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Isolation Of The Aqueous Phase Of Human Intestinal Contents During The Digestion Of A Fatty Meal.

H. P. Porter, D. R. Saunders
Published 1971 · Medicine

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Commonly used methods for isolating the aqueous phase of human intestinal contents after a fatty meal involve heating pooled samples of intestinal contents to 70 C to abolish lipolysis and spinning the heated sample at 100,000 g for 15 to 18 hr to separate the aqueous phase from the emulsion droplets. We found that this method introduces artifacts. (1) Heating batches of intestinal contents in a 70 C water bath increases free fatty acid markedly and alters phospholipid composition. (2) Spinning intestinal contents creates concentration gradients, which makes random sampling misleading because of the heterogeneous composition of the infranatant solution. A method is described which isolates the aqueous phase of intestinal contents by filtration through two series of Millipore filters. Clogging of filters is avoided by feeding a protein-free test meal. A 10-ml sample of intestinal contents can be passed within 30 sec through the first series of filters (mean pore diameter: 30,000; 8,000; 4,500; and 3,000 A). Release of free fatty acid in the resulting 3000-A filtrate is greatly reduced, because more than 95% of the triglyceride of intestinal contents is removed. Most of the turbidity in the 3000-A filtrates is removed when these filtrates are passed through the second series of filters (2200 and 1000 A). The resulting 1000-A filtrate is optically clear, or slightly turbid, and contains 89.7% of the bile salts of unfiltered intestinal contents. Experiments in which intestinal contents were labeled with radioactive cholesterol and bile salt indicate that the micellar association between these two lipids is not disrupted by filtration. Intestinal contents from the duodenojejunal junction of 9 normal subjects after a test meal contained 669 ± 106 (mean ± SD) μg of trypsin per ml and 8.6 ± 1.4 mm bile salts; the aqueous phase (1000-A filtrate) contained 7.7 ± 1.3 mm bile salts and 10.8 ± 3.5 mm free fatty acid.
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