Development and application of a calibration regression routine in conjunction with linear and non-linear chromatographic detector responses.

Linear regression is widely used in the calibration of chromatographic assays even in conjunction with some chromatographic detectors which show significant non-linearity in their response characteristics. A calibration routine, based upon the curve y = axlnx+bx+c is presented which describes the non-linear behaviour of some chromatographic systems, including electron capture, nitrogen-phosphorus and UV photometric detectors, and gives comparable results to weighted linear regression with assays showing linear concentration versus response relationships. The ratio of the coefficients a and b in the equation allows quantification of the deviation from linearity and provides a more sensitive indicator of linearity than the correlation coefficient often quoted with linear regression.

Evaluation of calibration routines in pharmaceutical and bioanalytical separations, with reference to Monte Carlo simulations.

A Monte Carlo method for the estimation of the bias and inter-assay component of precision attributable to given combinations of assay and calibration routine is described. Simulations are performed by a computer program which uses the actual concentration versus response and concentration versus intra-assay precision characteristics of the assay under investigation as its database. In operation, a calibration routine is selected, the calibrators defined and the analytical process of calibration followed by the analysis of unknowns is reproduced. In this way the mean bias and interassay component of precision at a number of pre-defined concentrations covering the analytical range of the assay is estimated. The program is intended for use by analysts as a practical aid in the selection and optimization of appropriate calibration routines in an experimental environment.

Determination of roxatidine in human plasma, urine and milk by capillary gas chromatography using nitrogen-selective detection.

An assay is described for the determination of roxatidine in human plasma, urine and milk by gas chromatography. Roxatidine is extracted from the basified matrix with dichloromethane and esterified with propionic anhydride prior to analysis of the extracts by capillary gas chromatography using a nitrogen-specific detector. Detection limits are 5 ng/ml for plasma and milk and 1 microgram/ml for urine, making the assay suitable for obtaining pharmacokinetic data from volunteer trials.

Determination of oxpentifylline and three metabolites in plasma by automated capillary gas chromatography using nitrogen-selective detection.

A procedure is described for the simultaneous determination of oxpentifylline and three metabolites in plasma by capillary gas chromatography. Plasma samples are acidified and extracted with chloroform. The carboxylic acid compounds are converted to their methyl esters and the hydroxylated metabolites to their O-trifluoroacetates prior to analysis by capillary gas chromatography using nitrogen-selective detection. The detection limits for the compounds are 5 ng/ml and the assay has been applied to the analysis of plasma from volunteer trials after oral administration of oxpentifylline.

Determination of oxpentifylline and four metabolites in plasma and urine by automated capillary gas chromatography using nitrogen-selective detection.

A capillary gas chromatographic assay for the determination of oxpentifylline and four hydroxylated metabolites is described. Oxpentifylline, metabolites and internal standards are extracted from basified plasma or urine with chloroform and the metabolites are converted to their O-trifluoroacetates before analysis by capillary gas chromatography using nitrogen-selective detection. A high throughput of samples is achieved by use of an autosampler and a split injection technique which allows the samples to be analysed isothermally. The detection limits of the compounds are in the range 2-10 ng/ml for plasma and 0.1-0.2 microgram/ml for urine. The assay has been applied to the analysis of plasma and urine samples after the oral administration of oxpentifylline.

The pharmacokinetics of oxpentifylline in man when administered by constant intravenous infusion.

Subjects were each given either a 25, 50 or 100 mg intravenous loading dose of oxpentifylline followed by an intravenous infusion at a constant rate of 1.5 mg/min for 3 h. Plasma levels of oxpentifylline were measured to obtain information on its pharmacokinetics and to establish which of the loading doses gave the most rapid attainment of the steady state plasma levels of intact drug. Oxpentifylline kinetics were best described by a two compartment model giving a characteristic dip in the plasma level versus time curves before steady state was reached when either the 50 or 100 mg loading doses, followed by the constant intravenous infusion, were given. The terminal half-life of oxpentifylline was 1.02 +/- 0.86 h, reflecting a very high clearance of the drug (approx. 3 000 to 6 000 ml/min). The high clearance could be attributed to extrahepatic metabolism occurring in blood which was observed in vitro using whole blood but not plasma. The clearance of a reduced metabolite of oxpentifylline was less than that of the intact drug, although the half-life was similar (0.83 +/- 0.18 h). Of the three loading doses tested, only the highest showed any side effects, these being transient and occurring within a 5 to 10 min period after dosing and appeared to correlate with the high initial plasma levels of the drug. The 25 mg loading dose gave initial plasma levels generally below the final steady state levels, whilst the 50 mg loading was the closest to giving immediate steady state plasma levels of oxpentifylline.

Determination of oxpentifylline and a metabolite, 1-(5'-hydroxyhexyl)-3,7-dimethylxanthine, by gas--liquid chromatography using a nitrogen-selective detector.

A gas chromatographic method for the determination of oxpentifylline and a metabolite, 1-(5'-hydroxyhexyl)-3,7-dimethylxanthine is described. Oxpentifylline, metabolite and internal standard are extracted from basified plasma into dichloromethane, then the metabolite and internal standard are converted to their O-trifluoroacetates. Analysis by gas--liquid chromatography using a nitrogen-selective detector allows quantification of oxypentifylline and 1-(5'-hydroxyhexyl)-3,7-dimethylxanthine down to levels of 3 ng/ml and 3--10 ng/ml, respectively. The assay had been applied to plasma samples from volunteers after both intravenous and oral administration of oxpentifylline. The need to separate plasma from erythrocytes immediately after venipuncture sampling to prevent further metabolism of oxpentifylline is emphasized.

Quantitative analysis of tiamenidine in human plasma by gas chromatography mass spectrometry of a dibenzyl derivative.

A gas chromatography mass spectrometry method has been developed and evaluated for the quantitative analysis of tiamenidine in plasma. Tiamenidine and internal standard are extracted from basified plasma, converted to dibenzyl derivatives by reaction with benzyl bromide and potassium t-butoxide in the presence of 18-crown-6 ether prior to analysis by selected ion monitoring. The method can be used over the range 0.2--10 ng ml-1 with a coefficient of variation of better than 20% at 1 ng ml-1.

Quantitative analysis of 6,11-dihydro-11-oxo-dibenz[b,e] oxepin-2-acetic acid (isoxepac) in plasma and urine by gas--liquid chromatography.

A method is described for the quantitative analysis of 6,11-dihydro-11-oxo-dibenz[b,e]-oxepin-2-acetic acid (isoxepac) in plasma and urine. Isoxepac and internal standard was analysed by gas-liquid chromatography using a flame ionization detector. The method is accurate and precise over the range 0.1--30 microgram/ml. The method has been applied to the analysis of plasma and urine from both healthy volunteers and patients receiving therapeutic oral doses of isoxepac.

Gas-liquid chromatographic determination of 1,3-dihydro-3-phenylspiro(isobenzofuran-1,4-piperidine), HP 505, in biological fluids using a nitrogen-specific detector.

A gas chromatographic method for the determination of 1,3-dihydro-3-phenylspiro[isobenzo-1,4-piperidine], HP 505, in plasma red blood cells and urine has been developed. HP 505 and internal standard are extracted from basified fluid with hexane and then back extracted into acetic acid. After re-extraction into hexane, HP 505 and internal standard are analysed by gas-liquid chromatography as the N-propionyl derivatives using a nitrogen-specific detector. Concentrations of HP 505 can be measured over the range 2-100 ng/ml plasma. The method has been applied to the analysis of biological fluids from volunteers receiving oral doses of HP 505.