Kiyoshi Banno

Analytical chiral separation of a new quinolone compound in biological fluids by high-performance liquid chromatography

Two methods for the separation of a new racemic quinolone compound, temafloxacin (TMFX), in biological fluids by high-performance liquid chromatography (HPLC) were studied. The first method was coupling of TMFX to S-(-)-N-1-(2-naphthyl sulfonyl)-2-pyrrolidine carbonylchloride (L-NSPC). The diastereomeric derivatives were separated on a silica gel column. The second method was separation on a chiral stationary phase with an ovomucoid conjugated to aminopropyl silica gel. Two enantiospecific methods gave a satisfactory result concerning both accuracy and precision, and the second method was superior to the first one for chromatographic separation. Furthermore, the pharmacokinetics of the enantiomers after oral administration of racemic TMFX to healthy volunteers was investigated by the second method.

Radioimmunoassay for imidapril, a new angiotensin-converting enzyme inhibitor, and imidaprilat, its active metabolite, in human plasma and urine

A radioimmunoassay (RIA) was investigated for the determination of imidapril and its active metabolite, imidaprilat, in human plasma and urine. Imidapril is a new angiotensin-converting enzyme inhibitor and an oral prodrug of imidaprilat. Imidapril was determined after conversion to imidaprilat with esterase. Antiserum was raised in rabbits against the p-amino derivative of imidaprilat conjugated to bovine serum albumin. Radioligand was prepared by iodination (125I) of the p-hydroxybenzoylamino derivative of imidaprilat. Cross-reactivities of anti-imidaprilat antiserum for imidapril, its metabolites and several cardiovascular drugs were low. The calibration range was 0.1-100 ng ml-1 using a 100 microliters of human plasma of urine. Intra- and inter-day variations of imidaprilat assay in plasma were 2.0-7.9 and 4.1-6.2%, respectively, and intra- and inter-day variations of imidapril assay in plasma were 5.4-10.7 and 7.9-18.1%, respectively. The variations of the assay in urine were a little smaller than those in plasma. The recovery of imidaprilat and imidapril spiked in plasma or urine samples was approximately 100%. A good correlation between RIA and high-performance liquid chromatograpy was observed for both plasma and urine samples. Furthermore, this method was applied to the determination of imidaprilat and imidapril in human plasma and urine samples, for the evaluation of the pharmacokinetics of imidapril in humans. From the results, it was demonstrated that the developed RIA was useful for the determination of imidaprilat and imidapril in human plasma and urine, and was applicable to pharmacokinetic studies in humans.

Simultaneous determination of pantothenic acid and hopantenic acid in biological samples and natural products by gas chromatography-mass fragmentography.

A method for the simultaneous determination of pantothenic acid and hopantenic acid in plasma samples was developed using gas chromatography-mass spectrometry with multiple ion detection. Plasma samples were directly purified without deproteinization on an ion-exchange resin, and the eluate was extracted with ethyl acetate under acidic conditions. The organic layer was evaporated to dryness under a stream of nitrogen, and the residue was dissolved in an internal standard solution. Pantothenic and hopantenic acids were converted into their trimethylsilyl derivatives by treating with bis(trimethylsilyl)trifluoroacetamide. Aliquots of this solution were injected into the gas chromatograph-mass spectrometer, which was equipped with a wide-bore fused-silica column (DB-17) and analysed by the multiple ion detection method. The detection limits for pantothenic acid and hopantenic acid in plasma were 1 ng/ml each at a signal-to-noise ratio of 5. This method was applied to a study of the assay of pantothenic acid and hopantenic acid in biological samples and natural products.

Assay of nicergoline and three metabolites in human plasma and urine by high-performance liquid chromatography-atmospheric pressure ionization mass spectrometry.

A method for the simultaneous determination of nicergoline and three of its metabolites in human plasma and urine has been developed using high-performance liquid chromatography-atmospheric pressure ionization mass spectrometry. Nicergoline and its metabolites were extracted from the plasma and urine samples with chloroform and separated on a reversed-phase ODS column. The eluents were led to the atmospheric pressure ionization interface and then analysed in the selected-ion monitoring mode. The detection limits of nicergoline and three of its metabolites were ca. 2 ng/ml in plasma and ca. 10 ng/ml in urine, at a signal-to-noise ratio of 4.

Determination of three metabolites of a new angiotensin-converting enzyme inhibitor, imidapril, in plasma and urine by gas chromatography-mass spectrometry using multiple ion detection

A specific and sensitive gas chromatographic-mass spectrometric method for the determination of three metabolites of the angiotensin-converting enzyme inhibitor, imidapril, in plasma and urine was developed. The metabolites were isolated from plasma and urine using a Bond Elut C18 solid-phase extraction cartridge. The isolated metabolites were converted to sensitive derivatives by pentafluorobenzyl bromide and heptafluoro-n-butyric acid anhydride. Following derivatization, the sample solutions were analysed by wide-bore column gas chromatography-mass spectrometry with multiple ion detection. The detection limits of the three metabolites were each 1 ng/ml in plasma and 5 ng/ml in urine. Analysis of the spiked plasma and urine samples demonstrated the good accuracy and precision of the method. This method was very useful for use in pharmacokinetic and bioavailability studies of the three metabolites of imidapril in humans.

Determination of diltiazem hydrochloride enantiomers in dog plasma using chiral stationary-phase liquid chromatography.

The separation and determination of d- and l-diltiazem hydrochloride in dog plasma by a two-column high-performance liquid chromatographic technique are described. Diltiazem hydrochloride and its metabolites were extracted from dog plasma and analyzed on a conventional column (Nucleosil 5C18) with a volatile buffer system. The column effluent of diltiazem hydrochloride was collected and evaporated. The enantiomeric ratio of the collected diltiazem was determined using a chiral column (Chiralcel OC). The method was accurate and sensitive.

Determination of thiamine in dried yeast by high-performance liquid chromatography using a clean-up column of CM-cellulose

Abstract Thiamine in dried yeast was cleaned up with a CM-cellulose column and determined by HPLC on a reversedphase ODS column with sodium 1-octanesulfonate as an ion-pairing agent and UV detection. Thiamine and phenacetin as an internal standard were monitored with UV detection at 254 nm. The calibration graph was linear in the range 0.25–4 μg/ml of thiamine (as thiamine hydrochloride) with a correlation coefficient of 0.998. Intra- and inter-day variations of thiamine in dried yeast were 6.5 and 6.9%, respectively. Thiamine was recovered in good yield (97.8–105.7%, n = 5). By using this method, the thiamine content in dried yeast was found to be 138–169 μg/g. These results were in good agreement with those of the conventional thiochrome method.

Radioimmunoassay for TA-0910, a new stable thyrotropin releasing hormone analogue and its metabolite, TA-0910 acid-type, in human plasma and urine

Radioimmunoassay (RIA) was investigated for the determination of TA-0910 and its main metabolite, TA-0910 acid-type, in human plasma and urine. TA-0910 is a new metabolically stable analogue of thyrotropin releasing hormone (TRH). Antiserum was raised in the rabbit against the 1-fluoro-2,4-dinitrophenyl derivative of TA-0910 or TA-0910 acid-type conjugated to keyhole limpet hemocyanin (KLH). The radioligand was prepared by iodination with 125I of the histidine imidazole ring of TA-0910 or TA-0910 acid-type. Cross-reactivities of anti-TA-0910 or TA-0910 acid-type antiserum for TA-0910, its metabolite and related compounds were low. The calibration range was 0.02-5 ng ml-1 using 100 microliters human plasma or urine. Inter-day variations of TA-0910 and TA-0910 acid-type assay in plasma were 3.5-15.5 and 1.8-9.4%, respectively. The variations of the assay in urine were the same as those in plasma. The recovery of TA-0910 and TA-0910 acid-type spiked in plasma or urine samples was approximately 100%. Furthermore, this method was applied to the determination of TA-0910 and TA-0910 acid-type in human plasma and urine samples, for the evaluation of the pharmacokinetics of TA-0910 in humans. From the results it was demonstrated that he developed RIA was useful for the determination of TA-0910 and TA-0910 acid-type in human plasma and urine, and was applicable to pharmacokinetic studies in humans.

Pharmacokinetic and pharmacodynamic study of imidaprilat, an active metabolite of imidapril, a new angiotensin-converting enzyme inhibitor, in spontaneously hypertensive rats

The pharmacokinetics and pharmacodynamics (PK/PD) of imidaprilat, an active metabolite of imidapril, a new angiotensin-converting enzyme (ACE) inhibitor, were investigated. Imidapril was infused subcutaneously for 4 weeks via an osmotic pump implanted under the skin in the back of male spontaneously hypertensive rats (SHRs). Plasma concentration of imidaprilat, systolic blood pressure (SBP), and plasma ACE activity were determined periodically. The plasma concentration of imidaprilat increased in proportion to the infusion rates and was maintained for 4 weeks. The SBP and ACE activity did not decrease in proportion to the infusion rates due to the saturation of the pharmacologic effects, but these actions also were maintained for 4 weeks. The PK/PD of imidaprilat were not influenced by aging of SHRs. The antihypertensive action in subcutaneous infusion of imidapril was as potent as that in oral administration at the same dose, although the maximum plasma concentration of imidaprilat in subcutaneous infusion was one-eightieth times of that in oral administration. The action was also maintained 28 times longer than that in oral administration, indicating that subcutaneous infusion is useful as an administration route. Furthermore, good correlation between plasma imidaprilat concentration and SBP was observed in subcutaneous infusion, indicating that plasma concentration may be a useful marker of pharmacologic action.

Liquid chromatographic analysis of penem antibiotic FCE22101 in biological fluids with a liquid-liquid extraction procedure

A sensitive high-performance liquid chromatographic method has been developed for the determination of penem antibiotic FCE22101 in plasma and urine. FCE22101 was extracted from plasma and urine with ethyl acetate. After evaporating, the sample solutions were analyzed by a reversed-phase high-performance liquid chromatographic system using a two-sided bracketing injection technique. The determination limit of FCE22101 was 5 ng/ml in plasma. Analysis of the spiked plasma samples demonstrated the good accuracy and precision of the method. The proposed method improved from five- to ten-fold on the analytical sensitivity in comparison with the most commonly ultrafiltration method.