To identify the active components of honeybee venom in production of inflammation and pain-related behaviors, five major peptidergic subfractions were separated, purified and identified from the whole honeybee venom. Among them, four active peptidergic components were characterized as apamin, mast-cell degranulating peptide (MCDP), phospholipase A(2) (PLA(2))-related peptide and melittin, respectively. All five subfractions were effective in production of local inflammatory responses (paw edema) in rats although the efficacies were different. Among the five identified subfractions, only MCDP, PLA(2)-related peptide and melittin were able to produce ongoing pain-related behaviors shown as paw flinches, while only apamin and melittin were potent to produce both thermal and mechanical hypersensitivity. As shown in our previous report, melittin was the most potent polypeptide in production of local inflammation as well as ongoing pain and hypersensitivity. To further explore the peripheral mechanisms underlying melittin-induced nociception and hypersensitivity, a single dose of capsazepine, a blocker of thermal nociceptor transient receptor potential vanilloid receptor 1, was treated s.c. prior to or after melittin administration. The results showed that both pre- and post-treatment of capsazepine could significantly prevent and suppress the melittin-induced ongoing nociceptive responses and thermal hypersensitivity, but were without influencing mechanical hypersensitivity. The present results suggest that the naturally occurring peptidergic substances of the whole honeybee venom have various pharmacological potencies to produce local inflammation, nociception and pain hypersensitivity in mammals, and among the five identified reverse-phase high pressure liquid chromatography subfractions (four polypeptides), melittin, a polypeptide occupying over 50% of the whole honeybee venom, plays a central role in production of local inflammation, nociception and hyperalgesia or allodynia following the experimental honeybee's sting. Peripheral transient receptor potential vanilloid receptor 1 is likely to be involved in melittin-produced ongoing pain and heat hyperalgesia, but not mechanical hyperalgesia, in rats.