The use of biofuels for transportation has grown substantially in the past decade in response to federal mandates and increased concern about the use of petroleum fuels. As biofuels become more common, it is imperative to assess their influence on mobile source emissions of regulated and hazardous pollutants. This assessment cannot be done without first obtaining a basic understanding of how biofuels affect the relationship between fuel properties, engine design, and combustion conditions. Combustion studies were conducted on biodiesel fuels from four feedstocks (palm oil, soybean oil, canola oil, and coconut oil) with two injection systems, mechanical and electronic. For the electronic system, fuel injection timing was adjusted to compensate for physical changes caused by different fuels. The emissions of nitrogen oxides (NO x ) and partial combustion products were compared across both engine injection systems. The analysis showed differences in NO x emissions based on hydrocarbon chain length and degree of fuel unsaturation, with little to no NO x increase compared with ultra-low sulfur diesel fuel for most conditions. Adjusting the fuel injection timing provided some improvement in biodiesel emissions for NO x and particulate matter, particularly at lower engine loads. The results indicated that the introduction of biodiesel and biodiesel blends could have widely dissimilar effects in different types of vehicle fleets, depending on typical engine design, age, and the feedstock used for biofuel production.