In vitro evaluation of the fermentation properties and potential prebiotic activity of caprine cheese whey oligosaccharides in batch culture systems

The prebiotic effect of oligosaccharides recovered and purified from caprine whey, was evaluated by in vitro fermentation under anaerobic conditions using batch cultures at 37ºC with human faeces. Effects on key gut bacterial groups were monitored over 24h by fluorescence in situ hybridisation (FISH), which was used to determine a quantitative prebiotic index score. Production of short-chain fatty acids (SCFAs) as fermentation end products was analysed by high-performance liquid chromatography (HPLC). Growth of Bifidobacterium spp was significantly higher (p ≥ 0.05) with the purified oligosaccharides compared to the negative control. Lactic and propionic acids were the main SCFAs produced. Antimicrobial activity of the oligosaccharides was also tested, revealing no inhibition though a decrease in Staphylococcus aureus and Escherichia coli growth. These findings indicate that naturally extracted oligosaccharides from caprine whey could be used as new and valuable source of prebiotics.

Gut microbial activity, implications for health and disease: the potential role of metabolite analysis

Microbial metabolism of proteins and amino acids by human gut bacteria generates a variety of compounds including phenol, indole, and sulfur compounds and branched chain fatty acids, many of which have been shown to elicit a toxic effect on the lumen. Bacterial fermentation of amino acids and proteins occurs mainly in the distal colon, a site that is often fraught with symptoms from disorders including ulcerative colitis (UC) and colorectal cancer (CRC). In contrast to carbohydrate metabolism by the gut microbiota, proteolysis is less extensively researched. Many metabolites are low molecular weight, volatile compounds. This review will summarize the use of analytical methods to detect and identify compounds in order to elucidate the relationship between specific dietary proteinaceous substrates, their corresponding metabolites, and implications for gastrointestinal health.

Human microbiota in health and disease

Each human body plays host to a microbial population which is both numerically vast (at around 1014 microbial cells) and phenomenally diverse (over 1,000 species). The majority of the microbial species in the gut have not been cultured but the application of culture-independent approaches for high throughput diversity and functionality analysis has allowed characterisation of the diverse microbial phylotypes present in health and disease. Studies in monozygotic twins, showing that these retain highly similar microbiota decades after birth and initial colonisation, are strongly indicative that diversity of the microbiome is host-specific and affected by the genotype. Microbial diversity in the human body is reflected in both richness and evenness. Diversity increases steeply from birth reaching its highest point in early adulthood, before declining in older age. However, in healthy subjects there appears to be a core of microbial phylotypes which remains relatively stable over time. Studies of individuals from diverse geopraphies suggest that clusters of intestinal bacterial groups tend to occur together, constituting ‘enterotypes’. So variation in intestinal microbiota is stratified rather than continuous and there may be a limited number of host/microbial states which respond differently to environmental influences. Exploration of enterotypes and functional groups may provide biomarkers for disease and insights into the potential for new treatments based on manipulation of the microbiome. In health, the microbiota interact with host defences and exist in harmonious homeostasis which can then be disturbed by invading organisms or when ‘carpet bombing’ by antibiotics occurs. In a portion of individuals with infections, the disease will resolve itself without the need for antibiotics and microbial homeostasis with the host’s defences is restored. The administration of probiotics (live microorganisms which when administered in adequate amounts confer a health benefit on the host) represents an artificial way to enhance or stimulate these natural processes. The study of innate mechanisms of antimicrobial defence on the skin, including the production of numerous antimicrobial peptides (AMPs), has shown an important role for skin commensal organisms. These organisms may produce AMPs, and also amplify the innate immune responses to pathogens by activating signalling pathways and processing host produced AMPs. Research continues into how to enhance and manipulate the role of commensal organisms on the skin. The challenges of skin infection (including diseases caused by multiply resistant organisms) and infestations remain considerable. The potential to re-colonise the skin to replace or reduce pathogens, and exploring the relationship between microbiota elsewhere and skin diseases are among a growing list of research targets. Lactobacillus species are among the best known ‘beneficial’ bacterial members of the human microbiota. Of the approximately 120 species known, about 15 are known to occur in the human vagina. These organisms have multiple properties, including the production of lactic acid, hydrogen peroxide and bacteriocins, which render the vagina inhospitable to potential pathogens. Depletion of the of the normal Lactobacillus population and overgrowth of vaginal anaerobes, accompanied by the loss of normal vaginal acidity can lead to bacterial vaginosis – the commonest cause of abnormal vaginal discharge in women. Some vaginal anaerobes are associated with the formation of vaginal biofilms which serve to act as a reservoir of organisms which persists after standard antibiotic therapy of bacterial vaginosis and may help to account for the characteristically high relapse rate in the condition. Administration of Lactobacillus species both vaginally and orally have shown beneficial effects in the treatment of bacterial vaginosis and such treatments have an excellent overall safety record. Candida albicans is a frequent coloniser of human skin and mucosal membranes, and is a normal part of the microbiota in the mouth, gut and vagina. Nevertheless Candida albicans is the most common fungal pathogen worldwide and is a leading cause of serious and often fatal nosocomial infections. What turns this organism from a commensal to a pathogen is a combination of increasing virulence in the organism and predisposing host factors that compromise immunity. There has been considerable research into the use of probiotic Lactobacillus spp. in vaginal candidiasis. Studies in reconstituted human epithelium and monolayer cell cultures have shown that L. rhamnosus GG can protect mucosa from damage caused by Candida albicans, and enhance the immune responses of mucosal surfaces. Such findings offer the promise that the use of such probiotic bacteria could provide new options for antifungal therapy. Studies of changes of the human intestinal microbiota in health and disease are complicated by its size and diversity. The Alimentary Pharmabiotic Centre in Cork (Republic of Ireland) has the mission to ‘mine microbes for mankind’ and its work illustrates the potential benefits of understanding the gut microbiota. Work undertaken at the centre includes: mapping changes in the microbiota with age; studies of the interaction between the microbiota and the gut; potential interactions between the gut microbiota and the central nervous system; the potential for probiotics to act as anti-infectives including through the production of bacteriocins; and the characterisation of interactions between gut microbiota and bile acids which have important roles as signalling molecules and in immunity. The important disease entity where the role of the gut microbiota appears to be central is the Irritable Bowel Syndrome (IBS). IBS patients show evidence of immune activation, impaired gut barrier function and abnormal gut microbiota. Studies with probiotics have shown that these organisms can exert anti-inflammatory effects in inflammatory bowel disease and may strengthen the gut barrier in IBS of the diarrhoea-predominant type. Formal randomised trials of probiotics in IBS show mixed results with limited benefit for some but not all. Studies confirm that administered probiotics can survive and temporarily colonise the gut. They can also stimulate the numbers of other lactic acid bacilli in the gut, and reduce the numbers of pathogens. However consuming live organisms is not the only way to influence gut microbiota. Dietary prebiotics are selectively fermented ingredients that can change the composition and/or activity of the gastrointestinal microbiota in beneficial ways. Dietary components that reach the colon, and are available to influence the microbiota include poorly digestible carbohydrates, such as non-starch polysaccharides, resistant starch, non-digestible oligosaccharides (NDOs) and polyphenols. Mixtures of probiotic and prebiotic ingredients that can selectively stimulate growth or activity of health promoting bacteria have been termed ‘synbiotics’. All of these approaches can influence gut microbial ecology, mainly to increase bifidobacteria and lactobacilli, but metagenomic approaches may reveal wider effects. Characterising how these changes produce physiological benefits may enable broader use of these tactics in health and disease in the future. The current status of probiotic products commercially available worldwide is less than ideal. Prevalent problems include misidentification of ingredient organisms and poor viability of probiotic microorganisms leading to inadequate shelf life. On occasions these problems mean that some commercially available products cannot be considered to meet the definition of a probiotic product. Given the potential benefits of manipulating the human microbiota for beneficial effects, there is a clear need for improved regulation of probiotics. The potential importance of the human microbiota cannot be overstated. ‘We feed our microbes, they talk to us and we benefit. We just have to understand and then exploit this.’ (Willem de Vos).

Phenolic acid intake, delivered via moderate champagne wine consumption, improves spatial working memory via the modulation of hippocampal and cortical protein expression/activation.

Aims: While much data exist for the effects of flavonoid-rich foods on spatial memory in rodents, there are no such data for foods/beverages predominantly containing hydroxycinnamates and phenolic acids. To address this, we investigated the effects of moderate Champagne wine intake, which is rich in these components, on spatial memory and related mechanisms relative to the alcohol- and energy-matched controls. Results: In contrast to the isocaloric and alcohol-matched controls, supplementation with Champagne wine (1.78 ml/kg BW, alcohol 12.5% vol.) for 6 weeks led to an improvement in spatial working memory in aged rodents. Targeted protein arrays indicated that these behavioral effects were paralleled by the differential expression of a number of hippocampal and cortical proteins (relative to the isocaloric control group), including those involved in signal transduction, neuroplasticity, apoptosis, and cell cycle regulation. Western immunoblotting confirmed the differential modulation of brain-derived neurotrophic factor, cAMP response-element-binding protein (CREB), p38, dystrophin, 2',3'-cyclic-nucleotide 3'-phosphodiesterase, mammalian target of rapamycin (mTOR), and Bcl-xL in response to Champagne supplementation compared to the control drink, and the modulation of mTOR, Bcl-xL, and CREB in response to alcohol supplementation. Innovation: Our data suggest that smaller phenolics such as gallic acid, protocatechuic acid, tyrosol, caftaric acid, and caffeic acid, in addition to flavonoids, are capable of exerting improvements in spatial memory via the modulation in hippocampal signaling and protein expression. Conclusion: Changes in spatial working memory induced by the Champagne supplementation are linked to the effects of absorbed phenolics on cytoskeletal proteins, neurotrophin expression, and the effects of alcohol on the regulation of apoptotic events in the hippocampus and cortex. Antioxid. Redox Signal. 00, 000-000.

Prebiotic feeding elevates central brain derived neurotrophic factor, N-methyl-D-aspartate receptor subunits and D-serine

The influence of the gut microbiota on brain chemistry has been convincingly demonstrated in rodents. In the absence of gut bacteria, the central expression of brain derived neurotropic factor, (BDNF), and N-methyl-d-aspartate receptor (NMDAR) subunits are reduced, whereas, oral probiotics increase brain BDNF, and impart significant anxiolytic effects. We tested whether prebiotic compounds, which increase intrinsic enteric microbiota, also affected brain BDNF and NMDARs. In addition, we examined whether plasma from prebiotic treated rats released BDNF from human SH-SY5Y neuroblastoma cells, to provide an initial indication of mechanism of action. Rats were gavaged with fructo-oligosaccharides (FOS), galacto-oligosaccharides (GOS) or water for five weeks, prior to measurements of brain BDNF, NMDAR subunits and amino acids associated with glutamate neurotransmission (glutamate, glutamine, and serine and alanine enantiomers). Prebiotics increased hippocampal BDNF and NR1 subunit expression relative to controls. The intake of GOS also increased hippocampal NR2A subunits, and frontal cortex NR1 and d-serine. Prebiotics did not alter glutamate, glutamine, l-serine, l-alanine or d-alanine concentrations in the brain, though GOSfeeding raised plasma d-alanine. Elevated levels of plasma peptide YY (PYY) after GOS intake was observed. Plasma from GOS rats increased the release of BDNF from SH-SY5Y cells, but not in the presence of PYY antisera. The addition of synthetic PYY to SH-SY5Y cell cultures, also elevated BDNF secretion. We conclude that prebiotic-mediated proliferation of gut microbiota in rats, like probiotics, increases brain BDNF expression, possibly through the involvement of gut hormones. The effect of GOS on components of central NMDAR signalling was greater than FOS, and may reflect the proliferative potency of GOS on microbiota. Our data therefore, provide a sound basis to further investigate the utility of prebiotics in the maintenance of brain health and adjunctive treatment of neuropsychiatric disorders.

Fat and fatty acid composition of cooked meat from UK retail chickens labelled as from organic and non-organic production systems

This study compared fat and fatty acids in cooked retail chicken meat from conventional and organic systems. Fat contents were 1.7, 5.2, 7.1 and 12.9 g/100 g cooked weight in skinless breast, breast with skin, skinless leg and leg with skin respectively, with organic meat containing less fat overall (P < 0.01). Meat was rich in cis-monounsaturated fatty acids, although organic meat contained less than did conventional meat (1850 vs. 2538 mg/100 g; P < 0.001). Organic meat was also lower (P < 0.001) in 18:3 n−3 (115 vs. 180 mg/100 g) and, whilst it contained more (P < 0.001) docosahexaenoic acid (30.9 vs. 13.7 mg/100 g), this was due to the large effect of one supermarket. This system by supermarket interaction suggests that poultry meat labelled as organic is not a guarantee of higher long chain n−3 fatty acids. Overall there were few major differences in fatty acid contents/profiles between organic and conventional meat that were consistent across all supermarkets.

The effects of tannin-containing ground pine bark diet upon nutrient digestion, nitrogen balance, and mineral retention in meat goats

Background Pine bark is a rich source of phytochemical compounds including tannins, phenolic acids, anthocyanins, and fatty acids. These phytochemicals have potential to significantly impact on animal health and animal production. The goal of this work is to measure the effects of tannins in ground pine bark as a partial feed replacement on feed intake, dietary apparent digestibility, nitrogen balance, and mineral retention in meat goats. Results Eighteen Kiko cross goats (initial BW = 31.8 ± 1.49 kg) were randomly assigned to three treatment groups (n = 6). Dietary treatments were tested: control (0 % pine bark powder (PB) and 30 % wheat straw (WS)); 15 % PB and 15 % WS, and 30 % PB and 0 % WS. Although dry matter (DM) intake and digestibility were not affected (P > 0.10) by feeding PB, neutral detergent fiber (linear; P = 0.01), acid detergent fiber (linear; P = 0.001) and lignin digestibility (linear; P = 0.01) decreased, and crude protein (CP) digestibility tended to decrease (P = 0.09) as PB increased in the diet, apparent retention of Ca (P = 0.09), P (P = 0.03), Mg (P = 0.01), Mn (P = 0.01), Zn (P = 0.01) and Fe (P = 0.09) also increased linearly. Nitrogen intake and fecal N excretion were not affected (P > 0.05) by addition of PB in the diet, but N balance in the body was quadratically increased (P < 0.01) in the 15 % PB diet compared to other diets. This may be due to more rumen escape protein and less excreted N in the urine with the 15 % PB diet. The study showed that a moderate level of tannin-containing pine bark supplementation could improve gastrointestinal nitrogen balance with the aim of improving animal performance. Conclusion These results suggest that tannin-containing PB has negative impact on fiber, lignin, and protein digestibility, but positively impacted on N-balance.

Impact of US Brown Swiss genetics on milk quality from low-input herds in Switzerland: interactions with grazing intake and pasture type

This study investigated the effect of, and interactions between, contrasting crossbreed genetics (US Brown Swiss [BS] × Improved Braunvieh [BV] × Original Braunvieh [OB]) and feeding regimes (especially grazing intake and pasture type) on milk fatty acid (FA) profiles. Concentrations of total polyunsaturated FAs, total omega-3 FAs and trans palmitoleic, vaccenic, α-linolenic, eicosapentaenoic and docosapentaenoic acids were higher in cows with a low proportion of BS genetics. Highest concentrations of the nutritionally desirable FAs, trans palmitoleic, vaccenic and eicosapentaenoic acids were found for cows with a low proportion of BS genetics (0-24% and/or 25-49%) on high grazing intake (75-100% of dry matter intake) diets. Multivariate analysis indicated that the proportion of OB genetics is a positive driver for nutritionally desirable monounsaturated and polyunsaturated FAs while BS genetics proportion was positive driver for total and undesirable individual saturated FAs. Significant genetics × feeding regime interactions were also detected for a range of FAs.

Oleaginous yeast Cryptococcus curvatus exhibits interplay between biosynthesis of intracellular sugars and lipids

Patterns of the biosynthesis ofmajor metabolites of the oleaginous yeast Cryptococcus curvatus NRRL Y-1511 were investigated during cultivation on sugar-based media. When lactose or sucrose was employed as substrate under nitrogen-limited conditions, the yeast strain accumulated high quantities of intra-cellular total sugars (ITS) at the beginning of fermentation (up to 68% w/w), with ITS values progressively decreasing to 20%, w/w, at the end of the fermentation. Decrease in ITS content and consumption of extracellular lactose led to a subsequent rise in lipid accumulation, reaching 29.8% in dry cell weight at 80 g/L of initial lactose concentration. Lactose was a more favorable substrate for lipid production than sucrose. In nitrogen-excess conditions, ITS were produced in significant quantities despite the continuous presence of nitrogen in the medium. Growth on lactose was not followed by secretion of extra-cellular b-galactosidase. High quantities of extra-cellular invertase were observed during growth on sucrose. The composition of ITS was highly influenced by the sugar used as substrate. Cellular lipids contained mainly palmitic and to lesser extent linoleic and stearic acids. This is the first report in the literature that demonstrates the interplay between the biosynthesis of intra-cellular total sugars and lipid synthesis for oleaginous yeast strains.

Rhodosporidium toruloides cultivated in NaCl-enriched glucose-based media: adaptation dynamics and lipid production

In the present report and for the first time in the international literature, the impact of the addition of NaCl upon growth and lipid production on the oleaginous yeast Rhodosporidium toruloides was studied. Moreover, equally for first time, lipid production by R. toruloides was performed under non-aseptic conditions. Therefore, the potentiality of R. toruloides DSM 4444 to produce lipid in media containing several initial concentrations of NaCl with glucose employed as carbon source was studied. Preliminary batch-flask trials with increasing amounts of NaCl revealed the tolerance of the strain against NaCl content up to 6.0% (w/v). However, 4.0% (w/v) of NaCl stimulated lipid accumulation for this strain, by enhancing lipid production up to 71.3% (w/w) per dry cell weight. The same amount of NaCl was employed in pasteurized batch-flask cultures in order to investigate the role of the salt as bacterial inhibiting agent. The combination of NaCl and high glucose concentrations was found to satisfactorily suppress bacterial contamination of R. toruloides cultures under these conditions. Batch-bioreactor trials of the yeast in the same media with high glucose content (up to 150 g/L) resulted in satisfactory substrate assimilation, with almost linear kinetic profile for lipid production, regardless of the initial glucose concentration imposed. Finally, fed-batch bioreactor cultures led to the production of 37.2 g/L of biomass, accompanied by 64.5% (w/w) of lipid yield. Lipid yield per unit of glucose consumed received the very satisfactory value of 0.21 g/g, a value amongst the highest ones in the literature. The yeast lipid produced contained mainly oleic acid and to lesser extent palmitic and stearic acids, thus constituting a perfect starting material for “second generation” biodiesel

The use of FT-IR as a screening technique for organic residue analysis of archaeological samples

A range of archaeological samples have been examined using FT-IR spectroscopy. These include suspected coprolite samples from the Neolithic site of Catalhoyuk in Turkey, pottery samples from the Roman site of Silchester, UK and the Bronze Age site of Gatas, Spain and unidentified black residues on pottery sherds from the Roman sites of Springhead and Cambourne, UK. For coprolite samples the aim of FT-IR analysis is identification. Identification of coprolites in the field is based on their distinct orange colour; however, such visual identifications can often be misleading due to their similarity with deposits such as ochre and clay. For pottery the aim is to screen those samples that might contain high levels of organic residues which would be suitable for GC-MS analysis. The experiments have shown coprolites to have distinctive spectra, containing strong peaks from calcite, phosphate and quartz; the presence of phosphorus may be confirmed by SEM-EDX analysis. Pottery containing organic residues of plant and animal origin has also been shown to generally display strong phosphate peaks. FT-IR has distinguished between organic resin and non-organic compositions for the black residues, with differences also being seen between organic samples that have the same physical appearance. Further analysis by CC-MS has confirmed the identification of the coprolites through the presence of coprostanol and bile acids, and shows that the majority of organic pottery residues are either fatty acids or mono- or di-acylglycerols from foodstuffs, or triterpenoid resin compounds exposed to high temperatures. One suspected resin sample was shown to contain no organic residues. and it is seen that resin samples with similar physical appearances have different chemical compositions. FT-IR is proposed as a quick and cheap method of screening archaeological samples before subjecting them to the more expensive and time-consuming method of GC-MS. This will eliminate inorganic samples such as clays and ochre from CC-MS analysis, and will screen those samples which are most likely to have a high concentration of preserved organic residues. (C) 2008 Elsevier B.V. All rights reserved.

Simulation of milk production by dairy cows fed sugarcane top-based diets with locally available supplements under Indian conditions

A model of sugarcane digestion was applied to indicate the suitability of various locally available supplements for enhancing milk production of Indian crossbred dairy cattle. Milk production was calculated according to simulated energy, lipogenic, glucogenic and aminogenic substrate availability. The model identified the most limiting substrate for milk production from different sugarcane-based diets. For sugarcane tops/urea fed alone, milk production was most limited by amino acid followed by long chain fatty acid availability. Among the protein-rich oil cake supplements at 100, 200 and 300 g supplement/kg total DM, cottonseed oil cake proved superior with a milk yield of 5.5, 7.3 and 8.3 kg/day, respectively. This was followed by mustard oil cake with 5.1, 6.5 and 7.6 kg/day, respectively. In the case of a protein-rich supplement (fish meal), milk yield was limited to 6.6 kg/day due to a shortage of long chain fatty acids. However, at 300 g of supplementation, energy became limiting, with a milk yield of 6.7 kg/day. Supplementation with rice bran and rice polishings at 100, 200 and 300 g restricted milk yield to 4.3, 4.9 and 5.5 and 4.5, 5.3 and 6.1 kg/day, respectively, and amino acids became the factor limiting milk production. The diet comprising basal sugarcane tops supplemented by leguminous fodder, dry fodder (e.g. rice or wheat straw) and concentrates at levels of 100, 200 and 300 g supplements/kg total diet DM proved to be the most balanced with a milk yield of 5.1, 6.7 and 9.0 kg/day, respectively.

Major advances in fundamental dairy cattle nutrition

Fundamental nutrition seeks to describe the complex biochemical reactions involved in assimilation and processing of nutrients by various tissues and organs, and to quantify nutrient movement (flux) through those processes. Over the last 25 yr, considerable progress has been made in increasing our understanding of metabolism in dairy cattle. Major advances have been made at all levels of biological organization, including the whole animal, organ systems, tissues, cells, and molecules. At the whole-animal level, progress has been made in delineating metabolism during late pregnancy and the transition to lactation, as well as in whole-body use of energy-yielding substrates and amino acids for growth in young calves. An explosion of research using multicatheterization techniques has led to better quantitative descriptions of nutrient use by tissues of the portal-drained viscera (digestive tract, pancreas, and associated adipose tissues) and liver. Isolated tissue preparations have provided important information on the interrelationships among glucose, fatty acid, and amino acid metabolism in liver, adipose tissue, and mammary gland, as well as the regulation of these pathways during different physiological states. Finally, the last 25 yr has witnessed the birth of "molecular biology" approaches to understanding fundamental nutrition. Although measurements of mRNA abundance for proteins of interest already have provided new insights into regulation of metabolism, the next 25 yr will likely see remarkable advances as these techniques continue to be applied to problems of dairy cattle biology. Integration of the "omics" technologies (functional genomics, proteomics, and metabolomics) with measurements of tissue metabolism obtained by other methods is a particularly exciting prospect for the future. The result should be improved animal health and well being, more efficient dairy production, and better models to predict nutritional requirements and provide rations to meet those requirements.

Effects of defoliation on blackcurrant infection by Armillaria species

Field inoculation trials and laboratory studies were used to investigate the effects of defoliation stress on potted black currant plants and the infection by English and African isolates of Armillaria. Defoliation has varying effects on the carbohydrate, fatty acids and amino acids contents of roots. All isolates of Armillaria tested infected black currant plants irrespective of stress treatment; with two of the test isolates, more of the infected plants were killed with defoliation treatment. Media supplemented with water extract from defoliated roots stimulated growth of isolates compared to media supplemented with extract from non-defoliated control root tissues. The differences observed in the pathogenic behaviour of isolates, may be of importance in the epidemiology of Armillaria infections.

Organic acid formation in the gas-phase ozonolysis of alpha-pinene

The mechanism of formation of pinonic and norpinonic acids from alpha-pinene ozonolysis has been investigated by studying the products of the ozonolysis of an enone derived from alpha-pinene using gas chromatography coupled to mass spectrometry.