Use of substituted quinone electron transfer agents in analytical determinations

• US 4,746,607 A
• Filed: 02/07/1985
• Issued: 05/24/1988
• Est. Priority Date: 02/07/1985
• Status: Expired due to Term

• Current Assignee: Clinical Diagnostic Systems Inc.
• Original Assignee: Eastman Kodak Company
• Inventors: Belly, Robert T., Mura, Albert J., Scensny, Patricia M., Lum, Vanessa R.
• Primary Examiner(s): Marantz, Sidney
• Assistant Examiner(s): Saunders, David A.
• Application Number:
  US06/699,374
• Time in Patent Office: 1,202 Days
• Field of Search: 422/56, 422/57, 422/60, 436/63, 436/93, 436/120, 436/169, 436/170, 436/172, 436/518, 436/904, 435/25, 435/26, 435/28, 435/29, 435/34, 435/805, 436/94
• US Class Current: 435/25
• CPC Class Codes: C12Q 1/04 Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor C12Q1/6897 takes precedence
• CPC Class Codes: C12Q 2304/26 Quinone; Quinol
• CPC Class Codes: G01N 33/52 Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72
• CPC Class Codes: G01N 33/521 Single-layer analytical elements
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 436/904 Oxidation - reduction indicators
• CPC Class Codes: Y10T 436/142222 Hetero-O [e.g., ascorbic acid, etc.]

Abstract

Certain substituted benzo- and naphthoquinone electron transfer agents are useful in analytical compositions, elements and methods, e.g. for determinations of living cells. These electron transfer agents are capable of being reduced by an analyte, and the reduced electron transfer agent, in turn, reduces another compound providing a detectable species (e.g. a dye). The reduction potential (E_1/2) of the electron transfer agents useful inthis invention is in the range of from about -320 to about +400 mV as measured in an aqueous buffer solution at pH 7.

Claims

1. A composition for the determination of an analyte in a liquid, said composition comprising:

(a) an at least partially substituted benzo- or naphthoquinone electron transfer agent (ETA) which is capable of being reduced by said analyte and has an E_1/2 of from about -320 to about +400 mV as measured in an aqueous buffer solution at pH 7,

(b) a reducible compound which provides a detectable species when reduced by said reduced ETA, and

(c) a buffer which maintains the pH of said composition at pH 9 or less,

said reducible compound being selected from the group consisting of

tetrazolium salts

dichloroindophenol dyes, and

compounds of the structure CAR--R¹)_n wherein CAR-- is an aromatic or quinone nucleus, R¹ is a moiety which comprises a shiftable detectable species, and n is 1 or 2, provided that said reducible compound is capable of being reduced at a pH of 9 or less to release said shiftable detectable species, and further provided that when R¹ is replaced with H, CAR--H)_n has an E1/2 of either at least about +100 mV when measured in water, or of at least about -650 mV when measured in acetonitrile.

2. A composition for the determination of an analyte in a liquid, said composition comprising:

(a) an at least partially substituted benzo- or naphthoquinone electron transfer agent (ETA) which is capable of being reduced by said analyte and has an E₁₇₈ of from about -320 mV to about +400 mV as measured in buffer at pH 7, and

(b) a reducible compound represented by the structure

CAR--R¹)_n wherein CAR-- is an aromatic or quinone nucleus, R¹ is a moiety which comprises a shiftable detectable species, and n is 1 or 2,

provided that said reducible compound is capable of being reduced at a pH of 9 or less to release said shiftable detectable species, and

further provided that when R¹ is replaced with H, CAR--H)_n has an E_1/2 of either at least about +100 mV when measured in water, or of at least about -650 mV when measured in acetonitrile.

3. The composition of claim 2 wherein said reducible compound is a reducible intramolecular nucleophilic displacement (RIND) compound having the structure CAR--R¹ wherein CAR-- is ##STR24## R² and R⁴ are independently hydrogen, alkyl, aryl or an electron withdrawing group,

R³ is R¹, hydrogen, alkyl, aryl or an electron withdrawing group, or R³ and R⁴, taken together, represent the atoms necessary to complete a fused carbocyclic ring,

R⁵ is alkylene of 1 or 2 carbon atoms,

R⁶ is alkyl, cycloalkyl or aryl,

Q is carbonyl or thiocarbonyl,

FRAG is a shiftable detectable species which provides a detectable species when released from said reducible compound, and

m is 0 or 1,

provided when R¹ is replaced with H, CAR--H has an E_1/2 of either at least about +100 mV when measured in water, or at least about -650 mV when measured in acetonitrile.

4. The composition of claim 2 wherein said ETA has the structure ##STR25## wherein R⁷ and R⁸ are independently hydrogen, alkyl, alkenyl, alkoxy, hydroxyalkyl, hydroxyalkoxy, alkoxyalkyl, alkoxyalkoxy, acetoxyalkyl, acetoxyalkoxy, aryl, alkaryl, a heterocycle or an heteroalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together supply the atoms to complete a 4- to 8-membered fused carbocyclic or heterocyclic ring,

provided at least one of R⁷, R⁸, R⁹ and R¹⁰ is not hydrogen.

5. The composition of claim 4 wherein R⁷ and R⁸ are independently hydrogen, alkyl, alkoxy or hydroxyalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together, supply the atoms to complete a 6- to 8-membered fused carbocyclic ring, and

said ETA has an E_1/2 of from about -185 mV to about +400 mV as measured in buffer at pH 7.

6. The composition of claim 2 wherein said ETA is 2,3,5-trimethyl-1,4-benzoquinone, 2,3-dimethyl-5-hydroxymethyl-1,4-benzoquinone or 2,3-dimethoxy-5-methyl-1,4-benzoquinone.

7. The composition of claim 3 wherein m is 1, at least two of R², R³ and R⁴ are independently electron withdrawing groups, or R³ and R⁴, taken together, represent the atoms necessary to complete a fused 5- to 7-membered carbocyclic ring, R⁵ is methylene, R⁶ is alkyl of 1 to 3 carbon atoms or phenyl, Q is carbonyl, and FRAG is a chromogen or fluorogen.

8. A dry analytical element for the determination of an analyte in a liquid, said element comprising an absorbent carrier material, an at least partially substituted benzo- or naphthoquinone electron transfer agent (ETA) which is capable of being reduced by said analyte and has an E_1/2 of from about -320 mV to about +400 mV as measured in buffer at pH 7, and a reducible compound which provides a detectable species when reduced by said reduced ETA.

9. The element of claim 8 wherein said ETA has the structure ##STR26## wherein R⁷ and R⁸ are independently hydrogen, alkyl, alkenyl, alkoxy, hydroxyalkyl, hydroxyalkoxy, alkoxyalkyl, alkoxyalkoy, acetoxyalkyl, acetoxyalkoxy, aryl, alkaryl, a heterocycle or an heteroalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together supply the atoms to complete a 4- to 8-membered fused carbocyclic or heterocyclic ring,

provided at least one of R⁷, R⁸, R⁹ and R¹⁰ is not hydrogen,

and said reducible compound is a reducible intramolecular nucleophilic displacement (RIND) compound having the structure CAR--R¹ wherein CAR-- is ##STR27## R² and R⁴ are independently hydrogen, alkyl, aryl or an electron withdrawing group,

R³ is R¹, hydrogen, alkyl, aryl or an electron withdrawing group, or R³ and R⁴, taken together, represent the atoms necessary to complete a fused carbocyclic ring,

R⁵ is alkylene of 1 or 2 carbon atoms,

R⁶ is alkyl, cycloalkyl or aryl,

Q is carbonyl or thiocarbonyl,

FRAG is a shiftable detectable species which provides a detectable species when released from said reducible compound, and

m is 0 or 1,

provided when R¹ is replaced with H, CAR--H has an E_1/2 of either at least about +100 mV when measured in water, or at least about -650 mV when measured in acetonitrile.

10. The element of claim 9 wherein at least two of R², R³ and R⁴ are independently electron withdrawing groups, or R³ and R⁴, taken together, represent the atoms necessary to complete a fused 5- to 7-membered carboxylic ring, R⁵ is methylene, R⁶ is alkyl of 1 to 3 carbon atoms or phenyl, Q is carbonyl, FRAG is a chromogen or fluorogen,

R⁷ and R⁸ are independently hydrogen, alkyl, alkoxy or hydroxyalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together, supply the atoms to complete a 6- to 8-membered fused carbocyclic ring,

and said ETA has an E_1/2 of from about -185 mV to about +400 mV measured in buffer at pH 7.

11. The element of claim 8 comprising an interactive composition for said analyte.

12. The element of claim 8 comprising a carbon source nutrient for living organisms.

13. A method for the determination of an analyte in a liquid, said method comprising the steps of:

A. at a pH of 9 or less, physically contacting a sample of said liquid with

an at least partially substituted benzo- or naphthoquinone electron transfer agent (ETA) which is capable of being reduced by said analyte and has an E_1/2 of from about -320 to about +400 mV as measured in an aqueous buffer solution at pH 7, and

a reducible compound which provides a detectable species upon reduction by said reduced ETA, and

B. detecting said detectable species,

said reducible compound being selected from the group consisting of

tetrazolium salts,

dichloroindophenol dyes, and

compounds of the structure CAR--R¹)_n wherein CAR-- is an aromatic or quinone nucleus, R¹ is a moiety which comprises a shiftable detectable species, and n is 1 or 2, provided that said reducible compound is capable of being reduced at a pH of 9 or less to release said shiftable detectable species, and further provided that when R¹ is replaced with H, CAR--H)_n has an E_1/2 of either at least about +100 mV when measured in water, or of at least about -650 mV when measured in acetonitrile.

14. The method of claim 13 to determine a non-living analyte in the presence of an interactive composition for said analyte.

15. The method of claim 13 to determine bacterial cells.

16. The method of claim 13 wherein said reducible compound is represented by the structure

CAR--R¹)_n wherein CAR-- is an aromatic or quinone nucleus, R¹ is a moiety which comprises a shiftable detectable species, and n is 1 or 2,

provided that said reducible compound is capable of being reduced at a pH of 9 or less to release said shiftable detectable species, and

further provided that when R¹ is replaced with H, CAR--H)_n has an E_1/2 of either at least about +100 mV when measured in water, or of at least about -650 mV when measured in acetonitrile.

17. The method of claim 16 wherein said reducible compound is a reducible intramolecular nucleophilic displacement (RIND) compound having the structure CAR--R¹ wherein CAR is ##STR28## R² and R⁴ are independently hydrogen, alkyl, aryl or an electron withdrawing group,

R³ is R¹, hydrogen, alkyl, aryl or an electron withdrawing group, or R³ and R⁴, taken together, represent the atoms necessary to complete a fused carbocyclic ring,

R⁵ is alkylene of 1 or 2 carbon atoms,

R⁶ is alkyl, cycloalkyl or aryl,

Q is carbonyl or thiocarbonyl,

FRAG is a shiftable detectable species which provides a detectable species when released from said reducible compound, and

m is 0 or 1,

provided when R¹ is replaced with H, CAR--H has an E_1/2 of either at least about +100 mV when measured in water, or at least about -650 mV when measured in acetonitrile, and

said ETA has the structure ##STR29## wherein R⁷ and R⁸ are independently hydrogen, alkyl, alkenyl, alkoxy, hyroxyalkyl, hydroxyalkoxy, alkoxyalkyl, alkoxyalkoxy, acetoxyalkyl, acetoxyalkoxy, aryl, alkaryl, a heterocycle or an heteroalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together supply the atoms to complete a 4- to 8-membered fused carbocyclic or heterocyclic ring,

provided at least one of R⁷, R⁸, R⁹ and R¹⁰ is not hydrogen.

18. The method of claim 17 wherein R⁷ and R⁸ are independently hydrogen, alkyl, alkoxy or hydroxyalkyl, and

R⁹ and R¹⁰ are independently selected from the substituents defined as R⁷ or R⁸, or taken together, supply the atoms to complete a 6- to 8-membered fused carbocyclic ring,

provided said ETA has an E_1/2 of from about -185 mV to about +400 mV as measured in an aqueous buffer solution at pH 7.

19. The method of claim 13 wherein step B is carried out within 60 minutes of step A.

20. A composition for the determination of an analyte in a liquid, said composition comprising:

(a) an at least a partially substituted naphthoquinone electron transfer agent (ETA) which is capable of being reduced by said analyte and has an E_1/2 of from about -320 to about +400 mV as measured in an aqueous buffer solution at pH 7, said naphthoquinone being represented by the structure;

##STR30## wherein SUB represents one or more electron withdrawing substituents, electron releasing substituents, or substituents which provide an aromatic or nonaromatic carbocyclic or heterocyclic ring fused with the naphthoquinone nucleus to provide said ETA with the desired E_1/2 value,

(b) a reducible compound which provides a detectable species when reduced by said reduced ETA, and

(c) a buffer which maintains the pH of said composition at pH 9 or less.