Fluorescence detection of cysteine and homocysteine

Patent metadata

Field	Value
Publication number	US-9709572-B2
Application number	US-201514950961-A
Country	US
Kind code	B2
Filing date	Nov 24, 2015
Priority date	Aug 3, 2011
Publication date	Jul 18, 2017
Grant date	Jul 18, 2017

Abstract

Embodiments of probes for selectively detecting compounds having a thiol group and an amino group, e.g., cysteine and/or homocysteine, are disclosed, along with methods and kits for detecting the compounds in neutral media with the probes. The probes have a structure according to the general formula

where R 1 -R 4 independently are hydrogen hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy, x is an integer from 0 to 4, and each R 5 independently is halogen, hydroxyl, thiol, thioether, lower aliphatic, or lower alkoxy. Embodiments of the disclosed probes are capable of undergoing condensation/cyclization reactions with cysteine and/or homocysteine. Cysteine and/or homocysteine can be selectively detected and identified by determining fluorescence emission of the probes at characteristic wavelengths.

First claim

We claim:

1. A probe having a chemical structure according to formula II

where R 1 is hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy;
R 2 -R 4 independently are hydrogen, hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy;
x is an integer from 0 to 4; and
each R 5 independently is halogen, hydroxyl, thiol, thioether, lower aliphatic, or lower alkoxy.

2. The probe of claim 1 where R 1 , R 3 , or R 4 is hydroxyl, thiol, thioether, or lower alkoxy.

3. The probe of claim 1 where R 1 is methoxy.

4. The probe of claim 1 where R 1 is methoxy, R 2 -R 4 are hydrogen, and x is 0.

5. The probe of claim 1 where the probe is capable of undergoing a condensation/cyclization reaction with a compound having a thiol group and an amino group.

6. The probe of claim 5 where the probe has a first fluorescence spectrum having an emission spectrum maximum at a first wavelength after condensation with the compound, and the probe has a subsequent fluorescence spectrum having an emission spectrum maximum at a second wavelength after cyclization, wherein the first and second wavelengths are different from one another.

7. The probe of claim 5 where the compound is cysteine, homocysteine, or a combination thereof.

8. A kit for detecting at least one compound having a thiol group and an amino group, comprising at least one probe according to general formula II

where R 1 is hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy, R 2 -R 4 independently are hydrogen, hydroxyl, halogen, thiol, thioether, lower aliphatic, or lower alkoxy, x is an integer from 0 to 4, and each R 5 independently is halogen, hydroxyl, thiol, thioether, lower aliphatic, or lower alkoxy.

9. The kit of claim 8 where R 1 is methoxy.

10. The kit of claim 8 where the probe is

11. The kit of claim 8 , further comprising a buffer solution at physiologic pH.

12. The kit of claim 11 , where the buffer solution is a phosphate solution at pH 7-8.

13. The kit of claim 11 , where the buffer solution further comprises a surfactant.

14. The kit of claim 13 , where the surfactant is a quaternary ammonium surfactant.

15. The kit of claim 13 , wherein the surfactant is cetyltrimethylammonium bromide.

16. The kit of claim 8 , further comprising a plurality of disposable containers in which a reaction between the probe and the at least one compound can be performed.

17. The kit of claim 16 , wherein an amount of the probe effective to undergo a detectable change in the probe's fluorescence emission spectrum when reacted with the at least one compound is premeasured into the plurality of disposable containers.

Assignees

• Univ Portland State

Inventors

• Strongin Robert
• Yang Xiaofeng

Classifications

• G01N21/6428

  Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" (in vivo A61B5/00; immunoassay G01N33/53) · CPC title

• Y10T436/173845

  Amine and quaternary ammonium · CPC title

• G01N33/6815 Primary

  containing sulfur, e.g. cysteine, cystine, methionine, homocysteine · CPC title

• C07D277/66

  with aromatic rings or ring systems directly attached in position 2 · CPC title

• G01N2021/6439

  with indicators, stains, dyes, tags, labels, marks · CPC title