• KSBB Journal
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• 제15권6호
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• pp.556-559
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• 2000

Cyclodextrin Glucanotransferase(EC 2.4.1.19 : 1,4-${\alpha}$-glucano) transferase, cyclizing; CGTase) can be separated and recovered in an aqueous two-phase system composed of poly(ethylene glycol)(PEG)/dextran and PEG/salt. In an aqueous two-phase system consisting of PEG 35000 (5%) and dextran T2000 (7%), all cell and debris were collected at the interphase. CGTase partitioned to the denser dextran phase at an yield of 83.4%. On the other hand, in an aqueous two-phase system consisting of PEG 35000 (10%) and sodium phosphate (15%), CGTase partitioned to the denser salt phase at an yield of 95.5%. In order to recover CGTase using an aqueous two-phase system, the PEG/salt system proved to be more efficient than the PEG/dextran system in terms of yield and cost.

• 한국미생물·생명공학회지
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• 제22권5호
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• pp.531-537
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• 1994

This study was undertaken with objective of optimizing the conditions of fermentation in an aqueous two-phase system which is composed of polyethylene glycol (PEG) 20000 and crude dextran (Dx). The data were obtained and analyzed using the Box-Wilson's experimental design protocol and the response surface methodology. To reach this end a multilinear polynomial regres- sion model was developed, which can be utilized for the purpose of optimizing the extractive fermentation. Optimum conditions for batch fermentation with aqueous two phase system were found to be at 4.2~5.4% PEG/3.2~4.2% Dx range. The composition of the center was 4.8% PEG/ 3.6% Dx. Optimum operating conditions for initial sugar concentration and fermentation time were approximately 160 g/l, and 21~22 hr, respectively. Fermentation in the aqueous two phase system composed of 5% PEG/4% Dx showed increase of 23% in ethanol concentration, of 9.5% in ethanol yield, and of 19% in ethanol productivity as compared to the case of fermentation of neat Jerusalem artichoke juice.

• Journal of Microbiology and Biotechnology
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• 제2권2호
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• pp.135-140
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• 1992

The partitioning of recombinant human interleukin-2(rhII-2) in PEG 8000-dextran 38800 aqueous two-phase system has been investigated using three different sources of rhIL-2. In the case of pure rhIL-2, the solubility in a PEG-dextran two-phase system was low and most of rhIL-2 was partitioned into the bottom phase. For the recovery of rhIL-2 from insoluble protein aggregates, the inclusion bodies of recombinant E. coli were solubilized by the treatment with sodium dodecyl sulfate (SDS). The addition of SDS significantly enhanced not only the solubility of rhIL-2 but also the partitioning of rhIL-2 to the top phase. When the ratio of SDS to rhIL-2 was 2.0, the partition coefficient(K) and the recovery yield(Y) at the top phase were 4.5 and 88%, respectively, at pH 6.8. In order to reduce the recovery steps further, SDS was directly added to the intact recombinant E. coli cells and then partitioned into the PEG/dextran aqueous two-phase system. The observed partition coefficient ($K{\cong{3.0$) and recovery yield ($Y{\geq}80%$ )of this method were comparable to the rhIL-2 recovery from insoluble protein aggregates. The results obtained in this work indicate that PEG-dextran two-phase partitioning might provide a simple way for the recovery and partial purification of recombinant proteins which are produced as inclusion bodies.

• Journal of Microbiology and Biotechnology
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• 제9권4호
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• pp.498-503
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• 1999

Protease produced from Bacillus sp. FSE-68 was isolated from Meju, a Korean fermented soybean starter, and was purified by a two-consecutive aqueous two-phase system. The change of partition coefficient (K) in the polyethylene glycol (PEG)/potassium phosphate buffer (PPB) aqueous two-phase system was measured at different pHs (6.0- 9.2), PPB concentrations (8-12%), and temperatures (4 and $20^{\circ}C$). As the PPB concentration in the aqueous two-phase system increased, the protease concentration in the top phase (PEG-rich phase) increased, thereby enhancing the partition coefficient. The minimum partition coefficient of the protease was achieved at pH 7.0, whereas that of the total protein was at pH 6.0. The biggest difference in partition coefficients of total protein and protease occurred at pH 6.0. It was interesting to note that the partition coefficient of protease decreased as the temperature increased. The optimum condition of the primary aqueous two-phase extraction of Bacillus sp. FSE-68 was pH 6.0, 14% (w/w) PPB, and 16% (w/w) PEG at $4^{\circ}C$, and the crude enzyme concentration in this system was 50% (w/w). The protease, which was concentrated in the top phase, was further mixed with 15% (w/w) PPB (pH 7.0) in the ratio of 1:1 at $20^{\circ}C$ to elute the bottom phase (PPB-rich phase). Using these steps, the purification fold achieved was 9.2 with a 44.7% yield.

• Journal of Microbiology and Biotechnology
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• 제9권5호
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• pp.589-592
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• 1999

Suspension cultures of Digitalis lanata were successfully performed in an aqueous two-phase system (ATPS) of 4.5％ polyethylene glycol (PEG) 20,000 and 2.8％ crude dextran. Cell growth in the medium containing an individual ATPS-forming polymer was inhibited due to the toxicity of PEG and a high viscosity of dextran. Formation of ATPS supported cell growth by showing a considerably decrease in viscosity and partitioning of cells into a PEG-lean dextran phase. It was found that an aqueous two-phase cultivation of plant cells in a stirred tank bioreactor could be successfully applied.

• 한국미생물·생명공학회지
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• 제26권1호
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• pp.55-60
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• 1998

수용성 이상계에서 효율적인 추출발효를 수행하기 위해서는 생체촉매와 생성물이 서로 반대 상에 우세하게 분배되어아 한다. 일반적으로 분자량이 낮은 생성물은 양쪽상에 비슷하게 분배되므로, 상의 조성 등을 조절하여 한쪽상의 체적을 증가시켜 최대한으로 생성물을 추출하여야 한다. 본 연구에서는 PEI와 HEC으로 구성된 수용성 이상계에서 L. helveticus와 젖산의 분배에 미치는 다가 음이온의 농도, pH, 배지의 농도 및 생육배지 농도 등의 영향을 검토하였다. PEI의 상대 이온으로 작용하는 phosphate, sulfate 음이온수의 증가는 PEI 상의 체적을 감소시켜 하부상에 분배되는 세포의 양(Cb)을 감소시켰으며, sulfate 농도의 영향은 phosphate에 비해 적었다. 검토한 범위에서 대략 93%이상의 세포가 HEC-rich상부상에 분배되었다. 이상계에 생육배지 농도를 달리하여 첨가했을 때 배지 중에 존재하는 phosphate와 sulfate 농도에 따라 비슷한 경향을 나타냈지만 낮은 농도로 인하여 그 영향은 크지 않았다. Phosphate 농도에 따른 젖산분배를 검토한 결과 농도가 증가할수록 분배계수는 감소하였으며, 대략 85%가량의 젖산이 PEI의 하부상에 분배되었다. 이 결과로부터 수용성 이상계에서 전하를 띤 저 분자량의 생성물의 추출은 반대 전하를 띠는 폴리머를 하나의 상형성 풀리머로 사용함으로써 증진되었음을 알 수 있었다. 따라서 L. helveticus 세포는 95% 정도가 상부상에 분배되고 젖산은 85%정도가 하부상에 우세하게 분배되므로 PEI/HEC 이상계는 치즈유청으로 부터의 젖산의 추출발효에 가능성이 높은 시스템으로 응용될 수 있는 것으로 생각된다.

• Journal of Microbiology and Biotechnology
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• 제7권5호
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• pp.345-351
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• 1997

An aqueous/organic two-phase reaction system was applied to the production of catechol using immobilized resting cells of Pseudomonas putida CY 400. Water/ethyl ether system was used because of high partition coefficient of catechol and thus to reduce the product inhibition and degradation. Among the tested immobilization carriers, polyacrylamide gel gave the highest catechol productivity. The immobilization seemed to protect the cells against solvent toxicity. From the simulation of reaction conditions based on two-phase models, it was found that there was an optimum acetate concentration at fixed benzoate and cell concentrations for the catechol productivity. A lower phase volume ratio (lower fraction of organic phase) gave a higher productivity. However, the substrate conversion was low at low phase volume ratio.

• Biotechnology and Bioprocess Engineering:BBE
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• 제2권2호
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• pp.97-100
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• 1997

Cyclodxtrin homologues(CDs), produced by cyclodextrin glycosyltransferase(CGTase), were simultaneously partitioned in aqueous two-phase system(ATPS). Partition coefficients of CDs were measured in PEG/dextran systems. Phosphate, citrate, sulfate were tested as salt. ATPS of PEG/salt and PEG/dextran had the partition coefficients of the CDs, larger than unity. However, PEG/dextran system was observed better than PEG/salt as CGTase activity decreased sharply with salt concentration. Enzymatic rection occurred mainly in PEG-rich bottom phase because of the low partition coefficient of CGTase. The resulting CDs transferred to the PEG-rich top phase, obeying the diffusional partition. In the ATPS of 7% PEG(M.W.40, 000), 7mg/ml of CDs were obtained in top phase at 4.5 hours.

• 한국미생물·생명공학회지
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• 제16권5호
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• pp.358-362
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• 1988

세포외 효소를 효율적으로 생산하기 위한 새로운 발효시스템으로서 수성 2상계를 시험하였다. 실험대상으로는 Bacillus amyloliquefaciens 균주에 의하여 $\alpha$-amylase를 생산하는 시스템을 선정하였다. Polyethylene glycol 5%, dextran 7%, 그리고 발효액이 88% 포함된 수성 2상계 발효시스템에 의하여 $\alpha$-amylase의 생합성은 25% 정도 증가하였으며 효소의 비활성화는 관찰되지 않았다. 또한 Polyethylene glycol 이 효소의 생합성을 증진시키는 것으로 관찰되었다. 효소의 생합성을 증가시키며 동시에 효소를 발효액으로부터 분리하기 위하여는 polyethylene glycol을 성장도중에, dextran은 효소의 활성이 최고에 달한 후에 발효액에 가하는 발효방법이 바람직하다.

• 한국미생물·생명공학회지
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• 제20권2호
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• pp.190-195
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• 1992

Polythylene glycol/dextran 수성 2단계에서 polyethylene glycol의분자량과 농도가 증가할수록 체적비는 증가하고 분배계수는 감소하였다. 또한 dextran의 분자량이 증가할 수록 체적비는 감소하고 분배계수는 증가하였다. 한편 dextran의 농도가 증가할수록 체적비와 분배계수는 감소하였다.