Abstract: The use of Highly Active Antiretroviral Therapy (HAART) has improved the prognosis of HIV-infected individuals. However, the beneficial effect of reduced risk of early death from opportunistic infections and other consequences of HIV infection is reduced because Antiretroviral Therapy (ART) also carries negative side effects, including hepatotoxicity. The study examines hepatotoxic effect of HAART and hepatitis co-infection in HIV patients. Alkaline phosphatase, alanine transaminase, aspartate transaminase, gamma glutamyl transferase were determined from the serum of 100 HAART-experienced and 50 HAART-naïve HIV infected subjects enzymatically using Junior Flexor autoanalyser. Serum proteins were assayed by the biuret reaction and bilirubin determination was based on the diazo coupling reaction of Ehrlich method. HBV surface antigen (HBsAg) was detected by enzyme-linked immunosorbent assay. Globulins, total proteins, alkaline phosphatase, alanine transaminase, aspartate transaminase and gamma glutamyl transferase were significantly (p<0.001) elevated in the HIV HAART-experienced patients than the controls. Forty six percent of the HAART-experienced developed hepatotoxicity. Based on bilirubin levels, 11% of the HAART-experienced had hepatotoxicity. Nine percent of the HAART-experienced were co-infected with hepatitis B and 77% of them developed hepatotoxicity whilst 22% of the HAART-naïves were co-infected with hepatitis B and 73% of these developed hepatotoxicity. Hepatotoxicity was negatively correlated with alkaline phosphatase, alanine transaminase, aspartate transaminase, gamma glutamyl transferase and total plasma proteins. HAART induced hepatotoxicity in HIV patients more than HAART-naïve. There is an increased risk of hepatotoxicity in HIV patients’ co-infection with hepatitis B virus and on antiretroviral therapy.