Effect of Sodium Selenite on the Metabolite Profile of Epichloë sp. Mycelia from Festuca sinensis in Solid Culture.

Selenium (Se) is an essential micronutrient with many beneficial effects for humans and other living organisms. Numerous microorganisms in culture systems enrich and convert inorganic selenium to organic selenium. In this study, Epichloë sp. from Festuca sinensis was exposed to increasing Na2SeO3 concentrations (0, 0.1, 0.2, 0.3, and 0.4 mmol/L) in Petri dishes with potato dextrose agar (PDA) for 8 weeks. Epichloë sp. mycelia were immediately collected after mycelial diameters were measured at 4, 5, 6, 7, and 8 weeks of cultivation, respectively. Gas chromatography-mass spectrometer (GC-MS) analysis was performed on different groups of Epichloë sp. mycelia. Different changes were observed as Epichloë sp. was exposed to different selenite conditions and cultivation time. The colony diameter of Epichloë sp. decreased in response to increased selenite concentrations, whereas the inhibitory effects diminished over time. Seventy-two of the 203 identified metabolites did not differ significantly across selenite treatments within the same time point, while 82 compounds did not differ significantly between multiple time points of the same Se concentration. However, the relative levels of 122 metabolites increased the most under selenite conditions. Specifically, between the 4th and 8th weeks, there were increases in 2-keto-isovaleric acid, uridine, and maltose in selenite treatments compared to controls. Selenium increased glutathione levels and exhibited antioxidant properties in weeks 4, 5, and 7. Additionally, we observed that different doses of selenite could promote the production of carbohydrates such as isomaltose, cellobiose, and sucrose; fatty acids such as palmitoleic acid, palmitic acid, and stearic acid; and amino acids such as lysine and tyrosine in Epichloë sp. mycelia. Therefore, Epichloë sp. exposed to selenite stress may benefit from increased levels of some metabolite compounds.

Fusion of cellobiose phosphorylase and potato alpha-glucan phosphorylase facilitates substrate channeling for enzymatic conversion of cellobiose to starch.

We previously reported an in vitro enzymatic pathway for conversion of nonfood cellulose to starch (PNAS,110 (18): 7182-7187, 2013), in which the two sequential enzymes cellobiose phosphorylase (CBP) from Clostridium thermocellum and potato alpha-glucan phosphorylase (PGP) from Solanum tuberosum were the two key enzymes responsible for the whole conversion rate. In this work CBP and PGP were fused to form a large enzyme and it turned out that the fusion protein could exhibit a good bifunctionality when PGP moiety was put at the N-terminus and CBP moiety at the C-terminus (designated as PGP-CBP). Although the coupled reaction rate of PGP-CBP was decreased by 23.0% compared with the free enzymes, substrate channeling between the two active sites in PGP-CBP was formed, demonstrated by the introduction of the competing enzyme of PGP to the reaction system. The potential of PGP-CBP fusion enzyme being applied to the conversion of cellulose to amylose was discussed.

Active Acupoints Differ from Inactive Acupoints in Modulating Key Plasmatic Metabolites of Hypertension: A Targeted Metabolomics Study.

The effect of active acupoints versus inactive acupoints in treating hypertension is not well documented. Metabolic phenotypes, depicted by metabolomics analysis, reflect the influence of external exposures, nutrition, and lifestyle on the integrated system of the human body. Therefore, we utilized high-performance liquid chromatography tandem mass spectrometry to compare the targeted metabolic phenotype changes induced by two different acupoint treatments. The clinical outcomes show that active acupoint treatment significantly lowers 24-hour systolic blood pressure but not diastolic blood pressure, as compared with inactive acupoint treatment. Furthermore, distinctive changes are observed between the metabolomics data of the two groups. Multivariate analysis shows that only in the active acupoint treatment group can the follow-up plasma be clearly separated from the baseline plasma. Moreover, the follow-up plasma of these two groups can be clearly separated, indicating two different post-treatment metabolic phenotypes. Three metabolites, sucrose, cellobiose, and hypoxanthine, are shown to be the most important features of active acupoint treatment. This study demonstrates that metabolomic analysis is a potential tool that can be used to efficiently differentiate the effect of active acupoints from inactive acupoints in treating hypertension. Possible mechanisms are the alternation of hypothalamic microinflammation and the restoration of host-gut microbiota interactions induced by acupuncture.

Effect of cellobiose supplementation and dietary soluble fibre content on in vitro caecal fermentation of carbohydrate-rich substrates in rabbits.

The in vitro caecal fermentation of five substrates low in starch and protein content [d-(+)-glucose (GLU), d-cellobiose (CEL), sugar beet pectin (PEC), sugar beet pulp (SBP) and wheat straw (WS)] was investigated using soft faeces from rabbits receiving different levels of cellobiose and soluble fibre as inoculum. A total of 24 rabbits were supplemented 3 levels of cellobiose in the drinking water (0.0, 7.5, 15.0 g/l) and fed two experimental diets containing either low soluble fibre (LSF) or high soluble fibre (HSF) levels (84.0 and 130 g/kg dry matter). All substrates were subjected to a two-step pepsin/pancreatin in vitro pre-digestion, and the whole residue was used as substrate for the in vitro incubations. Gas production was measured until 144 h, and volatile fatty acid (VFA) production was determined at 24 h incubation. Experimental treatments did not affect SBP fermentation and had only a subtle influence on fermentation of WS and GLU. In contrast, cellobiose supplementation × donors' diet interactions were detected for most gas production parameters for CEL. Both the fractional gas production (k) and maximal gas production rates were linearly increased (p ≤ 0.042) and the initial delay in the onset of gas production (Lag) linearly decreased (p < 0.001) by cellobiose supplementation with the HSF inoculum, with no differences between the 7.5 and 15.0 doses. In contrast, with the LSF inoculum cellobiose supplementation only affected k values, which were quadratically increased (p = 0.043) and had maximal values for the 7.5 dose. A quadratic effect (p ≤ 0.018) of cellobiose supplementation was observed for total VFA production at 24 h when CEL and PEC were fermented, obtaining the maximal VFA production for the 7.5 dose of cellobiose. Total VFA production for CEL was greater with LSF than with HSF inoculum (20.7 vs. 12.9 mmol/l; p = 0.014), but the opposite was found for WS (3.97 vs. 6.21 mmol/l; p = 0.005). The use of LSF inoculum for CEL fermentation sharply reduced acetate (p = 0.001) and increased butyrate proportions (p ≤ 0.001) compared with the HSF inoculum. A positive relationship between total VFA caecal concentrations in rabbits receiving the same experimental treatments and in vitro values was only observed when WS was used as substrate (r = 0.90; p = 0.015; n = 6). The results suggest that experimental factors influenced the fermentative activity of caecal digesta, but the observed response differed with the incubated substrate, being the CEL the most affected.

The effect of cellobiose on the health status of growing rabbits depends on the dietary level of soluble fiber.

The aim of this study was to examine whether the combination of dietary soluble fiber and cellobiose exerts a synergistic effect on growth performance, health status, fermentation traits, and immune response in rabbits. Six treatments in a 3 × 2 factorial arrangement were used: 3 cellobiose concentrations in drinking water (0.0, 7.5, and 15.0 g/L) × 2 dietary levels of soluble fiber (84.0 and 130 g/kg DM, for the low soluble fiber [LSF] and high soluble fiber [HSF] diets, respectively). A total of 318 young rabbits (53/treatment) were weaned at 34 d of age and had ad libitum access to feed and water. At 46 d of age, 9 rabbits/treatment were slaughtered and ileal and cecal digesta were collected to analyze VFA profile and the immune response in the cecal appendix mucosa. At 48 d of age, the cellobiose supplementation was withdrawn and the experimental diets were replaced by a standard commercial diet until 61 d of age. From 34 to 48 d of age, there was a linear increase of mortality with the level of cellobiose in the HSF group (0% vs. 17.1%; P = 0.017). In contrast, a quadratic effect of cellobiose level on mortality was observed in the LSF group, the rabbits offered 7.5-cellobiose showing the lowest mortality (5.7% vs. 21.4%; P = 0.030). Cellobiose level had a quadratic effect on ADFI, ADG, and G:F in this period (P ≤ 0.047), with the 7.5-cellobiose groups having the best growth performance. In contrast, only minor changes on these traits were observed from 48 d of age onwards. Cellobiose level influenced quadratically the ileal VFA concentrations (P = 0.014), showing the maximal value in the 7.5-cellobiose groups. In rabbits fed 7.5-cellobiose-LSF, a change of acetate to propionate, butyrate, and valerate was observed in the ileum. Increasing cellobiose levels reduced linearly cecal VFA concentrations in HSF fed rabbits, but no effect was detected in LSF groups (P = 0.046). The level of soluble fiber increased VFA concentrations in both the ileum (by 22%; P < 0.001), and the cecum (by 11%; P = 0.005). The relative gene expression of IL-6, IL-10, TNF-α, iNOS, MUC-1, and toll-like receptors (TLR-2 and TLR-4) in the cecal appendix increased linear and quadratically with increasing levels of cellobiose (P ≤ 0.063). In conclusion, in rabbits fed LSF diets, a dose of 7.5 g cellobiose/L drinking water would be recommended, whereas these levels of cellobiose supplementation should be avoided in rabbits fed HSF diets.

Cellulose-Derived Oligomers Act as Damage-Associated Molecular Patterns and Trigger Defense-Like Responses.

The plant cell wall, often the site of initial encounters between plants and their microbial pathogens, is composed of a complex mixture of cellulose, hemicellulose, and pectin polysaccharides as well as proteins. The concept of damage-associated molecular patterns (DAMPs) was proposed to describe plant elicitors like oligogalacturonides (OGs), which can be derived by the breakdown of the pectin homogalacturon by pectinases. OGs act via many of the same signaling steps as pathogen- or microbe-associated molecular patterns (PAMPs) to elicit defenses and provide protection against pathogens. Given both the complexity of the plant cell wall and the fact that many pathogens secrete a wide range of cell wall-degrading enzymes, we reasoned that the breakdown products of other cell wall polymers may be similarly biologically active as elicitors and may help to reinforce the perception of danger by plant cells. Our results indicate that oligomers derived from cellulose are perceived as signal molecules in Arabidopsis (Arabidopsis thaliana), triggering a signaling cascade that shares some similarities to responses to well-known elicitors such as chitooligomers and OGs. However, in contrast to other known PAMPs/DAMPs, cellobiose stimulates neither detectable reactive oxygen species production nor callose deposition. Confirming our idea that both PAMPs and DAMPs are likely to cooccur at infection sites, cotreatments of cellobiose with flg22 or chitooligomers led to synergistic increases in gene expression. Thus, the perception of cellulose-derived oligomers may participate in cell wall integrity surveillance and represents an additional layer of signaling following plant cell wall breakdown during cell wall remodeling or pathogen attack.

Citrobacter freundii as a test platform for recombinant cellulose degradation systems.

Cellulosic biomass represents a huge reservoir of renewable carbon, but converting it into useful products is challenging. Attempts to transfer cellulose degradation capability to industrially useful micro-organisms have met with limited success, possibly due to poorly understood synergy between multiple cellulases. This is best studied by co-expression of many combinations of cellulases and associated proteins. Here, we describe the development of a test platform based on Citrobacter freundii, a cellobiose-assimilating organism closely related to Escherichia coli. Standard E. coli cloning vectors worked well in Cit. freundii. Expression of cellulases CenA and Cex of Cellulomonas fimi in Cit. freundii gave recombinant strains which were able to grow at the expense of cellulosic filter paper or microcrystalline cellulose (Avicel) in a mineral medium supplemented with a small amount of yeast extract. Periodic physical agitation of the cultures was highly beneficial for growth at the expense of filter paper. This provides a test platform for the expression of combinations of genes encoding biomass-degrading enzymes to develop effective genetic cassettes for degradation of different biomass streams. SIGNIFICANCE AND IMPACT OF THE STUDY: Biofuels have been shown to be the best sustainable and alternative source of fuel to replace fossil fuels. Of the different types of feedstocks used for producing biofuels, lignocellulosic biomass is the most abundant. Converting this biomass to useful products has met with little success. Different approaches are being used and microbial platforms are the most promising and sustainable method. This study shows that Citrobacter freundii is a better test platform than Escherichia coli for testing various combinations of cellulases for the development of microbial systems for biomass conversion.

Whole-Cell Biocatalysis for Producing Ginsenoside Rd from Rb1 Using Lactobacillus rhamnosus GG.

Ginsenosides are the major active ingredients in ginseng used for human therapeutic plant medicines. One of the most well-known probiotic bacteria among the various strains on the functional food market is Lactobacillus rhamnosus GG. Biocatalytic methods using probiotic enzymes for producing deglycosylated ginsenosides such as Rd have a growing significance in the functional food industry. The addition of 2% cellobiose (w/v) to glucose-free de Man-Rogosa-Sharpe broths notably induced ß-glucosidase production from L. rhamnosus GG. Enzyme production and activity were optimized at a pH, temperature, and cellobiose concentration of 6.0, 40°C, and 2% (w/v), respectively. Under these controlled conditions, ß-glucosidase production in L. rhamnosus GG was enhanced by 25-fold. Additionally, whole-cell homogenates showed the highest ß-glucosidase activity when compared with disrupted cell suspensions; the cell disruption step significantly decreased the ß-glucosidase activity. Based on the optimized enzyme conditions, whole-cell L. rhamnosus GG was successfully used to convert ginsenoside Rb1 into Rd.

[Effect of D-cellobiose on oral bioavailability of gentiopicroside].

In this study, the effect of D-cellobiose on oral bioavailability of gentiopicroside (GPS) was investigate. The influence of D-cellobiose on GPS was achieved by calculating the residual GPS after being degraded with ß-glucosidase or intestinal flora, and the data demonstrated D-cellobiose could inhibit the degradation of GPS in intestines; in bioavailability experiment, D-cellobiose could significantly improve the oral bioavailability (P<0.05) of GPS at the mass ratio of 1∶5, 1∶10 (GPS-D-cellobiose). D-cellobiose applied in this study may improve the oral bioavailability of GPS through delaying the degradation in intestines.

Overexpression and characterization of a glucose-tolerant ß-glucosidase from T. aotearoense with high specific activity for cellobiose.

Thermoanaerobacterium aotearoense P8G3#4 produced ß-glucosidase (BGL) intracellularly when grown in liquid culture on cellobiose. The gene bgl, encoding ß-glucosidase, was cloned and sequenced. Analysis revealed that the bgl contained an open reading frame of 1314 bp encoding a protein of 446 amino acid residues, and the product belonged to the glycoside hydrolase family 1 with the canonical glycoside hydrolase family 1 (GH1) (ß/α)8 TIM barrel fold. Expression of pET-bgl together with a chaperone gene cloned in vector pGro7 in Escherichia coli dramatically enhanced the crude enzyme activity to a specific activity of 256.3 U/mg wet cells, which resulted in a 9.2-fold increase of that obtained from the expression without any chaperones. The purified BGL exhibited relatively high thermostability and pH stability with its highest activity at 60 °C and pH 6.0. In addition, the activities of BGL were remarkably stimulated by the addition of 5 mM Na(+) or K(+). The enzyme showed strong ability to hydrolyze cellobiose with a K m and V max of 25.45 mM and 740.5 U/mg, respectively. The BGL was activated by glucose at concentration varying from 50 to 250 mM and tolerant to glucose inhibition with a K i of 800 mM glucose. The supplement of the purified BGL to the sugarcane bagasse hydrolysis mixture containing a commercial cellulase resulted in about 20 % enhancement of the released reducing sugars. These properties of the purified BGL should have important practical implication in its potential applications for better industrial production of glucose or bioethanol started from lignocellulosic biomass.

Enhancement of beta-Glucosidase Activity from a Brown Rot Fungus Fomitopsis pinicola KCTC 6208 by Medium Optimization

beta-Glucosidase, which hydrolyzes cellobiose into two glucoses, plays an important role in the process of saccharification of the lignocellulosic biomass. In this study, we optimized the activity of beta-glucosidase of brown-rot fungus Fomitopsis pinicola KCTC 6208 using the response surface methodology (RSM) with various concentrations of glucose, yeast extract and ascorbic acid, which are the most significant nutrients for activity of beta-glucosidase. The highest activity of beta-glucosidase was achieved 3.02% of glucose, 4.35% of yeast extract, and 7.41% ascorbic acid where ascorbic acid was most effective. The maximum activity of beta-glucosidase predicted by the RSM was 15.34 U/mg, which was similar to the experimental value 14.90 U/mg at the 16th day of incubation. This optimized activity of beta-glucosidase was 23.6 times higher than the preliminary activity value, 0.63 U/mg, and was also much higher than previous values reported in other fungi strains. Therefore, a simplified medium supplemented with a cheap vitamin source, such as ascorbic acid, could be a cost effective mean of increasing beta-glucosidase activity.

Stability of amorphous silica-alumina in hot liquid water.

Herein, the hydrothermal stability of amorphous silica-alumina (ASA) is investigated under conditions relevant for the catalytic conversion of biomass, namely in liquid water at 200 °C. The hydrothermal stability of ASA is much higher than that of pure silica or alumina. Interestingly, the synthetic procedure used plays a major role in its resultant stability: ASA prepared by cogelation (CG) lost its microporous structure, owing to hydrolysis of the siloxane bonds, but the resulting mesoporous material still had a considerable surface area. ASA prepared by deposition precipitation (DP) contained a silicon-rich core and an aluminum-rich shell. In hot liquid water, the latter structure was transformed into a layer of amorphous boehmite, which protected the particle from further hydrolysis. The surface area showed relatively minor changes during the transformation. Independent of the synthetic method used, the ASAs retained a considerable concentration of acid sites. The concentration of acid sites qualitatively followed the changes in surface area, but the changes were less pronounced. The performance of different ASAs for the hydrolysis of cellobiose into glucose is compared.

Genomic, proteomic, and biochemical analyses of oleaginous Mucor circinelloides: evaluating its capability in utilizing cellulolytic substrates for lipid production.

Lipid production by oleaginous microorganisms is a promising route to produce raw material for the production of biodiesel. However, most of these organisms must be grown on sugars and agro-industrial wastes because they cannot directly utilize lignocellulosic substrates. We report the first comprehensive investigation of Mucor circinelloides, one of a few oleaginous fungi for which genome sequences are available, for its potential to assimilate cellulose and produce lipids. Our genomic analysis revealed the existence of genes encoding 13 endoglucanases (7 of them secretory), 3 ß-D-glucosidases (2 of them secretory) and 243 other glycoside hydrolase (GH) proteins, but not genes for exoglucanases such as cellobiohydrolases (CBH) that are required for breakdown of cellulose to cellobiose. Analysis of the major PAGE gel bands of secretome proteins confirmed expression of two secretory endoglucanases and one ß-D-glucosidase, along with a set of accessory cell wall-degrading enzymes and 11 proteins of unknown function. We found that M. circinelloides can grow on CMC (carboxymethyl cellulose) and cellobiose, confirming the enzymatic activities of endoglucanases and ß-D-glucosidases, respectively. The data suggested that M. circinelloides could be made usable as a consolidated bioprocessing (CBP) strain by introducing a CBH (e.g. CBHI) into the microorganism. This proposal was validated by our demonstration that M. circinelloides growing on Avicel supplemented with CBHI produced about 33% of the lipid that was generated in glucose medium. Furthermore, fatty acid methyl ester (FAME) analysis showed that when growing on pre-saccharified Avicel substrates, it produced a higher proportion of C14 fatty acids, which has an interesting implication in that shorter fatty acid chains have characteristics that are ideal for use in jet fuel. This substrate-specific shift in FAME profile warrants further investigation.

Exoproteome profiles of Clostridium cellulovorans grown on various carbon sources.

The cellulosome is a complex of cellulosomal proteins bound to scaffolding proteins. This complex is considered the most efficient system for cellulose degradation. Clostridium cellulovorans, which is known to produce cellulosomes, changes the composition of its cellulosomes depending on the growth substrates. However, studies have investigated only cellulosomal proteins; profile changes in noncellulosomal proteins have rarely been examined. In this study, we performed a quantitative proteome analysis of the whole exoproteome of C. cellulovorans, including cellulosomal and noncellulosomal proteins, to illustrate how various substrates are efficiently degraded. C. cellulovorans was cultured with cellobiose, xylan, pectin, or phosphoric acid-swollen cellulose (PASC) as the sole carbon source. PASC was used as a cellulose substrate for more accurate quantitative analysis. Using an isobaric tag method and a liquid chromatography mass spectrometer equipped with a long monolithic silica capillary column, 639 proteins were identified and quantified in all 4 samples. Among these, 79 proteins were involved in saccharification, including 35 cellulosomal and 44 noncellulosomal proteins. We compared protein abundance by spectral count and found that cellulosomal proteins were more abundant than noncellulosomal proteins. Next, we focused on the fold change of the proteins depending on the growth substrates. Drastic changes were observed mainly among the noncellulosomal proteins. These results indicate that cellulosomal proteins were primarily produced to efficiently degrade any substrate and that noncellulosomal proteins were specifically produced to optimize the degradation of a particular substrate. This study highlights the importance of noncellulosomal proteins as well as cellulosomes for the efficient degradation of various substrates.

Purification and physicokinetic characterization of a gluconolactone inhibition-insensitive ß-glucosidase from Andrographis paniculata nees. Leaf.

A gluconolactone inhibition-insensitive ß-glucosidase from Andrographis paniculata (Acanthaceae) leaves has been isolated, homogeneity purified, and characterized for its physicokinetic properties. The purified enzyme appeared to be a monomeric structure with native molecular weight about 60 kD. The enzyme exhibited optimum pH 5.5 and pI 4.0, meso-thermostability and high temperature optimum (55°C) for catalytic activity, with activation energy of 6.8 kcal Mol(-1). The substrate saturation kinetics studies of the enzyme revealed a Michaelis-Menten constant (Km) of 0.25 mM for pNPG and catalytic efficiency (Kcat/Km) of 52,400 M (-1) s(-1), respectively. Substrate specificity of the enzyme was restricted to ß-linked gluco-, manno- and fuco-conjugates. The gluconolactone inhibition insensitivity was evident from its very low inhibition at millimolar inhibitor concentrations. Interestingly, the enzyme showed geraniol transglucosylating activity with pNPG as glucosyl donor but not with cellobiose. The catalytic activity of the enzyme has been reported to be novel with respect to its activity and preferences from a medicinal plant resource.

Nitrogen and sulfur requirements for Clostridium thermocellum and Caldicellulosiruptor bescii on cellulosic substrates in minimal nutrient media.

Growth media for cellulolytic Clostridium thermocellum ATCC 27405 and Caldicellulosiruptor bescii bacteria usually contain excess nutrients that would increase costs for consolidated bioprocessing for biofuel production and create a waste stream with nitrogen, sulfur and phosphate. C. thermocellum was grown on crystalline cellulose with varying concentrations of nitrogen and sulfur compounds, and growth rate and ethanol production response curves were determined. Both bacteria assimilated sulfate in the presence of ascorbate reductant, increasing the ratio of oxidized to reduced fermentation products. From these results, a low ionic strength, defined minimal nutrient medium with decreased nitrogen, sulfur, phosphate and vitamin supplements was developed for the fermentation of cellobiose, cellulose and acid-pretreated Populus. Carbon and electron balance calculations indicate the unidentified residual fermentation products must include highly reduced molecules. Both bacterial populations were maintained in co-cultures with substrates containing cellulose and xylan in defined medium with sulfate and basal vitamin supplements.

Impairment of cellulose- and cellobiose-degrading soil Bacteria by two acidic herbicides.

Herbicides have the potential to impair the metabolism of soil microorganisms. The current study addressed the toxic effect of bentazon and 4-chloro-2-methylphenoxyacetic acid on aerobic and anaerobic Bacteria that are involved in cellulose and cellobiose degradation in an agricultural soil. Aerobic saccharide degradation was reduced at concentrations of herbicides above environmental values. Microbial processes (e.g. fermentations, ferric iron reduction) that were linked to anaerobic cellulose and cellobiose degradation were reduced in the presence of both herbicides at concentrations above and at those that occur in crop field soil. 16S rRNA gene transcript numbers of total Bacteria, and selected bacterial taxa (Clostridia [Group I], Planctomycetaceae, and two uncultivated taxa of Bacteroidetes) decreased more in anoxic than in oxic cellulose-supplemented soil microcosms in the presence of both herbicides. Collectively, the results suggested that the metabolism of anaerobic cellulose-degrading Bacteria was impaired by typical in situ herbicide concentrations, whereas in situ concentrations did not impair metabolism of aerobic cellulose- and cellobiose-degrading soil Bacteria.

Mariannaea samuelsii Isolated from a Bark Beetle-Infested Elm Tree in Korea

During an investigation of fungi from an elm tree infested with bark beetles in Korea, one isolate, DUCC401, was isolated from elm wood. Based on morphological characteristics and phylogenetic analysis of the internal transcribed spacer and 28S rDNA (large subunit) sequences, the isolate, DUCC401, was identified as Mariannaea samuelsii. Mycelia of the fungus grew faster on malt extract agar than on potato dextrose agar and oatmeal agar media. Temperature and pH for optimal growth of fungal mycelia were 25degrees C and pH 7.0, respectively. The fungus demonstrated the capacity to degrade cellobiose, starch, and xylan. This is the first report on isolation of Mariannaea samuelsii in Korea.

Enzymatic hydrolysis and simultaneous saccharification and fermentation of alkali/peracetic acid-pretreated sugarcane bagasse for ethanol and 2,3-butanediol production.

The enzymatic digestibility of alkali/peracetic acid (PAA)-pretreated bagasse was systematically investigated. The effects of initial solid consistency, cellulase loading and addition of supplemental ß-glucosidase on the enzymatic conversion of glycan were studied. It was found the alkali-PAA pulp showed excellent enzymatic digestibility. The enzymatic glycan conversion could reach about 80% after 24 h incubation when enzyme loading was 10 FPU/g solid. Simultaneous saccharification and fermentation (SSF) results indicated that the pulp could be well converted to ethanol. Compared with dilute acid pretreated bagasse (DAPB), alkali-PAA pulp could obtain much higher ethanol and xylose concentrations. The fermentation broth still showed some cellulase activity so that the fed pulp could be further converted to sugars and ethanol. After the second batch SSF, the fermentation broth of alkali-PAA pulp still kept about 50% of initial cellulase activity. However, only 21% of initial cellulase activity was kept in the fermentation broth of DAPB. The xylose syrup obtained in SSF of alkali-PAA pulp could be well converted to 2,3-butanediol by Klebsiella pneumoniae CGMCC 1.9131.

Supplementation of the diet of dairy cows with trehalose results in milk with low lipid peroxide and high antioxidant content.

The objective of this study was to investigate the effect of dietary supplementation with the disaccharides trehalose and cellobiose on antioxidant activity in rumen fluid, blood, and milk of dairy cows. Nine Holstein dairy cows housed in a free-stall barn were divided into 3 groups, with each group receiving a different dietary treatment (a control diet, a 1% trehalose-supplemented diet, or a 1% cellobiose-supplemented diet) following a 3x3 Latin square design. Feed intake and milk production increased in cows receiving the trehalose-supplemented diet compared with those receiving the control and cellobiose-supplemented diets. The total protozoa numbers in the rumen fluid of cows fed trehalose- or cellobiose-supplemented diets were greater than those of the control group. The C18:0 and C18:1 fatty acid content was increased in the milk of cows fed the trehalose-supplemented diet compared with that of the control group, and the C18:3n-3 fatty acid content in the milk of cows fed the cellobiose-supplemented diet was less than that of the control group. Plasma biochemical parameters were unchanged among the different treatments. In rumen fluid, 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity and superoxide dismutase activity were increased 2h after feeding in cows receiving the cellobiose-supplemented diet compared with the control group, and the concentration of thiobarbituric acid reactive substances in the rumen fluid of cows fed the cellobiose-supplemented diet was decreased. In contrast, the values of these parameters measured in the milk of cows fed the cellobiose-supplemented diet were no different from those of control cows. Dietary supplementation with trehalose did, however, bring about an improvement of the oxidative status of milk and blood in these animals compared with controls. These results provide the first evidence supporting the use of dietary disaccharides to decrease lipid peroxide levels and increase the antioxidant content of dairy cow milk. The findings suggest that disaccharides, particularly trehalose, might be useful as supplements for reducing oxidative stress and improving the quality of milk for human consumption, as well as possibly impairing the processes that give rise to lipid oxidation odor in dairy cow milk.