1,3-Propanediol production by Escherichia coli using genes from Citrobacter freundii atcc 8090.

Compared with chemical synthesis, fermentation has the advantage of mass production at low cost, and has been used in the production of various industrial chemicals. As a valuable organic compound, 1,3-propanediol (1,3-PDO) has numerous applications in the production of polymers, lubricants, cosmetics and medicines. Here, conversion of glycerol (a renewable substrate and waste from biodiesel production) to 1,3-PDO by E. coli bacterial strain carrying altered glycerol metabolic pathway was investigated. Two gene constructs containing the 1,3-PDO operon from Citrobacter freundii (pCF1 and pCF2) were used to transform the bacteria. The pCF1 gene expression construct contained dhaBCE genes encoding the three subunits of glycerol dehydratase, dhaF encoding the large subunit of the glycerol dehydratase reactivation factor and dhaG encoding the small subunit of the glycerol dehydratase reactivating factor. The pCF2 gene expression construct contained the dhaT gene encoding the 1,3-propanediol dehydrogenase. Expression of the genes cloned in the above constructs was under regulation of the T7lac promoter. RT-PCR, SDS-PAGE analyses and functional tests confirmed that 1,3-PDO synthesis pathway genes were expressed at the RNA and protein levels, and worked flawlessly in the heterologous host. In a batch flask culture, in a short time applied just to identify the 1,3-PDO in a preliminary study, the recombinant E. coli bacteria produced 1.53 g/L of 1,3-PDO, using 21.2 g/L of glycerol in 72 h. In the Sartorius Biostat B Plus reactor, they produced 11.7 g/L of 1,3-PDO using 24.2 g/L of glycerol, attaining an efficiency of 0.58 [mol1,3-PDO/molglycerol].

Conversion of glycerol to 1,3-propanediol by Citrobacter freundii and Hafnia alvei - newly isolated strains from the Enterobacteriaceae.

In this study, nearly 4000 bacterial strains from the family of Enterobacteriaceae isolated from different environments were screened for ability to convert glycerol to 1,3-propanediol (1,3-PD). The aim of the research was to isolate 1,3-PD producers from the natural environment, identify and characterize the best isolates. Three selective media were tested to usefulness in the isolation of bacteria from the family Enterobacteriaceae. Only, 28% of examined isolates could synthesize 1,3-PD from glycerol. 1,3-PD producing bacteria were identified by API 20E tests and 16S rRNA sequences to be Klebsiella pneumoniae, Klebsiella oxytoca, Citrobacter freundii and Hafnia alvei. It is the first time, when the fermentation glycerol to 1,3-PD by H. alvei was investigated. The selected strains (C. freundii AD119 and H. alvei AD27) were analyzed on a bioreactor scale under constant pH value 7.0 at temperature of 30°C and 37°C. After 40h in batch fermentation, H. alvei AD27 produced 11.3g/L of 1,3-PD at 37°C. For C. freundii AD119, the best results were obtained at temperature of 30°C. After 24h of fermentation, the 1,3-PD concentration reached above 23 g/L of 1,3-PD.

Application of fast liquid chromatography for antioxidants analysis.

BACKGROUND: An intensive development of the Fast Liquid Chromatography (FLC) has been recently observed. It makes possible to reduce time analysis and improve resolution as well as sensitivity. The aim of this study was to separate the chosen antioxidants optimization using the FLC method. MATERIAL AND METHODS: The three various procedures for antioxidants analysis were compared. Mobile phases containing aqueous solution of formic acid, acetic acid, acetonitrile, and methanol were tested. Limit of detection (LOD), limit of quantification (LOQ), linearity and repeatability of each procedures were determined. RESULTS: Developed procedure enabled to separate all analytes and allowed to get low LOD levels and good repeatability. This procedure was used for antioxidants analysis in buckwheat and buckwheat products. CONCLUSION: Fast Liquid Chromatography allows to reduce time analysis and obtain good validation parameters.

Vitamin B12 production from crude glycerol by Propionibacterium freudenreichii ssp. shermanii: optimization of medium composition through statistical experimental designs.

A two-step statistical experimental design was employed to optimize the medium for vitamin B(12) production from crude glycerol by Propionibacterium freudenreichii ssp. shermanii. In the first step, using Plackett-Burman design, five of 13 tested medium components (calcium pantothenate, NaH(2)PO(4)·2H(2)O, casein hydrolysate, glycerol and FeSO(4)·7H(2)O) were identified as factors having significant influence on vitamin production. In the second step, a central composite design was used to optimize levels of medium components selected in the first step. Valid statistical models describing the influence of significant factors on vitamin B(12) production were established for each optimization phase. The optimized medium provided a 93% increase in final vitamin concentration compared to the original medium.