Stereoselective high-performance liquid chromatographic assay for pirmenol enantiomers in dog plasma.

Pirmenol enantiomers in dog plasma were quantified using a stereospecific high-performance liquid chromatographic method with ultraviolet detection at 262 nm. Racemic pirmenol and internal standard, (+)-propranolol, were isolated from dog plasma by a three-step extraction procedure using toluene, 0.1 M hydrochloric acid and hexane, respectively. A chiral analytical column (Chiralcel OJ) was used with a mobile phase consisting of hexane-isopropanol-diethylamine (98.9:1.0:0.1). Linear calibration curves were obtained in the concentration range 0.0200-5.00 micrograms/ml for each enantiomer. Precision of the method, expressed as coefficient of variation for nine quality control samples, was 7.1% for (+)-pirmenol and 6.4% for (-)-pirmenol. Bias was +/- 2.2% for (+)-pirmenol and +/- 1.5% for (-)-pirmenol in quality control samples.     

Stereoselective determination of the enantiomers of methadone in plasma using high-performance liquid chromatography.

An enantioselective high-performance liquid chromatographic assay for the quantification of methadone in human and beagle plasma is described. The procedure involves extraction of methadone from alkalized plasma into hexane-isoamyl alcohol (99:1, v/v). Stereoselective separation was achieved with a silica column with covalently bound alpha 1-acid glycoprotein (Chiral-AGP) without any derivatization procedure. The detection wavelength was set at 215 nm. Using an internal standard provided reliable control of the extraction procedure as well as quantification of the enantiomers of methadone. The limit of quantification was found to be 2.5 ng/ml. The method was demonstrated to be sufficiently sensitive for stereoselective pharmacokinetic studies of methadone.     

Stereoselective high-performance liquid chromatographic assay for propranolol enantiomers in serum

A simple and sensitive stereoselective high-performance liquid chromatographic assay for the quantitation of propranolol enantiomers in serum is described. The method involves conversion of the propranolol enantiomers to diastereomeric urea derivatives by reaction with the clinical reagent (+)-phenylethylisocyanate, followed by chromatographic separation of the diastereomeric products. Conditions of the derivatization reaction were optimized to achieve rapid and quantitative yield with either of the enantiomers. Baseline resolution of the diastereomers was achieved on a reversed phase C8 column with an isocratic mobile phase. Fluorescence detection afforded an absolute on-column detection limit of 100 pg. The assay has been applied to pharmacokinetic studies in humans and small laboratory animals.     

Solid-phase extraction and direct high-performance liquid chromatographic determination of metoprolol enantiomers in plasma

A method is described by which underivatized metoprolol enantiomers in plasma can be quantitated by high-performance liquid chromatography with fluorescence detection. Samples are prepared for injection using a simple solid-phase extraction procedure which is essentially 100% efficient for all analytes. The metoprolol enantiomers are resolved using a cellulose tris(3,5-dimethylphenylcarbamate) chiral stationary phase and a hexane-ethanol-diethylamine mobile phase. Samples were tested for adaptability to autoinjection and found to be stable for at least 16 h after reconstitution in mobile phase. The automated method was determined to be precise and accurate for enantiomer concentrations as low as 10 ng base per ml and was successfully employed in a pharmacokinetic investigation.     

A stereospecific high-performance liquid chromatographic assay for the determination of ketoconazole enantiomers in rat plasma

A stereospecific high-performance liquid chromatographic assay was developed for the quantitation of ketoconazole enantiomers (KTZ) in rat plasma. After protein precipitation of 100 microL plasma using acetonitrile, a wash step was performed using hexane. The supernatant was removed and KTZ enantiomers and amiodarone, the internal standard, were extracted using liquid-liquid extraction with tert-butyl methyl ether. After transfer and evaporation of the organic layer, the residue was reconstituted in mobile phase and injected into the HPLC through a chiral column. The mobile phase consisted of hexane:ethanol:2-propanol with diethyl amine, pumped at 1.5 mL/min. All components eluted within 18 min. KTZ enantiomers were baseline resolved and peaks were symmetrical in appearance with no interferences. Calibration curves were linear over the range 62.5-5000 ng/mL of enantiomer. The intraday and interday CV% assessments were 相似文献   

Stereospecific high-performance liquid chromatographic assay of pirprofen enantiomers in rat plasma and urine.

A stereospecific high-performance liquid chromatographic method was developed for the assay of pirprofen enantiomers in rat plasma and urine. Following addition of internal standard (ketoprofen) and acidifier (L-ascorbic acid) to biological fluids, pirprofen was extracted into an isopropanol-isooctane (5:95) mixture. Diastereomers of pirprofen enantiomers, which were formed using L-leucinamide, were separated on a reversed-phase column with ultraviolet detection at 275 nm using 0.06 M KH2PO4-acetonitrile-triethylamine (64:36:0.1) as mobile phase. The limit of quantitation was 0.1 microgram/ml for each enantiomer, based on 100 microliters of rat plasma. No spontaneous oxidation of pirprofen to its pyrrole metabolite occurred during sample preparation and analysis. In three female rats which were dosed with 10 mg/kg racemic pirprofen orally, plasma concentrations of the enantiomers could be followed for 24 h. Pirprofen enantiomers in plasma were virtually unconjugated, and negligible concentrations of pyrrole metabolites were observed. Less than 10% of the total dose was recovered in urine as intact drug and its glucuroconjugates. The assay was found suitable for the study of the pharmacokinetics of pirprofen enantiomers in the rat.     

Isocratic high-performance liquid chromatographic determination of plasma adenosine

Summary Due to manifold physiological and cardioprotective actions of adenosine, the demand for a simple but accurate method to determine its concentration in plasma is increasing. The aim of this study was firstly to develop a simple isocratic method instead of the gradient elution or peak-shifting techniques used earlier and secondly to check conflicting data on the composition of stop-solution, added to the sample in order to prevent changes in adenosine concentration. Isocratic elution improved signal to noise ratio and concentrations of 100 mol L^–1 dipyridamole and 2.5 mol L^–1 erythro-9(2-hydroxy-3-nonyl)adenine in the blood sample effectively prevented both adenosine formation and degradation, even without the use of a 5-ecto-nucleotidase inhibitor. Lowering the concentration of dipyridamole to 25 mol L^–1 caused more than a tenfold increase of adenosine concentration in two out of five cases and even 100 mol L^–1 dipyridamole alone is not sufficient to inhibit adenosine deaminase in blood samples.     

Sensitive high-performance liquid chromatographic determination of pirenzepine in plasma

A high-performance liquid chromatographic method for the determination of pirenzepine in human plasma is reported using imipramine as an internal standard. The assay has a lower limit of detection of 2.5 ng/ml. The calibration function is found to be linear in the range from 5 ng/ml up to at least 100 ng/ml. Two sets of chromatographic conditions are described, which provide different chromatographic selectivities for the separation of the compounds of interest from other material present in a sample.     

Gas chromatographic analysis of ketamine and norketamine in plasma and urine: nitrogen-sensitive detection

A sensitive gas chromatographic method for quantitative analysis of ketamine and norketamine in human and animal biological fluids is described. The nitrogen-sensitive detection procedure used is more stable than electron-capture detection and reduced analysis time. The method used bromo-ketamine as an internal standard for quantitation and is linear from 10-25,000 ng/ml. No interferences were shown with drugs commonly associated with cardiac surgery with cardiopulmonary by-pass. This assay is sensitive, specific, using either native or derivatized drugs and can be used for routine analysis of ketamine and norketamine in plasma or urine.     

Activated alpha-alkyl-alpha-arylacetic acid enantiomers for stereoselective thin-layer chromatographic and high-performance liquid chromatographic determination of chiral amines

The separation of racemic benoxaprofen into the two benoxaprofen enantiomers by preparative high-performance liquid chromatography and the application of the activated enantiomers as derivatization reagents for the simultaneous stereoselective determination of chiral amines in biological material is described. Activated (+)- and (-)-benoxaprofen are both shown to be very sensitive and stable chiral fluorescence markers, applicable to thin-layer chromatography as well as to high-performance liquid chromatography.     

Characterization of the stereoselective biotransformation of ketamine to norketamine via determination of their enantiomers in equine plasma by capillary electrophoresis

A robust CE method for the simultaneous determination of the enantiomers of ketamine and norketamine in equine plasma is described. It is based upon liquid-liquid extraction of ketamine and norketamine at alkaline pH from 1 mL plasma followed by analysis of the reconstituted extract by CE in the presence of a pH 2.5 Tris-phosphate buffer containing 10 mg/mL highly sulfated beta-CD as chiral selector. Enantiomer plasma levels between 0.04 and 2.5 microg/mL are shown to provide linear calibration graphs. Intraday and interday precisions evaluated from peak area ratios (n = 5) at the lowest calibrator concentration are < 8 and < 14%, respectively. The LOD for all enantiomers is 0.01 microg/mL. After i.v. bolus administration of 2.2 mg/kg racemic ketamine, the assay is demonstrated to provide reliable data for plasma samples of ponies under isoflurane anesthesia, of ponies premedicated with xylazine, and of one horse that received romifidine, L-methadone, guaifenisine, and isoflurane. In animals not premedicated with xylazine, the ketamine N-demethylation is demonstrated to be enantioselective. The concentrations of the two ketamine enantiomers in plasma are equal whereas S-norketamine is found in a larger amount than R-norketamine. In the group receiving xylazine, data obtained do not reveal this stereoselectivity.     

Direct high-performance liquid chromatographic resolution of dihydropyridine enantiomers

A direct high-performance liquid chromatographic enantioseparation on a chiral stationary phase using a protein as chiral selector was achieved for some dihydropyridines with different structural groups (amine, amide, acid and hydroxyl groups). The protein involved was alpha 1-acid glycoprotein and the work was mainly carried out with an EnantioPac column. An improved version of this chiral column (Chiral-AGP) was compared with the previous one and gave interesting results.     

Analytical and preparative high-performance liquid chromatographic separation of the enantiomers of ifosfamide, cyclophosphamide and trofosfamide and their determination in plasma

A high-performance liquid chromatography chiral stationary phase (HPLC-CSP) based upon cellulose-tris(3,5-dimethylphenylcarbamate), the OD-CSP, was used to stereochemically resolve three racemic anticancer drugs: ifosfamide (IFF), cyclophosphamide (CTX) and trofosfamide (TFF). The observed enantioselectivities (alpha) were 1.45 (IFF), 1.21 (CTX) and 1.10 (TFF). The OD-CSP was also used on a preparative scale to isolate the enantiomers of IFF with an optical purity greater than 97%. In addition, an analytical assay was developed for the determination of the enantiomeric composition of IFF and CTX in plasma using achiral-chiral coupled column chromatography. A racemic form of another CSP, the D,L-naphthylalanine-CSP, was used as the achiral pre-column. The sample clean-up prior to HPLC analysis was realized by liquid-liquid extraction with chloroform. The correlation coefficient was 0.987 for both standard curves (20-130 micrograms/ml) and the controls were within 10% of the real value. The analysis time for each serum sample was less than 30 min.     

Stereoselective high-performance liquid chromatographic assay with fluorometric detection for the isomers of mivacurium in human plasma.

A high-performance liquid chromatographic assay with fluorometric detection was developed for the analysis of the stereoisomers of mivacurium, a new short-acting neuromuscular blocker, in plasma. The isomers were isolated from plasma by solid-phase extraction with C18 and anion-exchange cartridges. The extracts were chromatographed on a LiChrosphere 60 RP Select B column (125 mm x 4.6 mm I.D.) using a mobile phase of acetonitrile-water (4:6, v/v) containing 0.005 M octanesulfonic acid. The fluorescence excitation and emission wavelengths were 202 and 320 nm, respectively. The accuracy and precision of the assay, expressed as the percentage deviation of measured values from true values and the percentage coefficient of variation, respectively, were less than or equal to 10% at all concentrations except for the percentage coefficient of variation at the lower limit of quantitation (5 ng/ml). The assay has been successfully used for the analysis of plasma samples from a pharmacokinetic study in human volunteers.     

An improved high-performance liquid chromatographic determination of chlorpropamide in human plasma

Summary Samples were extracted with dichloromethane and the organic layer evaporated to dryness. The residue was dissolved in methanol, and 10 μl aliquot injected onto the column. Tolbutamide was used as the internal standard for chlorpropamide. The UV detector response was linear over the range 0–200 μg ml⁻¹ with a correlation coefficient of 0.999; detection limit: 0.002 μg ml⁻¹. Within-day and between-day assay variation was generally ≤7%. No interference from endogenous constituents was observed. The utility of the method was demonstrated by determining chlorpropamide in samples from six healthy volunteers following a single oral dose of 250 mg. The procedure is simple and requires small volumes of plasma.