To explore the structural characteristic and enzymatic mechanism for melanosis development of Litopenaeus vannamei during cold storage， the cell integrity in hepatopancreas tissue was monitored by histochemistry and transmission electron microscope. The results showed that the cell structure of hepatopancreas tissue raptured gradually as the increase of storage， thoroughly disintegrated on the 5th day. In addition， the image with low-field nuclear magnetic resonance indicated the fast increase in content of free water in tissue as the disruption of cell structure. Furthermore， the results of immunohistochemistry with Ca2+ fluorescence probe and serine protease demonstrated that Ca2+ and serine protease regularly migrated with disruption in cell structure and water migration. With combination of these driving forces， the pro-polyphenol oxidase was activated and the melanosis development was triggered and accelerated.