Diagnostic test for Niemann-Pick disease

Claim 1

A chromogenic method for determining the sphingomyelinase activity in an extract of human cells or tissues for use in the diagnostic testing for Niemann-Pick disease, comprising the steps of: a. forming an aqueous incubation mixture of said extract and a sphingomyelinase-specific chromogenic artificial substrate consisting of a 2-alkanoylamino-4-nitrophenyl phosphorylcholine-hydroxide having the formula ##STR5## wherein n is an integer from 10 to 18, inclusive; b. incubating said mixture for a determinate time period, whereby sphingomyelinase-catalyzed hydrolysis of said substrate into the corresponding 2-alkanoylamino-4-nitrophenol and phosphoryl-choline takes place in an amount proportional to the sphingomelinase activity in said extract; c. alkalinizing said mixture so as to convert all of said 2-alkanoylamino-4-nitrophenol formed in step (b) into the alkali salt thereof, said alkali salt imparting to said mixture a bright yellow color whose intensity is proportional to the amount of said substrate that has been hydrolyzed; d. precipitating and removing from said mixture the entire protein content in said mixture, thereby leaving a clear supernatant solution having the color developed in step (c); e. measuring said protein content which was removed from said mixture in step (d); and f. measuring the optical density of said clear supernatant solution to thereby quantitatively determine the amount of said substrate that has been hydrolyzed; whereby the sphingomyelinase activity in said extract may be determined as the number of nanomoles of said substrate that has been hydrolyzed per hour of said incubation time period per milligram of protein as measured in step (e).