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Streptozotocin-treated rats were used as models of type 1 diabetes to study the effects of dietary taurine on insulin- and adrenergic-stimulated 2-deoxyglucose uptake by isolated adipocytes. In addition to the well-established impairment of basal and insulin-stimulated 2-deoxyglucose uptakes in adipocytes prepared from streptozotocin-diabetic rats, the alpha-(phenylephrine) and beta-(isoproterenol) adrenergic stimulations of glucose uptake were also abolished. The insulin stimulation of glucose uptake in adipocytes was selectively abolished by the phosphatidylinositol 3-kinase inhibitor wortmannin, whereas that by the adrenergic agonists, phenylephrine and isoproterenol, was inhibited by prazosin and propranolol, respectively. Dietary taurine, 4 weeks before and 4 weeks after streptozotocin administration, prevented the loss of both insulin and adrenergic agonist stimulations of 2-deoxyglucose uptake, without affecting hyperglycaemia. Because insulin and adrenergic activations of glucose transport by adipocytes are coupled to different signalling pathways, it is unlikely that these effects of taurine are related to these disparate postreceptor mechanisms.

The loss of dopaminergic neurons of the substantia nigra in Parkinson's disease and in animal models of Parkinson's disease is associated with an imbalance in the activity of the so-called 'direct' and 'indirect' pathways of information flow through the basal ganglia. The aim of the present study was to determine whether the imbalance is reflected in changes in the release of GABA, aspartate and glutamate in the pathways using dual probe microdialysis in freely moving rats. Control and 6-hydroxydopamine-(6-OHDA)-lesioned rats were implanted with microdialysis probes in the neostriatum and substantia nigra or globus pallidus and the release of amino acids was analysed in the dialysates. Basal levels of amino acids were largely unaltered by the 6-OHDA lesion; however, the levels of GABA in the globus pallidus dialysates were significantly elevated in the lesioned rats, indicating an imbalance in favour of the indirect pathway. Administration of kainic acid to the neostriatum enhanced the release of GABA locally and in the distal probes in the substantia nigra and globus pallidus. In 6-OHDA-lesioned rats, stimulated release of GABA in the substantia nigra was abolished, indicating a reduction in transmission along the direct pathway. Thus, consistent with the direct-indirect pathway model of the basal ganglia, the 6-OHDA lesion results in an elevation of the basal release of GABA in the striatopallidal (indirect) pathway and a reduction in the evoked release of GABA in the striatonigral (direct) pathway. These imbalances may underlie, at least in part, the motor abnormalities of Parkinson's disease and in animal models of Parkinson's disease.

Hydatidosis is related to parasitical etiology, linked to Echinococcus granulosis and Echinococcus multilocularis. The authors describe an uncommon clinical case of old hydatidosis in an 80-year old woman, presenting hepatic skip , with localization to the right lung and kidney and burrowing to the posterior abdominal wall. Because of the patient's lack of compliance, percutaneous treatment with alcohol of the lumbar lesion with 95% ethanol is carried out under echographic guide, to improve quoad valetudinem prognosis. The authors indicate that the method target are: 1) multiple relapse lesions; 2) complicated anatomie area; 3) multiltle cyst; 4) marginal lesions. Moreover, this method shows important advantages such as be possibility of repetition, low invasivity, high patient's compliance, low rate of shock and dissemination and finally low cost.

Amino acids have received increased attention with regard to their thermoregulatory effects and possible role as neurotransmitters within the thermoregulatory system. The purpose of the present work was to evaluate in conscious rabbits the changes in cerebrospinal fluid (CSF) concentration of taurine, GABA, aspartate, and glutamate during exposure to high ambient temperature (50 min, 40 degrees C) to investigate their involvement in heat stress (HS). CSF and plasma osmolality and CSF concentrations of some cations and proteins were also determined. HS animals underwent transient hyperthermia and thereafter fully recovered. This was accompanied by a significant rise in CSF and plasma osmolality, CSF protein, calcium, taurine, and GABA. Artificial CSF osmolality measurements after addition of CaCl(2) or taurine demonstrated that the increased CSF osmolality after HS is accounted for, only in part, by the increased concentrations of either calcium and taurine. It is suggested that, during HS, taurine and GABA are released in the extracellular space of brain tissues in higher amounts, possibly to counteract the resulting hyperthermia.

We investigated whether heat-stress induced hyperthermia could enhance release of both endogenous taurine and GABA from nerve cells into the extracellular compartment, thus acting like endogenous cryogens. Conscious rabbits were exposed for 1 hr to 40 degrees C (heat stress) while cerebrospinal fluid (CSF) and plasma osmolality and the CSF concentrations of some cations, proteins as well as those of taurine and GABA were determined. Heat stress-induced hyperthermia was accompanied by a significant rise in CSF and plasma osmolality, CSF calcium, taurine and GABA levels. It is suggested that during heat stress taurine and GABA are released in the extracellular space of brain tissues in higher amounts, as compared to control conditions, to counteract the resulting hyperthermia, thus acting as cryogenic agents.

Intracerebral MD enables the retrieval of endogenous substances from the extracellular fluid (ECF) of the brain and has been demonstrated to be a sensitive technique for early detection of subtle vasospasm-induced neurometabolic abnormalities in patients with subarachnoid hemorrhage (SAH). The aim of this study was to monitor cortical extracellular concentrations of energy metabolism markers, such as glucose and lactate, neurotransmitter amino acids, such as glutamate, aspartate, GABA and taurine to identify any neurochemical patterns of cerebral ischemia. A prospective clinical study was conducted on a group of 16 patients with non-severe SAH operated on within 72 hours after initial bleeding. Following aneurysm clipping, an MD catheter was inserted in the cortical region where vasospasm could be expected to develop, and perfused with artificial CSF at 0.3 microl/min flow rate. Dialysate was collected every 6 hours and then analyzed on High Performance Liquid Cromatography (HPLC) for glucose, lactate, pyruvate, glutamate, aspartate, GABA and taurine. Mean ECF taurine concentrations ranged from 1.4 + 0.7 to 12.3 + 7.8 micromol/l in single patients: global mean value was 5.8 + 3.8 micromol/l. In this series, the highest absolute taurine value was 25.7 micromol/l, observed in a patient who developed clinical and radiological signs of cerebral ischemia. Nine patients presented clinical disturbances related to cerebral vasospasm. In this setting, representing a mild-to-moderate hypoxic condition, MD data demonstrated that lactate is the most sensitive marker of cellular energy imbalance. Increased lactate levels positively correlated with glutamate (P<0.0001), aspartate (P<0.0001), GABA (P<0.0001) and taurine (P<0.0001) concentrations. These results suggest that also in humans increased taurine levels reflect a condition of cellular stress. This study confirms that MD is a sensitive technique to reveal subtle metabolic abnormalities possibly resulting in cell damage.

Perturbed cellular calcium homeostasis has been implicated in both apoptosis and necrosis, but the role of altered mitochondrial calcium handling in the cell death process is unclear. Recently we found that taurine, a naturally occurring amino acid potentiates Ca2+ sequestration by rat liver mitochondria. These data, which accounted for the taurine antagonism on Ca2+ release induced by the neurotoxins 1-methyl-4-phenylpyridinium plus 6-hydroxy dopamine previously reported, prompted us to investigate the effects of taurine on the permeability transition (PT) induced experimentally by high Ca2+ plus phosphate concentrations. The parameters used to measure the PT were, mitochondrial swelling, cytochrome c release and membrane potential changes. The results showed that, whereas taurine failed to reverse changes of these parameters, cyclosporin A completely reversed them. Even though these results exclude a role in PT regulation under such gross insult conditions, they cannot exclude an important role for taurine in controlling pore-opening under milder more physiological PT-inducing conditions.

The aldehyde produced from the oxidative deamination of primary and secondary amines by the monoamine oxidases (EC 1.4.3.4; MAO) or the semicarbazide-sensitive amine oxidases (EC 1.4.3.6; SSAO) may be determined followed reaction at elevated temperatures with 2-diphenylacetyl-1,3-indandione-1-hydrazone (DIH), separation by high-performance chromatography liquid (C-8 column, eluting isocratically with acetonitrile, ammonium acetate, water) and fluorimetric detection (excitation and emission wavelengths 430 and 525 nm). The detection limits for benzaldehyde, p-hydroxybenzaldehyde and 2-phenylacetaldehyde were 125 nM, 150 and 62.3 nM, respectively. Thus the assay is appropriate for determination of amine oxidase activities towards benzylamine, 2-phenylethylamine and tyramine. The fluorescence of the DIH adduct with indole-3-aldehyde was strongly quenched, giving a relatively high detection limit (17.5 microM). The detection limit was lower (3.8 microM) when the absorbance at 430 nm was monitored. Enzyme activities determined by this procedure were shown to be linear with enzyme-protein concentration (rat liver mitochondria). The presence of 1-2 mM semicarbazide, necessary for determining MAO activities in samples also containing SSAO, did not adversely affect the derivatization reaction. The DIH-aldehyde adducts were sufficiently stable to permit their storage at low temperatures prior to assay. The product produced by reaction of 5-hydroxyindole acetaldehyde with DIH had no significant fluorescence and too low an absorbance at 430 nm to allow its determination for assay of activities towards 5-HT. This procedure can also measure succinic semialdehyde (detection limit 240 nM) and thus would be applicable to the determination of GABA transaminase activity.

The effects of neurotensin (NT) on the release of acetylcholine (ACh), aspartate (Asp), glutamate (Glu) and gamma-aminobutyric acid (GABA) from the hippocampus of freely moving rats were studied by transversal microdialysis. ACh was detected by High Performance Liquid Chromatography (HPLC) with electrochemical detection while GABA, glutamate and aspartate were measured using HPLC with fluorometric detection. Neurotensin (0.2 and 0.5 microM) administered locally through the microdialysis probe to the hippocampus produced a long-lasting and concentration-dependent increase in the basal extracellular levels of GABA and ACh but not of glutamate and aspartate. The increase in the extracellular levels of GABA and ACh produced by 0.5 microM neurotensin in the hippocampus reached a maximum of about 310% for GABA and 250% for ACh. This stimulant effect of NT was antagonized by the NT receptor antagonist SR 48692 (100 microg/kg, i.p.). Local infusion of tetrodotoxin (1 microM) decreased the basal release of ACh, GABA, Asp, Glu and prevented the 0.2 microM NT-induced increase in GABA and ACh release. The effect of NT on the release of ACh was blocked by the GABA(A) receptor antagonist bicuculline (2-10 microM). Our findings indicate for the first time that neurotensin plays a neuromodulatory role in the regulation of GABAergic and cholinergic neuronal activity in the hippocampus of awake and freely moving rats. The potentiating effects of neurotensin on GABA and ACh release in the hippocampus are probably mediated by (i) NT receptors located on GABAergic cell bodies and (ii) through GABA(A) receptors located on cholinergic nerve terminals.

The release of glutamate, aspartate, GABA, and taurine from the striatum of young (3 months), mature (12 months), and old (22 months), freely moving male rats was investigated by using a microdialysis fiber inserted transversally in the striatum. In old rats basal extracellular glutamate and aspartate levels were decreased vs. young rats (-38 and -49%, respectively). GABA and taurine levels were unmodified by age. In the presence of the adenosine receptor antagonist 8-phenyltheophilline (8-pT) at the concentration of 50 microM, both K(+)-evoked releases of glutamate and aspartate were more than doubled in young, but not in mature and old rats. 8-pT at the concentration of 500 microM significantly decreased glutamate basal levels and K(+)-evoked aspartate release in old rats only. GABA and taurine releases were not affected by 8-pT at either dose. Our findings indicate a modified adenosine modulation on glutamate and aspartate release in aged rats, that could result from a change in the balance between A1 and A2a adenosine receptor density or an alteration of A1 and A2a receptor-effector coupling.

A single oral dose of 90 mg kg-1 chlorimipramine or chlorpromazine, corresponding to 54.5 or 55.9 mumol, respectively, was given to male Sprague-Dawley rats and concentrations of parent drugs and their N-desmethyl metabolites measured by gas chromatography in plasma and major organs (brain, liver, lung, kidney, heart, spleen and peritoneal fat). In the case of chlorimipramine, N-desmethyl metabolite levels were consistently higher than those of the parent drug for the entire observation period of 24 h in all tissues except fat, while lower N-desmethyl metabolite/parent compound ratios were observed for chlorpromazine. N-Desmethyl metabolite kinetics of chlorimipramine appeared to be elimination-rate limited, while those of chlorpromazine were formation-rate limited. In all analysed organs, the maximu detectable drug+metabolite concentrations accounted for only 2.3 and 4.6% of the initial dose of chlorimipramine and chlorpromazine. Chlorpromazine treatment gave rise to an area under the total amount-time curve (AUC0-24) for parent drug+metabolites, 3.9-fold that for chlorimipraine. Closer scrutiny discloses a conversion ratio of parent compound to N-desmethyl metabolite of 1.1 for chlorpromazine and of 2.2 for chlorimipramine, indicating the greater efficiency of chlorimipramine metabolism in all compartments. The expected high conversion index found in the liver (2.3) reaches its maximum of 5.4 in the lung. Fractional data analysis of chlorimipramine and chlorpromazine distribution patterns revealed greater organ transfer for the N-desmethyl metabolites than for the more stably-located parent compounds. The N-desmethyl metabolites of chlorimipramine apparently moved from liver to lung, kidney and spleen, whereas N-desmethylchlopromazine moved preferentially to the brain and lung tissue. This single dose study of chlorimipramine and chlorpromazine kinetics, highlights the two distinct dispositional processes at work in the rat in all likelihood, attributable to different absorption patterns, to a slower metabolism and, thus, to the longer persistence of chlorpromazine.

In order to test the hypotheses that stimulation of non-N-methyl-D-aspartate (NMDA) receptors in the neostriatum causes the release of gamma-aminobutyric acid (GABA) from nigrostriatal neurones, dual microdialysis was carried out in the neostriatum and substantia nigra of freely moving rats. Application of kainic acid to the neostriatum caused a dose-dependent release of GABA both locally and, at the same time, from the ipsilateral substantia nigra. These effects were blocked by the non-NMDA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX). Direct application of kainic acid to the substantia nigra caused a DNQX-sensitive local release of GABA. It is concluded that excitatory amino acid receptor stimulation of the neostriatum releases GABA from striatonigral neurones and that stimulation of the substantia nigra causes the release from striatonigral terminals and/or the collaterals of nigrofugal neurones.

A single oral dose of 0.7 mg kg-1 chlorimipramine (n = 18) and chlorpromazine (n = 16) was given to each subject 45 days apart and plasma concentrations of parent drugs and their monodesmethyl and didesmethyl metabolites were measured by GC. Ingestion of chlorimipramine resulted in an area under the plasma concentration-time curve (AUC0-24) for parent drug plus metabolites 5-fold higher than that observed in the same subjects following chlorpromazine intake (600 +/- 87 and 124 +/- 14 ng mL-1, respectively). Plasma chlorimipramine levels reached a mean peak value of 43.8 ng mL-1, which occurred 2 h after administration. Desmethyl metabolite kinetics of chlorimipramine appeared to be elimination rate-limited and those of chlorpromazine appeared to be formation-rate-limited. The response to single doses of these two drugs in healthy subjects highlights the two distinct dispositional processes involved, thus offering pharmacokinetic explanation of the hitherto empirical discrepancy in dosage levels in chronic treatment.

Dehalogenaton and N-demethylation of chlorpromazine (CPZ) were studied in twelve psychotic inpatients orally treated for four weeks with a daily CPZ dose of 6.4 +/- 1.1 (S.E.) mg/kg body weight (1.5-14.1, range). Weekly drug and metabolite plasma concentrations were measured by a gas liquid chromatography-nitrogen/phosphorus detector method (GLC/NPD). Plasma concentrations of the parent compound CPZ, of the dehalogenated metabolite promazine (PZ) and of the N-dealkylated metabolites N-monodemethylated chlorpromazine (CPZ-nor1) and N-didemethylated chlorpromazine (CPZ-nor2) had already reached a steady state by the end of the first week of treatment. During the entire treatment period the major plasma component was found to be PZ. Patients (N = 6) on a low dose regimen (3.7 +/- 0.2 mg/kg body weight) showed significantly lower mean plasma concentrations of CPZ and nor metabolites than patients (N = 6) on a higher dose regimen (8.6 +/- 0.7 mg/kg body weight). PZ mean plasma concentrations, however, were not significantly different in the two groups of patients, indicating that the yield of the CPZ dechlorination pathway, as opposed to that of the N-demethylation pathway, was already maximal at the CPZ concentrations attained under the low dose regimen. Male patients (N = 5) exhibited significantly higher mean PZ plasma concentrations over the 4 week period of the study than female patients (N = 7).

The autoradiographic localization of radiolabelled taurine taken up in the rat substantia nigra in vivo together with conditions of release of the [3H]taurine taken up into brain slices were studied to determine whether they are consistent with the hypothesis that taurine may act as a neurotransmitter in the striatonigral pathway. At the light microscopic level the main cellular elements that became radiolabelled following the injection of [3H]taurine into the substantia nigra could be identified as glial cells. Electron microscope autoradiography confirmed that a subpopulation of glial cells including astrocytes, pericytes, and oligodendrocytes were radiolabelled and that neuronal perikarya were not radiolabelled. In addition, axonal elements including both terminal and preterminal boutons were found to have silver grains overlying them and were thus considered to be radiolabelled. This was supported by a quantitative analysis of the distribution of the silver grains; whereas glial elements had a significantly higher number of grains associated with them than with any other structure, axonal elements had a significantly greater number of grains than dendritic structures. Release of the preloaded [3H]taurine from superfused slices of substantia nigra occurred in response to veratridine, was calcium-dependent and was sensitive to inhibition by high magnesium concentrations or tetrodotoxin. Following the destruction of neurons in the striatum by ibotenic acid injections, although the weight of the ipsilateral substantia nigra was reduced, the uptake of [3H]taurine was not altered. In contrast to this, the veratridine-stimulated release was markedly attenuated, implying that the destruction of striatal neurons causes the loss of sites in the substantia nigra from which exogenous taurine is released. These results add further support to previous suggestions that taurine might act as a neurotransmitter or neuromodulator in the striatonigral pathway.

A rat intestine M-L fraction was exposed to several agents in order to obtain the solubilization of peroxidase. These included inorganic cations, detergents, hydrolytic enzymes and membrane depolarizing drugs. Inorganic cations and cationic detergents were effective in solubilizing the enzyme activity from the particulate fraction.

The ability of a partially purified rat ileum peroxidase to synthetize PGE2 and PGF2 alpha from arachidonic acid was researched. The enzyme preparation, when incubated anaerobically with H2O2 and arachidonic acid, produced a compound extractable with ethyl acetate which showed on TLC the same Rf as authentic PGE2. This compound, after elution from the gel, was able to contract the rat strip fundus preparation.

Nanogram amounts of chlorimipramine, chlorpromazine and their Nor1- and Nor2-metabolites were detected in plasma by GLC with nitrogen-sensitive detection. Two extraction procedures were compared. The use of Sep Pak C18 cartridges produced a higher degree of accuracy and precision with significant time and materials saving, as compared to the use of organic solvents in a three steps extraction procedure.

A method is described for assaying by GLC the mono- and di-N-demethyl derivatives of chlorimipramine. This was performed by N-acylating the compounds with pentafluoropropionic anhydride and pyridine.

A new method is described for solubilizing and partially purifying rat ileum peroxidase. The enzyme exhibited a different sensitivity to the inhibitory action of protamine at the different steps of the purification procedure. The only enzyme fraction obtained after gel filtration on Sephadex G-200 was inhibited by protamine.

The distribution of 2-t-butyl-4-methoxyphenol (BHA) and its conversion to 2,2'-dihydroxy-3,3'-di-t-butyl-5,5'-dimethoxydiphenyl (di-BHA) in rat tissues at different times (1-96 hr) following the intraperitoneal administration of a single dose of BHA (32 mg kg-1 body weight) were monitored by gas chromatography-mass spectrometry (GC/MS) analysis of both compounds. High BHA levels were found in the intestine and liver persisting up to 24 hours (5.5-20.7 and 1.8-3.3 micrograms g-1 wet weight, respectively). In these tissues, values of the area under the experimental concentration curve (AUC0-24) were 285 and 49 times higher, respectively, than those observed in plasma (945 ng mL-1 hr), AUC0-24 values in kidney, spleen, erythrocytes, and brain were 2-7 times higher, whereas values below those found in plasma were observed in lung and muscle. The metabolite di-BHA could be detected in the intestine, kidney, and spleen, amounting to 5-8% of BHA. These findings indicate that rat intestine is capable of transforming in vivo BHA into di-BHA even when the former compound is administered intraperitoneally and that this capacity is shared by the kidney and spleen.

A method is described for the determination of the antioxidant 3-tert.-butyl-4-hydroxy-anisole in rat plasma using high-resolution capillary gas chromatography-mass spectrometry with selective ion monitoring. Following the addition of the isomer 2-tert.-butyl-4-hydroxy-anisole, used as an internal standard, extraction was made with n-hexane and the extract derivatized with heptafluorobutyric anhydride. The gas chromatographic separation was carried out on a SE-52 fused silica capillary column and the derivatized 3-tert.-butyl-4-hydroxyanisole and its isomer detected by recording the intensities of their common fragment ion at m/e 361. The sensitivity of the method allowed the antioxidant to be measured in 0.1-ml rat plasma samples down to a level of 10 ng/ml with a high degree of specificity and accuracy. The method has been applied to a preliminary pharmacokinetic study in rats after oral dosage.

The distribution of BHA and its conversion to di-BHA in the everted rat gut preparation was monitored by GC/MS analysis of both compounds in the mucosal and serosal side solutions and in the ileum wall. Following 1 h incubation at 27 degrees C, BHA concentration in the mucosal side solution was found to decay from an initial value of 0.1 mM down to 0.033 mM, while in the ileum it reached a 0.35 mM concentration. The amount of di-BHA found at this time in the ileum wall accounted for the transformation of 5% of the amount of BHA present in this compartment.

The toxicity of butylated hydroxyanisole (BHA) was greatly enhanced in the rat after i.p. administration, the LD50 being two orders of magnitude higher than the oral administered dose. The compound was shown either "in vitro" or "in vivo" to impair the smooth muscle contractility. Study on the mechanism of toxic action of Butylated hydroxyanisole (BHA): "in vivo" and "in vitro" effects on mammalian smooth muscle.

Rat intestine peroxidase was solubilized from a M-L fraction with BaCl2 and purified by gel filtration. The peroxidase preparation obtained by this procedure was purified 150 folds as compared to the activity present in the crude homogenate.

N-Demethylation and dehalogenation of chlorpromazine (CPZ) were compared in six psychotic inpatients and in rats orally treated for 4 weeks with a daily CPZ dose of 5.4 (mean value) and 20 mg X kg-1 body weight, respectively, by measuring drug and metabolite plasma levels by means of a gas-liquid chromatography-nitrogen/phosphorus detector method. In patients the major plasma metabolite was found to be promazine (PZ), as identified by capillary GC-MS analysis. In rats, on the contrary, PZ represented only a small proportion of the compounds detected in plasma. The mean [PZ]/[CPZ] ratio after 4 weeks of treatment was 1.64 in patients and 0.08 in rats. The relative frequency of the N-demethyl metabolites in plasma, however, was similar in the two species. The mean [N-monodemethylated CPZ]/[CPZ] and [N-didemethylated CPZ]/[CPZ] ratios after 4 weeks of treatment were 0.45 and 0.24 in patients and 0.56 and 0.25 in rats, respectively. These findings suggest that dechlorination of CPZ in psychotic patients represents an important metabolic pathway.

Some kinetic properties of a partially purified rat ileum peroxidase were investigated. The enzyme was shown to be inhibited by sodium dodecylsulfate, poly-L-lysine, deferoxamine, sodium azide and by H2O2 concentrations higher than its Km.

Fifty depressed in-patients at two psychiatric units, one in Italy the other in England, were treated with clomipramine, either orally, or intravenously and orally. A comparison of clinical response with plasma levels of clomipramine and its metabolite, desmethylclomipramine, showed clear relationships especially in the case of desmethylclomipramine. In the intravenously-treated group this was linear, in the orally-treated group it was curvilinear. Plasma levels of desmethylclomipramine and administered clomipramine correlate highly. These findings, together with the fact that significant clinical improvement was observed in only 55% of the patients, suggest that titration of the administered dose to obtain more effective plasma levels of the metabolite might improve the clinical response to the drug in some patients.

Peroxidase partially purified from rat intestine exhibited a tendency to aggregate which was inversely related to I values of the medium. This enzyme preparation showed an optimum pH 7.5-9.0 and was inhibited by excess H2O2 and Fe complexing agents.

A GC/Ms method is described for the determination of the antioxidant 2-t-butyl-4-methoxyphenol (BHA) in plasma using Sep-Pak C18 cartridges for extraction. Picogram amounts of BHA could be detected in rat plasma with a high degree of specificity, accuracy and significant time and material saving.

Subchronic treatment of rats with the antidepressant drug chlorimipramine produced an accumulation of phospholipids (PL) which was particularly evident in lung and to a lesser extent in liver and spleen. The overall increase in PL was mostly sustained by phosphatidylcholine. Both chlorimipramine and its demethylated metabolite were preferentially accumulated by lung reaching levels of about two orders of magnitude higher than those found in plasma. A significant correlation between the percentage increase in phospholipid contents and drug metabolite tissue level was evidentiated.

Data obtained in this preliminary study show that in patients chronically treated either with Chlorimipramine (n = 6) or with Chlorpromazine (n = 2), significant amounts of the corresponding Nor1- and Nor2-metabolites were detected in plasma. The role of these metabolites in the overall therapeutic effect is under investigation.

A gas chromatographic method using nitrogen-selective detection for the quantitative determination of nanogram amounts of chlorimipramine, chlorpromazine and their nor1 and nor2 derivatives in plasma is described. Derivatization with trifluoroacetic anhydride of nor1 and nor2 metabolites allowed the chromatographic separation of these compounds. A three-step solvent extraction procedure was performed using n-heptane containing 1% isoamyl alcohol and n-hexane and compared with a plasma clean-up procedure using C18 Sep-Pak cartridges. The two procedures were characterized by similar degrees of precision. The use of C18 Sep-Pak cartridges, however, produced a significant time and material saving over the conventional extraction method.

The effect of adrenalectomy on the activities of monoamine oxidase (MAO), NADH cytochrome c reductase (NCR), succinate dehydrogenase, malate dehydrogenase, fumarase, NAD+ nucleosidase and acid phosphatase in homogenates of rat hearts was examined. Besides MAO only the NCR activity increased. However, both the total and the rotenone-insensitive NCR activities increased, with that of the rotenone-insensitive being about half of the total, which indicated that the effect of adrenalectomy was exerted on components of this enzyme localized on both the inner and outer membranes of the mitochondrion. The lack of effect on the other enzymes suggests that adrenalectomy has a relatively selective action on MAO and NCR, and does not work by a generalized increase in protein synthesis or by an effect on the FAD cofactor. The MAO increase was seen with a variety of substrates, and was due to a rise in Vmax without change in Km. The response to adrenalectomy in the summer differed from that seen in the winter. The possible reasons for these effects of adrenalectomy are discussed.

BHA LD50 was determined in mice and rats following i.p. or oral administration, using DMSO or olive oil as vehicle. When DMSO was used, BHA i.p. LD50 was about two orders of magnitude lower as compared to oral LD50. This difference was not so marked when BHA was administered in olive oil.

A method has been developed for the determination of 2-t-butyl-4-methoxyphenol (BHA) and its metabolite di-BHA in rat plasma and tissues using gas chromatography-mass spectrometry with selected ion detection. Deuterium labeled BHA-d3 and di-BHA-d6 were synthesized and added to the tissue specimens as internal standards before methylene chloride extraction. The extracted compounds were derivatized with trifluoroacetic anhydride and analyzed by selected ion monitoring. Rat plasma and intestine concentrations of BHA and di-BHA at different times (0.15-24 hr) following the oral administration of a single dose of BHA (2 g X kg-1 body weight) were determined. Both BHA and di-BHA were present in all the analyzed samples, their concentration peaking within 1 hr after treatment. While in the intestine BHA levels were about 10 times higher than those of di-BHA, in the plasma they were between 100 and 15 times higher. These findings indicate that rat intestine is capable of transforming in vivo BHA into di-BHA and suggest that this organ is the major site where this transformation occurs.