Conjugation with polyamines enhances the antitumor activity of naphthoquinones against human glioblastoma cells.

Glioblastoma multiform (GBM) is the most common and devastating type of primary brain tumor, being considered the deadliest of human cancers. In this context, extensive efforts have been undertaken to develop new drugs that exhibit both antiproliferation and antimetastasis effects on GBM. 1,4-Naphthoquinone (1,4-NQ) scaffold has been found in compounds able to inhibit important biological targets associated with cancer, which includes DNA topoisomerase, Hsp90 and monoamine oxidase. Among potential antineoplastic 1,4-NQs is the plant-derived lapachol (2-hydroxy-3-prenyl-1,4-naphthoquinone) that was found to be active against the Walker-256 carcinoma and Yoshida sarcoma. In the present study, we examined the effect of polyamine (PA)-conjugated derivatives of lapachol, nor-lapachol and lawsone on the growth and invasion of the human GBM cells. The conjugation with PA (a spermidine analog) resulted in dose-dependent and time-dependent increase of cytotoxicity of the 1,4-NQs. In addition, in-vitro inhibition of GBM cell invasion by lapachol was increased upon PA conjugation. Previous biochemical experiments indicated that these PA-1,4-NQs are capable of inhibiting DNA human topoisomerase II-α (topo2α), a major enzyme involved in maintaining DNA topology. Herein, we applied molecular docking to investigate the binding of PA-1,4-NQs to the ATPase site of topo2α. The most active molecules preferentially bind at the ATP-binding site of topo2α, which is energetically favored by the conjugation with PA. Taken together, these findings suggested that the PA-1,4-NQ conjugates might represent potential molecules in the development of new drugs in chemotherapy for malignant brain tumors.

The Hypnotic, Anxiolytic, and Antinociceptive Profile of a Novel µ-Opioid Agonist.

5'-4-Alkyl/aryl-1H-1,2,3-triazole derivatives PILAB 1-12 were synthesized and a pharmacological screening of these derivatives was performed to identify a possible effect on the Central Nervous System (CNS) and to explore the associated mechanisms of action. The mice received a peritoneal injection (100 µmol/kg) of each of the 12 PILAB derivatives 10 min prior to the injection of pentobarbital and the mean hypnosis times were recorded. The mean hypnosis time increased for the mice treated with PILAB 8, which was prevented when mice were administered CTOP, a µ-opioid antagonist. Locomotor and motor activities were not affected by PILAB 8. The anxiolytic effect of PILAB 8 was evaluated next in an elevated-plus maze apparatus. PILAB 8 and midazolam increased a percentage of entries and spent time in the open arms of the apparatus compared with the control group. Conversely, a decrease in the percentages of entries and time spent in the closed arms were observed. Pretreatment with naloxone, a non-specific opioid antagonist, prior to administration of PILAB 8 exhibited a reverted anxiolytic effect. PILAB 8 exhibited antinociceptive activity in the hot plate test, and reduced reactivity to formalin in the neurogenic and the inflammatory phases. These data suggest that PILAB 8 can activate µ-opioid receptors to provoke antinociceptive and anti-inflammatory effects in mice.

Electrospray ionization tandem mass spectrometry analysis of isopimarane diterpenes from Velloziaceae.

RATIONALE: The study of natural products by electrospray ionization tandem mass spectrometry (ESI-MS/MS) is an important strategy for the characterization of the major fragmentation reactions which can then help to determine the composition of complex mixtures. Application of ESI-MS/MS to a series of isopimarane diterpenes from Velloziaceae allowed the rationalization of their fragmentation mechanisms. METHODS: Velloziaceae diterpenes were isolated by silica gel column chromatography and investigated by ESI-MS/MS analysis. The fragmentation studies were performed on a quadrupole-time-of-flight instrument using N2 as the collision gas. To help rationalize the fragmentation pathways observed, the geometry and sites of reactivity of the diterpenes were obtained by theoretical calculations using the B3LYP/6-31 + G(d,p) model. Fragmentation mechanisms were proposed on the basis of the calculated protonation sites and product ions energies using density functional theory (DFT) methods. RESULTS: The presence of hydroxyl and carbonyl groups on the terpene core influences the protonation site observed. One compound showed a radical cation as the base peak. MS/MS spectra exhibit water elimination as the major fragmentation pathway (via two ways), either when protonation takes place on the oxygen atom, or through elimination after activation from hydrogen migration. After the elimination of water, the formation of an endocyclic double bond induces a sequential retro-Diels-Alder (RDA) reaction as the major fragmentation step. CONCLUSIONS: A thorough rational analysis of the fragmentation mechanisms of protonated Velloziaceae diterpenes was used to propose the dissociation mechanisms in ESI-MS/MS. The presence of esters in the side chain also influenced the intensity or occurrence of the observed protonated or cationized molecules in ESI-MS. These results will aid the identification of analogues in sample extracts in future metabolomics studies.

Design, synthesis, and evaluation of guanylhydrazones as potential inhibitors or reactivators of acetylcholinesterase.

Analogs of pralidoxime, which is a commercial antidote for intoxication from neurotoxic organophosphorus compounds, were designed, synthesized, characterized, and tested as potential inhibitors or reactivators of acetylcholinesterase (AChE) using the Ellman's test, nuclear magnetic resonance, and molecular modeling. These analogs include 1-methylpyridine-2-carboxaldehyde hydrazone, 1-methylpyridine-2-carboxaldehyde guanylhydrazone, and six other guanylhydrazones obtained from different benzaldehydes. The results indicate that all compounds are weak AChE reactivators but relatively good AChE inhibitors. The most effective AChE inhibitor discovered was the guanylhydrazone derived from 2,4-dinitrobenzaldehyde and was compared with tacrine, displaying similar activity to this reference material. These results indicate that guanylhydrazones as well as future similar derivatives may function as drugs for the treatment of Alzheimer's disease.

Antinociceptive and anti-inflammatory effects of a Geissospermum vellosii stem bark fraction.

Geissospermum vellosii (Pao pereira) is a Brazilian tree whose stem barks are rich in indole alkaloids that present intense anticholinesterase activity. The present study evaluated the effects of a stem bark fraction (PPAC fraction) and ethanolic extract (EE) of Pao pereira in classic murine models of inflammation and pain. The EE and PPAC fraction, both at a dose of 30 mg/kg, significantly reduced mice abdominal constriction induced by acetic acid by 34.8% and 47.5%, respectively. In the formalin test, EE (30 mg/kg) and PPAC fraction (30 and 60 mg/kg) inhibited only the second phase, by 82.8%, 84.9% and 100%, respectively. Compared with indomethacin, similar doses of EE or PPAC fraction were approximately twice as effective in causing antinociception. PPAC fraction was not effective in the hot plate test but reduced the inflammatory response at the second (50.6%) and third (57.8%) hours of rat paw edema induced by carrageenan. Antihyperalgesic activity was observed within 30 min with a peak at 2 h (60.1%). These results demonstrate that compounds in PPAC fraction have anti-inflammatory and antinociceptive activity by a mechanism apparently unrelated to the opioid system. Regardless of similar responses to indomethacin, the effects of PPAC fraction are mainly attributed to acetylcholine actions.

Anthranilic acids from isatin: an efficient, versatile and environmentally friendly method.

This paper describes the preparation of a series of 16 anthranilic acids in yields ranging from 51 to 97%, by treating the isatins with NaOH and H2O2. Independently of the nature of the substituent on the aromatic ring, the reactions were complete in 15 min at room temperature, whereas those of isatins containing a substituent on the nitrogen atom required longer reaction time for completion (45 min) under the same reaction conditions.

A Taxonomic Revision of the South American Trilobite Cockroaches of Parahormetica Brunner von Wattenwyl 1865 (Blattodea: Blaberidae), with Description of Parahormetica museunacional sp. nov. from the Atlantic Forest.

The taxonomically intricate genus of trilobite cockroaches, Parahormetica Brunner von Wattenwyl, 1865, is revised based on a comparative morphological analysis. The goals of this study are to review the nomenclature, propose hypotheses about specific delimitation, and provide diagnoses to allow identification of the taxonomic units in the genus. Based on the revised status of Parahormetica, we transferred Parahormetica hylaeceps Miranda-Ribeiro, 1936, and Parahormetica punctata Saussure, 1873, to the genus Bionoblatta Rehn, 1940. Therefore, the genus includes now four species of giant cockroaches which are predominantly distributed on the Atlantic Forest: Parahormetica bilobata (Saussure, 1864), Parahormetica cicatricosa Saussure, 1869, Parahormetica monticollis (Burmeister, 1838), and Parahormetica museunacional sp. nov. (holotype male deposited in DZUP: Brazil, Paraná). Diagnoses, key, distribution maps, images of living, non-type, and type specimens are made available. Our results make clear that the status and limits among Brachycolini genera pending a full revision.

Larval taxonomy of Macrothemis Hagen, 1868 (Odonata: Libellulidae), with descriptions of four larvae and a key to the fourteen known species.

The ultimate larval stadia of Macrothemis declivata, M. hemichlora, M. imitans imitans and M. tenuis are described and illustrated for the first time, based on material from Brazil. Six of the most used keys to larvae of libellulid genera of the New World are evaluated with respect to the correct identification of the 27 known larvae of Macrothemis, Brechmorhoga, Gynothemis and Scapanea. Macrothemis species were wrongly identified in more than 50% of the trials, being keyed as Brechmorhoga, Gynothemis and even Dythemis. The genus Macrothemis and its relatives need to be revaluated and adequately diagnosed based on larvae. A key to the 14 known larvae of species currently included in Macrothemis is presented.

New trifluoromethyl triazolopyrimidines as anti-Plasmodium falciparum agents.

According to the World Health Organization, half of the World's population, approximately 3.3 billion people, is at risk for developing malaria. Nearly 700,000 deaths each year are associated with the disease. Control of the disease in humans still relies on chemotherapy. Drug resistance is a limiting factor, and the search for new drugs is important. We have designed and synthesized new 2-(trifluoromethyl)[1,2,4]triazolo[1,5-a]pyrimidine derivatives based on bioisosteric replacement of functional groups on the anti-malarial compounds mefloquine and amodiaquine. This approach enabled us to investigate the impact of: (i) ring bioisosteric replacement; (ii) a CF3 group substituted at the 2-position of the [1,2,4]triazolo[1,5-a]pyrimidine scaffold and (iii) a range of amines as substituents at the 7-position of the of heterocyclic ring; on in vitro activity against Plasmodium falciparum. P. falciparum dihydroorotate dehydrogenase (PfDHODH) through strong hydrogen bonds. The presence of a trifluoromethyl group at the 2-position of the [1,2,4]triazolo[1,5-a]pyrimidine ring led to increased drug activity. Thirteen compounds were found to be active, with IC50 values ranging from 0.023 to 20 µM in the anti-HRP2 and hypoxanthine assays. The selectivity index (SI) of the most active derivatives 5, 8, 11 and 16 was found to vary from 1,003 to 18,478.

The mechanism of Sandmeyer's cyclization reaction by electrospray ionization mass spectrometry.

Using electrospray ionization (tandem) mass spectrometry (ESI-MS(/MS)) spectrometric experiments, the Sandmeyer reaction was monitored on-line, and key intermediates were intercepted and characterized for the first time. The mechanistic information provided by on-line ESI-MS(/MS) is in accordance with Sandmeyer's proposal, and was made possible by coupling a microreactor on-line to the ESI ion source, which allowed reactions to be screened from 0.7-2.0 s, identifying and characterizing all intermediates that were formed and consumed during the reaction.

Pharmacological activity of novel 2-hydroxyacetophenone isatin derivatives on cardiac and vascular smooth muscles in rats.

Isatin (1H-indole-2,3 dione) is an endogenous compound with biological activities. Many of its derivatives have pharmacological effects, including inhibition of cyclic guanosine monophosphate levels in cardiac tissue; sedative-hypnotic profiles; anticonvulsant, analgesic, antithermic, and anti-inflammatory activities; and anxiolytic, antimicrobial, and proapoptotic effects. Carbamates derived from isatin have a vasorelaxant profile. This study investigated the activity of 2 novel 2-hydroxyacetophenone derivatives of isatin (named MB101 and MB130) on the contractility of rat aorta and papillary muscles. Both compounds induced a concentration-dependent relaxation (5-100 µM) in the endothelium-intact aorta that was abolished by N-nitro-L-arginine methyl ester. Atropine, a muscarinic receptor antagonist, significantly prevented vasodilatation of 100 µM MB101. In contrast, atropine caused no significant alteration in MB130-induced vasorelaxation. Naloxone, a nonselective opioid receptor antagonist, completely prevented the relaxing effect of MB101 and MB130 at all concentrations. In papillary muscles, only MB130 induced a significant depression, and this contractile response was not altered by propranolol and atropine. Both the compounds reduced systolic and diastolic pressures in a dose-dependent manner in anesthetized rats. The 2-hydroxyacetophenones produced direct effects on vascular tonus through either muscarinic or opioid pathways. MB130 produced cardiac depression by opioid receptors and bradykinin because pretreatment HOE140 or with naloxone, an antagonist of type-2, bradykinin were able to partially block the decrease in twitch amplitude in papillary muscles induced by MB130. These findings provide information for designing new strategies for the treatment of cardiovascular disorders.

Anthraquinones from the bark of Senna macranthera.

2-acetyl physcion (2-acetyl-1,8-dihydroxy-6-methoxy-3-methyl-9,10-anthraquinone, 2), a rare anthraquinone, was isolated from Senna macranthera var. nervosa (Vogel) H.S. Irwin & Barneby (Fabaceae). The chemical structure was elucidated and all (1)H and (13)C NMR chemical shifts were assigned by NMR one- ((1)HNMR, {(1)H}-(13)CNMR, and APT-(13)CNMR) and two (COSY, NOESY, HMQC and HMBC) dimensional of this natural compound. Furthermore, the minor anthraquinones chrysophanol (3), chrysophanol-8-methyl ether (4) and physcion (5) were characterized by GC-MS analysis. The occurrence of the anthraquinones 3-5 confirms that S. macranthera is a typical representative of the genus Senna.

A rare case of spontaneous rupture of epithelioid angiomyolipoma.

A male patient, 40 years of age, arrived at our Institute with diffuse abdominal tenderness, right flank pain, hematuria and early stage of hemorrhagic shock with anemia and initial hypotension. The immediate clinical history revealed no significant previous trauma, only subsequently was reported inconstant pain in the right flank for 4-5 days with pallor and asthenia, signs and symptoms that the patient had not investigated. Abdominal CT scan with angiographic evaluation was performed showing right kidney mass and perirenal fluid collection by blood component. Immediate nephrectomy was performed and histopathological and further immunohistochemical study, revealed the epithelioid variant of angiomyolipoma.

Are publications on zoological taxonomy under attack?

Taxonomy is essential to biological sciences and the priority field in face of the biodiversity crisis. The industry of scientific publications has made extensive promotion and display of bibliometric indexes, resulting in side effects such as the Journal Impact Factor™ (JIF) mania. Inadequacies of the widely used indexes to assess taxonomic publications are among the impediments for the progress of this field. Based on an unusually high proportion of self-citations, the mega-journal Zootaxa, focused on zoological taxonomy, was suppressed from the Journal Citation Reports (JCR, Clarivate™). A prompt reaction from the scientific community against this decision took place exposing myths and misuses of bibliometrics. Our goal is to shed light on the impact of misuse of bibliometrics to the production in taxonomy. We explored JCR's metrics for 2010-2018 of 123 zoological journals publishing taxonomic studies. Zootaxa, with around 15 000 citations, received 311% more citations than the second most cited journal, and shows higher levels of self-citations than similar journals. We consider Zootaxa's scope and the fact that it is a mega-journal are insufficient to explain its high level of self-citation. Instead, this result is related to the 'Zootaxa phenomenon', a sociological bias that includes visibility and potentially harmful misconceptions that portray the journal as the only one that publishes taxonomic studies. Menaces to taxonomy come from many sources and the low bibliometric indexes, including JIF, are only one factor among a range of threats. Instead of being focused on statistically illiterate journal metrics endorsing the villainy of policies imposed by profit-motivated companies, taxonomists should be engaged with renewed strength in actions directly connected to the promotion and practice of this science without regard for citation analysis.

Synthesis of alpha- and beta-pyran naphthoquinones as a new class of antitubercular agents.

A series of alpha- and beta-pyran naphthoquinones (lapachones) have been synthesized and evaluated for their in-vitro antibacterial activity against Mycobacterium tuberculosis strain H37Rv (ATCC 27294) using the Alamar-Blue susceptibility test; the activity was expressed as the minimum inhibitory concentration (MIC) in microg/mL. The synthetic methodology consisted of the formation of methylene and aryl o-quinone methides (o-QMs) generated by Knoevenagel condensation of 2-hydroxy-1,4-naphthoquinone with formaldehyde and arylaldehydes. These o-QMs then undergo facile hetero Diels-Alder reactions with dienophiles in aqueous ethanol media. Some naphthoquinones exhibited inhibition with MIC values of 1.25 microg/mL, similar to that of pharmaceutical concentrations currently used in tuberculosis treatment. These results justify further research into the value of these quinones as part of an original treatment for tuberculosis.

Synthetic lapachol derivatives relax guinea-pig ileum by blockade of the voltage-gated calcium channels.

The present study was designed to further evaluate a possible spasmolytic activity of synthetic lapachol derivatives, norlapachol, alpha-norlapachone, beta-norlapachone and hydro-hydroxy-norlapachol (HH-norlapachol), on guinea-pig ileum. In guinea-pig ileum, except for norlapachol, all naphthoquinones inhibited the phasic contractions induced by carbachol or histamine. Even when the ileum was pre-contracted with KCl, carbachol or histamine, all naphthoquinones induced relaxation, suggesting that these naphthoquinones could be acting on the voltage-gated calcium channels (Ca(V)). As the tonic component this contraction is maintained mainly by the opening of the Ca(V), we hypothesized that these naphthoquinones might be acting on these channels. This hypothesis was confirmed by the observation that norlapachol (pD'2 = 4.99), alpha-norlapachone (pD'2 = 4.49), beta-norlapachone (pD'2 = 6.33), and HH-norlapachol (pD'2 = 4.53) antagonized the contractions induced by CaCl2 in depolarizing medium nominally without Ca2+. As beta-norlapachone was the most potent we decided to continue the study of its action mechanism. The fact that this naphthoquinone has inhibited the tonic contractions induced by S-(-)-Bay K8644 [EC50 = (1.6 +/- 0.30) x 10(-5) M] suggests that the Ca2+ channel involved belongs to the type L (Ca(V)1.2). In addition, in the functional level, the spasmolytic effect of beta-norlapachone does not involve participation of free radicals, since its curve of relaxation was unchanged in the presence of glutathione, an antioxidant agent.

(2E)-N-(3,5-Dibromo-4-methoxy-phen-yl)-2-(hydroxy-imino)acetamide.

The title compound, C(9)H(8)Br(2)N(2)O(3), is planar (r.m.s. deviation = 0.030 Å) with the exception of the terminal methyl group which lies out of the plane [1.219 (3) Å]. The conformation about the C=N double bond [1.268 (3) Å] is E. An intra-molecular N-H⋯N hydrogen bond occurs. Linear supra-molecular chains along the b axis mediated by O-H⋯O hydrogen-bonding inter-actions feature in the crystal structure. These chains are also stabilized by weak C-H⋯N contacts.

Thraulodes marianoi sp. nov., a remarkable new species of mayfly (Ephemeroptera: Leptophlebiidae) with dark wings from the southern Brazilian Atlantic Forest.

Thraulodes marianoi sp. nov. is described, illustrated and diagnosed based on a single male imago from a subtropical forest at Pico do Marumbi State Park, in the protected area of Mananciais da Serra, state of Paraná, Brazil. Based on the dark coloration of legs and the large dark spot at base of forewing, T. marianoi sp. nov. is similar to species of the niger-group (Peruvian Amazonia) and to T. basimaculatus Giordano Domínguez, 2005, a species endemic to Bolivia. The new species can be distinguished from all other species in the genus Thraulodes Ulmer, by the combination of following characteristics: (1) four cross-veins basal to bulla in forewing; (2) brown area covering about of proximal half of the forewing; (3) pleura violet; (4) middle and posterior femora yellowish washed with dark brown; (5) terga and sterna I-IV white washed with dark brown on posterior and lateral margin terga and sterna V-X dark brown; (6) penes short and wide with distolateral area, "ear-like" and poorly developed lateral pouch; (7) styliger plate triangular, median projection short and rounded on the apex. This new species represents the first record of the genus from the state of Paraná, Brazil.

Oral treatments with a flavonoid-enriched fraction from Cecropia hololeuca and with rutin reduce articular pain and inflammation in murine zymosan-induced arthritis.

ETHNOPHARMACOLOGICAL RELEVANCE: Cecropia Loefl. species (Urticaceae) are widely spread across the rainforest in tropical and subtropical regions of Central and South America. Inhabitants of different regions of Brazil employ leaves, fruits and sprouts of Cecropia hololeuca Miq. mainly as anti-inflammatory, anti-asthmatic, expectorant, fever suppressant, and against cough. AIM OF THE STUDY: To evaluate the antinociceptive and anti-inflammatory activities of an aqueous leaf extract of C. hololeuca in a murine model of zymosan-induced arthritis (ZIA) and characterize compounds contributing to these effects. MATERIALS AND METHODS: The crude aqueous extract of C. hololeuca (CAE) was obtained by infusion, screened for antinociceptive and anti-inflammatory activities, and fractionated (solvent partition; RP-2 and Sephadex G-25 column chromatography), yielding fractions that were chemically and pharmacologically investigated. TLC, HPLC-DAD, HPLC-DAD-ESI-MS/MS and NMR analyses were peformed. The antinociceptive activity was assessed by means of acetic acid-induced writhing, hot-plate and rota-rod tests. ZIA was used to evaluate the anti-arthritic activity of oral treatment with CAE, butanolic (BF) and aqueous fraction (AF), as well as the fractions obtained from BF (F2, F2-A and F2-B). Rutin, a flavonoid found in C. hololeuca, was also tested. Mechanical hypernociception, joint edema, local neutrophil recruitment and articular TNF-α quantification were performed to measure the severity of arthritis and identify the anti-inflammatory potential of C. hololeuca. RESULTS: CAE (0.03-1 g/kg, p.o.) showed a dose-related inhibitory effect on acetic acid-induced writhing test, but did not change the pain latency in the hotplate test, nor the first fall time on the rota-rod test. In addition, CAE (1 g/kg, p.o.) inhibited by 65% the mechanical hypernociception, 46% the joint edema, 54% the neutrophil recruitment and 53% the articular TNF-α concentration levels in ZIA. BF (0.4 g/kg, p.o.), AF (0.6 g/kg), F2 (0.1 g/kg) and F2-A (0.045 g/kg), but not F2-B (0.055 g/kg), inhibited the mechanical hypernociception, joint edema and neutrophil recruitment in ZIA. Rutin (0.001-0.03 g/kg, p.o.) produced dose-related inhibitory effects in the mechanical hypernociception, joint edema and neutrophil recruitment, and at 0.03 g/kg also inhibited articular TNF-α synthesis after intra-articular zymosan injection. Isoorientin, isovitexin, rutin and isoquercitrin were identified in the most active fraction (F2-A), along with luteolin and apigenin derivatives, tentatively identified as isoorientin-2″-O-glucoside and isovitexin-2″-O-glucoside. CONCLUSION: This study corroborates the popular use by oral route of aqueous preparations of C. hololeuca against joint inflammatory disorders, such as rheumatoid arthritis. Our results demonstrated for the first time that oral administration of rutin shows antinociceptive and anti-inflammatory effects in ZIA, indicating that this flavonoid is one of the immunomodulatory compounds involved in the anti-arthritic activity of C. hololeuca.

Acute effect of Copaifera reticulata Ducke copaiba oil in rats tested in the elevated plus-maze: an ethological analysis.

OBJECTIVES: Copaiba oil oleoresin exuded from Copaifera reticulata Ducke (CRD) is commonly used in anti-inflammatory, healing and anti-tumoral folk medicines. The purpose of this study was to investigate the putative anxiolytic effect of acute administration of CRD. METHODS: CRD was administered (100, 400 and 800 mg/kg, p.o.) to male Wistar rats submitted to the elevated plus-maze model of anxiety using an ethopharmacological analysis. KEY FINDINGS: In comparison with control rats, CRD increased the percentage of entries in the open arms over the entire dose range tested (vehicle, 33.6 +/- 4.5; CRD 100 mg/kg, 44.67 +/- 3.68; CRD 400 mg/kg, 47.2 +/- 2.3; CRD 800 mg/kg, 50.7 +/- 2.2) and the percentage of time spent in the open arms of the elevated plus-maze at the highest dose (800 mg/kg) (vehicle, 26.4 +/- 5.7; CRD 800 mg/kg, 52.0 +/- 2.7). A standard anxiolytic, diazepam (3 mg/kg, p.o.), was used as a positive control. In a similar way, diazepam increased the percentage of entries and time spent in the open arms when compared with vehicle (% open entries: vehicle, 45.4 +/- 1.3; diazepam, 50.7 +/- 1.9; % time spent in open arms: vehicle, 28.2 +/- 0.9; diazepam, 38.9 +/- 1.2). Regarding ethological measures, CRD at the highest dose (800 mg/kg) reduced peeping out (anxiety-related behaviour) (vehicle, 3.1 +/- 0.6; CRD, 0.9 +/- 0.2) and increased end-arm activity (vehicle, 0.2 +/- 0.2; CRD, 2.0 +/- 0.4), indicating an enhanced tendency of the rats to explore actively the potentially dangerous areas of the maze. Diazepam decreased peeping out (vehicle, 3.3 +/- 0.3; diazepam, 1.0 +/- 0.2) and flat-back approach (vehicle, 0.8 +/- 0.2; diazepam, 0.2 +/- 0.1) and increased end-arm activity (vehicle, 0.3 +/- 0.1; diazepam, 2.5 +/- 0.3) and head-dipping (vehicle, 8.2 +/- 0.4; diazepam, 12.0 +/- 0.5). CONCLUSIONS: These data showed, for the first time, that acute treatment with CRD copaiba oil produced a dose-dependent anxiolytic-like effect over the dose range tested, on conventional and ethological parameters, without adversely affecting general activity levels.