HPLC-DAD analysis, antinociceptive and anti-inflammatory properties of the ethanolic extract of Hyptis umbrosa in mice

Abstract

Hyptis umbrosa (syn. Mesosphaerum sidifolium) (Lamiaceae Family) has been used to treat several conditions such as gastrointestinal disorders, skin infections, nasal congestion, fever and cramps. The objective of this study was to evaluate the chemical composition, analgesic and anti-inflammatory profiles of ethanol extract from leaves of Hyptis umbrosa (EEB). HPLC-DAD was used to determine the fingerprint chromatogram of the extract. Male Swiss mice were orally pretreated with EEB (100, 200 or 400 mg/kg; 60 min before initiating algesic stimulation) and antinociceptive activity was assessed using the acetic acid-induced writhing model, formalin test and hyperalgesia induced by glutamate or capsaicin. Also, peritonitis was induced by the intrathoracic injection of carrageenan to quantify the total number of leukocytes. The presence of phenolic compounds in the extract was confirmed using HPLC-DAD. The treatment with EEB, at all doses, produced a significant analgesic effect against acetic acid-induced antinociceptive activity. In the formalin test, only the 400-mg/kg-dose of EEB had a significant effect in the first phase. However, all doses tested were able to reverse nociception in the second phase. The effect of all doses of EEB also showed a significant antinociceptive effect in the glutamate and capsaicin tests and inhibited the carrageenan-induced leukocyte migration to the peritoneal cavity. The present study suggests that the EEB possesses peripheral analgesic action and showed potential in reducing the spreading of the inflammatory processes. Also, it seems to be related with vanilloid and glutamate receptors.

Keywords: inflammatory pain; medicinal plants; nociception; phenolic compounds.

Table 1. Table 1: Gradient system used in the analysis through HPLC-DAD

Table 2. Results obtained after the analysis of the analytical standards

Figure 1. Figure 1: Chromatogram of H. umbrosa ethanolic extract (270 nm)

Figure 2. Effect of EEB of H. umbrosa on the acetic acid-induced writhing test in the absence and presence of naloxone in mice. Vehicle (control), EEB (100, 200 and 400 mg/kg) or morphine (MOR) were administered p.o. 1 h before acetic acid injection. Pre-treatment with naloxone (NAL, 1.5 mg/kg, i.p.) was performed 0.5 h before treatment (i.p.) with EEB (400 mg/kg), or MOR (3 mg/kg). Each column represents mean ± S.E.M. (n = 6, per group). *p < 0.05 or ***p < 0.001 versus control. ###p < 0.001 versus MOR group (ANOVA followed by Tukey's test).

Figure 3. Effects of EEB of H. umbrosa on the formalin-induced nociception in mice. Vehicle (control), EEB (100, 200 and 400 mg/kg) or morphine (MOR) were administered p.o. 1 h before formalin injection. (A) Represents the first phase and (B) represents second phase of formalin-induced nociception. Each column represents mean ± S.E.M. (n = 8, per group). *p < 0.01, ** p > 0.01 or ***p < 0.001 versus control (ANOVA followed by Tukey's test).

Figure 4. Effects of EEB of H. umbrosa on the glutamate-induced nociception in mice. Vehicle (control), EEB (100, 200 and 400 mg/kg) or morphine (MOR) were administered p.o. 1 h before glutamate injection. Each column represents mean ± S.E.M. (n = 8, per group). ***p < 0.001 versus control (ANOVA followed by Tukey's test).

Figure 5. Effects of EEB of H. umbrosa on the capsaicin-induced nociception in mice. Vehicle (control), EEB (100, 200 and 400 mg/kg) or morphine (MOR) were administered p.o. 1 h before capsaicin injection. Each column represents mean ± S.E.M. (n = 8, per group). **p < 0.01 or ***p < 0.001 versus control (ANOVA followed by Tukey's test).

Figure 6. Figure 6: Effect of EEB of H. umbrosa on leukocyte migration into the peritoneal cavity induced by carrageenan in mice. Groups of rats were pre-treated with vehicle (control), dexamethasone (Dexa, 2 mg/kg, s.c.) or EEB (100, 200 and 400 mg/kg) 60 min before carrageenan (500 µg/cavity, 500 µL, i.p.)-induced peritonitis. Cell counts were performed at the time 4 h after the injection of carrageenan. Each value represents the mean ± S.E.M. **p < 0.01 or ***p < 0.001 related to control group. ANOVA followed by Tukey's test.