In the earlier investigations on the animal fat biodiesel, the authors have found that blend 20 was suitable to run the engine. In the CI engines, operating input parameters, FIPs and FITs, are the significant parameters to improve the engine performance and emission characteristics. Biodiesel was prepared by the transesterification process and blended with 20% diesel fuel to get AFB20 blend and to carried out the experiments by varying the FIPs by 180, 200, 220 and 240 bars and FITs by 19°, 21°, 23°, 25° and 27° bTDCs. The experiments were investigated in the single-cylinder 4S CI engine. In the evaluation, it was found that the FIP of 200 bar and 180 bar consumes 0.96% lower BSFC compared to 240 & 220 bars and exhibits a higher BTE of 2.6%. 1.62% lower EGT was found in 240 bar compared to 180 bar. 0.86% lower UBHC and 24.67% lower smoke emissions were observed in 200 and 240 bars. Lower BSFC was found in the 23° IT and higher BTE was found in 19° IT. Lower EGT and UBHC emissions were found in 19° IT. Lower EGT and CO emissions were found in 19° CA and higher CO emission was found in 23° IT at full load.