• Analytical Science and Technology
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• v.21 no.2
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• pp.102-108
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• 2008

Reversed-phase high performance liquid chromatography (RP-HPLC) was used for the simultaneous determination of liquiritin (LQ), glycyrrhizic acid (GA) and glabridin from licorice. An optimized run condition was selected with a binary gradient elution of methanol-water which ramped 35/65 to 80/20 (vol. %) in 0.0-8.0 min and a flow rate of 1.0 mL/min. A good linearity was obtained between 0.2 mg/mL and 1.0 mg/mL for LQ and GA, and 0.01 mg/mL-0.2 mg/mL for glabridin with the relative standard deviations less than 0.90% (n=5). The developed method was successfully applied to determination of the three components from licorice samples. The mean recoveries of three components are 80.79% for liquiritin, 89.71% for glycyrrhizic acid and 72.50% for glabridin.

• The Korean Journal of Physiology
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• v.23 no.1
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• pp.79-87
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• 1989

Molecular property as well as biological activities of bombesin-like substance (BBS-LS) isolated from the skin of B. orientalis in Korea was compared with those of synthetic BBS-14. BBS-LS in the crude extract was applied on a Sephadex G-50, superfine column $(1.6{\times}112\;cm)$. On chromatogram, the first peak (3.5% of BBS-LS) was found ahead of synthetic GRP-27, and the second peak (rest) between synthetic GRP-27 and synthetic BBS-14. The main form of BBS-LS was successfully purified by using a column of alkaline alumina followed by sequential gel-filtrations on a column of Sephadex G-10, fine and a column of Sephadex G-50, superfine. Chromatographic analysis of the purified BBS-LS using a column of Sephadex G-50, superfine and reversed phase HPLC revealed that the main form of BBS-LS in the skin of B. orientalis could be distinctly different from either BBS-14 or GRP-27 in molecular size. The purified BBS-LS exerted biological activities quite identical to those of synthetic BBS-14. The results of the present investigation indicate that the skin of B. orientalis contains BBS-LS composed of two distinct forms. The main form of BBS-LS purified in the present study is heterogenous to both synthetic BBS-14 and GRP-27 in molecular size but identical to BBS in biological activities.

• Journal of Life Science
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• v.28 no.11
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• pp.1301-1315
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• 2018

We purified an antimicrobial peptide from the acidified hemocyte extract of Mytilus coruscus by $C_{18}$ reversed-phase high-performance liquid chromatography (RP-HPLC). The peptide was 4041.866 Da based on matrix-assisted laser desorption ionization time-of-flight mass spectrophotometer (MALDI-TOF/MS) and the 25 amino acids of the N-terminus sequence were identified. Comparison of this sequence of the purified peptide with the N-terminus sequences of other antimicrobial peptides revealed 100% identity with the mytilin B precursor of Mytilus coruscus. We also identified a 312 bp open-reading frame (ORF) encoding 103 amino acids based on the obtained amino acid residues. The nucleotide sequence of this ORF and the amino acid sequence also revealed 100% identity with the mytilin B precursor of Mytilus coruscus. We synthesized two antimicrobial peptides with an alanine residue in the C-terminus, and designated them mytilin B1 and B2. These two antimicrobial peptides showed antimicrobial activity against gram-positive bacteria, including Bacillus cereus and Streptococcus parauberis (minimal effective concentration, MECs $41.6-89.7{\mu}g/ml$), gram-negative bacteria, including Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Providencia stuartii, Pseudomonas aeruginosa, and Vibrio ichthyoenteri (MECs $7.4-39.5{\mu}g/ml$), and the fungus Candida albicans (MECs $26.0-31.8{\mu}g/ml$). This antimicrobial activity was stable under heat and salt conditions. Furthermore, the peptides did not exhibit significant hemolytic activity or cytotoxic effects. These results suggest that mytilin B could be applied as alternative antibiotic agent, and they add to the understanding of the innate immunity of hard-shelled mussels.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.15 no.7
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• pp.4682-4691
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• 2014

Banhabaekchulchhonma-tang is a traditional Korean herbal prescription with a range of pharmacological activities. In this study, the simultaneous analysis of 12 kinds of marker components, Homogentisic acid, Hesperidin, Naringin, Alisol A, Atractylenolide II, Atractylenolide III, Ginsenoside Rg1, Formononetin, Gastrodin, Berberine, Palmatine and 6-gingerol, in Banhabaekchulchhonma-tang was performed using high-performance liquid chromatography (RP-HPLC). The standard sample of commercial $C_{18}$ reversed phase-column using water (0.1% TFA) and acetonitrile as the mobile phase with a step gradient elution mode. The flow rate (1.0mL/min), injection volume ($10{\mu}L$) and column oven temperature ($40^{\circ}C$) at a 200, 220, 280 and 340nm wavelength was conducted. All calibration curves of the standard components showed good linearity ($r^2$ >0.999). In addition, the limit of detection (LOD) and limit of quantification (LOQ) ranged from 0.012 to $0.878{\mu}g/mL$ and 0.009 to $0.290{\mu}g/mL$. The precision intra-day and inter-day were ranged from 0.07 to 1.21% and 0.20 to 0.90%, respectively. The recoveries ranged from 97.17 to 108.40%. Each sample amount showed a very small change. These results highlight the efficient quality evaluation of Banhabaekchulchhonma-tang.

• Bulletin of the Korean Chemical Society
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• v.26 no.5
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• pp.729-734
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• 2005

A sensitive and selective method for quantitation of sofalcone and its active metabolite in human plasma has been established using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS). Plasma samples were transferred into 96-well plate using an automated sample handling system and spiked with 10 $\mu$L of 2 $\mu$g/mL $d_3$-sofalcone and $d_3$-sofalcone metabolite solutions (internal standard), respectively. After adding 0.5 mL of acetonitrile to the 96-well plate, the plasma samples were then vortexed for 30 sec. After centrifugation, the supernatant was transferred into another 96-well plate and completely evaporated at 40 ${^{\circ}C}$ under a stream of nitrogen. Dry residues were reconstituted with mobile phase and were injected into a $C_{18}$ reversed-phase column. The limit of quantitation of sofalcone and its metabolite was 2 ng/mL, using a sample volume of 0.2 mL for analysis. The reproducibility of the method was evaluated by analyzing 10 replicates over the concentration range of 2 ng/mL to 1000 ng/mL. The validation experiments of the method have shown that the assay has good precision and accuracy. Sofalcone and its metabolite produced a protonated precursor ion ([M+H]$^+$) of m/z 451 and 453, and a corresponding product ion of m/z 315 and 317, respectively. Internal standard ($d_3$-sofalcone and $d_3$-sofalcone metabolite) produced a protonated precursor ion ([M+H]$^+$) of m/z 454 and 456 and a corresponding product ion of m/z 315 and 317, respectively. The method has been successfully applied to a pharmacokinetic study of sofalcone and its active metabolite in human plasma.

• Journal of Life Science
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• v.26 no.3
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• pp.296-301
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• 2016

An antibacterial peptide from skin extract of the catfish Silurus asotus was purified and characterized. The acidified skin extract was put through a Sep-Pak C18 solid phase extraction cartridge using a stepwise gradient and divided into flow-through (F.T.), 10% methanol-elute (RM10), 60% methanolelute (RM60), and 100% methanol-elute (RM100) fractions. RM10, RM60, and RM 100 showed antimicrobial activity against Escherichia coli D31. On the other hand, the F.T. fraction did not show antimicrobial activity. Among the various fractions, RM 60 had the highest activity. RM 60 was partially purified on a cation exchange column (CM52) by a stepwise gradient. The ammonium acetate (pH 5.15) 0.02 M – 0.8 M fraction showed antimicrobial activity. Then an antimicrobial peptide was purified using a 0.6M fraction with strong antibacterial activity through a series of five C18 reversed-phase HPLC columns. For the characterization of the purified peptide, the molecular weight and amino acid sequence were analyzed by MALDI-TOF MS and Edman degradation. The molecular weight of this peptide was about 4182.1 [M+H]⁺. The amino acid sequence of this peptide was partially determined as follows: PALXXKARREAKVKF. These findings suggest that this peptide plays a significant role in the innate defense system of catfish skin.

• Journal of Life Science
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• v.27 no.1
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• pp.50-56
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• 2017

An antibacterial peptide was purified from an acidified gill extract of the pufferfish Takifugu pardalis. The acidified gill extract was put through a Sep-Pak C18 solid phase extraction cartridge using a stepwise gradient and divided into a flow-through (F.T.) and 60% methanol fraction (RM 60). Among the eluents, RM 60 had potent antibacterial activity against Bacillus subtilis KCTC 1021. RM 60 was partially purified on a cationic-exchange column (SP-5PW) by a linear gradient, and the antibacterial peptide was then further purified, using a series of cationic-exchange and $C_{18}$ reversed-phase HPLC columns. For characterization of the purified peptide, its molecular weight and amino acid sequence were analyzed by MALDI-TOF MS and Edman degradation. The molecular weight of the peptide was about 1171.6 Da. The amino acid sequence of the peptide was partially determined as: STKEKAPRKQ. A comparison of the N-terminal amino acid sequence of the purified peptide with that of other known polypeptides revealed high homology with the N-terminus of the histone H3 protein, which belongs to the histone H3 family. Thus, this peptide was designated as a puffer fish gill (PFG)-related antimicrobial peptide. This is the report to describe an antimicrobial function for the N-terminus of histone H3 of an animal species. The findings suggest that this peptide plays a significant role in the innate defense system of the pufferfish.

• Journal of the Korean Crystal Growth and Crystal Technology
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• v.17 no.2
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• pp.82-89
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• 2007

Glass-ceramics were fabricated by heat-treating a glass at $700^{\circ}C$/10hr which was obtained by melting a glass frit mixed with $40{\sim}80 wt%$ EAF dust at $1300^{\circ}C$/1hr. Dependence of crystal phase and bonding state change upon a compositional change and heat treatment condition were studied and the results was connected to the toxic characterization leaching procedure (TCLP) test data to investigate a chemical durability of the specimens. Increasing dust in a glass shifted the peak around $1000cm^{-1}$ to the lower frequency which was composed of two vibration peaks for the nonbridging oxygen at $960cm^{-1}$ and the bridging oxygen at $1050{\sim}1060cm^{-1}$. Also, the $B_2O_3$ structure of boroxol ring changed to a tetrhedral-, trigonal- and di-borate with dust addition. The Fe-O peaks in the glass-ceramics were observed which is consitent with XRD results of spinel formation. The surface of glass after TCLP test was severely cracked while there was no cracks on a glass-ceramics after TCLP test so the chemical durability of the glass-ceramics is superior than that of glass. The leaching concentration of Fe for the glass-ceramics containing EAF dust 80 wt% is 1/15 times lower than that of glass. The Zn leaching concentration fur the glass-ceramics containing dust < 70 wt% was higher than that of glass but its trend was reversed for the specimen of dust content > 80 wt% which could be concluded as correlated with occurrence of willemite phase.

• Journal of fish pathology
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• v.20 no.2
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• pp.139-145
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• 2007

To decrease stress in eel (Anguilla japonica) during its culture or transportation, aspirin (ASA) known as analgesic, antiinflammatory and antithrombic agent was administrated by dipping or oral routes. Concentrations of aspirin (ASA) and salicylic acid (SA) in eel plasma were simultaneously measured by a high performance liquid chromatography (HPLC). The plasma was acidified with 0.2 M HCl and 0.2 M orthophosphoric acid, and mixed with acetonitrile. ASA and SA extracted with acetonitrile were analyzed by the HPLC equipped with reversed phase Novapak C18 column (4 ㎛ silica, 150×4 mm) and UV detector(237 nm). The mobile phase consisted of 740 ㎖ water, 900 ㎕ orthophosphoric acid (85%) and 180 ㎖ acetonitrile. The retention times of ASA, SA and 2-methylbenzoic acid(MBA) were 4.8 min, 8.4 min and 11.5 min, respectively. The limit of quantification was 0.01 ㎍/㎖ for SA and 0.05 ㎍/㎖ for ASA. The mean recovery from eel plasma was 70.8～99.6% for ASA and 95.2～100.3% for SA. This HPLC method was applied to analyze ASA and SA of eel plasma after either dipping in a concentration of 20 ppm or feeding the feed supplemented with 50 ㎎/kg BW. Only SA was detected in eel plasma after the administration of ASA by dipping or oral routes because the drug was quickly decomposed into SA in eel plasma. The amount of SA in eel plasma reached the highest value at 3hr in dipping and 7 days in oral administration. When the ASA-administrated eel were kept in ASA free aquaria, 0.02-0.03 ㎍/㎖ of SA were detected 48 hr after the administration in both routes.

• Journal of Pharmaceutical Investigation
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• v.36 no.2
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• pp.89-95
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• 2006

A selective and sensitive reversed-phase HPLC method for the determination of pentoxifylline in human serum was developed, validated, and applied to the pharmacokinetic study of pentoxifylline. Pentoxifylline and internal standard, chloramphenicol, were extracted from the serum by liquid-liquid extraction with dichloromethane and analyzed on a Luna CI8(2) column with the mobile phase of acetonitrile-0.034 M phosphoric acid (25:75, v/v, adjusted to pH 4.0 with 10 M NaOH). Detection wavelength of 273 nm and flow rate of 0.8 mL/min were used. This method showed linear response over the concentration range of 10-500 ng/mL with correlation coefficients greater than 0.999. The lower limit of quantification using 0.5 mL of the serum was 10 ng/mL, which was sensitive enough for pharmacokinetic studies of pentoxifylline. The overall accuracy of the quality control samples ranged from 89.3 to 92.7% for pentoxifylline with overall precision (% C.V.) being 4.1-9.2%. The relative mean recovery of pentoxifylline for human serum was 105.8%. Stability (stock solution, short and long-term) studies showed that pentoxifylline was not stable during storage. But three freeze-thaw cycles and extracted serum samples were stable. This method showed good ruggedness (within 15% C.V.) and was successfully applied for the analysis of pentoxifylline in human serum samples for the pharmacokinetic studies of orally administered $Trental^{\circledR}$ tablet (400 mg pentoxifylline), demonstrating the suitability of the method.