Effects Of The Fibrous Root Of Rhizoma Coptidis And Its Main Alkaloids On Non-specific Immunity And Disease Resistance Of Common Carp

Objective:Rhizoma Coptidis(RC), an important antipyretic and wetnessdrying traditional Chinese medicine, has been widely used in veterinary clinics due to its antibacterial and antipyretic activities. However, to the best of our knowledge, the effect of RC on the immunity function of macrophages and non-specific immunity of common carp has not been reported. In this study, we focused on the utilization of its fibrous root(FRC), a by-product in the processing of RC, which was always abandoned in the field by farmers. In order to promote the utilization of the Rhizoma Coptidis, common carp were used as the animal model,then the effects of the FRC and its main alkaloids on non-specific immunity and disease resistance of common carp were investigated. Methods and results:(1) Contents of main alkaloids in the fibrous root of Rhizoma Coptidis(FRC) were measured by HPLC, then the main alkaloids from FRC were isolated by high speed counter-current chromatography.(2) The MICs of the extract of FRC and its main alkaloids against Aeromonas hydrophila and Vibrio parahemolyticus were tested to explore their antibacterial activities. Results found that FRC, total alkaloids(TA), BBR, Cop all showed antibacterial activity against Aeromonas hydrophila and Vibrio parahemolyticus. Among FRC alkaloids, the total alkaloids from FRC was the strongest, followed by berberine, coptisne and FRC.(3) The effects of the four alkaloids isolated from FRC on the non-specific immunity of mouse peritoneal macrophages in vitro were investigated. Results showed that the four alkaloids could increase the phagocytosis of macrophages in different level and berberine had the best effect. Berberine(BBR), coptisine(Cop) and palmatine(pal) had up-regulation effects on respiratory burst activity of mouse peritoneal macrophages. The results suggested that the four alkaloids have the potential non-specific immune pharmacological activities.(4) The effects of FRC and its main alkaloids on serum non-specific immunity parameters, respiratory burst and phagocytic activity of head kidney macrophage were investigated. The fish were randomly assigned to seven groups: normal control group(NC), groups treated with 12.5 g/kg FRC(FRC-L), 25 g/kg FRC(FRC-M), 50 g/kg FRC(FRC-H), 0.78 g/kg total alkaloids(TA), 0.78 g/kg berberine(BBR), and 0.78 g/kg coptisine(Cop), respectively. Results showed that, only the activities of myeloperoxidase(MPO), respiratory burst and phagocytic activity were significantly increased in FRC-M group when compared with NC(P < 0.05) after 14 consecutive days feeding. On the 21 th day, the activities of lysozyme(LZM), MPO and respiratory burst were enhanced by FRC-M, FRC-H, TA, BBR and Cop as compared with NC. For C3 and phagocytic activities, the groups of TA and BBR reached the significant differences.(5) To explore the molecular mechanisms for the disease resistance of FRC and its main alkaloids, the expression profiles of the immune-related genes(IL-1β, IL-10, C3, lysozyme-c and TNF-α mRNA) were examined. The results showed that the FRC-M, FRC-H, TA and BBR could significantly up-regulate the mRNA expression of IL-1β in the head kidney of the common carp. The mRNA level of TNF-α was significantly increased in FRC-L, FRC-M, FRC-H, TA, BBR and Cop group. The mRNA expression of lysozyme-c significantly increased in FRC-M,FRC-H, TA, BBR and Cop group when compared with NC. The mRNA expression of C3 mRNA was significantly up-regulated in the FRC-M, TA and BBR group when compared with the NC group, whereas IL-10 expression was significantly down-regulated in FRC-L, FRC-M and FRC-H group.(6) At the end of the experiments, the effects of FRC and its main alkaloids on the survival rate of common carp infected with Aeromonas hydrophila were investigated. Results showed that FRC-M, FRC-H, TA, BBR and Cop significantly enhanced the survival rate of common carp after challenged with Aeromonas hydrophila ten days, when compared to NC group. Conclusion:FRC could restrain bacteria growth, enhance the non-specific immunity and disease resistance of common carp and the main alkaloids contained in FRC may contribute to these effects. These results suggest that the FRC is a promising feed additive for common carp in aquaculture industry.