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Background Oral fluid is a widely studied matrix able to isolate the primary Cannabis constituent THC, facilitating its detection via mass spectrometry, and in most cases link these findings to recent drug use. As an alternative to liquid oral fluid, dried oral fluid spots (DOFS) is a simple and a low-cost sampling technique. It has shown improved stability compared to liquid samples, allowing for the possibility to preserve the specimens under various temperature and humidity conditions. The sampling strategy is straightforward and involves the application of a small quantity of oral fluid aliquot to a paper substrate that is set to air dry allowing for on-site collection at a large-scale demand. The goal of this study is to study THC and CBD extraction from DOFS, applying a previous established protocol for a LC–MS/MS qualitative method validation. Although other drugs of abuse have been included in DOFS methods, this is the first method validation including cannabinoids. An alternative oral fluid extraction method (WAX-S tips) is demonstrated to improve the recovery of the analytes. Methods A pool of blank oral fluid was used to prepare THC and CBD spiked DOFS samples for method validation and application. Spiked oral fluid was used to demonstrate WAX-S tips THC and CBD extraction. All samples were analyzed on a LC–MS/MS instrument. Results The qualitative method validation for THC and CBD confirmation in DOFS included method selectivity, matrix effects ( Conclusions THC and CBD extraction from DOFS showed low recoveries. However, the LC–MS/MS qualitative confirmation of THC and CBD in DOFS could improve cannabinoids screening in oral fluid, as it shows adequate LOD and stability over time. This method has potential for assisting the screening of drivers under possible drug influence by facilitating sample transportation and temporary storage in dried spot form. Additional research is suggested for WAX-S tips extraction and quantitative method validation.

A Ciência Forense é uma área multidisciplinar, que usa do conhecimento científico para a investigação de questões penais e cíveis. Dentro de suas mais diversas áreas, a Toxicologia e a Documentoscopia Forense necessitam do auxílio de métodos analíticos, como a cromatografia, a espectrometria de massas e a análise multivariada, para a análise de compostos diversos. Em Toxicologia Forense, novos métodos para a detecção das drogas de abuso anfetamina, metanfetamina, cocaína, cetamina, mitraginina e canabinoides (THC e CBD) foram desenvolvidos utilizando coleta de fluido oral em dispositivos de dried oral fluid spots (DOFS) em papel filtro, seguido de extração líquido-líquido e análise por LC-MS/MS. Enquanto que para cinco drogas de abuso foi realizada a validação de um método quantitativo, para os canabinoides foram encontradas dificuldades na extração do papel filtro, e foi proposto um método qualitativo. Ainda, foi realizada uma revisão bibliográfica a respeito dos métodos quantitativos para a análise de canabinoides em fluido oral. Na área da Documentoscopia Forense, uma nova metodologia não-destrutiva, de baixo custo e de fácil acesso, foi desenvolvida para a diferenciação entre marcas de canetas esferográficas azuis. Esse estudo envolveu a captura de imagens utilizando o aplicativo para celulares Photometrix PRO®, com posterior análise estatística multivariada, utilizando PCA, HCA e PLS-DA. Este método demonstra também a necessidade de que Peritos em Documentoscopia tenham conhecimentos de análise multivariada, para proceder com a correta interpretação dos resultados. Para valorizar o conhecimento das metodologias analíticas na análise de tintas de canetas, foram apresentadas duas revisões bibliográficas. A primeira objetivou a busca sistemática dos métodos analíticos já estudados para a diferenciação e para a datação de tintas de canetas esferográficas azuis e pretas. Foram encontrados diversos métodos cromatográficos, de espectroscopia, de espectrometria de massas e de análise multivariada. A segunda revisão buscou a abordagem dessas metodologias voltada para Peritos Brasileiros. Essa Tese de Doutorado destaca ainda a multidisciplinariedade das Ciências Forenses, bem como a necessidade da formação de recursos humanos adequada na Perícia Brasileira. Forensic Science is a multidisciplinary area, which uses scientific knowledge in judicial questions investigations. Among its many subareas, Forensic Toxicology and Documentoscopy demand the support of analytical methodologies for several different compounds’ evaluation, such as chromatographic methods, mass spectrometry and multivariate analysis. Inside Forensic Toxicology scope, new analytical methods were developed for different drugs of abuse detection (amphetamine, methamphetamine, cocaine, ketamine, mitragynine and cannabinoids THC and CBD), using oral fluid collection applied into dried oral fluid spots (DOFS), followed by liquid-liquid extraction and LC-MS/MS analysis. A quantitative method validation was proposed for all drugs, except for cannabinoids, which have shown difficulties in the filter paper extraction process. Therefore, a qualitative method validation is proposed for THC and CBD detection in DOFS. Considering this, a systematic review was conducted concerning cannabinoids quantitative analysis in oral fluid. In Forensic Documentoscopy, it was developed a new non-destructive methodology, of low-cost and easy access, to differentiate between blue ballpoint pen brands. This study used image capturing with Photometrix PRO®, an app for smartphones, followed by multivariate analysis (PCA, HCA and PLS-DA). This method emphasizes that Documentoscopy Experts need to understand multivariate analysis, so that they can proceed with adequate results interpretation. To highlight the need for analytical methodologies knowledge in pen inks analysis, two literature reviews are presented. The first one aimed to perform a systematic search for studied analytical methods used to differentiate or to date blue and black ballpoint pen inks. Several chromatographic, spectroscopy, mass spectrometry and multivariate analysis methods were found. Thus, the second review aimed to highlight the importance of those methods approaches for Brazilian Experts. Besides the technological advance achieved with new analytical methodologies, this Doctorate Thesis highlights Forensic Science as a multidisciplinary field, also reinforcing the need for adequate human resources in Brazil.