Regina Lúcia de Moraes Moreau

Solid-phase micro-extraction-gas chromatography-mass spectrometry and headspace-gas chromatography of tetrahydrocannabinol, amphetamine, methamphetamine, cocaine and ethanol in saliva samples

In the present work, a method was developed aiming at the serial detection of tetrahydrocannabinol (THC), amphetamine, methamphetamine, cocaine and ethanol in saliva. Saliva samples were submitted to an initial headspace procedure for ethanol determination by gas chromatography/flame ionization detector (GC-FID). After this step, two consecutive solid-phase micro-extractions (SPME) were carried out: THC was extracted by submersing a polydimethylsiloxane fiber (100 micro m) in the vial for 20 min; amphetamine, methamphetamine and cocaine were subsequently extracted after alkalinization. Derivatization of the amphetamines was carried out directly in the solution by adding 2 micro l of butylchloroformate. Gas chromatography-mass spectrometry (GC-MS) was used to identify the analytes in selected ion monitoring (SIM) mode. Confidence parameters of validation of the method were: recovery, linearity, intra- and inter-assay precision as well as limits of detection and quantification of the analytes. The limits of quantification (LOQ) obtained were: ethanol (0.010 g/l); amphetamine (5.0 ng/ml); methamphetamine (0.5 ng/ml); cocaine (5 ng/ml) and THC (5 ng/ml). The method proved to be highly precise (coefficient of variation<8%) for all detected substances.

Marijuana as Doping in Sports

A high incidence of positive cases for cannabinoids, in analyses for doping control in sports, has been observed since the International Olympic Committee (IOC) included them in the 1989 list of prohibited drugs under the title of classes of prohibited substances in certain circumstances. Where the rules of sports federations so provide, tests are conducted for marijuana, hashish or any other cannabis product exposure by means of urinalysis of 11-nor-delta-9-tetrahydro-cannabinol-9-carboxylic acid (carboxy-THC) the main metabolite of delta-9-tetrahydrocannabinol (THC). Concentrations >15 ng/mL (cut-off value) in confirmatory analytical procedures are considered doping. Cannabis is an illicit drug in several countries and has received much attention in the media for its potential therapeutic uses and the efforts to legalise its use.Studies have demonstrated that the use of cannabinoids can reduce anxiety, but it does not have ergogenic potential in sports activities. An increase in heart rate and blood pressure, decline of cardiac output and reduced psychomotor activity are some of the pharmacological effects of THC that will determine a decrease in athletic performance. An ergolytic activity of cannabis products has been observed in athletes of several different sport categories. In Brazil, analyses for doping control in sports, performed in our laboratories, have detected positive cases for carboxy-THC in urine samples of soccer, volleyball, cycling and other athletes.It is our intention to discuss in this article some points that may discourage individuals from using cannabis products during sports activities, even in the so-called permitted circumstances defined by the IOC and some sports federations.

Uso de esteróides anabólicos androgênicos por praticantes de musculação de grandes academias da cidade de São Paulo

To estimate the use of anabolic-androgenic steroids (AAS) among body builders of three - professionally equipped private gym in Sao Paulo, Brazil, body builders answered voluntary and anonimously a structured multiple itens questionnaire which was available for a week in these gym centers. The participants were informed in advance of the aim of the study. Of the 209 body builders attending (3% of the total), 19% had used AAS at some stage. The stratification of the results showed that 11% were composed of ex-user and 8% were of active users. Considering only male body builder the incidence of AAS was 24%. Stanozolol and nandrolone decanoate were the most commonly AAS used. The significant features of their profile were: the median age of users was 27 (25-29) years, predominantely males and motivation for using AAS was aesthetic with a high level of physical training. Food suplements as well as other drugs in association were used. Mainly AAS were bought in drugstores without prescriptions. Body builders considerered anabolic-androgenic steroids to be dangerous and that the adverse effects can be managed or avoided with other drugs or medical care. This study demonstrate the use of AAS among body builders and consequently the necessity of deep preventive and educationaly investigation among people exposed to AAS.

Alteração da relação testosterona: cortisol induzida pelo treinamento de força em mulheres

La razon entre testosterona y cortisol (T:C) es frecuentemente utilizada como indicador del nivel de stress impuesto por el ejercicio. Las alteraciones de las concentraciones de estas hormonas son las responsables por modular diversas respuestas inducidas por el entrenamiento, como son la hipertrofia y el aumento de la fuerza. El objetivo del presente estudio fue examinar la influencia del protocolo de entrenamiento de fuerza, conocido como series multiples (MS), sobre la ganancia de fuerza, la resistencia muscular localizada y la relacion entre las concentraciones de las hormonas catabolicas (cortisol) y anabolicas (testoterona). Para testar esta hipotesis, cinco jovenes del sexo feminino con un ano de experiencia en entrenamiento de fuerza fueron sometidas al protocolo MS. Las muestras de sangre fueron colectadas antes e imediatamente despues del ejercicio, en el primer dia y despues de ocho semanas de entrenamiento. Los tests de 1-RM y de repeticiones maximas fueron realizados tambien al inicio y al final despues de las ocho semanas del entrenamiento de fuerza. No fueron observadas alteraciones de la masa corporal, IMC, porcentaje de masa grasa, fuerza (1-RM) para los ejercicios supino, agachamiento y rosca directa. El numero de repeticiones maximas al 50% de 1-RM fue aumentando solamente apenas para el supino (p < 0,05). No se observo alteracion en la concentracion de la testosterona total. Con relacion a la concentracion plasmatica de cortisol despues de las ocho semanas de entrenamiento, en la situacion de reposo fue reducida (38% - p < 0,05). En consecuencia de la atenuacion de la secrecion de cortisol despues de ocho semanas de entrenamiento, la razon T:C presento elevacion del 20% de la situacion de reposo (p < 0,05). A pesar de no haber sido detectadas alteraciones funcionales en los tests de 1-RM de repeticiones maximas, el metodo MS indujo un cuadro hormonal favorable al anabolismo proteico.

Cafeína e performance em exercícios anaeróbios

The ergogenic effects of caffeine on performance in anaerobic activites are not clear yet, so are the mechanisms involved in this type of physical effort. The theories that have been trying to explain the ergogenic effects of caffeine during anaerobic activities are related to the effect of caffeine in some portion of the central nervous system (CNS), the propagation of neural signs between brain and neuromuscular junction, and also to the effect of caffeine on the skeletal muscle, facilitating the stimulation-contraction of the skeletal muscle. Some studies have been indicating increases of muscular strength accompanied by greater resistance to the installation of the process of muscular fatigue after caffeine ingestion. It has been suggested that it happens much more by the direct action of caffeine in CNS than by its action at peripherical level. Regarding maximum and submaximum exercises of short duration, the studies have been controversial, although most of them indicates that caffeine seems to improve the performance significantly in maximum exercises of short duration (<5 min), when not preceded by sub maximum prolonged exercises. However, these results need to be confirmed, as well as the mechanisms of action of caffeine in these types of efforts.

Non-Intentional Doping in Sports

Compulsory drug testing was introduced in 1968 by the International Olympic Committee. Since then, several doping cases have been reported in sports competition world wide. Positive results are based on the detection of prohibited substances, their metabolites and markers in biological (mainly urine) samples supplied by athletes. In some cases, the evidences were not contested and athletes admitted the use of banned substances. However, in other cases, athletes denied the use of doping to enhance performance and claimed to have inadvertently or passively absorbed the drug. Unfortunately, no current accepted analytical method is capable of distinguishing between a sample from a cheater and one from an athlete who was passively exposed to a doping agent.Athletes’ allegations have included the passive inhalation of drug smoke (e.g. marijuana) or the ingestion of food or products sold as nutritional supplements that contained prohibited substances. In the scientific literature, several studies have been performed to investigate the possibility of an accidental exposure being the reason for the appearance of detectable quantities of banned substances in urine samples. Based on these studies, this article discusses those cases where the athlete’s claims could be possible in generating a positive result in doping control and in which circumstances it would be improbable to happen.

Abnormal Neurovascular Control in Anabolic Androgenic Steroids Users

PURPOSE Previous studies showed that anabolic androgenic steroids (AAS) increase vascular resistance and blood pressure (BP) in humans. In this study, we tested the hypotheses 1) that AAS users would have increased muscle sympathetic nerve activity (MSNA) and reduced forearm blood flow (FBF) compared with AAS nonusers and 2) that there would be an association between MSNA and 24-h BP. METHODS Twelve AAS users aged 31 +/- 2 yr (means +/- SE) and nine age-matched AAS nonusers aged 29 +/- 2 yr participated in the study. All individuals were involved in strength training for at least 2 yr. AAS was determined by urine test (chromatography-mass spectrometry). MSNA was directly measured by microneurography technique. FBF was measured by venous occlusion plethysmography. BP monitoring consisted of measures of BP for 24 h. RESULTS MSNA was significantly higher in AAS users than that in AAS nonusers (29 +/- 3 vs 20 +/- 1 bursts per minute, P = 0.01). FBF (1.92 +/- 0.17 vs 2.77 +/- 0.24 mL x min(-1) x 100 mL(-1), P = 0.01) and forearm vascular conductance (2.01 +/- 0.17 vs 2.86 +/- 0.31 U, P = 0.02) were significantly lower in AAS users than that in AAS nonusers. Systolic (131 +/- 4 vs 120 +/- 3 mm Hg, P = 0.001), diastolic (74 +/- 4 vs 68 +/- 3 mm Hg, P = 0.02), and mean BP (93 +/- 4 vs 86 +/- 3 mm Hg, P = 0.005) and heart rate (74 +/- 3 vs 68 +/- 3 bpm, P = 0.02) were significantly higher in AAS users when compared with AAS nonusers. Further analysis showed that there was a significant correlation between MSNA and 24-h mean BP (r = 0.75, P = 0.002). CONCLUSIONS AAS increases MSNA and reduces muscle blood flow in young individuals. In addition, the increase in BP levels in AAS users is associated with augmented sympathetic outflow. These findings suggest that AAS increases the susceptibility for cardiovascular disease in humans.

Effects of different doses and schedules of diazepam treatment on lymphocyte parameters in rats

Benzodiazepines (BZD) are widely used for the treatment of anxiety. They enhance GABA-ergic neurotransmission through the binding on specific BDZ recognition sites, within the GABA(A) receptor-ion channel complex. However, recent studies showed that BZD also act on peripheral benzodiazepine receptor sites (PBR) or translocator protein 18 kDa (TSPO). Evidence for a direct immunomodulatory action for BZD emerged from studies that demonstrated the presence of TSPO on immune/inflammatory cells. The present study was designed to analyze the effects of diazepam on rat lymphocyte parameters, specifically on phenotype, cell proliferation and cell death. The effects of both acute and long-term (21 days) diazepam (1 and 10 mg/kg/day) administrations were evaluated. Results showed that diazepam (1 mg/kg) treatment did not change the immune parameters analyzed. However, both diazepam (10 mg/kg) acute and long-term treatments decreased the number of apoptotic cells; they also increased the percentage of T cytotoxic cells; decreased the percentage of B cells and increased the corticosterone serum levels. The induction of functional tolerance was suggested for the highest dose of diazepam (10 mg/kg), but not for the smaller dose (1 mg/kg) used, at least for diazepam effects on corticosterone serum levels. Diazepam effects were discussed as being related to the number of TSPO sites present on immune cells and/or to the increased levels of serum corticosterone observed after the treatments used.

Methylenedioxymethamphetamine (Ecstasy) Decreases Neutrophil Activity and Alters Leukocyte Distribution in Bone Marrow, Spleen and Blood

Objective: Looking for possible neuroimmune relationships, we analyzed the effects of methylenedioxymethamphetamine (MDMA) administration on neuroendocrine, neutrophil activity and leukocyte distribution in mice. Methods: Five experiments were performed. In the first, mice were treated with MDMA (10 mg/kg) 30, 60 min and 24 h prior to blood sample collection for neutrophil activity analysis. In the second experiment, the blood of naïve mice was collected and incubated with MDMA for neutrophil activity in vitro analysis. In the third and fourth experiments, mice were injected with MDMA (10 mg/kg) and 60 min later, blood and brain were collected to analyze corticosterone serum levels and hypothalamic noradrenaline (NA) levels and turnover. In the last experiment, mice were injected with MDMA 10 mg/kg and 60 min later, blood, bone marrow and spleen were collected for leukocyte distribution analysis. Results: Results showed an increase in hypothalamic NA turnover and corticosterone serum levels 60 min after MDMA (10 mg/kg) administration, a decrease in peripheral blood neutrophil oxidative burst and a decrease in the percentage and intensity of neutrophil phagocytosis. It was further found that MDMA (10 mg/kg) treatment also altered leukocyte distribution in blood, bone marrow and spleen. In addition, no effects were observed for MDMA after in vitro exposure both in neutrophil oxidative burst and phagocytosis. Conclusion: The effects of MDMA administration (10 mg/kg) on neutrophil activity and leukocyte distribution might have been induced indirectly through noradrenergic neurons and/or hypothalamic-pituitary-adrenal axis activations.

Differential behavioral outcomes of 3,4-methylenedioxymethamphetamine (MDMA-ecstasy) in anxiety-like responses in mice

Anxiolytic and anxiogenic-like behavioral outcomes have been reported for methylenedioxymethamphetamine (MDMA or ecstasy) in rodents. In the present experiment, we attempted to identify behavioral, hormonal and neurochemical outcomes of MDMA treatment to clarify its effects on anxiety-related responses in 2-month-old Balb/c male mice (25-35 g; N = 7-10 mice/group). The behavioral tests used were open field, elevated plus maze, hole board, and defensive behavior against predator odor. Moreover, we also determined striatal dopamine and dopamine turnover, and serum corticosterone levels. MDMA was injected ip at 0.2, 1.0, 5.0, 8.0, 10, or 20 mg/kg. MDMA at 10 mg/kg induced the following significant (P < 0.05) effects: a) a dose-dependent increase in the distance traveled and in the time spent moving in the open field; b) decreased exploratory activity in the hole board as measured by number of head dips and time spent in head dipping; c) increased number of open arm entries and increased time spent in open arm exploration in the elevated plus maze; d) increased time spent away from an aversive stimulus and decreased number of risk assessments in an aversive odor chamber; e) increased serum corticosterone levels, and f) increased striatal dopamine level and turnover. Taken together, these data suggest an anxiogenic-like effect of acute MDMA treatment, despite the fact that behavioral anxiety expression was impaired in some of the behavioral tests used as a consequence of the motor stimulating effects of MDMA.