Pepsin digest residue method

Recently, Noort et al developed a procedure that is based on straightforward isolation of adducted BuChE from plasma by means of affinity chromatography with a procainamide column, followed by pepsin digestion and LC/electrospray tandem MS analysis of a specific nonapeptide containing the phosphonylated active site serine-198 residue (5). This method surpasses the limitations of the fluoride-reactivation method, since it can also deal with dealkylated ( aged ) phosphonylated BuChE. The method allowed the positive analysis of several serum samples of Japanese victims of the terrorist attack in the Tokyo subway in 1995. Furthermore, the method could be applied for detection of ChE modifications induced by, e.g., diethyl paraoxon and pyridostigmine bromide, illustrating the broad scope of this approach. This new approach... 

Fidder and coworkers (50) developed a versatile procedure that identifies phosphylated butyrylcholi-nesterase. Adducted butyrylcholinesterase is isolated from plasma by affinity chromatography (procainamide column), digested with pepsin, and a nonapep-tide containing the phosphylated active-site serine residue detected using LC/ESI/MS/MS (quadrupole-TOF hybrid instrument). A C18 150 x 0.3-mm LC column was used, eluted with a gradient of water-acetonitrile-0.2 % formic acid. The method was applied successfully to casualties of sarin poisoning from the Tokyo subway attack (see Chapter 17). 

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