The concentration of chloride ions in rumen fluid is a useful measure of obstructive gastrointestinal disease in ruminants and camelids. However, rumen fluid is very different from other biological fluids in its bacterial populations, consistency, and concentrations of various anions. Two methods of determining the chloride concentration in biological fluids were compared using centrifuged and filtered rumen fluid containing different amounts of sodium chloride. Although coulometric titration and potentiometric electrode analysis yielded results that had a strong linear relationship, the results of potentiometry were consistently and significantly higher, by about 20 mEq/L. This difference was investigated further by analyzing a series of fluids containing different concentrations of sodium acetate. Acetate was detected as chloride (0.21 chloride molecules per acetate molecule) by potentiometry but not by coulometric titration. Therefore, the acetate concentration of rumen fluid was the most likely cause of the discrepancy between tests in the original trial. In conclusion, the coulometric procedure may be more accurate than the potentiometric procedure for measuring rumen chloride when the concentrations of possible confounding ions are unknown.