• Korean Journal of Food Science and Technology
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• v.29 no.1
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• pp.26-31
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• 1997

Bee venom (Apis mellifera) phospholipase $A_2$ solubilized in reversed micelles containing small amount of water stabilized by surfactant could catalyze the hydrolysis of dipalmitoyl phosphatidylcholine (DPPC). A sensitive and simple high performance liquid chromatographic (HPLC) methodology of phospholipase $A_2$ assay for the hydrolysis of DPPC was developed. Kinetic analysis of the phospholipase $A_2$-catalyzed reaction was found to be possible in reversed micelles. Among the surfactants and organic solvents tested, aerosol-OT and isooctane were most effective for the hydrolysis of DPPC in reversed micelles. Optimal temperature, optimal pH, $K_{m,app.},\;V_{max.,app.}$ and activation energy were determined to be $35{\sim}40^{\circ}C$, 7.0, 8.73 mM, 2.83 units/㎎ protein and 12.31 kcal/mole, respectively. The hydrolysis activity was dependent on water content and maximum activity was obtained at R value (=[water]/[aerosol-OT]) of 10.0.

• Journal of Yeungnam Medical Science
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• v.5 no.2
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• pp.37-45
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• 1988

Phosphoinositide-specific phospholipase C(PI-PLC) is a second messenger of signal transducer on cell membrane. In the previous study, PLC of bovine brain has been purified three isozymes. In this paper, uterus and seminal vesicle have been purified. Two peaks of PI-PLC activity were resolved when bovine uterus and seminal vesicle proteins were chromatographed on a DEAE and phenyl TSK 5PW HPLC column. Each two peak was compared with PI-PLC I, IT and ill from bovine brain and we got the retension time on HPLC. The peak fractions with PLC activity were tested homogeneity with brain PLC monoclonal antibodies(Mab). Mab-labeled affigels were bounded in the range of 73.8%~97.5% with PLC I, IT and III. Homogeneity of fractions were revealed that DEAE F-1 and phenyl F-1-I were highest level of PLC III in uterus and seminal vesicle and DEAE F-2 and phenyl F-2-I were mixed PLC I and II.

• Analytical Science and Technology
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• v.27 no.6
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• pp.277-283
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• 2014

The effects of concentration, pressure, and molecular sige on removing allergenic substance (phospholipase $A_2$) in bee venom by ultrafiltration were investigated. The membrane pore sizes were selected based on the molecular weight of the main compounds. The conditions of concentration and pressure were selected randomly. As results, we obtained the optimum condition (1 mg/mL, 20 psi, 10,000 dalton) for removing $PLA_2$ at constant concentration of melittin and apamin and confirmed the separation results by HPLC and SDS-PAGE.

• Journal of Acupuncture Research
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• v.34 no.1
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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.

• Journal of Physiology & Pathology in Korean Medicine
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• v.18 no.5
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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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• v.46 no.3
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• pp.251-256
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• 2003

The polyphenol fractions of Korean ginseng were purified using Sephadex LH-20, MCI gel, Bondapak $C_{18}$ TLC, and HPLC from the 60% acetone soluble fraction. Fraction I showed 48.16%, 79.71% and 43.55% inhibition at 150 ppm against lipid oxidation in the presence of copper ion, superoxide and hydrogen peroxidation. Electron donating abilities of fraction II showed 35.17% inhibition at 200 ppm. Fraction III showed 48.49% and 25% inhibition at 150 ppm against lipid oxidation in the presence of ferrous ion and hydroxy radical ion. The phospholipase $A_2$ inhibitory effect of fraction III was 48.9% at the concentration of $60\;{\mu}g/ml$. The cytotoxic effects of fraction II was the highest (73.29% at 0.25 mg/ml) among the tested polyphenol fractions.

• Biomolecules & Therapeutics
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• v.4 no.2
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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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• v.31 no.2
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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 $PLA_2$.

• Journal of Pharmacopuncture
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• v.10 no.2 s.23
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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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• v.20 no.2
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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.