• 생약학회지
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• 제45권4호
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• pp.300-314
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• 2014

An ultra-performance liquid chromatography-electrospray ionization-mass spectrometer (UPLC-ESI-MS) method was established for the simultaneous quantification of eighteen marker compounds in traditional Korean formula, Cheonwangbosimdan (CWBSD). In addition, we evaluated the antioxidant effects of CWBSD. Eighteen marker components were separated on a UPLC BEH $C_{18}$ analytical column ($2.1{\times}100mm$, $1.7{\mu}m$) and kept at $45^{\circ}C$ by gradient elution with 0.1% (v/v) formic acid in water and acetonitrile as mobile phase. The flow rate was 0.3 mL/min and the injection volume was $2.0{\mu}L$. The antioxidant activities of CWBSD were assessed by measuring free radical scavenging activities on 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and 1-1-diphenyl-2-picrylhydrazyl (DPPH). The calibration curves of all analytes showed good linearity (correlation coefficient ${\geq}0.9937$) within the test ranges. The limits of detection and quantification for the 18 marker compounds were 0.01-4.71 ng/mL and 0.03-14.13 ng/mL, respectively. The contents of the 18 compounds in CWBSD extract ranged from none to $1701.00{\mu}g/g$. The CWBSD showed the radical scavenging activity in a dose-dependent manner. The concentration required for 50% reduction ($RC_{50}$) against ABTS and DPPH radicals were $149.42{\mu}g/mL$ and $339.24{\mu}g/mL$.

• 생약학회지
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• 제47권4호
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• pp.366-373
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• 2016

Bulhwangeumjeonggisan (BHGJGS) is a traditional herbal formulation generally used in the treatment of cold and gastritis. BHGJGS consists of eight herbal plants; Atractylodis Rhizoma, Magnoliae Cortex, Citri Pericarpium, Glycyrrhizae Radix, Agastachis Herba, Pinelliae Rhizoma, Zingiberis Rhizoma and Zizyphi Fructus. Complete standardization of this formulation has not been done yet. So, a simple and accurate method was developed and validated using Ultra Performance Liquid Chromatography (UPLC) with Diode Array Detector (DAD) for the standardization of BHGJGS. UPLC conditions were optimized using a c18 RP-Amide column with mobile phase; 0.1% phosphate buffer and acetonitrile, detection wavelength; 210 and 325 nm. The linearities of calibration curves were acceptable ($R^2$>0.9994), and the limit of detection and quantification were within the ranges of 0.011-0.091 and $0.034-0.277{\mu}g/ml$ respectively. The relative standard deviation (RSD) of intra- and inter-day precisions were under 3.61%. The RSD of repeatability was under 0.68 %. The results of recovery test were 94.4-107.9%, and the RSD were under 4.6%. The developed method was used to find the contents of standard constituents in BHGJGS mix extract powder, and two commercial formulation (A and B). The data show that the developed method was specific, sensitive, accurate, and precise for analysis of BHGJGS components.

• Biomolecules & Therapeutics
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• 제15권2호
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• pp.118-122
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• 2007

This study was performed to evaluate the bioequivalency between the Osmetone$^{TM}$ Tablet (Myeongmoon Pharm. Co., Ltd.) as a test formulation and the Relafen$^{TM}$ Tablet (Handok Pharm. Co., Ltd.) as a reference formulation. Twenty-four healthy male volunteers were administered the formulations by the randomized Latin square crossover design, and the plasma samples were determined by a high performance liquid chromatography (HPLC) with Ultra-Violet (UV) detector. AUC$_t$, C$_{max}$ and T$_{max}$ were obtained from the time-plasma concentration curves, and log-transformed AUC$_t$ and C$_{max}$ and log-untransformed T$_{max}$ values for two formulations were compared by statistical tests and analysis of variation. AUC$_t$ was determined to be 897.8${\pm}$431.1 ug.hr/ml for the reference formulation and 902.3${\pm}$408.4 ug.hr/ml for the test formulation. The mean values of C$_{max}$ for the reference and test formulations were 24.2${\pm}$8.9 and 24.0${\pm}$9.5 ug/ml, respectively. The AUC$_t$ and C$_{max}$ ratios of the reference Relafen$^{TM}$ Tablet to the test Osmetone$^{TM}$ Tablet were +5.01% and -0.83%, respectively, showing that the mean differences were satisfied the acceptance criteria within 20%. The results from analysis of variance for logtransformed AUC$_t$ and C$_{max}$ indicated that sequence effects between groups were not exerted and 90% confidence limits of the mean differences for AUC$_t$ and C$_{max}$ were located in ranges from log 0.80 to log 1.25, satisfying the acceptance criteria of the KFDA bioequivalence. The Osmetone$^{TM}$ Tablet as the test formulation was considered to be bioequivalant to the Relafen$^{TM}$ Tablet used as its reference formulation, based on AUC$_t$ and C$_{max}$ values.

• 대한한의학방제학회지
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• 제18권1호
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• pp.145-154
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• 2010

Objective : Licorice has been used for treating digestive disorder and also recommended as a detoxification agent. Liquiritigenin, a component of licorice, has been reported to have various biological activities. In this study, we aimed to establish the analytical method for liquiritigenin content in licorice and the processing method for the enhancement of liquiritigenin content in licorice. Methods : Processing was accomplished by roasting licorice at $250^{\circ}C$ for indicated time periods (5-20 min). Analysis of liquiritigrnin from roasted licorice was conducted using UPLC(Ultra Performance Liquid Chromatography). Results : We established UPLC method for the analysis of liquiritigenin using water : acetonitrile gradient as mobile phase. Furthermore, we standardized the processing condition of licorice to enhance liquiritigenin content using UPLC method. Processing of licorice was accomplished by roasting at $250^{\circ}C$ for indicated time periods (5-20 min) and by pretreating with 50% of acetic acid or 30% ethanol for 24 h. By roasting licorice, the liquiritigenin contents in the licorice were increased. The best roasting time of licorice was 6 min, while roasting for the time above 8 min resulted in diminishing liquiritigenin contents. Moreover, pretreatment with 50% of acetic acid or 30% ethanol picked up liquiritigenin contents in roasted licorice. Conclusion : The adequate processing condition of licorice for the enhancement of liquiritigenin contents was obtained by pretreating licorice with 50% of acetic acid or 30% ethanol for 24 h and then by roasting at $250^{\circ}C$ for 6 min.

• 한국약용작물학회지
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• 제28권1호
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• pp.21-28
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• 2020

Background: Owing to its high efficiency in lipid and protein production, peanut (Arachis hypogaea L.) is considered one of most important crops world-wide. The kernels of peanuts are undoubtedly the most important product this plant, whereas the skin is almost completely neglected in nutraceutical terms. However, peanut skin contains potentially health-promoting phenolics and dietary fiber, and there is considerable potential for commercial exploitation. In this study, we evaluated the α-glucosidase inhibitory activity of an extract of peanut skin (PS). Methods and Results: The α-glucosidase inhibitory effects of 80% ethanol extracts of peanut (A. hypogaea L. 'Sinpalkwang') skin were evaluated and found to have a half-maximal inhibitory concentration (IC₅₀) value of 1.2 ㎍/㎖. Progress curves for enzyme reactions were recorded spectrophotometrically, and the inhibition kinetics revealed time-dependent inhibition with enzyme isomerization. Furthermore, using ultra-high performance liquid chromatography combined with quadrupole-orbitrap mass spectrometry, we identified 26 compounds in the peanut skin extract, namely, catechin, epicatechin, and 24 proanthocyanidins. Conclusions: The results suggest that peanut skin can be utilized as an effective source of α-glucosidase inhibition in functional foods and nutraceuticals.

• 대한한의학방제학회지
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• 제21권1호
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• pp.177-185
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• 2013

Objectives : Leejung-tang (Lizhong-tang) has been used for treatment of gastrointestinal disorders in Korea. In this study, we performed quantification analysis of five marker components, liquiritin, ginsenoside Rb1, ginsenoside Rg1, glycyrrhizin, and 6-gingerol in Leejung-tang using a ultra performance liquid chromatography- electrospray ionization-mass spectrometer (UPLC-ESI-MS). In addition, we evaluated antioxidant activity of Leejung- tang. Methods : The column for separation of five constituents used a UPLC BEH C18 ($100{\times}2.1mm$, $1.7{\mu}m$) maintained at $45^{\circ}C$. The mobile phase consisted of two solvent systems, 0.1% (v/v) formic acid in H2O (A) and CH3CN (B) by gradient flow. The flow rate was 0.3 mL/min with detection at mass spectrometer. The antioxidative activities conduct an experiment on 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activities of Leejung-tang. Results : Calibration curves of five marker compounds were acquired with r2 values > 0.99. The amount of the five compounds in Leejung-tang were 0.07 - 0.84 mg/g. The concentration required for 50% reduction (RC50) against ABTS radical was 119.02 ug/mL. In addition, the scavenging against DPPH radical of Leejung-tang was 11.4%, 14.5%, 19.8%, 29.6%, and 49.2% at 25 ug/mL, $50{\mu}g/mL$, $100{\mu}g/mL$, $200{\mu}g/mL$, and $400{\mu}g/mL$, respectively. Conclusions : The established LC-MS/MS method will be helpful to improve quality control of Leejung-tang. In addition, Leejung-tang is a potential antioxidant therapeutic agent.

• 한국작물학회:학술대회논문집
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• 한국작물학회 2017년도 9th Asian Crop Science Association conference
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• pp.31-31
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• 2017

Understanding the mechanism(s) to overcome or prevent seed ageing deterioration during storage is of fundamental interest to seed physiologists. Vitamin E (tocols) is known as a key metabolite to efficiently scavenge lipid peroxy radicals which cause membrane breakdown resulting in seed ageing. However, in rice research this hypothesis has been tested for very few lines only without considering intraspecific variation in genomic structure. Here, we present a correlation study between tocols and seed longevity using a diverse rice panel. Seeds of 20 rice accessions held in the International Rice Genebank at the International Rice Research Institute, representing aus, indica, temperate japonica and tropical japonica subpopulations, were used for tocols analysis (quantification of ${\alpha}$-, ${\beta}$-, ${\gamma}$-, ${\delta}$-tocopherol/tocotrienol by ultra performance liquid chromatography) and storage experiments at $45^{\circ}C$ and 10.9% seed moisture content (sample taken for germination testing every 3 days up to 60 days). To examine interactions between DNA sequences and phenotype, the 700k high-density single-nucleotide polymorphism marker data-set was utilized. Both seed longevity (time for viability to fall to 50%; $p_{50}$) and tocols content varied across subpopulations due to heterogeneity in the genetic architecture. Among eight types of tocol homologues, ${\alpha}$-tocopherol and ${\gamma}$-tocotrienol were significantly correlated with $p_{50}$ (negatively and positively, respectively). While temperate japonica varieties were most abundant in ${\alpha}$-tocopherol, indica varieties recorded 1.3 to 1.7-fold higher ${\gamma}$-tocotrienol than those of other subpopulations. It was highlighted that specific ratio of tocol homologues rather than total tocols content plays an important role in the seed longevity mechanism.

• 대한본초학회지
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• 제30권3호
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• pp.19-24
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• 2015

Objectives : HPL-1, a novel herbal medicine which is composed of five herbs such as Kalopanacis Cortex, Chaenomelis Fructus, Raphani Semen, Atractylodis Rhizoma and Pulvis Aconiti Tuberis Purificatum, was developed for treatment of osteoarthritis. This study is aimed to develop analytical method for consistent quality control of HPL-1 and validate chromatographic method. Methods : Chromatographic analysis was performed using ultra-high performance liquid chromatography - diode array detector (UHPLC-DAD) equipped with RP-amide column, column oven, and auto sampler. Marker compounds [protocatechuic acid, chlorogenic acid, liriodendrin, 3,5-dicaffeoylquinic acid, ${\beta}$-D-(3-O-sinapoyl)-fructofuranosyl-$\alpha$-D-(6-O-sinapoyl)glucopyranoside and benzoylmesaconine] were separated by step gradient elution of acetonitrile and 0.1% phosphoric acid/water. The method validation was evaluated by quantitative validation parameters of linearity, accuracy, precision, limit of detection (LOD) and limit of quantification (LOQ) according to KFDA guideline.Results : An optimized method for six marker compounds in HPL-1 was established by UHPLC-DAD. The correlation coefficient (R²) with each calibration curve was greater than 0.99. The LOD and LOQ were within the range of 0.008-0.090 and $0.023-0.274{\mu}g/mL$, respectively. The relative standard deviation (RSD) of intra- and inter-day variability were less than 4.0%. The result of recovery test was range from 93.3-106.3% with RSD < 4.0%.Conclusions : These results suggest that the quantitative UHPLC method is precise, accurate, effective for quality evaluation of HPL-1. The method may also contribute to improve quality of crude drug preparations used for treatment of various diseases.

• 생약학회지
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• 제49권1호
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• pp.76-83
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• 2018

A traditional herbal formula, Gyeji-tang has been used to treat the early colds, headache, chills, and fever in Asian countries. In this study, we were performed simultaneous determination of the 14 bioactive marker compounds, gallic acid, spinosin, paeoniflorin, albiflorin, liquiritin apioside, liquiritin, 6'''-feruloylspinosin, liquilitigenin, coumarin, cinnmamic acid, benzoylpaeoniflorin, cinnamaldehyde, glycyrrhizin, and 6-gingerol in Gyeji-tang using an ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS/MS). Analytical column was used a Waters Acquity UPLC BEH $C_{18}$ analytical column ($2.1{\times}100mm$, $1.7{\mu}m$) and maintained at $45^{\circ}C$ with a flow rate of 0.3 mL/min. The mobile phase consists of 0.1% (v/v) formic acid in water and acetonitrile with gradient elution. The MS analysis was conducted using multiple reaction monitoring in the positive and negative modes by a Waters ACQUITY TQD LC-MS/MS system. The calibration curves of 14 bioactive marker compounds showed linearity with correlation coefficients ${\geq}0.9798$. The limits of detection and quantification values were in the range of 0.11-6.66 ng/mL and 0.34-19.99 ng/mL, respectively. As a result of the analysis using the established LC-MS/MS method, the amounts of tested 14 compounds in the lyophilized Gyeji-tang sample were detected up to $85.7{\mu}g/g$. These results may be useful for quality assessment of a traditional herbal formulas.

• 생약학회지
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• 제49권3호
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• pp.270-277
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• 2018

Pyungwi-san has been used to treat the digestive system diseases, physconia, nausea, anorexia, and dyspepsia in Korea. In this study, an ultra-performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS/MS) method was optimized for simultaneous determination of the 14 marker components, spinosin, liquiritirn apioside, liquiritin, narirutin, 6'''-feruloylspinosin, hesperidin, liquiritigenin, glycyrrhizin, 6-gingerol, atractylenolide III, honokiol, atractylenolide II, magnolol, and atractylenolide I in Pyungwi-san extract. All analytes were separated on a Waters Acquity UPLC BEH $C_{18}$ analytical column ($2.1{\times}100mm$, $1.7{\mu}m$) with maintained at $45^{\circ}C$. The mobile phase consisted of 0.1% (v/v) aqueous formic acid and acetonitrile. The MS conditions were as follows: capillary voltage 3.3 kV, extractor voltage 3.0 V, RF lens voltage 0.3 V, source temperature $120^{\circ}C$, desolvation temperature $300^{\circ}C$, desolvation gas 600 L/h, cone gas 50 L/h and collision gas 0.14 mL/min. The coefficient of determination of 14 analytes was 0.9989-1.0000. The limits of detection and quantification values of the all analytes were 0.04-2.56 and 0.13-7.69 ng/mL, respectively. As a result of the analysis using the established LC-ESI-MS/MS method, the 5 components, spinosin, 6'''-feruloylspinosin, atractylenolide III, II, and I derived from Zizyphi Fructus and Atractylodis Rhizoma, were not detected in this extract. On the other hand, the 9 components except for the 5 components were 4.15-498.87 mg/kg in lyophilized Pyungwi-san extract. Among these components, glycyrrhizin, marker compound of Glycyrrhizae Radix et Rhizoma, was detected the most amount as a 498.87 mg/kg.