• Journal of Korean Society of Environmental Engineers
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• v.35 no.2
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• pp.109-114
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• 2013

Polyhydroxyalkanoates (PHAs) are synthesized by numerous bacteria as carbon and energy storage compounds and are raw materials for biocompatible plastics. In this paper, the effect of $CO_2$ concentrations on the growth of C. necator and the accumulation of Poly(3-hydroxybutyrate) (P(3HB)) are investigated by increasing the $CO_2$ concentration in the substrate gas mixture. During 6 d cultivation in a nitrogen-present mineral medium, the $CO_2$ concentration did not affect the growth of the cells, while the Poly(3- hydroxybutyrate) (P(3HB)) content decreased with increasing $CO_2$ concentrations from 1% to 20%. During 4 d cultivation in the nitrogen-limited medium, the P(3HB) accumulation was the greatest at 3% $CO_2$; however, the total amount of accumulated P(3HB) was the greatest at 1% $CO_2$, which decreased with increasing $CO_2$ concentrations. The results indicate that the gas mixture with 1% $CO_2$ is the most effective in both growing the cells and accumulating P(3HB) under our experimental conditions.

• Korean Chemical Engineering Research
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• v.54 no.5
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• pp.665-670
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• 2016

This study deals with the possible degradation of toluene and acetic acid when subjected to cell-free enzyme system from the toluene degrading bacteria Pseudomonas putida and acetic acid degrading bacteria Cupriavidus necator. P. putida produces toluene dioxygenase only under the existence of toluene in culture medium and toluene is degraded to cis-toluene dihydrodiol by this enzyme. C. necator produces acetyl coenzyme A synthetase-1 and converts acetic acid to acetyl CoA in order to synthesize ATP to need for growth or PHA which is biodegradable polymer. In case of toluene degradation, the experiment was conducted before and after production of toluene dioxygenase as this enzyme, produced by P. putida, is an inducible enzyme. Toluene was detected using gas chromatography (GC). Similar amount of toluene was found in control group and before production of toluene dioxygenase (experimental group 1). However, reduction in toluene was detected after the production of toluene dioxygenase (experimental group 2). Acetic acid was detected through application of gas chromatography-mass spectrometer (GC-MS). The results showed the acetic acid peak was not detected in the experimental group to apply cell-free enzyme system. These results show that the cell-free enzyme system obtained from P. putida and C. necator retained the ability to degrade toluene and acetic acid. However, P. putida needs to produce the inducible enzyme before preparation of the cell-free enzyme system.

• Journal of Microbiology and Biotechnology
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• v.25 no.11
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• pp.1936-1943
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• 2015

The present study developed an efficient and environmentally friendly method for recovering polyhydroxybutyrate (PHB) from Cupriavidus necator. Several non-halogenated solvents were tested and it was found that butyl acetate and ethyl acetate are powerful solvents for the biopolymer. Testing was performed to examine the effects of temperature (25℃ until temperature below solvent boiling points) and heating incubation time (0-60 min) on the two solvents. Butyl acetate had a higher recovery level (96%) and product purity (up to 99%) than ethyl acetate at 103℃ and a heating incubation time of 30 min. Under these conditions, PHB recorded the highest molecular weight of 1.4 × 10⁶ compared with the standard procedure (i.e., recovery using chloroform). The proposed strategy showed that butyl acetate is a good alternative to halogenated solvents such as chloroform for recovery of PHB.

• Environmental Engineering Research
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• v.20 no.3
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• pp.246-253
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• 2015

The effects of carbon-to-nitrogen (C/N) ratio in simulated rice straw hydrolysates using glucose and ammonium chloride on polyhydroxyalkanoates (PHA) accumulation by Cupriavidus necator was investigated. In general, PHA accumulation rate was higher under higher degrees of N-deficient conditions (e.g., C/N ratio of 360:1) than lower degrees of N-deficient conditions (e.g., C/N ratio of 3.6:1 and 36:1). Also, the most PHA accumulation was observed during the first 12 h after the PHA accumulation initiation. This study showed that the similar PHA accumulation could be achieved by using different accumulation periods depending on C/N ratios. N source presence was important for new cell production, supported by approximately ten times greater PHA accumulation under the N-deficient condition ($NH_4Cl$ 0.01 g/L) than the N-free (without $NH_4Cl$) condition after 96 h. C/N ratio of the rice straw hydrolysate was approximately 160:1, based on the glucose content, and this accumulated $0.36{\pm}0.0033g/L$ PHA with PHA content of $21{\pm}3.1%$ after 12 h. Since external C or N source addition for C/N ratio adjustment increases production cost, an appropriate accumulation period may be used for PHA accumulation from organic wastes, based on the PHA accumulation patterns observed at various C/N ratios and C and N concentrations.

• Journal of Microbiology and Biotechnology
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• v.29 no.3
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• pp.382-391
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• 2019

Many poultry eggs are discarded worldwide because of infection (i.e., avian flu) or presence of high levels of pesticides. The possibility of adopting egg yolk as a source material to produce polyhydroxyalkanoate (PHA) biopolymer was examined in this study. Cupriavidus necator Re2133/pCB81 was used for the production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) or poly(3HHx), a polymer that would normally require long-chain fatty acids as carbon feedstocks for the incorporation of 3HHx monomers. The optimal medium contained 5% egg yolk oil and ammonium nitrate as a nitrogen source, with a carbon/nitrogen (C/N) ratio of 20. Time course monitoring using the optimized medium was conducted for 5 days. Biomass production was 13.1 g/l, with 43.7% co-polymer content. Comparison with other studies using plant oils and the current study using egg yolk oil revealed similar polymer yields. Thus, discarded egg yolks could be a potential source of PHA.

• Journal of Korean Society of Environmental Engineers
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• v.30 no.11
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• pp.1161-1169
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• 2008

The removals of TCE and PCE vapor with or without a supply of toluene as a primary substrate were compared in a biofiltration process, and the variations of microbial communities associated with the removal were also investigated. As a result of investigations on the removals of TCE/PCE in a biofilter B within which TCE/PCE-acclimated sludge was attached on the surface of media without a supply of primary substrate, and those in another biofilter A where toluene-acclimated sludge was attached with a supply of toluene as a primary substrate, followings were found: (i) parts of microbes responsible to the decomposition of toluene vapor participate in the removal of chlorinated VOCs such as TCE and PCE, and (ii) effective biological removals of TCE and PCE vapor do not necessarily need cometabolism. Sequencing of 16S rDNA obtained from the band profile of DGGE (Denaturating Gradient Gel Electrophoresis), it was confirmed that: (i) uncultured alpha proteobacterium, uncultured Desulfitobacterium, uncultured Rhodobacteraceae bacterium, Cupriavidus necator, and Pseudomonas putida were found to be toluene-decomposing microbes, (ii) alpha proteobacterium HTCC396 is a TCE-removing microbe, (iii) Desulfitobacterium sp. is a PCE-decomposing microbe, and (iv) particularly, uncultured Desulfitobacterium sp. is probably a microbe decomposable not only toluene but also various chlorinated VOC vapor including TCE and PCE.

• Journal of Microbiology and Biotechnology
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• v.32 no.1
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• pp.110-116
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• 2022

Polyhydroxyalkanoates (PHAs) are emerging as alternatives to plastics by replacing fossil fuels with renewable raw substrates. Herein, we present the construction of engineered Escherichia coli strains to produce short-chain-length PHAs (scl-PHAs), including the monomers 4-hydroxyvalerate (4HV) and 3-hydroxyvalerate (3HV) produced from levulinic acid (LA). First, an E. coli strain expressing genes (lvaEDABC) from the LA metabolic pathway of Pseudomonas putida KT2440 was constructed to generate 4HV-CoA and 3HV-CoA. Second, both PhaAB enzymes from Cupriavidus necator H16 were expressed to supply 3-hydroxybutyrate (3HB)-CoA from acetyl-CoA. Finally, PHA synthase (PhaC_Cv) from Chromobacterium violaceum was introduced for the subsequent polymerization of these three monomers. The resulting E. coli strains produced four PHAs (w/w% of dry cell weight): 9.1 wt% P(4HV), 1.7 wt% P(3HV-co-4HV), 24.2 wt% P(3HB-co-4HV), and 35.6 wt% P(3HB-co-3HV-co-4HV).