• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.6
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• pp.482-493
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• 2005

Biomass is originally photosynthesized from inorgainic compounds such as $CO_2$, minerals, water and solar energy. Recent studies have shown that anaerobic bacteria have the ability to convert recalcitrant biomass such as cellullosic or chitinoic materials to useful compounds. The biomass containing agricultural waste, unutilized wood and other garbage is expected to utilize as feed, food and fuel by microbial degradation and other metabolic functions. In this study we isolated several anaerobic, cellulolytic and chitinolytic bacteria from rumen fluid, compost and soil to study their related enzymes and genes. The anaerobic and cellulolytic bacteria, Clostridium thermocellum, Clostridium stercorarium, and Clostridium josui, were isolated from compost and the chitinolytic Clostridium paraputrificum from beach soil and Ruminococcus albus was isolated from cow rumen. After isolation, novel cellulase and xylanase genes from these anaerobes were cloned and expressed in Escherichia coli. The properties of the cloned enzymes showed that some of them were the components of the enzyme (cellulase) complex, i.e., cellulosome, which is known to form complexes by binding cohesin domains on the cellulase integrating protein (Cip: or core protein) and dockerin domains on the enzymes. Several dockerin and cohesin polypeptides were independently produced by E. coli and their binding properties were specified with BIAcore by measuring surface plasmon resonance. Three pairs of cohesin-dockerin with differing binding specificities were selected. Two of their genes encoding their respective cohesin polypeptides were combined to one gene and expressed in E. coli as a chimeric core protein, on which two dockerin-dehydrogenase chimeras, the dockerin-formaldehyde dehydrogenase and the dockerin-NADH dehydrogenase are planning to bind for catalyzing $CO_2$ reduction to formic acid by feeding NADH. This reaction may represent a novel strategy for the reduction of the green house gases. Enzymes from the anaerobes were also expressed in tobacco and rice plants. The activity of a xylanase from C. stercorarium was detected in leaves, stems, and rice grain under the control of CaMV35S promoter. The digestibility of transgenic rice leaves in goat rumen was slightly accelerated. C. paraputrificum was found to solubilize shrimp shells and chitin to generate hydrogen gas. Hydrogen productivity (1.7 mol $H_2/mol$ glucos) of the organism was improved up to 1.8 times by additional expression of the own hydrogenase gene in C. paraputrficum using a modified vector of Clostridiu, perfringens. The hydrygen producing microflora from soil, garbage and dried pelletted garbage, known as refuse derived fuel(RDF), were also found to be effective in converting biomass waste to hydrogen gas.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.1
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• pp.63-70
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• 2018

Objective: The aim of the study was to isolate gossypol-degrading bacteria and to assess its potential for gossypol degradation. Methods: Rumen liquid was collected from fistulated cows grazing the experimental pasture. Approximately 1 mL of the rumen liquid was spread onto basal medium plates containing 2 g/L gossypol as the only source of carbon and was then cultured at $39^{\circ}C$ to isolate gossypol-degrading bacteria. The isolated colonies were cultured for 6 h and then their size and shape observed by microscope and scanning electron microscope. The 16S rRNA gene of isolated colonies was sequenced and aligned using National Center for Biotechnology Information-Basic Local Alignment Search Tool. The various fermentation conditions, initial pH, incubation temperature, inoculum level and fermentationperiod were analyzed in cottonseed meal (CSM). The crude protein (CP), total gossypol (TG), and free gossypol (FG) were determined in CSM after fermentation with isolated strain at $39^{\circ}C$ for 72 h. Results: Screening results showed that a single bacterial isolate, named Rumen Bacillus Subtilis (RBS), could use gossypol as a carbon source. The bacterium was identified by 16S rDNA sequencing as being 98% homologous to the sequence of Bacillus subtilis strain GH38. The optimum fermentation conditions were found to be 72 h, $39^{\circ}C$, pH 6.5, moisture 50%, inoculum level $10^7cell/g$. In the optimum fermentation conditions, the FG and TG content in fermented CSM decreased 78.86% and 49% relative to the control. The content of CP and the essential amino acids of the fermented CSM increased respectively, compared with the control. Conclusion: The isolation of a gossypol-degrading bacterium from the cow rumen is of great importance for gossypol biodegradation and may be a valuable potential source for gossypol-degradation of CSM.

• The Journal of Internal Korean Medicine
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• v.31 no.2
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• pp.224-241
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• 2010

Objectives : The aim of the current study was to investigate the effects of Sargassum (Sargassum pallidum (TURN.) C. AG.; SP) on the experimental colitis induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS) in mice. Methods : ICR mice were divided into 7 groups (NOR, CON, $SS50\times5$, $SP20\times3$, $SP50\times3$, $SP20\times5$, $SP50\times5$). TNBS processing was intrarectally applied to all experimental groups on the 3rd experiment day, except the normal group (NOR). For investigating the prophylactic effect, SP at doses of 20 mg/kg ($SP20\times5$) and 50 mg/kg ($SP50\times5$) were orally administered for 5 days. The SP at doses of 20 mg/kg ($SP20\times3$) and 50 mg/kg ($SP50\times3$) were orally administered for 3 days after the colitis induction in order to check the effect of treatment. As a positive control group, sulfasalazine 50 mg/kg ($SS50\times5$) was administrated. Macroscopic findings of epithelial tissue on mice were measured by colon length and macroscopic score. Histologic findings were also checked by crypt cell, epithelial cell, inflammatory cell and edema of submucosa. We measured the ability of SP to inhibit lipid peroxidation and myeloperoxidase activity. We also measured levels of the inflammatory markers, interleukin (IL)-$1\beta$ and cyclooxygenase-2 (COX-2), its transcription factor activation, phospho-NF-${\kappa}B$ (pp65), in the colon by enzyme-linked immunosorbent assay and immunoblot analysis. We measured activation of fecal bacterial enzyme, $\beta$-glucuronidase and degradation activation of fecal glycosaminoglycan (GAG), and hyaluronic acid. Results : Oral administration of SP on mice inhibited TNBS-induced colon shortening and myeloperoxidase activity in the colon of mice as well as IL-$1\beta$ and COX-2 expression. SP also inhibited TNBS-induced lipid peroxidation and pp65 activation in the colon of mice. SP inhibited $\beta$-glucuronidase activation and fecal hyaluronic acid degradation activation as well. Conclusions : SP could be a possible herbal candidate and preventive prebiotic agent for treating inflammatory bowel disease (IBD). Further experiments to differentiate effects of SP on IBD, such as other solutions and extracting times, might be promising.

• Journal of the Korean Society of Food Science and Nutrition
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• v.46 no.7
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• pp.857-867
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• 2017

This study was performed to encapsulate low molecular weight marine collagen and ${\gamma}$-aminobutyric acid (GABA)-producing lactic acid bacteria to inhibit degradation and improve survival rate during exposure to adverse conditions of the gastro-intestinal tract. Calcium-alginate method was used for the manufacture of a double-layered coated capsule. The inner core material was composed of collagen and lactic acid bacteria, and the coating materials were alginate and chitosan. The sizes and shapes of the double-coated capsule were affected mainly by centrifuge speed and pH. Manufactured capsules were observed with a scanning electron microscope and by confocal laser scanning microscopy to confirm the micromorphological changes of capsules and bacterial cells. As a result, double-layered coated capsules were not degraded at pH 1.2, whereas degradation occurred at pH 7.4. In addition, GABA and collagen were maintained in stable state at pH 1.2. Therefore, double-layered coated capsules developed in this study would not be degraded in the stomach and could be stably delivered to the small intestine to benefit intestinal and dermatic health.

• Journal of the Korean Society for Marine Environment & Energy
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• v.8 no.4
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• pp.179-185
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• 2005

Polycyclic aromatic hydrocarbons (PAHs) are a kind of toxic environmental pollutants and has been accumulated usually in marine sediments. Due to their potential hazardous to human, removal of PAHs from environments has been great concern. In the present study, the effect of microbial inoculation and the supplementation of mixed form cyclodextrin (M-CD) was assessed in the pre-sterilized or nonsterilized microcosms for optimizing operational conditions for ex situ bioremediation of sediments contaminated by PAHs. Activity of electron transport system (ETSA) was increased by the addition of M-CD regardless of inoculation of microorganisms in microcosms without sterilization. The degradation rate of PAHs in sterilized microcosms was app. 9-20% by the inoculation of single strain and 24-37% by the inoculation of microbial consortium supplemented with 1% M-CD, respectively. The degradation was not observed in microcosms without sterilization under the same conditions. The proportion of inoculated microorganisms also decreased in nonsterilized microcosms. Signals of inoculated bacteria were decreased to detection limit after 2 days in the microcosms without M-CD. In conclusion, microbial inoculation with appropriate carbon sources and removal of natural flora and grazers are required for the efficient ex situ bioremediation of sediments contaminated by PAHs in bioslurry reactor.

• Journal of The Korean Society of Grassland and Forage Science
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• v.42 no.4
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• pp.229-236
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• 2022

This study was conducted to evaluate the efficacy of adding the microbial inoculants to silage for reducing mycotoxins in rice straw silage. When a single agent of L. plantarum and a mixed agent of L. plantarum and S. cerevisiae were added in rice straw silage contaminated mycotoxins, it had an effect on silage fermentation and fiber degradation as well as mycotoxin reduction. Among the mycotoxins, only ochratoxin A and zearalenone were found in the test sample. Ochratoxin A and zearalenone showed a decreasing trend with the addition of silage inoculants compared to the control groups (38.11±2.22 and 633.67±50.30 ㎍/kg), and there was a significant difference at the mixed agents; 27.78±2.28 and 392.72±25.04 ㎍/kg, respectively (p<0.05). The pH was lower in the single agent and the mixed agent compared to the control (p<0.05). The concentration of lactic acid was higher in the single agent (11.73±0.31 mM) than in the control group (8.18±0.93 mM), and the highest concentration was 16.01±0.88 mM in the mixed agent (p<0.05). Acetic acid and propionic acid were found to be significantly lowered with the addition of silage inoculants (p<0.05). Total VFA was also lower at the addition of silage inoculants than the control group (p<0.05). The rumen in situ dry matter degradation of NDF and ADF was maintained at the highest levels of the mixed agent during the culture period, followed by the single agent and the control group at the lowest level. NDF and ADF degradation showed a significant difference at all time points after 12 and 24 hours of culture, respectively (p<0.05). The study results showed that the silage inoculants had the positive effects on quality increasing of rice straw silage; fermentative charateristics, fiber degradation and mycotoxins reduction. Ochratoxin A and zearalenone were greater reduction by adding bacterial inoculants of silage. Therefore it is considered that L. plantarum and S. cerevisiae will improve the quality and stability with remediation of mycotoxin in silage.

• Journal of Korean Society on Water Environment
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• v.16 no.4
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• pp.541-554
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• 2000

Experimental study of anaerobic digestion process combined with and without the submerged separation membrane was investigated by using laboratory-scale reactor at the hydraulic retention time of 0.5 day and 1.0 day. The removal efficiencies of carbohydrate at the digester without and with membrane were 84.4 to 86.8 % and 99.6 to 99.7 %, respectively, and the methane gas recovery efficiencies were 0.4 to 1.2 % and 12.3 to 28.7 %. According to the measurement by the most probable numbers method. the numbers of various groups of bacteria in the digesters with membrane were enumerated in the following ranges ; acidogens : $1.7{\times}10^9$ to $5.0{\times}10^9MPN/m{\ell}$, homoacetogens : $5.0{\times}10^7$ to $2.4{\times}10^8MPN/m{\ell}$, $H_2$-utilizing methanogens : $1.3{\times}10^7$ to $9.2{\times}10^8MPN/m{\ell}$, and acetate-utilizing methanogens : $2.3{\times}10^6$ to $2.0{\times}10^8MPN/m{\ell}$. The number of methanogens at the digester with membrane increased by approximately $10^2$ to $10^4$ times in comparison with that of the digester without membrane.

• KSBB Journal
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• v.26 no.3
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• pp.199-205
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• 2011

Using tetrameric ${\beta}$-galactosidase as a model protein, anchoring motives were screened in Bacillus subtilis spore display system. Eleven spore coat proteins were selected considering their expression levels and the location in the spore coat layer. After chromosomal single-copy homologous integration in the amyE site of Bacillus subtilis chromosome, cotE and cotG were chosen as possible spore surface anchoring motives with their higher whole cell ${\beta}$-galactosidase activity. PAGE and Wester blot of extracted fraction of outer layer of purified spore, which express CotE-LacZ or CotG-LacZ fusion verified the existence of exact size of fusion protein and its location in outer coat layer of purified spore. ${\beta}$-galactosidase activity of spore with CotE-LacZ or CotG-LacZ fusion reached its highest value around 16~20 h of culture time in terms of whole cell and purified spore. After intensive spore purification with lysozyme treatment and renografin treatment, spore of BJH135, which expresses CotE-LacZ, retained only 1~2% of its whole cell ${\beta}$-galactosidase activity. Whereas spore of BJH136, which has cotG-lacZ cassette in the chromosome, retained 10~15% of its whole cell ${\beta}$-galactosidase activity, proving minor perturbation of CotG-LacZ, when incorporated in the spore coat layer of Bacillus subtilis compared to CotE-LacZ. Usage of Bacillus subtilis WB700, of which 7 proteases are knocked-out and thereby resulting in 99.7% decrease in protease activity of the host, did not prevent the proteolytic degradation of spore surface expressed CotG-LacZ fusion protein.

• Textile Coloration and Finishing
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• v.19 no.1 s.92
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• pp.31-36
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• 2007

There are several factors in the degradation of textiles. The crucial factors in textile weakening are humidity, dust, smoke, sunlight, microorganisms and so on. Especially silk fabrics are more susceptible to microorganisms than other fabrics, because they are mainly consisted of proteins. In this study, we investigated the activities for degrading casein and silk fibers with 2 strains, Bacillus cereus TX1 and Pseudomonas fluorescens TX 2, isolated from domestic museums. They were compared to those of standard control strains, Klebsiella pneumoniae and Staphylococcus aureus, usually used for the antibiotic test of fabrics. The caseinolytic activities of K. pneumoniae and S. aureus were higher than those of isolated strains. But in the cases of silk fiber degrading, B. cereus TX 1 showed the highest activity on both silk 1 and silk 2. Therefore, caseinolytic activities were not coincident with the activity to degrade silk fibers. All strains degraded silk 1(strength retention 100%) better than silk 2(strength retention 50%). It means that bacteria mainly participate in the early stage of degrading silk fabrics, but as time goes by, the importance of bacteria for degrading silk fabrics would decreased. Even though the importance of bacteria may decrease, controlling bacterial activity is necessary to preserve historic silk fabrics.

• KSBB Journal
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• v.24 no.6
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• pp.556-562
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• 2009

The biological treatment of TNT-containing wastewater, known commonly as pink water, was investigated using a stirred tank reactor with Stenotrophomonas maltophilia OK-5 bacterial culture. S. maltophilia OK-5 exhibited effective degradation of TNT contained in pink water, completely degrading TNT (100 mg/L) within 6 days of incubation. The dark-red brown color derived from Hydride-Meisenheimer complex became more pronounced during the incubation period, which was determined quantitatively. High-pressure liquid chromatography was used to measure residual TNT, which also resolved the metabolic intermediates (i.e., 2,4-dinitrotoluene, 2,6-dinitrotoluene and 2,4-dinitro-6-hydroxytoluene). Gas chromatography-mass spectrometry was used to verify these intermediates. Quantification of the nitroreductase (pnrB) gene isolated from S. maltophilia OK-5 growing in pink water was performed with real-time PCR. The amount of pnrB gene copies increased to $10^3$-fold after 5 days of incubation time.