• Journal of Pharmacopuncture
• /
• v.9 no.3 s.21
• /
• pp.61-77
• /
• 2006

Objectives : Sweet bee venom is made by removing allergen from the bee venom through gel filtration chromatography and propionic acid/urea polyacrylamide gel electrophoresis. The aim of this study was to verify allergy inhibitory action in Sweet Bee Venom in which the allergy causing enzyme is removed. Methods : 95 healthy adult men and women were selected through a survey whom had never received the bee venom therapy in the past. The concentration of bee venom pharmacopuncture and Sweet BV pharmacopuncture was equally at 0.1mg/ml and the experiment was conducted as the double blind test. Experiment groups were classified into low dosage groups(0.1ml for both bee venom pharmacopuncture and Sweet BV) and high dosage groups where 0.4ml of respective administrations were rendered made observations for allergic responses. Results : Participants of the study was comprised of 71 men and 24 women with the average age of 29.0 years. According to results of the low dosage groups, Sweet BV group showed significant reduction in pain after 4 hours and 24 hours compared to the bee venom pharmacopuncture group. Other allergic responses were insignificant between the groups. For the high dosage groups, Sweet bee venom group showed reduction in pain after 30 minutes and 4 hours. Other allergic responses such as edema, itchiness, dizziness from hypersensitivity, and fatigue were significantly lower in the Sweet bee venom administered group after 30 minutes. Conclusions : As a result of removed allergen, Sweet bee venom significantly inhibits allergic responses both locally and throughout the body. This indicates wider and easier application of Sweet bee venom for the symptoms applicable to the bee venom pharmacopuncture. Further comparative studies should be conducted to yield more objective verification.

• Journal of Pharmacopuncture
• /
• v.16 no.2
• /
• pp.46-54
• /
• 2013

Objective: This study was undertaken to isolate a fibrin(ogen)olytic enzyme from the snake venom of Gloydius blomhoffii siniticus and to investigate the enzymatic characteristics and hemorrhagic activity of the isolated enzyme as a potential pharmacopuncture agent. Methods: The fibrinolytic enzyme was isolated by using chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and fibrin plate assay. The characteristics of the enzyme were determined by using fibrin plate assay, protein hydrolysis analysis, and hemorrhage assay. Its amino acid composition was determined. Results: The fibrin(ogen)olytic enzyme with the molecular weight of 27 kDa (FE-27kDa) isolated from G. b. siniticus venom consisted of two heterogenous disulfide bond-linked polypeptides with the molecular weights of 15 kDa and 18 kDa. When more than $20{\mu}g$ of FE-27kDa was applied on the fibrin plate, fibrinolysis zone was formed as indicating its fibrinolytic activity. The fibrinolytic activity was inhibited completely by phenylmethanesulfonylfluoride (PMSF) and ethylenediaminetetraacetic acid (EDTA) and partially by thiothreitol and cysteine. Metal ions such as $Hg^{2+}$ and $Fe^{2+}$ inhibited the fibrinolytic activity completely, but $Mn^{2+}$ did not. FE-27kDa preferentially hydrolyzed ${\alpha}$-chain of fibrinogen and slowly hydrolyzed ${\beta}$-chain, but did not hydrolyze ${\gamma}$-chain. High-molecular-weight polypeptides of gelatin were hydrolyzed partially into polypeptides with molecular weights of more than 45 kDa. A dosage of more than $10{\mu}g$ of FE-27kDa per mouse was required to induce hemorrhage beneath the skin. Conclusion: FE-27kDa was a serine proteinase consisting of two heterogeneous polypeptides, hydrolyzed fibrin, fibrinogen, and gelatin, and caused hemorrhage beneath the skin of mouse. This study suggests that the potential of FE-27kDa as pharmacopuncture agent should be limited due to low fibrinolytic activity and a possible side effect of hemorrhage.

• Journal of Sericultural and Entomological Science
• /
• v.34 no.1
• /
• pp.41-48
• /
• 1992

Dissolution and degradation of the parasporal crystal proteins produced from B. thuringiensis var. kurstaki, B. thuringiensis var. dendrolimus and B. thuringiensis var. aizawai were investigated. SDS-polyacrylamide gel electrophoresis analysis showed that the crystals contained major protein with molecular weight of approximately 134 kDa for B. thuringiensis var. aizawai 143 kDa for B. thuringiensis var. kurstaki and 149 kDa for B. thuringiensis var. dendrolimus, respectively. Crystals of three other strains were incubated alkali solutions at various pH or gut juice of Silkwarm Bombyx mori, Fall webworm Hyphantria cunea, and Common Cabbage worm Pieris rapae. When crystals of these strains were solubilized by alkali solutions, no major differences among strains B. thuringiensis could be detected. Among the strains studied, crystal protins (130-66 kDa) consist of protease resistant polypeptides in the 45-66 kDa size range when treated with gut juice of three insect species.

• International Journal of Industrial Entomology and Biomaterials
• /
• v.6 no.1
• /
• pp.33-44
• /
• 2003

The arylphorin was purified from the pupal haemolymph of the Chinese oak silkmoth, Antheraea pernyi, and characterized physiologically and biochemically, The protein was purified by a simple preparative polyacrylamide gel electrophoresis (PAGE) and subsequent diffusive elution. The preparation was shown to be homogeneous by 7.5% native-PAGE. The native molecular weight of arylphorin was 450 kDa with a 80 kDa single subunit, suggesting hexamer, The protein contained high amounts (18.3%) of aromatic amino acids, phenylalanine (9.7%) and tyrosine (8.6%). Therefore, the protein was identified as a kind of a storage protein referred to as an arylphorin. The protein was stained by Schiff's reagent, suggesting a glycoprotein. The protein contained 4.9% (w/w) sugar and mannose and N-acetylglucosamine were major components. Also, degradation of the protein was begun by heat treatment at 90 for 20 minutes. These results showed that the A. pernyi arylphorin in the study is hexamer associated with the six subunits consisting of a 80kDa single subunit, and is different from that of Kajiura et al. (1998) in the subunit composition.

• Korean Journal of Pharmacognosy
• /
• v.37 no.4 s.147
• /
• pp.221-228
• /
• 2006

A lectin was purified from Serrognathus platymelus castanicolor larvae and named as SPL. The purification was carried out by ion-exchange chromatography on DEAE Sephadex A-50 and gel filtration chromatography on Sephadex G-200. The purity of the protein was verified by polyacrylamide gel electrophoresis and the purified lectin agglutinated erythrocytes of rabbit and human A, B, O, AB. SPL was tested it's ability to enhance the expressions of cytokines, $IL-1\alpha$, IL-2, IL-6, $TNF\alpha$ and $IFN\gamma$ by human peripheral blood mononuclear cells (PBMC) obtained from healthy donors. mRNA analyses were performed by RT-PCR at the moment of 1, 4, 8, 24, 48, 72 and 96 h after stimulation of PBMC with purified SPL. The patterns of IL-2 band were slightly expressed from 24 h and the strongest band was appeared at 96 h. The expressions of $IL-1\alpha$ and IL-6 mRNA were strong from 1 to 8 h and those of $TNF\alpha$ were from 48 to 96 h. The mRNA encoding $IFN\gamma$ were not detected. The addition of SPL for macrophage cultures induced production of nitric oxide (NO) by cells in a dose-dependent manner. NO release was partially inhibited by $TNF\alpha$ antibodies. These results suggest that SPL has the ability to enhance cytokine expressions in PBMC and to induce the NO release by TNFa in macrophage cultures from PBMC cultures.

• The Plant Pathology Journal
• /
• v.22 no.1
• /
• pp.51-62
• /
• 2006

Three strains of Cucumber mosaic virus (CMV) have been found to cause a lethal disease, referred to as fern leaf syndromes and mild mosaic symptoms in tomato (Lycopersicon esculentum Mill.) crops grown in Kuwait. CMV strains were detected and identified based on host range, symptomatology, serology, electron microscopy, and ribonucleic acid (RNA) electrophoresis in polyacrylamide gels. A high degree of viral genomic heterogeneity was detected among CMV strains isolated in Kuwait, with no apparent correlation to symptomatology in tomato host plants. Two different virus satellites of 'CMV associated RNA 5', designated CARNA 5, were detected in two virus strains that caused both lethal disease and mild symptoms, designated CMV-D1 and CMV-S1 respectively. CARNA5 was not detected in the third CMV strain that caused fern leaf syndromes designated CMV-F. All the three isolated strains were serologically indistinguishable from each other and may belong to one serotype according to Ouchterlony gel diffusion tests. These strains transmitted via aphids (Myzus persicae Sulz) in a non-persistent manner. Physical properties of the virus strains were very similar where thermal inactivation test showed that virus withstood heating for 10 min at $70^{/circ}$, dilution end point was $10^{-4}$, and the longevity in vitro at room temperature was less than 5 days for all virus strains. CMV-D1 and CMV-F were the most devastating diseases spreading in both greenhouse and field-grown tomato where aborted flower buds failed on fruit setting due to the viral infection. This is the first report to isolate three different strains of CMV in Kuwait.

• Korean Journal of Poultry Science
• /
• v.24 no.2
• /
• pp.59-66
• /
• 1997

Optimum drying conditions to utilize porcine blood from slaughter house for blood meals, and the effects of blood meals on growth in broiler chicks were investigated. Moisture and protein con-tents of slaughter porcine blood were 79.8 and 16.4%, respectively. The protein contents of the flash dried blood meals at 80˚C were not different from those of the spray dried blood meals at 160 and 190˚C, but higher by 17% relative to those of the spray dried blood meals at 80 and 120˚C. Results from protein analysis by SDS-polyacrylamide electrophoresis showed that flash dried blood meals at 80˚C and spray dried blood meals at 160˚C were better than spray dried blood meals at 80, 120 and 190˚C in terms of protein quality. In Feeding Trial I with broiler chicks, body weights of chicks fed 2, 4 and 6% flash dried blood meal diets at 80˚C were increased at 35 days by 5.6, 7.9 and 4.0%, respectively, compared to control group(P<0.05). In Feeding Trial II, body weights of chicks fed 4 and 6% flash dried blood meal diets at 80˚C were increased at 42 days by 4.9 and 5.3%, respectively, compared to control group(P<0.05). Feed conversion ratios of chicks fed diets 4 and 6% flash dried blood meal diets at 80˚C were significantly improved at 42 days by 7.0 and 3.7%, respectively, compared to that of control group(P<0.05). The optimum drying condition of slaughter porcine blood seemed to be the flash drying method at 80˚C

• Parasites, Hosts and Diseases
• /
• v.30 no.4
• /
• pp.323-328
• /
• 1992

Analysis of siR isolates of Trichemenes veginalis was carried out with sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) and enzyme-linked immunoelectrotransfer blot (EITB). Trichloroacetic acid-treated antigens of the 6 isolates revealed 25 protein profiles ranging 12~170 kDa of molecular weight in SDS-PAGE. In EITB, the specific immunogenic bands were visualized at 51 kDa and 96 kDa when HY-1 antigen was probed with difFerent mice sera immunized with 6 isolates of T. vaginalis. The banding patterns with different sera showed isolate-to-isolate variability. In EITB, homologous antigen (HY-1) did not show any enhanced response in reacting to homologous antiserum(HY-1) when 6 isolates of T. vaginalis were probed with a single serum (HY-1). It is assumed that the different banding patterns of six isolates show isolate-to-isolate variability and immunogenic common bands in 41, 47, 74 and 9:k kDa on EITB may connote the important significance on immune response in T. vaginalis infection.

• Korean Journal of Veterinary Research
• /
• v.40 no.3
• /
• pp.551-561
• /
• 2000

Swine respiratory diseases have induced severe economic losses in swine industry worldwide. Several methods have been developed and applied to prevent and control the disease. However, those are still problematic in swine industry. Recently, the use of egg yolk antibodies with several advantages was introduced and applied to control diseases in animal as well as human. As the first step of the use of egg yolk antibodies in the control of the swine respiratory diseases, we investigated the immunogens of the causative agensts of the diseases and immune response in egg yolk of hens immunized with them. Bacterial antigens prepared from Bordetella bronchiseptica, Pasteurella multocida 3A and 4D, and Actinobacillus pleuropneumaniae serotype 2 and 5 were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blot and toxicity test in mice. The antigens were injected into laying hens in order to produce antibodies against them in egg yolk. After chickens were immunized three times in 2 weeks interval, the profile of antibody production was examined by ELISA. The production of antibody in egg yolk was started in 2 weeks after the first injection, reached peak in 6-8 weeks and maintained until 12 weeks. Of two adjuvants used in this study, ISA70 was more effective than aluminum hydroxide gel in enhancing immunogenecity, laying rates and safety in hens. These results suggested that egg yolk antibodies could be a good source for production of antibodies specific to pathogenic bacteria inducing respiratory diseases of swine.

• Korean Journal of Veterinary Research
• /
• v.36 no.2
• /
• pp.379-387
• /
• 1996

The objects of the present study were to establish the method of purification, subunit dissociation of verotoxin-2 (VT2) produced by Escherichia coli O157:H7, and to investigate the characteristics of purified verotoxin-2 such as molecular weight and composition of amino acid. The results were summerized as follows; Verotoxin-2 was extracted by addition of polymyxin B sulfate into bacterial cell lysate prepared from Escherichia coli O157:H7(KSC109). As an initial step, the bacterial cell lysate was precipitated with 30% saturated ammonium sulfate. The precipitated crude toxin was then subjected to anion-exchange, chromatofocusing and cation-exchange chromatography. Using this scheme, we obtained highly purified toxin with a specific activity of $1.1{\times}10^9$ $CD_{50}/mg$. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) for purified VT2 showed two protein bands. The upper band, approximately 32 Kd, was supposed as A subunit and the lower band, approximately 7.7 Kd, was supposed as B subunit. When the toxin was separated in the subunit-dissociating solution, two peaks emerged with retention times of 15 and 28 min by HPLC. These peaks represented A subunit and B subunit, respectively. The amino acid composition of purified VT2 were made up in order of glutamic acid, histamine, asparaginic acid, histidine, lysine, alanine and leucine etc. The largest amount among the amino acid composing VT2 was methionine.