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- Evaluation of the Cytotoxicity of Ayahuasca BeveragesPublication . Simão, Ana Y.; Gonçalves, Joana; Gradillas, Ana; García, Antonia; Restolho, José; Fernández, Nicolás; Rodilla, Jesus M.; Barroso, Mário; Duarte, Ana Paula; Cristóvão, Ana C.; Gallardo, EugeniaAyahuasca is a beverage consumed at shamanic ceremonies and currently has gained popularity on recreational scenarios. It contains beta-carboline alkaloids and N,N-dimethyltryptamine, which possesses hallucinogenic effects. Only a few studies have elicited the psychoactive effects and the dose of such compounds on neurological dopaminergic cells or animals. In this work, we aimed to study the cytotoxic effects of these compounds present in ayahuasca beverages and on five different teas (Banisteriopsis caapi, Psychotria viridis, Peganum harmala, Mimosa tenuiflora and Dc Ab (commercial name)) preparations on dopaminergic immortalized cell lines. Moreover, a characterization of the derivative alkaloids was also performed. All the extracts were characterized by chromatographic systems and the effect of those compounds in cell viability and total protein levels were analyzed in N27 dopaminergic neurons cell line. This is the first article where cytotoxicity of ayahuasca tea is studied on neurological dopaminergic cells. Overall, results showed that both cell viability and protein contents decreased when cells were exposed to the individual compounds, as well as to the teas and to the two mixtures based on the traditional ayahuasca beverages. View Full-Tex
- Deteção e quantificação dos principais constituintes de Ayahuasca com recurso a QuEChERS e HPLC-DADPublication . Gonçalves, Joana; Rosado, Tiago; Barroso, Mário; Restolho, José; Luís, Ângelo; Duarte, Ana Paula; Gallardo, EugeniaA Ayahuasca é uma bebida psicoativa originalmente consumida na bacia amazónica. Esta decocção possui N,N-dimetiltriptamina (DMT), responsável pelos efeitos psicoativos, e alcaloides β-carbolínicos, que inibem a Monoamina Oxidase A (MAO-A), permitindo que o DMT aceda ao sistema nervoso central. A legislação que regula o consumo de Ayahuasca é pouco rigorosa, uma vez que esta também é consumida em contexto religioso, levando muitas vezes a um consumo não controlado. Este estudo tem como objetivo a quantificação simultânea de DMT, tetrahidroharmina (THH), harmina, harmalina, harmol e harmalol em amostras de Ayahuasca. Assim, inicialmente foram comparadas três técnicas de extração: microextração líquido-líquido dispersiva (DLLME), microextração em seringa empacotada (MEPS) e Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS). Esta última mostrou ser a mais promissora, portanto foi submetida a um processo de otimização, onde foram testadas três variáveis (volume de solvente extrator, quantidade de amina primária secundária (PSA) e tempo de vórtex). As condições finais utilizadas foram 500 μL de solvente extrator, 85 mg de PSA e 4 segundos de agitação por vórtex. Posteriormente, foi utilizada cromatografia líquida de alta eficiência acoplada a um detetor de diode array (HPLC-DAD). O método analítico foi validado, apresentando linearidade entre 10 e 0,16 μg/mL (1 e 0,016 μg/mL para o DMT) com coeficientes de determinação superiores a 0,99. Os limites de deteção e quantificação foram de 0,16 μg/mL para as β-carbolinas e 0,016 μg/mL para o DMT e as eficiências de extração variaram entre 60 e 88 %. Este é o primeiro estudo que quantifica os principais constituintes de ayahuasca recorrendo à técnica de QuEChERS.
- Comparative study of sample preparation procedures to determine the main compounds in ayahuasca beverages by QuEChERS and high‐performance liquid chromatography analysisPublication . Gonçalves, Joana; Rosado, Tiago; Barroso, Mário; Restolho, José; Fernández, Nicolás; Luís, Ângelo; Gallardo, Eugenia; Duarte, Ana PaulaIntroduction : Ayahuasca is a psychoactive drink originally consumed by indigenous people of the Amazon. The lack of regulation of this drink leads to uncontrolled consumption, and it is often consumed in religious contexts. Objective :The aim of this work is to compare three miniaturised extraction techniques for extracting the main ayahuasca compounds from beverages. Methodology:Three sample pretreatment techniques were evaluated (dispersive liquid–liquid microextraction [DLLME], microextraction by packed sorbent [MEPS] and QuEChERS [Quick, Easy, Cheap, Effective, Rugged and Safe]) for the simultaneous extraction of N,N-dimethyltryptamine (DMT), tetrahydroharmine (THH), harmine, harmaline, harmol and harmalol from ayahuasca beverage samples. Then, the most promising technique (QuEChERS) was chosen to pre-concentrate the analytes, subsequently detected by high-performance liquid chromatography coupled to a diode array detector (HPLC-DAD). Results: The procedure was optimised, with the final conditions being 500 μL of extractor solvent, 85 mg of primary secondary amine (PSA) and 4 s of vortexing. The analytical method was validated, showing to be linear between 0.16 and 10 μg/mL for β-carbolines and between 0.016 and 1 μg/mL for DMT, with coefficients of determination (R2) between 0.9968 and 0.9993. The limit of detection (LOD) and lower limit of quantification (LLOQ) were 0.16 μg/mL for all compounds, except for DMT (0.016 μg/mL) and extraction efficiencies varied between 60.2% and 88.0%. Conclusion: The analytical methodology proved to be accurate and precise, with good linearity, LODs and LLOQs. This method has been fully validated and successfully applied to ayahuasca beverage samples.