There has been an extensive worldwide search for alternate fuels that fit with the existing infrastructure and would thus displace fossil-based resources. In metabolic engineering work at Argonne National Laboratory, strains of fuel have been designed that can be produced in large quantities by photosynthetic bacteria, eventually producing a heavy alcohol called phytol (C20H40O). Phytol’s physical and chemical properties (cetane number, heat of combustion, heat of vaporization, density, surface tension, vapor pressure, etc.) correspond in magnitude to those of diesel fuel, suggesting that phytol might be a good blending agent in compression ignition (CI) engine applications. The main reason for this study was to investigate the feasibility of using phytol as a blending agent with diesel; this was done by comparing the performance and emission characteristics of different blends of phytol (5%, 10%, 20% by volume) with diesel. The experimental research was performed on a single-cylinder engine under conventional operating conditions. Since phytol’s viscosity is much higher than that of diesel, higher-injection-pressure cases were investigated to ensure the delivery of fuel into the combustion chamber was sufficient. The influence of the fuel’s chemical composition on performance and emission characteristics was captured by doing an injection timing sweep. Combustion characteristics as shown in the cylinder pressure trace were comparable for the diesel and all the blends of phytol at each of the injection timings. The 5% and 10% blends show lower CO and similar NOx values. However, the 20% blend shows higher NOx and CO emissions, indicating that the chemical and physical properties have been altered substantially at this higher percentage. The combustion event was depicted by performing high-speed natural luminosity imaging using endoscopy. This revealed that the higher in-cylinder temperatures for the 20% blend are the cause for its higher NOx emissions. In addition, three-dimensional simulations of transient, turbulent nozzle flow were performed to compare the injection and cavitation characteristics of phytol and its blends. Specifically, area and discharge coefficients and mass flow rates of diesel and phytol blends were compared under corresponding engine operating conditions. The conclusion is that phytol may be a suitable blending agent with diesel fuel for CI applications.