2.6. Acetic Acid-Induced Writhing Test

The test procedure followed the method as described by Pingsusaen et al., 2015 [24]. Six groups (n = 6) of rats were assigned for this experimental model; Group A: vehicle (control, saline, p.o.), Group B: indomethacin (10 mg/kg, p.o.), Groups C–E: PBME (100, 200, and 400 mg/kg, p.o., respectively), Group F: PBME (400 mg/kg, p.o.) + naloxone (5 mg/kg, p.o., treated 30 min prior of PBME administration). Thirty minutes post-dosing, 0.6% acetic acid (10 mL/kg) was injected intraperitoneally. The number of cumulative writhes was documented for 30 min. Inhibition percentage was calculated as in following equation: Inhibition (%) = ((A_Control – B_Test)/ A_Control) × 100, where, A is the mean writhes in control group and B is the mean writhes in test groups.