Application of microalgae hydrolysate as a fermentation medium for microbial production of 2-pyrone 4,6-dicarboxylic acid

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Chlorine isotope fractionation during reductive dechlorination of chlorinated ethenes by anaerobic bacteria

Chlorine isotope fractionation during reductive dechlorination of trichloroethene (TCE) and tetrachloroethene (PCE) to cis-1,2-dichloroethene (cDCE) by anaerobic bacteria was investigated. The changes in the 37Cl/35Cl ratio observed during the one-step reaction (TCE to cDCE) can be explained by the regioselective elimination of chlorine accompanied by the Rayleigh fractionation. The fractionation factors (alpha) of the TCE dechlorination by three kinds of anaerobic cultures were approximately 0.994-0.995 at 30 degrees C. The enrichment of 37Cl in the organic chlorine during the two-step reaction (PCE to cDCE) can be explained by the random elimination of one chlorine atom in the PCE molecule followed by the regioselective elimination of one chlorine atom in the TCE molecule. The fractionation factors for the first step of the PCE dechlorination with three kinds of anaerobic cultures were estimated to be 0.987-0.991 at 30 degrees C using a mathematical model. Isotope fractionation during the first step would be the primary factor for the chlorine isotope fractionation during the PCE dechorination to cDCE. The developed models can be utilized to evaluate the fractionation factors of regioselective and multistep reactions.     

Novel poly-gamma-glutamate-processing enzyme catalyzing gamma-glutamyl DD-amidohydrolysis

The pgdS gene product of Bacillus subtilis, PgdS, cleaves poly-gamma-glutamate (PGA) in an endo-peptidase-like fashion. However, its catalytic property remains obscure. In this study, a simple assay for the PgdS enzyme using 1-fluoro-2,4-dinitrobenzene was developed, and some characteristics of PgdS, such as optimal pH, were examined. The enzyme was strongly inhibited by a thiol-modifying reagent, suggesting that it possesses essential cysteine residue(s) in catalysis. PgdS exhibited a high affinity to PGA that consisted mainly of D-glutamate residues, but no affinity to PGA composed only of L-glutamate residues (L-PGA). The enzyme processed DL-copolymer-type PGA (DL-PGA) with an average molecular mass of 1,000 kDa to a high-molecular-mass L-glutamate-rich fragment (average 200 kDa), the L-rich PGA fragment, and low-molecular-mass fragment composed mostly of D-glutamate residues (average 5 kDa), D-fragment. To deepen our understanding of the catalytic property of the PgdS enzyme, we analyzed the structures of the N- and C-terminal regions and found that D-glutamyl residues successively lie even at both ends of the L-rich PGA fragment. Our observations indicate that PgdS is a novel endo-peptidase that specifically cleaves the gamma-amide linkage between two D-glutamate residues in PGA, i.e., gamma-glutamyl DD-amidohydrolase. The enzyme is possibly useful in the biochemical processing of B. subtilis DL-PGA.     

Lacticaseibacillus paracasei K71 Alleviates UVB-Induced Skin Barrier Dysfunction by Attenuating Inflammation via Increased IL-10 Production in Mice

Scope

Ultraviolet B (UVB) radiation causes skin barrier dysfunction, leading to decreased water-holding capacity, impaired epidermal barrier function, and increased skin thickness. This study investigates the protective effects of oral administration of Lacticaseibacillus paracasei K71 against skin barrier dysfunction in UVB-irradiated mice.

Methods and results

Mice are fed diets with or without K71 and irradiated with UVB three times a week for 12 weeks. Oral administration of K71 suppresses UVB-induced decrease in stratum corneum water content, mitigates the increase of transepidermal water loss, and decreases epidermal thickness of the dorsal skin. Treatment with K71 reverses the upregulation of inflammatory cytokines and the activation of nuclear factor-κB induced by UVB irradiation and upregulates the expression of anti-inflammatory IL-10 in the dorsal skin. Notable upregulation of IL-10 is observed in the spleens of K71-treated mice. K71 treatment enhances IL-10 production in J774.1 macrophages; however, this enhancement is diminished by inhibiting K71 phagocytosis and TLR3. Furthermore, transfection using K71 RNAs significantly increases IL-10 production.

Conclusion

These results indicate that K71 may alleviate UVB-induced skin barrier dysfunction by attenuating inflammation via increasing IL-10 production and that K71 RNAs may induce IL-10 production in macrophages. Therefore, K71 may be beneficial for preventing skin barrier dysfunction.     

Potential of fermentation byproducts as nitrogen supplements for lactating dairy cows

Two feeding trials evaluated several byproducts from commercial amino acid fermentations as N supplements for lactating cows. Trial 1 was a replicated 5 x 5 Latin square that used 2-wk periods and 25 Holstein cows (five with ruminal cannulae) fed diets containing [dry matter (DM) basis] 28% alfalfa silage, 31% corn silage, 28% high moisture ear corn plus 4 percentage units of crude protein (CP) from: soybean meal, urea, commercial fermentation byproduct 1 or 2, or a blend of fermentation byproducts plus wheat middlings. Diets averaged 15.1% CP and 32% neutral detergent fiber. Intake of DM, body weight (BW) gain, and yield of milk and milk components were greatest for cows fed soybean meal; animal performance was similar with urea, byproduct 1 and the byproduct blend. Intake, BW change, and yield of milk and protein when cows were fed byproduct 2 were lower than when fed urea. Urine output (estimated with creatinine in spot urine samples) was greater on fermentation byproduct 1 and the byproduct blend. There were no differences due to N source in microbial synthesis (based on estimated purine derivative excretion), in situ digestion of alfalfa hay DM, or molar proportions of ruminal volatile fatty acids. Trial 2 was a replicated 5 x 5 Latin square using 2-wk periods and 10 Holstein cows fed diets containing (DM basis) 37% alfalfa silage, 28% corn silage, 29% high moisture ear corn plus 2 percentage units of CP from urea, fermentation byproduct 1, or one of three blends of fermentation byproducts plus wheat middlings. Except for greater DM intake in cows fed the byproduct blends, performance and urinary metabolite excretion did not differ because of N supplement. Relative to other fermentation byproducts and urea, byproduct 1 resulted in reduced milk urea N in both trials. Under the conditions of these trials, fermentation byproducts were less effective than soybean meal, and no more effective than urea, as N supplements.     

Isolation, characterization and identification of polyhydroxyalkanoate-accumulating bacteria from activated sludge

Two novel gram-positive bacteria capable of accumulating poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [poly(3HB-co-3HV)] were isolated from an anaerobic-oxic activated sludge system fed with acetate. Strains Lpha5 and Lpha7 are motile cocci, 1-2 microm in diameter, occurring singly or in pairs. These isolates have doubling times ranging from 0.4-1.7 d and can accumulate in high levels of poly(3HB-co-3HV) (up to 44.7% of cell dry weight) when grown on complex media. Furthermore, these two strains exhibited the rapid substrate uptake and accumulation of storage granules as observed in situ. Under aerobic conditions, about 14.4% (cell dry weight) polyhydroxyalkanoate and 82% (carbon dry weight) cellular carbohydrate were produced from acetate and glucose, respectively. Under anaerobic conditions, poly(3HB-co-3HV) and cellular carbohydrate accumulated when glucose but not acetate was fed. The result of analysis of 16S rRNA sequence revealed that both strains belong to the gram-positive high-G + C group, but are significantly different from their closest phylogenetic relatives, Dermatophilus sp. and Terrabacter sp., to warrant classification as a new species.     

Contamination levels of PCDDs, PCDFs and Co-PCBs in commercial baby foods in Japan

To examine dioxin contamination in commercial baby foods in Japan, congener analyses of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and coplanar polychlorinated biphenyls (Co-PCBs) were performed on 102 varieties of baby foods obtained from supermarkets in 2001-2002. The toxic equivalent quantity (TEQ) levels for dioxins in samples ranged from < 0.001 to 0.135 pg-TEQ/g wet weight when undetected or trace levels of congeners were taken as zero. Among 102 samples tested, 26 samples exceeded 0.010 pg-TEQ/g. The highest TEQ value was for "sardine, vegetables" (0.135 pg-TEQ/g), followed by "Japanese radish (daikon), sardine" (0.080 pg-TEQ/g). Thus, dioxins were detected at low levels in baby foods containing animal products such as fishes and/or dairy products.     

Enhanced antiviral activity of soybean β-conglycinin-derived peptides by acylation with saturated fatty acids

Abstract: Peptide mixtures prepared from soybean β‐conglycinin (7S‐peptides) were acylated with saturated fatty acids of different chain length (6C‐18C) in order to improve their antiviral activity against Feline calicivirus (FCV) strain F9 which is a typical norovirus surrogate. Among the fatty acids varieties, it was revealed that 7S‐peptides acylated with myristic and palmitic acids potently inhibited FCV replication. Myristorylation and palmitoylation of 7S‐peptides kept host cells viability at 91.51% and 98.90%, respectively. The infectivity of FCV on Crandell–Reese feline kidney cells was further determined after exposure of initial titer of 10^6.47 TCID₅₀/mL. Myristoylated and palmitoylated 7S‐peptides significantly (P < 0.006) reduced FCV infectivity as compared to native 7S‐peptides. Native 7S‐peptides showed 25% FCV inhibitory activity while myristoylated and palmitoylated 7S‐peptides exhibited 98.59% and 99.98% reduction in FCV infectivity, respectively. Myristoylated and palmitoylated 7S‐peptides demonstrated higher anti‐FCV activity in a wide range of concentration with complete reduction at 25 μg/mL. Surface hydrophobicity was significantly (P < 0.05) increased after attachment of long hydrocarbon fatty acids to 7S‐peptides as supported by changes in fluorescence intensity. Enzymatic hydrolysis together with acylation will give an insight into surface and physiological functional lipopeptides derived from soy β‐conglycinin.     

Multiresidue method for pesticides and veterinary drugs in bovine milk using GC/MS and LC/MS/MS

A simple, sensitive and selective method with gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/tandem mass spectrometry (LC/MS/MS) has been developed to detect 342 pesticides and veterinary drugs contaminating bovine milk at the maximum residue limits (MRLs) defined in the "positive list system". Sample preparation was performed by extracting the analytes with acetonitrile, followed by salting-out with sodium chloride. For some pesticides, the extract was further cleaned up by n-hexane partitioning and PSA cartridge column chromatography. GC/MS-EI or -NCI was used to determine pesticide residues, while LC/MS/MS-ESI was applicable to the determination of pesticide and veterinary drug residues. The variation of the recoveries of these drugs at MRL was relatively wide; however the relative standard deviations of the recovery of each drug were within 28%, suggesting that the present method is good enough for use as a screening test for contaminants at the MRLs. These results show that this method is useful for multiresidue analysis of numerous pesticides and veterinary drugs in bovine milk.