Intrajejunal Absorption of Liposomally Entrapped Insulin in Normal Man

Since the discovery of insulin, many investigators have explored the possibility of developing a n effective method for its oral administration. Attempts t o secure absorption of insulin from the gastrointestinal tract using adjuvants have been unsuccessful from a therapeutic point of view, but it has become generally accepted that insulin can be absorbed from the intestine of mammals and some hypoglycaemic activity of the absorbed insulin observed (Laskowski et al., 1959; Inouye & Mars, 1962; Speth & Christian, 1963; Engel et al., 1968; Galloway & Root, 1972). Patel & Ryman (1976, 1977) and Dapergolas et al. (1976) have demonstrated that liposomally entrapped insulin, when given orally, produces hypoglycaemia in both diabetic and normal rats. Thus liposomes not only seem t o protect insulin from proteolytic degradation in the stomach and intestine, but also act as structurally organized vehicles with endocrine therapeutic potential. This has led us t o investigate whether absorption of liposomally entrapped insulin occurs in the intestine of normal man. Liposomes containing insulin were prepared by the method of Patel & Ryman (1976) with the following modifications. Neutral liposomes were prepared by dissolving egg phosphatidylcholine/cholesteroI (1 : 5, w/w) in chloroform. To the dry lipid was added a mixture of crystalline ox insulin (2mg/ml) and bovine serum albumin (1 mgiml), prepared in 10mM-potassium phosphate containing 80m~-NaC1, p H 7.4 (phosphate/saline buffer). Liposomes thus formed were sonicated for 8 min, with cooling, a t 4°C using a 19mm titanium probe at 0.15nm in an MSE 150W sonicator. The sonicated liposomes were passed through a Sepharose 6B column, equilibrated with the phosphate/saline buffer, and the lipid fractions with insulin associated were collected and stored under N2 at 4°C. Preparations were transported from London t o Glasgow by rail and were received on the day of preparation and used shortly afterwards. For this pilot study, volunteers were healthy male medical graduates, who acted as their own controls. Liposomally entrapped insulin (20-200 units) a t body temperature were administered quickly into the jejunum by syringe through a radio-opaque tube 1.5mm in diameter. This tube was taken orally, retained by a small mercury-filled terminal balloon and located in the jejunum by image-intensified fluoroscopy. Each intubation was performed between 08:30 and 09:30h, the subject having fasted for 10h previously. Samples of venous blood were withdrawn and collected in lithium-heparin tubes. Blood glucose was assayed by the glucose oxidase method, and plasma insulin by immunoassay using Wellcome reagents. In the first subject 22 units of liposomally entrapped insulin were administered intrajejunally and there appeared to be a small but progressive rise in concentration of plasma immunoreactive insulin, but no significant change in blood glucose concentration. The following five subjects were given up to 200 units of liposomallyentrapped insulin. Again a small increase in the concentration of plasma immunoreactive insulin was observed, but generally insignificant changes were seen in blood glucose concentration. Control liposomes (without insulin) had no effect upon the concentration of either plasma immunoreactive insulin or of blood glucose. We have estimated that about 1 of the administered dose of insulin appears in the plasma by 60min in our normal subjects. This study used a range of insulin doses known to be effective parenterally and has not yielded overt clinical hypoglycaemia. Nevertheless, indications are that insulin is taken up by normal males from the liposomal complexes, but presently acts a t too