This study focuses on an analysis of the spray behavior, combustion, and emission characteristics of jet propellant-5 (JP-5) and biodiesel fuels with single-injection timing and multiple split injection strategies in a common rail direct injection (CRDI) single-cylinder diesel engine system. The analysis includes visualization of the spray and combustion. Multiple split injection strategies (e.g., double, triple, quadruple, and quintuple) were considered by equally distributing the fuel injection amount within the single-injection. Injection of biodiesel has a delayed start (0.2 ms) as well as shorter spray tip penetration compared with JP-5. As the fuel injection timing was approached to the top dead center (TDC), the engine performance and combustion efficiency improved. Retarding the injection timing contributed to an increase in carbon dioxide (CO2) (JP-5: max. 2.6% up, BD100: max. 1.5% up) and a decrease in carbon monoxide (CO) (JP-5: max. 93% down, BD100: max. 91% down) and nitrogen oxides (NOx) (JP-5: max. 83% down, BD100: max. 82% down). In comparison with JP-5, biodiesel showed disadvantages from the point of its combustion and emission characteristics for all injection timings. The quadruple-injection strategy, in which fuel injection was performed four times, showed excellent combustion, engine performance, and combustion efficiency. The CO2 emissions were highest with the quadruple-injection strategy (JP-5: 6.6%, BD100: 5.8%). The CO emissions of biodiesel decreased as the pulses of split injection extended, and a significant reduction of 83.8% was observed. NOx increased as the number of split injections increased (JP-5: max. 37% up, BD100: max. 52% up). JP-5 was a longer ignition delay than that of biodiesel from combustion flame visualization results. The final combustion in the multiple-injection strategy showed a typical diffusion combustion pattern.