• 미생물학회지
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• 제39권4호
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• pp.242-247
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• 2003

Murine encephalomyocarditis virus (EMCV)는 혈장유래의약품의 바이러스 안전성 검증을 위해 hepatitis A virus (HAV)의 모델 바이러스로 사용되어왔다. 근래에 혈액응고인자제제에 의한 HAV 감염사례가 보고되면서 혈장유래의약품의 HAV 안전성 검증에 대한 국제적인 규제가 강화되어가고 있다. 본 연구에서는 HAV와 EMCV의 바이러스 불활화 공정에 대한 민감도를 평가하여, 혈장유래의약품 제조공정에서 HAV 불활화 공정의 검증법을 표준화하고자 하였다. HAV와 EMCV의 바이러스 불활화 공정에 대한 민감도를 평가한 결과 HAV가 60$^{\circ}C$ 열처리, low pH 처리(pH 3.9), 0.1 M NaOH 처리, 동결건조 공정 모두에서 EMCV보다 더 저항성이 큰 것을 확인할 수 있었다. EMCV는 특히 열처리와 0.1 NaOH 처리에 민감하게 불활화 되었지만, HAV는 큰 저항성을 나타내었다. 열처리의 경우 2시간 안에 EMCV는 검출한계 이하로 감소하였지만, HAV는 5시간 후에 검출한계 이하로 감소하였다. 0.1 M NaOH 처리시 EMCV는 15분 안에 검출한계 이하로 감소하였지만, HAV는 120분 정도의 처리에도 감염성 바이러스가 검출되었다. pH 3.9에서 25$^{\circ}C$로 14일 동안 항온하였을 때 HAV와 EMCV의 log 감소인수는 각각 1.63, 3.84이었다. 또한 혈액응고 8인자 제조공정의 동결건조 과정에서 HAV와 EMCV의 log 감소인수는 각각 1.21, 4.57이었다. 이와 같은 결과는 혈장유래의약품 제조공정의 HAV 불활화 또는 제거 검증시 모델 바이러스로 사용된 EMCV의 검증 결과를 해석함에 있어 보다 신중함을 가져야 한다는 것을 보여준다. 또한 보다 정확한 HAV검증 결과를 얻고자 한다면 모델 바이러스인 EMCV 보다 HAV를 사용하는 것이 보다 더 타당하다고 사료된다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제7권6호
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• pp.340-346
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• 2002

The purpose of the present study was to examine the efficacy and mechanism of the PAB (para-amino benzamidine) affinity column chromatography, Viresolve NFP virus filtration, pasteurization (60$\^{C}$ heat treatment for 10 h), and lyophilization steps employed in the manufacture of urokinase from human urine as regards the removal and/or inactivation of the hepatitis A virus (HAV). Samples from the relevant stages of the production process were spiked with HAV and subjected to scale-down processes mimicking the manufacture of urokinase Samples were collected at each step, immediately titrated using a 50% tissue culture infectious dose (TCID$\_$50/), and the virus reduction factors evaluated. PAB chromatography was found to be an effective step for removing HAV with a log reduction factor of 3.24. HAV infectivity was rarely detected in the urokinase fraction, while most of the HAV infectivity was recovered in the unbound and wash fractions. HAV was completely removed during the Viresolve NFP filtration with a log reduction factor of $\geq$ 4.60. Pasteurization was also found to be an effective step in inactivating HAV where the titers were reduced from an initial titer of 7.18 log$\_$10/ TCID$\_$50/ to undetectable levels within 10 h of treatment. The log reduction factor achieved during pasteurization was $\geq$ 4.76. Lyophilization revealed the lowest efficacy for inactivating HAV with a log reduction factor of 1.48. The cumulative log reduction factor was $\geq$ 14.08. Accordingly, these results indicate that the production process for urokinase exhibited a sufficient HAV reducing capacity to achieve a high margin of virus safety.

• 한국동물위생학회지
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• 제16권1호
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• pp.20-33
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• 1993

The studies were conducted to determine the effects of preservatives such as sodium chloride, sodium nitrite, sodium benzoate and sorbic acid on the survival of L. monocytogenes with regard to interaction of temperature, heat and pH of the medium. Inactivation of L. monocytogenes Scott A was more predominent by combination of sodium chloride and the other preservatives than sodium chloride alone, and inactivation was more exhilarated at $4^{\circ}C$ than at $35${\circ}C.$ The organism was not inactivated when sodium chloride, sodium nitrite, sodium benzoate and sorbic acid were added to 3%, 100ppm, 0.1, or lower, respectively, but was inactivated in the concentration increased twice. In TSB(tryptic soy broth) at pH 5.0 or lower, the organism did not grow regadless of the kinds of preservatives, and inactivation effect particularly was prominent in the presence of sodium nitrite and sorbic acid. On the other hand, at pH 6.0 or higher L. monocytogenes gradually increased in numbers and the effects of inhibition was higher in the presence of sorbic acid than in the other preservatives. When the preservatives were added to the concentration commonly used, incubation in TSB at $4^{\circ}C$ gradually resulted in growth of the bacterium and the organism rapidly decreased in numbers at $20^{\circ}C\; or\; 35^{\circ}C$ after incubation for 1 week. When L. monocytogenes was inoculated in TSB containing various preservatives and heated at $55^{\circ}C$ for 30minutes, the organism decreased in numbers at all preservatives. Particularly, viability rate of the organism was the lowest as 0.07% in the presence of sorbic acid.

• Journal of Plant Biotechnology
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• 제42권1호
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• pp.43-48
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• 2015

바이로이드에 감염된 과수의 생산성 하락으로 인한 농업인의 소득이 점점 감소하고 있는 실정이다. 감에 감염이 가능한 바이로이드는 PVd(Persimmon viroid)와 CVd(Citrus viroid) 등이 존재하는 것으로 알려져 있다. 따라서 본 연구에서는 전라남도 감 재배 농가에서의 감 바이로이드 감염 현황을 확인하였고 열처리, 한냉처리, 항바이러스제 처리를 통해 무병화 효율성을 알아보았다. 전라남도 나주, 영암, 담양, 순천, 구례의 20농가에서 감 부유 품종 198개체를 샘플링하였다. RT-PCR 실험을 통해 감 바이로이드의 감염률을 확인해 본 결과 CVd 41%, PVd 34% 로 확인되었으며 이미 알려진 수준에 비해 높게 감염되어 있음을 알 수 있었다. 무병화 효율 실험은 PVd와 CVd가 감염된 감을 이용하였으며 처리 방법에 따라 열처리($38^{\circ}C$), 한냉처리($4^{\circ}C$), 항바이러스제(Ribavirin) 단독 처리 그룹과 열처리와 항바이러스제를 동시에 처리한 그룹 및 한냉처리 및 항바이러스제를 동시에 처리한 그룹으로 나누었으며 처리 시간에 따라 2주와 4주로 나누어 각각의 바이로이드 제거 효율을 확인하였다. 한냉 처리한 그룹의 경우 생존율이 100%에 무병화 효율 또한 67%의 높은 제거 효율을 확인할 수 있었고, 한냉 처리와 항바이러스제를 2주간 복합처리한 군에서도 67%의 높은 효율을 확인하였다. 결론적으로 한냉 처리가 가장 높은 무병화율을 보여 주었으며 본 연구결과를 통해 감 무병화 연구에 좋은 자료가 될 것으로 판단된다.

• 한국막학회:학술대회논문집
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• 한국막학회 1996년도 추계 총회 및 학술발표회
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• pp.18-21
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• 1996

Virus and DNA removal in bio-drug manufacturing processes has received a great deal of attention in recent years. Removing of a virus using a membrane process is a promising method, because inactivated virus can be removed from the bio-drug and the process can be used as an additional and security inactivation after the method of general heat-inactivation of the virus in the bio-drug. The FDA and the biopharmaceutical industry have recently announced strict guidelines for impurities of virus and DNA contamination. The regulatory guidelines on residual amounts of DNA in mammalian cell culture products require DNA contamination of less than 100 pg/dose. Therefore, permeation and rejection of DNA through the porous membranes have become important in the application of DNA removal in bio-drug manufacturing using membrane technology. In this study, the permeation of DNA and chromatin through regenerated cellulose hollow fibers that have a mean pore diameter of 15 nm was investigated.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.615-618
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• 2000

The purpose of present study was to examine the efficacy of PAB (para-amino benzamidine) affinity column chromatography, pasteurization ($60^{\circ}C$ heat treatment for 10 h), and lyophilization steps, employed in the manufacture of urokinase from human urine, in the removal and/or inactivation of urine-born viruses. Bovine herpes virus (BHV) and Murine encephalomyocarditis virus (EMCV) were selected for this study. Samples from the relevant stages of the production process were spiked with the viruses and the amount of virus in each fraction was quantified by 50% tissue culture infectious dose ($TCID_{50}$). BHV and EMCV were effectively partitioned from urokinase during PAB chromatography with the log reduction factors of 6.71 and 5.27, respectively. Pasteurization was a robust and effective step in inactivating BHV and EMCV, of which titers were reduced from initial titers of $8.65\;log_{10}\;TCID_{50}$ and $7.81\;log_{10}\;TCID_{50}$, respectively, to undetectable levels within 1 hour of treatment. The log reduction factors achieved during lyophilization were 2.06 for BHV and 4.54 for EMCV. These results indicate that the production process for urokinase has sufficient virus reducing capacity to achieve a high margin of virus safety.

• 대한의생명과학회지
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• 제23권3호
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• pp.230-237
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• 2017

Changes in the activity of alkaline phosphatase (ALP) isoenzymes and isoforms in human serum have a major diagnostic value, therefore the regulation of ALP activities is a valuable target for therapeutic interventions. To assess the pharmacological activity of retinoids, i.e., all-trans retinoic acid and 13-cis retinoic acid, their tissue-specific inhibitory effect on human serum ALP activity was elucidated by chemical inhibition methods, heat-sensitive inactivation, and wheat-germ lectin precipitation test. Retinoids showed significant inhibition of the total ALP activity in human serum at a concentration of 5 mM. All-trans retinoic acid (5 mM) and 13-cis retinoic acid (5 mM) inhibited ALP activities by up to 12% and 15%, respectively, compared to that by guanidine hydrochloride (200 mM). L-phenylalanine (100 mM) and urea (30 mM) had no further inhibitory effect on ALP activities in human serum pretreated with retinoids (5 mM). Retinoids significantly inhibited ALP activities by up to 20% compared with that of tetramisole (30 mM). The ALP activities in retinoid-pretreated serum remained unchanged after the heat inactivation process. These results suggest that retinoids are inhibitors of the intestinal ALP isoenzyme. Remarkably, retinoids revealed potent inhibitory activities against ALP in wheat-germ lectin precipitant serum, indicating that they also function as inhibitors of the bone ALP isoform. The results show that retinoids inhibit the specific tissue-derived human serum ALP activities, moreover, the inhibitory effect of retinoids against bone ALP activity suggests their clinical utility as monitoring and prevention of metastasis of bone cancer.

• 한국식품조리과학회지
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• 제14권5호
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• pp.492-497
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• 1998

The stabilizing effect of fructooligosanharide (FO) on hen＇s egg yolk immunoglobulin (yIgG) by heat and acid was investigated. The heat stability of yIgG at 70∼80$^{\circ}C$ was enhanced in a concentration-dependent manner by adding 0∼50% (w/v) FO to a yIgG solution. Acid-induced inactivation of yIgG was also suppressed in a concentration-dependent relationship by addition of FO. Addition of 50% FO almost completely stabilized yIgG at pH 3. The remarkable stablizing effect of FO on yIgG may enhance the use of yIgG as functional food ingredients.

• 한국축산식품학회지
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• 제32권6호
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• pp.713-717
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• 2012

The objective of this study was to evaluate the effect of sodium habituation on thermal resistance of Staphylococcus aureus in various ready-to-heat (RTH) sauces. The strain mixture of S. aureus strains KACC10768, KACC10778, KACC11596, KACC13236 and NCCP10862 was habituated up to 9% of NaCl. The inocula of NaCl-habituated and non-habituated S. aureus were inoculated in 5 g portions of pork cutlet, meat and Carbonara sauces at 7 Log CFU/g, and the samples were vortexed vigorously. The inoculated samples were then exposed to 60 and $70^{\circ}C$ in a water-bath, and survivals of total bacteria and S. aureus were enumerated on tryptic soy agar and mannitol salt agar, respectively, every 30 min for 120 min. At 60oC, the cell counts of total bacteria and the significant difference in survivals between sodium-habituated and non-habituated S. aureus were observed only in the Carbonara sauce; the tailing effect, which is the period of no reduction of bacterial cell counts, was observed in pork cutlet, meat and Carbonara sauces subjected to $60^{\circ}C$. At $70^{\circ}C$, total bacterial populations and sodium-habituated and non-habituated S. aureus cell counts in meat and Carbonara sauce also significantly decreased (p<0.05) after 30 min of heat treatment, followed by the obvious tailing effect. Sodium-habituated S. aureus cell counts in meat and Carbonara sauces were higher (p<0.05) than those of non-habituated S. aureus at $70^{\circ}C$. The results indicate that sodium habituation of S. aureus cells may increase the thermal resistance of the pathogen in RTH sauces; moreover, heating RTH sauces for a short time before serving may not sufficiently decrease the cell counts of S. aureus, particularly for sodium-habituated strain.

• Food Science and Biotechnology
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• 제15권6호
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• pp.838-843
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• 2006

The chemical processes involved in the formation of green pigment in crushed garlic cloves were investigated based on the principle of pink pigmentation in macerated onions. Intact greening and non-greening garlic cloves were either left untreated or heated at $90^{\circ}C$ for 3 min to inactivate enzyme activities. First, a colorless ether soluble compound referred to as color developer reacted with glycine (among all free amino acids) in garlic to form a second compound insoluble in ether. The latter compound then reacted with formaldehyde to yield the green colored pigment. Alliinase activity was necessary for the production of color developer and for the development of green pigment. In greening garlic that had been heat treated, green pigmentation did not proceed due to the heat-inactivation of alliinase, but the addition of alliinase solution into the garlic homogenates restored the pigmentation. However, this phenomenon was not observed in non-greening garlic with or without heat treatment. Finally, the mechanism of green pigment formation in crushed garlicis similar to that of pink pigment formation in macerated onions.