• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.17 no.5
• /
• pp.368-372
• /
• 1984

This paper deals with a rapid method for determination of ATP and its related compounds in fish and shellfish products using high performance liquid chromatography(HPLC). The HPLC used is a HPLC/ALC-224 equiped with UV-spectrophotometer (254 nm) as detector and integrator (Yanagimoto system-1000). The column used is a stainless steel tubing ($30.0\;cm{\times}3.9\;mm\;i.d.$) packed with ${\mu}-Bon-dapak\;C_{18}$. A mixture of $1\%$ triethylamine-phosphoric acid(pH6.5) was used as an eluent and the flow rate of the eluent was controlled at 2 ml/min. For the separation of ATP and its related compounds, a standard mixture of ATP, ADP, AMP, IMP, inosine and hypoxanthine was subjected to HPLC under the above mentioned conditions. Six peaks were obtained with retention times within 20 min, and elution order were hypoxanthine, IMP, inosine, AMP, ADP and ATP. But 5'-IMP and 5'-GMP fractions were not separated by this method. In generally, IMP content in boiled-dried fish and shellfish products purchased from the market was comparatively higher than that of other nucleotides. Especially, boiled-dried big eye herring marked higher value in IMP content than other boiled-dried ones. Hypoxanthine and inosine were major components of ATP-related compounds in dried products and seasoned-dried ones. And IMP content in seasoned-dried products was higher than that of dried ones. This fact is suggested that a part of IMP in seasoned-dried ones was derived from flavoring matter (MSG, 5'-IMP and 5'-GMP) which is added during the seasoning treatment.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.16 no.2
• /
• pp.140-146
• /
• 1983

Considering fermented sardine with $20\%$ salt as a reference, the fermented sardines were prepared with 8 or $10\%$ salt, $0.5\%$ lactic acid, $6\%$ sorbitol and $6\%$ ethanol. The taste compounds and TBA value as the index of lipid oxidation were analysed prior to fermentation and after 60 days fermentation in products. The major free amino acids in fermented sardine determined after 60 days fomentation were lysine, leucine, histidine, glutamic acid, arginine and alanine which occupied about $58\%$ of the total free amino acids while histidine and taurine occupied about $93.3\%$ of them in raw meat. Betaine and TMA increased while TMAO and total creatinine decreased filling the fermentation of 60 days. Addition of $0.02\%$ BHA was effective to retard rancidity of the product. The result of omission test showed that the major taste compounds of the fermented sardine with low salt contents were amino acid such as lysine, alanine, glycine, glutamic acid, leucine and nucleotides and their related compounds, and it is also suggested that betaine, total creatinine and TMAO acted as an auxiliary role.

• Journal of Ginseng Research
• /
• v.37 no.2
• /
• pp.176-186
• /
• 2013

In this study, we have investigated the effects of total saponin from Korean red ginseng (TSKRG) on thrombin-induced platelet aggregation. TSKRG dose-dependently inhibited thrombin-induced platelet aggregation with $IC_{50}$ value of about 81.1 ${\mu}g/mL$. In addition, TSKRG dose-dependently decreased thrombin-elevated the level of cytosolic-free $Ca^{2+}$ ($[Ca^{2+}]_i$), one of aggregation-inducing molecules. Of two $Ca^{2+}$-antagonistic cyclic nucleotides as aggregation-inhibiting molecules, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), TSKRG significantly dose-dependently elevated intracellular level of cAMP, but not cGMP. In addition, TSKRG dose-dependently inhibited thrombin-elevated adenosine triphosphate (ATP) release from platelets. These results suggest that the suppression of $[Ca^{2+}]_i$ elevation, and of ATP release by TSKRG are associated with upregulation of cAMP. TSKRG elevated the phosphorylation of vasodilator-stimulated phosphoprotein (VASP)-$Ser^{157}$, a cAMP-dependent protein kinase (A-kinase) substrate, but not the phosphorylation of VASP-$Ser^{239}$, a cGMP-dependent protein kinase substrate, in thrombin-activated platelets. We demonstrate that TSKRG involves in increase of cAMP level and subsequent elevation of VASP-$Ser^{157}$ phosphorylation through A-kinase activation to inhibit $[Ca^{2+}]_i$ mobilization and ATP release in thrombin-induced platelet aggregation. These results strongly indicate that TSKRG is a beneficial herbal substance elevating cAMP level in thrombin-platelet interaction, which may result in preventing of platelet aggregation-mediated thrombotic diseases.

• Journal of Korean Society of Forest Science
• /
• v.102 no.2
• /
• pp.223-228
• /
• 2013

Phytoplasmas were detected consistently in 42 mulberry cultivars showing dwarf disease using DNA analysis by amplification with phytoplasma universal primer pairs P1/P7 (about 1.8 kb and R16F2n/R2 (about 1.2 kb). The point mutation from 42 cultivars of mulberry tree was detected by single-strand conformation polymorphism (SSCP) analysis. The SSCP profiles were clearly observed from all of cultivars in 8% polyacrylamide gel, electrophoresizing for and running 8-15 hrs. at 150V, $10^{\circ}C$. The MD and JWB phytoplasma PCR products was mixed and electrophoresis was performed to detect their polymorphism. In this results, the SSCP profiles of all bands of MD and JWB were analyzed on single lane and were distinct in their each of band patterns. The SSCP analysis was possible to detect of 1.8 kb and 1.2 kb nucleotide size and near close band patterns were distinct by mix of two samples. Previously, it was only possible to detect of point mutation under 600 bp nucleotide sequence by SSCP analysis but this modification of SSCP technique was possible to detect clearly SSCP band patterns of about 1.8 kb and 1.2 kb nucleotides.

• Korean Journal of Agricultural Science
• /
• v.28 no.2
• /
• pp.78-84
• /
• 2001

This study was analyzed by PCR technique with specific primer in order to investigate the characteristics of ${\beta}$-lactoglobulin (${\beta}$-LG) gene 5'flanking region in Korean native goat. This work confirmed amplified product of 1,077 bp fragments obtained from the amplification of ${\beta}$-LG promoter from genomic DNA using PCR in Korean native goat. The nucleotide sequence of ${\beta}$-LG gene 5'flanking region in Korean native goat as compared with that of sheep ${\beta}$-LG were different at 46 base of 897 nucleotides, and showed high homology as about 94.9% each breed. Especially we confirmed that the difference of nucleotide sequences between Korean native goat and sheep were consisted of $T{\rightarrow}C$ substitution and $C{\rightarrow}T$ substitution reversely. As a consequences, the sequences of ${\beta}$-LG gene 5'flanking region showed a high homology between Korean native goat and sheep. Furthermore we should be studied that relationships between the control of gene expression and nucleotide sequences of transcription factor in Korean native goat.

• Journal of Ginseng Research
• /
• v.39 no.4
• /
• pp.354-364
• /
• 2015

Background: Intracellular $Ca^{2+}$($[Ca^{2+}]_i$) is a platelet aggregation-inducing molecule. Therefore, understanding the inhibitory mechanism of $[Ca^{2+}]_i$mobilization is very important to evaluate the antiplatelet effect of a substance. This study was carried out to understand the $Ca^{2+}$-antagonistic effect of total saponin from Korean Red Ginseng (KRG-TS). Methods: We investigated the $Ca^{2+}$-antagonistic effect of KRG-TS on cyclic nucleotides-associated phosphorylation of inositol 1,4,5-trisphosphate receptor type I ($IP_3RI$) and cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) in thrombin (0.05 U/mL)-stimulated human platelet aggregation. Results: The inhibition of $[Ca^{2+}]_i$ mobilization by KRG-TS was increased by a PKA inhibitor (Rp-8-BrcAMPS), which was more stronger than the inhibition by a cyclic guanosine monophosphate (cGMP)- dependent protein kinase (PKG) inhibitor (Rp-8-Br-cGMPS). In addition, Rp-8-Br-cAMPS inhibited phosphorylation of PKA catalytic subunit (PKAc) ($Thr^{197}$) by KRG-TS. The phosphorylation of $IP_3RI$ ($Ser^{1756}$) by KRG-TS was very strongly inhibited by Rp-8-Br-cAMPS compared with that by Rp-8-BrcGMPS. These results suggest that the inhibitory effect of $[Ca^{2+}]_i$ mobilization by KRG-TS is more strongly dependent on a cAMP/PKA pathway than a cGMP/PKG pathway. KRG-TS also inhibited the release of adenosine triphosphate and serotonin. In addition, only G-Rg3 of protopanaxadiol in KRG-TS inhibited thrombin-induced platelet aggregation. Conclusion: These results strongly indicate that KRG-TS is a potent beneficial compound that inhibits $[Ca^{2+}]_i$ mobilization in thrombin-platelet interactions, which may result in the prevention of platelet aggregation-mediated thrombotic disease.

• Korean Journal of Plant Resources
• /
• v.16 no.3
• /
• pp.230-237
• /
• 2003

This study was conducted to investigate the occurrence and characterization of tobacco mosaic virus(TMV) in Chinese foxglove isolated from the field of the Chonbuk province(Jinan, Jangsu, Jeongeup). TMV was detected in all three regions and confirmed positive reaction by ELISA test. In the host range test, Chenopodium amaranticola, Nicotiana glutinosa, N. tabacum cv. 'Bright yellow', N. tabacum cv. 'KY­57, Datura stramonium were locally infected with the virus. The virus produced mosaic symptom on inoculated leaves of N. tabacum cv. 'Samson'. However, Chenopodium quinoa, Glycine max, Raphanus sativus, Cucumis sativus, Cucurbita moschata, Brassica rape and Lycopersion esculentum did not show any symptoms. TMV particles were revealed as a stiff rod shape by transmission electron microscopic(TEM) and measured as 300 nm in length with 18 nm in diameter. Total RNA was extracted from showing symptom loaves infected with TMV and the reverse transcription­polymerase chain reaction (RT­PCR) obtained 531 bp DNA product of RNA with specific primer used. The capsid protein of TMV­RE showed higher amino acid sequence homology(97.7％) with TMV­To than with TMV­P(72.2％). The capsid protein of TMV­152 showed same amino acid sequence homology with TMV­F. The result of comparison of nucleotides sequence homology between TMV­RE strain and other TMV strain showed 94％ homology with others except TMV­P(67.3％) and TMV ­ C(68.6％).

• Journal of Life Science
• /
• v.17 no.11
• /
• pp.1497-1504
• /
• 2007

To investigate the function and secretion of human thrombopoietin (TPO) in mammalian cells, hTPO cDNA was cloned using human liver cDNA, and recombinant hTPO (rec-hTPO) was produced in CHO cell lines. In addition, six N-linked glycosylation sites were substituted for Ala to elucidate the role of each carbohydrate chain. To analyze the biological activity, rec-hTPO protein was injected subcutaneously. Blood was withdrawn for platelet determination. The metabolic clearance rate (MCR) was also analyzed at the 1, 4, 10 and 24 hr after tail vein injection. Wild-type TPO (WT) was efficiently secreted into the medium. However, a hTPO mutant with 116 deleted nucleotides detected by PCR cloning was not secreted. The N-linked glycosylation sites had nearly the same expression quantity as rec-hTPO WT apart from mutants 3 and 4. The glycosylation site of mutant 4 appeared to be an indispensable site for hTPO secretion. Also characterized was the biological activity through an injection with rec-hTPO (10 ng) to ICR mice (7 weeks). The result of the blood analysis showed a considerable increase in the platelet number six days after He injection. To analyze the pharmacokinetics, rec-hTPO was injected into the tail vein (5 ng). The result was 200 pg/ml 1hr after this injection. Following this, it dramatically decreased and virtually disappeared 10 hours after the injection. Thus, rec-hTPO may be a treatment for thrombopenia by the production of the high active rec-hTPO. In addition, hTPO can permit the development of potent new analogues that stimulate the platelet value.

• Journal of Life Science
• /
• v.27 no.9
• /
• pp.1003-1010
• /
• 2017

An open reading frame coding for mannanase predicted from the partial genomic sequence of Paenibacillus woosongensis was cloned into Escherichia coli by polymerase chain reaction amplification, and completely sequenced. This mannanase gene, designated man26AT, consisted of 3,162 nucleotides encoding a polypeptide of 1,053 amino acid residues. Based on the deduced amino acid sequence, Man26AT was identified as a modular enzyme, which included a catalytic domain belonging to the glycosyl hydrolase family 26 and two carbohydrate-binding modules, CBM27 and CBM11. The amino acid sequence of Man26AT was homologous to that of several putative mannanases, with identity of 81% for P. ihumii and identity of less than 57% for other strains of Paenibacillus. A cell-free extract of recombinant E. coli carrying the man26AT gene showed maximal mannanase activity at $55^{\circ}C$ and pH 5.5. The enzyme retained above 80% of maximal activity after preincubation for 1 h at $50^{\circ}C$. Man26AT was comparably active on locust bean gum (LBG), galactomanan, and kojac glucomannan, whereas it did not exhibit activity on carboxymethylcellulose, xylan, or para-nitrophenyl-${\beta}$-mannopyranoside. The common end products liberated from mannooligosaccharides, including mannotriose, mannotetraose, mannopentaose, and mannohexaose, or LBG by Man26AT were mannose, mannobiose, and mannotriose. Mannooligosacchrides larger than mannotriose were found in enzymatic hydrolyzates of LBG and guar gum, respectively. However, Man26AT was unable to hydrolyze mannobiose. Man26AT was intracellularly degraded into at least three active proteins with different molecular masses by zymogram.

• Korean Journal of Microbiology
• /
• v.45 no.4
• /
• pp.306-311
• /
• 2009

A new PCR method was developed to detect Pectobacterium carotovorum which is the causative agent of soft rot in Brassica pekinensis. A specific detection primer set based on Lytic murein transglycolase gene was designed and evaluated. Using ERB3_F (5'-TGC GAC ACC TCC TCA TCA CG-3') and ERB3_R (5'-CTT ATC ACG CTG TAA CCA GC-3') primers, 437 nucleotides long fragment was specifically amplified. The amplified products were observed in 52 out of 55 strains of P. carotovorum or Pectobacterium carotovorum subsp. carotovorum. On the other hand, no amplification was observed in 8 organisms including Chinese cabbage and potato. The optimal PCR condition for the ERB3_F/ERB3_R primer set was $58^{\circ}C$ for annealing and 15 mM for $MgCl_2$. With serially diluted templates, the specific PCR sensitivitie limit was $2\times10^3$ copies. Also, this method can be applied not only to DNA but also to field samples. This PCR method may be expected to be useful for specific detection of P. carotovorum.