The larval nutritional state of the honey bees is controlled by the Royal Jelly. In addition, it stimulates the regeneration of tissues in other organisms, which has been one of the reasons it uses for cosmetics and as a nutritional ingredient. For the manufacturing and storage of products containing Royal Jelly, heat processing has played a significant role but it has not yet been clarified as a technical factor. The aim of the research was to investigate the effects of heating on the physiological activity of Royal Jelly. We studied the proteins at the native and denatured conditions using heating by the electrophoresis methods. Also, the experiments in vitro were applied when the diets included the non-heated and heated at 52, and 72°C Royal Jelly. The study determined that heating it at 52°C within 5 minutes didn’t change the visible protein composition. This diet was sufficient for the rearing of larvae and hatching of 78.7% against 83.3% of A. mellifera honey bees fed by the diet with non-heated Royal Jelly in the in vitro experiment. However, the weights of the hatched honey bees had significant differences (α=0.05) and were 97.42±14.45 and 102.09±12.49 mg, respectively. This study adds to our understanding of the upper limit of the heat resistance of Royal Jelly proteins by demonstrating the physiological activity of the components after heating at 52°C. So, this temperature can be applied in the protocol of the manufacturing of the Royal Jelly products.