# US 9,434,975 B2 · Fluorogenic or fluorophoric compounds and uses thereof

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- Number: US 9,434,975 B2
- Document ID: US9434975B2
- Status: Granted
- Kind: B2
- Application number: 14/008,905
- Filed: 2012-03-05
- Published: 2014-07-03
- Granted: 2016-09-06
- Earliest claimed: 2011-03-30
- Assignees: 3M Innovative Properties Company
- Applicants: Stephen B. Roscoe; Shih-Hung Chou; Stephanie J. Moeller; Jesse D. Miller; Kurt J. Halverson; Jason W. Bjork
- Inventors: Stephen B. Roscoe; Shih-Hung Chou; Stephanie J. Moeller; Jesse D. Miller; Kurt J. Halverson; Jason W. Bjork
- Web page: https://patentgrep.com/patent/US9434975B2

## CPC

- 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)}
- C07D 311/16: substituted in position 7
- C07D 405/04: directly linked by a ring-member-to-ring-member bond
- C07D 409/04: directly linked by a ring-member-to-ring-member bond
- C07H 17/075: Benzo[b]pyran-2-ones
- C09B 57/02: Coumarine dyes
- C12Q 1/045: {Culture media therefor}
- G01N 33/525: {Multi-layer analytical elements}
- G01N 33/582: {with fluorescent label}
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C07D: HETEROCYCLIC COMPOUNDS (macromolecular compounds C08)
- C07H: SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS (derivatives of aldonic or saccharic acids C07C, C07D; aldonic acids, saccharic acids C07C59/105, C07C59/285; cyanohydrins C07C255/16; glycals C07D; compounds of unknown constitution C07G; polysaccharides, derivatives thereof C08B; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; sugar industry C13)
- C09B: ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES {, e.g. PIGMENTS}; MORDANTS; LAKES (fermentation or enzyme using processes to synthesise a desired chemical compound C12P)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Compounds are provided that are either fluorogenic or fluorophoric. Compositions and articles that include the compounds are also provided. Additionally, methods of detecting a microorganism using the compounds are provided. The compounds are fluorinated and can be used advantageously under acidic conditions.

## Claims

### Claim 1

A compound of Formula (I)
wherein
R1 is —(CO)—OR3, —(CO)—NR4R5, heterocyclic group, aryl, aralkyl, or cyano;
R2 is an enzyme-labile group;
R3 is alkyl, alkenyl, aryl, or aralkyl;
R4 and R5 are each independently hydrogen, alkyl, aryl, or aralkyl; and
either (a) X1 is hydrogen and X2 is fluoro or (b) X1 is fluoro and X2 is hydrogen, and wherein R2 is a sugar group, sugar acid group, amino sugar group, carboxyl-terminated amino acid group, carboxyl-terminated peptide group, ester group, ester-containing group, phosphate group, or sulfate group; and wherein either
R2 is a sugar group that is a monosaccaride group or a disaccharide group, or
R1 is —(CO)—OR3 where R3 is alkyl, or
R2 is a sugar group that is a monosaccaride group or a disaccharide group and R1 is —(CO)—OR3 where R3 is alkyl.

### Claim 2 (depends on 1)

The compound of claim 1, wherein R2 is a sugar group that is a monosaccharide group or disaccharide group.

### Claim 3 (depends on 1)

The compound of claim 1, wherein R1 is —(CO)—OR3 where R3 is an alkyl.

### Claim 4 (depends on 1)

The compound of claim 1, wherein R1 is a heterocyclic group having a five-member heterocyclic ring.

### Claim 5 (depends on 1)

The compound of claim 1, wherein R1 is a heterocyclic group having a five-member heterocyclic ring fused to a benzene ring.

### Claim 6

A composition comprising a mixture of a compound of Formula (I-1) and a compound of Formula (I-2);
wherein
R1 is —(CO)—OR3, —(CO)—NR4R5, heterocyclic group, aryl, aralkyl, or cyano;
R2 is hydroxyl or an enzyme-labile group;
R3 is alkyl, alkenyl, aryl, or aralkyl; and
R4 and R5 are each independently hydrogen, alkyl, aryl, or aralkyl.

### Claim 7 (depends on 1)

An article comprising:
a) a first layer that is a support layer;
b) a second layer adjacent to the support layer, wherein the second layer comprises a compound of Formula (I) according to claim 1.

### Claim 8 (depends on 7)

The article of claim 7, wherein the second layer comprises a mixture of a compound of Formula (I-1) and a compound of Formula (I-2)
wherein
R1 is —(CO)—OR3, —(CO)—NR4R5, heterocyclic group, aryl, aralkyl, or cyano;
R2 is hydroxyl or an enzyme-labile group;
R3 is alkyl, alkenyl, aryl, or aralkyl; and
R4 and R5 are each independently hydrogen, alkyl, aryl, or aralkyl.

### Claim 9 (depends on 7)

The article of claim 7, wherein the second layer further comprises a gelling agent, growth medium, or a combination thereof.

### Claim 10 (depends on 7)

The article of claim 7, wherein the second layer is dehydrated.

### Claim 11 (depends on 7)

The article claim 7, wherein the second layer further comprises polymeric particles.

### Claim 12 (depends on 11)

The article of claim 11, wherein the polymeric particles comprise an acrylate polymer or copolymer, styrene polymer or copolymer, cellulosic polymer or copolymer, dextran polymer or copolymer, derivatives thereof, or combinations thereof.

### Claim 13 (depends on 11)

The article of claim 11, wherein the polymeric particles have an average particle diameter in a range of 0.01 micrometers to 1 micrometer.

### Claim 14 (depends on 1)

A method of detecting a microorganism, the method comprising:
providing a compound of Formula (I) according to claim 1; and
incubating a test sample with the compound of Formula (I), wherein the test sample contains an enzyme that cleaves the R2 group of compound of Formula (I), resulting in the formation of a fluorophoric compound of Formula (Ib); and
exciting the fluorophoric compound of Formula (Ib) with a first wavelength of light and detecting light emitted at a second wavelength of light that is longer than first wavelength of light.

### Claim 15 (depends on 14)

The method of claim 14, wherein the step of incubating the test sample with the fluorogenic compound comprises adding the test sample in a liquid form to a dehydrated layer containing the fluorogenic compound of Formula (Ia) and a growth medium.

### Claim 16 (depends on 14)

The method of claim 14, wherein the method further comprises sequestering the compound of Formula (Ib) on polymeric particles.

### Claim 17 (depends on 16)

The method of claim 16, wherein the polymeric particles have an average particle diameter in a range of 0.01 micrometers to 1 micrometer.