• Journal of Life Science
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• v.27 no.10
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• pp.1145-1151
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• 2017

In this study, we report on a simple, efficient method for obtaining penicillin G amidase (PGA) from recombinant Escherichia coli using a formulation mixed with detergent and lysozyme. Research was conducted on the extraction efficiency of PGA from the periplasmic space in cells in terms of the type of detergent, detergent concentration, pH, reaction time, and temperature of permeabilization. The extraction yield of PGA in the formulated surfactant/lysozyme treatment was increased by approximately (55-65 U/ml) in comparison with that in the single surfactant treatment. The released PGA solution was concentrated and exchanged with buffer using an ultrafiltration (U/F) system. The yields of diatomite filtration, membrane filtration (M/F), and U/F were 69.7%, 93.8%, and 77.3%, respectively. A total of 212 KU of PGA was recovered. At the 25-L culture scale, the overall yield of extraction using the mixed surfactant/lysozyme method was 49.2%. The specific activity of extracted PGA was 11 U/mg in protein. The concentrated PGA solution was immobilized on microporous silica beads without further purification of PGA. The total immobilization yield of PGA on the resin was 48.7%, while the enzyme activity was 101 U/g. The immobilized PGA was successfully used to produce 6-APA from penicillin G. Our results indicated that a simple extraction method from periplasmic space in E. coli may be used for the commercial scale production of ${\beta}-lactam$ antibiotics using immobilized PGA.

• Journal of Microbiology and Biotechnology
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• v.3 no.1
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• pp.31-38
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• 1993

A bacterial strain KY-l isolated from sewage was able to produce an extracellular agarase(agarose 4-glycanohydrolase. EC 3.2.1.81). The strain KY-1 was identified as Cytophaga fermentans subsp. agarovorans based on its morphological and physiological characteristics. The agarase was purified by ammonium sulfate precipitation followed by DEAE-Sephadex A-50. Bio-Gel P-100. and CM-Cellulose column chromatography. The molecular weight of the purified enzyme was 24 kDa by SDS-polyacrylamide gel electrophoresis. The optimum temperature and pH for the enzyme activity were 30^{circ}C and 7.5, respectively. The enzyme activity was significantly inhibited in the presence of 0.1 mM $HgCl_2$. whereas it was elevated 3 times by $MnSO_4$ at 1 mM concentration. The Km value and Vmax were 16.67 mg/ml and 3.77 unit/ml.min. The agarase gene was cloned into Escherichia coli MC1061 using the plasmid vector pBR322. A 1.4 Kb DNA fragment of PstI-digested chromosomal DNA of C. fermentans KY-l was inserted into the PstI site of pBR322. expressed in the E. coli. and up to 60% of the total enzyme was extracellularly secreted. Enzymatic properties of the extracellular agarases produced by both the transformant and the donor were very similar in terms of optimal pH and temperature.

• Journal of Korean Society of Environmental Engineers
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• v.27 no.5
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• pp.549-554
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• 2005

Porous ceramic beads that had skeleton structure were impregnated with Ag from silver nitrate solution. Ag-impregnated porous ceramic beads were performed to evaluate the antibacterial properties on Escherichia coli and Staphylococcus aureus, also, compared with commercial silver-activated carbon on antibacterial activity. As concentration of silver nitrate solution increased, deposited-Ag contents of outer and inner surface of beads were increased. The size of silver particles supported on porous ceramic bead were range of $0.5{\sim}2.0\;{\mu}m$. The observed effects of the prepared Ag-impregnated beads on antibacterial activity are as follows : i) Antibacterial activity should be directly proportional to silver nitrate solution and reaction time. ii) The antibacterial activity against Escherichia coli was better than that against Staphylococcus aureus.

• Tuberculosis and Respiratory Diseases
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• v.43 no.4
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• pp.579-589
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• 1996

Background : Activation of neutrophil is critical for the clearance of microorganisms and toxic host mediators during sepsis. Unfortunately the activated neutrophil and its toxic byproducts can produce tissue injury and organ dysfunction. The leucocyte CD11/18 adhesion complex regulates neutrophil-endothelial cell adhesion, the first step in neutrophil migration to sites of injection and inflammation. To investigate the potential of neutrophil inhibition as a treatment strategy for sepsis, we evaluated the effects of monoclonal antibody against CD11b (MAb 1B6) in rats intrabronchial challenged with Escherichia coli. Methods : Animals were randomly assigned to receive monoclonal antibody against CD11b (1 mg/kg, sc) and bovine serum albumin(BSA, 1 mg/kg, sc) 6 hr before, at 0 and 6 hr after intrabronchial challenge of $20x10^9$ CFU/kg E. coli 0111. Animals were randomized to treat either 24, 60 or 90% oxygen after bacterial challenge and begining 4 hr after inoculation, all animals were received 100 mg/kg ceftriaxone qd for 3 days. Peripheral and alveolar neutrophil(by bronchoalveolar lavage) counts and lung injury parameters such as alveolar-arte rial $PO_2$ difference, wet to dry lung weight ratio and protein concentration of alveolar fluid were measured in survived rats at 12 hr and 96 hr. Results : Monoclonal antibody against CD11b decreased circulating and alveolar neutrophil especially more in 12 hr than in 96 hr The lung injury parameters of antibody-treated animals were not different from those of BSA-treated animals. but It was meaningless due to small number of survived animals. The early(6 hr) mortality rate was significantly increased in antibody-treated group(51%) compared to BSA-treated group(31%) (P=0.02) but late(from 12 hr to 72 hr) mortality rate was not different in antibody-treated group(44%) from BSA-treated group(36%) (P =0.089). Conclusion : Leucocyte CD11b/18 adhesion molecule is known to regulate neutrophil migration to the site of infection and inflammation. The monoclonal antibody against CD11b decreased alveolar neutrophil in rats with pulmonary sepsis and increased early mortality rate. Therefore, we can speculate that monoclonal antibody against CD11b blocks of alveolar recruitment of neutrophils, impairs host defense mechanism and increases early mortality rate of pulmonary sepsis in rat.

• Food Science and Preservation
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• v.16 no.4
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• pp.472-481
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• 2009

Response surface methodology (RSM) is frequently used for optimization studies. In the present work, RSM was used to determine the antimicrobial activitiesof grapefruit seed extract (GFSE) and a lactic acid mixture (LA) against Staphylococcus aureus, Bacillus cereus, Escherichia coli, Salmonella typhimurium, Pseudomonas fluorescens, and Vibrio parahaemolyticus. A central composite design was used to investigate the effects of independent variables on dependent parameters. One set of antimicrobial preparations included mixtures of 1% (w/w) GFSE and 10% (w/w) LA, in which the relative proportions of component antimicrobials varied between 0 and 100%. In further experiments, the relative proportions were between 20% and 100%. Antimicrobial effects against various microorganisms were mathematically encoded for analysis. The codes are given in parentheses after the bacterial names, and were S. aureus ($Y_1$), B. cereus ($Y_2$), E. coli ($Y_3$), S. typhimurium ($Y_4$), P. fluorescens ($Y_5$), and V. parahaemolyticus ($Y_6$). The optimum antimicrobial activity of the 1% (w/w) GFSE:10% (w/w) LA mixture against each microorganism was obtained by superimposing contour plots ofantimicrobial activities on measures of response obtained under various conditions. The optimum rangesfor maximum antimicrobial activity of a mixture with a ratio of 1:10 (by weight) GFSE and LA were 35.73:64.27 and 56.58:43.42 (v/v), and the optimum mixture ratio was 51.70-100%. Under the tested conditions (a ratio of 1% [w/w] GFSE to 10% [w/w] LA of 40:60, and a concentration of 1% [w/w] GFSE and 10% [w/w] LA, 70% of the highest value tested), and within optimum antimicrobial activity ranges, the antimicrobial activities of the 1% (w/w) GFSE:10% (w/w) LA mixture against S. aureus ($Y_1$), B. cereus ($Y_2$), E. coli ($Y_3$), S. typhimurium ($Y_4$), P. fluorescens ($Y_5$), and V. parahaemolyticus ($Y_6$) were 24.55, 25.22, 20.20, 22.49, 23.89, and 28.04 mm, respectively. The predicted values at optimum conditions were similar to experimental values.

• KSBB Journal
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• v.11 no.3
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• pp.380-387
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• 1996

The growth of recombinant E. coli and formation of the by-products were investigated. Glucose was consumed in approximately 8hours of cultivation when initial glucose concentration was 5g/1. When higher initial glucose concentrations were employed, it took approximately 16 to 20hours for the glucose to be consumed. When maltose or glucose was used as a carbon source, lactic acid and acetic acid were main by-products, while propionic acid and methanol production was not significant. Much by- products were formed at the higher initial carbon source concentrations. Higher concentration of lactic acid was observed when glucose was used, while higher concentration of acetic acid was observed when maltose was used. Lactic acid production affected the cell yield while acetic acid production affected the growth rate.

• Korean Journal of Veterinary Service
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• v.36 no.4
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• pp.311-320
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• 2013

The purpose of this study was to evaluate detecting methods and the relationship between tissues and blood for enrofloxacin and metabolic ciprofloxacin residues in broiler chickens. Two groups of broiler chickens were administrated via the drinking water with $50{\mu}g/mL$ and $100{\mu}g/mL$ of enrofloxacin for 5 days, respectively. The concentration of enrofloxacin and metabolic ciprofloxacin in tissues (muscle and kidney) and blood were measured during administration period (for 5 days) and withdrawal period (for 12 days) by high performance liquid chromatography (HPLC) method. Also, all samples were conducted for screening of residues by microbial method using E. coli for quinolone detection and immuno-chromatography method using Smart kit. The relationship between tissues (muscle and kidney) and blood for enrofloxacin and metabolic ciprofloxacin residues in broiler chickens was followed : The levels of enrofloxacin and metabolic ciprofloxacin residues in muscle and kidney were higher 2.9~3.2 folds, 3.6~3.8 folds more than the residues levels in blood, respectively. These results support we can predict the residues in muscle and kidney from the residues in blood. In comparison of detecting methods for antibiotic residues, microbial method using E. coli for quinolone detection and immuno-chromatography method using Smart kit could detect positive reaction at similar or lower concentration than violative concentration of enrofloxacin and metabolic ciprofloxacin in chicken tissues. These results support what two screening methods are useful for screening of quinolone detection in chickens.

• Food Science and Preservation
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• v.11 no.3
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• pp.319-324
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• 2004

The antimicrobial activity of the extract of citrus peel prepared by the method of hot water, ethanol and sugar was examined. The results showed that the extract of citrus peel prepared by hot water or ethanol did not have antimicrobial activity, but the extract by 10$\%$(w/v) sugar revealed the high antimicrobial activity. Extracted in 10%(w/v) sugar solution for 9 days, showed the highest antimicrobial activity against 8 strains of bacteria. The minimum inhibition concentration was found to be 0.5$\%$(v/v) against S. aureus, 1.5$\%$(v/v) against B. subtilis, M. luteus and E. coli, and 2.0$\%$(v/v) against S. mutans. The antimicrobial activity of the citrus peel extract was stable regardless of the treatment at 40 $\~$ 100 $^{\circ}C$C for 20 min and unstable response to the change of pH. The results suggested the development of citrus peel as heat-stable antimicrobial agents.

• Journal of Korean Society of Occupational and Environmental Hygiene
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• v.8 no.2
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• pp.231-241
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• 1998

To assess biological indoor air quality in hospital, concentrations of viable airborne microbes were determined at intensive care unit(ICU), patient room (PR), outpatient waiting room(OPWR) in hospitals of large(1000 beds), middle(500 beds), small(100 beds) hospitals, respectively. Gram positive bacteria, gram negative bacteria, fungi were sampled using suctional sampling method by RCS sampler (Reuter centrifugal air sampler) and RCS GK-A agar plate. In gram positive bacteria groups, CNS(Coagulase Negative Staphylococcus), Micrococcus, Lactobacillus, S. aureus, Enterococcus, St. viridans identified. In gram negative bacteria groups, A. baumannii, Kl. peumoniae and E. coli were identified, and Penicillium was identified in fugi groups. Results of the study were as follows. 1. The highest concentrations of airborne microbes was $971CFU/m^3$ at 5:00 PM in small hospital patient room, and average concentrations of airborne microbes in large, middle and small hospitals were $282CFU/m^3$, $289CFU/m^3$ and $625CFU/m^3$, respectively. Average concentrations of airborne microbes in office(control) was $90CFU/m^3$. Thus, the small hospital showed the worst condition. 2. Representatives of 8 different genera were identified in 150 samples. The most frequently isolated organisms were Staphylococcus (73.0%), Micrococcus (20.7%) and Lactobacillus (4.7%), respectively. Pathogenic microbes isolated were A. baumannii, E. coli, Enterococcus, Kl. peumoniae, S. aureus, St. viridans and Penicillium as fungi. In office, no pathogenic microbes were identified. Average concentrations of airborne pathogenic microbes in large, middle and small hospital were $5CFU/m^3$ (2%), $11CFU/m^3$ (4%) and $12CFU/m^3$ (2%), respectively. Thus, condition in a large hospital was better than those in a middle and a small hospital.

• Journal of the Korea Organic Resources Recycling Association
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• v.15 no.2
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• pp.98-108
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• 2007

In order to use food wastes for the source of fermented feed for pigs, this study was aimed to produce better culture inoculum by the aeration and addition of pig' s blood meal as sub nutrient. For the preparation of inoculum as bacterial strain, Lactobacillus brevis isolated from pig intestine, and a yeast Saccharomyces cerevisiae from strawberries were used. Molasses and whey were used as main ingredients for the culture solution as well as yeast extract and other ingredients as sub nutrients. As the experimental result, aeration showed a positive effect to enhance viable cell count or retarding death phase. Although sub nutrient yeast extracts were replaced with pig's blood meal, fermentation characteristics were almost similar to that of yeast extract. When the inoculum was stored at room temperature, L. brevis and S. cerevisiae maintained the viable cell concentration of approximately 8 log cfu/mL for 1 week. 2 Days after the culture solution was mixed with food waste, the number of unwanted bacteria had rapidly increased, but E.coli was not detected for 5 days.