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Congeneric species often share ecological niche space resulting in competitive interactions that either limit co-occurrence or lead to niche partitioning. Differences in fundamental nutritional niches mediated through character displacement or isolation during evolution are potential mechanisms that could explain overlapping distribution patterns of congenerics. We directly compared nutritional requirements and tolerances that influence the fundamental niche of mule (Odocoileus hemionus) and white-tailed deer (O. virginianus), which occur in allopatry and sympatry in similar realized ecological niches across their ranges in North America. Digestible energy and protein requirements and tolerances for plant fiber and plant secondary metabolites (PSMs) of both deer species were quantified using in vivo digestion and intake tolerance trials with six diets ranging in content of fiber, protein, and PSMs using tractable deer raised under identical conditions in captivity. We found that compared with white-tailed deer, mule deer required 54% less digestible protein and 21% less digestible energy intake per day to maintain body mass and nitrogen balance. In addition, they had higher fiber, energy, and dry matter digestibility and produced glucuronic acid (a byproduct of PSM detoxification) at a slower rate when consuming the monoterpene α-pinene. The mule deers’ enhanced physiological abilities to cope with low-quality, chemically defended forages relative to white-tailed deer might minimize potential competitive interactions in shared landscapes and provide a modest advantage to mule deer in habitats dominated by low-quality forages.

Accumulation of certain phenolics is a well‐known response of plants to enhanced UVB radiation (280–315 nm), but few experiments have compared the relative importance of different phenolic groups for UVB resilience. To study how an altered phenolic profile affects the responses and resilience of silver birch (Betula pendula) to enhanced UVB, we used RNA interference (RNAi) targeting dihydroflavonol reductase (DFR), anthocyanidin synthase (ANS), or anthocyanidin reductase (ANR) to change the accumulation of phenolics. The unmodified control line and RNAi‐modified plants were grown for 51 days under ambient or +32% enhanced UVB dose in a greenhouse. RNAi greatly affected phenolic profile and plant growth. There were no interactive effects of RNAi and UVB on growth or photosynthesis, which indicates that the RNAi and unmodified control plants were equally resilient. UVB enhancement led to an accumulation of foliar flavonoids and condensed tannins, and an increase in the density of stem glands and glandular trichomes on upper leaf surfaces in both the control and RNAi‐modified plants. Our results do not indicate a photoprotective role for condensed tannins. However, decreased growth of high‐flavonoid low‐tannin DFRi and ANRi plants implies that the balance of flavonoids and condensed tannins might be important for normal plant growth.

Proanthocyanidins (condensed tannins) are important in food chemistry, agriculture, and health, driving demand for improvements in structure determination. We used ultrahigh resolution Fourier transform-ion cyclotron resonance mass spectrometry (FT-ICR MS) methods to determine the exact composition of individual species in heterogeneous mixtures of proanthocyanidin polymers from Sorghum bicolor grain and Neptunia lutea leaves. Fragmentation patterns obtained with FT-ICR ESI MS-MS (electrospray ionization) confirmed structural details from thiolysis-high-performance liquid chromatography (HPLC)-diode array detection (DAD) and 1H-13C heteronuclear single quantum coherence (HSQC) NMR. We found that A-type linkages were characteristic of shorter polymers in predominantly B-linked proanthocyanidin. We suggest that supramolecular complex formation between proanthocyanidins and matrix components such as 2,5-dihydroxybenzoic acid was responsible for anomalous 152 dalton peaks, incorrectly assigned as 3-O-galloylation, when using FT-ICR matrix-assisted laser desorption ionization (MALDI-MS). Our data illustrate the power of the ultrahigh resolution FT-ICR methods but include the caveat that MALDI-MS must be paired with complementary analytical tools to avoid artifacts.

Despite active research, antiherbivore activity of specific plant phenolics remains largely unresolved. We constructed silver birch (Betula pendula) lines with modified phenolic metabolism to study the effects of foliar flavonoids and condensed tannins on consumption and growth of larvae of a generalist herbivore, the autumnal moth (Epirrita autumnata). We conducted a feeding experiment using birch lines in which expression of dihydroflavonol reductase (DFR), anthocyanidin synthase (ANS) or anthocyanidin reductase (ANR) had been decreased by RNA interference. Modification-specific effects on plant phenolics, nutrients and phenotype, and on larval consumption and growth were analyzed using uni- and multivariate methods. Inhibiting DFR expression increased the concentration of flavonoids at the expense of condensed tannins, and silencing DFR and ANR decreased leaf and plant size. E. autumnata larvae consumed on average 82% less of DFRi plants than of unmodified controls, suggesting that flavonoids or glandular trichomes deter larval feeding. However, larval growth efficiency was highest on low-tannin DFRi plants, indicating that condensed tannins (or their monomers) are physiologically more harmful than non-tannin flavonoids for E. autumnata larvae. Our results show that genetic manipulation of the flavonoid pathway in plants can effectively be used to produce altered phenolic profiles required for elucidating the roles of low-molecular weight phenolics and condensed tannins in plant–herbivore relationships, and suggest that phenolic secondary metabolites participate in regulation of plant growth.

Microorganisms play vital roles in modulating organic matter decomposition and nutrient cycling in soil ecosystems. The enzyme latch paradigm posits microbial degradation of polyphenols is hindered in anoxic peat leading to polyphenol accumulation, and consequently diminished microbial activity. This model assumes that polyphenols are microbially unavailable under anoxia, a supposition that has not been thoroughly investigated in any soil type. Here, we use anoxic soil reactors amended with and without a chemically defined polyphenol to test this hypothesis, employing metabolomics and genome-resolved metaproteomics to interrogate soil microbial polyphenol metabolism. Challenging the idea that polyphenols are not bioavailable under anoxia, we provide metabolite evidence that polyphenols are depolymerized, resulting in monomer accumulation, followed by the generation of small phenolic degradation products. Further, we show that soil microbiome function is maintained, and possibly enhanced, with polyphenol addition. In summary, this study provides chemical and enzymatic evidence that some soil microbiota can degrade polyphenols under anoxia and subvert the assumed polyphenol lock on soil microbial metabolism., It is thought that polyphenols inhibit organic matter decomposition in soils devoid of oxygen. Here the authors use metabolomics and genome-resolved metaproteomics to provide experimental evidence of polyphenol biodegradation and maintained soil microbial community metabolism despite anoxia.

The effects of condensed tannins on N dynamics in ruminants have been a topic of research for some time, but much less work has focused on their impacts on other nutrients in manure. A 4 × 4 Latin square trial was used to determine if intake of sericea lespedeza (Lespedeza cuneata; SL; a condensed tannin source), at 0, 10, 20, or 40% of the diet (as-fed basis), would affect concentrations of nutrients in manure and patterns of total excretion when offered with alfalfa (Medicago sativa; ALF) to sheep. With SL additions, average daily manure production increased linearly (P ≤ 0.01), from 40 to 50% of the diet mass. The concentrations of total C, total N, soluble P, total and soluble Na, total and soluble S, total and soluble Mn, and total and soluble B in feces increased (P ≤ 0.05) while soluble N, total Ca, total and soluble Mg, soluble Zn, total and soluble Fe, total and soluble Cu decreased (P ≤ 0.02). Total P, total and soluble K, soluble Ca, and total Zn were less affected (P > 0.05). Comparing diets containing 0 to 40% SL, average daily outputs of total C, total N, soluble P, soluble K, total and soluble Na, and total Mn increased linearly (P ≤ 0.01) by 42.0, 71.2, 93.3, 45.2, 111, 148, and 52.4 percentage points, respectively. Total K, total and soluble S, soluble Mn, and total and soluble B increased quadratically (P ≤ 0.02) by 26.1, 52.3, 26.7, 147, 100, and 19.5 percentage points, respectively. Conversely, outputs of soluble Zn and total Fe decreased linearly (P ≤ 0.01), by –51.5 and –24.8 percentage points, while total Ca, total and soluble Mg, soluble Fe, and soluble Cu decreased quadratically (P ≤ 0.05) by –15.7, –12.3, –40.0, –89.9, and –60.3 percentage points, respectively. Outputs of soluble N, total P, soluble Ca, total Zn, and total Cu remained unchanged (P ≥ 0.14). Ratios of manure outputs to feed inputs for C, N, K, and B increased (P ≤ 0.02) but those for P and Mg were unchanged (P ≥ 0.10). Ratios of soluble to total manure outputs (S:O) increased (P ≤ 0.01) for P, Ca, Na, Mn; decreased (P ≤ 0.05) for N, S, Mg, Zn, Fe, Cu, and B; and were unaffected by treatment (P ≤ 0.16) for K. Decreasing S:O ratios are consistent with the formation of complexes that adsorb these nutrients to insoluble fiber fractions of manure and could thus affect mineralization rates. This study suggests that dietary tannins, found in forages like SL, can alter the concentrations, total excretion rates and throughput efficiency of nutrients in manure.

(−)-Epigallocatechin is a chemoprotective polyphenolic compound found in green tea. Its low abundance in tea increases the cost and difficulty of obtaining sufficient (−)-epigallocatechin to conduct research on (−)-epigallocatechin-mediated health benefits. We developed a convenient and economical method for preparing (−)-epigallocatechin from a commercial green tea extract in a sufficient quantity to carry out mechanistic studies of the pure compound. Crude epigallocatechin gallate was isolated from the tea extract by silica gel column chromatography, and was then hydrolyzed with immobilized tannase. The (−)-epigallocatechin found in the hydrolyzed products was extracted into ethyl acetate and purified by Sephadex LH-20 to produce (−)-epigallocatechin. Very pure (−)-epigallocatechin (93.7 %) was obtained and its identity was confirmed by ESI-MS and HPLC. The overall yield was 10 % by mass of the initial green tea extract obtained as pure (−)-epigallocatechin. No specialized equipment was required, and the solvents and chromatographic supports used were inexpensive and readily available.

Phenolic plant secondary metabolites actively participate in a broad range of important reactions that affect livestock, plants and soil. In soil, phenolic compounds can affect nutrient dynamics and mobility of metals but their role in northern Great Plains agroecosystems is largely unknown. We evaluated the effects of three phenolic compounds on plant nutrient extractability in agricultural soil. Soil samples (0–10 cm) from pasture and cropped sites near Mandan, North Dakota, USA were treated with water (control) or aqueous solutions of increasingly complex compounds; benzoic acid (BA), gallic acid (GA), or β-1,2,3,4,6-penta-O-galloyl- d -glucose (PGG) at four concentrations (1.25, 2.5, 5 or 10 mg compound gram−1 soil). We measured extractable nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg) and manganese (Mn) in treatment supernatants and after a subsequent incubation in hot water (16 h, 80 °C). While significant quantities of nitrogen were extracted with water, each compound reduced N extracted in a concentration dependent manner with greatest response to BA (about 25%). However, PGG reduced the solubility of N only during the hot water incubation, suggesting its effects are mostly on organic forms of N. Unlike N, GA and PGG increased extraction of P, relative to water, while BA had less effect. Extraction of the major cations, K, Ca, and Mg, was strongly increased by BA and GA but unaffected by PGG. Extraction of Mn was increased most by treatments of GA but less so by BA. The effects of PGG on Mn were consistent with GA when expressed on a molar basis. These findings suggest plant secondary compounds affect nutrient dynamics in soil, and thus may be part of future management strategies to improve nutrient-use efficiency.

(−)-Epigallocatechin-3-gallate (EGCg) is a naturally occurring polyphenol found in plant-based foods and beverages such as green tea. Although EGCg can eliminate carbonyl species produced by glucose autoxidation and thus can inhibit protein glycation, it is also reported to be a pro-oxidant that stimulates protein glycation in vitro. To better understand the balance between antioxidant and pro-oxidant features of EGCg, we evaluated EGCg-mediated bioactivities in a human serum albumin (HSA)/glucose model by varying three different parameters (glucose level, EGCg concentration, and time of exposure to EGCg). Measurements of glycation-induced fluorescence, protein carbonyls, and electrophoretic mobility showed that the level of HSA glycation was positively related to the glucose level over the range 10 to 100 mM during a 21-day incubation at 37 °C and pH 7.4. Under mild glycemic pressure (10 mM), long exposure to EGCg enhanced HSA glycation, while brief exposure to low concentrations of EGCg did not. Under high glycemic pressure (100 mM glucose), long exposure to EGCg inhibited glycation. For the first time we showed that brief exposure to EGCg reversed glycation-induced fluorescence, indicating a restorative effect. In conclusion, our research identified glucose level, EGCg concentration, and time of exposure as critical factors dictating EGCg bioactivities in HSA glycation. EGCg did not affect HSA glycation under normal physiological conditions but had a potential therapeutic effect on HSA severely damaged by glycation.

The principal green tea polyphenol, (−)-epigallocatechin-3-O-gallate (EGCg), may provide chemoprotection against conditions ranging from cardiovascular disease to cancer. Binding to plasma proteins stabilizes EGCg during its transport to targeted tissues. This study explored the details EGCg binding to bovine serum albumin. Both fluorescence lifetime and intensity data showed that the hydrophobic pocket between subdomains IIA and IIIA is the binding site for EGCg. Fluorescence and circular dichroism were used to establish the roles of the flavan-3-ol and galloyl moieties of the EGCg in binding and to demonstrate a binding-dependent conformational change in the protein. Competitive binding experiments confirmed the location of binding, and molecular modeling identified protein residues that play key roles in the interaction. This model of EGCg–BSA interactions improves the understanding of the likely physiological fate of this green tea-derived bioactive polyphenol.

The beneficial effects of forages containing condensed tannins (CTs) on ruminants are well documented, but the chemical features of CT that yield benefits have not been defined. Some evaluations of limited numbers of highly purified compounds have resulted in positive correlations between CT molecular weight (MW) and biological activity, while others have failed to show a correlation. The objectives of this study were to determine if MW of CT could predict biological activity relative to protein precipitability. MW of condensed tannin, protein-precipitable phenolics (PPP), and the amount of protein bound (PB) were determined for nine species of warm-season perennial legumes. There was no correlation between PPP or PB and MW (R2 0.11 and R2 0.02, respectively). However, CT concentration did correlate with PPP and PB (R2 0.81 and R2 0.69, respectively). It was concluded that CT MW does not explain the variation in protein precipitation by CT from the forage legumes surveyed.

To elucidate the anti-venom mechanism of persimmon tannin, the interaction between a polymeric persimmon proanthocyanidin fraction (PT40) and phospholipase A2 (PLA2) or bovine serum albumin (BSA) were studied using a competitive binding assay and spectroscopic methods including Fourier transform infrared spectroscopy (FT-IR), circular dichroism (CD), and resonance light scattering (RLS) spectroscopy. The results revealed that PT40 has a higher affinity for PLA2 than for BSA at physiological pH and induced greater conformational changes in PLA2 than in BSA. PT40 covalently bound to PLA2 in a reaction probably involving Lys residues. We propose that the high affinity of PT40 for PLA2 and the covalent modification of PLA2 by PT40 may be responsible for the ability of the tannin to irreversibly inhibit PLA2 catalytic activity, to prevent edema, and to neutralize the lethality of Chinese cobra PLA2 in vivo.

The plant natural products known as polyphenols are found at micronutrient levels in fruits, vegetables, and plant-based beverages such as wine, tea, coffee and cocoa. Consumption of a fruit- and vegetable-rich diet, the "Mediterranean diet", has been epidemiologically related to health benefits especially for chronic diseases including diabetes, cardiovascular disease, and Alzheimer's disease. The abundance of polyphenols in plant-rich diets, and the potent bioactivities of polyphenols, provide indirect evidence for a role for polyphenols in maintaining good health. However, molecular mechanisms for therapeutic or preventative activity have not been demonstrated in vivo. We summarize the chemical classes of natural polyphenols, their bioactivities and bioavailability and metabolism. Because many polyphenols bind protein, we focus on the potential of protein binding to mediate the health-related effects of polyphenols. We discuss interactions with plasma proteins as the first target organ past the digestive tract for these orally-ingested compounds.

Polyphenols including tannins comprise a large percentage of plant detritus such as leaf litter, and affect soil processes including metal dynamics. We tested the effects of tannins on soil metal mobilization by determining the binding stoichiometries of two model polyphenols to Al(III) and Fe(III) using micelle-mediated separation and inductively coupled plasma optical emission spectroscopy (ICP-OES). By fitting the data to the Langmuir model we found the higher molecular weight polyphenol (oenothein B) was able to bind more metal than the smaller polyphenol (epigallocatechin gallate, EGCg). For example, oenothein B bound 9.43 mol Fe mol(-1), while EGCg bound 4.41 mol of Fe mol(-1). Using the parameters from the binding model, we applied the Langmuir model for competitive binding to predict binding for mixtures of Al(III) and Fe(III). Using the parameters from the single metal experiments and information about polyphenol sorption to soils we built a model to predict metal mobilization from soils amended with polyphenols. We tested the model with three natural soils and found that it predicted mobilization of Fe and Al with r(2)=0.92 and r(2)=0.88, respectively. The amount of metal that was mobilized was directly proportional to the maximum amount of metal bound to the polyphenol. The secondary parameter in each model was the amount of weak organically chelated Fe or Al that was in the soil. This study provides the first compound-specific information about how natural polyphenols interact with metals in the environment. We propose a model that is applicable to developing phytochelation agents for metal detoxification, and we discuss how tannins may play a role in metal mobilization from soils.

Many food and forage plants contain tannins, high molecular weight polyphenols that characteristically interact strongly with protein, forming complexes that affect taste, nutritional quality, and the health of the consumer. In the present study, the interaction between bovine serum albumin (BSA) and each of seven hydrolyzable tannins or epigallocatechin gallate was examined. The objective was to define the effect of tannin oxidation, measured as oxidative activity (browning) or as oxidizability (degradation monitored by HPLC), on the formation on highly stabilized tannin-protein complexes and to determine how the reaction depended on the pH conditions. Gel electrophoresis and MALDI-TOF-MS were used to assess the formation of tannin-protein complexes. The results showed that tannin oxidizability was directly correlated with the tendency of the tannins to form highly stabilized complexes with BSA at increased pH (7.6). However, at slightly lower pH (6.7), other tannin features, such as the size and flexibility of the tannin, appeared to dictate the formation of highly stabilized tannin-protein complexes.

Natural polyphenols may play an important role in aluminum detoxification in some plants. We examined the interaction between Al(3+) and the purified high molecular weight polyphenols pentagalloyl glucose (940 Da) and oenothein B (1568 Da), and the related compound methyl gallate (184 Da) at pH 4 and 6. We used spectrophotometric titration and chemometric modeling to determine stability constants and stoichiometries for the aluminum-phenol (AlL) complexes. The structures and spectral features of aluminum-methyl gallate complexes were evaluated with quantum chemical calculations. The high molecular weight polyphenols formed Al3L2 complexes with conditional stability constants (β) ∼ 1 × 10(23) at pH 6 and AlL complexes with β ∼ 1 × 10(5) at pH 4. Methyl gallate formed AlL complexes with β = 1 × 10(6) at pH 6 but did not complex aluminum at pH 4. At intermediate metal-to-polyphenol ratios, high molecular weight polyphenols formed insoluble Al complexes but methyl gallate complexes were soluble. The high molecular weight polyphenols have high affinities and solubility features that are favorable for a role in aluminum detoxification in the environment.

We investigated tannin–soil interactions by assessing the kinetics of sorption and sorption capacities, and their relationship to the chemical properties of six polyphenolic compounds and the textures of six soils. We developed a new extraction procedure for recovering tannins from soil samples by successive extraction with solvents of decreasing polarity. Sorption of polyphenolic compounds methyl gallate, catechin, oenothein B, pentagalloyl glucose, epigallocatechin gallate, and procyanidin dimer was determined using six soils with textures ranging from 7% silt–89% sand to 52% silt–22% sand. Sorption kinetics and capacity of the soils were determined using room temperature mixing with high performance liquid chromatography (HPLC) to determine polyphenol concentration. Tannins were extracted from soils loaded with known amounts of the model compounds using solvents with a range of polarities, and establishing recovery by HPLC. Multivariable regression was used to establish relationships between polyphenol properties and sorption, and between soil texture and sorption. Sorption of the polyphenols followed the Langmuir isotherm with unique binding capacity and kinetics of sorption for each compound. Tannin sorption was correlated to molecular weight and polarity. For an Ultisol pasture soil, up to 18.7 mg g−1 soil of the large, hydrophobic compound epigallocatechin gallate was bound compared to only 5.11 mg g−1 soil of the smaller, more polar compound methyl gallate. Kinetics of sorption also varied with sorption reaching equilibrium between 75 and 350 min. Silt and sand composition affected polyphenol sorption in a unique fashion for each polyphenol. Using sequential extraction with solvents ranging from polar (water) to non-polar (hexane), we extracted up to 42% of the material that had been loaded on the soil as a mixture of five polyphenols. This work demonstrates that the likely fate of tannins and related polyphenols from plant sources is rapid sorption, with little likelihood of release of unmodified tannins from the soil by leaching. Tight sorption impedes recovery and analysis of soil tannins, but better methods for extraction may improve our knowledge of tannins in the soil.

The antioxidant activities of high molecular weight persimmon condensed tannin (HMWPT) were evaluated in an ex vivo tissue system and in vivo . Addition of HMWPT to mouse liver homogenate protected the samples against auto-peroxidation or H 2 O 2 - or Fe 2+ /ascorbic acid-induced lipid peroxidation. The IC 50 values for HMWPT were 4.32 ± 0.20 μg/mL (auto-oxidation), 1.36 ± 0.40 μg/mL (H 2 O 2 -induced peroxidation) or 0.20 ± 0.09 mg/mL (Fe 2+ /ascorbic acid-induced peroxidation). Mice were oxidatively stressed with bromobenzene to test the antioxidant activity of HMWPT in vivo. An oral dose of HMWPT at 200 or 400 mg/kg HMWPT significantly ( P P ex vivo and in vivo .

The aim of this study was to assess the use of the protein precipitable phenolics assay as a method for estimating tannin in soil samples. This method is more selective than general methods such as Folin assays and is used to differentiate tannin from other phenolics in plant extracts. We evaluated the role of the protein as an agent that separated tannin from nontannin phenolics in soil by running the assay in the normal mode, with protein present, and in a modified format without the protein. We tested the method using native soils and soils amended with tannic acid. We found that this assay is not suitable for determining tannin in soil samples, because the protein precipitation step is incompatible with solid soil samples. We also assessed the background color of soil samples in the assay and established that background color is so high that reliable absorbance readings cannot be obtained.

Retention of tannins, produced by plants, could be important for managing soil organic matter and nutrient cycling. However, we know little about the comparative retention of different classes of tannins and related compounds or if soils have a maximum storage capacity for them. To address these questions, forest, and pasture loam soils, collected at 0–5 cm (surface) and 10–20 cm (subsurface), were repeatedly treated with water (Control) or solutions containing condensed and hydrolyzable tannins or related phenolic subunits (10 mg g−1 soil). Treatments included a polymeric flavonoid-based procyanidin from sorghum, catechin, tannic acid, β-1,2,3,4,6-penta-O-galloyl- d -glucose (PGG), gallic acid, and methyl gallate. After each application, soluble-C in supernatants was determined by oxidative-combustion infrared analysis and retention of treatment-carbon by soil was calculated as the difference between added and recovered soluble-C. An interaction between soil depth and treatment was evident through all applications with highest retention of both hydrophobic (PGG) and hydrophilic (procyanidin) tannins, compared to other phenolic compounds. For all treatments except gallic acid and methyl gallate, higher sorption occurred in surface soil, which contained more organic matter than subsurface soil. With each successive application, less additional treatment-C was retained by soil and the amount of C remaining in supernatants was correlated with the presence of phenolic substances. Cumulative retention by surface soil was more than 10.3, 8.5 and 6.4 mg C g−1 soil for PGG, tannic acid, and procyanidin, several times higher than the other compounds. Soluble-C extracted from treated soil, with cool water (23 °C), was 1–2 orders of magnitude greater than Control samples and highly correlated with Prussian Blue (PB) phenolics, indicating some retained treatment-C was only weakly held on the soil. The final extraction, with hot water (80 °C), removed more soluble-C, particularly from surface samples, that contained fewer PB phenolics per unit soluble-C than cool water extracts. After all extractions more than 85% of sorbed procyanidin-C was retained by samples compared to 81% of methyl gallate, 79% of PGG, 74% of tannic acid, 50% of catechin, and 40% of the gallic acid. Total C, measured in soil after all extractions, was close to expected values, confirming tannins and phenolic compounds had remained in soil and were not otherwise lost. Cation exchange capacity was increased about 30% in subsurface and forest samples by PGG, a hydrolyzable tannin, but decreased by 30% and 35% in surface and pasture soil, respectively, by its monomer, gallic acid.

Polyphenols are bioactive natural products that appear to act against a wide range of pathologies. Mechanisms of activity have not been established, but recent studies have suggested that some polyphenols bind to membranes. This study examined the interaction between lipid bilayers and three structurally diverse polyphenols. It was hypothesized that features of the polyphenols such as polarity, molecular size, molecular geometry, and number and arrangement of phenol hydroxyl groups would determine the tendency to interact with the bilayer. The examined compounds included a mixed polyphenol, (-)-epigallocatechin gallate (EGCg); a proanthocyanidin trimer comprising catechin-(4→8)-catechin-(4→8)-catechin (cat₃; and a hydrolyzable tannin, 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose (PGG). These polyphenols were incorporated at different levels into ²H-labeled 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) multilamellar vesicles (MLVs). ³¹P and ²H solid-state NMR experiments were performed to determine the dynamics of the headgroup region and the hydrophobic acyl chain region of the lipid bilayer upon addition of polyphenols. The chemical shift anisotropy (CSA) width of the ³¹P NMR spectra decreased upon addition of polyphenols. Addition of PGG induces a dramatic reduction on the CSA width compared with the control lipid bilayer sample, whereas addition of cat₃ barely reduces the CSA width. The ²H quadupolar splitting of the lipids also decreased upon addition of polyphenols. At the same concentration, PGG substantially reduced the quadrupolar splitting, whereas cat₃ barely reduced it when compared with the control sample. From a calculation of the order parameters of the acyl chain region of the lipid bilayer, it was concluded that the hydrophobic part of the lipid bilayer was perturbed by PGG, whereas cat₃ did not cause large perturbations. The data suggest that the polarity of the polyphenols affects the interaction between tannins and membranes. The interactions may relate to the biological activities of polyphenols.

The bioactivities and bioavailability of plant polyphenols including proanthocyanidins and other catechin derivatives may be affected by covalent reaction between polyphenol and proteins. Both processing conditions and gastrointestinal conditions may promote formation of covalent complexes for polyphenol-rich foods and beverages such as wine. Little is known about covalent reactions between proteins and tannin, because suitable methods for quantitating covalent complexes have not been developed. We established capillary electrophoresis methods that can be used to distinguish free protein from covalently bound protein-polyphenol complexes and to monitor polyphenol oxidation products. The methods are developed using the model protein bovine serum albumin and the representative polyphenol (-)epigallocatechin gallate. By pairing capillaries with different diameters with appropriate alkaline borate buffers, we are able to optimize resolution of either the protein-polyphenol complexes or the polyphenol oxidation products. This analytical method, coupled with purification of the covalent complexes by diethylaminoethyl cellulose chromatography, should facilitate characterization of covalent complexes in polyphenol-rich foods and beverages such as wine.

The literature on proanthocyanidins (tannins) in ecological systems is dominated by quantitative studies. Despite evidence that the qualitative characteristics (subunit type, polymer chain length) of these complex polyphenolics are important determinants of biological activity, little is known about genetic and environmental controls on the type of proanthocyanidins produced by plants. We tested the hypothesis that genetics, season, developmental stage, and environment determine proanthocyanidin qualitative characteristics by using four Populus "cross types" (narrowleaf [P. angustifolia], Fremont [P. fremontii], F1 hybrids, and backcrosses to narrowleaf). We used thiolysis and HPLC analysis to characterize the proanthocyanidins, and found that genetics strongly control composition. The narrowleaf plants accumulate mixed procyanidin/prodelphinidins with average composition epicatechin(11)-epigallocatechin(8)-catechin(2)-catechin((terminal)). Backcross genotypes produce mixed procyanidin/prodelphinidins similar to narrowleaf, while Fremont makes procyanidin dimers, and the F1 plants contain procyanidin heptamers. Less striking effects were noted for genotype × environment, while season and developmental zone had little effect on proanthocyanidin composition or chain length. We discuss the metabolic and ecological consequences of differences in condensed tannin qualitative traits.

Regular bouts of physical activity may cause changes in gene expression that accumulate over time and ultimately affect phenotypes, such as body weight, blood lipid profile and tumour development. Furthermore, acute activity may affect gene expression and phenotypes differently depending on whether the individual is regularly inactive or active. One-month-old male Sprague–Dawley rats (n= 72) were equally divided into SED (standard laboratory cage, n= 24), PA (large activity box, n= 24) and EX groups (exercise wheel inside standard cage, n= 24). At 3 months of age, half the animals from each group were killed at rest and the other half following 30 min of physical activity. The RNA was extracted from cardiac tissue, and microarray analysis was performed on 27,000 genes. Select gene results were validated using quantitative PCR. No gene expression differences occurred when comparing all 3-month-old groups at rest. A relatively small percentage of genes (1.9%) were differentially expressed (P PA > EX. Differences in gene expression levels in young physically inactive and active animals following acute activity have different regulatory roles in gene networks that affect health-related phenotypes.

Previous studies showed that a series of purified condensed tannins (CTs) from warm-season perennial legumes exhibited high variability in their modulation of methane production during in vitro rumen digestion. The molecular weight differences between these CTs did not provide correlation with either the in vitro CH4 production or the ability to precipitate bovine serum albumin. In an effort to delineate other structure-activity relationships from these methane abatement experiments, the structures of purified CTs from these legumes were assessed with a combination of methanolysis, quantitative thiolysis, 1H-13C HSQC NMR spectroscopy and ultrahigh-resolution MALDI-TOF MS. The composition of these CTs is very diverse: procyanidin/prodelphinidin (PC/PD) ratios ranged from 98/2 to 2/98, cis/trans ratios ranged from 98/2 to 34/66, mean degrees of polymerization ranged from 6 to 39, and % galloylation ranged from 0 to 75%. No strong correlation was observed between methane production and the protein precipitation capabilities of the CT towards three different proteins (BSA, lysozyme, and alfalfa leaf protein) at ruminal pH. However, a strong non-linear correlation was observed for the inhibition of methane production versus the antioxidant activity in plant sample containing typical PC- and PD-type CTs. The modulation of methane production could not be correlated to the CT structure (PC/PD or cis/trans ratios and extent of galloylation). The most active plant in methane abatement was Acacia angustissima, which contained CT, presenting an unusual challenge as it was resistant to standard thiolytic degradation conditions and exhibited an atypical set of cross-peak signals in the 2D NMR. The MALDI analysis supported a 5-deoxy flavan-3-ol-based structure for the CT from this plant.

We have developed a simple method for preparing and verifying suitable standards for the acid butanol assay from a readily available source. Phenolics were extracted from fresh apples with methanol, and sugars were removed from the crude extract by treatment with Amberlite resin before fractionating the proanthocyanidins into ethyl acetate. The ethyl acetate fraction was chromatographed on Toyopearl TSK HW-50F to yield about 50 mg of procyanidin dimer and 35 mg of trimer from 1 kg fresh apple fruit. The purity and identity of the standards was easily confirmed by using ESI-MS. In the acid butanol assay, the pure dimer, trimer and purified Sorghum procyanidin had similar color yields on a mass basis, and produced about three times more color than purified quebracho tannin. This new standard overcomes problems associated with overestimation of tannin due to use of the unreactive quebracho tannin standard. Use of the new standard will enable accurate comparisons of tannin levels between laboratories and will standardize comparisons between species, thus promoting our understanding of the role of condensed tannins in plants.

Almonds, pistachios, and walnuts grown in California have an aggregate value of over $3.3 billion, with a large proportion of the crop being exported. Mycotoxin contamination affects this value, because importing nations, and in particular the European Community (EC), apply strict regulatory levels for aflatoxins, formed by infection of the product by Aspergillus species. Conformance with such limits can best be achieved by capitalising on natural resistance factors inherent to the nuts. Differential resistance of tree nut species to aflatoxin formation, with walnuts being least susceptible, led to research which established that the hydrolysable tannins in walnut pellicle were responsible for this effect. This observation has now been extended to show that the addition to Aspergillus flavus growth media of natural antioxidants present in tree nuts, such as hydrolysable tannins, flavonoids and phenolic acids, decrease maximum aflatoxin by as much as 99.8%. There is evidence that aflatoxin formation is exacerbated by oxidative stress on the fungus and therefore compounds capable of relieving such stress should limit aflatoxin accumulation. The addition of tertiary-butyl hydroperoxide to the media induced oxidative stress in A. flavus and stimulated peak aflatoxin production with high levels being maintained, rather than declining as in control samples. Aflatoxin formation was significantly inhibited by incorporation into the media of the antioxidant, tannic acid. A similar effect was seen for drought-stressed cultures, with increased aflatoxigenesis being reduced in the presence of gallic or caffeic acids. Increasing antioxidant natural products in tree nuts may restrict the ability of A. flavus to biosynthesise aflatoxins, thus achieving conformity with regulatory limits.

Tropical forests throughout the world are home to a guild of small ruminants that consume fruit as a substantial portion of their diet. Because the rumen is relatively inefficient at digesting nonstructural carbohydrates and only slowly digests cellulose, the feeding adaptations of frugivorous ruminants are enigmatic. We examined the nutritional value of wild figs to blue duikers, one of the smallest and most frugivorous ruminants, through chemical analyses and a series of digestion trials with six species of wild African figs. These figs were high in fat, protein, cell wall, lignin and Ca : P ratios, low in sugar and starch, and high in unextractable, fibre-bound tannins when compared with many other fruits. The fibre-bound tannins and protein caused protein digestibility and nitrogen balance to be consistently low or negative. The high fibre content of the figs allowed duikers to only digest 30–60% of energy contained in the figs. However, duikers were able to consume enough digestible energy to maintain body mass during 4-day trials. Therefore, a ruminant digestive system is beneficial to mammals eating high fibre, high-tannin tropical fruit like figs, especially if the mammal is small enough to harvest a sufficient amount to meet its daily energy requirements and has adaptations for reducing the effects of tannins on protein digestibility. Resume Les forets tropicales du monde entier abritent toute une faune de petits ruminants dont les fruits constituent une part non negligeable de l’alimentation. Comme le rumen est relativement peu efficace pour digerer les hydrates de carbone non structuraux et ne digere que lentement la cellulose, les adaptations alimentaires des ruminants frugivores sont encore enigmatiques. Nous avons examine la valeur nutritionnelle des figues sauvages pour le cephalophe bleu, un des ruminants les plus petits et parmi les plus frugivores, au moyen d’analyses chimiques et d’une serie d’essais de digestion avec six especes de figues sauvages africaines. Ces figues etaient riches en graisses, en proteines, en parois cellulaires, en lignine, et leur rapport Ca/P etait eleve; elles avaient un contenu faible en sucre et en amidon, et beaucoup de tanins impossibles a extraire, lies aux fibres, compares a de nombreux autres fruits. Les tanins lies aux fibres et les proteines faisaient que la digestibilite des proteines et l’equilibre azoteetaient en permanence faibles ou negatifs. Le contenu en fibres eleve des figues ne permettait aux cephalophes de digerer que 30 a 60% de l’energie contenue dans les figues. Cependant, pendant les quatre jours du test, les cephalophes ont pu consommer suffisamment d’energie digestible pour conserver leur masse corporelle. C’est pourquoi le systeme digestif des ruminants est benefique pour les mammiferes qui consomment des fruits tropicaux riches en fibres et en tanins, comme les figues, specialement si le mammifere est assez petit pour pouvoir en trouver une quantite suffisante pour repondre a ses besoins quotidiens en energie, et qu’il possede des adaptations qui lui permettent de reduire les effets des tanins sur la digestibilite des proteines.

Some tannins, plant-derived polyphenolic compounds, can rapidly affix to soil and affect the solubility of labile soil-N but a more complete understanding of the nature and persistence of tannin-soil interactions is needed. Forest and pasture soils from two depths were treated for 1 h with cool (23 °C) water (Control) or solutions that added 10 mg g−1 soil tannic acid (TA), an imprecisely defined mixture of galloyl esters, gallic acid (GA), a phenol, or β-1,2,3,4,6-penta-O-galloyl- d -glucose (PGG), a hydrolyzable tannin. Soluble-C and N, in treatment supernatants, was measured to uncover evidence for sorption of treatments or effects on extraction of soil-N. Significant amounts of soluble-C, added with treatments, were not recovered in supernatants indicating sorption of nearly 90% of the PGG-C, about 75% of the TA-C but less than 25% of the GA-C in surface soil. Disappearance of soluble-C from treatment supernatants was accompanied by a corresponding reduction of total phenolic content. Treatments added a negligible amount of N to soil; but while PGG and TA reduced soluble-N, in extracts from surface soil, GA had little effect. Soluble-N in extracts was composed mainly of organic-N. Effects of tannins persisted in surface soil through 12 washings with hot water (80 °C), suggesting the formation of stable complexes with soil. The proportion of initial soil-C and N remaining after all extractions was higher in samples treated with PGG or TA than either the Control or GA treatment. We conclude PGG readily sorbs to soil and reduces the solubility of soil organic-N unlike GA, its simple monomeric constituent. These differences could be especially important near the surface where quantities of soil organic matter and biological activity are comparatively large and most easily affected by management.

Research that connects ecosystem processes to genetic mechanisms has recently gained significant ground, yet actual studies that span the levels of organization from genes to ecosystems are extraordinarily rare. Utilizing foundation species from the genus Populus, in which the role of condensed tannins (CT) has been investigated aboveground, belowground, and in adjacent streams, we examine the diverse mechanisms for the expression of CT and the ecological consequences of CT for forests and streams. The wealth of data from this genus highlights the importance of form and function of CT in large-scale and long-term ecosystem processes and demonstrates the following four patterns: (1) plantspecific concentration of CT varies as much as fourfold among species and individual genotypes; (2) large within-plant variation in CT occurs due to ontogenetic stages (that is, juvenile and mature), tissue types (that is, leaves versus twigs) and phenotypic plasticity in response to the environment; (3) CT have little consistent effect on plant‐herbivore interactions, excepting organisms utilizing woody tissues (that is, fungal endophytes and beaver), however; (4) CT in plants consistently slow rates of leaf litter decomposition (aquatic and terrestrial), alter the composition of heterotrophic soil communities (and some aquatic communities) and reduce nutrient availability in terrestrial ecosystems. Taken together, these data suggest that CT may play an underappreciated adaptive role in regulating nutrient dynamics in ecosystems. These results also demonstrate that a holistic perspective from genesto-ecosystems is a powerful approach for elucidating

We investigated the effects of long-term acclimation of Eucalyptus nitens seedlings to ultraviolet-A (UV-A) irradiation (320–400 nm) on phenolic compounds (gallotannins, stilbenes, and flavonols), photochemical efficiency, and chlorophyll and carotenoid contents. Seedlings were raised under four nutrient regimes, ranging from low to high application rates, in an environment that included or excluded UV-A irradiance. Our aims were: to classify phenolic compounds that absorb in the UV-A and their relative contribution to total UV-A absorption; to identify how phenolic compounds respond to UV-A exposure and exclusion, and to determine how plant nutrient status affects acclimation of photo-and pigment-chemistry to UV-A exposure and exclusion. Gallotannins contributed to only a minor fraction of total absorption within the lower range (320–360 nm) of the UV-A spectrum. Stilbene and flavonol compounds dominated absorption within the 320–360 and 360–400 nm ranges, respectively. Contents of gallotannin were generally high in UV-A-exposed seedlings. Although there was a significant effect of UV-A on contents of stilbenes, a general response (across nutrient treatment comparisons) was not evident. Contents of flavonols were not affected by UV-A exposure. Contents of gallotannin, stilbene, and flavonols decreased from low to high nutrient-application treatments. There were no effects of UV-A on photochemical efficiency or pigment-chemistry.

The objective of the present investigation was to examine the oral astringency and protein-binding activity of four structurally well-defined tannins, namely, procyanidin [epicatechin16(4--8)catechin], pentagalloyl glucose (1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranose), castalagin, and grandinin, representing the three main structural categories of tannins, the proanthocyanidins, the gallotannins, and the ellagitannins. Astringency threshold and dos/response were determined by the half-tongue test using a trained human panel. Protein-binding stoichiometry and relative affinity were determined using radioiodinated bovine serum albumin in precipitation or competitive binding assays. Procyanidin and pentagalloyl glucose were perceived as highly astringent compounds and had relatively steep dose/response curves, but castalagin and grandinin had a lower mass threshold for detection. In vitro, procyanidin was the most effective protein-precipitating agent and grandinin the least. Increasing the temperature increased protein precipitation by the hydrolyzable tannins, especially grandinin. All four polyphenols had higher relative affinities for proline-rich proteins than for bovine serum albumin.

Plants synthesize a diversity of tannin structures but little is known about whether these different types have different oxidative activities in herbivores. Oxidative activities of hydrolyzable and condensed tannins were compared at pH 10 with two methods: EPR spectrometry was used to quantify semiquinone radicals in anoxic conditions and a spectrophotometric assay was used to measure the rate of browning of phenolics oxidized in ambient oxygen conditions. A little-studied group of hydrolyzable tannins (ellagitannins) contained the most active tannins examined, forming high concentrations of semiquinone radicals and browning at the highest rates. On average, galloyl glucoses and high-molecular-weight gallotannins had intermediate to low oxidative activities. Condensed tannins generally formed low levels of semiquinone radicals and browned most slowly. The results suggest that ellagitannin-rich plants have active oxidative defenses against herbivores, such as caterpillars, whereas the opposite may hold true for plants that contain predominantly condensed tannins or high-molecular-weight gallotannins.

A short-term isometric exercise protocol was tested in ten hypertensive individuals to determine its efficacy as a high blood pressure-reducing intervention.The study was a prospective case study of 10 hypertensive individuals (8 men, 2 woman, mean age = 52 + 5 years) who underwent six weeks of isometric exercise training (three sessions/week).Blood pressure, blood lipids and markers of oxidative stress were monitored before, during and following the isometric intervention. Electron spin resonance spectroscopy was used to directly measure radicals in the blood samples.After six weeks, systolic blood pressure decreased an average 13 mm Hg (p0.05) from a mean blood pressure of 146 to 133 mm Hg, a level that is below the usual 140 mm Hg hypertension threshold. Blood lipids were unchanged, but markers of oxidative stress were affected, with a dramatic decrease in exercise-induced oxygen centered radicals (-266%), (p0.05) and an increased resting whole blood glutathione:oxidized glutathione (+61%) in hypertensive adults following six weeks of isometric exercise.Six weeks of isometric exercise training was effective in lowering systolic but not diastolic blood pressure in pre-hypertensive and hypertensive individuals, and enhanced antioxidant protection is a likely underlying mechanism.

In Saccharomyces cerevisiae, the trans-membrane helix of Qcr8p, the ubiquinone binding protein of complex III, contributes to the Q binding site. In wild-type cells, residue 62 of the helix is non-polar (proline). Substitution of proline 62 with a polar, uncharged residue does not impair the ability of the cells to respire, complex III assembly is unaffected, ubiquinone occupancy of the Q binding site is unchanged, and mitochondrial ubiquinone levels are in the wild-type range. Substitution with a +1 charged residue is associated with partial respiratory competence, impaired complex III assembly, and loss of cytochrome b. Although ubiquinone occupancy of the Q binding site is similar to wild-type, total mitochondrial ubiquinone doubled in these mutants. Mutants with a +2 charged substitution at position 62 are unable to respire. These results suggest that the accumulation of ubiquinone in the mitochondria may be a compensatory mechanism for impaired electron transport at cytochrome b.

Cognitive performance is sensitive to both neural and non-neural changes induced by physical activity and inactivity. This study investigated whether access to physical activity outside a standard laboratory animal cage affected cognitive performance as measured by navigation of a spatial maze. It also examined gene expression in heart tissue for genes associated with cardiovascular function given recent reports of cognitive impairment associated with hyperlipidemia. Furthermore, we measured expression of neural-regulatory genes typically expressed in brain, but also found in cardiac tissue. Male Sprague-Dawley rats (n = 72) were separated into three groups having different access to physical activity: none outside a standard cage, twice-weekly physical activity, and every other day exercise on a running wheel. Compared with a sedentary group, spatial maze performance was enhanced in animals that had access to physical activity, either twice-weekly in a large box or every other day on a running wheel. Both the cardiovascular and neural-related genes expressed in the heart were distinguished by access to physical activity. Several genes that are associated with heart rate, cholesterol biosynthesis, blood pressure, and cell adhesion regulation, including GJA1, FDFT1, EDN1, and CD36, differed in animals based on access to physical activity. Neural-related genes expressed in cardiac tissue associated with neurite outgrowth, neuroplasticity, and neurogenesis including RTN4, HOMER2, ACTB, NCDN, KIF5B, and HMGB2, were expressed differently among the three groups. Significant shifts in ten cardiovascular and neural-related gene expressions in cardiac tissue were associated with physical activity and may have influenced learning and performance on a spatial maze.

To better understand the biochemical consequences when polymeric polyphenols serve as biological antioxidants, we studied how reaction pH (pH 2.1-7.4) and protein affected the oxidation of pentagalloyl glucose (PGG) by NaIO4 in aqueous solution. PGG oxidation produced an o-semiquinone radical intermediate, which tended to form polymeric products at pH values below 5, and o-quinones at higher pH. The model protein bovine serum albumin promoted the formation of quinone even at low pH. Two other polyphenols, procyanidin (epicatechin16-(4--8)-catechin) and epigallocatechin gallate, had similar pH-dependent oxidation patterns.