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A procedure was developed for extraction of 'free' condensed tannins (CT) using a mixture of acetone/water/diethyI ether (4.7 : 2.0 : 3.3), followed by extraction of protein-bound and fibre-bound CT using boiling sodium dodecyl sulphate containing 2-mercaptoethanol (SDS). CT concentrations in all three fractions were determined by a modified butanolLHC1 procedure. Separate standard curves using purified CT in water or SDS solution were utilised for analysis of extractable C T (water standards) and protein-bound and fibre-bound CT (SDS standards). The method accurately predicted the concentration of CT added to forage extracts. CT extractable in acetone/water/diethyI ether comprised, on average, 68 YO of total CT in a range of freeze dried forage legume samples, with most of the remainder being bound to protein. When total CT concentration was low (0.6-3.0 YO DM). a lower proportion was extractable (33-35 O h ) . In protein concentrate meals containing CT, the extractable. proteinbound and fibre-bound components comprised 15, 60 and 25% respectively of total CT. Total CT concentration in the forages Lorus cornicukutus and Coronillu ouriu was considered appropriate for ruminant nutrition (2.1 and 3.0 YO DM), whilst CT concentration in the forage of Dorycnium spp (13-19 % DM) was more suitable for soil conservation purposes. The substantial CT concentration in cottonseed meal (1.6% DM) may be involved in the high resistance of proteins in this product to ruminal degradation. CT concentration was indistinguishable from zero in perennial ryegrass forage, in barley and triticale grains and in soya bean meal (0.1 YO DM).

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1. Vegetative secondary growth Lotus pedunculatus was cut daily, and fed fresh at hourly intervals (600 g dry matter (DM)/d) to three groups each of three sheep fitted with permanent cannulas into the rumen and duodenum. Lotus fed to two of the groups was sprayed with low and high rates of polyethylene glycol (PEG; molecular weight 3350), which specifically binds the condensed tannins (CT). Nutrient intake and faecal excretion were measured directly, duodenal flows estimated from continuous intraruminal infusion of inert ruthenium phenanthroline (Ru-P) and CrEDTA markers, and rumen pool sizes measured at slaughter.2. Dietary concentrations of total reactive CT (i.e. that not bound to PEG) were 95,45 and 14 g/kg DM, whilst the corresponding values for free CT were 15, 5 and 2 g/kg DM.3. Increasing dietary reactive CT concentration linearly increased duodenal flows of non-ammonia nitrogen, but linearly decreased the apparent digestibility of energy and organic matter, and rumen digestion of hemicellulose but not of cellulose. Rumen digestion as a proportion of total digestion was increased by the higher PEG rate for organic matter, energy, pectin and lignin.4. High dietary CT concentration was associated with increased N retention. Rumen ammonia concentration and pool size showed only a slight decline on this diet, indicating that there must have been increased recycling of N into the rumen.5. Increasing dietary reactive CT concentration had no effect on the rate at which carbohydrate constituents were degraded in the rumen per unit time (FDR), but increased the rate at which their undegraded residues (FOR) left the rumen per unit time. The latter appeared to be the principal mechanism by which rumen digestion as a proportion of total digestion was reduced at high dietary CT concentrations. From a comparison of FDR and FOR of carbohydrate components in lotus and Brassica oleracea diets, it was concluded that hemicellulose digestion was rate-limiting for rumen cell-wall digestion, probably due to bonding with lignin. However, the considerable post-rumen digestion of hemicellulose was not associated with post-rumen lignin digestion.6. It was concluded that a desired concentration of CT in Lotus sp. should represent a balance between the positive effect of CT in improving the efficiency of N digestion and their negative effect in depressing rumen carbohydrate digestion. A recommended concentration is 3 W O g/kg DM.From measurements of duodenal non-ammonia nitrogen (NAN) flow it has been estimated that the absorption of essential amino acids was limiting the output of high producing ruminants consuming fresh forages ad lib., and this has been verified through post-rumen supplementation studies with protein (Barry 198 1, 1982; Beever & Siddons, 1986). From a review of New Zealand (NZ) literature, Barry & Reid (1986) concluded that the presence of condensed tannins (CT) uniformly distributed throughout leaf and stem tissue in forage plants would increase amino acid supply through CT reacting with plant proteins by reve...

Three experiments were conducted to determine the fate of condensed tannins (CT) during digestion in sheep. CT were measured as extractable, protein-bound and fibre-bound fractions using the butanol-HC1 procedure. In Expt 1, purified CT were added to digesta from different parts of the digestive tract obtained from a pasture-fed sheep. Recoveries of CT after 0 and 4 h of anaerobic incubation at 39" averaged: rumen 78-9 and 575 % ; abomasum 50.9 and 490 YO ; duodenum 64.4 and 46.0 YO and ileum 43.4 and 38.8 %. In Expt 2, ['4C]CT was given per ubomusum over a 6 5 h period at 15 min intervals to a sheep previously fed on Lotus peduncuZutus (which contains CT). The sheep was killed at the end of the period and 92.4 % of the label was recovered. Virtually all of the label was in the digesta, and none was detected in the blood, so that the CT-carbon appeared not to be absorbed from the small intestine. In Expt 3, rumen, abomasal and ileal digesta and faeces samples from sheep fed on Lotus pedunculutus were analysed for CT and CT flow along the digestive tract calculated from reference to indigestible markers. Values were low in all digesta samples, indicating disappearance of CT across the rumen and small intestine, and CT recovery in faeces was only about 15% of intake. However, the I4C results from Expt 2 suggested that little if any CT-carbon was absorbed and the low recoveries in Expt 1 are considered to be a consequence of either conformational changes to the CT molecule such that it is no longer detectable by colorimetric methods, an inability of the analytical method to release bound CT for the butanol-HC1 assay, or interference from other digesta constituents. It is concluded that the butanol-HC1 method of CT analysis is appropriate for quantifying CT in herbages but not in digesta or faeces, and that a substantial part of CT released during protein digestion in the small intestine may not be detectable by normal CT analytical methods.['4C]Condensed tannin: Digesta tannins : Tannin digestion Condensed tannins (CT) are polyphenolic compounds occurring in a wide range of plants eaten by ruminants (Jones et al. Sarkar et al. 1976; Terrill et al. 1992b). They may be either beneficial or detrimental to the animals' nutrition depending on concentration in the forage, astringency and pH-dependent protein-binding characteristics. The concentration of CT may be as high as 200g/kg plant dry matter (DM) but most of the herbages ingested by ruminants contain 2&100gCT/kg DM (Terrill et al. 1992b). Evaluation of the role played by CT in ruminant nutrition has been hindered by analytical difficulties, especially the quantification of CT in digesta and faecal material. There are two commonly used procedures for CT quantification; the vanillin-HC1 method (Broadhurst & Jones, 1978) and the n-butanol-HC1 method (Bate-Smith 1973; Porter et al. 1986). Both methods are semi-quantitative (Mangan, 1988) so that considerable variation can exist

Variableregions of the 16s ribosomal RNA have been frequently used as the target for DNA probes to identify microorganisms. In some situations, however, there is very little sequence variation observed between the 16s rRNA genes of closely related microorganisms. This study presents a general method to obtain species-specific probes using the spacer (intergenic) region between the 16s and 23s rRNA genes. The overall strategy is analogous to that which has previously been developed for the variable regions of the 16s rRNA genes. Sequence analysis of the 16s rRNA and 23s rRNA coding sequences flanking the spacer regions resulted in the design of PCR primers that can be used to amplify the spacer regions of a wide range of eubacterla.Sequencing the amplified spacer region then gives rise to the information that can be used to select specific DNA sequences for use as a DNA probe or for the generation of specific PCR primers to a microorganism of interest. In this study the approach to develop specific DNA markers for members of the genus Clostridium is described in detail. A specific DNA oligonucleotide probe and PCR primers have been designed for Clostridium perfringens that distinguish it from other organisms in the genus. Theapplication of DNA probe methodology to the identification and detection of microorganisms is becoming well established in the field of diagnostic bacteriology. Fundamental to such a technology is the ability to define suitable nucleic acid sequences that identify a particular microorganism or group of related microorganisms. DNA probes have been used successfully in the identification and detection of microorganisms./1,21 In general, nucleic acid sequences that are used as DNA probe targets for microorganisms fall into five main categories: (1) DNA sequences that code for antigens, (2) DNA sequences that code for toxins, (3) DNA sequences identified by differential hybridization using total DNA probes from related species against a DNA bank made from the microorganism of interest, (4) unique plasmid-borne DNA sequences, and (5) ribosomal RNA (rRNA) sequences.In the case of the first four categories, the means by which the DNA targets are selected involve laborious cloning methodologies and frequently a signficant amount of biochemical or immunological data. Subsequently, cross-reactivity, nonspecificity, and the possibility of the loss of the target sequences due to recombination and deletion events, especially with relation to plasmid-borne sequences, results, in some cases, in a limited usefulness of these probes, rRNAs in general have been the main targets for the generation of DNA markers for microorganisms, and we have used this region as the target for DNA probes for a number of microorganisms. (2) The major disadvantage these sequences have as candidates for DNA markers is that the "variable" regions can almost be identical when closely related microorganisms are examined, resulting in the need for finely defined stringency conditions when using such probes to detect a microorganism of...

Chicory (Cichorium intybus) is perhaps best known for the extract of its roots used as an ingredient in ‘coffee substitute’ beverages. It is less well known as a grazed forage for ruminants. Thomas et al. (1952) reported the high content of some major and minor trace minerals in chicory grown in the UK, and commented on its use in pasture mixtures as a source of these minerals. Chicory was first mentioned in New Zealand (NZ) literature as an animal forage by Cockayne (1915), but a long period then elapsed before Lancashire (1978) reported its excellent value for forage production under rotational grazing in dry summer conditions. Plant selection then followed and the cultivar ‘Grasslands Puna’ was approved for commercial release as a grazed forage plant in 1985 (Rumball 1986). The use of Puna chicory has now spread throughout NZ and the variety is also being used commercially in Australia, North America and South America and is being evaluated in parts of Europe and Asia (W. Green, personal communication). Chicory is a herb, whereas other temperate forages used for ruminant production are either grasses or legumes. This paper reviews work on the chemical composition, nutritive value and feeding value of chicory relative to perennial ryegrass (Lolium perenne) and to red clover (Trifolium pratense), a legume that, like chicory, is used as a forage for dry summer conditions. Throughout this paper, feeding value is defined as the animal production response to grazing a forage under unrestricted conditions (Ulyatt 1973), with its components being voluntary feed intake (VFI), the digestive process and the efficiency of utilization of digested nutrients; the latter two comprise nutritive value/dry matter (DM) eaten.