• Proceedings of the Korean Society of Applied Pharmacology
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• 1995.04a
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• pp.114-114
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• 1995

이 연구에서는 혈전증 치료에 사용되는 혈전 응해제를 국내 독사독으로부터 개발하기 위한 실험을 실시하였다. 그 내용을 요약하면 다음과 같다. 국내에 서식하는 독사인 Agkistrodon blomoffi brevicaudus, Agkistrodon caliginosus와 Agkistrodon saxatilis에서 각각 사독을 채취하여 fibrin plate 방법으로 fibrin 분해능을 조사하여 Agkistrodon blomoffi brevicaudus의 독이 분해능이 가장 우수함을 밝혔다. 이와 같은 사실에 기초하여 A. blomoffi brevicaudus의 독으로부터 p-Aminobenzamidine affinity chromatography와 DEAE ion-exchange chromatography를 이용하여 분자량이 50.8 kDa인 황성 단백질을 정제하였다. 위에서와 같은 방법으로 정제한 단백질은 fibrin 분해능이 우수하고 fibrinogen의 ${\gamma}$ chain은 분해하지 않으나 B$\beta$ chain을 $A\alpha$ chain에 비하여 보다 선택적으로 분해하는 단백분해 효소임을 증명하였다. 이 정제 효소의 Fibrin에 대한 분해능은 266$\mu\textrm{g}$/${\mu}\ell$의 농도에서 Plasmin 1.0 unit (=3.0 WHO unit)보다 높게 나타났다. 정제된 효소는 chromogenic substrate인 N-Benzoyl-Phe-Val-Arg-pNA와 N-p-Tosyl- Gly-Pro-Arg-pNA의 arginine carboxyl side를 분해하고 pH 7.5에서 최대 활성을 보이며, Vmax는 5.46 umo1/1ㆍmin이고, Km 값은 0.20mM이며, 그리고 Cu$^{2+}$, $Zn^{2+}$, soybean trypsin inhibtor에 의해 25～50% 정도, serine proteinase inhibitor인 phenylmethylsulfonyl floride에 의해 80%정도 활성이 억제되는 특성이 있음을 규명하였다.

• Microbiology and Biotechnology Letters
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• v.15 no.2
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• pp.135-139
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• 1987

Aspergillus sp. MK-24 was cultured at 3$0^{\circ}C$ in the medium consisting of 2% glucose, 0.2% NaNO$_3$, 0.02% $K_2$HPO$_4$, 0.02% MgSO$_4$ 7$H_2O$, 0.02% KCl, and at initial pH of 5.0. The production of the inhibitor on venom proteinase reached to the maximum in 7 days. Sodium nitrate or potassium nitrate as a nitrogen source was favorable. The production of inhibitor was not affected by the addition of most of the inorganic salt used but depressed by lead, zinc, cobalt, mercuric or silver salts.

• The Korean Journal of Pharmacology
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• v.12 no.2 s.20
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• pp.33-41
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• 1976

Agkistrodon halys (Crotalidae) is the only species of poisonous snakes in Korea, and is divided into three subspecies; Agkistrodon bromhoffii brevicaudus, Agkistrodon calaginosus and Agkistrodon saxatilis. With the three venoms, the pharmacological actions on the cardiovascular system and intestine as well as some toxicological characteristics were studied. In addition, the precipitin test in an agar gel medium was employed for immunological comparison of the venoms and the sera of envenomed patients. The results obtained were as follows: Lyophilized venoms contained solids of $211{\sim}273mg/ml$, and LD50 to mice were 1.73 and 0.86 mg/kg in venoms of Agkistrodon bromhoffii brevicaudus obtained on July and October respectively, and 0.40 and 0.32 mg/kg in Agkistrodon calaginosus and the venoms of Agkistrodon saxatilis obtained on October was 2.29 mg/kg. Isoelectric focusing of lyophilized snake venoms showed 19 to 22 protein fractions and 2 to 3 isoamylase fractions. Acute irreversible hypotension was caused by the intravenous injection of large doses of venoms in rabbits and cats, but at the small doses, acute hypotension followed by slow recovery. Little changes of cardiac movements by the venom injection despite of marked hypotension were showed except bradycardia and arrhythmia prior the death. Also no changes on the isolated rabbit atria by the snake venoms were noted. The hypotensive effect of the snake venoms was prevented by the bilateral vagotomy or atropine pretreatment (1 mg/kg), but they did not affect when already the hypotension has undergone. In the isolated rabbit duodenum, small doses of venom increased the phasic movement, while large doses decreased after spastic contraction. With the injection of venoms in dog, strong contraction of gall-bladder was caused and it was not blocked by the pretreatment with phenoxybenzamine (10 mg/kg) or atropine (1.4 mg/kg). In the venoms of Agkistrodon bromhoffii brevicaudus and Agkistrodon calaginosus, at least 5 antigenic components were detected, and four of them were shared in common with each other. Polyvalent antivenin (Wyeth Lab. USA) had three common precipitating antibodies with the venom of Agkistrodon bromhoffii brevicaudus and Akistrodon calaginosus. In the serum of envenomed patients, no precipitating antibodies were seen to the venoms and little changes in serum protein, GOT and GPT were observed. In conclusion, the snake venoms obtained in Korea were highly toxic and caused chiefly the vascular collapse leading to death. This vascular collapse was resulted largely by cholinergic effects, and not cardiotoxin of venoms. In human, it is likely that precipitating antibodies to venom were not produced by an envenomed incidence to poisonous snakes.

• Korean Journal of Veterinary Service
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• v.32 no.3
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• pp.201-207
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• 2009

Porcine transmissible gastroenteritis virus (TGEV), porcine epidemic diarrhea virus (PEDV) and rotaviruses are considered as the most important causative agents of diarrhea in piglets. The study established 3 method vaccination programs to prevent PEDV. A (LL)group inoculated twice vaccinations on 2-weeks-interval during the late term of pregnant sows with PEDV live vaccine. The B (LKK) group was applied that one time single PEDV live vaccine at the pre-mate followed by the TGEV PEDV combined inactivated vaccine (twice vaccination on 2-weeks interval at the third-trimester). C (KK) group was applied to sow which inoculated twice vaccination on 2-weeks-interval during the late term of pregnant sows with by the TGEV, PEDV combined inactivated vaccine. As the result of SN test on sows in the pig farm before vaccination, antibody titers was showed 9/45 (20.0%). By comparison with the serum neutralizing antibody titers against PEDV of the vaccination programs after PEDV of the vaccination, A group and B group vaccination method was higher than those of C group in sows. In the piglets up to 2 weeks of age, A group was showed antibody titers of 17/22 (81.8%) that showed 2-128, and B group was showed antibody titers of 30/37 (81.1%) that showed 2-512, and C group was showed antibody titers of 14/28 (50.0%) that showed 2-32. On the other hand, PEDV antibody titers were tested for the survey. As the results of SN test, Aujeszky's disease survey in 54 pig farms from november 2005 to august 2006, antibody titers of 47/286 (16.4%) showed above 2. Five breeding farms were antibody titers of 38/77 (49.4%), Wanggung zone farms antibody titers of 59/85 (69.4%). In pigs farms vaccinated the first of twice PEDV live vaccine, and after 6 month, the second of twice TGEV PEDV combined inactivated vaccine (LLKK, 256-1024 titer) method was higher than those of vaccinated twice the early term of pregnant, and twice the late term of pregnant sows of PEDV live vaccine (LLLL, 32 titer).

• Microbiology and Biotechnology Letters
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• v.15 no.2
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• pp.129-134
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• 1987

Aspergillus sp. (MK-24) producing a biological active substance that inhibited the venom proteinase activity was isolated from soil. The substance also inhibited the activity of trypsin and coagulation of blood, but did not inhibit papain, $\alpha$-chymotrypsin and pepsin. The substance was partially purified from culture filtrate by precipitaion with acetone, and by chromatography of DEAE-Sepadex A-50 column and Amberlite IRC-50 ion exchange. The inhibitory substance was stable in the wide pH range from 2.0 to 12.0 at 37$^{\circ}C$, but not stable at $65^{\circ}C$ in the alkaline pH. Only 12% of the activity was decreased by the heat treatment at 10$0^{\circ}C$ for two hours. The inhibition on venom proteinase (Agkistrodon bromohoffi brevicaudus) was a mixed type. The inhibitory activity depended on the preincubation time and completely depressed by cupric, zinc and cobalt ions. The inhibition on the venom proteinase was appeared strongly on casein but not on ovalbumin or hemoglobin as a substrate.

• Journal of Acupuncture Research
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• v.32 no.1
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• pp.79-88
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• 2015

Objectives : The purpose of this research was to investigate the cytotoxic effect of Natural Killer(NK)-92 cell and Snake Venom, and to elucidate its mechanism on human lung carcinoma cell A549. Methods : In order to figure out whether Snake Venom enhances the cytotoxic effect of NK-92 cell in A549 cell, Cell Viability Assay was conducted. Also, in order to observe the changes of Caspase-3 and Caspase-8, both of which are proteinases that advance apoptosis, and the changes of TNRF and DR3, which are Death Receptors of the extrinsic pathway of apoptosis, Western Blot Analysis was conducted. By conducting RT-PCR analysis, we have tried to confirm Perforin, Granzyme B, and GADPH, all of which are cytotoxic-related proteins. Lastly, in order to observe the effect of Snake Venom on NO formation within human lung carcinoma cells, NO determination was conducted. Results : 1. After conducting Cell Viability Assay, Snake Venom enhanced the cytotoxic effect of NK-92 cell and inhibited the growth of A549. 2. Western Blot Analysis caused proteinases Caspase-3 and Caspase-8, which advance apoptosis, to increase in the combined treatment group, but not in treatment groups that focused only on either Snake Venom or NK-92 cell in A549 lung carcinoma cells. 3. Western Blot Analysis caused an expression of TNFR2 and DR3, both of which are Death Receptors of the apoptosis extrinsic pathway, in the combined treatment group, but not intreatment groups that focused only on either Snake Venom or NK-92 cell in A549 human lung carcinoma cells. 4. After conducting NO determination, NO formation within A549 cell showed no significant changes in both treatment groups that focused NK-92 cell and combined treatment group. 5. After conducting RT-PCR, the expression of Granzyme B and Perforin, which are cytotoxic-related proteins within A549 human lung carcinoma cells, showed growth in the combined treatment group, but not the treatment group that focused only on NK-92 cell. Conclusion : It has been indicated that, when it comes to the A549 cell, Snake Venom enhances the increase of Death Receptor expression and continuous apoptosis reaction, leading to the enhancement of the cancer cell cytotoxic effect of the NK-92 cell. It is expected that Snake Venom can be used with the NK-92 cell for further lung cancer treatment.

• Journal of Practical Agriculture & Fisheries Research
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• v.20 no.1
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• pp.89-96
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• 2018

The object of this study was to evaluate the change of antibody titers on virus strains after inoculation with commercial killed equine influenza (EI) vaccines in horses. Serum antibodies of 20 Thoroughbred yearlings were detected using hemagglutination inhibition test for 41 weeks. Second vaccination is inoculated 4 weeks after the initial vaccination. Most of antibody titers were not increased until 4 weeks after first vaccination. The highest titers were detected 6-10 weeks after vaccination. The titers were decreased slowly and maintained for 16 weeks after inoculation. We could barely detect the antibody 41 weeks after vaccination in most cases. Vaccine anergia were appeared in 3 horses (15%) but it depended on virus strains. A/Equine/La Plata/93(H3N8) strain that induce high and durable antibody responses was the most effective among three strains. This study presents the first comprehensive data on the endurance of antibody titers against EI. Our data also suggests that yearlings should be inoculated three times in order to maintaining optimal antibody titers against EI. We speculate the causes of anergia were vaccine break down or individual specificity. Further research is needed to investigate immunological unresponsiveness. This was the first study on strain of equine vaccine in Korea.

• The Journal of Dong Guk Oriental Medicine
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• v.6 no.1
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• pp.177-203
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• 1997

Various kinds of medicinal herbs and prescribed herb medicines which have usually been used for treatment of reducing fever, purging intense heat, and detoxication were secreened for antibacterial activity against Proteus vulgaris, S. aureus, and E. faecalis which causing puerperal infection. Minimal inhibitory concentrations of water and ethanol-soluble extracts of Hwangyon(C. japonica), Omae(P. mume), and Omiza(S. chinensis) were determined and using thin-layer chromatography the extracts of Hwangyon and Omiza were separated. The results obtained were as follow : 1. The water-soluble extracts of Keumeunhwa(L. japonica), Chiza(G. jasminoides), Hwangyon(C. japonica), Dansam(S. miltiorrhiza), Omae(P. mume) and Omiza(S. chinensis) showed antibacterial activities against P. vulgaris ; among them, the extracts of Hwangyon, Omae, and Omiza showed high antibacterial activities. The water-soluble extracts of Keumeunhwa(L. japonica), Hwangyon(C. japonica), Hwangkeum(S. baicalensis), Hwangbaek(P. amurense), Dansam(S. miltiorrhiza), Daehwang(R. coreanum), Omae(P. mume), Omiza(S. chinensis), Gingseng(P. ginseng), and Gamcho(G. uralensis) showed antibacterial activities against S. aureus : among them, the extracts of Hwangyon and Omiza showed high antibacterial activities. The water-soluble extracts of Banggi(C. trilobus), Daehwang(R. coreanum), Omae(P. mume), Omiza(S. chinensis), and Gamcho(G. uralensis) inhibited the cell growth of E. faecalis : among them, Omae and Omiza showed high antibacterial activities. The extract of Hwangyon(C. Japonica) and Omae(P. mume) showed no inhibition against E. coli. However, the extract of Omiza(S. chinensis) inhibited cell growth of both E. coli and B. subtilis. 2. The water- and ethanol-soluble extract of Hwangyon(C. Japonica) showed antibacterial activities against P. vulgaris and S. aureus, those of Omae(P. mume) against P. vulgaris and E. faecalis, and those of Omiza(S. chinensis) against all species tested, P. vulgaris, S. aureus, and E. faecalis. With the exception that ethanol-soluble extract of Hwangyon(C. japonica) showed much higher antibacterial activity against S. aureus than water-soluble one, antibacterial activities of both water-and ethanol-soluble ones were similar to each other of other two medicinal herbs. 3. When the prescribed herb medicines were tested, Sambohwan showed antibacterial activities against P. vulgaris and S. aureus. No prescribed herb medicine inhibitied the cell growth of E. faecalis. 4. Minimal inhibitory concentrations(MICs) of both water- and ethanol-soluble extracts of Hwangyon(C. japonica), Omae(P. mume), and Omiza(S. chinensis) against P. vulgaris were $2.5mg/m{\ell}$, $10mm{\ell}$, and $20mg/m{\ell}$, respectively. Those of both Hwangyon(C. japonica) and Omiza(S. chinensis) against S. aureus were $1.25mg/m{\ell}$ and $10mg/m{\ell}$, respectively. MICs of water-soluble extracts of Omae(P. mume) and Omiza(S. chinensis) against E. faecalis were $2.5m{\ell}$ and $5m{\ell}$, and those of ethanol-soluble extracts were $5m{\ell}$ and $10m{\ell}$, respectively Except for those of E. faecalis the cell growth of P. vulgaris and S. aureus were inhibited by much lower concentration of ethanol-soluble extracts used. As a result, the antibacterial compounds against P. vulgaris S. aureus, and E. aecalis are contained in the extracts of Hwangyon(C. japonica), Omae(P. mume), Omiza(S. chinensis), the prescribed herb medicine, Sambohwan, and might be used for treatment of puerperal infection. Further study should be carried out to identify which compounds affect the cell growth inhibition of P. vulgaris S. aureus, and E. faecalis.