S. A. Angarita2, T. A. Russell2, P. Ruchala3, S. Duarte2, I. A. Elliott2, J. P. Whitelegge3, A. Zarrinpar1 1University Of Florida,Surgery,Gainesville, FL, USA 2University Of California – Los Angeles,Surgery,Los Angeles, CA, USA 3University Of California – Los Angeles,Pasarow Mass Spectrometry Laboratory,,Los Angeles, CA, USA
Introduction: Intestinal barrier integrity loss plays a key role in the development and perpetuation of disease states such as inflammatory bowel and celiac disease. It also crucial to the onset of sepsis and multiple organ failure in situations of intestinal hypoperfusion, including trauma and major surgery, or in the setting of abnormal blood flow such as portal hypertension. A variety of tests have been developed to assess intestinal epithelial cell damage, intestinal tight junction status, and the consequences of intestinal barrier integrity loss, i.e. increased intestinal permeability. These methods suffer from a lack of sensitivity, a prolonged period of specimen collection, or high expense. We have developed a technique to measure the concentration of the nonabsorbable food dye FD&C Blue #1 from the blood and sought to apply this technique to assess its utility in measuring intestinal barrier function in humans.
Methods: Four healthy volunteers and ten subjects in the intensive care unit were recruited in accordance with an IRB approved protocol. Subjects were given 0.5 mg/kg Blue #1 orally or per nasogastric tube as an aqueous solution of diluted food coloring (0.5 mg/mL). Five blood specimens were drawn per subject (5 mL/draw): 0 hour – prior to dose, 1 hour, 2 hours, 4 hours, and 8 hours. The plasma was then extracted with an acidified mixture of isopropanol and acetonitrile. The organic extracts were then analyzed by high performance liquid chromatography/mass spectrometry looking for the presence of the unmodified dye.
Results: This study was performed in two phases. Phase one attempted to establish the lower limit of detection and measure the baseline/normal intestinal absorption of Blue #1. To do so four healthy subjects were recruited. We found no detectable absorption. In Phase 2, ten patients in the intensive care unit were recruited. Six patients met criteria for septic shock (identified by a vasopressor requirement to maintain a mean arterial pressure of 65 mm Hg or greater in the absence of a hypovolemic or cardiogenic etiology). The septic patients demonstrated significantly greater absorption of Blue #1 after 2 hours and 8 hours.
Conclusion: We have developed a novel, easy-to-use method to measure intestinal permeability. The method utilizes a food grade non-absorbable dye that can be detected by mass spectrometry analysis of patient blood at multiple time points following oral consumption. This method would allow for the measurement of the intestinal permeability of patients at risk for sepsis, organ failure, or other conditions where loss of function of the intestinal barrier could lead to adverse symptoms or secondary effects.
