Korea Hydro & Nuclear Power Company, Ltd., has operated the Wolsong Tritium Removal Facility (WTRF) since 2007 to reduce tritium concentrations in the moderator and coolant of the Wolsong nuclear power plant. As a result of the WTRF operation, the concentration of tritium in the moderator and coolant significantly decreased from 2320 to 9.3 GBq/kg. In particular, the tritium concentrations of the radioactive waste directly affected by radioactivity in the moderator and coolant were reduced by up to 99% during the WTRF operation. For this purpose, a chemical separation and quantification method for tritium separation was developed, and its average recovery yield was 98%.