Abstract: The androgenic gland was collected from male Macrobrachium rosenbergii,then was homogenized with 0.1mol / L NaAc soluhon. The homogenate was centrifgated 15min at 12000 r /min, the upper layer clear solution was adjusted to pH4.5,then was centrifugated again and the precipitate was collected. This precipitate was separated with SDS-PAGE method. The crude product of the isopada AGH analogue (Androgenic Gand Hormone) of M. rosenbergii was collected from the protein belt which molecular weight range from 17k Da to 18k De by SDS-PAGE method. The applications of new techniques such as HPLC, and MALDL-TOF-MS were carried out.Matrix-assisted laser desorption ionization time-of -flight mass spectrometry and HPLC-ESI-MS methods may provide a more sensitive and accurate determination for the molecular weight of AGH. The molecular weight of the isopada AGH analogue is 17480u as identified by HPLC-ESI-MS and 17436?10u by capillary electrophoresis.