• Biomolecules & Therapeutics
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• v.6 no.3
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• pp.289-295
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• 1998

The bioequivalence of two nilvadipine products was evaluated in 16 normal male volunteers (age 22-32 yr, body weight 57-80 kg) following sidle oral dose. Test product was Overca $l_{R}$ tablet (Choong-Wae Pharm. Corp., Korea) and reference product was Nivadi $l_{R}$ tablet (Hyundai Pharm. Corp., Korea). Both products contain 4 mg of nilvadipine. One tablet of the test or the reference product was administered to the volunteers, respectively, by randomized two period cross-over study (2$\times$2 Latin square method). The determination of nilvadipine was accomplished using a validated capillary column GC with electron-capture detection. As a result of the assay validation, the quantiflcation of nilvadipine in human plasma by this technique was possible down to 0.5 ng/ml using 1 ml of plasma. Absolute overall recovery from five replicate analyses of nilvadipine-spiked sample were 88.4$\pm$ 10.24% (mean$\pm$ 5.D.) for human plasma of 10 ng/ml. The coefficients of variation (C.V.) were less than 20% and the actual concentration of nilvadipine measured by GC ranged from 80 to 99% in all plasma. Average drug concentrations at each sampling time and pharmacokinetic parameters calculated were not significantly different between two products (p>0.05); the area under the curve from time zero to 8 hr (AUCo-$_{8 hr}$) (22.8$\pm$5.90 vs 22.2$\pm$6.10 ng . hr/ml), maximum plasma concentration ( $C_{max}$) (10.0$\pm$2.85 vs 9.3$\pm$3.28 ng/ml) and time to reach maximum plasma concentration ( $T_{max}$) (1.2$\pm$0.31 vs 1.3 $\pm$0.47 hr). The differences of mean AU $Co_{8hr}$ $C_{max}$, and $T_{max}$ between the two products (2.25, 7.65, and 10.30%, respectively) were less than 20%. The power (1-$\beta$) and treaeent difference (7) for AU $Co_{8hr}$, and $C_{max}$ were more than 0.8 and less than 0.2, respectively. Although the power for Tmax was under 0.8, Tm\ulcorner of the two products was not significantly different from each other (p>0. 05). These results suggest that the bioavailability of Overeat tablet is not significantly different from that of Nivadil tablet. Therefore, two products are bioequivalent based on the current results.sults.lts.lts.lts.

• Bulletin of the Korean Chemical Society
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• v.24 no.5
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• pp.559-565
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• 2003

An analytical method the determination of benzo(a)pyrene (BaP) and its hydroxylated metabolites, 1-hydroxybenzo(a)pyrene (1-OHBaP), 3-hydroxybenzo(a)pyrene (3-OHBaP), benzo(a)pyrene-4,5-dihydrodiol (4,5-diolBaP) and benzo(a)pyrene-7,8-dihydrodiol (7,8-diolBaP), in rat urine and plasma has been developed by HPLC/FLD and GC/MS. The derivatization with alkyl iodide was employed to improve the resolution and the detection of two mono hydroxylated metabolites, 1-OHBaP and 3-OHBaP, in LC and GC. BaP and its four metabolites in spiked urine were successfully separated by gradient elution on reverse phase ODS $C_{18}$ column (4.6 mm I.D., 100 mm length, particle size 5 ㎛) using a binary mixture of MeOH/H₂O (85/15, v/v) as mobile phase after ethylation at 90 ℃ for 10 min. The extraction recoveries of BaP and its metabolites in spiked samples with liquid-liquid extraction, which was better than solid phase extraction, were in the range of 90.3- 101.6% in n-hexane for urine and 95.7-106.3% in acetone for plasma, respectively. The calibration curves has shown good linearity with the correlation coefficients (R²) varying from 0.992 to 1.000 for urine and from 0.996 to 1.000 for plasma, respectively. The detection limits of all analytes were obtained in the range of 0.01-0.1 ng/mL for urine and 0.1-0.4 ng/mL for plasma, respectively. The metabolites of BaP were excreted as mono hydroxy and dihydrodiol forms after intraperitoneal injection of 20 mg/kg of BaP to rats. The total amounts of BaP and four metabolites excreted in dosed rat urine were 3.79 ng over the 0-96 hr period from adminstration and the excretional recovery was less than 0.065% of the injection amounts of BaP. The proposed method was successfully applied to the determination of BaP and its hydroxylated metabolites in rat urine and plasma for the pharmacokinetic studies.

• BMB Reports
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• v.33 no.2
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• pp.112-119
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• 2000

Three kinds of serine protease inhibitors, members of the Bowman-Birk trypsin inhibitor, were purified from Dolichos lablab seeds and named Dolichos protease inhibitor 1, 2 and 3 (DI-1, DI-2 and DI-3), respectively. Each inhibitor showed a single band with gel mobility at around 15.9, 12.1 and 14.6 kDa on 20% SDS-PAGE under reducing conditions. To characterize inhibitory specificity, the inhibition constant (Ki) for these inhibitors was measured against several known serine proteases. All three Dolichos protease inhibitors (DI-1, DI-2 and DI-3) inhibited the activity of trypsin and plasmin, but had no effect on thrombin and kallikrein (either for human plasma kallikrein or for porcine pancreas kallikrein). DI-1 inhibited chymotrypsin most effectively (Ki = $3.6{\times}10^{-9}\;M$), while DI-2 displayed inhibitory activity for porcine pancreatic elastase (Ki = $6.2{\times}10^{-8}\;M$). Pre-treatment of the 33 mg/kg of DI-mixture (active fractions from $C_{18}$ open column chromatography that included DI-1, DI-2 and DI-3) inhibited the induction of pseudomonal elastase-induced septic hypotension and prevented an increase in bradykinin generation in pseudomonal elastase-treated guinea pig plasma. Also, the increase of kallikrein activity, by injection of pseudomonal elastase, was inhibited by the pretreatment of the DI-mixture in a guinea pig. Since the DI-mixture had no inhibitory effect on kallikrein activity when Z-Phe-Arg-MCA was used as a substrate in vitro, its inhibitory activity in the pseudomonal elastase-induced septic hypotension model might not be due to a direct inhibition of plasma kallikrein in the activation cascade of the Hageman factor and prekallikrein system. These results suggest that the Dolichos DI-mixture might be used as an inhibitor in pathogenic bacterial protease-induced septic shock.

• Environmental Analysis Health and Toxicology
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• v.29
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• pp.18.1-18.9
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• 2014

Objectives The purpose of this study was to determine a separation method for each arsenic metabolite in urine by using a high performance liquid chromatography (HPLC)-inductively coupled plasma-mass spectrometer (ICP-MS). Methods Separation of the arsenic metabolites was conducted in urine by using a polymeric anion-exchange (Hamilton PRP X-100, $4.6mm{\times}150mm$, $5{\mu}m$) column on Agilent Technologies 1260 Infinity LC system coupled to Agilent Technologies 7700 series ICP/MS equipment using argon as the plasma gas. Results All five important arsenic metabolites in urine were separated within 16 minutes in the order of arsenobetaine, arsenite, dimethylarsinate, monomethylarsonate and arsenate with detection limits ranging from 0.15 to $0.27{\mu}g/L$ ($40{\mu}L$ injection). We used G-EQUAS No. 52, the German external quality assessment scheme and standard reference material 2669, National Institute of Standard and Technology, to validate our analyses. Conclusions The method for separation of arsenic metabolites in urine was established by using HPLC-ICP-MS. This method contributes to the evaluation of arsenic exposure, health effect assessment and other bio-monitoring studies for arsenic exposure in South Korea.

• Journal of Pharmaceutical Investigation
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• v.35 no.6
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• pp.431-436
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• 2005

A high-performance liquid chromatographic method was validated for quantitation of nadolol in human plasma. Nadolol and internal standard, pindolol, were extracted with tert-butyl methyl ether after addition of 10 M sodium hydroxide solution. The analytes were separated on a reverse phased C18 column using a mobile phase consisting of 0.05 M ammonium phosphate monobasic buffer, acetonitrile and methanol (81: 17:2 v/v/v) and detected using a fluorescence detector (excitation wavelength 230 nm, emission wavelength 294 nm). The method was specific and sensitive enough to detect as low as 3 ng/mL of nadolol in human plasma. Linear calibration range was 3-150 ng/mL with correlation coefficient greater than 0.999. The overall accuracy was in the range of 96.8 to 103% and precision C.V.(%) 7.30 to 12.2%. The recovery was approximately 100% and stability was confirmed during storage and sample preparation. The present HPLC method was successfully applied to study bioavailability after oral administration of 80 mg of nadolol in healthy Korean subjects. The mean $AUC_{t}$ was $1968{\pm}397\;ng{\cdot}hr/mL$ and $C_{max}$ of $186{\pm}79.3\;ng/mL$ was reached at $3.5{\pm}0.76\;hr$. The mean $t_{1/2}$ of nadolol was $17.3{\pm}2.59\;hr$.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.10
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• pp.1498-1504
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• 2005

Competitive direct ELISA was developed to detect gentamicin residues. Mice immunized with gentamicin-keyhole limpet hemocyanin (KLH) conjugate developed good antiserum titers, which gradually increased with booster injections, indicating immunization was successfully processed. Monoclonal antibody against gentamicin was prepared using hybridoma cells cloned by limit dilution of fused cells. IgG was purified from ascites fluid of hybridoma cell-injected mice through ammonium sulfate precipitation and Sephadex G-25 gel filtration. After the gel filtration, fractions of high antibody titer were further purified through affinity chromatography on protein A/G column. Monoclonal antibody against gentamicin was confirmed as IgG1, which has kappa light chain. Cross-reactivities ($CR_{50}$) of gentamicin monoclonal antibody to other aminoglycosides (kanamycin, neomycin, and streptomycin) were less than 0.005%, indicating the monoclonal antibody was highly specific for gentamicin. Standard curve constructed through competitive direct ELISA showed measurement range (from 80 to 20% of B/$B_0$ ratio) of gentamicin was between 1 and 40 ng/ml, and 50% of B/$B_0$ ratio was about 4 ng/ml. The gentamicin concentration rapidly increased to 1,300 ng/ml after the intramuscular administration up to 2 h, then sharply decreased to less than 300 ng/ml after 4 h of withdrawal, during which the elimination half-life ($t_{1/2}$) of gentamicin in the rabbit plasma was estimated to be 1.8 h. Competitive direct ELISA method developed in this study using the prepared monoclonal antibody is highly sensitive for gentamicin, and could be useful for detecting gentamicin residues in plasma of live animals.

• Journal of Pharmaceutical Investigation
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• v.28 no.3
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• pp.171-175
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• 1998

KHPC-002 $[(trans-l-diaminocyclohexane-bis-l,2(diphenylphosphinoethane)platinum)\;{\cdot}2NO_3]$ and $KHPC-006[(cis-diaminocyclohexane-bis-1,2(diphenylphosphinoethane)platinum)\;{\cdot}2NO_3]$ were synthesized as candidates for third platinum antitumor agent. Before their pharmacokinetic study, we optimized the analytical condition with HPLC and identified the major metabolites in the rat plasma. HPLC analysis by $C_{18}$ reverse-phase column showed that standard peak of both compounds appeared rapidly at around 1 minutes, whereas metabolites of KHPC-002 and KHPC-006 which were extracted from plasma after single I.V. administration in rats or incubation for 24 hr at $37^{\circ}C$ showed retention time of $10{\sim}11$ minutes. These metabolites were identified as the major compound by Matrix Associated Laser Deposition/Ionization (MALDI), which only lose the 2 molecules of $NO_3$. Based on these results, we suggest that the major metabolites of KHPC-002 and KHPC-006 were [trans-l-diamino-cyclohexane-bis-l,2(diphenylphosphinoethane)platinum] and [cis-diaminocyclohexane-bis-l.2(diphenylphosphinoethane)platinum], respectively.

• Analytical Science and Technology
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• v.8 no.3
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• pp.229-236
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• 1995

Multiple-step gradient method was used for the analysis of free amino acids in physiological fluids by high-performance liquid chromatography with diode array detector on the Amino Quant $C_{18}$ column under the condition of pH 7.2 of buffer solutions. Plasma samples of normal Korean people and abnormal Korean people who have schizophrenia were subjected to derivatization with o-phthalaldehyde in the presence of 3-mercaptopropionic acid. Quantitative analysis of amino acids in physiological fluids by internal standard method gave highly reproducible results within a relative standard deviation of less than 2~6%. And amino acids amounts of physiological fluids of Korean people gave some different results from those of foreigners. There was large differences in tyrosine amount between normal and abnormal man.

• YAKHAK HOEJI
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• v.45 no.4
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• pp.378-386
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• 2001

A purified histone Hl protein, p961, which plays a role in mediating the condensation of DNA into chromatin, was recently proved as an arthritis-suppressing agent in the mouse CIA model. The pharmacokinetics of p961 was carried out in rats and rabbits. The rat's blood, bile and urine samples were serially collected from the femoral vein, common bile duct, and bladder respectively, after bolus i.v. injection at low (10 mg/kg) and high (30 mg/mg) doses. The rabbit's blood samples were also collected from the marginal ear vein after bolus i.v. injection at a dose 10 mg/kg. p961 and its major metabolite in the physiological samples were analyzed by reverse-phase HPLC using a Yydac C4 protein column and a multistep water-acetonitrile gradient containing 0.24% trifluoroacetic acid. The major pharmacokinetic parameters (AUC, $C_{max}$, MRT, $t_{1}$2/, $V_{ss}$ and Cl) were estimated from the time course of plasma p961 and metabolite concentrations using WinNonlin. A two-compartment model was chosen for p961 as the most appropriate pharmacokinetic model. After i.v. injection of p961 at doses of 10 mg/kg and 30 mg/kg, more than 80% of p961 was removed rapidly from the plasma within 15 min. The plasma half-life of p961 in rats and rabbits was found not to exceed 12 min. p961 (22448.9 mol wt) was rapidly cleaved to 21612 mot wt fragment and the breakdown product appeared rapidly in the circulation with no lag phase. p961 and metabolite were not detected in rat urine and bile....

• Bulletin of the Korean Chemical Society
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• v.33 no.1
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• pp.177-182
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• 2012

A new composition of standardized Scutellariae Radix extract (HPO12) was developed for treatment of Alzheimer's disease. For the preclinical pharmacokinetic study of HPO12, a rapid, sensitive, and selective LCMS/MS method was developed and validated for the simultaneous determination of 4 bioactive compounds, baicalein, baicalin, wogonin, and wogonoside. After extraction with ethylacetate, chromatographic analysis was performed on a Thermo $C_{18}$ column ($150mm{\times}2.1mm$, $3{\mu}m$) with a mobile phase consisting of 0.1% formic acid (A) and 0.1% formic acid in 95% acetonitrile (B) by using gradient elution at a flow rate of $250{\mu}L/min$. Analytes introduced to a mass spectrometer were monitored by multiple reaction monitoring (MRM) in positive ion mode. Using $25{\mu}L$ of plasma sample, the method was validated over the following concentration ranges: 25-5000 ng/mL for baicalein, 20-40000 ng/mL for baicalin, 1-1000 ng/mL for wogonin, and 5-10000 ng/mL for wogonoside. The intra- and inter-day precision and accuracy of the quality control samples at the 4 concentrations showed $\leq$ 13.7% relative standard deviation (RSD) and 86.6-105.5% accuracy. The method was successfully applied to determine the concentrations of baicalein, baicalin, wogonin, and wogonoside in rat plasma after intraperitoneal and oral administrations of HPO12.