A brief overview of the toxic sphingomyelinase Ds of brown recluse spider venom and other organisms, and simple methods to detect production of its signature cyclic ceramide phosphate.

A special category of phospholipase Ds (PLD) in the venom of the brown recluse spider ( Loxosceles reclusa ) and several other Sicariid spiders accounts for the dermonecrosis and many of the other clinical symptoms of envenomation. Related proteins are produced by other organisms including fungi and bacteria. These PLDs are often referred to as sphingomyelinase Ds (SMase D) because they cleave sphingomyelin (SM) to choline and "ceramide phosphate." The lipid product has actually been found to be a novel sphingolipid: ceramide 1,3-cyclic phosphate (Cer1,3P). Since there are no effective treatments for the injury induced by the bites of these spiders, SMase D/PLDs are attractive targets for therapeutic intervention, and some of their features will be described in this minireview. In addition, two simple methods are described for detecting the characteristic SMase D activity using a fluorescent SM analog, (N-[12-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]dodecanoyl]-SM (C12-NBD-SM), that is cleaved to C12-NBD-Cer1,3P, which is easily separated from other potential metabolites by thin-layer chromatography and visualized under ultraviolet light. Besides confirming that C12-NBD-Cer1,3P is the only product detected upon incubation of C12-NBD-SM with brown recluse spider venom, the method was also able to detect for the first time very low levels of activity in venom from another spider, Kukulcania hibernalis The simplicity of the methods makes it relatively easy to determine this signature activity of SMase D/PLD. Significance Statement The sphingomyelinase D/PLDs that are present in the venom of the brown recluse spider and other sources cause considerable human injury, but detection of the novel sphingolipid product, ceramide 1,3-cyclic phosphate, is not easy by previously published methods. This minireview describes a simple method for detection of this activity that will be useful for studies of its occurrence in spider venoms and other biological samples, perhaps including lesions from suspected spider bites and infections.