• Microbiology and Biotechnology Letters
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• v.48 no.4
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• pp.506-514
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• 2020

4-Hydroxybenzyl alcohol (4-HB alcohol) is one of the major active components of Gastrodia elata Blume, with beneficial effects on neurological disorders such as headache, convulsive behavior, and dizziness. Here, we developed a metabolically engineered Corynebacterium glutamicum strain able to produce 4-HB alcohol from 4-hydroxybenzoate (4-HBA). First, the strain APS963 was obtained from the APS809 strain via the insertion of aroK from Methanocaldococcus jannaschii into the NCgl2922-deleted locus. As carboxylic acid reductase from Nocardia iowensis catalyzes the reduction of 4HBA to 4-hydroxybenzaldehyde (4-HB aldehyde), we then introduced a codon-optimized car gene into the genome of APS963, generating the GAS177 strain. Then, we deleted creG coding for a putative short-chain dehydrogenase and inserted ubiCpr encoding a product-resistant chorismate-pyruvate lyase into the pcaHG-deleted locus. The resulting engineered GAS355 strain accumulated 2.3 g/l 4-HB alcohol with 0.32 g/l 4-HBA and 0.3 g/l 4-HB aldehyde as byproducts from 8% glucose after 48 h of culture.

• Journal of Life Science
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• v.34 no.10
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• pp.682-690
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• 2024

This study aimed to produce medium-chain-length polyhydroxyalkanoates (mcl-PHA) using oil extracted from spent coffee grounds (SCG) as a carbon source. The highest oil yields, 14.40%(w/w) and 14.49%(w/w), were achieved with an extraction time of 2 hr and an SCG/n-hexane ratio of 15:1. The fatty acid composition of the extracted SCG oil (EOC) primarily consisted of linoleic acid (45.08%), palmitic acid (35.56%), oleic acid (8.62%), and stearic acid (7.62%). Pseudomonas sp. HY61, isolated from soil, was used to biosynthesize mcl-PHA composed of 3-hydroxyhexanoate, 3-hydroxyoctanoate, 3-hydroxydecanoate, and 3-hydroxydodecanoate using EOC. Under optimal flask conditions, the highest cell growth and mcl-PHA accumulation were observed with 5 g/l EOC, 72 hr, with NH₄Cl as the nitrogen source, at pH 7, 25℃, and a C/N ratio of 15:1. Batch fermentation resulted in a dry cell weight (DCW) of 0.92 g/l and mcl-PHA of 0.17 g/l after 72 hr. In contrast, fed-batch fermentation, with additional supplementation of EOC and NH₄Cl at 18, 30, and 42 hr, increased the DCW to 2.85 g/l and mcl-PHA accumulation to 0.67 g/l, approximately three times higher than batch fermentation. Gas chromatography confirmed that the mcl-PHA structure was consistent across both fermentation modes. The synthesized mcl-PHA had higher molecular weight and thermal decomposition temperatures compared to other mcl-PHA, suggesting its potential applications in packaging and biomedical fields. This study offers an economical approach for mcl-PHA production using waste EOC and Pseudomonas sp. HY61.

• KSBB Journal
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• v.22 no.5
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• pp.288-296
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• 2007

Monascus pilosus (KCCM 60160) in submerged culture was optimized based on culture medium and fermentation conditions. Monacolin-K (Iovastatin), one of the cholesterol lowing-agent which was produced by Monascus pilosus may maintain a healthy lipid level by inhibiting the biosynthesis of cholesterol. Plackett-Burman design and response surface method were employed to study the culture medium for the desirable monacolin-K production. As a result of experimental designs, optimized production medium components and concentrations (g/L) were determined on soluble starch 96, malt extract 44.5, beef extract 30.23, yeast extract 15, $(NH_4)_2SO_4$ 4.03, $Na_2HPO_4{\cdot}12H_2O$ 0.5, L-Histidine 3.0, $KHSO_4$ 1.0, respectively. Monacolin-K production was improved about 3 times in comparison with shake flask fermentation of the basic production medium. The effect of agitation speed (300, 350, 400 and 450 rpm) on the monacolin-K production were also observed in a batch fermenter. Maximum monacolin-K production with the basic production medium was 68 mg/L when agitation speed was 500 rpm. And it was found that all spherical pellets (average diameter of $1.0{\sim}1.5mm$) were dominant during fermentation. Based on the results, the maximum production of 185 mg/L of monacolin-K with the optimized production medium was obtained at pH (controlled) 6.5, agitation rate 400 rpm, aeration rate 1 vvm, and inoculum size 3%.

• KSBB Journal
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• v.23 no.6
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• pp.463-469
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• 2008

During the last decade, monacolin-K biosynthesized by fermentation of red yeast rice (Monascus strains) was proved to have an efficient cholesterol lowering capability, leading to rapid increase in the market demand for the functional red yeast rice. In this study, the production medium composition and components were optimized on a shake flask scale for monacolin-K production by Monascus pilosus (KCCM 60160). The effect of three different soybean flours on the monacolin-K production were studied in order to replace the nitrogen sources of basic production medium (yeast extract, malt extract and beef extract). Among the several experiments, the production medium with dietary soybean flour to replace a half of yeast extract was very good for monacolin-K production. Plackett-Burman experimental design was used to determine the key factors which are critical to produce the biological products in the fermentation. According to the result of Plackett-Burman experimental design, a second order response surface design was applied using yeast extract, beef extract and $(NH_4)_2SO_4$ as factors. Applying this model, the optimum concentration of the three variables was obtained. The maximum monacolin-K production (369.6 mg/L) predicted by model agrees well with the experimental value (418 mg/L) obtained from the experimental verification at the optimal medium. The yield of monacolin-K was increased by 67% as compared to that obtained with basic production medium in shake flasks.

• Journal of Life Science
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• v.31 no.1
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• pp.90-99
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• 2021

In this study, to reduce the production cost of poly(3-hydroxyalkanoates) (PHA), optimal cell growth and PHA biosynthesis conditions of the isolated strain Pseudomonas sp. EML8 were established using waste frying oil (WFO) as the cheap carbon source. Gas chromatography (GC) and GC mass spectrometry analysis of the medium-chain-length PHA (mcl-PHAWFO) obtained by Pseudomonas sp. EML8 of WFO indicated that it was composed of 7.28 mol% 3-hydrxoyhexanoate, 39.04 mol% 3-hydroxyoctanoate, 37.11 mol% 3-hydroxydecanoate, and 16.58 mol% 3-hydroxvdodecanoate monomers. When Pseudomonas sp. EML8 were culture in flask, the maximum dry cell weight (DCW) and the mcl-PHAWFO yield (g/l) were showed under WFO (20 g/l), (NH4)2SO4 (0.5 g/l), pH 7, and 25℃ culture conditions. Based on this, the highest DCW, mcl-PHAWFO content, and mcl-PHAWFO yield from 3-l-jar fermentation was obtained after 48 hr. Similar results were obtained using 20 g/l of fresh frying oil (FFO) as a control carbon source. In this case, the DCW, the mcl-PHAFFO content, and the mcl-PHAFFO yields were 2.7 g/l, 62 wt%, and 1.6 g/l, respectively. Gel permeation chromatography analysis confirmed the average molecular weight of the mcl-PHAWFO and mcl-PHAFFO to be between 165-175 kDa. Thermogravimetric analysis showed decomposition temperature values of 260℃ and 274.7℃ for mcl-PHAWFO and mcl-PHAFFO, respectively. In conclusion, Pseudomonas sp. EML8 and WFO could be suggested as a new candidate and substrate for the industrial production of PHA.

• Journal of Life Science
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• v.32 no.7
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• pp.510-522
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• 2022

In this study, to biosynthesize PHA with properties more similar to polypropylene, a Bacillus sp. EMK-5020 strain that biosynthesized poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) was isolated from soil. Bacillus sp. EMK-5020 strain biosynthesized PHBV containing 1.3% 3-hydroxyvalerate (3HV) using reducing sugar contained in Makgeolli lees enzymatic hydrolysate (MLEH) as a single carbon source. As the amount of propionic acid, which was added as a second carbon source, increased, the content of 3HV also increased. PHBV containing up to 48.6% of 3HV was synthesized when 1.0 g/l of propionic acid was added. Based on these results, the strain was cultured for 72 hr in a 3 l fermenter using reducing sugar in MLEH (20 g/l) and propionic acid (1 g/l) as the main and secondary carbon sources, respectively. As a result, 6.4 g/l DCW and 50 wt% of PHBV (MLEH-PHBV) containing 8.9% 3HV were biosynthesized. Through gel permeation chromatography and thermogravimetric analysis, it was confirmed that the average molecular weight and the decomposition temperature of MLEH-PHBV were 152 kDa and 273℃, respectively. In conclusion, the Bacillus sp. EMK-5020 strain could biosynthesize PHBV containing various 3HV fractions when MLEH and propionic acid were used as carbon sources, and PHBV-MLEH containing 8.9% 3HV was confirmed to have higher thermal stability than standard PHBV (8% 3HV).