# US 2010/0168236 A1 · Production of peracids using an enzyme having perhydrolysis activity

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- Number: US 2010/0168236 A1
- Document ID: US20100168236A1
- Status: Published application
- Kind: A1
- Application number: 12/722,302
- Filed: 2010-03-11
- Published: 2010-07-01
- Earliest claimed: 2005-12-13
- Assignees: E. I. Du Pont de Nemours and Company
- Inventors: Robert Dicosimo; John E. Gavagan; Mark S. Payne; Frederick B. Cooling, III
- Web page: https://patentgrep.com/patent/US20100168236A1

## CPC

- A01N 37/02: Saturated carboxylic acids or thio analogues thereof; Derivatives thereof
- A01N 37/16: containing the group [IMAGE cpc-sch-A01N-0934.gif]; Thio analogues thereof
- A01N 37/36: containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a singly bound oxygen or sulfur atom attached to the same carbon skeleton, this oxygen or sulfur atom not being a member of a carboxylic group or of a thio analogue, or of a derivative thereof, e.g. hydroxy-carboxylic acids
- A01N 59/00: Biocides, pest repellants or attractants, or plant growth regulators containing elements or inorganic compounds
- A61L 2/186: Peroxide solutions
- C12P 7/40: containing a carboxyl group {including Peroxycarboxylic acids}
- C12P 7/62: Carboxylic acid esters
- A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A61L: METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES (preservation of bodies or disinfecting characterised by the agents employed A01N; preserving, e.g. sterilising, food or foodstuffs A23; preparations for medical, dental or toiletry purposes A61K)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}

## Abstract

A process is provided for producing peroxycarboxylic acids from carboxylic acid esters. More specifically, carboxylic acid esters are reacted with an inorganic peroxide, such as hydrogen peroxide, in the presence of an enzyme catalyst having perhydrolysis activity. The present perhydrolase catalysts are classified as members of the carbohydrate esterase family 7 (CE-7) based on the conserved structural features. Further, disinfectant formulations comprising the peracids produced by the processes described herein are provided.

## Claims

### Claim at position 1

1-9. (canceled)

### Claim 10

A process to disinfect a hard surface or inanimate object using an enzymatically produced peroxycarboxylic acid composition, said process comprising:
a) providing a set of reaction components comprising:
1) at least one substrate selected from the group consisting of:
i) esters having the structure
[X]mR5
wherein X=an ester group of the formula R6C(O)O;
R6=C1 to C7 linear, branched or cyclic hydrocarbyl moiety, optionally substituted with hydroxyl groups or C1 to C4 alkoxy groups, wherein R6 optionally comprises one or more ether linkages for R6=C2 to C7;
R5=a C1 to C6 linear, branched, or cyclic hydrocarbyl moiety optionally substituted with hydroxyl groups; wherein each carbon atom in R5 individually comprises no more than one hydroxyl group or no more than one ester group; wherein R5 optionally comprises one or more ether linkages;
m=1 to the number of carbon atoms in R5; and
wherein said esters have a solubility in water of at least 5 ppm at 25° C.;
ii) glycerides having the structure
wherein R1=C1 to C7 straight chain or branched chain alkyl optionally substituted with an hydroxyl or a C1 to C4 alkoxy group and R3 and R4 are individually H or R1C(O); and
iii) acetylated saccharides selected from the group consisting of acetylated monosaccharides, acetylated disaccharides, and acetylated polysaccharides;
2) a source of peroxygen; and
3) an enzyme catalyst comprising an enzyme, wherein said enzyme has perhydrolase activity and comprises a polypeptide having at least 95% identity to the amino acid sequence set forth in SEQ ID NO:90;
b) combining said reaction components under suitable aqueous reaction conditions whereby a peroxycarboxylic acid is produced; and
c) contacting said hard surface or inanimate object with the peroxycarboxylic acid whereby said surface or said inanimate object is disinfected.

### Claim 11 (depends on 10)

The process of claim 10 further comprising diluting said peroxycarboxylic acid produced by said combining of said reaction components.

### Claim 12 (depends on 10)

The process of claim 10 wherein said enzyme is encoded by a nucleic acid molecule that hybridizes to SEQ ID NO: 88 or SEQ ID NO: 89 under the following conditions: 0.1×SSC, 0.1% SDS at 65° C. and washed with 2×SSC, 0.1% SDS at 65° C., followed by a second wash with 0.1×SSC, 0.1% SDS at 65° C.

### Claim 13 (depends on 10)

The process of claim 10 wherein the enzyme having perhydrolase activity comprises the amino acid sequence set forth in SEQ ID NO: 90.

### Claim 14 (depends on 10)

The process of claim 10 wherein the peroxycarboxylic acid is produced at a concentration of at least 20 ppm within about 5 minutes to about 2 hours of combining the reaction components.

### Claim 15 (depends on 10)

The process of claim 10 wherein the substrate is monoacetin; diacetin; triacetin; monopropionin; dipropionin; tripropionin; monobutyrin; dibutyrin; tributyrin; glucose pentaacetate; xylose tetraacetate; acetylated xylan; acetylated xylan fragments; 13-D-ribofuranose-1,2,3,5-tetraacetate; tri-O-acetyl-D-galactal; tri-O-acetyl-glucal; monoesters or diesters of 1,2-ethanediol, 1,2-propanediol, 1,3-propanediol, 1,2-butanediol, 1,3-butanediol, 2,3-butanediol, 1,4-butanediol, 1,2-pentanediol, 2,5-pentanediol, 1,6-pentanediol, 1,2-hexanediol, 2,5-hexanediol, 1,6-hexanediol; or a mixture thereof.

### Claim 16 (depends on 10)

The process of claim 10 wherein the peroxycarboxylic acid produced is peracetic acid, perpropionic acid, perbutyric acid, perlactic acid, perglycolic acid, permethoxyacetic acid, per-β-hydroxybutyric acid, or a mixture thereof.

### Claim 17 (depends on 10)

The process of claim 10 wherein the peroxycarboxylic acid produced is peracetic acid.

### Claim 18 (depends on 10)

The process of claim 10 wherein the hard surface or the inanimate object is contacted with the peroxycarboxylic acid within about 5 minutes to about 168 hours of combining said reaction components.

### Claim 19 (depends on 10)

The process of claim 10 wherein the hard surface or the inanimate object is contacted with the peroxycarboxylic acid within about 5 minutes to about 48 hours of combining said reaction components.

### Claim 20 (depends on 10)

The process of claim 10 wherein the hard surface or the inanimate object is contacted with the peroxycarboxylic acid within about 5 minutes to about 2 hours of combining said reaction components.

### Claim 21 (depends on 10)

The process according to claim 10 wherein a concentration of viable biological contaminants on the hard surface or the inanimate object is reduced at least 3-log.

### Claim 22 (depends on 10)

The process according to claim 10 wherein a concentration of viable biological contaminants on the hard surface or the inanimate object is reduced at least 5-log.

### Claim 23 (depends on 10)

The process of claim 10 wherein the enzyme catalyst is in the form of a microbial cell, a permeabilized microbial cell, a microbial cell extract, a partially purified enzyme, or a purified enzyme.

### Claim 24 (depends on 10)

The process of claim 10 wherein the enzyme catalyst lacks catalase activity.

### Claim 25

A process to disinfect a hard surface or inanimate object using an enzymatically produced peroxycarboxylic acid composition, said process comprising combining on said hard surface or inanimate object under suitable aqueous reaction conditions a set of reaction components comprising:
1) at least one substrate selected from the group consisting of:
i) esters having the structure
[X]mR5
wherein X=an ester group of the formula R6C(O)O;
R6=C1 to C7 linear, branched or cyclic hydrocarbyl moiety, optionally substituted with hydroxyl groups or C1 to C4 alkoxy groups, wherein R6 optionally comprises one or more ether linkages for R6=C2 to C7;
R5=a C1 to C6 linear, branched, or cyclic hydrocarbyl moiety optionally substituted with hydroxyl groups; wherein each carbon atom in R5 individually comprises no more than one hydroxyl group or no more than one ester group; wherein R5 optionally comprises one or more ether linkages;
m=1 to the number of carbon atoms in R5; and
wherein said esters have a solubility in water of at least 5 ppm at 25° C.;
ii) glycerides having the structure
wherein R1=C1 to C7 straight chain or branched chain alkyl optionally substituted with an hydroxyl or a C1 to C4 alkoxy group and R3 and R4 are individually H or R1C(O); and
iii) acetylated saccharides selected from the group consisting of acetylated monosaccharides, acetylated disaccharides, and acetylated polysaccharides;
2) a source of peroxygen; and
3) an enzyme catalyst comprising an enzyme wherein said enzyme has perhydrolase activity and comprises a polypeptide having at least 95% identity to the amino acid sequence set forth in SEQ ID NO:90;
thereby generating peroxycarboxylic acid, whereby said hard surface or inanimate object is disinfected.

### Claim 26 (depends on 25)

The method of claim 25 wherein said peroxycarboxylic acid is diluted.

### Claim 27 (depends on 25)

The process of claim 25 wherein enzyme is encoded by a nucleic acid molecule that hybridizes to SEQ ID NO: 88 or SEQ ID NO: 89 under the following conditions: 0.1×SSC, 0.1% SDS at 65° C. and washed with 2×SSC, 0.1% SDS at 65° C., followed by a second wash with 0.1×SSC, 0.1% SDS at 65° C.

### Claim 28 (depends on 25)

The process of claim 25 wherein the enzyme having perhydrolase activity comprises the amino acid sequence set forth in SEQ ID NO: 90.

### Claim 29 (depends on 25)

The process of claim 25 wherein the peroxycarboxylic acid is produced at a concentration of at least 20 ppm within about 5 minutes to about 2 hours of combining the reaction components.

### Claim 30 (depends on 25)

The process of claim 25 wherein the substrate is monoacetin; diacetin; triacetin; monopropionin; dipropionin; tripropionin; monobutyrin; dibutyrin; tributyrin; glucose pentaacetate; xylose tetraacetate; acetylated xylan; acetylated xylan fragments; β-D-ribofuranose-1,2,3,5-tetraacetate; tri-O-acetyl-D-galactal; tri-O-acetyl-glucal; monoesters or diesters of 1,2-ethanediol, 1,2-propanediol, 1,3-propanediol, 1,2-butanediol, 1,3-butanediol, 2,3-butanediol, 1,4-butanediol, 1,2-pentanediol, 2,5-pentanediol, 1,6-pentanediol, 1,2-hexanediol, 2,5-hexanediol, 1,6-hexanediol; or a mixture thereof.

### Claim 31 (depends on 25)

The process of claim 25 wherein the peroxycarboxylic acid produced is peracetic acid, perpropionic acid, perbutyric acid, perlactic acid, perglycolic acid, permethoxyacetic acid, per-β-hydroxybutyric acid, or a mixture thereof.

### Claim 32 (depends on 25)

The process of claim 25 wherein the peroxycarboxylic acid produced is peracetic acid.

### Claim 33 (depends on 25)

The process according to claim 25 wherein a concentration of viable biological contaminants on the hard surface or the inanimate object is reduced at least 3-log.

### Claim 34 (depends on 25)

The process according to claim 25 wherein a concentration of viable biological contaminants on the hard surface or the inanimate object is reduced at least 5-log.

### Claim 35 (depends on 25)

The process of claim 25 wherein the enzyme catalyst is in the form of a microbial cell, a permeabilized microbial cell, a microbial cell extract, a partially purified enzyme, or a purified enzyme.

### Claim 36 (depends on 25)

The process of claim 25 wherein the enzyme catalyst lacks catalase activity.

### Claim 37

(canceled)