John H. Vine

Plasma level of chlormethiazole and two metabolites after oral administration to young and aged human subjects

SummaryThe plasma concentration of chlormethiazole and two of its metabolites has been measured in three young and three aged human subjects following administration of a single oral dose of chlormethiazole. A sensitive analytical method based on gas chromatography-mass spectrometry using the selective ion monitoring mode of operation was developed to permit quantitation of the plasma levels. The time course of the plasma concentration of chlormethiazole and metabolites showed wide inter-subject variation, particularly between the young and elderly subjects. Absorption of chlormethiazole was rapid in the subjects of both groups as assessed by the time taken to reach the peak plasma concentration. The mean peak plasma level of chlormethiazole was more than five times greater in the elderly (2.90±1.56 µg/ml) than in the young (0.55±0.58 µg/ml) subjects. The plasma level of chlormethiazole was consistently higher in the aged subjects and this was reflected by the larger area under the plasma curve in aged (7.62±5.37 µg.h/ml) than in young (0.94±0.66 µg.h/ml) individuals. Decreased pre-systemic elimination by the liver has been suggested as an important factor contributing to the higher plasma level in the elderly. Estimates of absolute systemic availability, calculated by reference to previous intravenous studies, were greater for the elderly subjects. The distribution of chlormethiazole in whole blood from six young and six elderly human subjects was investigated in vitro. The unbound fraction of chlormethiazole in plasma increased significantly from 0.308±0.035 in young subjects to 0.403±0.067 in the elderly. Distribution of the drug in whole blood was different for the two age groups; the fraction of drug distributed to plasma water was significantly greater and the fraction in blood cells was significantly less in the aged.

The disposition of biphenylacetic acid following topical application

SummaryPlasma and synovial fluid concentrations of biphenylacetic acid were determined following application of 3 g of 3% biphenylacetic acid gel to one knee of patients suffering from rheumatoid arthritis.The mean peak plasma concentration was 34 ng/ml. Synovial fluid concentrations tended to follow plasma concentrations but at a somewhat lower level, the mean peak synovial fluid concentration was 21 ng/ml. The average ratio of synovial fluid AUC (0–24 h) to plasma AUC (0–24 h) was 0.58, r=0.97.Where patients had bilateral effusions, the concentration in the ipsilateral knee at each time point examined was not significantly different to that in the contralateral knee, suggesting that absorption was initially into the plasma and subsequently into the synovium.

Effects of serial subculture in vitro on the endogenous levels of indole-3-acetic acid and abscisic acid and rootability in microcuttings of ‘Jonathan’ apple

Proliferating axillary shoots of the difficult-to-root apple cultivar ‘Jonathan’ acquired an enhanced ability to form adventitious roots with increasing number of subcultures in vitro. The transition between the difficult-to-root and the easy-to-root condition occurred at the fourth subculture.Endogenous levels of free IAA and ABA in shoot tissues were analysed by gas chromatography/mass spectrometry/single ion monitoring (GC/MS/SIM) using negative ion chemical ionisation. Tissues from the mother plants grown in the glasshouse contained more IAA and ABA than those from tissue-culture material. After establishment in vitro there was no variation in the IAA content throughout the subcultures but a decrease in ABA content was observed after the fourth transfer. The IAA/ABA ratio increased from 0.2 in difficult-to-root shoots from the initial culture up to 0.7 in easy-to-root shoots from the long-term subculture.

Seasonal changes in endogenous ABA and IAA and the influence of applied ABA and auxin in relation to shoot growth and abscission in Valencia Orange (Citrus sinensis (L.) Osbeck)

Seasonal changes in endogenous IAA and ABA were measured by gas chromatography/mass-spectrometry. Highest concentrations of ABA occurred in leaves. There was a major ABA peak in early spring (up to 1360 ng g−1 dw). Levels were low in summer (90 ng g−1 dw). There was a minor ABA peak in autumn. Endogenous IAA in leaves was highest in winter/spring (up to 76 ng g−1 dw). Applied ABA promoted abscission of leaves and shoots while applied NAA delayed abscission. The main peak in leaf-ABA content was followed by extensive shoot abscission. The involvement of ABA and IAA in regulation of flush growth was not clear.

Endogenous indole-3-acetic acid and abscisic acid in apple microcuttings in relation to adventitious root formation

The effects of applying indole-3-butyric acid (IBA) for periods up to 48 h were examined in difficult-to-root microcuttings (from newly-established cultures) and in easy-to-root microcuttings (from long-term subcultures) of ‘Jonathan’ apple (Malus X domestica Borkh). In easy-to-root material, 20% of the microcuttings produced roots in the absence of IBA, while 6 h exposure to 10 μM IBA gave 100% rooting of microcuttings. In contrast, root formation in difficult-to-root material was IBA-dependent. Maximum rooting of these microcuttings (50%) required 24 h exposure to 10 μM IBA.Variation in the endogenous levels of free indole-3-acetic acid (IAA) during the course of root induction was similar in microcuttings of both types but there were marked differences in endogenous abscisic acid (ABA) levels. In easy-to-root microcuttings ABA remained at a constant low level, but in difficult-to-root material ABA exhibited marked fluctuations and was present at higher concentrations than in easy-to-root microcuttings.

Quantitation of doxapram in blood, plasma and urine

Methods for the quantitation of doxapram in blood, plasma and urine have been developed. Following extraction, gas-liquid chromatography was used to separate doxapram from basic metabolites. Doxapram was detected by mass spectrometry for blood and plasma assays, and by flame ionisation for urine assays. The limit of reliable quantitation in blood and plasma was 10 ng and in urine 500 ng, the coefficients of variation being 6.37%, 1.72% and 2.31% respectively. To illustrate the clinical applicability of the assay methods, plasma, blood and urine levels were monitored in a premature newborn following an intravenous infusion of doxapram.

Metabolism and Urinary Excretion of Benzhexol in Humans

1. Benzhexol and three of its metabolites excreted in urine in man have been investigated by g.l.c.--mass spectrometry. 2. Three isomeric hydroxylated metabolites were identified as the 1-(hydroxycyclohexyl)-1-phenyl-3-piperidinopropan-1-ols. 3. The amounts of benzhexol and its identified metabolites have been semiquantitatively determined after a single oral dose in two healthy adults. Approx. 56% of the dose was excreted as the hydroxylated metabolites. The levels of benzhexol excreted were too low to be measured by the techniques used.

The Identification of Eight Hydroxylated Metabolites of Etidocaine by Chemical Ionization Mass Spectrometry

1. Eight hydroxylated metabolites of etidocaine have been identified in urine of man by g.l.c.-chemical ionization mass spectrometry, using methane and a mixture of methane and deuterium oxide as reactant gases. 2. The metabolites identified were N-(2,6-dimethyl-3- and 4-hydroxyphenyl-)-2-(N,N-ethylpropylamino)butyramides, N-(2,6-dimethyl-3- and 4-hydroxyphenyl)-2-aminobutyramides, N-(2,6-dimethyl-3- and 4-hydroxyphenyl)-2-(N-ethylamino)butyramides, and N-(2,6-dimethyl-3- and 4-hydroxyphenyl)-2-(N-propylamino)butyramides. 3. The eight metabolites represent about 10% of the oral dose of etidocaine.