Avaliação da Função Renal e Alterações Morfológicas em Ratos Tratados com Dipirona em Diferentes Doses / Evaluation of Renal Function and Morphological Changes in rats Treated with Dipyrone in Different Dosing

A dipirona é um fármaco muito utilizado para o tratamento da febre e da dor, entretanto, seus efeitos em doses altas podem levar a danosirreversíveis e até letais. Na presente investigação foram utilizados 32 ratos albinos machos da linhagem Wistar. Um grupo (1) recebeu uma dose única, 5.000 mg/Kg de dipirona por gavagem e o outro grupo (2) administração uma vez ao dia, por quatro dias consecutivos, de 600mg/Kg de dipirona. Ao final de quatro dias, para o grupo 2 e após 3 horas de ingestão do medicamento para o grupo 1, os animais foram eutanasiados em câmara de CO2, conforme a aprovação do Comitê de Ética Animal do Centro Universitário Barão de Mauá (CEPAn). Foram avaliados os parâmetros ureia e creatinina e a histopatologia renal. Os resultados dos escores hemorragia glomerular, congestão renal e processo inflamatório foram estatisticamente significativos nos três parâmetros analisados para dose única; congestão renal e processo inflamatório para dose múltipla. O marcador de função renal, ureia apresentou diferença estatística significativa com a dose única. Os resultados do modelo experimental atestam que a metodologia aplicada foi capaz de demonstrar as alterações histopatológicas do grupo tratado com dipirona dose única assim como alterações na dosagem de ureia e a preservação das características normais do grupo controle. O mesmo não ocorreu com o grupo tratado com doses múltiplas.

Dipyrone is a drug widely used for the treatment of fever and pain, but their effects at high doses can lead to irreversible and even lethal damages.In the present study it was used 32 male albino rats of Wistar strain. First group received a single dose 5.000 mg/kg by gavage dipyrone and the second given once daily for four consecutive days of 600 mg/kg Dipyrone. At the end of four days, for group 2 and 3 hours after ingestion of the drug for group 1, the animals were euthanized in a CO2 chamber, as the approval of the Baron of Maua University Center (Cepan) Animal Ethics Committee. The urea and creatinine parameters and renal histopathology were evaluated. The results of the scores glomerular bleeding, renal congestion and inflammation were statistically significant in all parameters analyzed for single dose; renal congestion and inflammation to multiple dose. The marker of renal function, urea statistically significant difference with the single dose. The results of the experimental model show that the methodology was able to demonstrate the histopathological changes in the group treated with single dose dipyrone as well as changes in the levels of urea and the preservation of the normal characteristics of the control group. This did not occur with the group treated with multiple doses.

Avaliação hepática em ratos Wistar expostos ao chá de Ayahuasca / Hepatic evaluation in Wistar rats exposed to Ayahuasca tea

O chá de Ayahuasca é empregado no xamanismo, na terapia antidrogas e indevidamente em recreações alucinógenas. O estudo teve como objetivo avaliar o fígado, tanto histológica como bioquimicamente, por se tratar do órgão responsável pela metabolização desse chá. Foram utilizados ratos Wistar (n=24), machos, adultos, divididos em dois grupos (controle e tratado). Foi administrada uma dose diária quatro vezes superior à utilizada no contexto religioso, por quatro dias consecutivos. Não houve alteração morfológica pela análise qualitativa, porém, na análise quantitativa do grupo tratado ocorreu redução do volume do hepatócito e de seu núcleo. Na análise bioquímica observou-se uma redução da Gama-Glutamil-Transferase e da Fosfatase Alcalina. De acordo com a dose e o tempo de exposição não foram encontradas alterações morfológicas nesse estudo. O estudo morfométrico mostrou alterações significativas no volume do hepatócito e de seu núcleo, onde ocorreu diminuição do volume do grupo tratado em relação ao grupo controle. Observou-se uma redução estatisticamente significativa nos parâmetros da Gama-Glutamil-Transferase e Fosfatase Alcalina. Nesse experimento foi observada uma redução do volume dos hepatócitos, sem ocorrer, entretanto, alterações na função e na estrutura hepática.

Ayahuasca tea is employed in Xamanism, in anti-drug therapy and illicitly in hallucinogenic recreations. Current study assesses the liver histologically and biochemically since it is an organ involved in the metabolism of the tea. Twenty-four male adult Wistar rats were divided into two groups (control and treated). A daily dose four times higher than that used within a religious context was given, during four consecutive days. There was no qualitative morphological change but a reduction of hepatocyte volume and its nucleus occurred when the quantitative analysis of the treated group was performed. Biochemical analysis showed a decrease in gamma-glutamyltransferasis and Alkaline Phosphatase. There were no morphological alterations for dose and exposure time in current study. Morphometric analysis showed significant alterations in hepatocyte volume and its nucleus where a decrease in volume of the treated group occurred when compared to that of control. A significant statistical decrease was detected in the gamma-glutamyl-transferasis and alkaline phosphatase parameters. Current analysis also detected a decrease in hepatocyte volume without alterations in hepatic functions and structure.

L-arginine, a nitric oxide precursor, reduces dapsone-induced methemoglobinemia in rats

Dapsone use is frequently associated to hematological side effects such as methemoglobinemia and hemolytic anemia, which are related to N-hydroxylation mediated by the P450 enzyme system. The aim of the present study was to evaluate the influence of L-arginine supplementation, a precursor for the synthesis of nitric oxide, as single or multiple dose regimens on dapsone-induced methemoglobinemia. Male Wistar rats were treated with L-arginine at 5, 15, 30, 60 and 180 mg/kg doses (p.o., gavage) in single or multiple dose regimens 2 hours prior to dapsone administration (40 mg/kg, i.p.). The effect of the nitric oxide synthase inhibitor L-NAME was investigated by treatment with multiple doses of 30 mg/kg (p.o., gavage) 2 hours before dapsone administration. Blood samples were collected 2 hours after dapsone administration. Erythrocytic methemoglobin levels were assayed by spectrophotometry. The results showed that multiple dose supplementations with 5 and 15 mg/kg L-arginine reduced dapsone-induced methemoglobin levels. This effect is mediated by nitric oxide formation, since the reduction in methemoglobin levels by L-arginine is blocked by simultaneous administration with L-NAME, a nitric oxide synthase inhibitor.

O uso da dapsona é frequentemente associado a efeitos adversos hematológicos, como a metemoglobinemia e anemia hemolítica, ambos relacionados com a N-hidroxilação mediada pelo sistema P450. O objetivo do estudo foi avaliar a influência da suplementação de L-arginina, um precursor da síntese de óxido nítrico, administrado em regime de dose única ou múltipla na metemoglobinemia induzida pela dapsona. Ratos machos Wistar foram tratados com L-arginina (po, gavagem) em dose única ou múltipla de 5, 15, 30, 60 e 180 mg/kg 2 horas antes da administração de dapsona (40 mg/kg, ip). O efeito do L-NAME, um inibidor de óxido nítrico sintase (NOS), foi avaliado através do tratamento com doses múltiplas de 30 mg/kg. Amostras de sangue foram coletadas duas horas após a administração de dapsona. A concentração de metemoglobina eritrocitária foi analisada por espectrofotometria. Os resultados mostraram que a suplementação em dose múltipla de 5 e 15 mg/kg de L-arginina reduziu os níveis de metemoglobina induzida pela dapsona. Este efeito é mediado pela formação de óxido nítrico, uma vez que a redução nos níveis de metemoglobina pela L-arginina é bloqueada pela administração simultânea de L-NAME, um inibidor da óxido nítrico sintase.

Rifampicin determination in plasma by stir bar-sorptive extraction and liquid chromatography.

A sensitive and reproducible stir bar-sorptive extraction and high performance liquid chromatography-UV detection (SBSE/HPLC-UV) method for therapeutic drug monitoring of rifampicin in plasma samples is described and compared with a liquid:liquid extraction (LLE/HPLC-UV) method. This miniaturized method can result in faster analysis, higher sample throughput, lower solvent consumption and less workload per sample while maintaining or even improving sensitivity. Important factors in the optimization of SBSE efficiency such as pH, temperature, extraction time and desorption conditions (solvents, mode magnetic stir, mode ultrasonic stir, time and number of steps) were optimized recoveries ranging from 75 to 80%. Separation was obtained using a reverse phase C(8) column with UV detection (254nm). The mobile phase consisted of methanol:0.25N sodium acetate buffer, pH 5.0 (58:42, v/v). The SBSE/HPLC-UV method was linear over a working range of 0.125-50.0microgmL(-1). The intra-assay and inter-assay precision and accuracy were studied at three concentrations (1.25, 6.25 and 25.0microgmL(-1)). The intra-assay coefficients of variation (CVs) for all compounds were less than 10% and all inter-CVs were less than 10%. Limits of quantification were 0.125microgmL(-1). Stability studies showed rifampicin was stable in plasma for 12h after thawing; the samples were also stable for 24h after preparation. Based on the figures of merit results, the SBSE/HPLC-UV proved to be adequate to the rifampicin analyses from therapeutic to toxic levels. This method was successfully applied to the analysis of real samples and was as effective as the LLE/HPLC-UV method.

Quantification of carbamazepine, carbamazepine-10,11-epoxide, phenytoin and phenobarbital in plasma samples by stir bar-sorptive extraction and liquid chromatography.

A sensitive and reproducible stir bar-sorptive extraction and high-performance liquid chromatography-UV detection (SBSE/HPLC-UV) method for therapeutic drug monitoring of carbamazepine, carbamazepine-10,11-epoxide, phenytoin and phenobarbital in plasma samples is described and compared with a liquid:liquid extraction (LLE/HPLC-UV) method. Important factors in the optimization of SBSE efficiency such as pH, extraction time and desorption conditions (solvents, mode magnetic stir, mode ultrasonic stir, time and number of steps) assured recoveries ranging from 72 to 86%, except for phenytoin (62%). Separation was obtained using a reverse phase C18 column with UV detection (210nm). The mobile phase consisted of water:acetonitrile (78:22, v/v). The SBSE/HPLC-UV method was linear over a working range of 0.08-40.0microgmL(-1) for carbamazepine, carbamazepine-10,11-epoxide and phenobarbital and 0.125-40.0microgmL(-1) for phenytoin, The intra-assay and inter-assay precision and accuracy were studied at three concentrations (1.0, 4.0 and 20.0microgmL(-1)). The intra-assay coefficients of variation (CVs) for all compounds were less than 8.8% and all inter-CVs were less than 10%. Limits of quantification were 0.08microgmL(-1) for carbamazepine, carbamazepine-10,11-epoxide and phenobarbital and 0.125microgmL(-1) for phenytoin. No interference of the drugs normally associated with antiepileptic drugs was observed. Based on figures of merit results, the SBSE/HPLC-UV proved adequate for antiepileptic drugs analyses from therapeutic levels. This method was successfully applied to the analysis of real samples and was as effective as the LLE/HPLC-UV method.

Reliable HPLC method for therapeutic drug monitoring of frequently prescribed tricyclic and nontricyclic antidepressants.

A new high-performance liquid chromatography method is presented for the determination of 10 frequently prescribed tricyclic and nontricyclic antidepressants: imipramine, amitriptyline, clomipramine, fluoxetine, sertraline, paroxetine, citalopram, mirtazapine, moclobemide and duloxetine. The simple and accurate sample preparation step, consisted of liquid:liquid extraction with recoveries ranging between 72% and 86%, except for moclobemide (59%). Separation was obtained using a reverse phase Select B column under isocratic conditions with UV detection (230 nm). The mobile phase consisted of 35% of a mixture of acetonitrile/methanol (92:8, v/v) and 65% of 0.25 mol L(-1) sodium acetate buffer, pH 4.5. The standard curves were linear over a working range of 2.5-1000 ng mL(-1) for moclobemide, 5-2000 ng mL(-1) for citalopram, duloxetine, fluoxetine, 10-2000 ng mL(-1) for sertraline, imipramine, paroxetine, mirtazapine and clomipramine. The intra-assay and inter-assay precision and accuracy were studied at three concentrations (50, 200, and 500 ng mL(-1)). The intra-assay coefficients of variation (CVs) for all compounds were less than 8.8%, and all inter-CVs were less than 10%. Limits of quantification were 2.5 ng mL(-1) for moclobemide, 5 ng mL(-1) for citalopram, duloxetine and amitriptyline, and 10 ng mL(-1) for mirtazapine, paroxetine, imipramine, fluoxetine, sertraline, and clomipramine. No interference of the drugs normally associated with antidepressants was observed. The method has been successfully applied to the analysis of real samples, for the drug monitoring of ten frequently prescribed tricyclic and non-tricyclic antidepressant drugs.

Ranitidine treatment inducing methemoglobinemia in male Wistar rats

Drug idiosyncrasy is an adverse event of unknown etiology that occurs in a small fraction of people taking a drug. The histamine-2(H2)-receptor antagonist ranitidine causes idiosyncratic reactions in patients. To investigative the hypothesis that ranitidine could induce hematological toxic effects, the drug was administered intraperitoneally(ip) to two of six groups of 200-220 g male Wistar rats (n=6). Group I received as single dose of saline solution (NaCl, 200 µL) Group II received 200 µL of NaCl, ip, at 0, 24, 48 and 72 h, Group III (controls of the vehicle) received as single dose of dimethyl sulfoxide (DMSO, 200 µL), ip, Group IV (controls of the vehicle) received 200 µL of DMSO, ip, at 0, 24, 48, and 72 h, Group V received a single dose of 100 mg/kg of ranitidine in 200 µL of DMSO) ip, Group VI received 50 mg/kg of ranitidine, in 200 µL of DMSO, ip, at 0, 24, 48, and 72 h...

[Correlation between dose/plasma concentration and assessment of hepatic and renal changes in Wistar rats treated with the ROM scheme]. / Correlação entre dose/concentração plasmática e avaliação de alterações hepáticas e renais em ratos Wistar tratados com o esquema ROM.

Leprosy, a chronic granulomatous infectocontagious disease transmitted by Mycobacterium leprae, continues to be prevalent today, especially in underdeveloped countries and its paucibacillary form with a single lesion is being treated with rifampicin (600mg), ofloxacin (400mg) and minocycline (100mg) administered as a single dose (ROM scheme). Thus, the objective of the present study was to investigate the dose/plasma concentration correlation versus biochemical changes occurring in male Wistar rats receiving a single dose of rifampicin, ofloxacin and minocycline in mono- and polytherapy. Rifampicin and ofloxacin showed an increased concentration in plasma when administered in polytherapy, whereas minocycline was reduced, probably due to interference with its biotransformation and excretion. Biochemical analyses showed that rifampicin is probably responsible for hepatic and renal changes and that the medicamentous interactions involving the drug require individualized studies, especially when the drug is associated with ofloxacin and minocycline therapy.

Correlação entre dose/concentração plasmática e avaliação de alterações hepáticas e renais em ratos Wistar tratados com o esquema ROM / Correlation between dose/plasma concentration and assessment of hepatic and renal changes in Wistar rats treated with the ROM scheme

A hanseníase, doença crônica, granulomatosa, infecto-contagiosa, transmitida pelo Mycobacterium leprae, ainda se mantém prevalente nos dias atuais, principalmente em países subdesenvolvidos e a sua forma paucibacilar com lesão única, vem sendo tratada através da administração de rifampicina (600mg), ofloxacina (400mg) e minociclina (100mg), em dose única (esquema ROM). Assim, o objetivo deste trabalho foi investigar a correlação dose/concentração plasmática versus alterações bioquímicas na administração da rifampicina, ofloxacina e minociclina a ratos machos Wistar, em regime de dose única em mono e politerapia. Concluímos que a rifampicina e a ofloxacina sofreram um aumento na concentração plasmática quando administrados em politerapia, enquanto que a minociclina sofreu uma redução, provavelmente por interferências na biotransformação e excreção. Constatamos através das análises bioquímicas que a rifampicina provavelmente é a responsável por alterações hepáticas e renais, e que as interações medicamentosas envolvendo o fármaco exigem estudos individualizados principalmente quando o fármaco é usado associado a ofloxacina e minociclina.

Effects of H(2)-receptor antagonists on dapsone-induced methaemoglobinaemia in rats.

Dapsone (DDS) (4,4'diaminodiphenylsulfone), the drug of choice for the treatment of leprosy, frequently induces haemolytic anaemia and methaemoglobinaemia. N-hydroxylation, one of the major pathways of biotransformation, has been constantly related to the methaemoglobinaemia observed with the use of the drug. In order to determine the reversible inhibition of this toxicologic bioactivation pathway without changing the detoxification pathways of the drug or cytosolic acetylation, cimetidine (CIM), ranitidine and famotidine were administered in combination with DDS to male Wistar rats weighing 200-220 g. The animals were divided into nine groups of eight: group 1 received a single dose of 40 mg kg (-1) DDS in dimethylsulfoxide (DMSO) and groups 2-4 received the same treatment as group 1 but after the administration of a single dose of 100, 150 and 200 mg kg (-1) CIM, respectively, injected 2 h prior DDS administration. Groups 5-9 received the same treatment as group 2 but after the treatment of ranitidine (50 and 100 mg kg (-1) intraperitoneally (i.p.) in 200 microl DMSO) and famotidine (10, 50 and 100 mg kg (-1) i.p. in 200 microl DMSO), respectively. The animals were then anaesthetized with ether and blood was collected from the aorta for the determination of plasma DDS and monoacetyldapsone concentrations by HPLC and later for the determination of methaemoglobinaemia by spectrophotometry. CIM showed a higher affinity for cytochrome P-450 than famotidine and ranitidine. The results obtained showed the potentiality of the pharmacological effects of DDS with a low risk of adverse reactions, especially methaemoglobinaemia, which is dose dependent.

Efeito hemoprotetor da cimetidina na metemoglobinemia induzida pela dapsona em regime de dose única em ratos / Hemoprotective effect of cimetidine single dosis schedule on dapsone induced methemoglobinemia in rats

A dapsona (4,4-diaminodifenilsulfona), quimioterápico bacteriostático utilizado no tratamento da hanseníase, vem sendo associada a intercorrências clínicas, principalmente devido à sua hemotoxicidade caracterizada por metemoglobinemia e anemia hemolítica. A N-hidroxilação, uma das principais vias de biotransformação da dapsona, vem sendo associada constantemente a quadros de metemoglobinemia decorrentes de sua utilização. Com o objetivo de verificar-se a inibição reversível da via de bioativação toxicológica, sem alterar as vias de destoxificação do composto, a acetilação citosólica, a cimetidina foi administrada concomitantemente à dapsona em ratos machos Wistar, com peso variando entre 200 e 220 g, divididos em 8 grupos (n=6 por grupo), em estudo de dose única...

Efeito de antagonistas de receptores H2 na metemoglobinemia induzida pela dapsona em ratos / ?

Dapsona, farmaco bacteriostatico utilizado no tratamento da hanseniase, vem sendo associado a intercorrencias clinicas, principalmente devido a sua hemotoxicidade. A N-hidroxilacao, uma das principais vias de biotransformacao da dapsona vem sendo associada constantemente a quadros de metemoglobinemia decorrentes do uso da dapsona. Com objetivo de se verificar a inibicao reversivel desta via de bioativacao toxicologica, sem alterar os caminhos de detoxificacao do farmaco, a aceltilacao citosolica, a cimetidina, ranitidina e famotidina foram administradas concomitantemente a dapsona em ratos machos Wistar, com peso variando entre 200 e 250g divididos em 30 grupos (n=6), em estudo de doses unicas e multiplas. Apos as administracoes, os ratos foram submetidos a coleta de sangue