Intestinal Permeability Measurement Protocol

Intestinal permeability was assessed as previously described (33 (link)). In brief, all subjects were asked to avoid consuming lactulose, sugar substitutes, diet foods, and foods that might contain mannitol or sucralose for 72 hours before the study visit. Subjects then fasted overnight. In the morning, the subjects orally ingested a test solution. The solution was a cocktail of 2 g of mannitol, 7.5 g of lactulose, and 2 g of sucralose in 300 mL of water. The subjects fasted for 2 hours after the start of urine collection and then were asked to eat normally except for refraining from lactulose, sugar substitutes, and sucralose or mannitol–containing products, during the 24-hour urine collection period. Subjects were asked to empty their bladder before consuming the test solution. After consuming the solution, subjects collected their urine for 24 hours. Two urine collections were recorded: 0–5 hours and 0–24 hours. Urine volumes were recorded and stored until analysis. Five-hour urinary lactulose, mannitol, and lactulose-to-mannitol (L/M) ratio are primarily markers of small bowel permeability; and 24-hour urinary sucralose and lactulose excretion are markers of total gut permeability, with sucralose primarily representing colonic permeability (34 (link)). This is because of both sucralose and lactulose being able to permeate through both the small and large intestines (colon). However, sucralose is not fermented by colonic bacteria, whereas 75% of lactulose and mannitol are fermented by colonic bacteria (35 (link)).
To examine the intestinal permeability immediately after exposure to an injurious agent (aspirin), we did a second assessment of intestinal permeability after an aspirin challenge, as we have previously published (18 (link)). In brief, 2 weeks after the baseline collection, subjects were asked to do a second collection and were given the sugar cocktail along with 4 tablets of aspirin each containing 325 mg for the aspirin challenge. Urine samples were analyzed by gas chromatography as previously described (33 (link)). The intestinal permeability was measured using an Agilent 6890 GC equipped with a flame ionization detector.