Extracellular manganese peroxidases （MnPs） produced by native and mutant strains of Trametes versicolor IBL‐04 （EB‐60, EMS‐90） were purified by ammonium sulphate precipitation and dialysis, followed by ion‐exc...Extracellular manganese peroxidases （MnPs） produced by native and mutant strains of Trametes versicolor IBL‐04 （EB‐60, EMS‐90） were purified by ammonium sulphate precipitation and dialysis, followed by ion‐exchange and gel‐permeation chromatography. The purified enzymes elucidated a single band in the 43‐kDa region on sodium dodecyl sulphate‐polyacrylamide gel electrophoresis. The optimum pH and temperature of the purified enzymes were found to be 5.0 and 40 °C, respec‐tively. Mutant strain MnPs exhibited a broader active pH range and higher thermal stability than native MnP. Purified MnPs from selected mutants showed almost identical properties to native MnP in electrophoresis, steady‐state kinetics, and metal ion and endocrine‐disrupting compound （EDC） degradation efficiency. Although the fastest reaction rates occurred with Mn2＋, MnPs displayed the highest affinity for ABTS, methoxyhydroquinone, 4‐aminophenol and reactive dyes. MnP activity was significantly enhanced by Mn2＋and Cu2＋, and inhibited in the presence of Zn2＋, Fe2＋, ethylene‐diaminetetraacetic acid and cysteine to various extents, with Hg2＋ as the most potent inhibitory agent. MnPs from all sources efficiently catalyzed the degradation of the EDCs, nonylphenol and triclosan, removing over 80%after 3 h of treatment, which was further increased up to 90%in the presence of MnP‐mediator system. The properties of T. versicolor MnPs, such as high pH and ther‐mal stability, as well as unique Michaelis‐Menten kinetic parameters and high EDC elimination effi‐ciency, render them promising candidates for industrial exploitation.展开更多
Separation and purification of dodecanedioic acid (DDDA) from its homologous compounds were studied experimentally by falling film crystallization (FFC). The influences of various operation parameters, including cryst...Separation and purification of dodecanedioic acid (DDDA) from its homologous compounds were studied experimentally by falling film crystallization (FFC). The influences of various operation parameters, including crystallizing time, flow rate of melt and temperature of glycerine bath, on purity of DDDA and crystallizing rate were investigated. Over 99% (by mole) DDDA was obtained for a feed composition of 96% (by mole). The main factors affecting the separation efficiency are flow rate of melt and temperature of glycerine bath. The crystallizing layer of DDDA was further purified by sweating and blasting. A set of optimized operation data are provided for better understanding the mechanism of heat and mass transfer in FFC, and for further industrial application of DDDA purification process.展开更多
A new technique was developed for the integrated processing of cell disruption and aqueous two-phase extraction in a high-speed bead mill to separate intracellular proteins from microbial cells. The process was named ...A new technique was developed for the integrated processing of cell disruption and aqueous two-phase extraction in a high-speed bead mill to separate intracellular proteins from microbial cells. The process was named as simultaneous cell disruption and aqueous two-phase extraction (SDATE). Advantages, such as high cell disruption efficiency, biochemical activities preservation of proteins, cell debris elimination, and prelimiary purification of the target protein were being claimed. When this technique was employed for isolating recombinant Tumor Necrosis Factor (TNF) from E. coli, overall protein concentration and TNF activity were found to have been increased. More than 95% of TNF was partitioned into the top phase and all cell debris were in the bottom phase. The partition coefficient was greater than 3 and the TNF purification factor was greater than 6. It is shown that less separation steps were being utilized in the new technique, meaning a reduction in separation time and less process extractors required.展开更多
The title compound Cu3PSe4 was synthesized by the reaction of CuCl, P2Se5 and Se in a molar ratio of 1:1:1 at 500 C and structurally characterized by X-ray crystallography. The crystal belongs to orthorhombic, space g...The title compound Cu3PSe4 was synthesized by the reaction of CuCl, P2Se5 and Se in a molar ratio of 1:1:1 at 500 C and structurally characterized by X-ray crystallography. The crystal belongs to orthorhombic, space group Pmn21 with cell parameters: a = 7.685(2), b = 6.656(1), c = 6.377(1) , V = 326.2(1) 3, Z = 2, Dc = 5.472 g/cm3, Mr = 537.43, F(000) = 476, m = 32.12 mm-1, R = 0.0642, wR = 0.1481 and S = 1.037. The 3-D structure can be regarded as constructed from the alternately stacking of [Cu(2)Se4] tetrahedral layers and Cu(1)PSe tetrahedral layers along the b direction, in which the Cu(2)Se layer is comprised of corner-sharing [Cu(2)Se4] tetrahedra along the a and c directions, and the Cu(1)PSe layer is consisted of alternately corner-sharing [Cu(1)Se4] tetrahedra and [PSe4] tetrahedra along the a and c directions.展开更多
Our previously described environmental DNA extraction method has been widely used in environmental microbial community analysis. However, residual humic substances may remain with obtained environmental DNA, which int...Our previously described environmental DNA extraction method has been widely used in environmental microbial community analysis. However, residual humic substances may remain with obtained environmental DNA, which interferes downstream molecular analyses. To remedy this situation, two DNA extraction buffers （PIPES and Tris-HCl） and four purification strategies including our new modified low melting point gel purification method and three commercial kits from QIAEX, Omega and Promega were evaluated with diverse soil samples. The PIPES buffer （pH 6.5） is found to be more effective for removing the humic substances, but it leads to lower DNA yield and causes more severe DNA shearing than using the Tris-HC1 buffer （pH 8.0）. Gel purification and the Promega purification kit achieve much higher DNA recoveries than QIAEX or Omega kit, and higher purity of DNA is obtained by gel purification than by the Promega kit with both DNA extraction buffers mentioned above. Considering all results together, two alternative methods for DNA extraction and purification are proposed： one uses Tris-HCl buffer extraction and gel purification as the primary approach when the amount of soil or biomass is not a major concern, and the other uses PIPES buffer extraction and the Promega kit purification when severe DNA shearing and/or limited biomass occurs. Purified DNA samples by both methods are amenable for use as templates for whole community genome amplifications and PCR amplifications of bacterial 16S rRNA genes. It is demonstrated that these two alternative methods could be applied to a wide variety of environmental samples.展开更多
To observe the effect of Gardenia extract ZG on the adsorption quantity of herpes simplex virus type 1 （HSV-1） so as to explore the mechanism of its antiviral activity, fluorescein isothiocyanate （FITC） was used a...To observe the effect of Gardenia extract ZG on the adsorption quantity of herpes simplex virus type 1 （HSV-1） so as to explore the mechanism of its antiviral activity, fluorescein isothiocyanate （FITC） was used as the fluorescent probe to label viruses and heparin sodium was used as control. Meanwhile, the effect of Gardenia extract ZG on the adsorption quantity on the surface of Hep-2 cells was determined by flow cytometry. It was demonstrated that adsorption of HSV-1 on the surface of Hep-2 cells exhibited the character of saturation and specificity and heparin sodium could prevent attachment of viruses on these ceils. These results are in accord with those reported previously. It was also proved that the manner of drug-use prior to adsorption or simultaneous use of drug and adsorption was better than adsorption prior to drug-use, and the inhibition rates of the former and latter manner were 84.76% and 82.92% respectively. Three manners of drug-use with Gardenia extract ZG were all effective to reduce the adsorption quantity of viruses, especially the manner of simultaneous use of drug and adsorption with an adsorption inhibition rate of 68.46%. From the above observation, it is apparent that the mechanism of anti-viral activity of Gardenia extract ZG may be via several steps involved in the HSV-1 adsorption.展开更多
Plant peroxidase extracted from Coccinia grandis Lin. （Ivy gourd） was purified from crude extract by ammonium sulfate precipitation, ion-exchange chromatography, and size exclusion chromatography. The purified enzym...Plant peroxidase extracted from Coccinia grandis Lin. （Ivy gourd） was purified from crude extract by ammonium sulfate precipitation, ion-exchange chromatography, and size exclusion chromatography. The purified enzyme preparation exhibited a specific activity of 6106.63 p.mol.minl.mg protein1, while purification fold and yield were 17.45 and 34.70%, respectively. The purified peroxidase was homogenous as judged by native and sodium dodecyl sulfate （SDS） polyacrylamide gel electrophoresis. The molecular weight as determined by gel filtration and SDS-polyacrylamide gel electrophoresis was 45 kD, which suggested that the purified peroxidase contained only one subunit. The apparent Km and Vmax values of the enzyme against phenol were 93 p.M and 561 μmol.min^-1.mg protein^-1, respectively. The temperature and pH optimum for purified peroxidase were 45℃ and pH 6.0, respective. However, it was stable at 30-60℃ and pH 4.0-8.0. The presence of metal ions such as Cu2＋ and Ca2＋ enhanced peroxidase activity. On the other hand, Cr3＋ and Hg2＋ strongly inhibited the enzyme activity at 500 p.M. Sodium dodecyl sulfate reduced a half of peroxidase activity at approximately 3 mM. Ivy gourd was stability in the presence of each urea concentrations. The affinity of the enzyme with different substrates showed as the highest relative activities on gallic acid followed by catechin, ascorbic acid and caffeic acid, respectively.展开更多
OBJECTIVE: To study the antiviral activities of clemastanin B(CB), epigoitrin, phenylpropanoids portion(PEP) and the mixture of phenylpropanoids, alkaloids and organic acid fractions(PEP+ALK+OA)from Banlangen(Radix Is...OBJECTIVE: To study the antiviral activities of clemastanin B(CB), epigoitrin, phenylpropanoids portion(PEP) and the mixture of phenylpropanoids, alkaloids and organic acid fractions(PEP+ALK+OA)from Banlangen(Radix Isatidis).METHODS: The experiment consisted of four parts:therapeutic action, prophylaxsis action, inhibition of virus attachment, and direct virucidal action. Cytopathic effect(CPE) and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium(MTT) were used to assess antiviral activity.RESULTS: CB, epigoitrin, PEP and PEP + ALK + OA fractions from Banlangen(Radix Isatidis) extract significantly increased the viability of MDCK cells pre-infected with the virus compared with the virus control group in all the dilutions(P < 0.01). Pretreated with either pure compounds or chemical frac-tions of Banlangen(Radix Isatidis) extract in all the dilutions significantly improved the viability of MDCK cells(P < 0.01). The inhibition of virus absorption to the host cells by CB, epigoitrin and PEP was in a dose dependent manner.CONCLUSION: CB, epigoitrin, PEP and PEP+ALK+OA exert their anti-influenza activity by inhibiting the virus multiplication, prophylaxsis and blocking the virus attachment. The primary mode of action of PEP and PEP + ALK + OA is the inhibition of virus replication. The inhibitory effects on virus attachment and multiplication are the main modes for epigoitrin.展开更多
基金a part of a research project entitled "The development of immobilized ligninolytic enzymes for industrial applications" supported by Higher Education Commission (HEC), Islamabad, Pakistan
文摘Extracellular manganese peroxidases （MnPs） produced by native and mutant strains of Trametes versicolor IBL‐04 （EB‐60, EMS‐90） were purified by ammonium sulphate precipitation and dialysis, followed by ion‐exchange and gel‐permeation chromatography. The purified enzymes elucidated a single band in the 43‐kDa region on sodium dodecyl sulphate‐polyacrylamide gel electrophoresis. The optimum pH and temperature of the purified enzymes were found to be 5.0 and 40 °C, respec‐tively. Mutant strain MnPs exhibited a broader active pH range and higher thermal stability than native MnP. Purified MnPs from selected mutants showed almost identical properties to native MnP in electrophoresis, steady‐state kinetics, and metal ion and endocrine‐disrupting compound （EDC） degradation efficiency. Although the fastest reaction rates occurred with Mn2＋, MnPs displayed the highest affinity for ABTS, methoxyhydroquinone, 4‐aminophenol and reactive dyes. MnP activity was significantly enhanced by Mn2＋and Cu2＋, and inhibited in the presence of Zn2＋, Fe2＋, ethylene‐diaminetetraacetic acid and cysteine to various extents, with Hg2＋ as the most potent inhibitory agent. MnPs from all sources efficiently catalyzed the degradation of the EDCs, nonylphenol and triclosan, removing over 80%after 3 h of treatment, which was further increased up to 90%in the presence of MnP‐mediator system. The properties of T. versicolor MnPs, such as high pH and ther‐mal stability, as well as unique Michaelis‐Menten kinetic parameters and high EDC elimination effi‐ciency, render them promising candidates for industrial exploitation.
文摘Separation and purification of dodecanedioic acid (DDDA) from its homologous compounds were studied experimentally by falling film crystallization (FFC). The influences of various operation parameters, including crystallizing time, flow rate of melt and temperature of glycerine bath, on purity of DDDA and crystallizing rate were investigated. Over 99% (by mole) DDDA was obtained for a feed composition of 96% (by mole). The main factors affecting the separation efficiency are flow rate of melt and temperature of glycerine bath. The crystallizing layer of DDDA was further purified by sweating and blasting. A set of optimized operation data are provided for better understanding the mechanism of heat and mass transfer in FFC, and for further industrial application of DDDA purification process.
基金Supported by the National Natural Science Foundation of China(No.295256O9 and 29736180).
文摘A new technique was developed for the integrated processing of cell disruption and aqueous two-phase extraction in a high-speed bead mill to separate intracellular proteins from microbial cells. The process was named as simultaneous cell disruption and aqueous two-phase extraction (SDATE). Advantages, such as high cell disruption efficiency, biochemical activities preservation of proteins, cell debris elimination, and prelimiary purification of the target protein were being claimed. When this technique was employed for isolating recombinant Tumor Necrosis Factor (TNF) from E. coli, overall protein concentration and TNF activity were found to have been increased. More than 95% of TNF was partitioned into the top phase and all cell debris were in the bottom phase. The partition coefficient was greater than 3 and the TNF purification factor was greater than 6. It is shown that less separation steps were being utilized in the new technique, meaning a reduction in separation time and less process extractors required.
基金This work was supported by the National Natural Science Foundation of China (20001007 20131020) Natural Science Foundation of the Chinese Academy of Sciences (KJCX2-H3) and Fujian Province (2000F006)
文摘The title compound Cu3PSe4 was synthesized by the reaction of CuCl, P2Se5 and Se in a molar ratio of 1:1:1 at 500 C and structurally characterized by X-ray crystallography. The crystal belongs to orthorhombic, space group Pmn21 with cell parameters: a = 7.685(2), b = 6.656(1), c = 6.377(1) , V = 326.2(1) 3, Z = 2, Dc = 5.472 g/cm3, Mr = 537.43, F(000) = 476, m = 32.12 mm-1, R = 0.0642, wR = 0.1481 and S = 1.037. The 3-D structure can be regarded as constructed from the alternately stacking of [Cu(2)Se4] tetrahedral layers and Cu(1)PSe tetrahedral layers along the b direction, in which the Cu(2)Se layer is comprised of corner-sharing [Cu(2)Se4] tetrahedra along the a and c directions, and the Cu(1)PSe layer is consisted of alternately corner-sharing [Cu(1)Se4] tetrahedra and [PSe4] tetrahedra along the a and c directions.
基金Project(51104189)supported by the National Natural Science Foundation of ChinaProject(2010CB630901)supported by the National Basic Research Program of China+1 种基金Project(1343-77341)supported by the Graduate Education Innovative Program of Central South University,ChinaProject(DOE-ER64125)supported by Department of Energy,Office of Science under the Environmental Remediation Science Program of the United States
文摘Our previously described environmental DNA extraction method has been widely used in environmental microbial community analysis. However, residual humic substances may remain with obtained environmental DNA, which interferes downstream molecular analyses. To remedy this situation, two DNA extraction buffers （PIPES and Tris-HCl） and four purification strategies including our new modified low melting point gel purification method and three commercial kits from QIAEX, Omega and Promega were evaluated with diverse soil samples. The PIPES buffer （pH 6.5） is found to be more effective for removing the humic substances, but it leads to lower DNA yield and causes more severe DNA shearing than using the Tris-HC1 buffer （pH 8.0）. Gel purification and the Promega purification kit achieve much higher DNA recoveries than QIAEX or Omega kit, and higher purity of DNA is obtained by gel purification than by the Promega kit with both DNA extraction buffers mentioned above. Considering all results together, two alternative methods for DNA extraction and purification are proposed： one uses Tris-HCl buffer extraction and gel purification as the primary approach when the amount of soil or biomass is not a major concern, and the other uses PIPES buffer extraction and the Promega kit purification when severe DNA shearing and/or limited biomass occurs. Purified DNA samples by both methods are amenable for use as templates for whole community genome amplifications and PCR amplifications of bacterial 16S rRNA genes. It is demonstrated that these two alternative methods could be applied to a wide variety of environmental samples.
基金This subject was supported by the grants from the National Natural Science Foundation of China （ No. 30371754）.
文摘To observe the effect of Gardenia extract ZG on the adsorption quantity of herpes simplex virus type 1 （HSV-1） so as to explore the mechanism of its antiviral activity, fluorescein isothiocyanate （FITC） was used as the fluorescent probe to label viruses and heparin sodium was used as control. Meanwhile, the effect of Gardenia extract ZG on the adsorption quantity on the surface of Hep-2 cells was determined by flow cytometry. It was demonstrated that adsorption of HSV-1 on the surface of Hep-2 cells exhibited the character of saturation and specificity and heparin sodium could prevent attachment of viruses on these ceils. These results are in accord with those reported previously. It was also proved that the manner of drug-use prior to adsorption or simultaneous use of drug and adsorption was better than adsorption prior to drug-use, and the inhibition rates of the former and latter manner were 84.76% and 82.92% respectively. Three manners of drug-use with Gardenia extract ZG were all effective to reduce the adsorption quantity of viruses, especially the manner of simultaneous use of drug and adsorption with an adsorption inhibition rate of 68.46%. From the above observation, it is apparent that the mechanism of anti-viral activity of Gardenia extract ZG may be via several steps involved in the HSV-1 adsorption.
文摘Plant peroxidase extracted from Coccinia grandis Lin. （Ivy gourd） was purified from crude extract by ammonium sulfate precipitation, ion-exchange chromatography, and size exclusion chromatography. The purified enzyme preparation exhibited a specific activity of 6106.63 p.mol.minl.mg protein1, while purification fold and yield were 17.45 and 34.70%, respectively. The purified peroxidase was homogenous as judged by native and sodium dodecyl sulfate （SDS） polyacrylamide gel electrophoresis. The molecular weight as determined by gel filtration and SDS-polyacrylamide gel electrophoresis was 45 kD, which suggested that the purified peroxidase contained only one subunit. The apparent Km and Vmax values of the enzyme against phenol were 93 p.M and 561 μmol.min^-1.mg protein^-1, respectively. The temperature and pH optimum for purified peroxidase were 45℃ and pH 6.0, respective. However, it was stable at 30-60℃ and pH 4.0-8.0. The presence of metal ions such as Cu2＋ and Ca2＋ enhanced peroxidase activity. On the other hand, Cr3＋ and Hg2＋ strongly inhibited the enzyme activity at 500 p.M. Sodium dodecyl sulfate reduced a half of peroxidase activity at approximately 3 mM. Ivy gourd was stability in the presence of each urea concentrations. The affinity of the enzyme with different substrates showed as the highest relative activities on gallic acid followed by catechin, ascorbic acid and caffeic acid, respectively.
基金the National Natural Science Foundation of China Grant(No.81073023)the Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions(No.ysxk-2010)2013 Program sponsored for scientific innovation research of college graduate in Jiangsu province(No.CXZZ13_0631)
文摘OBJECTIVE: To study the antiviral activities of clemastanin B(CB), epigoitrin, phenylpropanoids portion(PEP) and the mixture of phenylpropanoids, alkaloids and organic acid fractions(PEP+ALK+OA)from Banlangen(Radix Isatidis).METHODS: The experiment consisted of four parts:therapeutic action, prophylaxsis action, inhibition of virus attachment, and direct virucidal action. Cytopathic effect(CPE) and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium(MTT) were used to assess antiviral activity.RESULTS: CB, epigoitrin, PEP and PEP + ALK + OA fractions from Banlangen(Radix Isatidis) extract significantly increased the viability of MDCK cells pre-infected with the virus compared with the virus control group in all the dilutions(P < 0.01). Pretreated with either pure compounds or chemical frac-tions of Banlangen(Radix Isatidis) extract in all the dilutions significantly improved the viability of MDCK cells(P < 0.01). The inhibition of virus absorption to the host cells by CB, epigoitrin and PEP was in a dose dependent manner.CONCLUSION: CB, epigoitrin, PEP and PEP+ALK+OA exert their anti-influenza activity by inhibiting the virus multiplication, prophylaxsis and blocking the virus attachment. The primary mode of action of PEP and PEP + ALK + OA is the inhibition of virus replication. The inhibitory effects on virus attachment and multiplication are the main modes for epigoitrin.
