• 한국식품과학회지
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• 제29권1호
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• pp.26-31
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• 1997

물을 극미량 함유하는 미수계 효소반응계인 aerosol-OT/이소옥탄으로 구성된 역미셀계내에서 레시친의 일종인 DPPC (dipalmitoyl phosphatidylcholine)의 가수분해반응을 phospholipase $A_2$가 촉매할 수 있었다. 또한 phospholipase $A_2$에 의한 가수분해반응을 정량분석하기 위한 고성능 액체 크로마토그래피에 의한 예민하고 간편한 방법을 고안하였는 바, 이에 의하여 aerosol-OT/이소옥탄 역미셀계내에서 phospholipase $A_2$에 의한 가수분해반응의 동력학적인 분석이 가능하였다. 한편, 여러가지 유화제와 유기용매 중 aerosol-OT와 이소옥탄일 때 효소반응이 가장 효과적이었으며, 반응 적정온도는 $35{\sim}40^{\circ}C$, 적정 pH는 7.0이었다. 또한 역미셀계내에서의 가수분해반응은 수분 함량변화에 민감하였으며, R-값(=[water]/[aerosol-OT])이 10.0일 때 가장 높은 효소활성을 나타내었다. 효소반응을 위한 최적조건하에서의 $K_{m,app.}$ 및 $V_{max.,app.}$는 각각 8.73 mM, 2.83 units/㎎ protein이었으며, 역미셀계내에서 phospholipase $A_2$에 의한 DPPC가수분해반응의 활성화에너지는 12.31 kcal/mole로 산출되었다.

• Journal of Yeungnam Medical Science
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• 제5권2호
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• pp.37-45
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• 1988

Phospholipase C는 second messenger로서 세포 밖의 signal transduction에 중요한 효소 이다. 본 연구에서 소의 뇌, 자궁 및 고환에서 high performance liquid chromatography(HPLC)를 이용하여 phospholipase C(PLC)를 추출하였으며 뇌에서 3개의 isozyme(F-1, F-2, F-3), 자궁과 고환에서 각각 2개의 isozyme을 얻었고 HPLC에서의 retension time을 구하였다. Homogeneity 검사를 위하여 소 brain의 각 isozyme I, II 및 III에 대한 PLC-monoclonal antibody(Mab)를 affigel에 label 시켰고 결합능(binding capacity)는 73.8~97.5%였다. PLC-Mab와 자궁 및 고환의 PLC isozyme과의 homogeneity 검사에서 binding capacity는 자궁의 F-1은 PLC III가 주이며 F-2는 DEAE column에서는 II가 주이나 phenyl column에선 I과 II가 주이고 고환에선 F-1은 PLC III가 주이고 F-2에서는 PLC II가 주인 것으로 나타났다.

• 분석과학
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• 제27권6호
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• pp.277-283
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• 2014

한외여과법을 이용하여 봉독의 알러지 원인성분인 $PLA_2$를 제거하기 위하여 압력, 농도, 분자크기의 영향을 조사하였다. 봉독의 주요성분 분자량을 바탕으로 한외여과막의 투과크기를 선정하고 농도와 압력을 달리하였다. 그 결과, melittin과 apamin 함량은 유지되면서 $PLA_2$를 제거하는 최적조건(1 mg/mL, 20 psi, 10,000 dalton)을 찾았으며, 이를 HPLC와 SDS-PAGE로 확인하였다.

• Journal of Acupuncture Research
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• 제34권1호
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• pp.31-38
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• 2017

Objectives : The purpose of this study was to analyze the components of the clinically used bee venom (BV) pharmacopuncture. Methods : Two kinds of bee venom pharmacopuncture (BV-I and II), three kinds of separate purification BV (SPBV) pharmacopuncture (SPBV-I, II, and III), and apitoxin were investigated in this study. We performed a component analysis of melittin, apamin, and phospholipase $A_2$ using high-performance liquid chromatography (HPLC). Results :1. BV-I contained approximately 40% more melittin than BV-II did. 2. In the three separate purification BV pharmacopuncture, SPBV-I, SPBV-II, and SPBV-III, phospholipase $A_2$ content decreased remarkably. 3. The melittin content in SPBV-I increased by 5% compared to that in BV-I. 4. The amount of melittin in apitoxin was similar to that in SPBV-I. Conclusion : The compositions of the BV pharmacopuncture and separate purification BV pharmacopuncture changed depending on the collection method and concentration. Therefore, it is necessary to choose the most suitable BV for each specific medical treatment target. Furthermore, research into the composition of BV may be needed for its safe and effective use.

• 동의생리병리학회지
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• 제18권5호
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• pp.1322-1330
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• 2004

This experimental studies were performed in order to prove the effect of Phospholipase A₂(PLA₂) Herbal-acupuncture by using rats that had neuronal damage due to the Middle Cerebral Artery Occulsion(MCAO). We observed the change of extracellular concentrations(μM) Of dopamine, DOPAC, HVA, HIAA, glutamate, aspartate, GABA, glysine, taurine, alanine, and tyrosine as extracted by vivo microdialysis, in the PLA₂ Herbal-acupuncture administrated rats(240-260g, Sprague-Dawley) subjected to the MCAO. The dialysates were extracted three times before the MCAO and six times after the MCAO every 20 minutes, and ana lysed by highperformance liquid chromatography(HPLC). PLA₂ Herbal-acupuncture significantly inhibited glutamate and tyrosine which are stimulant neurotransmitters at brain ischemia, and it significantly decreased glycine, GABA, taurine, and alanine which are inhibitory neurotransmitters at brain ischemia. PLA₂ Herbal-acupuncture may prevent delayed neuronal death(DND) in selectively vulnerable focal areas of the brain effectively.

• Applied Biological Chemistry
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• 제46권3호
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• pp.251-256
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• 2003

한국산 인삼을 60% acetone으로 추출하여 Sepadex LH-20 gel column chromatography, MCI-CHP 20 gel column chromatography, ${\mu}-Bondapack\;C_{18}$ gel column chromatography을 이용하여 polyphenol 분획물을 3종류를 분리하여 항산화 효과, phospholipase $A_2$ 저해효과 및 암세포 증식억제 효과를 검토하였다. 항산화 효과에서 분획물 I은 150 ppm에서 $Cu^{2+},\;KO_2$ 그리고 $H_2O_2$에 대하여 각각 48.16%, 79.71%, 43.55%를 나타내었고 DPPH 수소공여능은 분획물 II가 200 ppm에서 35.17%의 라디칼소거능을 보였다. 분획물 III는 $Fe^{2+}$, OH가 존재 시 150 ppm에서 48.49%, 25%의 항산화효과를 나타내었다. Phospholipase $A_2$ 저해활성은 분획물 III이 $60\;{\mu}/ml$에서 48.9%의 활성을 나타냈다. HT 29 cell에 대한 인삼의 세포 독성은 분획물 II에서 0.25 mg/ml에서 73.29%의 가장 높은 억제능을 나타내었다.

• Biomolecules & Therapeutics
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• 제4권2호
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• pp.148-153
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• 1996

Activation of the T lymphocytes results in a variety of early biochemical events ultimately leading to cell proliferation and lymphokine production. Stimulation of the signal transduction cascade in T cells through the T cell receptor coincides with activation of the phosphatidylinositol-phospholipase C (PI-PLC) pathway. Therefore, we have established a model system to screen immune-simulator that can increase the hydrolysis of phosphoinositides in human T cell leukemia Jurkat cells. As a result of screening from herbal medicine extract, 4 extracts (O1ibanum, Ephedrae Herba, Real Gar, Saussureae Radix) were found 14 increase the production of inositol phosphates. All the active fraction from the four kinds of extract were fluted in a different retention time on C-18 HPLC and these active fraction also showed difference in cell specificity. And all the active fractions increased DNA synthesis in T cell. Therefore, it is suggested that the active fraction among 4 extracts might contain a compound having different properties one another.

• BMB Reports
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• 제31권2호
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• pp.189-195
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• 1998

Methylmercury (MeHg), which is widely distributed in the environment, is well known for both its acute and chronic poisoning effects on the human health; however, the precise biochemical mechanisms by which this compound elicits its toxicity in a cellular level are still poorly understood. To examine whether MeHg-induced liver injury involves activation of Phospholipase $A_2$ ($PLA_2$), the $PLA_2$ activity of control and MeHg-administrated livers was measured. MeHg stably enhanced a liver form of cytosolic $PLA_2$ activity, which exhibited several biochemical properties similar to those of the 100 kDa $cPLA_2$, except in its elution profile of a DEAE-5PW HPLC, and it migrated as a molecular weight of 80 kDa in Western blot analysis. This blotting analysis also indicated that the MeHg-induced enhancement of the activity could be due to the increase in the amount of the enzyme protein rather than a stable modification of the enzyme such as phosphorylation. Our data also showed the higher myeloperoxidase activity in MeHg-administrated liver than in the control, suggesting that this increase in the amounts of the 80 kDa $PLA_2$ and its activity may be resulted from infiltration of neutrophils into the liver during a hepatic injury process such as MeHg-induced inflammation. Taken together, these data suggest that MeHg-induced liver injury may be mediated by activation of the 80 kDa form of liver cytosolic $PLA_2$.

• 대한약침학회지
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• 제10권2호통권23호
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• pp.81-86
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• 2007

Objectives : This study was conducted to carry out quantitative evaluation and safety of Sweet Bee Venom. Methods : Content analysis was done using HPLC, measurement of LD$^{50}$ was conducted intravenous, subcutaneous, and intramuscular injection to the ICR mice. Results : 1. According to HPLC analysis, removal of the enzymes containing phospholipase A2 was successfully rendered on Sweet Bee Venom. And analyzing melittin content, Sweet Bee Venom contained 12% more melittin than Bee Venom. 2. LD$^{50}$ of ICR mice with Sweet Bee Venom was more than 20mg/kg in subcutaneous injection and intravenous injection, between 15mg/kg and 20mg/kg in muscular injection. 3. LD$^{50}$ of ICR mice with Bee Venom was between 6 and 9mg/kg in subcutaneous injection and intravenous injection, and more than 9mg/kg in muscular injection. Conclusion : Above results indicate that Sweet Bee Venom was more safe than Bee Venom and the process of removing enzymes was well rendered in Sweet Bee Venom.

• The Korean Journal of Physiology and Pharmacology
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• 제20권2호
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• pp.177-183
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• 2016

We fortuitously observed a human neutrophil intracellular free-calcium concentration ($[Ca^{2+}]_i$) increasing activity in the commercially available phosphodiesterase I (PDE I), which is actually dried crude venom of Crotalus atrox. As this activity was not observed with another commercially available pure PDE I, we tried to find out the causative molecule(s) present in 'crude' PDE, and identified Lys49-phospholipase A2 (Lys49-PLA2 or K49-PLA2), a catalytically inactive protein which belongs to the phospholipase A2 family, by activity-driven three HPLC (reverse phase, size exclusion, reverse phase) steps followed by SDS-PAGE and LC-MS/MS. K49-PLA2 induced $Ca^{2+}$ influx in human neutrophils without any cytotoxic effect. Two calcium channel inhibitors, 2-aminoetoxydiphenyl borate (2-APB) ($30{\mu}M$) and SKF-96365 ($20{\mu}M$) significantly inhibited K49-PLA2-induced $[Ca^{2+}]_i$ increase. These results suggest that K49-PLA2 modulates $[Ca^{2+}]_i$ in human neutrophils via 2-APB- and SKF-96365-sensitive calcium channels without causing membrane disruption.