scholarly journals Technical-Economical Approach for PHB Production by Ralstonia Eutropha Strain Using Concentrated Vinasse as Carbon Source and Other Biotechnological Applications

2021 ◽  
Author(s):  
Manuella Silverio ◽  
Rosane Piccoli ◽  
João Reis ◽  
José Gregorio Gomez ◽  
Antonio Baptista
Keyword(s):  
Carbon Source ◽  
Ralstonia Eutropha ◽  
Culture Media ◽  
Dry Weight ◽  
Global Market ◽  
Cellular Growth ◽  
Production Costs ◽  
E Coli ◽  
Phb Production ◽  
Threonine Production

Abstract The Brazilian ethanol industry is one of the most important in the global market, however these important industrial activities have been generating significant amounts of vinasse and its management has become costly for distilleries. In this study, the aim was to evaluate concentrated and in natura vinasse as basal culture media for biotechnological processes. Different bacteria and processes were assessed: L-threonine production by E. coli THR14, with glucose as carbon source; PHB production by halophilic strain Halomonas sp. HG03, with sucrose as carbon source; and PHB biosynthesis by R. eutropha L359PCJ, which used glycerol from vinasse as carbon source. Strains were evaluated firstly in shake flasks cultivations using vinasse-based media. E. coli THR14 had no statistical difference for biomass and L-threonine concentrations among control and vinasse-based treatments (up to 50% v v-1 of in natura vinasse). Halomonas sp. HG03 and R. eutropha L359PCJ were cultivated in mineral media diluted by in natura (50% and 75% v v-1) and concentrated (50% and 75% v v-1) vinasses. Higher vinasse concentrations resulted in higher cellular growth rather than PHB accumulation for both bacteria. In vinasse-based treatments, Halomonas sp. HG03 had PHB content between 19.6 – 75.2% and R. eutropha L359PCJ, 48.4 – 68.5%. 50% (v v-1) of concentrated vinasse was the most attractive condition for PHB production by both bacteria. Further experiments in CSTR bioreactors used this nutritional condition and R. eutropha L359PCJ had PHB content of 66.3%, concentrations of residual cell dry weight (rCDW) = 9.4 g L-1 and PHB = 18.6 g L-1, with YX/S = 0.16 g gGLYCEROL-1, YP/S = 0.32 g gGLYCEROL-1 and 0.25 gPHB Lh-1. Halomonas sp. HG03 had PHB content of 45.7%, rCDW = 9.8 g L-1, PHB = 8.3 g L-1 and YX/S = 0.18 g gSUCROSE-1, YP/S = 0.16 g gSUCROSE-1 and 0.12 gPHB Lh-1. Finally, cost reductions of PHB production by R. eutropha L359PCJ with concentrated vinasse-based medium were evaluated in silico by using SuperPro Designer. As a partial source of glycerol and other nutrients for PHB production by R. eutropha L359PCJ, vinasse reduced overall production costs by 13%. Simulated processes that used concentrated vinasse-based media combined with improvements of PHB productivity and higher cellular densities had production costs between US$ 3.9 – 7.5/kgPHB and 2.6 – 7.3 years of payback time.

scholarly journals Electromagnetic Fields (0.04 to 0.39) mT effect on cellular growth cycles of Saccharomyces cerevisiae wine strains

2019 ◽  
Vol 17 (2) ◽  
pp. 196
Author(s):  
Eliseo Amado-González ◽  
Alveiro Álvarez Ovallos ◽  
Alfonso Quijano Parra
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Electromagnetic Fields ◽  
Biomass Production ◽  
Exposure Time ◽  
Culture Media ◽  
Dry Weight ◽  
Cellular Growth ◽  
Biomass Growth ◽  
Yeast Saccharomyces Cerevisiae ◽  
Growth Cycles

Low frecuency electromagnetic fields effect (EMF) on growth cycles of yeast Saccharomyces cerevisiae wine strains Rv1 and Rhône were studied.  A cylindrical coil induced magnetic fields with inductions up to 0,39 mT. Exposure time to EMF varied between (1 – 10) min at 30 °C.  The biomass growth were monitored in the reactor culture media (yeast extract + by measurement optical density from (0 to 32) h. The biomass was found by dry weight. After yeast expose to the different EMF, the number of growth cycles decreased from 4 cycles to 2 or 1. However, the biomass production increased almost 50 %.  The best biomass production was found at 0.39 mT and 10 min exposure time.  Keywords: Electromagnetic fields, Saccharomyces cerevisiae, biomass production, RV1

scholarly journals Impact of Ralstonia eutropha's Poly(3-Hydroxybutyrate) (PHB) Depolymerases and Phasins on PHB Storage in Recombinant Escherichia coli

2014 ◽  
Vol 80 (24) ◽  
pp. 7702-7709 ◽  
Author(s):  
Jessica Eggers ◽  
Alexander Steinbüchel
Keyword(s):  
Escherichia Coli ◽  
Ralstonia Eutropha ◽  
Model Organism ◽  
Dry Weight ◽  
Growth Behavior ◽  
Carbon Limitation ◽  
Content Type ◽  
Phb Depolymerase ◽  
E Coli ◽  
The Impact

ABSTRACTThe model organism for polyhydroxybutyrate (PHB) biosynthesis,Ralstonia eutrophaH16, possesses multiple isoenzymes of granules coating phasins as well as of PHB depolymerases, which degrade accumulated PHB under conditions of carbon limitation. In this study, recombinantEscherichia coliBL21(DE3) strains were used to study the impact of selected PHB depolymerases ofR. eutrophaH16 on the growth behavior and on the amount of accumulated PHB in the absence or presence of phasins. For this purpose, 20 recombinantE. coliBL21(DE3) strains were constructed, which harbored a plasmid carrying thephaCABoperon fromR. eutrophaH16 to ensure PHB synthesis and a second plasmid carrying different combinations of the genes encoding a phasin and a PHB depolymerase fromR. eutrophaH16. It is shown in this study that the growth behavior of the respective recombinantE. colistrains was barely affected by the overexpression of the phasin and PHB depolymerase genes. However, the impact on the PHB contents was significantly greater. The strains expressing the genes of the PHB depolymerases PhaZ1, PhaZ2, PhaZ3, and PhaZ7 showed 35% to 94% lower PHB contents after 30 h of cultivation than the control strain. The strain harboringphaZ7reached by far the lowest content of accumulated PHB (only 2.0% [wt/wt] PHB of cell dry weight). Furthermore, coexpression of phasins in addition to the PHB depolymerases influenced the amount of PHB stored in cells of the respective strains. It was shown that the phasins PhaP1, PhaP2, and PhaP4 are not substitutable without an impact on the amount of stored PHB. In particular, the phasins PhaP2 and PhaP4 seemed to limit the degradation of PHB by the PHB depolymerases PhaZ2, PhaZ3, and PhaZ7, whereas almost no influence of the different phasins was observed ifphaZ1was coexpressed. This study represents an extensive analysis of the impact of PHB depolymerases and phasins on PHB accumulation and provides a deeper insight into the complex interplay of these enzymes.

scholarly journals Studies on the lipid content and phosphate requirement of glucose- and acetate-grown Escherichia coli

1968 ◽  
Vol 110 (4) ◽  
pp. 775-781 ◽  
Author(s):  
A. P. Damoglou ◽  
E A Dawes
Keyword(s):  
Escherichia Coli ◽  
Energy Source ◽  
Carbon Source ◽  
Lipid Content ◽  
Dry Weight ◽  
The Other ◽  
Ammonium Salts ◽  
Phosphate Limitation ◽  
E Coli ◽  
High Phosphate

1. The phosphate requirement, i.e. the concentration of inorganic orthophosphate that just ceases to be limiting for growth, of Escherichia coli N.C.T.C. 5928 was determined for growth in ammonium–salts media containing glucose or acetate as the carbon and energy source, and compared with that of six other strains of E. coli. 2. The phosphate requirement for E. coli N.C.T.C. 5928 growing on acetate was about ten times that for growth on glucose, but this difference was not observed with any of the other strains. 3. After about 40 generations' growth on acetate with phosphate limitation in a chemostat, the phosphate requirement of the cells gradually decreased until it was equivalent to that of the glucose-grown organism; a single passage through glucose batch culture sufficed to restore the original high phosphate requirement, indicating a permeability phenomenon. 4. The lipid content of E. coli N.C.T.C. 5928 grown on glucose or acetate was measured isotopically by fractionation of cells grown on inorganic [32P]orthophosphate and gravimetrically after extraction from the cells by three different methods; change of carbon source from glucose to acetate did not affect the lipid content, which remained constant at 8–9% of the bacterial dry weight.

scholarly journals Study of pH Changes in Media during Bacterial Growth of Several Environmental Strains

Proceedings ◽  
2018 ◽  
Vol 2 (20) ◽  
pp. 1297 ◽  
Author(s):  
Rubén Sánchez-Clemente ◽  
María Isabel Igeño ◽  
Ana G. Población ◽  
María Isabel Guijo ◽  
Faustino Merchán ◽  
...  
Keyword(s):  
Carbon Source ◽  
Pseudomonas Putida ◽  
Bacterial Growth ◽  
Extracellular Ph ◽  
Cellular Growth ◽  
Exponential Growth Phase ◽  
Extracellular Medium ◽  
Pseudomonas Putida Kt2440 ◽  
E Coli ◽  
Initial Ph

The effect of pH on bacterial cell-growth and the evolution of extracellular pH triggered by bacterial growth has been monitored for three bacterial strains, Escherichia coli ATCC 25922 and Pseudomonas putida KT2440 as reference strains, and Pseudomonas pseudoalcaligenes CECT 5344 because of its capacity to assimilate cyanide as the sole nitrogen source under alkaline conditions. In a first instance, the influence of the initial pH in the growth curve has been texted in LB-medium adjusted to pH 6, 7 and 8, for E. coli and P. putida, and 7.5, 8.25 and 9 for P. pseudoalcaligenes. Although the initial pH were different, the pH of the extracellular medium at the end of the stationary phase converged to a certain pH that is specific for each bacterium. Similar experiments were carried out in minimal medium with glucose as the carbon source. In this case, the pHs of the culture of both Pseudomonadaceae strains were almost constant, whereas it suddenly dropped during the exponential growth phase of E. coli. When the initial pH was 6 the extracellular pH fell sharply to 4.5, which irreversibly prevented further cellular growth. Nevertheless, at higher initial pH values subsequent cellular growth of E. coli restored the medium to the initial pHs values. Finally, in all cases the evolution of the pH has been shown to depend on the carbon source used. Among the sources used, cellular growth with glucose or glycerol did not affect the extracellular pH, whereas citrate caused the alkalinization of the media. This phenotype is in concordance with computational predictions, at least in the case of the genome-scale metabolic model of Pseudomonas putida KT2440.

Growth of a Methane-Utilizing Mixed Culture HD6T on Methanol and Poly-β-Hydroxybutyrate Biosynthesis

2010 ◽  
Vol 160-162 ◽  
pp. 171-175 ◽  
Author(s):  
Jing Dong ◽  
Jia Ying Xin ◽  
Ying Xin Zhang ◽  
Lin Lin Chen ◽  
Hong Ye Liang ◽  
...  
Keyword(s):  
Mixed Culture ◽  
Mineral Salt Medium ◽  
Biomass Concentration ◽  
Dry Weight ◽  
Maximum Growth ◽  
Maximum Growth Rate ◽  
Production Costs ◽  
Lag Phase ◽  
Phase Duration ◽  
Phb Production

Methane-utilizing mixed culture HD6T was successfully cultivated in a brief non-sterile process using methanol as a sole carbon and energy source for the production of poly-β-hydroxybutyrate(PHB). Shake-flask experiments showed HD6T could grow well in the mineral salt medium with the addition of methanol exposed to the air directly. This non-sterile process and the use of cheap substrates (methanol) can reduce the production costs of PHB. It was found that HD6T grew better and PHB production in a more effective way with an initial liquid methanol concentration of 0.15%(v/v).The lag phase duration, the maximum growth rate, the biomass concentration and the PHB yield, for the optimal conditions were, respectively, 12.03h, 0.04h-1(OD600), 1.54g/l(dry weight), 0.424g/l(dry weight). Methane-utilizing mixed culture HD6T appears to be a promising organism for PHB production.

scholarly journals Ralstonia eutropha H16 Encodes Two and Possibly Three Intracellular Poly[d-(−)-3-Hydroxybutyrate] Depolymerase Genes

2003 ◽  
Vol 185 (13) ◽  
pp. 3788-3794 ◽  
Author(s):  
Gregory M. York ◽  
Joachim Lupberger ◽  
Jiamin Tian ◽  
Adam G. Lawrence ◽  
JoAnne Stubbe ◽  
...  
Keyword(s):  
Ralstonia Eutropha ◽  
Dry Weight ◽  
Complete Sequence ◽  
Inverse Pcr ◽  
Degenerate Primers ◽  
Phb Depolymerase ◽  
Phb Production ◽  
Ralstonia Metallidurans

ABSTRACT Intracellular poly[d-(−)-3-hydroxybutyrate] (PHB) depolymerases degrade PHB granules to oligomers and monomers of 3-hydroxybutyric acid. Recently an intracellular PHB depolymerase gene (phaZ1) from Ralstonia eutropha was identified. We now report identification of candidate PHB depolymerase genes from R. eutropha, namely, phaZ2 and phaZ3, and their characterization in vivo. phaZ1 was used to identify two candidate depolymerase genes in the genome of Ralstonia metallidurans. phaZ1 and these genes were then used to design degenerate primers. These primers and PCR methods on the R. eutropha genome were used to identify two new candidate depolymerase genes in R. eutropha: phaZ2 and phaZ3. Inverse PCR methods were used to obtain the complete sequence of phaZ3, and library screening was used to obtain the complete sequence of phaZ2. PhaZ1, PhaZ2, and PhaZ3 share ∼30% sequence identity. The function of PhaZ2 and PhaZ3 was examined by generating R. eutropha H16 deletion strains (ΔphaZ1, ΔphaZ2, ΔphaZ3, ΔphaZ1ΔphaZ2, ΔphaZ1ΔphaZ3, ΔphaZ2ΔphaZ3, and ΔphaZ1ΔphaZ2ΔphaZ3). These strains were analyzed for PHB production and utilization under two sets of conditions. When cells were grown in rich medium, PhaZ1 was sufficient to account for intracellular PHB degradation. When cells that had accumulated ∼80% (cell dry weight) PHB were subjected to PHB utilization conditions, PhaZ1 and PhaZ2 were sufficient to account for PHB degradation. PhaZ2 is thus suggested to be an intracellular depolymerase. The role of PhaZ3 remains to be established.

scholarly journals Low-quality animal by-product streams for the production of PHA-biopolymers: fats, fat/protein-emulsions and materials with high ash content as low-cost feedstocks

2020 ◽  
Author(s):  
Victoria Saad ◽  
Björn Gutschmann ◽  
Thomas Grimm ◽  
Torsten Widmer ◽  
Peter Neubauer ◽  
...  
Keyword(s):  
Ralstonia Eutropha ◽  
Low Cost ◽  
Carbon Sources ◽  
Dry Weight ◽  
Ash Content ◽  
Production Costs ◽  
Medium Chain Length ◽  
Current Production ◽  
Copolymer Poly ◽  
By Products

Abstract Objective The rapid accumulation of crude-oil based plastics in the environment is posing a fundamental threat to the future of mankind. The biodegradable and bio-based polyhydroxyalkanoates (PHAs) can replace conventional plastics, however, their current production costs are not competitive and therefore prohibiting PHAs from fulfilling their potential. Results Different low-quality animal by-products, which were separated by thermal hydrolysis into a fat-, fat/protein-emulsion- and mineral-fat-mixture- (material with high ash content) phase, were successfully screened as carbon sources for the production of PHA. Thereby, Ralstonia eutropha Re2058/pCB113 accumulated the short- and medium-chain-length copolymer poly(hydroxybutyrate-co-hydroxyhexanoate) [P(HB-co-HHx)]. Up to 90 wt% PHA per cell dry weight with HHx-contents of 12–26 mol% were produced in shake flask cultivations. Conclusion In future, the PHA production cost could be lowered by using the described animal by-product streams as feedstock. Graphical abstract

scholarly journals Sugar-cane juice induces pectin lyase and polygalacturonase in Penicillium griseoroseum

1998 ◽  
Vol 29 (4) ◽  
pp. 246-250 ◽  
Author(s):  
Rosana Cristina Minussi ◽  
Juliana Rocha Lopes Soares-Ramos ◽  
Jorge Luiz Cavalcante Coelho ◽  
Daison Olzany Silva
Keyword(s):  
Yeast Extract ◽  
Sugar Cane ◽  
Culture Media ◽  
Cellulase Activity ◽  
Dry Weight ◽  
Pectin Lyase ◽  
Production Costs ◽  
Cane Juice ◽  
Sugar Cane Juice ◽  
Penicillium Griseoroseum

The use of other inducers as substitutes for pectin was studied aiming to reduce the production costs of pectic enzymes. The effects of sugar-cane juice on the production of pectin lyase (PL) and polygalacturonase (PG) by Penicillium griseoroseum were investigated. The fungus was cultured in a mineral medium (pH 6.3) in a rotary shaker (150 rpm) for 48 h at 25oC. Culture media were supplemented with yeast extract and sucrose or sugar-cane juice. Sugar-cane juice added singly to the medium promoted higher PL activity and mycelial dry weight when compared to pectin and the use of sugar-cane juice and yeast extract yielded levels of PG activity that were similar to those obtained with sucrose-yeast extract or pectin. The results indicated that, even at low concentrations, sugar-cane juice was capable of inducing pectin lyase and polygalacturonase with no cellulase activity in P. griseoroseum.

Some biochemical effects of 4-deoxy-4-fluoro-D-glucose on Escherichia coli

1977 ◽  
Vol 55 (8) ◽  
pp. 911-915 ◽  
Author(s):  
N. F. Taylor ◽  
Li-Yu Louie
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Catabolite Repression ◽  
Dry Weight ◽  
Resting Cells ◽  
Phosphotransferase System ◽  
E Coli ◽  
Biochemical Effects ◽  
Inhibition Of Growth ◽  
Uncompetitive Inhibitor

The uptake of 4-deoxy-4-fluoro-D-glucose (4FG), without subsequent catabolism, by resting cells of Escherichia coli (ATCC 11775) is 0.06 mg/mg dry weight. In frozen–thawed cells of this organism, 4FG is a substrate for the phosphoenolpyruvate phosphotransferase system with a rate of phosphorylation twice that found for the isomeric 3-deoxy-3-fluoro-D-glucose. 4FG is not a carbon source for growth of this organism and it inhibits the extent of growth of cells in the presence of glucose. The inhibition of growth of E. coli K12 on lactose by 4FG is also observed and this is considered to be consistent with the fact that 4FG is an uncompetitive inhibitor of β-galactosidase (EC 3.2.1.23) activity and that 4FG or 4-deoxy-4-fluoro-D-glucose-6-phosphate repress β-galactosidase synthesis. These results support the view that catabolite repression may be produced by compounds which are not necessarily metabolised further than hexose-6-phosphates.

scholarly journals Kinetic Studies and Biochemical Pathway Analysis of Anaerobic Poly-(R)-3-Hydroxybutyric Acid Synthesis in Escherichia coli

2005 ◽  
Vol 71 (2) ◽  
pp. 713-720 ◽  
Author(s):  
Ross Carlson ◽  
Aaron Wlaschin ◽  
Friedrich Srienc
Keyword(s):  
Escherichia Coli ◽  
Kinetic Studies ◽  
Dry Weight ◽  
Acid Synthesis ◽  
Control Culture ◽  
Biochemical Network ◽  
Hydroxybutyric Acid ◽  
E Coli ◽  
Phb Production ◽  
Acetate Accumulation

ABSTRACT Poly-(R)-3-hydroxybutyric acid (PHB) was synthesized anaerobically in recombinant Escherichia coli. The host anaerobically accumulated PHB to more than 50% of its cell dry weight during cultivation in either growth or nongrowth medium. The maximum specific PHB production rate during growth-associated synthesis was approximately 2.3 ± 0.2 mmol of PHB/g of residual cell dry weight/h. The by-product secretion profiles differed significantly between the PHB-synthesizing strain and the control strain. PHB production decreased acetate accumulation for both growth and nongrowth-associated PHB synthesis. For instance under nongrowth cultivation, the PHB-synthesizing culture produced approximately 66% less acetate on a glucose yield basis as compared to a control culture. A theoretical biochemical network model was used to provide a rational basis to interpret the experimental results like the fermentation product secretion profiles and to study E. coli network capabilities under anaerobic conditions. For example, the maximum theoretical carbon yield for anaerobic PHB synthesis in E. coli is 0.8. The presented study is expected to be generally useful for analyzing, interpreting, and engineering cellular metabolisms.

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