Ceramidase catalyzes the hydrolysis of the N-acyl linkage between the fatty acid and sphingosine base in ceramide to produce sphingosine and a free fatty acid. Three isoforms of ceramidases (acid, neutral, and alkaline) have been characterized based on differences in their catalytic pH optimum.[1][2] C12 NBD dihydro ceramide is a fluorescent ceramide analog that contains a saturated bond in the C-4/C-5 position of the sphingosine backbone. C-12 NBD ceramide, which contains the C-4/C-5 double bond, is a fluorescent ceramidase substrate that can be used for the measurement of alkaline and neutral ceramidase activity from a variety of sources.[3] C12 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 C-4/C-5 sphingosine bond in C-16-ceramide results in approximately a 10-fold reduction in efficiency as a substrate for rat brain ceramidase compared to the unsaturated substrate. [4] However, experimental characterization of C12 NBD dihydro ceramide as a ceramidase substrate needs to be performed.
Reference:
[1]. Tani, M., Okino, N., Mori, K., et al. Molecular cloning of the full-length cDNA encoding mouse neutral ceramidase. A novel but highly conserved gene family of neutral/alkaline ceramidases. The Journal of Biological Chemisty 275(15), 11229-11234 (2000).
[2]. Ferlinz, K., Kopal, G., Bernardo, K., et al. Human acid ceramidase. Processing, glycosylation, and lysosomal targeting. The Journal of Biological Chemisty 276(38), 35352-35360 (2001).
[3]. Tani, M., Okino, N., Mitsutake, S., et al. Specific and sensitive assay for alkaline and neutral ceramidases involving C12-NBD-ceramide. J. Biochem. 125(4), 746-749 (1999).
[4]. Bawab, S.E., Usta, J., Roddy, P., et al. Substrate specificty of rat brain ceramidase. Journal of Lipid Research 43, 141-148 (2002).