# US 5,756,090 A · Oxygen activatable formulations for disinfection or sterilization

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- Number: US 5,756,090 A
- Document ID: US5756090A
- Status: Granted
- Kind: A
- Application number: 08/457,524
- Filed: 1995-06-01
- Granted: 1998-05-26
- Earliest claimed: 1991-02-21
- Assignees: EOE, Inc.
- Inventors: Robert C. Allen
- Web page: https://patentgrep.com/patent/US5756090A

## CPC

- A61L 2/186: Peroxide solutions
- A61K 31/198: Alpha-amino acids, e.g. alanine or edetic acid [EDTA] (betaine A61K31/205; proline A61K31/401; tryptophan A61K31/405; histidine A61K31/4172; peptides not degraded to individual amino acids A61K38/00)
- A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
- A61L 2/16: using chemical substances
- A61L 2/22: Phase substances, e.g. smokes or aerosols
- A61L 12/082: {in combination with specific enzymes}
- A61L 12/126: {neutralised with catalase or peroxidase}
- A61L 15/38: containing enzymes
- A61L 15/46: Deodorants or malodour counteractants, e.g. to inhibit the formation of ammonia or bacteria
- A61P 27/02: Ophthalmic agents
- A61P 31/04: Antibacterial agents
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C12N 9/0065: {acting on hydrogen peroxide as acceptor (1.11)}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61L 2103/15: Laboratory, medical or dentistry appliances, e.g. catheters or sharps
- A61L 2202/122: Chambers for sterilisation
- A61L 2300/102: Metals or metal compounds, e.g. salts such as bicarbonates, carbonates, oxides, zeolites, silicates
- A61L 2300/106: Halogens or compounds thereof, e.g. iodine, chlorite
- A61L 2300/11: Peroxy compounds, peroxides, e.g. hydrogen peroxide
- A61L 2300/254: Enzymes, proenzymes
- A61L 2300/404: Biocides, antimicrobial agents, antiseptic agents
- 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)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Methods and compositions are disclosed for producing air-activated, i.e., oxygen (O.sub.2) activated, disinfectant-sterilent solutions. Solutions containing a haloperoxidase (i.e., a halide:hydrogen peroxide (H.sub.2 O.sub.2) oxidoreductase, such as myeloperoxidase, eosinophil peroxidase or lactoperoxidase) plus a halide or combination of halides (i.e., chloride, bromide and/or iodide), an oxidase (i.e., a substrate: O.sub.2 oxidoreductase) capable of generating H.sub.2 O.sub.2, and a substrate specific for that oxidase, are separately prepared under aerobic conditions, but all of the component solutions are made anaerobic prior to final combination and mixing. The anaerobic formulations are dispensed into containers capable of maintaining the anaerobic condition (e.g., pressurized canisters). Dispensing the solution at the time of use exposes the formulation to air (i.e., O.sub.2) which activates its disinfectant-sterilent properties. O.sub.2 is the rate limiting component for oxidase generation of H.sub.2 O.sub.2. Under aerobic conditions the oxidase catalyzes the oxidation of its substrate and the reduction of O.sub.2 to generate H.sub.2 O.sub.2. In turn, H.sub.2 O.sub.2 serves as substrate for haloperoxidase which catalyzes the oxidation of halide to hypohalous acid. Hypohalous acid reacts with an additional H.sub.2 O.sub.2 to generate singlet molecular oxygen (.sup.1 O.sub.2). Hypohalous acid (e.g., hypochlorous acid) and especially .sup.1 O.sub.2 are potent microbicidal agents. Both haloperoxidase generation of hypohalous acid and its reactive consumption to yield .sup.1 O.sub.2 are dependent on the availability of H.sub.2 O.sub.2. A high rate of H.sub.2 O.sub.2 generation does not result in the accumulation of hypohalous acid, but instead results in a high rate of .sup.1 O.sub.2 production. The microbicidal capacity and toxicity of .sup.1 O.sub.2 are limited by the half-life of this metastable electronically excited reactant, and as such, disinfectant-sterilent activity is temporally defined by and confined to the dynamics of oxidant generation.

## Claims

### Claim 1

A hermetically sealed container comprising, under substantially anaerobic conditions, an antimicrobial formulation comprising a haloperoxidase, a halide, a peroxide generating agent capable of generating peroxide upon exposure to oxygen, and an antimicrobial activity enhancing agent of the formula: ##STR5## wherein R.sub.1 is hydrogen, an unsubstituted or hydroxy or amino substituted straight or branched chain alkyl group having from 1 to 6 carbon atoms, and R.sub.2 is hydrogen or a straight or branched chain alkyl group having from 1 to 6 carbons, and means for releasing the formulation from the container.

### Claim 2 (depends on 1)

The container of claim 1 wherein the haloperoxidase is selected from the group consisting of myeloperoxidase, eosinophil peroxidase and lactoperoxidase.

### Claim 3 (depends on 1)

The container of claim 1 wherein the halide is selected from the group consisting of chloride, bromide, iodide and combinations thereof.

### Claim 4 (depends on 1)

The container of claim 1, wherein the peroxide generating agent is an enzyme capable of oxidizing a substrate and reducing oxygen to hydrogen peroxide.

### Claim 5 (depends on 4)

The container of claim 4 wherein the enzyme is glucose oxidase and the substrate is glucose.

### Claim 6 (depends on 1)

The container of claim 1 wherein the haloperoxidase is myeloperoxidase.

### Claim 7 (depends on 1)

The container of claim 1 wherein the haloperoxidase is eosinophil peroxidase and the halide is selected from the group consisting of bromide, iodide and combinations thereof.

### Claim 8 (depends on 1)

The container of claim 1 wherein the haloperoxidase is lactoperoxidase and the halide is selected from the group consisting of bromide, iodide and combinations thereof.

### Claim 9 (depends on 6)

The container of claim 6 which comprises at least 0.01 pmol/ml of myeloperoxidase in a liquid carrier.

### Claim 10 (depends on 6)

The container of claim 6 which comprises from 0.1 pmol/ml to 500 pmol/ml of myeloperoxidase.

### Claim 11 (depends on 7)

The container of claim 7 which comprises at least 0.01 pmol/ml of eosinophil peroxidase in a liquid carrier.

### Claim 12 (depends on 7)

The container of claim 7 which comprises from 0.1 pmol/ml to 500 pmol/ml of eosinophil peroxidase.

### Claim 13 (depends on 1)

The container of claim 1 wherein the halide is chloride and the formulation comprises from 100 nmol/ml to 300 .mu.mol/ml chloride.

### Claim 14 (depends on 1)

The container of claim 1 wherein the halide is bromide and the formulation comprises from 10 nmol/ml to 50 .mu.mol/ml bromide.

### Claim 15 (depends on 1)

The container of claim 1 wherein the halide is iodide and the formulation comprises from 1 nmol/ml to 5 .mu.mol/ml iodide.

### Claim 16 (depends on 1)

The container of claim 1 wherein the peroxide generating agent comprises a peroxide producing oxidase effective to generate from 1 pmol to 50 .mu.mol peroxide per ml per minute when in the presence of a substrate from the oxidase.

### Claim 17 (depends on 1)

The container of claim 1 wherein the peroxide generating agent comprises an amount of glucose oxidase effective to generate from 1 pmol to 50 .mu.mol peroxide per ml per minute when in the presence of D-glucose.

### Claim 18

A hermetically sealed container which comprises, in the substantial absence of oxygen, an air activatable disinfectant-sterilent solution comprising:
(a) from 0.1 pmol/ml to 500 pmol/ml of a haloperoxidase selected from the group consisting of myeloperoxidase, eosinophil peroxidase, lactoperoxidase and combinations thereof;
(b) a halide selected from the group consisting of 100 nmol/ml to 300 .mu.mol/ml chloride, 10 nmol/ml to 50 .mu.mol/ml bromide, 1 nmol/ml to 5 .mu.mol/ml iodide and combinations thereof;
(c) from 0.005 .mu.mol/ml to 50 .mu.mol/ml of an .alpha.-amino acid selected from the group consisting of glycine, alanine, valine, leucine, isoleucine, serine, threonine, lysine and the alkyl esters of any members of the group;
(d) from 0.05 units/ml to 10 units/ml of an enzyme capable of oxidizing a substrate and reducing oxygen to hydrogen peroxide; and
(e) from 0.1 .mu.mol/ml to 10 .mu.mol/ml of a substrate for the enzyme;
in a liquid carrier; and means for releasing the disinfectant-sterilent solution from the container.

### Claim 19

A hermetically sealed container which comprises, in the substantial absence of oxygen, an air activatable ophthalmic solution comprising:
(a) from 0.1 .mu.mol/ml to 500 pmol/ml of a haloperoxidase selected from the group consisting of myeloperoxidase, eosinophil peroxidase, lactoperoxidase and combinations thereof;
(b) a halide selected from the group consisting of 100 nmol/ml to 300 .mu.mol/ml chloride, 10 nmol/ml to 50 .mu.mol/ml bromide, 1 nmol/ml to 5 .mu.mol/ml iodide and combinations thereof;
(c) from 0.005 .mu.mol/ml to 50 .mu.mol/ml of an .alpha.-amino acid selected from the group consisting of glycine, alanine, valine, leucine, isoleucine, serine, threonine, lysine and the alkyl esters of any members of the group;
(d) from 0.05 units/ml to 10 units/ml of an enzyme capable of oxidizing a substrate and reducing oxygen to hydrogen peroxide; and
(e) from 0.1 .mu.mol/ml to 10 .mu.nmol/ml of a substrate for the enzyme;
in a liquid carrier; and means for releasing the ophthalmic solution from the container.

### Claim 20 (depends on 13)

The container of claim 13 wherein the antimicrobial activity enhancing agent is an .alpha.-amino acid selected from the group consisting of glycine and the l- or d-enantiomers of alanine, valine, leucine, isoleucine, serine, threonine, lysine, phenylalanine, tyrosine and the alkyl esters thereof.

### Claim 21 (depends on 1)

The container of claim 1 wherein the microbicidal composition is hermetically sealed in the container for dispensing as a liquid, a foam or a gel.

### Claim 22 (depends on 1)

The container of claim 1 wherein the microbicidal composition is impregnated in a tangible substrate.