The dual sugar test (lactulose plus mannitol) is an unreliable indicator of feed withdrawal-associated changes in intestinal permeability in broiler chickens

Abstract

Despite its importance in poultry research, there is lack of standardized and practical techniques to measure intestinal permeability in a noninvasive manner. Therefore, this research sought to standardize a procedure using lactulose (Lac) and mannitol (Man) to measure intestinal barrier function in broilers. Twenty-one-day-old male and female Ross 308 birds were orally gavaged (either 2 mL/kg BW or fixed 3 mL per bird) with a solution containing 5 to 25 g Lac and 1 to 5 g Man dissolved in pure water to reach 100 mL of final solution. Feed withdrawal (FW; 2-24-h duration) prior to dosing with Lac and Man (LacMan) was mainly used to induce graded intestinal permeability. Blood samples were collected at 60-, 90-, or 120-min after LacMan dosing using serum or plasma (K2EDTA and/or Na-Heparin) blood tubes. Lac and Man concentrations were quantified by HPLC. Plasma samples collected 90-min after LacMan dosing elicited the least variable response (22.4% vs. 22.8% or 23.4% CV when compared with 60- and 120-min sampling time-points, respectively), and both markers were detectable after administering a solution containing the lowest concentration of Lac and Man. However, analytical problems arose when using Na-Heparin anticoagulant as high glucose levels interfered with Lac quantification. Upon improving the chromatographic technique, it became evident that a 24-h FW increased (P < 0.01) Lac concentrations. In the last trial, a more severe glucose interference was observed, resulting in no Lac detection within an entire treatment group. Twelve hours of FW increased (P < 0.01) Man concentration in the plasma of birds receiving the solution containing 3 g Man, but had no effect on the birds receiving the solution containing 5 g Man. A 24-h FW did not affect the Man concentrations in birds receiving the solution containing either 3 or 5 g Man. With inconsistency of Lac detection throughout our trials, it was concluded that the combination of Lac and Man is an unreliable marker to predict intestinal barrier function in broilers.