Evaluation of the nicotine metabolite ratio in smoking patients treated with varenicline and bupropion.

Background: Smoking is the leading cause of preventable death worldwide. It is responsible for several types of cancer, cardiovascular diseases, and diseases of the reproductive system, among others. Therefore, advances in research are increasingly necessary in order to make smoking cessation treatment more effective. Some studies have investigated the association of the nicotine metabolite ratio (NMR) with general characteristics and treatment outcomes. In the present study, the main aim was to evaluate the NMR in smoking patients from an Assistance Program of a tertiary cardiology hospital. Methodology: Serum samples were collected from 185 patients at T0 (while patients were still smoking and before starting pharmacological treatment). Cotinine and hydroxycotinine analytes were measured using liquid-chromatography tandem mass-spectrometry (LC-MS/MS). By looking at the relationship between hydroxycotinine and cotinine, we can obtain the NMR, with which it is possible to classify patients into slow metabolizers (NMR < 0.31), as well as normal or fast metabolizers (NMR ≥ 0.31). Results: From 185 patients, 55 were considered slow metabolizers and 130 as normal/fast. The metabolite averages were associated with the number of cigarettes smoked per day (p < 0.001 for cotinine and 0.023 hydroxycotinine). However, we were unable to analyze the association of the NMR with general and clinical characteristics of patients under smoking cessation treatment. Conclusion: We were able to evaluate the NMR, and to observe categories of metabolizers in Brazilian patients under pharmacological treatments. Thus, this study can contribute to the indication of a form of analysis, which might form part of the customization of smoking cessation treatments and, consequently, improve the success rates.

Liquid chromatography-tandem mass spectrometry method for simultaneous quantification of neurotransmitters in rat brain tissue exposed to 4'-fluoro-α-PHP.

The combination of different advanced analytical techniques makes it possible to determine the concentrations of neurotransmitters in various biological matrices, providing a complex and comprehensive study of the effects of psychoactive substances. The present study aimed to develop and validate a fast and simple analytical method for the determination of acetylcholine, serotonin, γ-aminobutyric acid, glutamate, dopamine and metabolites in rats brain tissue by liquid chromatography coupled to tandem mass spectrometry. The brain was homogenized and aliquots of the sample, dopamine-d4 , and acetone were added to a tube and then vortexed and centrifuged. The supernatant was collected and dried. The residue was reconstituted and injected. The LLOQ ranged from 0.001 to 1 µg/g; the intra-run precision ranged from 0.47 to 11.52%; the inter-run precision ranged from 0.68 to 17.54%; and the bias ranged from 89.10 to 109.60%. As proof of concept, the method was applied to animals exposed to the synthetic cathinone 4'-fuoro-α-pyrrolidinohexanophenone (300 mg/kg). In addition, the workflow proved to be simple, rapid and useful to estimate the concentration of neurotransmitters. This analytical tool can be used to support the investigation of the changes in the neurochemical profile for the characterization of the mechanism of action of psychoactive substances, as well as both neurological and psychiatric diseases.

Green sample preparations for the bioanalysis of drugs of abuse in complex matrices.

The sample preparation is the most critical step involved in the bioanalytical process. When dealing with green analytical chemistry, sample preparation can be even more challenging. To fit the green analytical chemistry principles, efforts should be made toward the elimination or reduction of the use of toxic reagents and solvents, minimization of energy consumption and increased operator safety. The simplest sample preparations are more appropriate for liquid biological matrices with little interfering compounds such as urine, plasma and oral fluid. The same does not usually occur with complex matrices that require more laborious procedures. The present review discusses green analytical approaches for the analyses of drugs of abuse in complex biological matrices, such as whole blood, breast milk, meconium and hair.

Investigation on the relationship between violent death, cocaine abuse and single nucleotide polymorphisms / Estudo da relação entre morte violenta, uso de cocaína e polimorfismos de nucleotídeo único

Violence is a dreadful phenomenon spread throughout the world, resulting in unfortunate events that can ultimately cause death. It is known that some countries play a much worrying role in this scenario than others. Brazil is one of them. The present study has focused on identifying the use of cocaine within 105 postmortem cases arriving at the Institute of Legal Medicine of São Paulo (IML-SP) through analytic toxicological methods and latter applying genetic testing to see whether the presence of certain single nucleotide polymorphisms (SNPs) is more predominant within users rather than non-users, which would help to better understand one's susceptibility to abuse the drug. Both blood and hair samples have been analysed through ultra-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) in order to distinguish between recent or chronic cocaine use among violent individuals whose violence has ultimately leaded to their death. Two dilute-and-shoot methods have been validated and used for this purpose, and the final residue was analysed through the UPLC-ESIMS/ MS system. From the 105 postmortem cases, a rather high proportion of cocaine and its metabolites was found. A chronic use of the drug was denoted in 53% of the cases, which were positive for cocaine and benzoylecgonine, followed by 43% for norcocaine, 40% for cocaethylene and 13% for anhydroecgonine methyl ester, in hair. As for blood, reflecting the use of cocaine prior to death, 51% of the cases have shown to be positive for benzoylecgonine, followed by 41% for cocaine, 23% for cocaethylene and 20% for norcocaine. These findings suggest a probable association between the use of the drug and risky/violent behaviours. Genetic wise, a significant difference has been observed for SNP rs4263329 from the BCHE gene in its dominant model, with higher frequencies of the genotypes A/G and G/G seen in cocaine users rather than non-users (OR=8.91; 95%CI=1.58-50.21; ρ=0.01). Likewise, also SNP rs6280 from the DRD3> gene presented a significant association in both its additive and dominant model, suggesting that the C allele may be playing a role in cocaine use as both genotypes T/C and C/C were significantly more frequent in users than non-users. This association was not lost when adjusted for covariants using logistic regression (OR=4.96; 95%CI=1.07; ρ=0.04). Finally, a statistically significant association (ρ = 0.003) was also encountered among individuals with both A/G and G/G genotypes within SNP rs4263329 and the use of cocaine HCl (f(A/G+G/G)=44.7%) versus crack-cocaine (f(A/G+G/G)=7.7%) and nonusers (f(A/G+G/G)=16.2%). In conclusion, this study has found significant associations within two SNPs related to cocaine use, however, due to several inherent limitations, these must be confirmed by further studies with a higher number of subjects and within a more controlled setting. Definite assumptions may not be made at this point and future researches are to be conducted

A violência é um fenômeno aterrador espalhado por todo o mundo, resultando em eventos que podem, em última instância, causar a morte. Sabe-se que, em alguns países esse cenário é mais preocupante que em outros. O Brasil é um deles. O presente estudo teve como objetivo identificar o uso de cocaína em 105 casos postmortem provenientes do Instituto de Medicina Legal de São Paulo (IML-SP) por meio de métodos toxicológicos analíticos e posterior aplicação de testes genéticos para verificar se a presença de determinados polimorfismos de nucleotídeo único (SNPs) é mais predominante dentro dos usuários do que dos não usuários, o que explicaria uma possível suscetibilidade de um indivíduo ao abuso da droga. Amostras de sangue e cabelo foram analisadas através de cromatografia líquida de ultra-eficiência acoplada a espectrometria de massas e ionização por electrospray (UPLC-ESI-MS/MS) para distinguir entre uso recente ou crônico de cocaína entre indivíduos violentos cuja violência levou à sua morte. Para tal, dois métodos de extração baseados na técnica de "dilute-and-shoot" foram validados e utilizados para esse fim, e o resíduo final foi analisado através de um sistema UPLC-ESI-MS/MS. Dos 105 casos postmortem, foi encontrada uma proporção significativa de cocaína e seus produtos de biotransformação. O uso crônico da droga foi denotado em 53% dos casos, sendo estes positivos para cocaína e benzoilecgonina, seguidos de 43% para norcocaína, 40% para cocaetileno e 13% para anidroecgonina metil éster, no cabelo. Quanto ao sangue, refletindo o uso de cocaína antes da morte, 51% dos casos mostraram-se positivos para benzoilecgonina, seguido de 41% para cocaína, 23% para cocaetileno e 20% para norcocaína. Esses dados corroboram a hipótese provável da relação entre o uso da droga e comportamentos de risco/violentos. Quanto à genética, uma diferença significativa foi observada para o SNP rs4263329 do gene BCHE em seu modelo dominante, com maiores frequências dos genótipos A/G e G/G vistos em usuários de cocaína ao contrário de não usuários (OR=8,91; 95%IC=1,58-50,21; ρ=0,01). Da mesma forma, também o SNP rs6280 do gene DRD3 apresentou uma associação significativa tanto no seu modelo aditivo quanto dominante, sugerindo que o alelo C pode estar desempenhando um papel no uso de cocaína, pois ambos os genótipos T/C e C/C foram significativamente mais frequentes nos usuários do que não usuários. Essa associação não foi perdida quando ajustada para co-variáveis usando regressão logística (OR=4,96; 95%IC=1,07; ρ=0,04). Finalmente, uma associação estatisticamente significativa (ρ=0,003) também foi encontrada entre indivíduos com ambos os genótipos A/G e G/G dentro do SNP rs4263329 e o uso de cocaína HCl (f(A/G + G/G)=44,7%) versus crack (f(A/G + G/G)=7,7%) e não usuários (f(A/G + G/G)=16,2%). Em conclusão, este estudo encontrou associações significativas em dois SNPs relacionados ao uso de cocaína, no entanto, devido a várias limitações inerentes, estas devem ser confirmadas por mais estudos com um maior número de indivíduos e dentro de um cenário mais controlado. Hipóteses definitivas não poderão ser feitas neste momento e futuras pesquisas devem ser conduzidas