Anal fistula, characterized by abnormal tracts between the perianal skin and the anal canal, presents challenges in treatment because of its diversity and complexity. This study investigates the use of non-thermal plasma as a postsurgical therapy for anal fistula, aiming to promote healing and tissue regeneration. A specialized plasma reactor was designed to apply non-thermal plasma within the anorectal cavity practically. Non-thermal plasma treatment was administered to 20 patients including 10 undergoing fistulectomies and 10 undergoing fistulotomies. The average duration of non-thermal plasma application in the operating room was shorter for fistulotomies. The pain reported the day after surgery was similar in both groups. Improvements in the number of evacuations starting from the day after surgery, as well as the assessment of stool quality using the Bristol scale, indicated satisfactory intestinal recovery. Fistulotomy patients exhibited faster wound healing times. These findings underscore the efficacy of non-thermal plasma as a postoperative therapy for anal fistula, enhancing healing and recovery outcomes without increasing complication risks.