Irene Mueller-Harvey

Linkage of p-coumaroyl and feruloyl groups to cell-wall polysaccharides of barley straw

Abstract Treatment of cell walls of barley straw with Oxyporus “cellulase” (a mixture of polysaccharide hydrolases) released compounds containing p-coumaroyl and feruloyl groups bound to carbohydrates, two of which were identified as O-[5-O-(trans-p-coumaroyl)-α- l -arabinofuranosyl]-(1→3)-O-β- d -xylopyranosyl-(1→4)- d -xylopyranose (PAXX) and O-[5-O-(trans-feruloyl)-α- l -arabinofuranosyl]-(1→3)-O-β- d -xylopyranosyl-(1→4)- d -xylopyranose (FAXX).

Analysis of hydrolysable tannins

This review deals with the main issues relating to the analysis of hydrolysable tannins (HTs). It covers briefly their distribution in the plant kingdom and describes their main structural features. HTs have often been ignored because they are apparently more difficult to analyse than condensed tannins. Meaningful analytical data are critically dependent on appropriate sample preparation, storage and extraction techniques. This requires some understanding of the reactivities of hydrolysable tannins. Mixtures of HTs have been measured by general tannin assays, such as precipitation with metals or proteins, and by colorimetric assays for total phenols. Some HTs can also be measured by more specific colorimetric tests. Although colorimetric assays are widely used for tannin analyses, they generally do not provide accurate quantitative data. At best, they provide data for comparative purposes. Thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) are useful tools to screen samples for the different types of tannins, hydrolysable or condensed tannins. In addition, there are physicochemical techniques, such as nuclear magnetic resonance (NMR) and mass spectrometry (MS), which are used to identify pure tannin compounds. More recently, other MS techniques have been developed that are capable of yielding molecular weights of crude tannin mixtures.

Interactions of tea tannins and condensed tannins with proteins.

Binding parameters for the interactions of four types of tannins: tea catechins, grape seed proanthocyanidins, mimosa 5-deoxy proanthocyanidins, and sorghum procyanidins (mDP=17), with gelatin and bovine serum albumin (BSA) have been determined from isothermal titration calorimetry data. Equilibrium binding constants determined for the interaction with gelatin were in the range 10(4) to 10(6) M(-1) and in the order: sorghum procyanidins > grape seed proanthocyanidins > mimosa 5-deoxy proanthocyanidins > tea catechins. Interaction with BSA was generally weaker, with equilibrium binding constants of < or =10(3)M(-1) for grape seed proanthocyanidins, mimosa 5-deoxy proanthocyanidins and tea catechins, and 10(4)M(-1) for the sorghum procyanidins. In all cases the interactions with proteins were exothermic and involved multiple binding sites on the protein. The data are discussed in relation to the structures and the known nutritional effects of the condensed tannins.

Potential of legume-based grassland-livestock systems in Europe: a review.

European grassland-based livestock production systems face the challenge of producing more meat and milk to meet increasing world demands and to achieve this using fewer resources. Legumes offer great potential for achieving these objectives. They have numerous features that can act together at different stages in the soil–plant–animal–atmosphere system, and these are most effective in mixed swards with a legume proportion of 30–50%. The resulting benefits include reduced dependence on fossil energy and industrial N-fertilizer, lower quantities of harmful emissions to the environment (greenhouse gases and nitrate), lower production costs, higher productivity and increased protein self-sufficiency. Some legume species offer opportunities for improving animal health with less medication, due to the presence of bioactive secondary metabolites. In addition, legumes may offer an adaptation option to rising atmospheric CO2 concentrations and climate change. Legumes generate these benefits at the level of the managed land-area unit and also at the level of the final product unit. However, legumes suffer from some limitations, and suggestions are made for future research to exploit more fully the opportunities that legumes can offer. In conclusion, the development of legume-based grassland–livestock systems undoubtedly constitutes one of the pillars for more sustainable and competitive ruminant production systems, and it can be expected that forage legumes will become more important in the future.

Determination of the substrates for sulphate-reducing bacteria within marine and estuarine sediments with different rates of sulphate reduction

The substrates used by sulphate-reducing bacteria in sediment slurries from Loch Eil, Loch Etive and the Tay estuary were determined by selectively inhibiting sulphate reduction with 20 mM-molybdate and measuring the resultant substrate accumulation. Substrate accumulation was linear after molybdate addition, and the rate of accumulation closely matched sulphate reduction rates, indicating that metabolic pathways other than those specifically involving sulphate reduction were not affected by the inhibitor. In sediments from all three sites acetate was a major substrate, although the percentage of sulphate reduced due to acetate oxidation varied considerably among the sites (Tay estuary, 35%; Loch Eil, 64%; Loch Etive, 100%). In addition to acctate, 17 individual substrates were shown to be involved in sulphate reduction to varying extents in the Tay estuary and Loch Eil sediments; these included lactate, H2, propionate, iso- and n-butyrate, iso- and n-valerate, 2-methylbutyrate and amino acids. At both sites propionate accounted for between 6 and 12% of sulphate reduction. Butyrate (n- and iso-), iso-valerate and 2-methylbutyrate were of approximately equal importance at each site and together accounted for 13 and 11%, respectively, of the sulphate reduction in the Tay estuary and Loch Eil sediments. Lactate was only importnat in the Tay estuary sediments, where it accounted for 43% of sulphate reduction. The rate of accumulation of amino acids was greatest in the Tay estuary sediments, but the contribution of amino acids to sulphate reduction was higher in the Loch Eil (9%) than in the Tay estuary sediments (2%). Of the 21 individual amino acids that were measured there was a linear increase in nine; the most important of these were serine, glutamate and arginine. In general, when sulphate reduction rates were high the substrates for this process were more varied than when rates were low. Combining the results of two experiments and assuming complete degradation of the individual substrates, almost all the sulphate reduction could be accounted for at each site (Tay estuary, 101%; Loch Eil, 98%; Loch Etive, > 100%).

Potential of legume-based grassland-livestock systems in Europe.

European grassland-based livestock production systems face the challenge of producing more meat and milk to meet increasing world demands and to achieve this using fewer resources. Legumes offer great potential for achieving these objectives. They have numerous features that can act together at different stages in the soil–plant–animal–atmosphere system, and these are most effective in mixed swards with a legume proportion of 30–50%. The resulting benefits include reduced dependence on fossil energy and industrial N-fertilizer, lower quantities of harmful emissions to the environment (greenhouse gases and nitrate), lower production costs, higher productivity and increased protein self-sufficiency. Some legume species offer opportunities for improving animal health with less medication, due to the presence of bioactive secondary metabolites. In addition, legumes may offer an adaptation option to rising atmospheric CO2 concentrations and climate change. Legumes generate these benefits at the level of the managed land-area unit and also at the level of the final product unit. However, legumes suffer from some limitations, and suggestions are made for future research to exploit more fully the opportunities that legumes can offer. In conclusion, the development of legume-based grassland–livestock systems undoubtedly constitutes one of the pillars for more sustainable and competitive ruminant production systems, and it can be expected that forage legumes will become more important in the future.

Characterisation of tannins and in vitro protein digestibility of several Lotus corniculatus varieties

Seven birdsfoot trefoil (BFT) varieties (Lotus corniculatus) grown in Sweden, were harvested at the 50% flowering stage and analysed for tannins by the radial diffusion and HCl–butanol methods. The flavan-3-ol composition of different BFT tannins was determined by HPLC. Tannins were isolated and examined for their molecular weight distributions by HPLC gel permeation chromatography (GPC) and MALDI-TOF mass spectrometry. Ruminal protein degradability was determined in vitro and related to tannin chemistry. Tannin concentrations of the BFT varieties were generally low and ranged between 0.3–1.0% (radial diffusion assay) and 0.2–1.7% (HCl–butanol assay) on a DM basis. The delphinidin:cyanidin ratios showed considerable variation ranging from 16:84 to 33:67 amongst the seven varieties. GPC analysis revealed small differences between the varieties with most of the variation occurring in the relative proportions of the higher molecular weight tannins. MALDI-TOF mass spectrometry of tannins from two varieties gave well-resolved spectra of tetramers, pentamers and hexamers. Oligomers up to the decamers were also detectable. Each of these oligomers had a subset of structures incorporating catechin/epicatechin (CE) and gallocatechin/epigallocatechin (GE) units. Some homopolymers containing CE units only (i.e. procyanidins), but none with GE units only (i.e. prodelphinidins), were detected. Most mixed CE/GE oligomers of all sizes contained one or two GE units. There were significant differences (P≤0.05) in vitro N-degradability between four varieties. The data suggest that degradability of the soluble proteins in birdsfoot trefoil were negatively correlated to tannin concentrations (R2=0.93) despite the fact that their overall concentrations were very low.

In situ analysis and structural elucidation of sainfoin (Onobrychis viciifolia) tannins for high-throughput germplasm screening

A rapid thiolytic degradation and cleanup procedure was developed for analyzing tannins directly in chlorophyll-containing sainfoin ( Onobrychis viciifolia ) plants. The technique proved suitable for complex tannin mixtures containing catechin, epicatechin, gallocatechin, and epigallocatechin flavan-3-ol units. The reaction time was standardized at 60 min to minimize the loss of structural information as a result of epimerization and degradation of terminal flavan-3-ol units. The results were evaluated by separate analysis of extractable and unextractable tannins, which accounted for 63.6-113.7% of the in situ plant tannins. It is of note that 70% aqueous acetone extracted tannins with a lower mean degree of polymerization (mDP) than was found for tannins analyzed in situ. Extractable tannins had between 4 and 29 lower mDP values. The method was validated by comparing results from individual and mixed sample sets. The tannin composition of different sainfoin accessions covered a range of mDP values from 16 to 83, procyanidin/prodelphinidin (PC/PD) ratios from 19.2/80.8 to 45.6/54.4, and cis/trans ratios from 74.1/25.9 to 88.0/12.0. This is the first high-throughput screening method that is suitable for analyzing condensed tannin contents and structural composition directly in green plant tissue.

The effect of drying treatment on the fodder quality and tannin content of two provenances of Calliandra calothyrsus Meissner

The effects of freeze drying and of air drying at 30 °C on the nutritive value of Calliandra calothyrsus leaves were evaluated, using material from two provenances grown together under controlled greenhouse conditions. Freshly harvested leaf of each provenance was compared with the dried treatments in terms of in vitro digestibility, content of extractable, protein-bound and fibre-bound proanthocyanidin (condensed tannin) and protein precipitation activity of the extractable fraction, as well as crude protein and fibre content. The effect of long-term (12 months) storage of fresh-frozen and dried leaf was also evaluated. Provenance was found to have a major effect on most of the traits, and large differences between the provenances in proanthocyanidin structure were found by high-performance liquid chromatography, with the tannins of one provenance consisting mainly of catechin/epicatechin subunits (producing cyanidin on treatment with butanol/HCl), and those of the other mainly gallocatechin/epigallocatechin subunits (producing delphinidin with butanol/HCl). This also resulted in large differences in colour yield in the butanol/HCl assay for proanthocyanidins, necessitating separate standards for each provenance. Both drying treatments tended to increase digestibility, though the effect was only significant for one provenance. Air drying, but not freeze drying, increased proanthocyanidin binding to protein, but not to fibre, in both provenances. Long-term freezing reduced proanthocyanidin content by about half and should be avoided in tannin research. The study suggests that careful air drying, without excessive heating, may not reduce the nutritive value of C calothyrsus, though this needs validation through feeding studies with ruminants. © 2000 Society of Chemical Industry

Sainfoin (Onobrychis viciifolia): A beneficial forage legume

The Onobrychis genus comprises a few agronomically important forage legume species, with sainfoin ( Onobrychis viciifolia ) being the most widespread. O. viciifolia has a long history of traditional culture worldwide, but its use has declined in western countries over the last decades. It suffers from low productivity and is more difficult to maintain than other legumes but is known to have valuable characteristics such as palatability and drought tolerance. Recent studies suggest that it has several other highly beneficial properties due to its unique tannin and polyphenol composition. Condensed tannins present in Onobrychis species have been shown to confer anthelmintic properties, increase protein utilization and prevent bloating; they may also have the potential to reduce greenhouse gas emissions. Positive effects on wildlife and honey production could also be advantageous in the context of sustainable farming. Modern breeding programmes have not been a priority, leading to a lack of genetic knowledge in comparison to extensively used forage legumes. It is expected that potential for O. viciifolia improvements could be achieved by rigorous characterization of the available germplasm and utilization of characters derived from close relatives of the genus. Breeding priorities for the future would include enhanced germination and improved early establishment, allied to the best anthelmintic properties observed in some varieties.