Abstract: Objective: To explore the on-line detecting on the anti-oxidant activity of the alkaloids from the stems of Dendrobium huoshanense C. Z. Tang et S. J. Cheng by HPLC-UV-CL system,and provide references on researching the potency material base of D. huoshanense and on quick discovery of antioxidants from Traditional Chinese Medicine. Method: Applying the on-line HPLC-UV-CL analysis with Luminol-perhydrol chemiluminescence system as the donor of hydroxyl radicals to evaluate the anti-oxidant activity of the constituents of the total alkaloid extraction from D.huoshanense.The alkaloids were identified by knocking-out the alkaloid components using phosphomolybdic acid and iodine-potassium iodide as precipitation reagent. Result:Total four kinds of D.huoshanense alkaloids( the HPLC peak 6,peak 10,peak 11 and peak 14) were checked out and peak 14 and peak 11 showed remarkable hydroxyl-radical-cleaning activity with the calibration clearance values 10.94% and 4.95%,respectyively.No effect of the other two D.huoshanense alkaloids( peak 6 and peak 10)on the chemiluminescence value of Luminol-perhydrol system were determined by the established on-line HPLCUV-CL method. Conclusion: The on-line HPLC-UV-CL system could be used to evaluate the cleaving hydroxyl radicals activity of D.huoshanense alkaloids.