Bioethanol and plant oil-derived biodiesel are generally considered first generation biofuels. More sustainable and cost effective new biofuels are being designed and produced using modern tools of metabolic engineering and synthetic biology. These new microbial fuels have great potential to become viable alternatives and supplements for petroleum-derived liquid transportation fuels. MAN Latin America has worked in cooperation with REG Life Sciences, a North American industrial biotechnological company, to help in the development of high quality fuels for automotive purposes.The aim of this paper is to present the test engine results of a novel microbially produced fatty acid methyl ester (FAME), under the banner of UltraClean™ Diesel, in a Proconve P7 (Euro V) MAN D0834, diesel engine. Described are a comprehensive performance and emissions evaluation as well as an interpretation of the primary fuel properties. The test engine was operated with 100% standard Brazilian diesel (S50), blend of 20% of UltraClean™ Diesel with 80% of S50 and 100% UltraClean™ Diesel. Tests were conducted in Brazil using an original 4.58 liter, 166 kW engine, equipped with common rail fuel injection system, external cooled EGR, and an oxidation catalytic converter.