Abstract: Major Royal Jelly Proteins ( MRJPs) was extracted by centrifugation from royal jelly ( RJ) .The orthogonal experiment optimized with four parameters, ratio of solution to royal jelly, pH, extracting time and ionic strength was implemented.The quality of MRJPs, the trans-10-hydroxy-decenoic and total sugar in by-product were analyzed.Orthogonal experiment showed the factors affecting MRJPs extraction rate in descending order of size were: ionic strength > pH > extracting time > ratio of solution to RJ.81.14% of MRJPs was obtained from RJ under the optimum extraction conditions ( the ratio of solution to RJ 8mL /g, pH8, extracting time 8h, ionic strength 0.5mol /L) .Analysis of SDS-PAGE and ELISA of extracted MRJPs showed that it possessed molecular weight ranged from 42.1 to 97.4ku, and that its purity reached 71.7%.The by- product contained 2.91% of trans-10- hydroxy-2- decenoic acid and1.26% of total sugar, respectively. The results provided essential scientific knowledge for development of MRJPs and utilization of its by-products.