[ Objective] The research aimed to get the optimized separation and purification conditions of the hirudin produced from Bacillus subtilis DB403 （pUBH5）. [Method] Through the systemic pretreatment, preliminary chrom...[ Objective] The research aimed to get the optimized separation and purification conditions of the hirudin produced from Bacillus subtilis DB403 （pUBH5）. [Method] Through the systemic pretreatment, preliminary chromatography and fine chromatography. [Result]The optimized separation and purification conditions were that： Supernatant was treated by trichloroacetic acid, then by ultrafiltration desalt and anion exchange chromatography. Strong anion Q F. F. was better than weak anion DEAE F.F. The proper balanced solution was Tris-HCI （ pH 8.0）. The proper conductivity was 6 ms/cm. The maximum applied sample was 240 ATU/ml to matrix of strong anion Q F. F. This optimized procedure was magnified in strong anion exchange HiPrep 16/10Q with the 90% recovery and 70.2% purity. The purification of gel filtration of Sephacryl S-100 to hirudin was not relative to flow rate within certain scope. The application size of sample was 10 ml. The purity checked by HPLC was 95.1%, and the recovery was 93%, and the band of SDS-PAGE was single. [ Conclusion] The research provided the reference of the further industrialization separation and purification of hiruin.展开更多
The aim of this study was to elucidate the mode of action of two antimicrobial compounds, nisin, which is hydrophilic, and the lipophilic compound carvacrol. For that purpose, both antimicrobials were loaded into Oil-...The aim of this study was to elucidate the mode of action of two antimicrobial compounds, nisin, which is hydrophilic, and the lipophilic compound carvacrol. For that purpose, both antimicrobials were loaded into Oil-in-/Water nanoemulsions, and their antimicrobial activity against Bacillus subtilis was investigated. The experimental results indicated that the O/W nanoemulsions loaded with both, nisin and carvacrol, having Sauter mean diameter （d3,2） of around 120 nm, were considerably stable under the conditions investigated （up to 1 week storage at room temperature）. The interfacial tension between nisin aqueous solutions and soybean oil could be reduced up to 12 mN/m, as compared to that of pure water and soybean oil （21 mN/m）, so that demonstrating the potential of nisin to be used as emulsifier. The results obtained for O/W nanoemulsions loaded with both antimicrobial compounds indicated a reduction of nearly two log cycles （around 100 times） on B. subtilis population upon incubation for 24 h, compared to the blank （10 mM phosphate buffer）.展开更多
In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY- 3, which produced a bacteriocin-like substance （BLS）, was characterized to be a strain of Baci...In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY- 3, which produced a bacteriocin-like substance （BLS）, was characterized to be a strain of Bacillus subtillis by biochemical profiling and 16S rDNA sequence analysis. It is the first time to report that Bacillus subtilis from Jiaozhou Bay sediments could produce a BLS. The BLS of B. subtillis SLYY-3 exhibited strong inhibitory activity against gram-positive bacteria （including Staphylococcus aureus and B. subtillis） and some fimgi （including Penicillium glaucum, Aspergillus niger and Aspergillus flavus）. The antimicrobial activity was detected from culture in the exponential growth phase and reached its maximum when culture entered into stationary growth phase. It was thermo-tolerant even when being kept at 100~C for 60 min without losing any activity and stable over a wide pH range from 1.0 to 12.0 while being inactivated by proteolytic enzyme and trypsin, indicating the proteinaceous nature of the BLS. The BLS was purified by precipitation with hydrochloric acid （HC1） and gel filteration （Sephadex G-100）. SDS-PAGE analysis of the extracellular peptides of SLYY-3 revealed a bacteriocin-like protein with a molecular mass of 66 kDa. Altogether, these characteristics indicate the potential of the BLS for food industry as a protection against pathogenic and spoilage microorganisms.展开更多
The nitrogen source requirements for riboflavin production by ccpA mutant Bacillus subtilis 24A1/pMX45 were optimized using linear regression. The optimal medium components considered included 8% glucose as carbon sou...The nitrogen source requirements for riboflavin production by ccpA mutant Bacillus subtilis 24A1/pMX45 were optimized using linear regression. The optimal medium components considered included 8% glucose as carbon source, 2% yeast powder, 0.05% MgSO4 ·7H2O, and four types of nitrogen sources ： 0.1% yeast extract, 2% soybean powder, 1% corn plasm, and 0.2% （ NH4 ） 2 HPO4 in shake flask tests. Predictive ellipsoid was applied to determining the response values under the optimal levels for riboflavin production and glucose consumption. The optimal concentrations of the four types of nitrogen sources can remedy ammonium assimilative defection of ccpA mutant. Under the optimal conditions, the riboflavin yield increases to more than 5.0 g/L and 8%, glucose can be consumed completely after 60 h.展开更多
A riboflavin operon （rib operon） derived from Bacillus subtilis 368 was modified on structure and the resulting operons were expressed in various strains of Escherichia coli. The results showed that the optimization...A riboflavin operon （rib operon） derived from Bacillus subtilis 368 was modified on structure and the resulting operons were expressed in various strains of Escherichia coli. The results showed that the optimization of the rib operon and the host strain used for expression are two main factors affecting the riboflavin production. Replacing the promoterl and rfn box of the rib operon with a strong constructive promoter spol drastically increased the expression of the rib genes. When E. coli JM109 was used as the host strain, the highest riboflavin production reached 95.3μg/mL （about eight times higher than that of the unmodified r/b operon）. In addition, when tetracycline （20 μg/mL） was used as the selective pressure, compared with the ampicillin resistant transformants, a higher riboflavin yield was obtained in tetracycline resistant host strain.展开更多
The purpose of this work is to perform a detailed study of carbohydrate specificity of the new extracellular bacilli lectins which is considered to determine mechanisms of the lectins action. Sources of lectins were b...The purpose of this work is to perform a detailed study of carbohydrate specificity of the new extracellular bacilli lectins which is considered to determine mechanisms of the lectins action. Sources of lectins were bacterial strains from Ukrainian collection of microorganisms. The optimized protocol of bacilli lectins isolation and purification included precipitation with ammonium sulfate with subsequent gel filtration chromatography on Sepharose CL-6B. Hemagglutinating activity of bacilli lectins and their fine carbohydrate specificity to sialic acids and their derivatives as well as sialic asid-containing and asialic glycoconjugates were studied. The ability of extracellular bacilli lectins to discriminate a- and 13-conformation of carbohydrate molecule and the type of connection between the monomers was determined. Studied lectins showed the most affinity to glycoconjugates containing both types of sialic acids （N-acetylneuraminic acid （Neu5Ac） and N-glycolylneuraminic acid （NeuGc）） and it is supposed to be a basis of their diagnostic and analytical potential.展开更多
Paraquat(PQ),a highly effective herbicide,is widely used worldwide.PQ poisoning can cause multiple organ failure,in which the lung is the primary target organ.After PQ poisoning,the patient mortality rate is as high a...Paraquat(PQ),a highly effective herbicide,is widely used worldwide.PQ poisoning can cause multiple organ failure,in which the lung is the primary target organ.After PQ poisoning,the patient mortality rate is as high as 90%,and there is currently no specific antidote.The main clinical treatment is the use of glucocorticoids and cyclophosphamide for pulse therapy,but its effectiveness and safety are still uncertain.We investigated the effectiveness and safety of immunosuppressive pulse therapy with glucocorticoids and cyclophosphamide to evaluate the treatment value in patients with acute PQ poisoning.This meta-analysis,combined with seven trials that enrolled a total of 426 patients,showed that immunosuppressive pulse therapy with glucocorticoids and cyclophosphamide for PQ poisoning significantly reduced mortality of the study group(59.3%,134/226) compared with the control group(81.0%,162/200).There was no significant difference of hepatitis or renal failure between the control and study groups,indicating that immunosuppressive pulse therapy was relatively safe.Several patients were reported to have leukopenia and returned to normal after 1-2 weeks without any abnormalities.Two cases of non-fatal sepsis were reported and considered to be a side effect of the immunosuppressive pulse therapy.Thus,immunosuppressive pulse therapy can efficiently reduce the mortality of PQ poisoning and it is relatively safe.展开更多
基金Supported by 863 Program of China(2006AA03Z0453)NaturalScience Research Program of Higher Education of Jiangsu Province(09KJB230001)+1 种基金973 Program of China(2009CB724700)AndSchool Foundation of Jiangsu University(08JDG009)~~
文摘[ Objective] The research aimed to get the optimized separation and purification conditions of the hirudin produced from Bacillus subtilis DB403 （pUBH5）. [Method] Through the systemic pretreatment, preliminary chromatography and fine chromatography. [Result]The optimized separation and purification conditions were that： Supernatant was treated by trichloroacetic acid, then by ultrafiltration desalt and anion exchange chromatography. Strong anion Q F. F. was better than weak anion DEAE F.F. The proper balanced solution was Tris-HCI （ pH 8.0）. The proper conductivity was 6 ms/cm. The maximum applied sample was 240 ATU/ml to matrix of strong anion Q F. F. This optimized procedure was magnified in strong anion exchange HiPrep 16/10Q with the 90% recovery and 70.2% purity. The purification of gel filtration of Sephacryl S-100 to hirudin was not relative to flow rate within certain scope. The application size of sample was 10 ml. The purity checked by HPLC was 95.1%, and the recovery was 93%, and the band of SDS-PAGE was single. [ Conclusion] The research provided the reference of the further industrialization separation and purification of hiruin.
文摘The aim of this study was to elucidate the mode of action of two antimicrobial compounds, nisin, which is hydrophilic, and the lipophilic compound carvacrol. For that purpose, both antimicrobials were loaded into Oil-in-/Water nanoemulsions, and their antimicrobial activity against Bacillus subtilis was investigated. The experimental results indicated that the O/W nanoemulsions loaded with both, nisin and carvacrol, having Sauter mean diameter （d3,2） of around 120 nm, were considerably stable under the conditions investigated （up to 1 week storage at room temperature）. The interfacial tension between nisin aqueous solutions and soybean oil could be reduced up to 12 mN/m, as compared to that of pure water and soybean oil （21 mN/m）, so that demonstrating the potential of nisin to be used as emulsifier. The results obtained for O/W nanoemulsions loaded with both antimicrobial compounds indicated a reduction of nearly two log cycles （around 100 times） on B. subtilis population upon incubation for 24 h, compared to the blank （10 mM phosphate buffer）.
基金supported by the National Science and Technology Support Program (No. 2011BAD14B04)Project of Shandong Province Higher Educational Science and Technology Program (J14LE59)+1 种基金Applied & Basic Research Foundation of Qingdao (No. 12-1-4-3(3)-jch)Science & Technology Project of AQSIQ (No. 2012IK176)
文摘In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY- 3, which produced a bacteriocin-like substance （BLS）, was characterized to be a strain of Bacillus subtillis by biochemical profiling and 16S rDNA sequence analysis. It is the first time to report that Bacillus subtilis from Jiaozhou Bay sediments could produce a BLS. The BLS of B. subtillis SLYY-3 exhibited strong inhibitory activity against gram-positive bacteria （including Staphylococcus aureus and B. subtillis） and some fimgi （including Penicillium glaucum, Aspergillus niger and Aspergillus flavus）. The antimicrobial activity was detected from culture in the exponential growth phase and reached its maximum when culture entered into stationary growth phase. It was thermo-tolerant even when being kept at 100~C for 60 min without losing any activity and stable over a wide pH range from 1.0 to 12.0 while being inactivated by proteolytic enzyme and trypsin, indicating the proteinaceous nature of the BLS. The BLS was purified by precipitation with hydrochloric acid （HC1） and gel filteration （Sephadex G-100）. SDS-PAGE analysis of the extracellular peptides of SLYY-3 revealed a bacteriocin-like protein with a molecular mass of 66 kDa. Altogether, these characteristics indicate the potential of the BLS for food industry as a protection against pathogenic and spoilage microorganisms.
文摘The nitrogen source requirements for riboflavin production by ccpA mutant Bacillus subtilis 24A1/pMX45 were optimized using linear regression. The optimal medium components considered included 8% glucose as carbon source, 2% yeast powder, 0.05% MgSO4 ·7H2O, and four types of nitrogen sources ： 0.1% yeast extract, 2% soybean powder, 1% corn plasm, and 0.2% （ NH4 ） 2 HPO4 in shake flask tests. Predictive ellipsoid was applied to determining the response values under the optimal levels for riboflavin production and glucose consumption. The optimal concentrations of the four types of nitrogen sources can remedy ammonium assimilative defection of ccpA mutant. Under the optimal conditions, the riboflavin yield increases to more than 5.0 g/L and 8%, glucose can be consumed completely after 60 h.
文摘A riboflavin operon （rib operon） derived from Bacillus subtilis 368 was modified on structure and the resulting operons were expressed in various strains of Escherichia coli. The results showed that the optimization of the rib operon and the host strain used for expression are two main factors affecting the riboflavin production. Replacing the promoterl and rfn box of the rib operon with a strong constructive promoter spol drastically increased the expression of the rib genes. When E. coli JM109 was used as the host strain, the highest riboflavin production reached 95.3μg/mL （about eight times higher than that of the unmodified r/b operon）. In addition, when tetracycline （20 μg/mL） was used as the selective pressure, compared with the ampicillin resistant transformants, a higher riboflavin yield was obtained in tetracycline resistant host strain.
文摘The purpose of this work is to perform a detailed study of carbohydrate specificity of the new extracellular bacilli lectins which is considered to determine mechanisms of the lectins action. Sources of lectins were bacterial strains from Ukrainian collection of microorganisms. The optimized protocol of bacilli lectins isolation and purification included precipitation with ammonium sulfate with subsequent gel filtration chromatography on Sepharose CL-6B. Hemagglutinating activity of bacilli lectins and their fine carbohydrate specificity to sialic acids and their derivatives as well as sialic asid-containing and asialic glycoconjugates were studied. The ability of extracellular bacilli lectins to discriminate a- and 13-conformation of carbohydrate molecule and the type of connection between the monomers was determined. Studied lectins showed the most affinity to glycoconjugates containing both types of sialic acids （N-acetylneuraminic acid （Neu5Ac） and N-glycolylneuraminic acid （NeuGc）） and it is supposed to be a basis of their diagnostic and analytical potential.
基金Project supported by the Foundation of Key Discipline Construction of Zhejiang Province for Traditional Chinese Medicine(No.2017-XK-A36)the Medical and Health Science Foundation of Zhejiang Province(No.2019327552)+1 种基金the Foundation of Key Research Project of Zhejiang Province for Traditional Chinese Medicine(No.2019ZZ014)the Foundation of Science and Technology Department of Zhejiang Province for Beneficial Technology Research of Social Development(No.2015C33146),China
文摘Paraquat(PQ),a highly effective herbicide,is widely used worldwide.PQ poisoning can cause multiple organ failure,in which the lung is the primary target organ.After PQ poisoning,the patient mortality rate is as high as 90%,and there is currently no specific antidote.The main clinical treatment is the use of glucocorticoids and cyclophosphamide for pulse therapy,but its effectiveness and safety are still uncertain.We investigated the effectiveness and safety of immunosuppressive pulse therapy with glucocorticoids and cyclophosphamide to evaluate the treatment value in patients with acute PQ poisoning.This meta-analysis,combined with seven trials that enrolled a total of 426 patients,showed that immunosuppressive pulse therapy with glucocorticoids and cyclophosphamide for PQ poisoning significantly reduced mortality of the study group(59.3%,134/226) compared with the control group(81.0%,162/200).There was no significant difference of hepatitis or renal failure between the control and study groups,indicating that immunosuppressive pulse therapy was relatively safe.Several patients were reported to have leukopenia and returned to normal after 1-2 weeks without any abnormalities.Two cases of non-fatal sepsis were reported and considered to be a side effect of the immunosuppressive pulse therapy.Thus,immunosuppressive pulse therapy can efficiently reduce the mortality of PQ poisoning and it is relatively safe.
