Dissertations / Theses on the topic 'Salvinorin A'

Thesis (M.S.)--Boston University PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you.
Salvia divinorum (S. divinorum) is a psychoactive plant from the Lamiaceae (mint) family originating in the Oaxaca region of Mexico. The plant's psychoactive compound, salvinorin A (sal A) has been found to be unique to the microscopic glands of S. divinorum and exhibit powerful psychoactive properties similar to lysergic acid diethylamide (LSD) at concentrations as low as 200 µg. A similar compound, salvinorin B (sal B), is also found in the glands of S. divinorum and does not have psychoactive properties, although is considered a precursor to sal A. The plant is traditionally used by the Mazatec Indians for divination and healing rituals but has since been brought to the United States (U.S.) where it has become a popular drug of abuse. The plant is abused like other psychoactive plants such as mescaline, psilocybin mushrooms, and cannabis, and is currently not regulated by federal laws. In anticipation of legislation regulating S. divinorum, sal A, and sal B, forensic analysis methods must be improved to allow for the rapid and accurate identification of plant material, such as fresh or dried leaves, and the active compounds. Currently, analysis of S. divinorum relies on the identification of sal A from extracts of leaf material, a time consuming process requiring sophisticated instrumentation. The development of preliminary tests such as a macroscopic/microscopic analysis or colorimetric spot test would allow for the rapid analysis of S. divinorum plant material, sal A, and sal B, and would allow for elimination of non- S. divinorum material from further time-consuming analyses. This research focused on examining the botanical features of S. divinorum plant material at the macroscopic and microscopic level for the ability to discriminate S. divinorum from other plants in the same taxonomic family and common household herbs. Additionally, a colorimetric test utilizing in-house preparations of the Ehrlich's reagent was evaluated for its potential to be used as a presumptive test for S. divinorum, sal A, and sal B. [TRUNCATED]

Salvinorin A [(2S,4aR,6aR,7R,9S,10aS,10bR)-9-(acetyloxy)-2-(3-furanyl)-dodecahydro-6a,10b-dimethyl-4,10-dioxo-2H-naptho[2,1-c]pyran-7-carboxylic acid methyl ester] is a trans-neoclerodane diterpene from the leaves of the hallucinogenic Mexican sage Salvia divinorum and has been identified as the principal psychoactive component in this plant of traditional spiritual importance. Salvinorin A is the most potent naturally occurring hallucinogen found so far and is reported to act selectively as a ƒÛ-opioid receptor agonist. Synthetic modification of the natural product has contributed to a number of proposed pharmacophores to identify the key structural features necessary for biological activity and a direct strategy for the asymmetric synthesis of the natural product is desirable since it allows access to a more diverse range of analogues. An ambitious retrosynthetic study of salvinorin A indicated the C(3)-heterosubstituted furan as an appropriate starting material for a Diels-Alder approach towards the ketone ring of the natural product. An expedient and high yielding methodology for the preparation of 3-furylamines is described, allowing the flexible introduction of alkyl substituents in the C(5) position. Optically pure ephedrine isomers have been explored as chiral amine auxiliaries and have been successfully attached as 3-furylamine substituents using the general methodology described. The 3-furylamines are electron rich dienes that are highly reactive towards Diels-Alder cycloaddition reactions with methyl acrylate. Diastereoisomers of the 7-oxanorbornane species methyl 1-methyl-5-oxo-7-oxa-bicyclo[2.2.1]heptane-2-carboxylate were prepared as new compounds from the hydrolysis of Diels-Alder cycloadducts and are functionalised bicyclic intermediates to access the ketone of the natural product. Diels-Alder reactions between the non-racemic (2S)-ephedrine-derived furans and methyl acrylate gave spiro-oxazolidine adducts that underwent hydrolysis to give the desired ketone. X-ray crystallography data for the derivatised cycloadduct established diastereoselectivity in favor of the (1S,4S)-enantiomer, as desired for the asymmetric natural product synthesis. A procedure for the ether cleavage of methyl 1-methyl-5-oxo-7-oxa-bicyclo[2.2.1]heptane-2-carboxylate was required to access the convergent precursor methyl 5-acetoxy-2-methyl-4-oxocyclohex-2-enecarboxylate. Successful C-O cleavage was achieved using Lewis-acid catalysis with BBr3 followed by mixing with the hindered base 2,4,6-collidine to yield methyl 5-hydroxy-2-methyl-4-oxocyclohex-2-enecarboxylate albeit only at high dilution. Acetylation proceeded in excellent yield in the same reaction vessel to give methyl 1-methyl-5-oxo-7-oxa-bicyclo[2.2.1]heptane-2-carboxylate in excellent yield. The devised synthetic pathway is shown to successfully construct the ketone ring of salvinorin A and stereoselectivity for the (1S,4S)-enantiomer can be achieved using the ephedrine derived furans as desired for the asymmetric natural product synthesis. The ƒÔ-lactone ring 6-(furan-3-yl)-5,6-dihydro-4-methyl-3-vinylpyran-2-one was derived from rudimentary precursors as a convergent reagent to introduce the lactone ring of salvinorin A. A short synthesis for the racemic compound is described starting from the aldol reaction between 3-furaldehyde and acetone to give the 3-furfurol, 4-(furan-3-yl)-4-hydroxybutan-2-one in quantitative yield. The 3-furfurol was reacted to form the ƒÑ-bromovinyl ester, 1-(furan-3-yl)-3-oxobutyl 2-bromobut-3-enoate using a deconjugation/esterification protocol with 2-bromobut-3-enoyl chloride. Intramolecular ring closure to the ƒÔ-lactone was achieved using a Reformatsky reaction and dehydration under acidic conditions yielded the racemic convergent precursor 6-(furan-3-yl)-5,6-dihydro-4-methyl-3-vinylpyran-2-one in high yield. A possible strategy for joining the ketone and lactone fragments for the total synthesis of salvinorin A is proposed.

Salvinorin A is a non-nitrogenous, selective kappa opioid receptor agonist with potent hallucinogenic properties. Because Salvinorin A has no basic nitrogen, it does not readily adhere to the “message-address” concept of selectivity for the opioid receptors. Therefore, a better understanding of how salvinorin A and its analogs interact with the kappa opioid receptor may shed some light on how salvinorin A obtains its potency and selectivity. The structure-affinity relationships (SAFIR) of salvinorin A and its analogs along with a discussion of the selectivity of the opioid receptors, is presented. A fragment of salvinorin A, methyl-3-acetoxy-4-oxocyclohexanecarboxylate, was synthesized to determine if the B, C and D rings are or are not necessary for binding to the opioid receptors. The fragment was found not to bind to the kappa, delta or mu receptor which reinforces the importance of the B, C and D rings in the binding of salvinorin A to the kappa opioid receptor. Homology models of the kappa, delta and mu opioid receptors were constructed based on inactive bovine rhodopsin, light-activated bovine rhodopsin and the human beta-2 adrenergic receptors. The program MODELLER was also used to construct the kappa opioid receptor. Two comparative molecular field analysis (CoMFA) studies are then presented which compared three different types of alignment methods. The alignment methods employed included a receptor-docked alignment in which the salvinorin A analogs were docked into a model of the kappa opioid receptor using the program GOLD. The docked poses for this alignment were chosen based on their similarity to our postulated model of salvinorin A in the kappa opioid receptor. In our model the furan oxygen forms hydrogen bonds with Q115(2.60) and Y320(7.43), the methoxy oxygen of the C-4 position ester group may form a hydrogen bond with Y312(7.35) and the methyl group of the C-2 position acetoxy moiety forms a hydrophobic interaction with Y313(7.36). These interactions are consistent with mutagenesis studies. The other alignment methods employed were a FlexS alignment and a realignment of the receptor-docked poses using the Fit Atoms function within SYBYL. Only the receptor-docked alignment method resulted in robust and predictive CoMFA models which indicates that the analogs may bind to the kappa opioid receptor in a similar but non-identical way. In addition, information from the CoMFA models based on the receptor-docked alignment led to a postulated binding mode for a set of amine analogs of salvinorin A which were not part of the original data set. Docking studies have the positively charged C-2 position amine group interacting with E209(XL2.49) while the furan oxygen and C-4 position ester group interacts with the same residues as in our model of salvinorin A in the kappa opioid receptor. The studies presented here not only support our postulated model of salvinorin A binding to the kappa opioid receptor but may also explain the trend of the beta epimers of the amine analogs to have a higher affinity than the corresponding alpha epimers. Site-directed mutagenesis studies could provide data to support or refute the postulated models of the amines docked in the kappa opioid receptor presented here.

Gold was considered for a long time to be an inert metal and was only in 1986 that the first homogeneous gold-catalyzed transformation was reported. In our laboratory, we isolated a surprisingly stable vinyl complex that resulted from an unexpected 1,2-silyl migration while working on a gold(I)-catalyzed reaction for the synthesis of polyprenylated polycyclic acylphloroglucinols (PPAPs). We herein report the isolation of a variety of organogold species where we could control the silyl migration based on the nature of the silyl group installed on the terminal alkyne. Silyl groups bearing an aromatic ring inhibited the silyl migration while the aliphatic silyl group afforded the 1,2-silyl migrated adduct. After mechanistic investigation of this intriguing migration, we believe that this process goes through a relatively rare gold vinylidene intermediate. More than 15 organogold complexes were isolated in good yield and characterized by x-ray crystallography. Investigation of their reactivity led to the formation of C(sp3)-C(sp2) bonds using electrophilic reagents without the use of Pd-based catalysts. We have also developed a new gold(I)-catalyzed dehydro Diels-Alder reaction using a simple monocyclic silyl enol ether. This methodology proceeds effectively with a wide scope by the use of [JackiephosAu(NCMe)]SbF6 in toluene. This methodology was then applied to the synthesis of magellanine, an architecturally complexed angular natural product isolated in 1976 from the club moss Lycopodium Magellanicum. The key step precursor was rapidly constructed via a Mitsunobu/Diels-Alder reaction that generated the requisite carboxaldehyde. The dehydro Diels-Alder reaction afforded the molecular skeleton of magellanine diastereoselectively in 91% yield. The synthesis was successfully accomplished in 11 steps demonstrating the ability of the gold(I) salt to rapidly construct complex molecules. Since the discovery of salvinorin A, a lot of efforts were exerted in order to optimize the biological activity for treatment of central nervous system disorders. Development of a new synthetic routes to salvinorins are essential to afford novel functionalized analogues. The decalin framework of salvinorin A was assembled with a Diels-Alder reaction with Et2AlCl followed by a gold(I)-catalyzed 6-endo-dig carbocyclization with [JohnphosAu(NCMe)]SbF6. Further functionalization afforded an elaborated intermediate which possesses the correct stereochemistry of the natural product. Following these promising results, efforts are currently in progress for the completion of the total synthesis.

While there are a variety of therapeutics that interact with the opioid receptor system, they are not without side effects; including constipation, dysphoria and respiratory depression. A better understanding of the opioid receptor system may yield therapeutic agents with a limited side effect profile. The neoclerodane diterpene, salvinorin A, appears to interact at opioid receptors through a unique mode of action. A better understanding of its interactions with opioid receptors will yield valuable information about the opioid system. In order to probe further how salvinorin A interacts at opioid receptors, a series of novel analogues modified at the C-2 and furan ring were synthesized and evaluated for their ability to interact at opioid receptors. Synthetic methods were identified to modulate the furan ring, including the synthesis of Diels-Alder cycloadducts and phenyl rings derived from a reductive elimination. The cycloadducts are one of the first reported examples of Diels-Alder chemistry being applied to modify a neoclerodane while the phenyl ring analogues are the first to have aromatic rings directly off the salvinorin A core. C-2 sulfonate analogues were found to interact differently then their ester counterparts at opioid receptors while several of the cycloadduct analogues maintained affinity and efficacy demonstrating the furan is not required for opioid receptor activity. These findings demonstrate that salvinorin A is amenable for chemical modification, illustrating its potential as a novel scaffold for the development of opioid ligands.

In this study, we have addressed the serotonergic and the opioid system within the CNS. Both systems are of outmost importance in the etiology of disease states, especially mental disorders. In our investigation of the serotonergic system, we have synthesized novel enantiomerically pure 6-aryl-3-amino- and 8-aryl-3-aminochromans as ligands for the 5-HT7 receptor. One reason for the lack of understanding of the physiological functionality of the serotonin 5-HT7 receptor, the most recently discovered member of the serotonin receptor family, is the absence of partial agonists and agonists. In this series, we have identified partial agonists with more than189 fold selectivity over the 5-HT1A receptor and one agonist with 29 fold greater selectivity over the serotonin 5-HT1A receptor. Thus the present series constitutes a starting point for developing highly selective ligands for the 5-HT7 receptor. In our investigation of the opioid system, our focus has been on the natural product salvinorin A, which is a highly selective kappa opioid receptor agonist. In the total synthesis of salvinorin A, we have accomplished the synthesis of a key intermediate, 6-(3-furyl)-4-methyl-5,6-dihydro-pyran-2-one via ring closing metathesis. Furthermore, synthetic methodologies have been developed as a part of the total synthesis. Several lipases have been screeened for their ability to generate enantiomerically pure 1-(3-Furyl)-3-buten-1-ol via bio-catalyzed hydrolysis of the corresponding acetate. The lipase from Pseudomonas fluorescens was identified as having stereoselectivity high enough to generate a % ee value above 98%. We have also developed a route for the introduction of a hydroxyl functionality in the γ position of α,β-unsaturated cyclic ketones by the regioselective oxidation of 1-silyloxy-1,3-dienes using dimethyldioxirane. We have initiated the investigation of the pharmacophore responsible for the kappa opioid activity by synthesizing simplified analogues of salvinorin A. A synthetic route providing easy access to simplified analogues of salvinorin A have been established.

In this study, we have addressed the serotonergic and the opioid system within the CNS. Both systems are of outmost importance in the etiology of disease states, especially mental disorders.

In our investigation of the serotonergic system, we have synthesized novel enantiomerically pure 6-aryl-3-amino- and 8-aryl-3-aminochromans as ligands for the 5-HT₇ receptor. One reason for the lack of understanding of the physiological functionality of the serotonin 5-HT₇ receptor, the most recently discovered member of the serotonin receptor family, is the absence of partial agonists and agonists. In this series, we have identified partial agonists with more than189 fold selectivity over the 5-HT_1A receptor and one agonist with 29 fold greater selectivity over the serotonin 5-HT_1A receptor. Thus the present series constitutes a starting point for developing highly selective ligands for the 5-HT₇ receptor.

In our investigation of the opioid system, our focus has been on the natural product salvinorin A, which is a highly selective kappa opioid receptor agonist. In the total synthesis of salvinorin A, we have accomplished the synthesis of a key intermediate, 6-(3-furyl)-4-methyl-5,6-dihydro-pyran-2-one via ring closing metathesis. Furthermore, synthetic methodologies have been developed as a part of the total synthesis. Several lipases have been screeened for their ability to generate enantiomerically pure 1-(3-Furyl)-3-buten-1-ol via bio-catalyzed hydrolysis of the corresponding acetate. The lipase from Pseudomonas fluorescens was identified as having stereoselectivity high enough to generate a % ee value above 98%. We have also developed a route for the introduction of a hydroxyl functionality in the γ position of α,β-unsaturated cyclic ketones by the regioselective oxidation of 1-silyloxy-1,3-dienes using dimethyldioxirane. We have initiated the investigation of the pharmacophore responsible for the kappa opioid activity by synthesizing simplified analogues of salvinorin A. A synthetic route providing easy access to simplified analogues of salvinorin A have been established.

Los datos disponibles en la actualidad indican que los efectos psicoactivos de la salvinorina A, el principio activo de la planta Salvia divinorum, se deberían al efecto agonista ejercido por este compuesto sobre el receptor kappa opioide. Sin embargo, hasta la fecha no se había atribuido a la neurotransmisión opioide ningún papel relevante en la regulación de la percepción. Las intensas modificaciones sensoriales referidas por los usuarios de la Salvia divinorum y las observadas en el laboratorio tras administración de salvinorina A se encuentran muy alejadas de las meras alteraciones somático-disfóricas inducidas por otros agonistas kappa. Por el contrario, los efectos de la salvinorina A se asemejarían en gran medida a los inducidos por los alucinógenos clásicos. Sin embargo, estos últimos actúan a través de un mecanismo neuroquímico totalmente distinto. Sustancias como la psilocibina, la LSD o la dimetiltriptamina no interactúan con el sistema opioide si no que presentan actividad agonista sobre el receptor 2A de la serotonina. Los dos estudios experimentales que se presentan en esta memoria se diseñaron con el fin de esclarecer esta paradoja. En el primero se procedió a evaluar de forma exhaustiva el perfil de efectos subjetivos inducido por la salvinorina A, haciendo especial hincapié en las similitudes y diferencias con el de los alucinógenos serotoninérgicos. También se evaluaron los efectos de la salvinorina A sobre la interocepción, un aspecto poco estudiado hasta el momento, y que revela claves importantes sobre los cambios en la percepción mediados por la activación de los receptores kappa opioides. En el segundo estudio, mediante la utilización de dos antagonistas selectivos, se investigó la participación del receptor kappa opioide y del receptor 2A de la serotonina en la farmacología general de la salvinorina A. También se evaluó los efectos endocrinos y autonómicos de la salvinorina A, así como su farmacocinética. Los dos estudios de interacción farmacológica realizados demuestran la participación de la neurotransmisión opioidérgica, en lugar de serotoninérgica, en los efectos de la salvinorina A en los seres humanos. Estos resultados son consistentes con acciones agonistas de la salvinorina A en los receptores kappa opioides.
Currently available data indicate that the psychoactive effects of salvinorin A, the active ingredient of the plant Salvia divinorum, are due to the agonist effect exerted by this compound on the kappa opioid receptor. However, to date, no relevant role in the regulation of perception has been attributed to opioid neurotransmission. The intense sensory modifications referred by the users of Salvia divinorum and those observed in the laboratory after administration of salvinorin A are far from the mere somaticdysphoric alterations induced by other kappa agonists. On the contrary, the effects of salvinorin A would closely resemble those induced by classical hallucinogens. However, the latter act through a totally different neurochemical mechanism. Substances such as psilocybin, LSD or dimethyltryptamine do not interact with the opioid system but have agonist activity on the serotonin 2A receptor. The two experimental studies presented in this thesis were designed to clarify this paradox. In the first one, the profile of subjective effects induced by salvinorin A was thoroughly evaluated, with special emphasis on the similarities and differences with that of the serotonergic hallucinogens. The effects of salvinorin A on interoception have also been evaluated, an aspect that has been little studied so far and that reveals important clues about the changes in perception mediated by the activation of kappa opioid receptors. In the second study, through the use of two selective antagonists, the participation of the kappa opioid receptor and serotonin 2A receptor in the general pharmacology of salvinorin A was investigated. In the second study we also analyzed its endocrine and autonomic effects, as well as its pharmacokinetics. The two pharmacological interaction studies carried out demonstrate the participation of opioidergic neurotransmission, instead of serotonergic, in the effects of salvinorin A in humans. These results are consistent with salvinorin A agonist actions in the kappa opioid receptors.

La salvinorine A est un néoclérodane naturel possédant une forte affinité pour les récepteurs opioïdes kappa. Tout comme elle, de nombreux terpénoïdes bioactifs naturels, tels que l’acide hardwickiique, possèdent une chaîne latérale 2-(furan-3-yl)-éthyle en position 9. L’introduction de cette chaîne sur le squelette décaline est actuellement reportée dans la littérature en 3 à 8 étapes formelles et nécessitent souvent une dérivatisation préalable des autres fonctionnalités, diminuant ainsi la possibilité d’accéder à plusieurs analogues à partir d’un seul intermédiaire. L’objectif de ce travail a été de développer une méthode d’introduction efficace et diastéréosélective de ce motif à partir de la dicétone de Wieland-Miescher C(9)-méthylée (DWM). Ceci a été envisagé afin d’accéder à un intermédiaire commun peu fonctionnalisé pouvant ensuite être dérivatisé en de nombreux analogues naturels ou synthétiques. Pour cela, plusieurs stratégies ont été envisagées et testées mais l’introduction du motif désiré n’a pas été possible. L’une d’elle a tout de même permis de synthétiser de manière fortuite un nouveau composé, l’α-iodocétone de la DWM. Des essais de dérivatisation ont été effectués et sont toujours en cours de développement. Une stratégie d’aldolisation de Mukaiyama en présence de l’éther d’énol silylé de la DWM et de divers acétals a également été mise au point et permet d’accéder de manière diastéréosélective en 2 étapes à 36 nouveaux aldols simples, silylés, méthylés ou bien encore benzylés comportant différentes chaînes latérales en position C(9) avec des rendements compris entre 9 et 97%. Des fonctionnalisations ultérieures ont ensuite été entrepris sur certains intermédiaires dans l’optique de synthétiser deux sesquiterpénoïdes naturels, le (+)-auréol et la (+)-strongyline A
The natural neoclerodane salvinorin A exhibits a great affinity for kappa opioid receptors. Several other bioactive natural terpenoids, such as hardwickiic acid, are also bearing a 2-(furan-3-yl)-ethyl chain at position 9. The introduction of this chain is usually performed with 3 to 8 formal synthetic steps. Derivatization of other functionalities is often mandatory in order to meet the chemoselectivity requirements, and is preventing the possibility to access multiple analogues. The aim of this work was to develop an efficient and diastereoselective method to introduce this lateral chain at C(9) position of the C(9)-methylated Wieland-Miescher diketone (DWM) in order to access many natural or synthetic analogues from a common intermediate. Several strategies were designed, but the introduction of the ethyl-3-furyl chain remained unsuccessful. Nevertheless, the new α-iodoketone of the DWM was obtained by serendipity during this work. Experiments in order to derive this compound are still under investigation. A Mukaiyama aldol reaction strategy was set up and permit a diastereoselective access to 36 new aldols (simple, silylated, methylated or benzylated), bearing different lateral chains at C(9) position, with 9 to 97% yield. Further functionalization of two aldols were undertaken in order to synthesize two natural sesquiterpenoids, the (+)-aureol and the (+)-strongylin A

A presente dissertação está integrada na unidade curricular estágio do Mestrado Integrado em Ciências Farmacêuticas da Universidade da Beira Interior. Encontra-se dividida em dois capítulos, sendo que o primeiro corresponde à componente de investigação e o outro ao estágio curricular realizado na Farmácia Diamantino (Fundão). A Salvia divinorum (S. divinorum) é uma planta psicoativa original e tradicionalmente utilizada para fins medicinais e espirituais pelo povo Mazateca da região mexicana de Oaxaca. No entanto, de acordo com a Organização Mundial de Saúde (OMS), ao longo dos últimos anos tem sido cada vez mais consumida enquanto droga recreativa, uma vez que não é controlada na maioria dos países e é facilmente acessível na Internet e em smart shops. O principal constituinte da S. divinorum, a salvinorina A (SA), é o responsável pelos seus efeitos psicoativos. Trata-se de um composto não-nitrogenado com elevada afinidade para o recetor ?-opióide (KOR) e, contrariamente aos alucinogénios clássicos, não tem afinidade para o recetor de serotonina. O KOR está envolvido na perceção da dor, controlo motor e do humor e liga-se a compostos do tipo opióide no sistema nervoso central (SNC) e, por essa razão, pode constituir um importante alvo terapêutico no tratamento de várias desordens e patologias do SNC, incluindo a doença de Alzheimer e a esquizofrenia. Além disso, nos últimos anos, o consumo de S. divinorum enquanto droga de abuso tem também vindo a aumentar em todo o mundo e, embora haja alguma literatura sobre este tema, pouco se sabe sobre os efeitos desta planta e seus constituintes no organismo. Neste contexto, considerou-se de extrema importância clarificar o estado de arte relacionado com a S. divinorum e os seus constituintes e verificar os seus potenciais efeitos no metabolismo, quer em termos terapêuticos quer toxicológicos. Assim sendo, esta revisão sumariza e aprofunda o conhecimento atual sobre as propriedades biológicas da S. divinorum, destacando as descobertas clínicas, in vitro e in vivo descritas na literatura. Pretende-se ainda identificar e propor futuras linhas de investigação relativamente aos efeitos biológicos desta planta, particularmente ao nível do SNC. O estágio curricular em farmácia comunitária decorreu entre 26 de janeiro e 13 de junho de 2015 e foi realizado na Farmácia Diamantino, no Fundão. Este estágio permitiu-me conhecer o funcionamento de uma farmácia comunitária, bem como desempenhar todas as tarefas e responsabilidades associadas a um farmacêutico em contexto de farmácia comunitária.
This dissertation is included in the curricular unit of Estágio of the curricular plan of the master’s integrated degree in pharmaceutical sciences from the University of Beira Interior. It is divided into two main chapters, which the first is related to the research work and the second to the curricular internship held at Farmácia Diamantino (Fundão). Salvia divinorum (S. divinorum) is a psychoactive plant traditionally used for medicinal and spiritual purposes by the Mazatec people of Oaxaca, Mexico. Although, according to the World Health Organization (WHO), over the last years it has been also increasingly consumed as a recreational drug, as it is not controlled in most countries and it is easily available in the internet and smart shops. The main constituent of S. divinorum, Salvinorin A (SA), is the responsible for its psychoactive effects. SA is a unique non-nitrogenous compound with high affinity for kappa-opioid receptor (KOR) and, contrarily to the classical hallucinogens, it has no action on the serotonin receptor. KOR is involved in pain perception, motor control and mood and binds to opioid-type compounds in the central nervous system (CNS) and for that reason, it could constitute an important therapeutic target for treatment of several CNS disorders, including Alzheimer’s disease and schizophrenia. Furthermore, in the last few years, the consumption of S. divinorum as a drug of abuse has been increasing worldwide and, although some literature regarding this topic, little is known about S. divinorum and its effects on organisms. In this context, we found of utmost importance to clarify the state of the art related to S. divinorum and its constituents and to ascertain the potential properties and effects in metabolism, both as a therapeutic and a toxic agent. This review summarizes and updates the current knowledge about the biological properties of S. divinorum, highlighting the clinical, in vitro and in vivo findings, and intends to identify and to propose future directions for further research regarding the biological effects of this plant, particularly in CNS. The curricular internship in community pharmacy occurred between january 26th and june 13th of 2015 at the Farmácia Diamantino, located in Fundão. This internship allowed me to know the functioning of a community pharmacy, as well as to develop the tasks and to have the responsibilities of a pharmacist in a community pharmacy context.

碩士
高雄醫學大學
藥學研究所
100
Recently, use of recreational drugs has become a new trend among young drug users. Besides the well-known drugs such as marijuana, amphetamine, 3,4-methylenedioxymethamphetamine (MDMA) and ketamine, Salvia divinorum (Lamiaceae), which contains a hallucinogenic ingredient, salvinorin A, has been abused as a new phenomenon among young drug users. The mechanism of salvinorin A is a selective agonist at κ-opioid receptors and distinct from the classical hallucinogens. Hallucinatory effect of salvinorin A in Salvia divinorum is similar to marijuana, LSD and ketamine. In the recent years, use of Salvia divinorum has increased in the many countries. The prevalence of Salvia divinorum use in young drug users is higher than other recreational drugs. Salvia divinorum is becoming a serious concern for its the potential harm. Currently, dry leaves of Salvia divinorum and its related “concentrated extract” products have been sold through the internet in Taiwan. Salvia divinorum may become a recreational drug in Taiwan, because it has not been regulated. In this study, we used high performance liquid chromatography (HPLC) to detect whether salvinorin A was present in substantial amounts among Salvia divinorum and endemic species of Salvia in Taiwan. The dry leaves of Salvia divinorum and endemic species of Salvia were extracted with methanol and examined on a C-18 column by isocratic elution with mobile phase of acetonitrile : water (35:65, v/v) at flow-rate of 1.5 mL/min and UV detection. Results from our study indicated that although salvinorin A was detected in Salvia divinorum, it was not found in endemic Salvia species of Taiwan under the collected conditions. Therefore, the endemic species of Salvia in Taiwan may not possess hallucinogenic potential. According to three major indicators for control of addictive drugs, i.e., addiction potential, abuse potential and liability of social harms, evaluation of the potential harms indicated that the effect of Salvia divinorum was similar to other hallucinogenic drug. Based on the results, we suggested that Salvia divinorum and its related “concentrated extract” products may need close scrutiny in Taiwan.

As novas substâncias psicoativas (NPS) são, segundo a Observatório Europeu da Droga e da Toxicodependência (EMCDDA), todo e qualquer novo estupefaciente ou nova droga psicotrópica na sua forma pura, ou por preparar, que não é controlada pela Convenção Única das Nações Unidas de 1961 sobre os estupefacientes nem pela Convenção das Nações Unidas de 1971, mas que possa representar uma ameaça para a saúde pública comparativamente às substâncias listadas nessas convenções. Pelo seu crescente consumo a nível mundial e elevado desconhecimento do seus efeitos no organismo, as NPS têm-se tornado cada vez mais uma questão preocupante e, em 2012, foram adotadas medidas preventivas em Portugal relativamente às NPS, sendo que uma das substâncias que passou a integrar esta lista foi a Salvia divinorum (S. divinorum) e o seu componente bioativo, a salvinorina A (SA). A S. divinorum é uma planta que apresenta propriedades alucinogénias a qual, até meados dos anos 60, era utilizada unicamente pelos Mazatecas (região de Oaxaca, México). A substância nela presente que lhe confere estas propriedades, e que, por isso, tem despertado um grande interesse junto da comunidade científica, é o diterpeno neoclerodano SA. A SA é distinta de todos os outros alucinogénios conhecidos por possuir um mecanismo de ação único, é um agonista seletivo dos recetores kappa opióides (KOR). Apesar do alvo principal da S. divinorum, e da SA, ser o sistema nervoso central (SNC), até a data, ainda não foram pesquisados os seus potenciais efeitos a nível periférico, em particular a nível hepático, o órgão responsável pelo metabolismo de grande parte de compostos endógenos e xenobióticos. Assim, com este estudo pretendeu-se determinar in vitro a citotoxicidade e analisar a expressão relativa de certos genes relevantes no organismo resultantes da incubação de células hepáticas (Hep G2 e WRL-68) com várias concentrações de SA durante vários períodos de tempo. Os genes estudados foram: apolipoproteina B100 (Apo B100); carboxilesterase 1 (Ces 1); citocromo C (Cit. C); citocromo P450 1A2 (CYP450 1A2); citocromo P450 2D6 (CYP450 2D6); citocromo P450 3A4; glucuronosiltranferase 1A1 (UGT 1A1). Os resultados obtidos mostraram que, de modo geral, 50 µM de SA são citotóxicos para ambas as linhas celulares, sendo que 10 µM apenas mostraram diminuir a viabilidade celular das células Hep G2 após 72 h de incubação indicando que, aparentemente, as células Hep G2 são mais sensíveis à SA, comparando com as WRL-68. Pelo contrário, 0,1 e 1 µM não promoveram qualquer alteração na viabilidade celular relativa em ambas as linhas celulares. Relativamente ao efeito da exposição de células Hep G2 e WRL-68 a 1 µM de SA durante vários períodos de incubação (12, 24 e 72 h) verificou-se que, no geral, a SA tem impactos em termos de expressão relativa do mRNA de vários genes, de modo dependente quer do tempo de incubação quer da linha celular. Além disso, não foi detetada qualquer expressão do gene Ces 1 nas células WRL-68, indicando que o mesmo não é expresso nesta linha celular. Em suma, a SA aparenta ter um perfil toxicológico seguro porém em termos de regulação da expressão de genes importa salientar que, de um modo geral, pode exercer efeitos positivos ou negativos, consoante o gene em análise, o tempo de incubação com o composto e o tipo celular. Assim, e por se tratar do primeiro estudo que avaliou o potencial papel da SA na expressão hepática de certos genes, são necessárias investigações futuras para confirmar que os efeitos apresentados neste estudo são observados in vivo e diretamente devidos ao consumo de SA.
According to European Monitoring Centre for Drugs and Drug Addiction (EMCDDA), the new psychoactive substances (NPS) are any new narcotic or psychotropic drug, in its pure form, or in preparation, that is uncontrolled by the United Nations Drug Convention of 1961 or 1971, but which may represent a public health threat comparable to that posed by substances listed in these conventions. As its consumption is growing worldwide and its effects in organism still largely unknown, NPS had become increasingly a matter of concern and, in 2012, Portugal adopted some preventive actions regarding NPS, and included both the whole plant and its main bioactive compound, salvinorin A (SA), in the list of NPS. S. divinorum is a plant with hallucinogenic properties that, till the mid-60’s, was used only by Mazatecas (Oaxaca region, Mexico). The responsible substance for its hallucinogenic properties is SA, which is a neoclerodane diterpene different from the others hallucinogenic substances known, as it presents a unique mechanism of action. In fact, SA is a selective agonist of kappa opioid receptors (KOR). Despite the fact that the main target of S. divinorum, and SA, is the central nervous system, to date, there are no studies assessing its putative peripheral effects, in particular in the liver, the main responsible organ for the metabolism of several endogenous substances and xenobiotics. With this study we intended to evaluate the in vitro cytotoxicity and to analyze the relative gene expression of certain genes relevant to organism caused by the exposure of hepatic cells (Hep G2 and WRL-68) to different concentrations of SA during various periods of time. The genes included in this study were: apolipoprotein B100 (Apo B100), carboxylesterase 1 (Ces 1), cytochrome C (Cyt C), cytochrome P450 1A2 (CYP450 1A2), cytochrome P450 2D6 (CYP450 2D6), cytochrome P450 3A4 and glucuronosyltransferase 1A1 (UGT 1A1). Overall, data obtained revealed that 50 µM of SA is cytotoxic for both cell lines, while 10 µM only promoted a decrease in the cellular viability in Hep G2 cells after a 72 h incubation. Thus, it seems that, apparently, Hep G2 cells are more sensitive to SA, comparing with WRL-68. Instead, 0,1 and 1 µM did not promoted any alteration in the relative cellular viability in both cell lines. Regarding the effects of 1 µM SA exposure in Hep G2 and WRL-68 at during various periods of time (12, 24 and 72 h) results showed that, overall, SA has a significant impact in the mRNA relative expression of various genes, depending on the time of incubation, concentration and the cell line. Besides that, we also found that Ces 1 was not detected in WRL-68 cells pointing out that it is not expressed in this cell line. SA revealed to have a safe toxicological profile yet, the effects of SA in the regulation of gene expression are important to highlight as they could exert positive or negative effects depending in the gene of study, time of exposure and cell line. Finally, because this is the first study that shows the SA potential effects in the liver, further research is needed to confirm if that they are directly related to SA consumption. Thus, a new research line raised and future studies should be conducted in order to achieve the in vitro protein expression of the genes and to confirm these findings in in vivo studies (rats).