In Greece, biodiesel, in the form of Fatty Acid Methyl Esters (FAME), is produced since 2005 andcurrently mixed in the Diesel fuel at about 5.5% vol., this percentage being slowly but steadily increasing.The production capacity of existing biodiesel factories in Greece can supply the required quantities toincrease the biodiesel blending percentage up to 15%, provided that the necessary vegetable oil andrecycled oil quantities should become available and prices be favorable. According to Greek legislation, atleast 30% of the raw material must be domestically produced. This need is covered mainly by recycledcooking oils and only secondarily by energy crops in Northern Greece. The production of biodiesel fromlocal factories in Greece was regulated by means of distributing an annual quantity of biodiesel among thefactories, based on an algorithm that is modified every year. Automotive manufacturers allow running ofmodern diesel powered passenger cars on blends of biodiesel up to B20‐B30, provided that certainadditional maintenance measures are taken, including more frequent fuel and oil filter changes, along withinspection of engine oil level and the fuel lines and injection system components for possible leaks. In orderto ensure customer’s acceptance, standardization and quality assurance are key factors for the marketintroduction of biodiesel as transport fuel. This PhD thesis examines the effect of biodiesel fuel onperformance, efficiency, emissions and injection system durability of conventional and modern Dieselengines under steady state and transient operating conditions.These include a better understanding of the engine ECU operation with a high biodiesel (FAME) blend inmodern common‐rail injection Diesel engine, the study of the biodiesel combustion analyzing the capturedindicator graphs, the investigation of the filter regeneration behavior by means of the infraredthermography on a small filter module and in‐house computational tool and the study of biodieselatomization and injection characteristics by means of the Laser Doppler Velocimetry technique.In the first part of the Thesis, a better understanding of the engine ECU operation with a high biodiesel(FAME) blend in modern common‐rail injection Diesel engines is attempted. This understanding is appliedto the explanation of the observed effects on the engine performance and emissions characteristics withthis engine category. The results are compared to the relatively scarce works reported in the specializedliterature for the specific engine category and high blending ratios. Also, the effect of biodiesel blends inthe transient fuel system and engine operation is examined. Furthermore, the effect of prolonged engineoperation at a high biodiesel blend on the fuel system and the engine components is investigated.In the second part of the Thesis, the effect of low percentage biodiesel blends on the diesel filter loadingand regeneration behavior of a conventional, single cylinder Diesel engine with low pressure injectionsystem is presented. A fuel additive is employed in order to perform filter regeneration at relatively lowexhaust temperatures. The effect of the biodiesel on the filter regeneration is further investigated bymeans of infrared thermography on a small filter module and in‐house computational tool. In addition, theeffect of biodiesel on injection characteristics is investigated by means of Laser Doppler Velocimetrytechnique.Τhe results enhance our understanding of the engine ECU behavior with the high percentage biodieselblends. Certain modifications to engine ECU maps and control parameters are proposed and discussed,aimed at improvement of transient performance of modern engines run on high percentage biodieselblends. However, a high pressure pump failure that was observed after prolonged engine operation withthe B70 blend, hints to the use of more conservative biodiesel blending in fuel that should not exceed 30%for common rail engines.The observed benefits as regards the DPF extend also to the faster evolution of regeneration and thelower overall filter wall temperatures with favorable effects on the filter durability.