Seung-Woon Myung

Determination of amphetamine, methamphetamine and dimethamphetamine in Human Urine by Solid-Phase Microextraction (SPME)-Gas Chromatography/Mass Spectrometry

A simple and rapid assay method for three stimulant drugs (amphetamine, methamphetamine, and dimethamphetamine) in human urine using solid-phase microextraction was developed. In solid-phase microextraction, the drugs were equilibrated between the adsorbent coated-fiber and aqueous sample matrix. After adsorption of the analytes, the fiber was directly transferred to the injector of a gas chromatograph, where the analytes were thermally desorbed and subsequently separated by the gas chromatograph and detected by mass spectrometer. The solid-phase microextraction method, which did not require solvents, was found to be a fast and simple analytical method. We optimized the solid-phase microextraction technique, for factors such as the NaCl salt effect (30%), pH effect (pH=12.4), equilibration time (30 min), desorption time (1 min) and coated-fiber type (100 microm poly(dimethylsiloxane)) and detected the stimulants in human urine, obtained from human subjects. The detection limits of each drug were below 1-10 ng/ml. The developed method can be applied to the abused drug test.

Absorption, distribution, metabolism, and excretion of CKD-732, a novel antiangiogenic fumagillin derivative, in rats, mice, and dogs

The pharmacokinetics of CKD-732 (6-0-4-[dimethyl-aminoethoxy)cinnamoyl]-fumagillol hemioxalate) was investigated in male SD rats and beagle dogs after bolus intravenous administration. The parent compound and metabolites obtained fromin vitro andin vivo samples were determined by LC/MS. The main metabolite was isolated and identified as anN-oxide form of CKD-732 by NMR and LC/MS/MS. CKD-732 was metabolized into eitherM11 or others by rapid hydroxylation, demethylation, and hydrolysis. The blood level following the intravenous route declined in first-order kinetics with T1/2β values of 0.72~0.78 h for CKD-732 and 0.92~1.09 h forM11 in rats at a dose of 7.5-30 mg/kg. In dogs, T1/2β values of CKD-732 andM11 were 1.54 and 1.79 h, respectively. Moreover, AUC values increased dose dependently for CKD-732 andM11 in rats and dogs. The CLtot and Vdss did not change significantly with increasing dose, indicating linear pharmacokinetic patterns. The excretion patterns through the urine, bile, and feces were also examined in the animals. The total amount excreted in urine, bile, and feces was 2.13% for CKD-732 and 1.29% forM11 in rats, and 1.58% for CKD-732 and 2.28% forM11 in dogs.

Gas chromatographic profiling and pattern recognition analysis of urinary organic acids from uterine myoma patients and cervical cancer patients

An efficient organic acid profiling and pattern recognition method is described for the correlation between urinary organic acid profiles and uterine cervical cancer. After methoximation of keto acids in alkalinized urine samples, all free organic acids were recovered by a dual solid-phase extraction procedure, followed by conversion to tert.-butyldimethylsilyl derivatives for the profiling analysis by dual-capillary column gas chromatography (GC) with subsequent screening for acids by retention index (I) library matching. A total of 50 organic acids were positively identified in urine samples (0.25 ml) from 12 uterine myoma (benign tumor group) and 14 uterine cervical cancer (malignant tumor group) patients studied. When the GC profiles were simplified to their corresponding organic acid I spectra in bar graphical form, characteristic patterns were obtained for each average of benign and malignant tumor groups. Stepwise discriminant analysis performed on the GC data selected 16 acids as the variables discriminating between the two groups. Canonical discriminant analysis applied to these 16 variables correctly classified 26 urine samples into two separate clusters according to tumor types in the canonical plot.

Determination of homocysteine and its related compounds by solid-phase microextraction-gas chromatography-mass spectrometry.

The purpose of this study was to develop a simple and accurate analytical method to determine homocysteine (Hcy), cysteine (Cys), and methionine (Met) in aqueous samples. Until now, the most frequently used method for the assay of Hcy, Cys, and Met has been high-performance liquid chromatography with fluorescence detection after fluorescent tagging. The newly developed method involves the employment of the SPME (solid-phase microextraction) technique together with GC-MS. For application to a gas chromatographic system, alkyl formate derivatives were prepared in the form of N(O,S)-alkoxycarbonyl alkyl ester with the analytes in the aqueous samples. The optimum derivatizing regent for N(O,S)-alkoxycarbonyl alkyl ester was chosen by comparing the efficiency of the derivatized analytes in a GC through the SPME method and liquid-liquid extraction. The optimum conditions of the SPME system for the analytes derivatized with N(O,S)-ethoxycarbonyl propyl ester in the aqueous matrix were pH 3.0 and no salt, and 30 min equilibration time using an 85 microm PA (polyacrylate) fiber. The developed method is inexpensive, easy and rapid.

Solid-phase microextraction for the determination of pethidine and methadone in human urine using gas chromatography with nitrogen–phosphorus detection

A simple and rapid analytical method is presented for the determination of pethidine (meperidine) and methadone in human urine using solid-phase microextraction (SPME) and gas chromatography with nitrogen-phosphorus detection (GC-NPD). After the analytes had been partitioned between an extracting phase and the aqueous sample matrix, the needle of the coating fiber assembly was injected directly into the GC injector. The analytes were thermally desorbed in the heated injector (240 degrees C) and subsequently separated and detected by the GC-NPD system. The factors influencing the SPME method, such as the salt (NaCl) effect (15%), pH (pH 11), and equilibration time (30 min), were optimized. The calibration graphs for urine samples showed a good linearity. The detection limit was below 1 ng ml-1 for both drugs.

Identification of IY81149 and its metabolites in the rat plasma using the on-line HPLC/ESI mass spectrometry

Reversed-phase high-performance liquid chromatography/mass spectrometry (HPLC/MS) with an electrospray ionization (ESI) interface was applied to the identification of metabolites of IY 81149 in the rat plasma. Fragments obtained using collision-induced dissociation (CID) in both positive and negative modes were utilized to elucidate the structure of metabolites. The eluent from the conventional HPLC column was split and directly introduced into an ESI-mass spectrometer for the identification of the structures. The CID technique allowed the sensitive identification of sulfonyl-IY81149 and hydroxy-IY81149 from the rat plasma.

Gas chromatographic-mass spectrometric analysis of mercaptan odorants in liquefied petroleum gas and liquefied natural gas

Abstract A gas chromatographic–mass spectrometric method for the determination of mercaptan odorants (dimethyl sulfide, tert .-butylmercaptan, tetrahydrothiophene) in natural gas has been developed. The gas sample filled in a 5 l Tedlar bag was introduced into the 0.5 ml volume of a sampling loop, separated on a 50 m capillary column coated with 5% phenylmethylsilicone and detected by a mass spectrometer. Natural gas samples collected from 124 sites were analyzed and the concentration of added odorants was found to be between 9.7 and 66.2 ppm (w/w). The detection limit of each odorant was below 1 ppm (w/w). The advantages of the developed technique were lower detection limits, elimination of interference peaks by introducing into the sampling loop system and selected-ion monitoring mode, and reduced total analysis time. The intra-day and inter-day precision of the established method was R.S.D. n =5).

Characterization of amiodarone metabolites and impurities using liquid chromatography/atmospheric pressure chemical ionization mass spectrometry

Using the high performance liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry (HPLC/APCI-MS/MS) technique, together with established trends from the literature, the structures of metabolites and impurities of amiodarone, an anti-arrhythmic drug, have been assigned. By comparing analyses of products of incubation with rat liver microsomes with controls in which glucose 6-phosphate dehydrogenase was omitted, metabolites could be distinguished from impurities. Structures for the two proposed metabolites and four impurities are proposed.

Genotoxicity study of bojungchisup-tang, an oriental herbal decoction-in vitro chromosome aberration assay in Chinese hamster lung cells andin vivo supravital-staining micronucleus assay with mouse peripheral reticulocytes

The toxicity evaluation of oriental herbal drugs is of great concern at present. Bojungchisuptang (BCST, in Korean), a decocted medicine of oriental herbal mixture, is now well used in clinic at oriental hospitals for the treatment of edema of several diseases in practice. However, the toxicity of the oriental herbal decocted medicines such as genetic toxicity is not well defined until now. In this respect, to clarify the genetic toxicity of BCST,in vitro chromosome aberration assay with Chinese hamster lung (CHL) fibroblasts andin vivo supravital micronucleus assay with mouse peripheral reticulocytes were performed in this study. In the chromosome aberration assay, we used 5,000 μg/ml BCST as maximum concentration because no remarkable cytotoxicity in CHL cells was observed both in the presence and absence of S-9 metabolic activation system. No statistical significant differences of chromosome aberrations were observed in CHL cells treated with 5,000, 2,500 and 1,250 μg/ml BCST for 6 hour both in the presence and absence of S-9 metabolic activation. However, very weak positive result (6.5∼8.0% aberration) of BCST was obtained in the absence of S-9 metabolic activation system at 5,000 μg/ml BCST when treated for 24 hour, i.e. 1.5 normal cell cycle time. And also,in vivo clastogenicity of BCST was studied by acridine orange-supravital staining micronucleus assay using mouse peripheral reticulocytes. We used 2,000 mg/kg as the highest oral dose in this micronucleus assay because no acute oral toxicity of BCST was observed in mice. The optimum induction time of micronucleated reticulocytes (MNRETs) was determined as 36 hours after oral administration of 2,000 mg/kg BCST. No significant differences of MNRETs between control and BCST treatment groups were observedin vivo micronucleus assay. From these results, BCST revealed very weak positive result in chromosome aberration assayin vitro with CHL cells and no clastogenicity in micronucleus assayin vivo.

Determination of carphedon in human urine by solid-phase microextraction using capillary gas chromatography with nitrogen–phosphorus detection

Carphedon is a phenyl derivative of nootropil and is effective in increasing physical endurance and cold resistance, and is used for amnesia treatment. Carphedon was extracted from human urine samples by solid-phase microextraction with a 65 microns carbowax-divinylbenzene-coated fiber. This analysis was performed by using capillary gas chromatography with nitrogen-phosphorus detection and optimized at pH 9.6, 30% NaCl, immersion time 10 min and desorption in the GC injector at 250 degrees C for 3 min. The regression equation for carphedon showed good linearity in the range from 0.1 to 10 micrograms ml-1 for human urine samples. The limit of detection was 0.01 microgram ml-1. The developed method is more sensitive and simpler in sample preparation than liquid-liquid extraction and can be applied to doping analysis for stimulants.