Fluorescein-based metal sensors, and methods of making and using the same

• US 20050112769A1
• Filed: 08/27/2004
• Published: 05/26/2005
• Est. Priority Date: 09/05/2003
• Status: Active Grant

• Current Assignee: Massachusetts Institute of Technology
• Original Assignee: Massachusetts Institute of Technology
• Inventors: Lippard, Stephen J., Nolan, Elizabeth Marie
• Granted Patent: US 7,615,377 B2
• Time in Patent Office: Days
• US Class Current: 436/81
• CPC Class Codes: G01N 31/22 using chemical indicators G01N31/02 takes precedence
• CPC Class Codes: G01N 33/5308 for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
• CPC Class Codes: Y10T 436/24 Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry

Abstract

The present invention is directed, in part, to fluorescein-based ligands for detection of metal ions, and methods of making and using the same.

Claims

1. A fluorescein-based ligand, comprising a ligand having the following structure:

2. The fluorescein-based ligand of claim 1, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

3. The fluorescein-based ligand of claim 2, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 10%.

4. The fluorescein-based ligand of claim 3, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 50%.

5. The fluorescein-based ligand of claim 4, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 100%.

6. The fluorescein-based ligand of claim 1, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 1 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

7. The fluorescein-based ligand of claim 1, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 10 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

8. The fluorescein-based ligand of claim 1, wherein said ligand has one or more K substituents, with each K independently being one of the following:

linear or branched alkyl, alkenyl, linear or branched aminoalkyl, linear or branched acylamino, linear or branched acyloxy, linear or branched alkoxycarbonyl, linear or branched alkoxy, linear or branched alkylaryl, linear or branched hyrdoxyalkyl, linear or branched thioalkyl, acyl, amino, hydroxy, thio, aryloxy, arylalkoxy, hydrogen, alkynyl, halogen, cyano, sulfhydryl, carbamoyl, nitro, trifluoromethyl, amino, thio, lower alkoxy, lower alkylthio, lower alkylamino, nitro, phenoxy, benzyloxy, hydrogen, amine, hydroxyl, alkoxyl, carbonyl, acyl, formyl or sulfonyl.

9. The fluorescein-based ligand of claim 1, wherein Q is —

OZ and Z2 forms a different tautomer with the fluorescein core.

10. The fluorescein based-ligand of claim 1, wherein said ligand has the following structure:

11. The fluorescein-based ligand of claim 10, wherein said ligand has one or more K substituents, with each K independently being either —

Cl or —

F.

12. The fluorescein-based ligand of claim 10, wherein said ligand has one of the following structures:

13. The fluorescein-based ligand of claim 10, wherein upon addition of said ligand to an aqueous solution containing Hg²⁺ at about pH 7.0, the fluorescence of the fluorescein-based ligand decreases.

14. The fluorescein-based ligand of claim 13, wherein said ligand has the following structure:

15. The fluorescein-based ligand of claim 1, wherein said ligand has the following structure:

16. The fluorescein-based ligand of claim 10, wherein the fluorescein structure of said ligand is in a different isomeric form.

17. The fluorescein-based ligand of claim 1, wherein said ligand forms a chelating agent upon complexation of said ligand with a metal ion.

18. The fluorescein-based ligand of claim 1, wherein said ligand forms a bidentate or tridentate chelating agent upon said complexation.

19. A fluorescein-based ligand, comprising a ligand having the following structure:

20. The fluorescein-based ligand of claim 19, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

21. The fluorescein-based ligand of claim 20, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 10%.

22. The fluorescein-based ligand of claim 21, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 50%.

23. The fluorescein-based ligand of claim 22, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 100%.

24. The fluorescein-based ligand of claim 19, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 10 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

25. The fluorescein-based ligand of claim 19, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 1 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

26. The fluorescein-based ligand of claim 19, wherein said ligand has one of the following structures:

27. The fluorescein-based ligand of claim 26, wherein upon addition of said ligand to an aqueous solution containing Hg²⁺ at about pH 7.0, the fluorescence of the fluorescein-based ligand decreases.

28. A coordination complex, comprising a metal ion coordinated to one of the fluorescein-based ligands claimed above.

29. The coordination complex of claim 28, wherein said metal ion is a heavy metal ion.

30. The coordination complex of claim 28, comprising Hg²⁺ coordinated to one of the fluorescein-based ligands claimed above.

31. A method of detecting, and optionally quantifying the concentration of, a metal ion in a sample, comprising:

a. adding to a sample a fluorescein-based ligand comprising one of the following structures;

wherein;

Z is hydrogen or any hydroxyl-protecting group;

Q is O, S or Se;

K is optionally one or more substituents of the indicated aromatic ring that do not preclude detecting a metal ion by fluorescence;

Y is O, S, Se, NR, or C(CH3)3, wherein R is an alkyl and R and the methyl groups of C(CH3)2 are optionally substituted; and

Z2 is N, HOOCCH₂CH₂C—

, HOOC—

CH═

CH—

C—

, (2-carboxyphenyl)-C—

, (2-sulfophenyl)-C—

, (2-carboxy-3,4,5,6-tetrachlorophenyl)-C—

, (2-carboxy-4-nitrophenyl)-C—

, (2-carboxy-5-nitrophenyl)-C—

, (2-carboxy-4-aminophenyl)-C—

, (2-carboxy-5-aminophenyl)-C—

, (2,4-dicarboxyphenyl)-C—

, (2,5-dicarboxylphenyl)-C—

, (2,4,5-tricarboxyphenyl)-C—

, or other substituted (2-carboxyphenyl)-C-derivative; and

V is a Lewis base; and

R′

is H, alkyl, aryl or V, wherein at least one V comprises at least one sulfur-containing moiety;

b. measuring the fluorescence of said ligand in said sample; and

c. determining whether a metal ion is present in said sample, and optionally the concentration of said metal ion in said sample.

32. The method of claim 31, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

33. The method of claim 32, wherein the fluorescence of the fluorescein-based ligand increases by at least about 10%.

34. The method of claim 33, wherein the fluorescence of the fluorescein-based ligand increases by at least about 50%.

35. The method of claim 34, wherein the fluorescence of the fluorescein-based ligand increases by at least about 100%.

36. The method of claim 31, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 10 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

37. The method of claim 31, wherein upon addition of about 1 μ

M of said ligand to an aqueous solution containing Hg²⁺ at about 1 ppm and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.

38. The method of claim 31, wherein said sample is soil.

39. The method of claim 31, wherein said sample is water.

40. The method of claim 31, wherein said sample is in vitro.

41. The method of claim 31, further comprising measuring the fluorescence of said ligand in said sample at a different concentration of said ligand.

42. The method of claim 31, wherein one of said metal ions is a heavy metal ion.

43. The method of claim 31, wherein one of said metal ions is Hg²⁺.

44. A diagnostic kit for a metal ion, comprising:

a. a fluorescein-based ligand comprising one of the following structures;

wherein;

Z is hydrogen or any hydroxyl-protecting group;

Q is O, S or Se;

K is optionally one or more substituents of the indicated aromatic ring that do not preclude detecting a metal ion by fluorescence;

Y is O, S, Se, NR, or C(CH3)3, wherein R is an alkyl and R and the methyl groups of C(CH3)2 are optionally substituted; and

Z2 is N, HOOCCH₂CH₂C—

, HOOC—

CH═

CH—

C—

, (2-carboxyphenyl)-C—

, (2-sulfophenyl)-C—

, (2-carboxy-3,4,5,6-tetrachlorophenyl)-C—

, (2-carboxy-4-nitrophenyl)-C—

, (2-carboxy-5-nitrophenyl)-C—

, (2-carboxy-4-aminophenyl)-C—

, (2-carboxy-5-aminophenyl)-C—

, (2,4-dicarboxyphenyl)-C—

, (2,5-dicarboxylphenyl)-C—

, (2,4,5-tricarboxyphenyl)-C—

, or other substituted (2-carboxyphenyl)-C-derivative;

V is a Lewis base;

R′

is H, alkyl, aryl or V, wherein at least one V comprises at least one sulfur-containing moiety; and

b. instructions for using said ligand to detect a metal ion in a sample.

45. The kit of claim 44, wherein upon addition of about 500 nM of said ligand to an aqueous solution containing Hg²⁺ at about 2 ppb and at about pH 7.0, the fluorescence of the fluorescein-based ligand increases by at least about 5%.