Abstract: The functional groups analysis methods,namely barium chloride (BaCl2) method and calcium acetate (Ca(CH3COO)2) method, were used to determine the content of functional groups of theabrownine for the first time, and suitable operating methods were established to determine the contents of total acidic groups, carboxyl groups and phenolic hydroxyl groups. The optimum conditions of determination were obtained as follows: for the BaCl2 method, sample weight 0.02-0.08 g, precipitation time 30-60 min, concentration of BaCl2 0.75 mol/L;for the Ca(CH3COO)2 method, sample weight 0.05-0.1 g, precipitation time 20 min, concentration of Ca(CH3COO)2 1 mol/L. The total acidic groups content of theabrownine from Pu-erh tea is 7.780 mmol/g, of which phenolic hydroxyl groups is 5.805 mmol/g and carboxyl groups is 1.975 mmol/g. The results indicate that phenol hydroxyl groups are the most important acidic groups in theabrownine. Both BaCl2 method and Ca(CH3COO)2 method are simple and require neither elaborate equipment nor special skills. With satisfactory results, these methods appear to be suitable for use in the routine analysis of functional groups in theabrownine.