RESUMEN
BACKGROUND: In a previous study a new hydrosoluble nail lacquer (P-3051) containing 8% ciclopirox (CPX) showed higher nail penetration compared to a water-insoluble 5% amorolfine (MRF) lacquer. To our knowledge, in vivo human data on a similar topic are not available. OBJECTIVES: To compare fingernail penetration of P-3051 with that of MRF reference in humans and to evaluate their predicted efficacy against Trichophyton rubrum and Candida parapsilosis. METHODS: Single centre, randomized, multiple dose, open label, within subjects study. Test and reference were self-applied to all fingernails of either hand for 28 days. At baseline and after 15 and 25 days, the nail free edge was collected for analysis. Efficiency coefficients were calculated for T. rubrum and C. parapsilosis as ratios of nail concentration/minimum inhibitory concentration. The coefficients were classified as very high, high or poor. RESULTS: Nail concentrations after 15 days were 2.82 ± 0.58 µg/mg for CPX and 0.64 ± 0.11 µg/mg for MRF. At day 25 there was a non-significant decline (1.85 ± 0.31 µg/mg, P = 0.077) for CPX and a highly significant (0.13 ± 0.03 µg/mg, P = 0.0002) 80% decline for MRF. Efficiency coefficients were very high/high in all subjects treated with P-3051 against both T. rubrum and C. parapsilosis; they were significantly lower for MRF reference against both pathogens at both observation points. CONCLUSIONS: P-3051 exhibited better penetration and higher predicted efficacy after in vivo multiple application to human fingernails when compared to MRF reference. These in vivo data are in good agreement with our previous in vitro study.
Asunto(s)
Morfolinas/uso terapéutico , Uñas/metabolismo , Onicomicosis/prevención & control , Piridonas/uso terapéutico , Adulto , Ciclopirox , Humanos , Masculino , Persona de Mediana Edad , Morfolinas/administración & dosificación , Morfolinas/farmacocinética , Piridonas/administración & dosificación , Piridonas/farmacocinética , Valores de ReferenciaRESUMEN
On the basis of a structure-activity study of a new series of anthracycline disaccharides, we recently identified a doxorubicin analogue (MEN 10755) with a promising antitumor activity. In the present study, to better support the pharmacological interest of MEN 10755, we extended the preclinical evaluation of antitumor efficacy to a large panel of 16 human tumor xenografts, which originated from different clinicopathological types. Tumors with typical multidrug-resistant phenotype were excluded because MEN 10755 was found unable to overcome resistance mediated by transport systems. In the doxorubicin-responsive series, MEN 10755 exhibited a higher activity in three of five tumors, as documented by a more marked tumor growth inhibition and an increased value of log-cell kill. In the series of doxorubicin-resistant tumors, MEN 10755 was found effective in 6 of 11 tumors (1 breast, 3 lung, and 2 prostate carcinomas). The overall response rates were 31% and 69% for doxorubicin and MEN 10755, respectively. The improvement in drug efficacy was also supported by a substantial increase in the long-term survivor rate of animals implanted with responsive tumors. Most of the tumors refractory to doxorubicin and responsive to MEN 10755 were characterized by overexpression of the antiapoptotic protein Bcl-2. In one of these tumors (MX-1 breast carcinoma), we examined the ability of MEN 10755 to induce phosphorylation of Bcl-2 after a single treatment with therapeutic doses. The results indicated that, unlike doxorubicin, MEN 10755 induced protein phosphorylation. A similar modification was produced by Taxol, which is known to be very effective against the tumor. The correlation between drug efficacy and Bcl-2 phosphorylation may underly a peculiar feature related to improvement of efficacy of the disaccharide analogue. In conclusion, the present study supports some favorable features of the novel doxorubicin analogue in terms of both efficacy and tolerability with comparison to doxorubicin, although the improvement is somewhat tumor- and schedule-dependent.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinoma/tratamiento farmacológico , Disacáridos/uso terapéutico , Doxorrubicina/análogos & derivados , Animales , Western Blotting , Carcinoma/metabolismo , Doxorrubicina/uso terapéutico , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
A variety of cytotoxic agents effective as antitumor drugs are known to kill tumor cells through induction of apoptosis as the most relevant modality of cell death. A specific role for the protein Bcl-2 in the cell death pathway induced by antimicrotubule agents has been proposed, because Bcl-2 phosphorylation occurs in response to microtubule damage. In this study, we compared efficacy, apoptosis, and Bcl-2 phosphorylation in the Bcl-2-overexpressing MX-1 human breast carcinoma xenograft after treatment with cytotoxic agents characterized by different mechanisms of action. We demonstrated that, in addition to antimicrotubule agents, effective DNA-damaging agents were also able to induce Bcl-2 phosphorylation irrespective of the type of genotoxic lesion. A comparison of effects of drugs belonging to the same class but endowed with a different antitumor activity (i.e. cisplatin versus a novel multinuclear platinum complex and doxorubicin versus a disaccharide analogue) showed a correlation between drug efficacy, apoptotic response, and Bcl-2 phosphorylation. In conclusion, overexpression of Bcl-2 did not counteract the apoptotic effects of a number of cytotoxic agents and could not be regarded as a mechanism of cellular resistance. Since Bcl-2 phosphorylation is a common event in response to different types of cytotoxic damage and is not only related to microtubule dysfunction, we suggest that many cell death pathways converge on Bcl-2 and protein phosphorylation is a step of the signaling cascade activated by diverse stimuli and likely related to the onset of drug-induced apoptosis.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Animales , Neoplasias de la Mama/genética , Daño del ADN/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Humanos , Ratones , Ratones Desnudos , Microtúbulos/efectos de los fármacos , Trasplante de Neoplasias , Fosforilación , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
The first antibiotic discovered, penicillin, appeared on the market just after the Second World War. Intensive research in subsequent years led to the discovery and development of cephalosporins, aminoglycosides, tetracyclines and rifamycin. The chemotherapeutic quinolones and the more recently discovered fluoroquinolones have added promising new therapeutic weapons to fight the microbial challenge. The major role pharmacokinetics has played in developing these compounds should be highlighted. Plasma concentration-time profiles and the therapeutic activity evoked by these compounds allow the therapeutic window, doses and dose turnovers to be appropriately defined as well as possible dose adjustment to be made in renal failure. The pharmacokinetics of antimicrobial agents were initially explored by using microbiological methods, but these lack specificity. The HPLC technique with UV, fluorometric, electrochemical and, in some cases, mass spectrometry detection has satisfactory solved the problem of antimicrobial agent assay for pharmacokinetic, bioavailability and bioequivalence purposes alike. Indeed, in these studies, plasma concentrations of the given analyte must be followed up for a period > or = 3 times the half-life, which calls for specific sensitive assays. In the review, the authors have described the analytical methods employed in the pharmacokinetics of antibiotics, including some chemotherapeutic agents which are used in medical practice as alternatives to antibiotics. The pharmacokinetic characteristics of each class of drugs are also briefly described, and some historical and chemical notes on the various classes are given.
Asunto(s)
Antibacterianos/análisis , Antibacterianos/farmacocinética , Cromatografía , Animales , HumanosRESUMEN
This paper describes a HPLC method for the determination of Pidotimod (3-L-pyroglutamyl-L-thiazolidine-4-carboxylic acid; PGT/1A), a new biological response modifier, in plasma. The column was an Aminex Ion Exclusion HPX 874 with a PRP precolumn, the mobile phase was 0.05% sulfuric acid-acetonitrile (88:12, v/v), the flow rate was 0.6 ml/min, the detection wavelength was 210 nm. Plasma (1 ml) was added with internal standard (Oxiracetam, concentration 400 micrograms/ml) (50 microliters) and 35% perchloric acid (100 microliters). The supernatant (0.5 ml) was added with mobile phase (0.5 ml) and, after centrifugation, injected into the column. The retention times of Pidotimod and Oxiracetam were 16.5 and 13.8 min. respectively. The method was validated for recovery, accuracy and reproducibility. The results after oral administration of 800 mg of Pidotimod in male volunteers were also given. This method is better than that previously described because it utilizes an internal standard and reaches a lower detection limit.
Asunto(s)
Factores Inmunológicos/sangre , Ácido Pirrolidona Carboxílico/análogos & derivados , Tiazoles/sangre , Adulto , Cromatografía Líquida de Alta Presión , Humanos , Factores Inmunológicos/farmacocinética , Masculino , Ácido Pirrolidona Carboxílico/sangre , Ácido Pirrolidona Carboxílico/farmacocinética , Tiazoles/farmacocinética , TiazolidinasRESUMEN
Preparation of (15 S)-hydroxy derivative (1-b), a key intermediate in the synthesis of PG like compounds, by reduction the corresponding enone (2), is described. High yields in (S)-isomer was obtained by means of chiral phase-transfer catalyst: (-)-N-(1-dodecyl)-N-methylephedrinium bromide. Eleven ammonium quaternary salts derived from (-)-N-methylephedrine were prepared and tested as catalyst in the reduction of enone (2) with NaBH4. Synthesis of enone (2), from phosphonate (6) (via Wadsworth-Emmons reaction) is also described.
Asunto(s)
Compuestos de Bifenilo/química , Ciclopentanos/química , Lactonas , Prostaglandinas Sintéticas/síntesis química , Catálisis , Espectroscopía de Resonancia Magnética , Prostaglandinas Sintéticas/químicaRESUMEN
An automated high-performance liquid chromatographic method for the direct injection of diltiazem plasma samples has been developed. It involves automatic injection of plasma samples (200 microliter) on a C18 pre-column (40 micron), clean-up of the pre-column with water to remove proteins and salts and transfer of the analytes to the analytical column. During the chromatographic process, the pre-column is reset with respect to the analytical column and flushed with different solvents to remove endogenous contaminants. The analysis is performed on a C18 column coupled to an ultraviolet detector. The whole process (on-line clean-up and chromatography) takes ca. 12 min and is completely automated. The detection limit of the method is ca. 2 ng/ml with 200-microliter aliquots of plasma sample. Very good results were obtained for the day-to-day and within-day reproducibilities (5.9 and 4.3%, respectively, in the concentration range 10-200 ng/ml in plasma).
Asunto(s)
Diltiazem/sangre , Administración Oral , Autoanálisis , Cromatografía Líquida de Alta Presión , Humanos , Masculino , Espectrofotometría UltravioletaRESUMEN
A high-performance liquid chromatographic method has been developed for the determination of a new cephalosporin antibiotic in plasma, urine and saliva (mixed saliva) using normal-phase technique and an NH2 bonded-phase column. The eluent mixture was a combination of acetonitrile and an aqueous solution of ammonium carbonate. The rapid method involved precipitation of protein from fluids by means of acetonitrile followed by automatic injection of the supernatant. The detection limit was 0.4 micrograms/ml for plasma, 3 micrograms/ml for urine and 0.03 micrograms/ml for saliva using UV detection.
Asunto(s)
Cefotaxima/análogos & derivados , Saliva/análisis , Acetonitrilos , Cefotaxima/análisis , Cefotaxima/sangre , Cefotaxima/orina , Ceftriaxona , Cromatografía Líquida de Alta Presión/métodos , Humanos , Espectrofotometría Ultravioleta , Factores de TiempoRESUMEN
A high-performance liquid chromatographic method has been developed for the determination of progabide and its main acid metabolite in blood, serum and plasma. The assay involved a single and rapid extraction of drug and metabolite into toluene from the biological matrix buffered at pH 4.8, evaporation of the organic phase, and chromatography of the extracts on a silica column with UV detection. SL 81 0142 was used as internal standard. The method was specific for unchanged drug and metabolite and had a sensitivity of ca. 50 ng/ml of biological fluid for both the compounds. The method was successfully applied to the analysis of progabide and its metabolite in biological fluids of patients administered orally with progabide for clinical pharmacokinetic studies and drug monitoring.
Asunto(s)
Ácido gamma-Aminobutírico/análogos & derivados , Biotransformación , Líquidos Corporales/análisis , Cromatografía Líquida de Alta Presión , Estabilidad de Medicamentos , Electroquímica , Humanos , Cinética , Solventes , Espectrofotometría Ultravioleta , Ácido gamma-Aminobutírico/análisis , Ácido gamma-Aminobutírico/sangreRESUMEN
A simple and rapid high-performance liquid chromatographic method was developed for the determination of vincamine in human plasma. Plasma samples were buffered at pH 9 and after extraction with tert.-butyl methyl ether back-extracted into 0.017 M orthophosphoric acid. Propranolol was used as the internal standard. An aliquot was injected on to a high-performance liquid chromatographic system using a C18 reversed-phase column and an acetonitrile-phosphate buffer containing triethylamine (30:70) as mobile phase. Detection was performed with an ultraviolet detector at 273 nm. The method had good accuracy and precision and the detection limit (0.3 ng/ml with a signal-to-noise ratio of 3:1) allowed the assessment of vincamine concentrations in plasma in pharmacokinetic studies on healthy human volunteers.
Asunto(s)
Vincamina/sangre , Cromatografía Líquida de Alta Presión , Humanos , Estándares de Referencia , Soluciones , Espectrofotometría Ultravioleta , Vincamina/farmacocinéticaRESUMEN
Zofenopril is a pro-drug designed to undergo metabolic hydrolysis yielding the active free sulfhydryl compound zofenoprilat, which is an angiotensin converting enzyme (ACE) inhibitor, endowed also with a marked cardioprotective activity. A simple, highly sensitive specific LC-MS-MS method was developed for the determination of zofenopril and zofenoprilat in human plasma. In order to prevent oxidative degradation of zofenoprilat and its internal standard, their free sulfhydryl groups were protected by treatment with N-ethylmaleimide (NEM), which produced the succinimide derivatives. The compounds and their corresponding fluorine derivatives, used as internal standards, were extracted from plasma with toluene. The reconstituted dried extracts were chromatographed and then monitored by a triple-stage-quadrupole instrument operating in the negative ion spray ionization mode. The method was validated over the concentration range of 1-300 ng/ml for zofenopril and 2-600 ng/ml for zofenoprilat. Inter- and intra-assay precision and accuracy of both zofenopril and zofenoprilat were better than 10%. The limit of quantitation was 1 ng/ml with zofenopril and 2 ng/ml with zofenoprilat. Extraction recovery proved to be on average 84.8% with zofenopril and 70.1% with zofenoprilat. Similar recoveries were shown by the above two internal standards. The method was applied to measure plasma concentrations of zofenopril and zofenoprilat in 18 healthy volunteers treated orally with zofenopril calcium salt at the dose of 60 mg.
Asunto(s)
Captopril/análogos & derivados , Captopril/sangre , Cromatografía Liquida/métodos , Espectrometría de Masas/métodos , Inhibidores de la Enzima Convertidora de Angiotensina/sangre , Calibración , Captopril/metabolismo , Estabilidad de Medicamentos , Humanos , Control de Calidad , Estándares de Referencia , Reproducibilidad de los ResultadosRESUMEN
A sensitive capillary gas chromatographic method was developed for the determination of fengabine (a GABAergic antidepressant drug) and some of its metabolites in plasma samples. The method involves a single and rapid liquid-liquid extraction of the parent drug and metabolites from plasma buffered at pH 5, evaporation of the organic phase under nitrogen, derivatization to tert.-butyldimethylsilyl ethers and esters and automatic gas chromatography on a fused-silica, silicone-bonded capillary column coupled to an electron-capture detector. The detection limit for fengabine and other compounds is lower than 1 ng/ml in plasma; the method was successfully applied to pharmacokinetic and drug monitoring clinical studies and tested on more than 2000 biological samples and was found not to suffer from endogenous or exogenous interferences.
Asunto(s)
Clorofenoles/sangre , Fenómenos Químicos , Química , Clorofenoles/metabolismo , Cromatografía de Gases , HumanosRESUMEN
The role of the site selectivity of topoisomerase II poisoning in the cytotoxic activity of anthracyclines has not been established. In this article, we have thus studied the levels and persistence of double-stranded DNA breaks (DSB) along with the cytotoxic activity in human leukemic HL60 cells of seven anthracyclines, including doxorubicin, daunorubicin, and idarubicin, as well as sugar-modified analogues characterized by an altered sequence specificity. Epimerization at the 3' position of the sugar moiety markedly affected the biological activity; indeed, a dramatic reduction of drug effects was evident for 3'-deamino-3'-epi-hydroxy-4'-deoxy-4'-amino-daunorubicin. The studied analogues could be gathered into three groups based on the DSB/cytotoxicity ratio. At equitoxic concentrations: (a) parent drugs and 3'-deamino-3'-epi-hydroxy-4'-deoxy-4'-amino-daunorubicin endowed with the same sequence specificity stimulated low DSB levels; (b) 3'-epi-daunorubicin and 3'-deamino-4'-deoxy-4'-epi-amino-idarubicin, which have a different sequence specificity, and teniposide (a structurally unrelated poison) stimulated higher amounts of DSB; and (c) 4-demethoxy-3'-deamino-3'-hydroxy-4'-epi-doxorubicin stimulated the highest DSB levels. For the last agent, a faster rate of cleavage resealing, which is consistent with a reduced DNA binding affinity, could account for the increased DSB/cytotoxicity ratio compared with parent drugs. However, for other analogues, the observed differences in DSB persistence/resealing could not completely explain the different DSB/cytotoxicity ratios. The results thus suggest that the cytotoxic potency of anthracyclines may be the result of an interplay of the level, the persistence, and the genomic localization of topoisomerase II-mediated DNA cleavage.
Asunto(s)
Antibióticos Antineoplásicos/toxicidad , Daño del ADN , ADN-Topoisomerasas de Tipo II/metabolismo , ADN de Neoplasias/efectos de los fármacos , ADN de Neoplasias/metabolismo , Antineoplásicos Fitogénicos/toxicidad , Apoptosis/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Células HL-60/efectos de los fármacos , Células HL-60/metabolismo , Células HL-60/patología , Humanos , Cinética , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Especificidad por Sustrato , Tenipósido/toxicidadRESUMEN
S-Naproxen betainate sodium salt monohydrate (naproxen-beta Na, CAS 104124-26-7, Aprenin) in 550 mg capsules (corresponding to 327 mg of naproxen) was administered to 24 healthy volunteers (12 males and 12 females) b.i.d. to steady state in order to check its bioavailability, food interaction and tolerability. Plasma concentrations of naproxen were measured by a well validated HPLC method with fluorimetric detection as a morning pre-dose on days 1 to 6 and in timed samples in three different situations, as follows: a) after the morning dose on day 7 in a fasting status, b) after the evening dose and dinner on day 7 and c) after the morning dose of day 8, taken after a high-fat content breakfast. Pharmacokinetic parameters were evaluated from plasma concentrations by non-compartmental analysis to describe the above three situations. The steady state was reached early, namely by the second day of treatment. The extent of absorption did not differ in the three situations tested, whereas the rate of absorption was fastest in fasting conditions, lowest with the evening dose and intermediate after the high-fat content breakfast. The slow absorption rate of the evening dose was attributed to a circadian rhythm and should allow therapeutically active levels early in the morning, when arthritis pain is particularly tedious. In the three situations explored Cmax, Cmin and AUC were associated with CV % values ranging from 11.7 to 17.2%, which are very low and rare in pharmacokinetic trials. This low variability should allow an accurate estimate of the therapeutic effect expected. Tolerability was checked by objective and subjective symptoms, including vital signs, blood/urine biochemical parameters and occult blood in stools, and proved to be very good. From the comparison of these data with those previously published by other authors who have administered 500 mg of naproxen b.i.d., pre-dose concentrations in a steady state proved to be similar, despite the different doses administered, whereas Cmax and AUC obtained in this study were marginally lower. The kind of food interaction was the same as previously described in literature with naproxen.
Asunto(s)
Antiinflamatorios no Esteroideos/efectos adversos , Antiinflamatorios no Esteroideos/farmacocinética , Naproxeno/efectos adversos , Naproxeno/farmacocinética , Adolescente , Adulto , Área Bajo la Curva , Disponibilidad Biológica , Cromatografía Líquida de Alta Presión , Grasas de la Dieta , Ayuno , Femenino , Interacciones Alimento-Droga , Humanos , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Espectrometría de FluorescenciaRESUMEN
Chlorodesmethyldiazepam (I) concentrations were followed for 72 h in the plasma of four volunteers given 2 mg of the drug orally. The drug appears to be well absorbed, reaching peak plasma levels of 20--30 ng ml-1 1--2 h after administration; 72 h after administration, plasma concentrations are still measurable (5 ng ml-1). The analytical method involved a single extraction of I from the plasma into benzene followed by centrifugation, evaporation, and gas-chromatographic analysis of the samples.
Asunto(s)
Ansiolíticos , Benzodiazepinas , Diazepam/análogos & derivados , Nordazepam/análogos & derivados , Administración Oral , Adulto , Biotransformación , Humanos , Masculino , Nordazepam/administración & dosificación , Nordazepam/sangre , Factores de TiempoRESUMEN
5 healthy volunteers (3 males, 2 females, aged 22-35 years) were given 600 mg ditazole (Ageroplas) every 12 h (9.00 a.m. and 9.00 p.m.) for 10 days. Venous blood was collected from all volunteers at 9.00 and 11.00 a.m. on days 0, 3, 5, 7 and 10. The threshold concentrations of adrenaline inducing two distinct waves of platelet aggregation (Born's method) within 3 min were determined each time. Blood levels of ditazole were measured by a gas-chromatographic technique using a nitrogen-phosphorus selective detector. The average blood levels of the drug ranged between 0.84 and 1.30 microgram/ml at 9.00 a.m., and between 1.93 and 2.85 microgram/ml at 11.00 a.m. Inhibition of platelet aggregation (expressed by a grading system in arbitrary units) ranged between 1.6 and 2.0 at 9.00 a.m. and between 2.0 and 2.4 at 11.00 a.m. The fluctuations of blood levels and platelet aggregation inhibitory activity of ditazole observed during the study period were virtually the same. It is suggested that the treatment schedule used in the present study results in blood levels of ditazole sufficient to reveal any consistent alteration of platelet function in normal subjects.
Asunto(s)
Oxazoles/sangre , Agregación Plaquetaria/efectos de los fármacos , Adulto , Epinefrina/farmacología , Femenino , Humanos , Masculino , Oxazoles/farmacología , Factores de TiempoRESUMEN
The percutaneous absorption of oxatomide gel at 5 per cent concentration was studied after single and repeated administration (85 mg b.i.d.) in six male and six female healthy volunteers, aged 25.7 +/- 0.8 years (mean +/- SEM) weighing 64.4 +/- 4.5 kg and the results compared with those obtained following a single oral dose (30 mg). The measurement of oxatomide was by means of a new sensitive and specific HPLC assay with limits of detection of 0.2 ng ml-1 in plasma and 1.0 ng ml-1 in urine. Poor percutaneous absorption was confirmed by the peak plasma concentrations which were 5.03 +/- 0.79 ng ml-1 following application of the gel for 7 days and 10.08 +/- 1.29 ng ml-1 following oral administration; the corresponding amounts of unchanged oxatomide recovered from 24 h urine collections were 1.42 +/- 0.39 micrograms and 3.93 +/- 0.92 micrograms.
Asunto(s)
Antagonistas de los Receptores Histamínicos H1/farmacocinética , Piperazinas/farmacocinética , Administración Cutánea , Administración Oral , Adulto , Esquema de Medicación , Femenino , Geles , Antagonistas de los Receptores Histamínicos H1/administración & dosificación , Antagonistas de los Receptores Histamínicos H1/sangre , Humanos , Masculino , Piperazinas/administración & dosificación , Piperazinas/sangre , Estándares de Referencia , Reproducibilidad de los Resultados , Absorción CutáneaRESUMEN
A high-performance liquid chromatographic method has been developed for the simultaneous determination of alpidem and its metabolites in human plasma. The method involved a single extraction of the parent drug and metabolites into diethyl ether from alkalinized plasma, evaporation of the organic solution and chromatography of the extracts on a C18 column coupled to a fluorimetric detector. An internal standard was used for the quantitative determination of the compounds. The method was selective for alpidem and three of its metabolites and has a limit of detection of less than 1 ng ml-1 for all the compounds. Since the chromatographic run took more than 20 min, the chromatographic process was fully automated and performed overnight.
Asunto(s)
Ansiolíticos/sangre , Anticonvulsivantes/sangre , Imidazoles/sangre , Piridinas/sangre , Ansiolíticos/orina , Anticonvulsivantes/orina , Biotransformación , Humanos , Imidazoles/orina , Indicadores y Reactivos , Cinética , Piridinas/orina , Espectrometría de FluorescenciaRESUMEN
The biochemical basis of the anti-proliferative effect of exogenous glutathione was investigated in A2780 ovarian carcinoma cells. Previous observations have implicated gamma-glutamyl transpeptidase-mediated pro-oxidant reactions as a primary mechanism of the extracellular effects of glutathione. In 2 cell lines (A2780 and IGROV-1), glutathione led to H(2)O(2) production, but only A2780 cells, characterized by low expression of detoxification enzymes, were sensitive to the thiol compound. In A2780 cells, glutathione exposure resulted in DNA single-strand break formation, as measured by alkaline elution. Glutathione-induced DNA damage generated significant levels of apoptosis in A2780 cells, but not in IGROV-1 cells. The capability of glutathione to induce apoptosis was associated with cleavage of poly(ADP-ribose)polymerase and with generation of a low-molecular-weight form of the pro-apoptotic protein bax. In A2780 cells, glutathione exposure was followed by p21 and Bax induction and p53 up-regulation, as expected for genotoxic stress. Consistently, analysis of cell-cycle perturbations demonstrated the occurrence of G(2)M accumulation after exposure to glutathione, similar to what was observed for H(2)O(2). Taken together, these results indicate that the cytotoxic effect of extracellular glutathione, related to membrane metabolism, is mediated by production of H(2)O(2) leading to DNA damage and a cellular response involving p53. These findings might also provide insights into the cellular and molecular determinants of chemosensitivity to DNA damaging agents, as oxidative stress is implicated in p53-dependent apoptosis.
Asunto(s)
Apoptosis/efectos de los fármacos , Daño del ADN , Glutatión/farmacología , Peróxido de Hidrógeno/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2 , Ciclo Celular/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Ciclinas/biosíntesis , Ciclinas/genética , ADN de Neoplasias/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes p53 , Inhibidores de Crecimiento/farmacología , Humanos , Inactivación Metabólica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Estrés Oxidativo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteínas Proto-Oncogénicas/biosíntesis , Proteínas Proto-Oncogénicas/genética , Células Tumorales Cultivadas/efectos de los fármacos , Proteína p53 Supresora de Tumor/biosíntesis , Proteína X Asociada a bcl-2RESUMEN
This paper describes a new sensitive gas chromatographic method with electron capture detector to assay estazolam in human plasma, which has been developed and validated for pharmacokinetic purposes. The drug and the internal standard (triazolam) were extracted from plasma buffered at pH 9.0 into toluene and analysed on a widebore DB 17 column. The calibration curve covered the 1.0-200 ng ml-1 range with a mean determination coefficient of 0.9996. The quantification limit was 1.0 ng ml-1. This method was used to investigate the bioequivalence of a new formulation of estazolam in drops (test) and the formulation in tablets (reference, ESILGAN). Both formulations were administered at a single dose of 2 mg in a clinical trial carried out on 24 healthy volunteers consisting of 12 males and 12 females, following a crossover randomised design in two periods with wash-out. The test and the reference formulations proved to be fully bioequivalent according to operating guidelines, namely through 90% confidence intervals in the 0.80-1.25 range.