin liposomes. The contents of total phenolics in the tannin fractions of walnuts, hazelnuts and almonds were 550, 329 and 83 mg catechin eq/g, respectively. The electrophoretic profiles of hazelnut and almond tannin fractions were similar, in contrast to the walnut profile. All analyzed fractions exhibited strong antioxidant properties. The antioxidant capacity of lipid‐soluble (ACL) compounds determined by PCL method was the highest for the fraction isolated from walnuts – 7.35 mmol Trolox eq/g. The DPPH radical and the ABTS radical cation were scavenged by the walnut tannin fraction with a higher efficacy than by the two other fractions. EC₅₀ values of the DPPH method were 1.8 times higher for the hazelnut fraction and 2.3 times higher for the almond fraction when compared to the walnut tannins. In turn, the total antioxidant activity values were 8.17, 2.82 and 1.98 mmol Trolox eq/g for the walnut, hazelnut and almond fractions, respectively. On the other hand, in both lipid models applied, lower antioxidant activity of walnut tannins than of hazelnut tannins was noted. The antioxidant effect of almond tannins was weaker or similar than that of walnut tannins in the β‐carotene‐linoleic acid emulsion and the L ‐α‐lecithin liposomal system, respectively.     

Tannin assays in ecological studies Precipitation of ribulose-1,5-bisphosphate carboxylase/oxygenase by tannic acid,quebracho, and oak foliage extracts

Tannic acid and quebracho precipitate many times their weight of the abundant leaf protein, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPC). The use of this protein in protein precipitation assays for tannin content is described. Extracts of mature foliage of pin, bur, and black oak precipitate 2.01, 0.69, and 0.09 mg RuBPC/ mg (dry wt) of leaf powder extracted, respectively, at pH 6.1. From these measurements it can be calculated that all three of these oak species have sufficient tannins to precipitate all of the RuBPC present in their foliage. At mildly alkaline pHs, however (pH ? 7.5), RuBPC is not precipitated by tannins. Since RuBPCis the most abundant protein present in photosynthetic tissues, often constituting as much as 50% of the soluble proteins and 25% of the total proteins in leaf tissue, the interactions of this protein with tannins are highly relevant to an evaluation of the role of tannins as antiherbivore, digestibility-reducing substances. Our measurements provide no basis for arguing that differences in tannin levels in different species reflect differences in the digestibility of leaf proteins or that tannins have any effect whatsoever upon the digestibility of leaf protein under conditions which normally prevail in most insects' guts. These findings emphasize the need to test more of the assumptions underlying contemporary interpretations of the importance of tannins in plant herbivore interactions.