• KSBB Journal
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• 제21권1호
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• pp.1-10
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• 2006

식물세포배양으로부터 항암제 paclitaxel의 생산을 위해 회수 및 정제는 산업적 공정에 있어서 필수적이다. 본 총설은 식물세포배양으로부터 고순도, 고수율의 paclitaxel 생산을 위한 대량 회수 및 정제 방법을 기술하고자 한다. 또한 이러한 분리 및 정제 공정은 추출, 전 처리, 정제, 제품화 단계를 총괄하여 최종 제품의 요구조건들 즉, 순도, 잔류용매, 제품형태, 불순물 함량, 엔도톡신 함량 등을 충족시킬 수 있도록 최적화되어야 한다. 이러한 관점에서 본 총설은 산업화 단계에서의 의약품 생산 및 품질관리에 상당히 유용하게 활용될 수 있을 것으로 판단된다.

• Biomolecules & Therapeutics
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• 제6권4호
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• pp.383-388
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• 1998

The present study was aimed to investigate the preventive effects of rebamipide on the multiple organ dysfunction in a rat model of circulatory shock induced by bacterial endotoxin (E. coli lipopolysaccharide; LPS) in comparison with that of methotrexate. Endotoxemia for 6 hours resulted in little change in the levels of hemoglobin and neutrophils. However, treatment with methotrexate decreased significantly the numbers of circulating neutrophils. Significant increases in serum alanine aminotransferase (ALT,958 $\pm$ 250 lU/L, p<0.001) and aspartate aminotransferase (AST, 1350 $\pm$ 295 lU/L, p<0.001) levels induced by endotoxemia were significantly decreased by rebamipide and methotrexate. The increased level of lactic acid dehydrogenase (LDH) by LPS (2850 $\pm$ 467 lU/L, p<0.05) was significantly inhibited by rebamipide, but not by methot.elate. The elevated serum creatinine (1.2$\pm$0.1, p.0.05) and urea levels (55.3$\pm$6.5 mg/dL, p.0.01) by LPS were also decreased by rebamipide, but not by methotrexate. In line with these results, the plasma concentration of tumor necrosis factor-$\alpha$ (TNF-7,167 $\pm$ 20 pg/mL) was significantly increased upon injection of endotoxin at 1 hour by 1570$\pm$100 pg/mL, and declined to 312$\pm$35 pg/mL at 6 hours. The TNF-$\alpha$ level at 6 hours was significantly decreased by rebamipide to 207$\pm$8 pg/mL (P<0.05). Taken together, it is summarized that rebamipide inhibits the development of multiple ogran dysfunction by inhibition of neutrophil activation in association with inhibition of TNF-$\alpha$ formation in a murine model of endotoxemia.

• Asian-Australasian Journal of Animal Sciences
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• 제30권12호
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• pp.1784-1795
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• 2017

Objective: The study examined the effect of intravenous administration of bacterial endotoxin-lipopolysaccharide (LPS) -on the nocturnal secretion of melatonin and on the expression of enzymes of the melatonin biosynthetic pathway in the pineal gland of ewes, taking into account two different photoperiodic conditions: short-night (SN; n = 12) and long-night (LN; n = 12). Methods: In both experiments, animals (n = 12) were randomly divided into two groups: control (n = 6) and LPS-treated (n = 6) one. Two hours after sunset, animals received an injection of LPS or saline. Blood samples were collected starting one hour after sunset and continuing for 3 hours after the treatment. The ewes were euthanized 3 hours after LPS/saline treatment. The concentration of hormones in plasma was assayed by radioimmunoassay. In the pineal gland, the content of serotonin and its metabolite was determined by HPLC; whereas the expression of examined genes and protein was assayed using real-time polymerase chain reaction and Western Blot, respectively. Results: Endotoxin administration lowered (p<0.05) levels of circulating melatonin in animals from LN photoperiod only during the first hour after treatment, while in ewes from SN photoperiod only in the third hour after the injection. Inflammation more substantially suppressed biosynthesis of melatonin in ewes from SN photoperiod, which were also characterised by lower (p<0.05) cortisol concentrations after LPS treatment compared with animals from LN photoperiod. In the pineal gland of ewes subjected to SN photoperiod, LPS reduced (p<0.05) serotonin content and the expression of melatonin biosynthetic pathway enzymes, such as tryptophan hydroxylase and arylalkylamine-N-acetyltransferase. Pineal activity may be disturbed by circulating LPS and proinflammatory cytokines because the expression of mRNAs encoding their corresponding receptors was determined in this gland. Conclusion: The present study showed that peripheral inflammation reduces the secretion of melatonin, but this effect may be influenced by the photoperiod.

• Journal of Microbiology and Biotechnology
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• 제17권11호
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• pp.1862-1867
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• 2007

An individual's immune response is critical for host protection from many different pathogens, and the responsiveness can be assessed by the amount of cytokine production upon stimulating bacterial components such as lipopolysaccharide (LPS). The difference between individuals in their peripheral blood mononuclear cells (PBMC) responsiveness to LPS, a Gram-negative endotoxin, was investigated from 27 healthy individuals. We observed a large variation in $IFN{\gamma}$ production among different individuals. The PBMC of the consistently three highest and three lowest $IFN{\gamma}$ producers were investigated. Since previous studies described that a single point mutation in the coding region of TLR2 and TLR4 is linked to the individual responsiveness to pathogenic bacterial infections, we first examined the known point mutations in the coding region of $TLR2^{Pro681His}$, $TLR4^{Pro714His}$ located in the cytoplasmic regions of the Toll-like domain as well as $TLR4^{Asp299Gly}$ located in the extracellular region. None of these mutations were associated with an individual's responsiveness to LPS, despite the presence of $TLR4^{Asp299Gly}$ mutation. Further investigation revealed that the variation of PBMC responsiveness to LPS among healthy individuals was due to constitutive expression levels of TLR4 and TLR2. This result is consistent with an aging-related low expression of Toll-like receptors in the mouse model of LPS responsiveness. The present study therefore suggests that the constitutive expression levels of TLR2 and TLR4 may contribute to the individual response to LPS.

• Journal of Microbiology and Biotechnology
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• 제19권1호
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• pp.108-112
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• 2009

A simplified method for the purification of cholera toxin was developed. The 569B strain of Vibrio cholerae, a recognized hyper-producer of cholera toxin, was propagated in a bioreactor under conditions that promote the production of the toxin. The toxin was separated from the bacterial cells using 0.2-${\mu}m$ crossflow microfiltration, the clarified toxin was passed through the membrane into the permeate, and the bacterial cells were retained in the retentate. The 0.2-${\mu}m$ permeate was then concentrated 3-fold and diafiltered against 10 mM phosphate buffer, pH 7.6, using 30-kDa crossflow ultrafiltration. The concentrated toxin was loaded onto a cation exchange column, the toxin was bound to the column, and most of the impurities were passed unimpeded through the column. The toxin was eluted with a salt gradient of phosphate buffer, pH 7.0, containing 1.0 M NaCl. The peak containing the toxin was assayed for cholera toxin and protein and the purity was determined to be 92%. The toxin peak had a low endotoxin level of $3.1\;EU/{\mu}g$ of toxin. The purified toxin was used to prepare antiserum against whole toxin, which was used in a $G_{M1}$ ganglioside-binding ELISA to determine residual levels of toxin in an oral inactivated whole-cell cholera vaccine. The $G_{M1}$ ganglioside-binding ELISA was shown to be very sensitive and capable of detecting as little as 1 ng/ml of cholera toxin.

• 한국약용작물학회지
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• 제15권6호
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• pp.451-456
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• 2007

Curcumin, a polyphenolic antioxidant purified from turmeric, has been known to possess various biological activities such as anti-oxidative, anti-inflammatory and anti-cancer effects. In this study, we have explored anti-inflammatory effect of curcumin using Gram (-) bacterium-derived endotoxin (lipopolysaccharide: LPS) and macrophage cell line RAW264.7. Curcumin suppressed NO production in LPS-activated RAW264.7 cells in a dose-dependent manner, Curcumin also blocked the activation of $NF-{\kappa}B$ but not AP-1 according to luciferase assay. Furthermore, this compound suppressed the phosphorylation of a series of intracellular signaling components such as Src, JAK-2, Akt, IKK and $I{\kappa}B{\alpha}$ under LPS stimulation in a time dependent manner, Therefore, our data suggest that curcumin was able to protect the host from Gram(-) bacterial-infection-mediated inflammatory symptoms.

• 대한약학회:학술대회논문집
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• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2
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• pp.329.2-330
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• 2002

This study was intended to establish test methods equivalent to those of "Interferon alfa-2 concentrated solution" monograph in European Pharmacophoeia(EP). Two recombinant interferon alfa concentrated solutions manufactured in Korea were tested according to the monograph of EP. Tests of identification(biological activity. isoelectric focusing. SDS-PAGE under reducing condition, peptide mapping). related proteins. impurities of moducular masses differing from that of interferon alfa-2(SDS-PAGE under reducing and non-reducing condition). bacterial endotoxin, protein, potency, host-cell-derrved proteins. and host-cell-derived DNA were performed in the laboratories of manufacrues and division of biotechnology. KFDA. The results of this study showed that specitications of interfenon alfa concentrated solutions manufactured in Korea were within the aceptance criteria of EP. Based on the study. specitications and test methods for interferon alfa concentrated solution can be established according to the monograph of EP suggesting the revision of Minimum requirements for biological products

• 생명과학회지
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• 제32권12호
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• pp.956-961
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• 2022

식물에서 재조합 단백질을 생산하는 것은 여러 가지 장점이 있다. 식물은 인간 병원체에 감염되지 않으며, 박테리아와 달리 내독소를 생산하지 않는다. 엽록체 형질전환은 핵 형질전환에 비해 안정적으로 많은 유전자를 발현시킬 수 있는 등 다양한 이점이 있다. 항균펩타이드(AMP)는 많은 동물들이 가지고 있는 선천면역의 일종으로, 소량이라도 항균력을 가지며, 기존 항생제와 다르게 쉽게 내성균이 생기지 않는다. 항균펩타이드인 moricin은 누에나방의 한 종류인 Bombyx mori에서 분리되었으며, C-말단은 염기성 아미노산이 모여 있고, N-말단은 α-helix 구조를 가지고 있다. Moricin을 생산할 때 SUMO와 6xHis tag를 융합하여 사용하였다. 발현된 moricin의 용해성과 안정성을 높이기 위해 SUMO를, 발현된 moricin을 정제하기 위하여 6xHis tag를 이용하였다. 본 연구에서 담배 엽록체와 대장균에서 항균펩타이드를 발현하기 위한 형질전환벡터를 제작하였다. 또한, 엽록체와 박테리아의 전사 및 번역의 유사성을 이용하여 대장균에서 단백질의 발현을 확인하였다. 발현된 moricin을 Ni 컬럼 및 SUMOase를 처리하여 정제하고 agar diffusion assay를 이용하여 항균 활성을 확인하였다.

• KSBB Journal
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• 제15권3호
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• pp.219-225
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• 2000

Bacillus thuringiensis를 생물농약으로 이용하기 위해서는 고농도의 포자형성에 의한 높은 살충성의 8 -endotoxin를 생산하는 것이 중요하다. 따라서 본 연구에서는 B. thuringiensis의 고농도 배양에 의한 높은 포자형성 수율을 얻기 위하여 40%의 용존산 소량과 $28^{\circ}C$에서 여랴 가지 방법의 유가식 배양법을 검토하였다. 최종 배양액의 glucose 농도가 50 g/L가 되도록 하는 경우의 유가식 배양에서는 continuos fed-batch culture의 linear gradient f feeding 법에 의하여 $9.37{\times}109$ cell/mL의 최대 생존 세포 수와 8 8.33 X 109 spore/mL의 최대 포자 수를 얻었으며, 이때의 포자 형 성율은 88.9% 이었다 최종 배양액의 glucose 농도가 100 m/L가 되도록 하는 경우의 유가식 배양에서는 intermittent fed-batch culture의 pH-stat 법에 의하여 $1.35{\times}1010$cell/mL의 최대 생존 세포 수와 $1.35{\times}1010$ spore/mL의 최대 포자 수를얻었으며, 이 때의 포자 형성율은 97.8% 이엇다.

• 한국환경과학회지
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• 제5권5호
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• pp.555-560
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• 1996

미생물 세포벽의 독성 물질 (내독소)을 incubator, centrifuge와 UV-Vis spectrophotometer를 이용하여 추출한 후 지방산 조성을 Gas Chromatography로 분석하였으며 독성 물질의 치사 독성과 발열 활성을 측정, 그 결과를 서로 비교하였다. 지방산 조성 분석 결과 V. vulnificus의 독성 물질 (내독소)에 대해 tetradecanoic acid가, E. coli게 대 해서는 dodecanoic acid, 그리고 S. typhimurium에 대해서는 decanoic acid가 주 지방산이었으며 이들 세 지방산이 세가지 독성 물질에서 대부분을 차지하였다 (70% 이상). 지방산 조성에서 특이한 점은 tetra-decanoic acid가 V. vulnificus에 대해 주 지방산이라는 점과 세가지 독성 물질 모두 hexadecanoic acid의 양이 매우 적다는 것이다 (2% 이하). V. vulnificus의 독성 물질의 mice 에서의 치사 독성 (LD50이 52.5 mg/kg)은 E. soli(56.5mg/kg)와 비슷하였으나 S. typhimunum (37.5mg/kg) 보다는 약하였으며 V. vulnificus의 독성 물질의 rabbit에서의 발열 활성은 E. coli보다 강하였으나 5S typhimurium 보다는 약하였다.