Post-mortem high-energy phosphate and glycolytic changes in two skeletal muscles of the ox [proceedings].

to normal rats deprived of food for 6h, and the stomach contents were collected after 15min. About 10-12% of the radioactivity of the dose administered was found in the stomach contents, which, when passed through the Sepharose 6B column, gave 65-75 % of the radioactivity in the lipid peak fractions (4-10) collected just after the void volume, as shown in Fig. 1 . The position of this peak is identical with that observed in the preparation and separation of liposomally entrapped insulin from free insulin, and it corresponds to the liposomes. We have concluded, therefore, that most of the insulin in still entrapped within liposomes in the stomach contents so examined. The remaining 25-35 % of the radioactivity found in the fractions between 18 and 30 (Fig. 1) we assume to be hydrolysed products of the '2sl-labelIed insulin, as the free '251-labelled insulin is normally eluted between fractions 13 and 20. Moreover, the radioactivity in fractions 18 to 30 (Fig. 1) is not precipitated with 10% (w/v) trichloroacetic acid. Most of this radioactivity probably originates from the '251-labelled insulin associated with the outside surface of the liposomes, which was not truly entrapped during the preparation of insulin-containing liposomes or from degraded liposomes. In both cases such insulin would be readily hydrolysed by the proteolytic enzymes of the stomach. To investigate the action of intestinal enzymes on liposomally entrapped 1Z51-labelled insulin and to compare it with the free hormone, incubations with rat intestinal washings were carried out. Although extensive degradation (as assessed by loss of trichloroacetic acid precipitability) of free insulin occurs (60% in Smin), as would be expected, very little degradation of liposomally entrapped insulin occurred under similar conditions (1 5 % in 25 min). Further, the rapid degradation of the free insulin in the presence of rat intestinal washings was inhibited in the presence of liposomes containing no insulin. A similar effect was observed when purified trypsin was incubated with liposomally associated or entrapped insulin; here again, the degradation of the hormone was much less than that obtained by incubation without liposomes. Thus these preliminary studies show that some at least of the liposomes given by the oral route to rats remain intact in the stomach, and thus protect the entrapped protein from proteolytic digestion by gastric enzymes. Proteolytic activity of the intestine appears to be inhibited by liposomes.