Methods for determining peroxidately active substances

Claim 5

The method of claim 2 wherein said peroxidatively active substance is selected from the group consisting of complexed transition metal ions, hemes, hemoproteins, and peroxidases.

Claim 6

The method of claim 2 wherein said F-L-Q is of the formula: ##STR36## 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 7

The method of claim 2 wherein said hydrogen donor is selected from the group consisting of benzidines, p-aminophenols, 4-aminoantipyrenes, and p-phenylenediamines.

Claim 8

The method of claim 2 wherein said hydrogen donor is of the formula: ##STR37## 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.

Claim 9

The method of claim 6 wherein said Q is of the formula: ##STR38## wherein: R 8 and R' 8 are 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, phosphorous, or halogen; or two R 8 s and/or two R' 8 s are taken together to form a ring or rings being substituted or unsubstituted, a single ring or part of a fused ring system; A is independently OH, N(R 9 ) 2 or SH (thio), wherein each R 9 is independently selected from 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, or phosphorous and is bonded to nitrogen at a saturated carbon atom; or A is taken together with R 8 or R' 8 and/or with another R 9 to form a ring or rings being substituted or unsubstituted, a single ring or part of a fused ring system; L is a linking group having atoms selected from the group consisting of carbon, nitrogen, oxygen, sulfur and phosphorus, wherein O, N, S (sulfur atom), and P when present are bound to at least a carbon atom and may be bound to one or more of each other and wherein the length of said linking group is sufficient to permit quenching of F by Q by energy transfer; and F is contained in a moiety selected from the group consisting of fluorescein, rhodamine, acridine, rosamine, naphthylamine, coumarin, benzoxazole, benzoxdiazole, pyrene, stilbene, phenanthridine, anthracene, merocyanine, and perylene moieties.

Claim 10

The method of claim 7 wherein said peroxidatively active catalyst is selected from the group consisting of complexed metal ions, hemes, hemoproteins, and peroxidases.

Claim 11

The method of claim 7 wherein R 8 and R' 8 are independently lower alkyl, N(lower alkyl) 2 , NH(lower alkyl), amino substituted lower alkyl, --COOR 13 , --CN, --Cl, --Br, --I, --NO 2 , --SO 2 R 13 , --PO 3 (R 13 ) 2 , or --C(O)N(R 13 ) 2 wherein R 13 is independently H, alkyl, carboxyalkyl, or amino substituted lower alkyl.

Claim 12

The method of claim 7 wherein said hydrogen donor is selected from the group consisting of benzidines, p-aminophenols, 4-aminoantipyrenes, and p-phenylenediamines.

Claim 13

A compound of the formula F-L-Q wherein F is a fluorescer capable of producing a fluorescent signal, and is contained in a moiety selected from the group consisting of fluorescein, rhodamine, acridine, rosamine, naphthylamine, coumarin, benzoxazole, benzoxdiazole, anthracene, merocyanine, and perylene moieties; Q is a quencher capable of quenching said signal when linked to F, and is contained in a moiety selected from the group consisting of aniline, phenol, and thiophenol moieties; and L is a bond or linking group containing a bond and is contained in a functional group selected from the group consisting of ethers, thioethers, disulfides, esters, amides, and carbonates; wherein said bond is capable of being cleaved by an oxidation-reduction reaction of a peroxidatively active substance (PAS) with a substrate of said PAS and a hydrogen donor wherein said cleavage of said bond substantially reduces said quenching.

Claim 14

A compound of the formula F-L-Q wherein F is a fluorescer capable of producing a fluorescent signal, and is contained in a moiety selected from the group consisting of fluorescein, rhodamine, acridine, rosamine, naphthylamine, coumarin, benzoxazole, benzoxdiazole, anthracene, merocyanine, and perylene moieties; Q is a quencher capable of quenching said signal when linked to F, and is contained in a moiety selected from the group consisting of aniline, phenol, and thiophenol moieties; and L is a bond or linking group containing a bond and is contained in a functional group selected from the group consisting of ethers, thioethers, disulfides, esters, amides, and carbonates; wherein said bond is capable of being cleaved by an oxidation-reduction reaction of a peroxidatively active substance (PAS) with a substrate of said PAS and a hydrogen donor wherein said cleavage of said bond substantially reduces said quenching.

Claim 15

The compound of claim 11 wherein said peroxidatively active substance is selected from the group consisting of complexed transition metal ions, hemes, hemoproteins, and peroxidases.

Claim 16

The compound of claim 11 wherein said F-L-Q is of the formula: ##STR39## 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 17

The compound of claim 11 wherein said hydrogen donor is selected from the group consisting of benzidines, p-aminophenols, 4-aminoantipyrenes, and p-phenylenediamines.

Claim 18

The compound of claim 11 wherein said hydrogen donor is of the formula: ##STR40## 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.