The literature shows that information about the physical, chemical, and cell toxicity properties of particulate matter (PM) from diesel vehicles is not rich as the existence of a remarkable number of studies about the combustion, performance, and emissions of diesel vehicles using renewable liquid fuels, particularly biodiesels and alcohols. Also, the PM analyses from combustion of spent coffee ground biodiesel have not been comprehensively explored. Therefore, this research is presented. Pure diesel, 90% diesel + 10% biodiesel, and 90% diesel + 9% ethanol + 1% biodiesel, volume bases, were tested under a fast idle condition. STEM, SEM, EDS, Organic Carbon Analyzer, TGA/DSC, and Raman Spectrometer were employed for investigating the PM physical and chemical properties, and assays of cell viability, cellular reactive oxygen species, interleukin-6, and tumor necrosis factor-alpha were examined for investigating the PM cell toxicity properties. It is found that the application of both biodiesel and ethanol has the potential to change the PM properties, while the impact of ethanol is more than biodiesel on the changes. Regarding the important aspects, biodiesel can be effective for better human health (due to a decrease in cell death (−60.8%)) as well as good diesel particulate filter efficiency (due to lower activation energy (−7.6%) and frequency factor (−83.2%)). However, despite a higher impact of ethanol on the reductions in activation energy (−24.8%) and frequency factor (−99.0%), this fuel causes an increase in cell death (84.1%). Therefore, biodiesel can be an appropriate fuel to have a positive impact on human health, the environment, and emissions catalysts performance, simultaneously.