RESUMO
Copaifera pubiflora Benth oleoresin (CPO) is used as an anti-inflammatory, wound healing, and antimicrobial. This paper reports the cytotoxic, anti-inflammatory, and antinociceptive activities of CPO. CPO (10 mg/kg) did not affect locomotor capacity in the open-field and rotarod tests and was not cytotoxic to CHO-k1, THP-1, and L929 cell lines. It was active in the formalin test at 3 mg/kg by 86 ± 3% and 96 ± 3%, respectively, for the first and second phases. At 10 mg/kg, CPO inhibited 90 ± 7%, the pain in the mechanical hyperalgesia test. In the tail-flick test, CPO at 3 mg/kg affected the tail-flick latencies in mice by 77 ± 20%, which in combination with naloxone was only partially reduced. At 3 mg/kg CPO inhibited 80 ± 12% the carrageenan-induced paw edema, and at 3 mg/kg it reduced by 91 ± 5% the nociception on acetic acid-induced abdominal writhing. Therefore, CPO possesses anti-inflammatory and antinociceptive activities.
Assuntos
Analgésicos , Fabaceae , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Edema/induzido quimicamente , Edema/tratamento farmacológico , Camundongos , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêuticoRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Copaifera species folkloric names are "copaíbas, copaibeiras, copaívas or oil stick", which are widely used in Brazilian folk medicine. Among all ethnopharmacological applications described for Copaifera spp oleoresins, their anti-inflammatory effect stands out. However, the knowledge of anti-inflammatory and antinociceptive properties of Copaifera pubiflora Benth is scarce. AIM OF THE STUDY: To investigate the cytotoxic, anti-inflammatory, and antinociceptive activities of C. pubiflora oleoresin (CPO), and its major compound ent-hardwickiic acid (HA). MATERIAL AND METHODS: The phosphatase assay was used to evaluate the cytotoxicity of CPO and HA in three different cell lines. CPO and HA doses of 1, 3, and 10 mg/kg were employed in the biological assays. The assessment of motor activity was performed using open-field and rotarod tests. Anti-inflammatory activity of CPO and HA was assessed through luciferase assay, measurement of INF-γ, IL-1ß, IL-6, IL-10, and TNF-α in a multi-spot system with the immortalized cell line THP-1, zymosan-induced arthritis, and carrageenan-induced paw edema. Acetic acid-induced abdominal writhing and formalin tests were undertaken to evaluate the antinociceptive potential of CPO and HA. In addition, the evaluation using carrageenan was performed to investigate the effect of CPO in pain intensity to a mechanical stimulus (mechanical hyperalgesia), using the von Frey filaments. A tail-flick test was used to evaluate possible central CPO and HA actions. RESULTS: In the cytotoxicity evaluation, CPO and HA were not cytotoxic to the cell lines tested. CPO and HA (10 mg/kg) did not affect animals' locomotor capacity in both open-field and rotarod tests. In the luciferase assay, CPO and HA significantly reduced luciferase activity (p < 0.05). This reduction indicates a decrease in NF-κB activity. HA and CPO decreased INF-γ, IL-1ß, IL-6, IL-10, and TNF-α at 24 and 72 h in the multi-spot system. In zymosan-induced arthritis, CPO and HA decreased the number of neutrophils in the joint of arthritic mice and the number of total leukocytes (p < 0.05). In experimental arthritis HA significantly decreased joint swelling (p < 0.05). CPO and HA also increased the mechanical threshold during experimental arthritis. HA and CPO significantly inhibited the carrageenan-induced paw edema, being the doses of 10 mg/kg the most effective, registering maximum inhibitions of 58 ± 8% and 76 ± 6% respectively, p < 0.05. CPO and HA reduced the nociceptive behavior in both phases of formalin at all tested doses. The highest doses tested displayed inhibitions of 87 ± 1% and 72 ± 4%, respectively, p < 0.001, in the first phase, and 87 ± 1% and 81 ± 2%, respectively, p < 0.001, in the second phase. Oral treatment of CPO and HA (1, 3, 10 mg/kg) significantly reduced the nociceptive response in acetic acid-induced abdominal writhings, and the 10 mg/kg dose was the most effective with maximum inhibitions of 86 ± 2% and 82 ± 1%, respectively, p < 0.001. Both HA and CPO significantly decreased the intensity of mechanical inflammatory hyper-nociception on carrageenan-induced hyperalgesia at all tested doses, and 10 mg/kg was the most effective dose with maximum inhibitions of 73 ± 5% and 74 ± 7%, respectively, p < 0.05.CPO increased the tail-flick latencies in mice, and concomitant administration of naloxone partially reduced its effect. CONCLUSIONS: CPO and HA may inhibit the production of inflammatory cytokines by suppressing the NF-κB signaling pathway, resulting in anti-inflammatory and antinociceptive activities.
Assuntos
Analgésicos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Artrite Experimental/tratamento farmacológico , Diterpenos/uso terapêutico , Edema/tratamento farmacológico , Fabaceae/química , Extratos Vegetais/uso terapêutico , Ácido Acético/toxicidade , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Artrite Experimental/induzido quimicamente , Comportamento Animal/efeitos dos fármacos , Brasil , Carragenina/toxicidade , Linhagem Celular , Citocinas/metabolismo , Diterpenos/isolamento & purificação , Diterpenos/farmacologia , Edema/induzido quimicamente , Formaldeído/toxicidade , Humanos , Hiperalgesia/induzido quimicamente , Hiperalgesia/tratamento farmacológico , Locomoção/efeitos dos fármacos , Medicina Tradicional , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/metabolismo , Extratos Vegetais/farmacologia , Zimosan/toxicidadeRESUMO
The microorganisms that constitute the oral microbiome can cause oral diseases, including dental caries and endodontic infections. The use of natural products could help to overcome bacterial resistance to the antimicrobials that are currently employed in clinical therapy. This study assessed the antimicrobial activity of the Copaifera pubiflora oleoresin and of the compounds isolated from this resin against oral bacteria. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assays provided values ranging from 6.25 to > 400⯵g/mL for the C. pubiflora oleoresin and its isolated compounds. The fractional inhibitory concentration index (FICI) assay showed that the oleoresin and chlorhexidine did not act synergistically. All the tested bacterial strains formed biofilms. MICB50 determination revealed inhibitory action: values varied from 3.12-25⯵g/mL for the oleoresin, and from 0.78 to 25⯵g/mL for the ent-hardwickiic acid. Concerning biofilm eradication, the C. pubiflora oleoresin and hardwickiic acid eradicated 99.9 % of some bacterial biofilms. Acid resistance determination showed that S. mutans was resistant to acid in the presence of the oleoresin and ent-hardwickiic acid at pH 4.0, 4.5, and 5.0 at all the tested concentrations. Analysis of DNA/RNA and protein release by the cell membrane demonstrated that the oleoresin and hardwiickic acid damaged the bacterial membrane irreversibly, which affected membrane integrity. Therefore, the C. pubiflora oleoresin and ent-hardwickiic acid have potential antibacterial effect and can be used as new therapeutic alternatives to treat oral diseases such as dental caries and endodontic infections.