Protein eletropherogram of haemolymph from Apis mellifera L. bees submitted to royal jelly production

Abstract

The objective of this research was to study the eletropherogram of the proteins of the haemolymph in Apis mellifera L. worker bees by utilizing eletroforese in polyacrylamide gel. Haemolymph samples were obtained from 18 beehives of africanized honey bees and their hybrids, italian and carniolas, which were submitted to royal jelly production, either with or without access to 20% pollen protein. Haemolymph sample from recently emerged adult bees aged, 12, 18 and 24 days was obtained. These samples were extracted by introducing a 50 μL micropipette tip into the dorsal heart of each bee, through the intrasegmental membrane, between the third and the fourth abdomen segments. In order to inhibit the tirosinase activity, haemolymph samples were treated with three drops of fenyltio-ureia. The dosage absorbance rate of protein for each sample were determined by the use of the spectrophotometer and the biochemical biureto method. The absorbance rate was measured at 540 nm. The PhastSystem and 8-25% gradient mini polyacrylamide gels were used in the sodium dodecyl sulfate polyacrylamide electrophoresis (SDS-PAGE) technique. Five hundred ng of protein were applied to each well. After one run of the electrophoresis, the gels were stained with silver according to the manufacturus specifications. In the haemolymph of newly emerged adult bees, the electrophoresis pattern is evenly distributed among protein bands of low (60 - 80 Kd), medium (100 - 150 Kd), and high (200 - 250 Kd) molecular weight, with a total of 27 bands. With the advance of age, the haemolymph electrophoretic pattern stabilizes in protein bands of high and medium molecular weight, with a total of 15 to 17 bands. No differences among treatments were observed (P > 0.05).