Methods for determining peroxidatively active substances

Claim 2

A kit for detecting a peroxidatively active substance (PAS) in a sample or medium suspected of containing said PAS, said kit comprising in packaged combination: a compound of the formula F-L-Q wherein F is a fluorescer capable of producing a fluorescent signal, Q is a quencher capable of quenching by energy transfer said signal when linked to F, and L is a bond or linking group containing a bond wherein said bond is capable of being cleaved by an oxidation-reduction reaction of said PAS with a substrate of said PAS and a hydrogen donor wherein said cleavage substantially reduces said quenching; and a hydrogen donor.

Claim 3

The kit of claim 1 wherein said bond is contained in a functional group selected from the group consisting of ethers, thioethers, disulfides esters, amides, and carbonates.

Claim 4

The kit of claim 1 wherein said fluorescer is contained in a moiety selected from the group consisting of fluorescein, rhodamine, acridine, rosamine, naphthylamine, coumarin, benzoxazole, benzoxdiazole, anthracene, merocyanine, and perylene moieties.

Claim 5

The kit of claim 1 wherein said peroxidatively active substance is selected from the group consisting of complexed transition metal ions, hemes, hemoproteins, and peroxidases.

Claim 6

The kit of claim 1 wherein said quencher is contained in a moiety selected from the group consisting of aniline, phenol, and thiophenol moieties.

Claim 7

The kit of claim 1 wherein said F-L-Q is of the formula: ##STR35## wherein: X is O, S (sulfur atom), or NR 1 wherein R 1 is H, alkyl, or aryl, with the proviso that O or S (sulfur atom) be bound to carbon atoms; Z is C or N; b is 0 or 1; a and c are independently 0, 1 or 2 with the proviso that the ring contain 5 to 7 atoms; R is independently H or a substituent having from 1 to 50 atoms other than hydrogen, which atoms are independently selected from the group consisting of carbon, oxygen, nitrogen, sulfur, phosphorus, and halogen, or two R's can be taken together to form a ring, being substituted or unsubstituted, a single ring or part of a fused ring system, wherein one R is OH, SH (thio), or amino nitrogen bound to a site on the ring, wherein said site is separated from D by an even number of ring atoms excluding X; and D is O or S (sulfur atom) wherein said O or S (sulfur atom) is bonded to F or Q with the proviso that the two electron oxidation potential of said compound of the formula F-L-Q is greater than that of said hydrogen donor.

Claim 8

The kit of claim 1 wherein said hydrogen donor is selected from the group consisting of benzidines, p-aminophenols, 4-aminoantipyrenes, and p-phenylenediamines.

Claim 9

The kit of claim 1 wherein said hydrogen donor is of the formula: ##STR36## wherein: A is a hydroxyl group, an amino group, --NHR 3 , or --NR 3 R 4 , wherein R 3 and R 4 are independently an alkyl group having 1 to 5 carbon atoms, a cycloalkyl group having 3 to 6 carbon atoms, a hydroxyalkyl group having 1 to 5 carbon atoms, or an acylaminoalkyl group having 1 to 5 carbon atoms; or R 3 and R 4 may combine to form a ring which is unfused or fused, substituted or unsubstituted, a single ring, or part of a multi-ring system; R 1 and R 2 are independently a hydrogen atom, an alkyl group having 1 to 5 carbon atoms, an alkoxy group having 1 to 5 carbon atoms, a halogen atom, a carboxyl group, a methanesulfonyl group or a phenyl group; or R 1 and R 2 may combine together to form a cycloalkene, an aromatic ring or a heterocyclic ring; and n is 1 to 5.