Reeta Davis - Academia.edu (original) (raw)
Papers by Reeta Davis
Bioresource technology, 2015
This study demonstrates the use of a mannitol rich ensiled grass press juice (EGPJ) as a renewabl... more This study demonstrates the use of a mannitol rich ensiled grass press juice (EGPJ) as a renewable carbon substrate for polyhydroxyalkanoates (PHA) production in shaking flask experiments and fed-batch stirred tank reactor cultivations. Fed-batch cultivations of Burkholderia sacchari IPT101 using EGPJ as sole carbon source produced 44.5g/L CDW containing 33% polyhydroxybutyrate (PHB) in 36h, while Pseudomonas chlororaphis IMD555 produced a CDW of 37g/L containing 10% of medium chain length polyhydroxyalkanoates (mcl-PHA) in 34h. PHB and mcl-PHA extracted from B. sacchari IPT101 and P. chlororaphis IMD555, grown on EGPJ, had a molecular weight of 548kg/mol and 115.4kg/mol, respectively. While mcl-PHA can be produced from EGPJ, PHB production is more interesting as there is a 4-fold higher volumetric productivity compared to mcl-PHA.
Biotechnology Letters, 2015
Recent developments in biocatalysis, where implementation beyond the laboratory has been demonstr... more Recent developments in biocatalysis, where implementation beyond the laboratory has been demonstrated, are explored: the use of transglutaminases to modify foods, reduce allergenicity and produce advanced materials, lipases for biodiesel production, and transaminases for biochemical production. The availability and application of enzymes at pilot and larger scale opens up possibilities for further improvements of biocatalyst-based processes and the development of new processes. Enzyme production, stability, activity, re-use, and product retrieval are common challenges for biocatalytic processes. We explore recent advances in biocatalysis within the process chain, such as protein engineering, enzyme expression, and biocatalyst immobilization, in the context of these challenges.
Applied microbiology and biotechnology, 2014
A mathematically based fed-batch bioprocess demonstrated the suitability of using a relatively ch... more A mathematically based fed-batch bioprocess demonstrated the suitability of using a relatively cheap and renewable substrate (butyric acid) for Pseudomonas putida CA-3 high cell density cultivation. Butyric acid fine-tuned addition is critical to extend the fermentation run and avoid oxygen consumption while maximising the biomass volumetric productivity. A conservative submaximal growth rate (μ of 0.25 h(-1)) achieved 71.3 g L(-1) of biomass after 42 h of fed-batch growth. When a more ambitious feed rate was supplied in order to match a μ of 0.35 h(-1), the volumetric productivity was increased to 2.0 g L(-1) h(-1), corresponding to a run of 25 h and 50 g L(-1) of biomass. Both results represent the highest biomass and the best biomass volumetric productivity with butyrate as a sole carbon source. However, medium chain length polyhydroxyalkanoate (mcl-PHA) accumulation with butyrate grown cells is low (4 %). To achieve a higher mcl-PHA volumetric productivity, decanoate was supplie...
Biotechnology and bioengineering, Jan 13, 2014
High Cell Density (HCD) cultivation of bacteria is essential for the majority of industrial proce... more High Cell Density (HCD) cultivation of bacteria is essential for the majority of industrial processes to achieve high volumetric productivity (g L(-1) h(-1) ) of a bioproduct of interest. This study developed a fed batch bioprocess using glucose as sole carbon and energy source for the HCD of the well described biocatalyst Pseudomonas putida KT2440 without the supply of oxygen enriched air. Growth kinetics data from batch fermentations were used for building a bioprocess model and designing feeding strategies. An exponential followed by linearly increasing feeding strategy of glucose was found to be effective in maintaining biomass productivity while also delaying the onset of dissolved oxygen (supplied via compressed air) limitation. A final cell dry weight (CDW) of 102 g L(-1) was achieved in 33 h with a biomass productivity of 3.1 g L(-1) h(-1) which are the highest ever reported values for P. putida strains using glucose without the supply of pure oxygen or oxygen enriched air. ...
Indian Journal of Microbiology, 2009
Rhizobium meliloti produced a copolymer of short chain length polyhydroxyalkanoate (scl-PHA) on s... more Rhizobium meliloti produced a copolymer of short chain length polyhydroxyalkanoate (scl-PHA) on sucrose and rice bran oil as carbon substrates. Recombinant Escherichia coli (JC7623ABC1J4), bearing PHA synthesis genes, was used to synthesize short chain length-co-medium chain length PHA (scl-co-mcl-PHA) on glucose and decanoic acid. Fourier transform infrared spectroscopy (FTIR) spectra of the PHAs indicated strong characteristic bands at 1282, 1723, and 2934 cm -1 for scl-PHA and at 2933 and 2976 cm -1 for scl-co-mcl-PHA polymer. Differentiation of polyhydroxybutyrate (PHB) and polyhydroxybutyrate-co-hydroxyvalerate-P(HB-co-HV) copolymer was obseverd using FTIR, with absorption bands at 1723 and 1281 for PHB, and at 1738, 1134, 1215 cm -1 for HV-copolymer. The copolymers were analyzed by GC and 1 H NMR spectroscopy. Films of polymer blends of PHA produced by R. meliloti and recombinant E. coli were prepared using glycerol, polyethylene glycol, polyvinyl acetate, individually (1:1 ratio), to modify the mechanical properties of the fi lms and these fi lms were evaluated by FTIR and scanning electron microscopy.
Bioresource Technology, 2013
Antonie van Leeuwenhoek, 2008
Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the b-oxidation pathway with PHA biosy... more Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the b-oxidation pathway with PHA biosynthesis in Pseudomonas aeruginosa. The use of antisense technique and over-expression to delineate the role of two of these enzymes, PhaJ1 and PhaJ4 forms the basis of this study. It has been observed that P. aeruginosa recombinant with phaJ1 antisense construct, fed with different fatty acids, produces PHA with less hydroxy octanoate (7-11% reduction) and a proportionate increase in other monomer fractions, compared to that of the control. Recombinants bearing phaJ4 antisense construct are found to contain less hydroxy decanoate (10-11% reduction) and more or less equal amount of hydroxy octanoate, compared to that of the control. P. aeruginosa produced PHA with more hydroxy octanoate and decanoate (6-17% increase), respectively, when PhaJ1 and PhaJ4 have been overexpressed individually or along with PhaC1. PhaJ1 and PhaJ4 are found to be involved mainly in the production of hydroxy octanoyl CoA and hydroxy decanoyl CoA, respectively, in P. aeruginosa. The strongest accumulation of hydroxy octanoate and hydroxy decanoate has been observed along with hydroxy butyrate, in PHA, produced by E. coli, when PhaC1 has been co-expressed with PhaJ1 and PhaJ4, respectively. We have demonstrated, for the first time, the polymerization of hydroxy butyryl CoA monomers in recombinant E. coli by PhaC1 of P. aeruginosa.
Antonie van Leeuwenhoek, 2008
Expression of Pseudomonas aeruginosa genes PHA synthase1 (phaC1) and (R)-specific enoyl CoA hydra... more Expression of Pseudomonas aeruginosa genes PHA synthase1 (phaC1) and (R)-specific enoyl CoA hydratase1 (phaJ1) under a lacZ promoter was able to support production of a copolymer of Polyhydroxybutyrate (PHB) and medium chain length polyhydoxyalkanoates (mcl-PHA) in Escherichia coli. In order to improve the yield and quality of PHA, plasmid bearing the above genes was introduced into E. coli JC7623, harboring integrated b-ketothiolase (phaA) and NADPH dependent-acetoacetyl CoA reductase (phaB) genes from a Bacillus sp. also driven by a lacZ promoter. The recombinant E. coli (JC7623ABC1J1) grown on various fatty acids along with glucose was found to produce 28-34% cellular dry weight of PHA. Gas chromatography and 1 H Nuclear Magnetic Resonance analysis of the polymer confirmed the ability of the strain to produce PHB-co-Hydroxy valerate (HV)-co-mcl-PHA copolymers. The ratio of short chain length (scl) to mcl-PHA varied from 78:22 to 18:82. Addition of acrylic acid, an inhibitor of b-oxidation resulted in improved production (3-11% increase) of PHA copolymer. The combined use of enzymes from Bacillus sp. and Pseudomonas sp. for the production of scl-co-mcl PHA in E. coli is a novel approach and is being reported for the first time.
Applied Microbiology and Biotechnology, 2014
A two step biological process for the conversion of grass biomass to the biodegradable polymer me... more A two step biological process for the conversion of grass biomass to the biodegradable polymer medium chain length polyhydroxyalkanoate (mcl-PHA) was achieved through the use of anaerobic and aerobic microbial processes. Anaerobic digestion (mixed culture) of ensiled grass was achieved with a recirculated leach bed bioreactor resulting in the production of a leachate, containing 15.3 g/l of volatile fatty acids (VFAs) ranging from acetic to valeric acid with butyric acid predominating (12.8 g/l). The VFA mixture was concentrated to 732.5 g/l with a 93.3 % yield of butyric acid (643.9 g/l). Three individual Pseudomonas putida strains, KT2440, CA-3 and GO16 (single pure cultures), differed in their ability to grow and accumulate PHA from VFAs. P. putida CA-3 achieved the highest biomass and PHA on average with individual fatty acids, exhibited the greatest tolerance to higher concentrations of butyric acid (up to 40 mM) compared to the other strains and exhibited a maximum growth rate (μ MAX =0.45 h −1 ). Based on these observations P. putida CA-3 was chosen as the test strain with the concentrated VFA mixture derived from the AD leachate. P. putida CA-3 achieved 1.56 g of biomass/l and accumulated 39 % of the cell dry weight as PHA (nitrogen limitation) in shake flasks. The PHA was composed predominantly of 3-hydroxydecanoic acid (>65 mol%).
Bioresource technology, 2015
This study demonstrates the use of a mannitol rich ensiled grass press juice (EGPJ) as a renewabl... more This study demonstrates the use of a mannitol rich ensiled grass press juice (EGPJ) as a renewable carbon substrate for polyhydroxyalkanoates (PHA) production in shaking flask experiments and fed-batch stirred tank reactor cultivations. Fed-batch cultivations of Burkholderia sacchari IPT101 using EGPJ as sole carbon source produced 44.5g/L CDW containing 33% polyhydroxybutyrate (PHB) in 36h, while Pseudomonas chlororaphis IMD555 produced a CDW of 37g/L containing 10% of medium chain length polyhydroxyalkanoates (mcl-PHA) in 34h. PHB and mcl-PHA extracted from B. sacchari IPT101 and P. chlororaphis IMD555, grown on EGPJ, had a molecular weight of 548kg/mol and 115.4kg/mol, respectively. While mcl-PHA can be produced from EGPJ, PHB production is more interesting as there is a 4-fold higher volumetric productivity compared to mcl-PHA.
Biotechnology Letters, 2015
Recent developments in biocatalysis, where implementation beyond the laboratory has been demonstr... more Recent developments in biocatalysis, where implementation beyond the laboratory has been demonstrated, are explored: the use of transglutaminases to modify foods, reduce allergenicity and produce advanced materials, lipases for biodiesel production, and transaminases for biochemical production. The availability and application of enzymes at pilot and larger scale opens up possibilities for further improvements of biocatalyst-based processes and the development of new processes. Enzyme production, stability, activity, re-use, and product retrieval are common challenges for biocatalytic processes. We explore recent advances in biocatalysis within the process chain, such as protein engineering, enzyme expression, and biocatalyst immobilization, in the context of these challenges.
Applied microbiology and biotechnology, 2014
A mathematically based fed-batch bioprocess demonstrated the suitability of using a relatively ch... more A mathematically based fed-batch bioprocess demonstrated the suitability of using a relatively cheap and renewable substrate (butyric acid) for Pseudomonas putida CA-3 high cell density cultivation. Butyric acid fine-tuned addition is critical to extend the fermentation run and avoid oxygen consumption while maximising the biomass volumetric productivity. A conservative submaximal growth rate (μ of 0.25 h(-1)) achieved 71.3 g L(-1) of biomass after 42 h of fed-batch growth. When a more ambitious feed rate was supplied in order to match a μ of 0.35 h(-1), the volumetric productivity was increased to 2.0 g L(-1) h(-1), corresponding to a run of 25 h and 50 g L(-1) of biomass. Both results represent the highest biomass and the best biomass volumetric productivity with butyrate as a sole carbon source. However, medium chain length polyhydroxyalkanoate (mcl-PHA) accumulation with butyrate grown cells is low (4 %). To achieve a higher mcl-PHA volumetric productivity, decanoate was supplie...
Biotechnology and bioengineering, Jan 13, 2014
High Cell Density (HCD) cultivation of bacteria is essential for the majority of industrial proce... more High Cell Density (HCD) cultivation of bacteria is essential for the majority of industrial processes to achieve high volumetric productivity (g L(-1) h(-1) ) of a bioproduct of interest. This study developed a fed batch bioprocess using glucose as sole carbon and energy source for the HCD of the well described biocatalyst Pseudomonas putida KT2440 without the supply of oxygen enriched air. Growth kinetics data from batch fermentations were used for building a bioprocess model and designing feeding strategies. An exponential followed by linearly increasing feeding strategy of glucose was found to be effective in maintaining biomass productivity while also delaying the onset of dissolved oxygen (supplied via compressed air) limitation. A final cell dry weight (CDW) of 102 g L(-1) was achieved in 33 h with a biomass productivity of 3.1 g L(-1) h(-1) which are the highest ever reported values for P. putida strains using glucose without the supply of pure oxygen or oxygen enriched air. ...
Indian Journal of Microbiology, 2009
Rhizobium meliloti produced a copolymer of short chain length polyhydroxyalkanoate (scl-PHA) on s... more Rhizobium meliloti produced a copolymer of short chain length polyhydroxyalkanoate (scl-PHA) on sucrose and rice bran oil as carbon substrates. Recombinant Escherichia coli (JC7623ABC1J4), bearing PHA synthesis genes, was used to synthesize short chain length-co-medium chain length PHA (scl-co-mcl-PHA) on glucose and decanoic acid. Fourier transform infrared spectroscopy (FTIR) spectra of the PHAs indicated strong characteristic bands at 1282, 1723, and 2934 cm -1 for scl-PHA and at 2933 and 2976 cm -1 for scl-co-mcl-PHA polymer. Differentiation of polyhydroxybutyrate (PHB) and polyhydroxybutyrate-co-hydroxyvalerate-P(HB-co-HV) copolymer was obseverd using FTIR, with absorption bands at 1723 and 1281 for PHB, and at 1738, 1134, 1215 cm -1 for HV-copolymer. The copolymers were analyzed by GC and 1 H NMR spectroscopy. Films of polymer blends of PHA produced by R. meliloti and recombinant E. coli were prepared using glycerol, polyethylene glycol, polyvinyl acetate, individually (1:1 ratio), to modify the mechanical properties of the fi lms and these fi lms were evaluated by FTIR and scanning electron microscopy.
Bioresource Technology, 2013
Antonie van Leeuwenhoek, 2008
Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the b-oxidation pathway with PHA biosy... more Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the b-oxidation pathway with PHA biosynthesis in Pseudomonas aeruginosa. The use of antisense technique and over-expression to delineate the role of two of these enzymes, PhaJ1 and PhaJ4 forms the basis of this study. It has been observed that P. aeruginosa recombinant with phaJ1 antisense construct, fed with different fatty acids, produces PHA with less hydroxy octanoate (7-11% reduction) and a proportionate increase in other monomer fractions, compared to that of the control. Recombinants bearing phaJ4 antisense construct are found to contain less hydroxy decanoate (10-11% reduction) and more or less equal amount of hydroxy octanoate, compared to that of the control. P. aeruginosa produced PHA with more hydroxy octanoate and decanoate (6-17% increase), respectively, when PhaJ1 and PhaJ4 have been overexpressed individually or along with PhaC1. PhaJ1 and PhaJ4 are found to be involved mainly in the production of hydroxy octanoyl CoA and hydroxy decanoyl CoA, respectively, in P. aeruginosa. The strongest accumulation of hydroxy octanoate and hydroxy decanoate has been observed along with hydroxy butyrate, in PHA, produced by E. coli, when PhaC1 has been co-expressed with PhaJ1 and PhaJ4, respectively. We have demonstrated, for the first time, the polymerization of hydroxy butyryl CoA monomers in recombinant E. coli by PhaC1 of P. aeruginosa.
Antonie van Leeuwenhoek, 2008
Expression of Pseudomonas aeruginosa genes PHA synthase1 (phaC1) and (R)-specific enoyl CoA hydra... more Expression of Pseudomonas aeruginosa genes PHA synthase1 (phaC1) and (R)-specific enoyl CoA hydratase1 (phaJ1) under a lacZ promoter was able to support production of a copolymer of Polyhydroxybutyrate (PHB) and medium chain length polyhydoxyalkanoates (mcl-PHA) in Escherichia coli. In order to improve the yield and quality of PHA, plasmid bearing the above genes was introduced into E. coli JC7623, harboring integrated b-ketothiolase (phaA) and NADPH dependent-acetoacetyl CoA reductase (phaB) genes from a Bacillus sp. also driven by a lacZ promoter. The recombinant E. coli (JC7623ABC1J1) grown on various fatty acids along with glucose was found to produce 28-34% cellular dry weight of PHA. Gas chromatography and 1 H Nuclear Magnetic Resonance analysis of the polymer confirmed the ability of the strain to produce PHB-co-Hydroxy valerate (HV)-co-mcl-PHA copolymers. The ratio of short chain length (scl) to mcl-PHA varied from 78:22 to 18:82. Addition of acrylic acid, an inhibitor of b-oxidation resulted in improved production (3-11% increase) of PHA copolymer. The combined use of enzymes from Bacillus sp. and Pseudomonas sp. for the production of scl-co-mcl PHA in E. coli is a novel approach and is being reported for the first time.
Applied Microbiology and Biotechnology, 2014
A two step biological process for the conversion of grass biomass to the biodegradable polymer me... more A two step biological process for the conversion of grass biomass to the biodegradable polymer medium chain length polyhydroxyalkanoate (mcl-PHA) was achieved through the use of anaerobic and aerobic microbial processes. Anaerobic digestion (mixed culture) of ensiled grass was achieved with a recirculated leach bed bioreactor resulting in the production of a leachate, containing 15.3 g/l of volatile fatty acids (VFAs) ranging from acetic to valeric acid with butyric acid predominating (12.8 g/l). The VFA mixture was concentrated to 732.5 g/l with a 93.3 % yield of butyric acid (643.9 g/l). Three individual Pseudomonas putida strains, KT2440, CA-3 and GO16 (single pure cultures), differed in their ability to grow and accumulate PHA from VFAs. P. putida CA-3 achieved the highest biomass and PHA on average with individual fatty acids, exhibited the greatest tolerance to higher concentrations of butyric acid (up to 40 mM) compared to the other strains and exhibited a maximum growth rate (μ MAX =0.45 h −1 ). Based on these observations P. putida CA-3 was chosen as the test strain with the concentrated VFA mixture derived from the AD leachate. P. putida CA-3 achieved 1.56 g of biomass/l and accumulated 39 % of the cell dry weight as PHA (nitrogen limitation) in shake flasks. The PHA was composed predominantly of 3-hydroxydecanoic acid (>65 mol%).