Antinociceptive activity—Formalin test

The mice were placed in an open Plexiglas observation chamber for 30 min to acclimate to their surroundings. They were then removed and gently restrained while 20 μL of 2.5% formalin (1:100 dilution of stock formalin solution, 37% formaldehyde in 0.9% saline) was injected subcutaneously into the dorsal surface of the hind paw using a 30 gauge needle. Following injection, the mice were returned to the observation chamber for a 30 min observation period. The nociceptive score was determined by counting the time the animal spent licking the injected paw during the early phase (0–10 min) and the late phase (10–30 min) [28]. The effects of ME (1.95–125 mg/kg) and its ether, ethyl acetate and chloroform fractions (100 mg/kg) were evaluated on the formalin test. Next, gallic acid, methyl gallate and 1,2,6-tri-O-galloyl-β-d-glucopyranose (0.19–200 mg/kg) obtained from the ethyl ether fraction were also evaluated. All treatments were administered once, by intraperitoneal route, 40 min before the injection of formalin. Indomethacin (10 mg/kg) and morphine (5 mg/kg) were used as the reference drugs. Vehicle group was treated with saline or 5% DMSO plus saline, as appropriate.