Post mortem changes: challenges in drug analysis in the diagnosis of deaths from intoxication

In forensic toxicology, alternative matrices and sampling sites are required for a correlation of antemortem and postmortem concentrations with the least possible error. Postmortem redistribution phenomena and biochemical changes inherent to these processes are possible, and represent interferences in these analyses. This study aimed to perform a bibliographic review through Pubmed database within a 10-year period of time, using the keywords: forensic analysis AND redistribution. We observed that for quantitative analyses the preferred matrix is blood from peripheral vessels, and when it is not available, vitreous humor is a great specimen for choice.

STABILITY-INDICATING LC METHOD FOR THE QUANTIFICATION OF MIDAZOLAM ACTIVE PHARMACEUTICAL INGREDIENT AND IN PHARMACEUTICAL FORMULATIONS

A stability-indicating LC method was validated for the quantification of midazolam (MDZ) active pharmaceutical ingredient (API) and in pharmaceutical formulations. Isocratic chromatography was performed on C18 column with mobile phase containing methanol/acetonitrile/water (45:35:20 v/v/v) with 0.4% of triethylamine pH 6.5. The validation included specificity, linearity, accuracy, precision and robustness. In specificity, after acid, basic, neutral, oxidant and thermal degradation, it was found that the concentration of MDZ decreased substantially, with the appearance of peaks representatives of the degradation products, proving the stability-indicating power of the method. The response was linear in the range 50.0 – 250.0 µg.mL-1, with 11.73 µg.mL-1 and 3.87 µg.mL-1 as LOQ and LOD, respectively. Recoveries ranged between 98.68 and 100.41%. The relative standard deviation values for intra and interday precision were 1.11%, 0.82% and 1.47%, respectively. The tablets and injections containing MDZ were approved in the assay and content uniformity. The method can be adopted by pharmacopeias and for routine quality control for analysis of MDZ API, tablets and injection

Lead evaluation in children's lipsticks through atomic absorption spectrometry

Lead is a metal with recognized toxicity and it is known that it may be a contaminant in lipsticks. In Brazil, the Health Regulatory Agency (ANVISA) determines that the maximum limit allowed for the presence of lead in lipsticks is 20 ppm. Children are more vulnerable to lead toxicity. The objective of this study was to evaluate the presence of lead in infant lipsticks. Nineteen samples from four different brands sold in Brazil were evaluated. After sample extraction, analyses were performed by graphite furnace atomic absorption spectrometry. Lead was not detected in any of the tested lipsticks. Considering the presence of lead in adult makeup, the present study reinforces the need to use products intended for children considering kids are more vulnerable to lead toxic effects

SILYMARIN ELICITS PARTIAL PROTECTION AGAINST METHOTREXATE-INDUCED HEPATOTOXICITY IN WISTAR RATS

Objective:  Methotrexate (MTX) is the gold standard in rheumatoid arthritis treatment; however, its continued use is related to hepatotoxicity. Thus, the aim of the present work is to evaluate the protective role of silymarin (SLM) against MTX-induced hepatotoxicity.Methods:  Male Wistar rats were treated by gavage for 4 weeks as followed:  Control (saline solution, daily), MTX (900µg/kg, once a week and daily with saline solution), SLM (50 mg/kg, daily) and MTX+SLM (MTX 900µg/kg, once a week, SLM 50 mg/kg, daily).Results:  MTX rats presented macro-and microscopic changes in the liver that was not counteracted by SLM. SLM was able to prevent the significant increase in lipid peroxidation observed in the hepatic tissue of MTX rats. SLM+MTX presented a decrease in the hepatic non-proteic thiols compared to control; suggesting SLM favors detoxification through glutathione conjugation. It was also seen changes in the relative mass of the spleen and lungs of MTX rats.Conclusions:  SLM protects partially against MTX-induced hepatotoxicity.Â

Potencial antimicrobiano e caracterização fitoquímica de extratos da casca do caule de Drimys brasiliensis Miers (Winteraceae)

Introdução: na área da saúde, a resistência aos antimicrobianos tem se constituído um grave problema, fazendo com que a busca por alternativas farmacológicas assuma grande importância. Produtos de origem natural, como extratos de plantas, têm sido pesquisados para uso potencial no tratamento antibacteriano. Drimys brasiliensis Miers (Winteraceae) é uma árvore nativa do Brasil que tem sido utilizada como medicinal. Objetivo: investigar o perfil fitoquímico e a atividade antimicrobiana de amostras obtidas a partir de extratos da casca do caule de Drimys brasiliensis. Metodologia:  frações de características químicas variadas foram obtidas a partir de extratos da casca do caule da planta. Realizou-se o screening fitoquímico destas frações através de reações gerais de caracterização e cromatografia em camada delgada. As amostras foram testadas por difusão em ágar e por microdiluição contra bactérias Gram-positivas e Gram-negativas e contra uma levedura. Resultados: o perfil fitoquímico mostrou diferenças entre as frações analisadas. Alcaloides foram detectados nas frações mais apolares. Taninos e flavonoides estavam presentes nas amostras mais polares e obtidas sem o uso de calor. As amostras com maior atividade antimicrobiana foram as obtidas com os solventes hexano e diclorometano, de características mais apolares. Destaca-se a atividade contra Bacillus cereus, Staphylococcus epidermidis, Staphylococcus aureus resistente à meticilina e Candida albicans. Conclusão: extratos da casca do caule de Drimys brasiliensis apresentam potencial antimicrobiano, com destaque para as frações mais apolares contra bactérias gram positivas

Mitochondrial Cumulative Damage Induced by Mitoxantrone: Late Onset Cardiac Energetic Impairment

Mitoxantrone (MTX) is a chemotherapeutic agent, which presents late irreversible cardiotoxicity. This work aims to highlight the mechanisms involved in the MTX-induced cardiotoxicity, namely the effects toward mitochondria using in vivo and in vitro studies. Male Wistar rats were treated with 3 cycles of 2.5 mg/kg MTX at day 0, 10, and 20. One treated group was euthanized on day 22 (MTX22) to evaluate the early MTX cardiac toxic effects, while the other was euthanized on day 48 (MTX48), to allow the evaluation of MTX late cardiac effects. Cardiac mitochondria isolated from 4 adult untreated rats were also used to evaluate in vitro the MTX (10 nM, 100 nM, and 1 lM) direct effects upon mitochondria functionality. Two rats of MTX48 died on day 35, and MTX treatment caused a reduction in relative body weight gain in both treated groups with no significant changes in water and food intake. Decreased levels of plasma total creatine kinase and CK-MB were detected in the MTX22 group, and increased plasma levels of lactate were seen in MTX48. Increased cardiac relative mass and microscopic changes were evident in both treated groups. Considering mitochondrial effects, for the first time, it was evidenced that MTX induced an increase in the complex IV and complex V activities in MTX22 group, while a decrease in the complex V activity was accompanied by the reduction in ATP content in the MTX48 rats. No alterations in mitochondria transmembrane potential were found in isolated mitochondria from MTX48 rats or in isolated mitochondria directly incubated with MTX. This study highlights the relevance of the cumulative MTX-induced in vivo mitochondriopathy to the MTX cardiotoxicity.This work was supported by the Fundação para a Ciência e Tecnologia (FCT)—project (EXPL/DTP-FTO/0290/ 2012)—QREN initiative with EU/FEDER financing through COMPETE— Operational Programme for Competitiveness Factors. LGR, VMC, and RJD-O thank FCT for their PhD Grant (SFRH/BD/63473/ 2009) and Post-doc Grants (SFRH/BPD/63746/2009) and (SFRH/ BPD/36865/2007), respectively. The authors are grateful to Fundação para a Ciência e Tecnologia for grant no. Pest C/EQB/LA0006/2011

Eugenia pyriformis ('uvaia') intoxication of sheep / Intoxicação por Eugenia pyriformis ('uvaia') em ovinos

This study describes the clinical and pathological picture of a disease followed by death in sheep raised for subsistence, characterized by difficulty in locomotion, sternal decubitus, severe dyspnea, opisthotonos, and pedaling movements, followed by death. The disease occurred in February 2020 in the northern state of Rio Grande do Sul, Brazil, due to the consumption of the fruits of Eugenia pyriformis ('uvaia'). All sheep that became ill later died, resulting in morbidity of 68.75% and mortality of 100%. An autopsy revealed the presence of E. pyriformis fruits with or without seeds in the rumen, in addition to the diet provided to the animals. The main histological findings consisted of edema and perivascular cuffs in the frontal telencephalon, nephrosis and glomerular necrosis, in addition to glomerulosclerosis. One sheep had heightened urea and creatinine and lower levels of non-protein thiols. Deaths ceased once animals were prevented from accessing E. pyriformis. These findings confirm that this plant was responsible for the deaths of the animals and point to the possibility of E. pyriformis having neurotoxic and nephrotoxic action on sheep

Effect of reciprocating instrumentation on chlorhexidine substantivity on human dentin: chemical analysis followed by confocal laser microscopy

The present research analyzed the reciprocating instrumentation associated to chlorhexidine (CHX) substantivity as its correlation with E. faecalis viabilityin ex vivoroot canals. Eighty extracted single-rooted human teeth were used, being 40 to high-performance liquid chromatography (HPLC) and 40 to confocal laser scanning microscopy (CLSM). In both, teeth were decoronated and the cervical third was prepared. In the CLSM analysis, the root canals were inoculated with E. faecalisfor 14 days. Samples were divided into 4 groups (n=10) according to instrumentation technique: no instrumentation and irrigation with distilled water (control); manual instrumentation (K-File); rotary instrumentation (ProTaper Next); and reciprocating instrumentation (Reciproc R25). Two percent chlorhexidine was applied as irrigating substance in experimental groups. Longitudinal grooves resulted in 2 halves root and 20 proof bodies in each group. Samples were divided by chance in two groups (n=10) and theoutcomes were evaluated after two days and one week. The retained chlorhexidine and live cells after instrumentation techniques in each evaluation time was measured by HPLC and CLSM, respectively. Specific analysis was applied for experimental tests (p≤0.05). Both rotary as well as reciprocating techniques significantly reduced the amount of chlorhexidine on dentin in all observation periods (p0.05). Reciprocating instrumentation does not interfere on chlorhexidine substantivity

N-acetylcysteine+nimesulide : an association strategy aiming to prevent nimesulide-induced hepatotoxicity

Introduction: Nimesulide is a potent anti-inflammatory with rapid and long-lasting effects, but also with a high risk of hepatotoxicity. Objective: This work aimed to prevent nimesulide-induced hepatotoxicity through the association of nimesulide with a hepatoprotective agent. Materials and Methods: First, we tested three hepatoprotective agents: N-acetylcysteine, L-carnitine, and Gingko biloba extract in an in vitro hepatic cell model. Both N-acetylcysteine and G. biloba showed promisor results. We selected N-acetylcysteine to continue the studies in an animal model. In vivo study was performed using male Wistar rats divided in 4 groups: control, nimesulide (100mg/kg/day), nimesulide (100mg/kg/day) + N-acetylcysteine (100mg/kg/day) and N-acetylcysteine alone (100mg/kg/day). Treatments were given by gavage, daily, for 15 days. Results: Animals receiving nimesulide alone showed lower body weight gain compared to control. Body weight gain in the nimesulide + N-acetylcysteine group was higher than nimesulide alone, evidencing lower toxicity. However, the body weight gain of the nimesulide + N-acetylcysteine group was still lower than the control animals. Animals treated with nimesulide alone presented an increased relative mass of heart, liver, and spleen and significant hepatic damage seen in microscopy when compared to other groups. N-acetylcysteine co-administered with nimesulide prevented the increased heart mass, but the same was not true with liver and spleen. Conclusions: This work evidence partial protection elicited by the association of N-acetylcysteine and nimesulide against nimesulide-induced hepatotoxicity.Introdução: A nimesulida é um potente anti-inflamatório com efeitos rápidos e duradouros, mas também com alto risco de hepatotoxicidade. Objetivo: Este trabalho teve como objetivo prevenir a hepatotoxicidade induzida pela nimesulida por meio da associação de nimesulida a um hepatoprotetor. Materiais e Métodos: Primeiro, testamos três possíveis hepatoprotetores: N-acetilcisteína, L-carnitina e extrato de Gingko biloba em um modelo de células hepáticas in vitro. N-acetilcisteína e G. biloba apresentaram bom potencial hepatoprotetor. Selecionamos a N-acetilcisteína para continuar os estudos em um modelo animal. O estudo in vivo foi realizado com ratos Wistar machos divididos em 4 grupos: controle, nimesulida (100 mg/kg/dia), nimesulida (100 mg/kg/dia) + N-acetilcisteína (100 mg/kg/dia) e N-acetilcisteína isolada (100 mg/kg/dia). Os tratamentos foram realizados por gavagem, diariamente, por 15 dias. Resultados: Os animais que receberam nimesulida isoladamente apresentaram menor ganho de peso corporal em comparação ao controle. O ganho de peso corporal no grupo nimesulida + N-acetilcisteína foi maior que o nimesulida isolado, evidenciando menor toxicidade. No entanto, o ganho de peso corporal do grupo nimesulida + N-acetilcisteína ainda era menor do que os animais controle. Os animais tratados com nimesulida isoladamente apresentaram aumento da massa relativa do coração, fígado e baço e dano hepático significativo observado na microscopia quando comparados a outros grupos. A N-acetilcisteína co-administrada com nimesulida impediu o aumento da massa cardíaca, mas tal fato não ocorreu com o fígado e o baço. Conclusões: Este trabalho evidencia proteção parcial provocada pela associação de N-acetilcisteína e nimesulida contra hepatotoxicidade induzida por nimesulida.Introducción: La nimesulida es un potente antiinflamatorio con efectos rápidos y duraderos, pero también con un alto riesgo de hepatotoxicidad. Objetivo: Este estudio tuvo como objetivo prevenir la hepatotoxicidad inducida por nimesulida combinando nimesulida con un medicamento hepatoprotector. Materiales y métodos: Primero, probamos tres candidatos: N-acetilcisteína, L-carnitina y extracto de Gingko biloba en un modelo de células hepáticas in vitro. N-acetilcisteína y G. biloba mostraron buen potencial. A continuación, seleccionamos N-acetilcisteína para continuar los estudios en un modelo animal. El estudio in vivo se realizó con ratas Wistar machos divididas en 4 grupos: control,nimesulida (100 mg/kg/día), nimesulida (100 mg/kg/día) + N-acetilcisteína (100 mg/kg/día) y N-acetilcisteína aislada (100 mg/kg/día). Los tratamientos se realizaron por sonda, diariamente, durante 15 días. Resultados: Los animales que recibieron nimesulida sola mostraron menos aumento de peso corporal en comparación con el control. El aumento de peso corporal en el grupo de nimesulida + N-acetilcisteína fue mayor que el de nimesulida sola, mostrando menos toxicidad. Sin embargo, el aumento de peso corporal del grupo nimesulida + N-acetilcisteína fue aún menor que el de los animales de control. Los animales tratados con nimesulida sola mostraron un aumento en la masa relativa del corazón, el hígado y el bazo y un daño hepático significativo observado al microscopio en comparación con otros grupos. La N-acetilcisteína administrada conjuntamente con nimesulida evitó el aumento de la masa cardíaca, pero no sucedió lo mismo con el hígado y el bazo. Conclusiones: Este trabajo muestra una protección parcial causada por la asociación de N-acetilcisteína y nimesulida contra la hepatotoxicidad inducida por la nimesulida