The distribution of ^(125)I recombinant E. coli L-asparaginase in tissues ororgans and the excretion in urine, feces and bile were studied with in vivo radioactive tracertechnique. The amount of radioactivity excreted...The distribution of ^(125)I recombinant E. coli L-asparaginase in tissues ororgans and the excretion in urine, feces and bile were studied with in vivo radioactive tracertechnique. The amount of radioactivity excreted in urine, feces and bile within 24 h afterintravenous administration of ^(125)I recombinant E. col L-asparaginase to rats was 68.95% ,4.44%and 5.36% of the dose respectively. ^(125)I recombinant E. coli L-asparaginase in plasma samples wasdetermined. The levels of structural intact molecule in plasma samples were evaluated by SDS-PAGEand bio-imaging analyzer system. Pharmacokinetic parameters were assessed with a model-dependentmethod. The concentration-time curves of recombinant E. coli L-asparaginase after intravenousinjection at 1 250 IU·kg^(-1), 2 500, IU·kg^(-1), 5 000 IU·kg^(-1) to rats were consistent withthe two-compartment model. The first and terminal elimination t_(1/2) were 0.52 ~ 0.63 h and 2.39 ~2.76 h respectively. AUC was linearly related to the doses. The results of distribution in tissuesor organs and excretion in urine suggested that the metabolites of the enzyme were cleared bymechanisms of urinary excretion. Pharmacokinetics parameters of recombinant E. coli L- asparaginasein rats are warranted for the design of future clinical trials.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
3,4-Dihydroxy-2-butanone 4-phosphate （DHBP） and GTP are the precursors for riboflavin biosynthesis. In this research, improving the precursor supply for riboflavin production was attempted by overexpressing ribB and...3,4-Dihydroxy-2-butanone 4-phosphate （DHBP） and GTP are the precursors for riboflavin biosynthesis. In this research, improving the precursor supply for riboflavin production was attempted by overexpressing ribB and engineering purine pathway in a riboflavin-producing Escherichia colt strain. Initially, ribB gene was overexpressed to increase the flux from ribulose 5-phosphate （Ru-5-P） to DHBP. Then ndk and grnk genes were overexpressed to enhance GTP supply. Subsequently, a R419L mutation was introduced into purA to reduce the flux from IMP to AMP. Finally, co-overexpression of mutant purF and prs genes further increased riboflavin production. The final strain RF18S produced 387.6 mg riboflavin · L-1 with a yield of 44.8 mg riboflavin per gram glucose in shake-flask fermentations. The final titer and yield were 72.2% and 55.6% higher than those of RF01S, respectively. It was concluded that simultaneously engineering the DHBP synthase and GTP biosynthetic pathway by rational metabolic engineering can efficiently boost riboflavin production in E. coll.展开更多
基金ProjectsupportedbytheNationalNinth FivePlanKeyProjectFoundation No 96 90 2 0 1 2 5
文摘The distribution of ^(125)I recombinant E. coli L-asparaginase in tissues ororgans and the excretion in urine, feces and bile were studied with in vivo radioactive tracertechnique. The amount of radioactivity excreted in urine, feces and bile within 24 h afterintravenous administration of ^(125)I recombinant E. col L-asparaginase to rats was 68.95% ,4.44%and 5.36% of the dose respectively. ^(125)I recombinant E. coli L-asparaginase in plasma samples wasdetermined. The levels of structural intact molecule in plasma samples were evaluated by SDS-PAGEand bio-imaging analyzer system. Pharmacokinetic parameters were assessed with a model-dependentmethod. The concentration-time curves of recombinant E. coli L-asparaginase after intravenousinjection at 1 250 IU·kg^(-1), 2 500, IU·kg^(-1), 5 000 IU·kg^(-1) to rats were consistent withthe two-compartment model. The first and terminal elimination t_(1/2) were 0.52 ~ 0.63 h and 2.39 ~2.76 h respectively. AUC was linearly related to the doses. The results of distribution in tissuesor organs and excretion in urine suggested that the metabolites of the enzyme were cleared bymechanisms of urinary excretion. Pharmacokinetics parameters of recombinant E. coli L- asparaginasein rats are warranted for the design of future clinical trials.
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.
基金supported by National High-tech R&D Program of China [2012AA02A702, 2012AA022103]
文摘3,4-Dihydroxy-2-butanone 4-phosphate （DHBP） and GTP are the precursors for riboflavin biosynthesis. In this research, improving the precursor supply for riboflavin production was attempted by overexpressing ribB and engineering purine pathway in a riboflavin-producing Escherichia colt strain. Initially, ribB gene was overexpressed to increase the flux from ribulose 5-phosphate （Ru-5-P） to DHBP. Then ndk and grnk genes were overexpressed to enhance GTP supply. Subsequently, a R419L mutation was introduced into purA to reduce the flux from IMP to AMP. Finally, co-overexpression of mutant purF and prs genes further increased riboflavin production. The final strain RF18S produced 387.6 mg riboflavin · L-1 with a yield of 44.8 mg riboflavin per gram glucose in shake-flask fermentations. The final titer and yield were 72.2% and 55.6% higher than those of RF01S, respectively. It was concluded that simultaneously engineering the DHBP synthase and GTP biosynthetic pathway by rational metabolic engineering can efficiently boost riboflavin production in E. coll.
