Study On The Separation And Analgesic Mechanlism Of The Activity Analgesic Composition From Earthworm

The earthworm has a variety of active ingredients, and the study of earthworms analgesic ingredients has become a hot spot gradually. It is reported that people has isolated from the earthworm extracts,having analgesic activity, but the the earthworms analgesic component analgesic mechanism of action has not been reported. This study is to isolated and purified earthworms analgesic active ingredient, and the analgesic mechanism of preliminary exploration.1Content and methods(1) Separation and Purification:Fresh earthworm tissue homogenates were centrifuged, through3KD and50KD ultrafiltration tube retention, and then gel filtration chromatography through Sephadex G-50, CM Sephadex C-50cation exchange chromatography, high performance liquid chromatography separation and purification, validate and screen the analgesic activity of the fractions at all levels with the injury model of sciatic nerve of rat.(2) Study on Analgesic mechanism:Through the experiment of the peripheral parts (acetic acid writhing, formalin induced pain experiment), the experiments of central parts of the analgesic (hot-plate, hot bath flick), opioid receptor-dependent experiments, earthworms analgesic components of experimental pain model rat monoamine neurotransmitters affect earthworms and the affect of analgesic ingredients NOS activity on the experimental pain model mice, discovering the analgesic effect and its mechanism.2The results(1) Through the homogenizer, high-speed centrifugation,ultrafiltration tube entrapped Sephadex G-50gel filtration chromatography, four protein peak was separated, the EAEC-I and EAEC-IV has significant analgesic activity by rat sciatic nerve injury model filter; the EAEC-I the EAEC-IV components respectively over CM Sephadex C-50cation exchange chromatography, the the EAEC-I was isolated of the two components, and the EAEC-IV three components, EAEC-IA and EAEC-IVB has significant analgesic activity after sciatic nerve injury model validation,, and then through the high-performance liquid chromatography, four peaks were isolated, EAEC-IA1and EAEC-IVB2, has provenobvious analgesic activity. Through Tricine-SDS-PAGE protein electrophoresis measured And molecular weight of the EAEC-IA1and EAEC-IVB2were21.6KD28.2KD.(2) Earthworm analgesic ingredient can significantly inhibit acetic acid-induced writhing response and formalin induced pain model II pain in a dose-dependent manner, indicating that earthworms and analgesic ingredients peripheral analgesic effect; earthworm analgesic composition can extend smallthermal latency of the the rat pain response and thermal bath mice, but was not significant difference in analgesic component of earthworm certain analgesic effect, the effect is not obvious; experiment, earthworms by the opioid receptor-dependent analgesic ingredient townthe pain effect does not depend on the opioid receptor; earthworm analgesic composition may be provided by reducing pain model in rat serum NE and5-HT content to play the outer peripheral analgesic played by raising the content of NE and5-HT in the brain tissue the central analgesic effect of; earthworm analgesic composition can significantly reduce pain model mice serum and brain tissue NOS activity, analgesic effect.3The Conclusion(1)In this study, a significant analgesic effect ingredient EAEC-IA1the EAEC-IVB2was isolated and purified from the earthworm extract, and the molecular weight isolated and purified were21.6KD28.2KD.(2) Earthworms analgesic composition has significant outer peripheral analgesic effect by decreasing peripheral5-HT, NE content, reducing peripheral NOS activity, reducing NO production; earthworms analgesic ingredients also have some analgesic effect by increase central5-HT, reduce the content of NE, improve NOS activity; earthworm analgesic ingredient analgesic effect of opioid receptor-dependent.