The determination in human plasma of 1-hydroxy-2-naphthoic acid following administration of salmeterol xinafoate.

The clinical development of salmeterol xinafoate, the 1-hydroxy-2-naphthoic acid salt of salmeterol, a potent long acting beta 2 agonist bronchodilator, has required the development of a method for the determination of 1-hydroxy-2-naphthoic acid (HNA), in human plasma. A sensitive, accurate and precise method was, therefore, required to enable the pharmacokinetic profile to be established. HNA was determined in human plasma using a semi-automated procedure with solid-phase extraction using an automated analytical sample processor (AASP) and high-performance liquid chromatography (HPLC) with fluorescence detection. The method was sensitive to 10 ng ml-1. The method is specific for HNA with respect to endogenous plasma components and has been shown to be robust, accurate and precise. Over four independent assay runs, the relative standard deviations (RSD) of the quality control samples (QC) were 1.6, 2.4 and 5.5% at 180, 100 and 40 ng ml-1, respectively. A pharmacokinetic profile of HNA in man has been established from a single dose kinetic study in healthy volunteers following an oral dose of 500 micrograms salmeterol xinafoate, equivalent to 225 micrograms HNA. Maximum plasma concentrations attained at 1 h after dosing ranged between 35.3 and 66.8 ng ml-1 and were within the calibration range of the assay.

Dose selection for toxicity studies: a protocol for determining the maximum repeatable dose.

1. A three-stage protocol is described for a dose-ranging study which defines the maximum repeatable dose (MRD) and provides a preview of the toxicology of new, pharmacologically active, substances before commencing the first formal regulatory toxicity studies, usually of 2 or 4 weeks duration. 2. Additionally, a range of toxicokinetic (TK) data relevant to protocol design for formal studies is generated. 3. Stage A is a dose incrementation process in which the MRD is provisionally determined and basic TK values generated. 4. In stage B the animals are dosed daily for at least 7 d, the MRD is substantiated and a wider range of TK data obtained. 5. In stage C, each of the dose levels identified for a formal study is administered once to investigate the relationship of doses to TK data. 6. This protocol can be completed using as few as 24 rats or six dogs (or primates). 7. Selection of dose levels for the first formal studies can be greatly aided by the results of a well-designed dose-ranging study including TK data. 8. For particularly toxic substances, the findings of studies based on this protocol have frequently been sufficiently clear to warrant early termination of their development.

Combined use of an automated sample processor and a polymer-based high-performance liquid chromatographic column to determine the pharmacokinetics of labetalol in man.

An improved high-performance liquid chromatographic (HPLC) assay has been developed for the analysis of labetalol in human plasma. The method is based on the combined use of an automated sample processor, reversed-phase analysis on a microparticulate polymer-based HPLC column and fluorescence detection. The pH stability of the polymeric column packing material allowed the use of a mobile phase adjusted to pH 9.5, which was optimal for the fluorescence of labetalol. Assay validation was undertaken over the labetalol concentration range 2-100 ng/ml. Calibration curves were essentially linear, and the mean coefficient of variation was 5.3%. The assay has been used for the analysis of clinical samples in support of pharmacokinetic studies.

Determination of ranitidine and its metabolites in human urine by reversed-phase ion-pair high-performance liquid chromatography.

A method using ion-pair high-performance liquid chromatography is presented for determining ranitidine, ranitidine N-oxide, ranitidine S-oxide and desmethyl ranitidine in the urine from four volunteers, given on separate occasions an intravenous and oral dose of 100 mg ranitidine. This method has been used to study the metabolism and pharmacokinetics of ranitidine by man. It was found that the elimination half-life of ranitidine ranged from 110-246 min. The mean renal clearance of ranitidine in these four volunteers was 512 ml/min.

Viral depuration by assaying individual shellfish.

A study was carried out to further evaluate the practicability of viral depuration by assaying individual shellfish. The Northern quahaug and a strain of the type 1 attenuated poliovirus were used as the working model. Two types of depuration systems were employed: the small experimental tanks and a pilot-size tank with a capacity of approximately 24 bushels (836 liters) of shellfish. Volumes of the individual shellfish samples were found uniform throughout the experiments when a prior selection for the weight of the shellfish was made. There was also no significant difference in volumes of the individual samples during the course of depuration (24 to 96 hr). Under controlled hydrographic conditions, however, the uptake of virus in individual shellfish varied considerably. In general, the individual variability reached 10- to 100-fold. This wide variation would explain the variability of viral contents obtained in pooled samples during depuration as reported previously. During a later phase of depuration, although a great majority of shellfish were free of the virus, a few still harbored minimal amounts of contaminants. The presence of virus in some of the shellfish after various periods of depuration would, theoretically, be obscured by the pooling of the sampled shellfish. Further examination of the negative samples by assaying larger quantities than those routinely used revealed that a few still contained virus. To simulate naturally polluted shellfish as closely as technically possible, shellfish were polluted with minimal amounts of virus. The shellfish were cleansed more rapidly by the depuration process than were those polluted with more virus. Since the naturally polluted shellfish were shown to contain less virus than those studied in the laboratory, it is anticipated that the former type of shellfish may be cleansed more readily by this process within a reasonable period of time. Justification for a field trial of depuration in this country is presented.

Determination of salbutamol in human plasma and urine by high-performance thin-layer chromatography.

A rapid, accurate and sensitive method for the determination of salbutamol in plasma and urine is described. Salbutamol is extracted using solid-phase techniques and converted to an indoaniline dye by reaction with dimethyl-p-phenylenediamine. The indoaniline is separated using high-performance thin-layer chromatography and quantified by absorption microdensitometry at 650 nm. The method is sensitive down to 20 ng/ml in urine and to 1 ng/ml in plasma and provides data in good agreement with that obtained by gas chromatography--mass spectrometry. The method can be used for analysis of pharmacokinetic studies.