M. Franklin

Simultaneous measurement of venlafaxine and its major metabolite, oxydesmethylvenlafaxine, in human plasma by high-performance liquid chromatography with coulometric detection and utilisation of solid-phase extraction

Venlafaxine, oxydesmethylvenlafaxine and an internal standard (paroxetine) were extracted from plasma by a solid-phase extraction technique. Chromatography was performed using isocratic reversed-phase high-performance liquid chromatography (HPLC) with coulometric endpoint detection. The standard curves were linear over the range 0-200 ng/ml for both venlafaxine and oxydesmethylvenlafaxine in plasma. The mean inter- and intra-assay coefficients of variation over the range of the standard curves were less than 10%. The absolute recovery averaged 74% for venlafaxine and 67% for oxydesmethylvenlafaxine. The sensitivity was 0.5 ng for both the analytes. Plasma profiles of the analytes following oral administration of venlafaxine, are presented.

Simultaneous determination of catecholamines in rat brain tissue by high-performance liquid chromatography.

A novel and highly sensitive method has been developed for the determination of catecholamines [noradrenaline (NA), dopamine (DA), serotonin (5-HT) and their metabolites 5-hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA)] in brain tissue. The method uses isocratic reversed-phase HPLC with amperometric end-point detection. The calibration curve was linear over the range 10-150 pg on-column. The assay limits of detection for NA, DA, 5-HT, 5-HIAA and HVA were 3.8, 3.8, 6.8, 5 and 7.5 pg on-column, respectively. The mean inter- and intra-assay relative standard deviations (RSDs) over the range of the standard curve were less than 5%. The absolute recoveries averaged 99.1%, 99.5%, 97.7%, 99.5% and 98.8% for NA, DA, 5-HT, 5-HIAA and HVA, respectively.

The neuroendocrine effects of oral imipramine

The release of growth hormone, prolactin and cortisol following oral imipramine was studied in nine fit young men. Imipramine 100 mg, but not 40 mg, led to reliable rises in the circulating levels of all three hormones in the majority of subjects. These responses are likely due to the enhancement of central noradrenergic and serotonergic function as a result of reuptake inhibition. The safety, sensitivity and reliability of these responses make imipramine 100 mg orally a potentially valuable neuroendocrine challenge test.

Effect of a long-term low tryptophan diet on the prolactin responses to the 5-HT1A and 5-HT2C agonists, 8-OH-DPAT and mCPP in the male rat.

The study was undertaken to assess the long term effects of tryptophan (TRP) depletion through diet on the prolactin (PRL) responses to the serotonin (5-hydroxytryptophan, 5-HT) agonists m-chlorophenyl-piperazine (mCPP) and 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) in the male rat. The low TRP diet caused significant reductions in both plasma total TRP and brain cortical 5-HT content together with a significant increase in the PRL responses to mCPP. In contrast the PRL responses to 8-OH-DPAT in animals on the low TRP diet for 1week and 6 weeks were similar to control rats. However, a small but significant increase in PRL was observed at 2 min after dosing in the 1-week group. At the same time the 3H-8-OH-DPAT binding parameters, Kd and Bmax, were similar in both control and TRP depleted animals. The results conform that long-term TRP depletion causes a deficiency of brain TRP and a subsequent reduction of brain 5-HT.This is associated with an enhanced PRL response to mCPP probably resulting from a functional up-regulation of post-synaptic 5-HT2Creceptors. The small or transient changes brought about by long-term TRP depletion on post-synaptic 5-HT1Areceptors, suggests that these receptors may be less susceptible to 5-HT depleting effects than the 5-HT2Csubtype.

Determination of sumatriptan succinate in human plasma by high-performance liquid chromatography with coulometric detection and utilization of solid-phase extraction

Sumatriptan succinate (the analyte) and naloxone (the internal standard) were extracted from plasma with a solid-phase extraction technique. Chromatography and detection were performed by isocratic reversed-phase high-performance liquid chromatography with coulometric end-point detection. The standard curve was linear over the range 0-100 ng/ml of sumatriptan succinate in plasma. The reproducibility (as defined by the coefficient of variation, C.V.) over the range of the standard curve was 4.9-7.3%. The recovery averaged 83%. The sensitivity was 0.25 ng of sumatriptan on column (allowing a concentration of 0.5 ng/ml to be determined from a 1-ml plasma sample volume). Plasma profiles of the analyte following subcutaneous (s.c.) administration in eight normal male volunteers, are presented.

Sub-chronic treatment with an extract of Hypericum perforatum (St John's wort) significantly reduces cortisol and corticosterone in the rat brain

Extracts of Hypericum perforatum (HP) have been shown to be effective for the treatment of mild to moderate depression. Its mode of action has not been fully elucidated. An increase in plasma and cerebrospinal fluid (CSF) cortisol is frequently observed in depression. Studies have suggested that HP might alter brain cortisol and corticosterone through its effect on multidrug transporter glycoprotein (Pgp). We investigated the effect of sub-chronic treatment with an extract of HP (LI 160) on brain levels of corticosterone and cortisol in the rat. Results show that HP significantly reduced corticosterone and cortisol in brain frontal cortex tissue. These changes are not reflected in serum. These findings may be important with respect to HPs mode of antidepressant action.

Simultaneous quantitation of buspirone and its major metabolite 1-(2-pyrimidinyl)piperazine in human plasma by high-performance liquid chromatography with coulometric detection

Buspirone is a member of the azapirone group of anxiolytic drugs and has one major metabolite, 1-(2-pyrimidinyl)piperazine (1-PP). The analyte, its metabolite and the internal standard were extracted from plasma utilizing solid-phase extraction columns. Chromatography was performed using isocratic reversed-phase high-performance liquid chromatography with coulometric end-point detection. The calibration graph was linear over the range 0-50 ng ml-1 of plasma. The lower limits of quantitation for buspirone and 1-PP were 0.5 and 2 ng ml-1, respectively, when 1 ml of plasma was extracted. The intra-assay relative standard deviations (RSD) over the range of the calibration graph varied from 4 to 12.5% for buspirone and 1-PP. The inter-assay RSD was 6.9% for 1-PP and 9.6% for buspirone. The recovery averaged 96% for buspirone and 66% for 1-PP. Plasma profiles of buspirone and 1-PP following oral dosing are presented.

Plasma fluphenazine and prolactin levels in schizophrenic patients during treatment with low and high doses of fluphenazine enanthate

The relationship between plasma prolactin (PRL) and drug levels in patients receiving neuroleptic drugs is of special interest in view of evidence that the PRL elevation induced by these durgs reaches its maximum at sub-therapeutic doses.Plasma PRL and fluphenazine (FPZ) levels were measured by radioimmunoassay (RIA) in each of 11 chronic schizophrenics (nine men, two women) during two 4-week periods of treatment with FPZ enanthate at a ‘High Dose’ (250 mg per week) and a ‘Low Dose’ (12.5 mg per week) given in random order.Plasma PRL levels were above normal in 9 of 11 subjects during the last week of Low Dosage. High Dosage resulted in PRL levels significantly greater than found during Low Dose treatment in 9 of 11 patients. Thus the PRL response had not reached its “ceiling” during Low Dosage in most patients.A significant correlation between PRL and FPZ levels was found in seven subjects; evidence that immunoreactive FPZ levels relate to an effect caused by blockade of dopamine receptors. The plasma FPZ pattern between injections during week 1 of Low Dosage was remarkably stable; High Doses produced an initial drug peak at 1–2 h and a secondary peak occurring on days 2–3 followed by a return to preinjection levels by day 7.

The effects of a low tryptophan diet on brain 5 -HT metabolism and 5-HT-mediated neuroendocrine responses in the male rat

The study was undertaken to assess the effects of periods of dietary tryptophan (TRP) depletion on (i) plasma total and free TRP together with brain TRP, 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5- HIAA) and (ii) the prolactin (PRL) responses to the 5-HT releasing agent, D-fenfluramine (FEN), and the 5-HT agonist, m-chlorophenylpiperazine (mCPP). The TRP-deficient diet caused significant reductions in plasma total and free TRP for a short-lived period; however, it caused longer lasting and significant reductions in brain TRP, 5-HT and 5-HIAA content. Following periods of dietary TRP depletion, plasma PRL responses to FEN were significantly reduced for 14 days but had normalised by day 21. In contrast, the PRL responses to mCPP were significantly enhanced after 6 and 21 days, the response at 14 days being similar to control. The result suggests that periods of dietary invoked TRP depletion cause a deficiency of brain TRP content which subsequently leads to a fall in brain 5-HT. This is associated with a reduced PRL response to FEN and an enhanced PRL response to mCPP, the latter possibly resulting from functional up-regulation of post-synaptic 5-HT receptors.

Determination of hypericin in plasma by high-performance liquid chromatography

Hypericin, a polycyclic dianthroquinone, is one of the characteristic ingredients of Hypericum perforatum extracts (St. Johns wort, HP), which has antidepressant effects. Hypericin and the internal standard (I.S.), dansylamide, were extracted from plasma utilizing solid-phase extraction (SPE). Chromatography was performed using isocratic reversed-phase high-performance liquid chromatography (HPLC) with fluorescence end-point detection. The calibration curve was linear over the range 5-100 ng per ml of plasma. The sensitivity for hypericin was 75 pg on column. Mean inter- and intra-assay coefficients of variation (C.V.s) over the range of the standard curve were less than 10%. The absolute recovery for hypericin averaged 72.6%.