Presence of sialic acid in prothoracic glands of Galleria mellonella (Lepidoptera).

The presence of sialic acid (SA) in prothoracic glands (PGs) of Galleria mellonella was determined by the methods of electron microscopy (EM), histochemistry, spectrophotometry (SP) and electronic ionization (EI)-mass spectroscopy. Histochemical observations were carried out by the cationic dye ruthenium red (RR), staining with and without neuraminidase digestion in the larval stage. Neuraminidase-sensitive SA was demonstrated by the decrease in the amount of RR-binding following neuraminidase digestion. The total amount of SA was found to be 0.09016 mg g(-1) in dry tissue by spectrophotometric determination. EI-mass spectroscopy results confirmed the EM and SP observations. The fragmentation scheme derived from EI-mass analysis exhibited the presence of the lactonized form of Neu5Gc7, 9Ac(2). On the basis of the various pieces of evidence described above, it was firmly concluded that Neu5Gc7, 9Ac(2) molecules were present in PGs of G. mellonella.

Storage of rat liver for plasma membrane isolation.

We described a procedure for the preservation of rat liver which makes possible the isolation of plasma membranes after 10 days storage at -70 degrees C. The yield of plasma membranes obtained from the liver tissue kept at -70 degrees C for 10 days (3.43 +/- 0.08 mg protein/10 g wet liver) was not different statistically (P > 0.05) from the yield of freshly obtained plasma membranes (3.32 +/- 0.05 mg protein/10 g wet liver). However, a significantly low yield (2.65 +/- 0.08; P < 0.01) was obtained from 90 days stored rat liver when compared with the immediate isolation. Plasma membrane Na+, K+ ATPase and 5'nucleotidase activities of the stored liver for 10 days were not different statistically (P > 0.05) from the enzyme activities of the freshly isolated membrane fractions. In contrast there was a significant decrease (p < 0.0001) in the activities of both plasma membrane Na+, K+ ATPase and 5'nucleotidase activities of 90 days stored rat liver at -70 degrees C when compared with immediate isolation. Considering the electron microscopic findings; we observed that the preservation of the integrity of the plasma membrane fractions obtained from fresh and frozen livers for 10 and 90 days seemed to be parallel to the biochemical results. Therefore we suggest that, storage of rat liver tissue for 10 days make feasible to maintain the experimental design and give convenience for obtaining intact plasma membrane fractions.