Zonulin elucidated as a thoroughly known protein, is capable of modulating the gut integrity of intercellular connections. Intestinal permeability and its modulation by zonulin have been well-defined. Zonulin levels could increase in response to several stimuli, i.e. infection/gluten ingestion. Even if the latter occurs, zonulin signals into the body for elevating the permeability of the gut lining, permitting larger molecules to pass through. All aforementioned conditions initiate inflammation. In the present prospective field study, the aim was to determine the specificity of zonulin as a noninvasive selected biomarker of gut barrier function to identify and debug calves suffering from diarrhea. Furthermore, another purpose was to define the appearance of leaky gut (LeaG) among calves with diarrhea. By use of commercially available Bovine Zonulin ELISA test kits with a well-designed methodology all 11 diseased and other relevant healthy calves gave positive test results. Circulating zonulin levels (ng/mL) expressed as (±SEM), there were significant differences (p<0.001) between healthy (26.43±3.528) and diarrheic calves (57.97±4.250). As a preliminary conclusion, it should not be unwise to draw the hypothesis that zonulin levels debug diarrheic calves from healthy ones. Further studies are warranted.