A chemiluminescent substrate of a hydrolytic enzyme having the following
general Formula I is disclosed, as follows: Lumi-M-P Formula I where
"Lumi" is a chemiluminescent moiety capable of producing light (a) by
itself, (b) with MP attached and (c) with M attached. Examples of Lumi
include chemiluminescent acridinium compounds, benzacridinium compounds,
quinolinium compounds, isoquinolinium compounds, phenanthridinium
compounds, and lucigenin compounds, spiroacridan compounds, luminol
compounds and isoluminol compounds. M is a multivalent heteroatom having
at least one lone pair of electrons selected from oxygen, nitrogen and
sulfur, directly attached to the light emitting moiety of Lumi at one end
and to P at the other end. P is a group that can be readily removed by
hydrolytic enzymes. An enzymatic reaction utilizing the above compound is
the following: ##STR00001## where HE is a hydrolytic enzyme. Lumi-M is a
chemiluminescent product having physical and/or chemical properties
different from those of Lumi-M-P.