Molecular analytical release tags and their use in chemical analysis

• US 5,360,819 A
• Filed: 03/11/1985
• Issued: 11/01/1994
• Est. Priority Date: 02/01/1982
• Status: Expired due to Term

• Current Assignee: Northeastern University
• Original Assignee: Northeastern University
• Inventors: Giese, Roger W.
• Primary Examiner(s): Killos, Paul J.
• Application Number:
  US06/710,318
• Time in Patent Office: 3,522 Days
• Field of Search: 514/538, 514/423, 546/226, 562/465, 562/840, 560/60, 568/355, 568/647
• US Class Current: 514/538
• CPC Class Codes: C07D 207/46 with hetero atoms directly attached to the ring nitrogen atom
• CPC Class Codes: G01N 33/532 Production of labelled immunochemicals
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: G01N 33/78 Thyroid gland hormones , e.g. T3, T4, TBH, TBG or their receptors

Abstract

A release tag reagent suitable for use in the chemical analysis of a substance to be detected, which substance contains reactive groups, such as for, but not limited to gas phase detection groups, which reagent comprises three covalently bonded groups: a signal group which on release provides a ketone signal compound to be detected, a release group which may be cleaved to release the ketone signal group, which release group contains, for example, a vic glycol or an olefin group and a reactivity group which is reactive with a reactive group of the substance to be detected.

Claims

1. A release tag compound for use in chemical analysis, said compound having the generalized formula:

space="preserve" listing-type="equation">S--R.sub.e --R.sub.x

wherein;

S is a signal group comprising a moiety selected from the group consisting of alkyl radicals of 1-12 carbon atoms, alicyclic radicals of 3-7 carbon atoms, and phenyl radicals, said moiety containing at least one halogen, said signal group having the property that upon cleavage of said release tag compound at the R_e group, signal group S is released in the form of a ketone which is sufficiently volatile for determination in the gas phase;

R_e is a release group capable of undergoing cleavage to release signal group S, and comprises a linkage selected from the group consisting of ##STR90## R_x is a reactivity group comprising a reactive functional group capable of bonding said release tag compound to a labelable substance;

and further,

in the release tag compound having the release group of formula i, the R_x group is bonded to the oxygen atom of the formula and at least one signal group S is bonded to the carbon atom of the formula;

in the release tag compound having the release group of formula ii, the R_x and S groups are both bonded to the carbon atom of the formula, and R_e further comprises a keto, olefinic, alkynyl, or ester functionality located in a beta position relative to the OH group of the formula; and

in the release tag compound having the release group of formula iii, the carbonyl group of the formula bears a moiety comprising the R_x reactivity group, at least one signal group S is bonded to the non-carbonyl carbon atom of the formula, and an alkyl or aryl group is bonded to the non-carbonyl oxygen atom of the formula.

2. A release tag compound of claim 1 wherein release group R_e has formula iii and reactivity group R_x contains nitrogen, oxygen, bromine, or iodine.

3. A release tag compound of claim 1 wherein release group R_e has formula iii, and the carbonyl group of the formula bears a moiety having the formula W(CH₂)_n R_x wherein W is an oxygen atom or a CH₂ group, n is 1-3, and R_x comprises a reactivity qroup.

4. A release tag compound of claim 3 wherein R_x is --(CH₂)₂ CO₂ H.

5. The compound ##STR91## wherein R₁ is H or CH₃,

R₂ is a reactivity group, and

R₃ is F or C₁ -C₆ alkoxy.

6. The compound ##STR92## wherein R₁ is H or CH₃,

R₂ is a reactivity group, and

R₃ is F or C₁ -C₆ alkoxy.

7. The compound ##STR93##

8. The compound ##STR94## wherein R₁, R₂, and R₃ are C₁ -C₆ alkyl.

9. The compound ##STR95##

10. The compound ##STR96##

11. The compound ##STR97## wherein R₁ is C₁ -C₆ alkyl or phenyl,

R₂ is H or C₁ -C₃ alkyl,

R₃ is H or C₁ -C₃ alkyl, and

R₄ is F or C₁ -C₆ alkoxy.

12. The compound ##STR98##

13. The compound ##STR99## wherein R is propyl or isopropyl.

14. An assay comprising the steps ofcontacting an analyte-containing sample with:

a) a release tag compound comprising;

a signal group, S, a release group Re, and a reactivity group Rx, group S being covalently linked to group Re, and group Re being further covalently linked to group Rx to define structure S-Re-Rx for said compound;

Rx further comprising a functional group capable of forming a covalent bond with another molecule;

Re further comprising a functionality selected from the group consisting of β

-hydroxy ketones, β

-hydroxy olefins, β

-hydroxy alkynes, β

-hydroxyesters, hemiketal esters, hemiketal carbonates, epoxides, alkylidenimes, β

-aminoalcohols, β

-(alkylamino)-alcohols, aikylidene-protected diols, α

-hydroxy-ketos, and α

-amino-ketos;

S further comprising a halogenated electron-absorbing organic group;

groups S and Re being mutually selected such that upon cleavage of said release tag at release group Re, signal group S is released in a volatile form suitable for electron capture determination in the gas phase;

or with

b) a molecular conjugate comprising a covalently linked combination of said release tag compound and a labelable substance;

2) incubating said contacted analyte-containing sample; and

3) determining the amount of released said signal group.

15. A release tag compound for use in chemical analysis, said compound having the generalized formula ##STR100## wherein:

S is a signal group comprising a moiety selected from the group consisting of alkyl radicals of 1-12 carbon atoms, alicyclic radicals of 3-7 carbon atoms, and phenyl radicals;

said moiety containing fluorine;

said signal group having the property that upon cleavage of said release tag compound at the R_e group, signal group S is released in the form of a ketone which is sufficiently volatile for determination in the gas phase;

R_e is a release group capable of undergoing cleavage to release signal group S, and comprises a linkage having the generalized formula ##STR101## wherein X is selected from the group consisting of;

a) two OH radicals, to form a vicinal diol;

b) a covalent bond, to form an olefin;

c) an oxygen atom, to form an epoxide;

d) an NH group, to form an alkylidenimine;

e) one OH and one N(R)₂ radical wherein each R is H or an alkyl group, forming an α

-hydroxy amine;

f) one OH and one ketone carbonyl, forming an α

-hydroxy ketone;

g) one ketone carbonyl and one N(R)₂ radical wherein each R is H or an alkyl group, forming an α

-keto amine; and

h) an alkylidene-protected diol; and

R_x is a reactivity group comprising a reactive functional group capable of bonding said release tag compound to a labelable substance.

16. A release tag compound for use in chemical analysis, said compound having the formula ##STR102## wherein R₁ is H, or an alkyl, aromatic, or alkyl-aromatic group;

R₃ is a signal group selected from the group consisting of ##STR103## wherein m is 3-5, G is H, alkyl, haloalkyl, halogen, or alkoxy group, n is 1-2, p is 0-1, and R₁₆ and R₁₇ are each H or an alkyl group;

R₄ is an alkyl group;

the ##STR104## portion of said release tag compound is a release group wherein X is selected from the group consisting of;

a) two OH groups, to form a vicinal diol;

b) a chemical bond, to form an olefin;

c) an oxygen atom, to form an epoxide;

d) an NH group, to form an alkylidenimine group;

e) one OH and one N(R)₂ radical wherein each R is H or an alkyl group, forming an a-hydroxy amine;

f) one OH and one ketone carbonyl, forming an α

-hydroxy ketone;

g) one ketone carbonyl and one N(R)₂ radical wherein R is H or an alkyl group, forming an α

-keto amine; and

h) an alkylidene-protected diol;

said release group being capable of undergoing cleavage to release the signal group as part of a ketone having the formula R₃ COR_4;

Y is a linking moiety selected from the group consisting of;

a chemical bond, a carbonyl group, an alkyl group, and an aromatic group; and

Z is a reactivity group comprising a reactive functional group capable of covalently bonding said release tag compound to a labelable substance.

17. A release tag compound for use in chemical analysis, said compound having the formula ##STR105## wherein R₁ is H, or an alkyl, aromatic, or alkyl-aromatic group;

R₃ and R₄ each comprises at least one moiety selected from the group consisting of alkyl radicals of 1-12 carbon atoms, alicyclic radicals of 3-7 carbon atoms, and phenyl radicals;

at least one said moiety containing at least one halogen or nitro substituent and being designated a signal group;

R₃ also including a halophenoxy or alkoxyphenoxy radical in the position alpha to the carbon atom joining R₃ and R₄ ; and

R₄ also comprising an alkyl radical;

the ##STR106## portion of said release tag compound is a release group wherein X is selected from the group consisting of;

a) two OH groups, to form a vicinal diol;

b) a chemical bond, to form an olefin;

c) an oxygen atom, to form an epoxide;

d) an NH group, to form an alkylidenimine group;

e) one OH and one N(R)₂ radical wherein each R is H or an alkyl group, forming an α

-hydroxy amine;

f) one OH and one ketone carbonyl, forming an α

-hydroxy ketone;

g) one ketone carbonyl and one N(R)₂ radical wherein R is H or an alkyl group, forming an α

-keto amine; and

h) an alkylidene-protected diol;

said release group being capable of undergoing cleavage to release the signal group as part of a ketone having the formula R₃ COR₄ ;

Y is a linking moiety selected from the group consisting of;

a chemical bond, a carbonyl group, an alkyl group, and an aromatic group; and

Z is a reactivity group comprising a reactive functional group capable of covalently bonding said release tag compound to a labelable substance.

18. The compound ##STR107## wherein R₃ is F or C₁ -C₆ alkoxy.

19. The compound m-pentafluorobenzyloxy-β

-methyl-cinnamic acid.

20. The compound m-pentafluorobenzyloxy-β

-methyl-cinnamoyl chloride.