Kaisa Poutanen

Interlaboratory testing of methods for assay of xylanase activity

Abstract Twenty laboratories participated in a collaborative investigation of assays for endo-1,4-β-xylanase activity based on production of reducing sugars from polymeric 4- O -methyl glucuronoxylan. The substrates and methods already in use in the different laboratories were first recorded and the apparent activities obtained using these methods in the analysis of a distributed enzyme sample were compared. The standard deviation of the results reported in this analysis was 108% of the mean. Significant reduction in interlaboratory variation was obtained when all the participants used the same substrate for activity determination, each with their own assay procedure. The level of agreement was further improved when both the substrate and the method procedure were standardized. In a round robin testing of a single substrate and method, including precise instructions for enzyme dilution, the standard deviation between the results after the rejection of two outliers was 17% of the mean. This figure probably reflects the inherently poor reproducibility of results when using only partially soluble, poorly defined and rather impure polymeric substrates. The final level of variation was however low enough to allow meaningful comparison of results obtained in different laboratories when using the standardized assay substrate and method procedure. Fifteen laboratories also participated in preliminary testing of an assay based on the release of dyed fragments from 4- O -methyl glucuronoxylan dyed with Remazol Brilliant Blue dye. High values of the coefficients of correlation indicated good linearity between the amount of dyed fragments released and enzyme concentration. The relative standard deviations of the results obtained by fifteen laboratories were about 30% for an optimum range of xylanase activity in the reaction mixture.

IMPACT OF DIETARY POLYPHENOLS ON CARBOHYDRATE METABOLISM

Polyphenols, including flavonoids, phenolic acids, proanthocyanidins and resveratrol, are a large and heterogeneous group of phytochemicals in plant-based foods, such as tea, coffee, wine, cocoa, cereal grains, soy, fruits and berries. Growing evidence indicates that various dietary polyphenols may influence carbohydrate metabolism at many levels. In animal models and a limited number of human studies carried out so far, polyphenols and foods or beverages rich in polyphenols have attenuated postprandial glycemic responses and fasting hyperglycemia, and improved acute insulin secretion and insulin sensitivity. The possible mechanisms include inhibition of carbohydrate digestion and glucose absorption in the intestine, stimulation of insulin secretion from the pancreatic β–cells, modulation of glucose release from the liver, activation of insulin receptors and glucose uptake in the insulin-sensitive tissues, and modulation of intracellular signalling pathways and gene expression. The positive effects of polyphenols on glucose homeostasis observed in a large number of in vitro and animal models are supported by epidemiological evidence on polyphenol-rich diets. To confirm the implications of polyphenol consumption for prevention of insulin resistance, metabolic syndrome and eventually type 2 diabetes, human trials with well-defined diets, controlled study designs and clinically relevant end-points together with holistic approaches e.g., systems biology profiling technologies are needed.

Oat β-glucan and xylan hydrolysates as selective substrates for Bifidobacterium and Lactobacillus strains

Abstract Novel oligomers that resist digestion in the upper gut were prepared from oat mixed-linked β-glucan and xylan by enzymatic hydrolysis with lichenase of Bacillus subtilis and xylanase of Trichoderma reesei respectively. The low-molecular-mass hydrolysis products of β-glucan and xylan were compared with fructooligomers and raffinose in their ability to provide growth substrates for probiotic (Lactobacillus and Bifidobacterium) and intestinal (Bacteroides, Clostridium and Escherichia coli) strains in vitro. A degradation profile of each carbohydrate and total sugar consumption were analysed with HPLC, and bacterial growth rate with an automatic turbidometer, the Bioscreen C system. β-Glucooligomers and xylooligomers both enhanced the growth of health-promoting probiotic strains as compared with intestinal bacterial growth, but not to a significant level. Raffinose stimulated the probiotic strains significantly, whereas fructooligomers induced high average growth for intestinal bacteria also.

Sourdough and cereal fermentation in a nutritional perspective

Use of sourdough is of expanding interest for improvement of flavour, structure and stability of baked goods. Cereal fermentations also show significant potential in improvement and design of the nutritional quality and health effects of foods and ingredients. In addition to improving the sensory quality of whole grain, fibre-rich or gluten-free products, sourdough can also actively retard starch digestibility leading to low glycemic responses, modulate levels and bioaccessibility of bioactive compounds, and improve mineral bioavailability. Cereal fermentation may produce non-digestible polysaccharides, or modify accessibility of the grain fibre complex to gut microbiota. It has also been suggested that degradation of gluten may render bread better suitable for celiac persons. The changes in cereal matrix potentially leading to improved nutritional quality are numerous. They include acid production, suggested to retard starch digestibility, and to adjust pH to a range which favours the action of certain endogenous enzymes, thus changing the bioavailability pattern of minerals and phytochemicals. This is especially beneficial in products rich in bran to deliver minerals and potentially protective compounds in the blood circulation. The action of enzymes during fermentation also causes hydrolysis and solubilisation of grain macromolecules, such as proteins and cell wall polysaccharides. This changes product texture, which may affect nutrient and non-nutrient absorption. New bioactive compounds, such as prebiotic oligosaccharides or other metabolites, may also be formed in cereal fermentations.

Two major xylanases of Trichoderma reesei

Abstract The two main xylanases produced by Trichoderma reesei were purified to electrophoretic homogeneity by ion-exchange and gel chromatography. The enzymes had isoelectric points of 9.0 and 5.5 and their molecular masses were 20 and 19 kDa, respectively. The purified xylanases were most probably distinct gene products, as they were also serologically dissimilar and had different proteolytic cleavage patterns. The pI 9.0 xylanase tolerated higher pH values and temperatures than the pI 5.5 xylanase. Both enzymes clearly preferred polymeric substrates to xylo-oligosaccharides. The substrate preference for polymeric xylans decreased with decreasing substitution (decreasing solubility). The apparent K m values for different xylans varied from 3.0 to 6.8 mg ml -1 for the pI 9.0 xylanase and from 14.8 to 22.3 mg ml -1 for the pI 5.5 xylanase. The pI 9.0 xylanase also showed transxylosidase activity with xylotetraose and xyloheptaose as substrates.

Characteristics of Trichoderma reesei ?-xylosidase and its use in the hydrolysis of solubilized xylans

SummaryThe β-xylosidase (EC 3.2.1.37) of Trichoderma reesei was purified and its characteristics and use in the hydrolysis of steamed birch xylan were studied. The enzyme was a glycoprotein with a molecular weight of 100000 as determined by SDS-gel electrophoresis and its isoelectric point was 4.7. The pH optimum was 4.0 and temperature optimum 60°C. β-Xylosidase was competitively inhibited by xylose and the inhibition constant was 2.3 mM. The purified enzyme also showed α-arabinofuranosidase activity.

Viscosity of Oat Bran-Enriched Beverages Influences Gastrointestinal Hormonal Responses in Healthy Humans

Viscous fibers, including beta-glucan in oat bran, favorably affect satiety as well as postprandial carbohydrate and lipid metabolism. However, effects of fiber viscosity on modulation of satiety-related gut hormone responses are largely unknown. We examined the effects of modified oat bran, with or without its natural viscosity, on sensations of appetite and satiety-related gastrointestinal (GI) hormone responses to establish the relevance of viscosity of beta-glucan in oat bran. Twenty healthy, normal-weight participants (16 female, 4 male, aged 22.6 +/- 0.7 y) ingested 2 isocaloric (1250 kJ) 300-mL oat bran beverages with low or high viscosity (carbohydrates, 57.9 g; protein, 7.8 g; fat, 3.3 g; fiber, 10.2 g) after a 12-h fast in randomized order. Viscosity of the low-viscosity oat bran beverage was reduced by beta-glucanase treatment. Blood samples were drawn before and 15, 30, 45, 60, 90, 120, and 180 min after beverage consumption. The oat bran beverage with low viscosity induced a greater postprandial increase in satiety (P = 0.048) and plasma glucose (P < 0.001), insulin (P = 0.008), cholecystokinin (P = 0.035), glucagon-like peptide 1 (P = 0.037), and peptide YY (P = 0.051) and a greater decrease in postprandial ghrelin (P = 0.009) than the beverage with high-viscosity oat bran. Gastric emptying as measured by paracetamol absorption was also faster (P = 0.034) after low-viscosity oat bran beverage consumption. In conclusion, viscosity differences in oat beta-glucan in a liquid meal with identical chemical composition strongly influenced not only glucose and insulin responses, but also short-term gut hormone responses, implying the importance of food structure in the modulation of postprandial satiety-related physiology.

The HEALTHGRAIN Cereal Diversity Screen : Concept, Results, and Prospects

One hundred and fifty bread wheat lines and 50 other lines of small-grain cereals (spelt, durum wheat, Triticum monococcum, Triticum dicoccum, oats, rye, and barley) were selected for diversity in their geographical origin, age, and characteristics. They were grown on a single site in Hungary in 2004-2005, harvested, milled, and analyzed for a range of phytochemicals (tocols, sterols, phenolic acids, folates, alkylresorcinols) and fiber components that are considered to have health benefits. Detailed analyses of these components in the different species are reported in a series of accompanying papers. The present paper discusses the comparative levels of the bioactive components in the different species, showing differences in both ranges and mean amounts. Furthermore, detailed comparisons of the bread wheat lines show that it is possible to identify lines in which high levels of phytochemicals and dietary fiber components are combined with good yield and processing quality. This means that commercially competitive lines with high levels of bioactive components are a realistic goal for plant breeders.

Process-induced changes on bioactive compounds in whole grain rye.

Manufacturing of healthy wholegrain foods demands knowledge of process-induced changes in macro-, micro- and non-nutrients. The high content of dietary fibre is a challenge in relation to good product texture and sensory quality. The stability and bioavailability of bioactive compounds have a marked influence on the health effects of cereal foods. It was confirmed that sterols, folates, tocopherols and tocotrienols, alkylresorcinols, lignans, phenolic acids and total phenolics are concentrated in the bran layers of the rye grain, and are only present at low levels in the flour endosperm. The levels of folate and easily-extractable phenolic compounds increase in germination and sourdough baking, but there are negligible changes in the levels of sterols, lignans and alk(en)ylresorcinols. The levels of tocopherols and tocotrienols are reduced during the sourdough fermentation. In conclusion, many of the bioactive compounds in wholegrain rye are stable during food processing, and their levels can even be increased with suitable processing.

In vitro fermentation of polysaccharides of rye, wheat and oat brans and inulin by human faecal bacteria

The in vitro fermentabilities of rye, wheat and oat brans and of a commercial fibre preparation, inulin, were compared. The brans were first digested enzymatically to remove starch and protein. The digested brans and inulin were then fermented with human faecal inoculum. The progress of fermentation was studied by following the consumption of carbohydrates and the production of short-chain fatty acids and gases. Inulin, a short fructose polymer, was consumed significantly faster than the more complex carbohydrates of cereal brans. Carbohydrates of oat bran (rich in β-glucan) were consumed at a higher rate than those of rye and wheat brans (rich in arabinoxylan). In all brans, glucose was consumed faster than the other main sugars, arabinose and xylose, and arabinose was degraded only slightly. The total production of short-chain fatty acids was slightly higher with oat bran than with rye and wheat brans and inulin. In the fermentation of inulin, relatively more butyric acid and less propionic acid were produced than in the fermentation of brans. The decrease in pH was also greater in the case of inulin. Wheat bran led to a slightly slower gas formation than rye and oat brans. Formation of gases was fastest and greatest in the case of inulin. In conclusion, rye, wheat and oat brans were fermented in a rather similar way. Fermentation of the brans was different from that of inulin. Cereal brans might serve as a more balanced source of dietary fibre supplement than gas-producing, readily fermentable polysaccharides such as inulin. © 2000 Society of Chemical Industry