Poly‐3‐hydroxybutyrate production by Azotobacter vinelandii strains in batch cultures at different oxygen transfer rates

BACKGROUND Poly-3-hydroxybutyrate (PHB) is a polyester that can be accumulated by Azotobacter vinelandii. This polymer is a biodegradable thermoplastic material used for producing plastics for packaging, biobased films, and biocompatible implants. RESULTS PHB produced by wild-type A. vinelandii and a mutant OP strain were evaluated at different agitation rates. The oxygen transfer rate (OTR) evolution was characterized in cultures grown at 300, 400, 500 and 600 rpm, showing for the first time the OTR evolution in cultures of mutant OP. Under the conditions evaluated, the cultures were limited by oxygen. In the cultures grown with the OP strain, lower OTR and specific oxygen uptake rate were obtained, indicating a lower necessity for oxygen for growth in this mutant. A higher amount of PHB can be produced by decreasing the OTR. A maximum PHB of 79% on dry weight was observed in the cultures with the wild-type strain, whereas the highest PHB productivity (0.18 g L−1 h−1) was obtained in cultures of A. vinelandii OP conducted at 600 rpm. CONCLUSION The amount of PHB produced during the period of oxygen limitation can be controlled by the OTR, opening the possibility to evaluate this parameter as a criterion for scaling-up PHB production by A. vinelandii. © 2015 Society of Chemical Industry

[1]  G. A. Ritchie,et al.  The role of oxygen limitation in the formation of poly- -hydroxybutyrate during batch and continuous culture of Azotobacter beijerinckii. , 1972, The Biochemical journal.

[2]  E. Heinzle,et al.  Oxygen supply strongly influences metabolic fluxes, the production of poly(3-hydroxybutyrate) and alginate, and the degree of acetylation of alginate in Azotobacter vinelandii , 2013 .

[3]  G. Espín,et al.  flhDC, but not fleQ, regulates flagella biogenesis in Azotobacter vinelandii, and is under AlgU and CydR negative control , 2008, Microbiology.

[4]  L. Savenkova,et al.  Effect of phosphate supply and aeration on poly-β-hydroxybutyrate production in Azotobacter chroococcum , 1999 .

[5]  A. Anderson,et al.  Occurrence, metabolism, metabolic role, and industrial uses of bacterial polyhydroxyalkanoates. , 1990, Microbiological reviews.

[6]  L. Savenkova,et al.  Effect of iron and aeration on superoxide dismutase and catalase activity of PHB-producing Azotobacter chroococcum , 2009 .

[7]  W. Page,et al.  Alginate formation in Azotobacter vinelandii UWD during stationary phase and the turnover of poly-beta-hydroxybutyrate. , 2001, Microbiology.

[8]  A. Díaz‐Barrera,et al.  Alginate production and alg8 gene expression by Azotobacter vinelandii in continuous cultures , 2012, Journal of Industrial Microbiology & Biotechnology.

[9]  E. Galindo,et al.  High production of poly-β-hydroxybutyrate (PHB) by an Azotobacter vinelandii mutant altered in PHB regulation using a fed-batch fermentation process , 2014 .

[10]  A. Romo‐Uribe,et al.  Biosynthesis of poly-β-hydroxybutyrate (PHB) with a high molecular mass by a mutant strain of Azotobacter vinelandii (OPN) , 2013, Annals of Microbiology.

[11]  A. Secchi,et al.  Influence of oxygen transfer rate on the accumulation of poly(3-hydroxybutyrate) by Bacillus megaterium , 2013 .

[12]  F. García-Ochoa,et al.  The effect of hydrodynamic stress on the growth of Xanthomonas campestris cultures in a stirred and sparged tank bioreactor , 2013, Bioprocess and Biosystems Engineering.

[13]  A. Zeng,et al.  Effect of Oxygen on Formation and Structure ofAzotobacter vinelandii Alginate and Its Role in Protecting Nitrogenase , 2000, Applied and Environmental Microbiology.

[14]  A. Moir,et al.  Biosynthesis of poly-beta-hydroxybutyrate (PHB) is controlled by CydR (Fnr) in the obligate aerobe Azotobacter vinelandii. , 2001, FEMS microbiology letters.

[15]  D. Emerich,et al.  Analysis of Poly-β-Hydroxybutyrate in Rhizobium japonicum Bacteroids by Ion-Exclusion High-Pressure Liquid Chromatography and UV Detection , 1983 .

[16]  E. Galindo,et al.  Role of oxygen in the polymerization and de-polymerization of alginate produced by Azotobacter vinelandii , 2015 .

[17]  E. Galindo,et al.  The oxygen transfer rate influences the molecular mass of the alginate produced by Azotobacter vinelandii , 2007, Applied Microbiology and Biotechnology.

[18]  W. Babel,et al.  Physiology, regulation, and limits of the synthesis of poly(3HB). , 2001, Advances in biochemical engineering/biotechnology.

[19]  Tajalli Keshavarz,et al.  Production of polyhydroxyalkanoates: the future green materials of choice , 2010 .

[20]  G. Sawers,et al.  Regulation of Cytochrome bd Expression in the Obligate Aerobe Azotobacter vinelandii by CydR (Fnr) , 2000, The Journal of Biological Chemistry.

[21]  W. Page,et al.  Control of polyhydroxyalkanoate synthesis in Azotobacter vinelandii strain UWD , 1994 .

[22]  C. Williams,et al.  Bacterial synthesis of biodegradable polyhydroxyalkanoates , 2007, Journal of applied microbiology.

[23]  P. Prasertsan,et al.  Effect of aeration and agitation rates and scale- up on oxygen transfer coefficient, kLa in exopolysaccharide production from Enterobacter cloacae WD7 , 2006 .

[24]  Jochen Büchs,et al.  Evolution of the specific power consumption and oxygen transfer rate in alginate-producing cultures of Azotobacter vinelandii conducted in shake flasks , 2007 .

[25]  F. García-Ochoa,et al.  Specific oxygen uptake rate as indicator of cell response of Rhodococcus erythropolis cultures to shear effects , 2015 .

[26]  V. Skulachev,et al.  Concept of Aging as a Result of Slow Programmed Poisoning of an Organism with Mitochondrial Reactive Oxygen Species , 2013 .

[27]  V. Deretic,et al.  Characterization of the genes coding for the putative sigma factor AlgU and its regulators MucA, MucB, MucC, and MucD in Azotobacter vinelandii and evaluation of their roles in alginate biosynthesis , 1996, Journal of bacteriology.

[28]  J. Büchs,et al.  Device for sterile online measurement of the oxygen transfer rate in shaking flasks. , 2001, Biochemical engineering journal.

[29]  G. L. Miller Use of Dinitrosalicylic Acid Reagent for Determination of Reducing Sugar , 1959 .

[30]  J. Oelze,et al.  Respiratory protection of nitrogenase in Azotobacter species: is a widely held hypothesis unequivocally supported by experimental evidence? , 2000, FEMS microbiology reviews.

[31]  F. García-Ochoa,et al.  Bioreactor scale-up and oxygen transfer rate in microbial processes: an overview. , 2009, Biotechnology advances.

[32]  J. Büchs,et al.  Characterisation of the gas-liquid mass transfer in shaking bioreactors. , 2001, Biochemical engineering journal.