C-12 NBD-dihydro-Ceramide compound is a fluorescent ceramide analog which contains a saturated bond in the C4-C5 position of the sphingosine backbone. C-12 NBD ceramide, is a fluorescent ceramidase substrate which can be used for the measurement of alkaline and neutral ceramidase activity from a variety of sources. C-12 NBD-dihydro-ceramide may exhibit reduced activity in ceramidase assays compared to C-12 NBD ceramide. This is based on the observation that saturation of the C4-C5 sphingosine bond in C16-ceramide results in approximately a 10-fold reduction in efficiency as a substrate for rat brain ceramidase compared to the unsaturated substrate. Yet, experimental characterization of C-12 NBD-dihydro-ceramide as a ceramidase substrate needs to be performed.