CA1226114A - Method for disinfecting a contact lens - Google Patents

Abstract

METHOD FOR DISINFECTING A CONTACT LENS

ABSTRACT

The method comprises a bactericide having a limited period of bacteriological activity with the bactericide comprising three components including a peroxide, a peroxides and a source of donor molecules adopted to act as a substrate for the peroxidase; storing the three components in a nonreacting state to maintain the bactericide inactive and admixing the three components in a liquid carrier to cause a catalyzed reaction by said peroxides for generating free radicals from the source of donor molecules and immersing the contact lens into the solution simultaneously with the admixture.

Description

isle TROD FOR DISINFECTING A CONTACT LETS

This invention relate to a method of disinfecting a contact lens without applying heat.
9~Ck~GROUND
FED regulations requires that a contact lens be sterilized upon removal prom an eye and before replacement in an eye. Soft contact lenses may be sterilized by one of two methods. The first method is the use of heat sterilization. Heat sterilization keenest of boiling soft contact lenses in a saline solution fur a period of about 45-50 minutes. The wearer may then remove the lens directly from the saline environment and place the contact lent in the eye.
Cold sue ilization consists ox disinfecting the contact lens in the absence of heat using a sterilizing solution containing a disinfectant which chemically sterilizes the contact lens.
Commercially available cold sterilization systems or contact lenses have two major problems: they irritate the eye to some extent and they require an inconYenien~ amount of time to worn, usually about your hour The most commonly used active component of cold sterilization systems today it ChlDrhe:~idine which can toxic and irritating to the user even after rinsing the lens.
The art has long sought a method Do disinfecting a contact lens in the absence Do heat; a method not based on the use of chlorhe~idine; a method which is nontoxic and non-irritating to the eye; a method which will use a single solution; a method which works rapidly a method simple to use; a method easy to convey; a method easy to store.

page 1 HOD FUR DISINFECTING CONTACT LENS

Hydrogen peroxide has been used in the past for disinfecting contact lenses but has been largely discarded because of the irritation it causes two the eye. This irritation it due to the high concentration of hydrogen purred required to be even minimally effective. We believe that high concentrations of hydrogen purred are required in conventional systems due to the slow rate at which bactericidal components are generated from hydrogen peroxide. The present invention uses a catalyzed system supplied in powder or pill form which continuously generates bactericidal free radicals over a controlled time period when dissolved in an aqueous carrier.
The present invention uses a peroxide based system in a non-reacting dry state which it activated by the user to form a bactericide having a limited period of bacteriological activity with the bactericide including a purred a peroxides and a source of donor molecules capable of acting a a substrate for the peroxides. The present invention overcome the most obvious objection to the Us of hydrogen peroxide alone sin e the concentration of peroxide in this system is several orders of magnitude less than that Llsed in the prior art end care ~ormulatiDn of the proportion a the components of the bactericide results in the consumption of a substantial percentage of the peroxide initially exposed to the lens. The present invention results in a concentration of peroxide Jo low as to cause no irritation to the eye and therefore Utah the criteria of an ideal clinical product.
Hydrogen peroxide dissociates into free radicals which ore nave :2 THUD FOR DISINFECTING CONTACT LETS

known to be bactericidal. The rate at which free radical species are generated from the uncatalyzed decomposition of ydrogen peroxide determines the bactericidal efficacy of this lo) compound. Enzyme catalyzed reactions are known to occur 10 to I
times as rapidly as the corresponding non-enzymatic reactions. In accordance with the present invention an enzyme, periods ha been selected to catalyze the reduction of hydrogen purred for generating free radicals. The enzyme perD~:idase catalyzes the transfer of electrons from donor molecules to acceptor molecules periods. When an electron i 5 removed from e donor molecule, this molecule it transfnr~ed into a bactericidal fret radical. The free radicals generated in this process are generated at greatly elevated rates relative to the rate at which free radicals are generated from the non-en-ymatic dissociation ox purred. The present invention allows for the sterilization of a contact lens in minutes instead of hours and therefore better suits the criteria of an ideal clinical product Paradises are classified as enzymes why h act to reduce hydrogen purred The different types Do paradises are distinguished by the donor molecules they use; donor molecules supply electrons which periods donates to hydrogen purred.
In accordance with the present invention a peroxides it used to generate free radicals from donor molecules. The donor molecules must be capable of acting as a substrate for periods in generating such free radicals. The method of the present invention teaches a practical means to control the generation of free radical species prom a bactericide having a limited period of bacteriological activity. The bactericide of the present page I

SHOD FOX DISINFECTING Q CONTACT LENS

invention is formed by combining three components, vim. a a peroxide a purred and a source of donor molecules The bactericide will continuously generate free radicals over a define period of time depending upon the concentration level of each component in the bactericide.
The duration of free radical production and the amD~nt of free radicals produced can be controlled by careful formulation of the three components comprising the system As long as the enzymatic reduction of hydrogen peroxide continues, free radicals will be generated. The free radicals being generated have an extremely short lifetime and as such lust be contlnL:ou~ly generated to prolong the period of bacteriological activity. The duration of the reaction, and therefore it bactericidal lifetime, is controller via the formulation. Other factors remaining constant, the lunger the reaction occurs the greater the bactericidal of f east vines.
The method of the present invention teaches how to maintain the bactericide in a non reacting state; how to activate the bactericide at the critical moment when sterilization of the contact lens is desired and how to control the generation of tree radicals over a preselected time period to complete the sterilization ox the lens using a minimum concentration of each component so as not to result in eye irritation. Integral to the success of the invention it the subject of patient compliance in the use of the invention. Given a product which requires admixture of several components at different concentrations, it is assumed by industry that patient compliance and therefore page 4 THUD FOR DISINFECTING CONTACT LENS

successful use so such a product will be low. Therefore, of critical importance to the present invention it storage of the bactericide in a non reacting state and preferably on powder or pill form 50 as to activate the bactericide in a liquid carrier simultaneously with the immersion on the contact lens. Inherent in the powder or pill compositiorl is ease of storage, ease of use and a high level of patient compliance which suit the criteria for an ideal clinical product. In fact, it is only via the fDrmulatiDn of these components in a powder or pill which allows feasibility of this approach for contact lens steril2ation.
Thy concentration and nature of donor molecule in the bactericide is of paramount importance since the donor molecules -or products thereof are transformed in the reaction into the bactericidal agents the reaction of the donor molecules with the enzyme is the slowest step in the reaction mechanism Many donor molecules may be used either along or in combination with other donor molecules. Same donor molecules aloe preferred substrates relative to the type of peroxides selected) and within such a group of donor molecules some are preferred due to interactions that donor molecules may have with bacteria or the surrounding matrix which will enhance or detract prom their bactericidal activity. Some donor molecules will be effective against only a limited number of bacterial strains Chile other donor molecules wit 1 l exert broader bactericidal efficacy. Some donor molecules will be effective against only comparatively law concentrations of bacteria while other donor molecules will exert bactericidal action over a large rang of bacterial concentrations.
The present~lnvention incorporates a peroxide a peroxides - Daze S

I' HOD FOR DISINFECTING A CONTACT LENS

and Dunn molecule which are readily available inexpensive and easily incorporated into a powder or pill form and therefore suits the criteria of on ideal clinical product. It has been discovered in accordance with the present invention that a minimum concentration level of donor molecules ecstasy below which the bactericide is ineffective for disinfecting a contact lens it the rate of free radical production is too low to be characterized as having any noticeable bactericidal effectiveness. This minimum level for the donor molecule is at least 1~0 x 10 molar. The minimum amount of peroxides to achieve noticeable bactericidal effectiveness is n. I x 10 units per ml; for hydrogen peroxide the minimum concentration level to achieve noticeable bactericidal effectiveness i 5 10 Mel en.
The minimum concentration levels for effective bactericidal action are critically important to the invention so nice a critical aspect of sterilization solutions is the irritation caused to certain users Hydrogen peroxide is recognized to be an eye irritant. The level of pursued left exposed to the lens after the reaction has occurred can be controlled by carefully selecting the initial concentration of the three components and the time for which the reaction will occur. It it important to this invention that the concentrations of the three components be carefully selected to cause most Do the peroxide initially exposed to the contact lens to be consumed during the reaction thus reducing the possibility for irritation. That i I the components can be formulated so that the concentration Do page I
..
r SHOD FOX DISINFECTING CONTACT LENS

hydrogen peroxide limit the duration and rate of the generation of tree radicals. When this it done it allows for a dramatic depletion ox the initial concentration of hydrogen peroxide over the concentration initially exposed to the contact lens. This type of formulation is achieved by insuring that the concentration of donor molecule is large enough 50 that its concentration does not influence the rate of the generation of tree radicals and that the concentration ox periods it large enough to reduce all of the hydrogen purred which is thermodynamically capable ox being reduced, This formulation it then pseudo-fir~t order with respect to the donor molecule and driven largely to thermodynamic equilibrium by the concentration of periods, In general the preparation used for cold sterile anion of contact lenses should have maximum concentration levels ox about Owe milligram per ml or the enzyme, Owe milligram per ml for the donor molecule and 0.03'~. for hydrogen peroxide.

SEYMOUR DO THE INVENTION
The present invention provides a practical method for producing bactericidal free radicals for disinfecting contact lens over a controlled tire period by subjecting the contact lens to a mixture containing periods a peroxide and source of preselected donor molecules with the components formulated within predetermined concentration levels to cause a substantial proportion of the purred tug be consumed within such time period. The threw components ox this invention mutt be stored in a nonre~cting state preferably as a powder or pill until use.

- page 7 HOD FOR DISINFECTING CONTACT LENS

cording to the invention bactericidal free ridiculous are continuously generated in a short time period at an effective concentration by dissolving a combination of a peroxides a purred and preselected donor molecules in a liquid to rapidly and efficiently form bactericidal free radical 5. The components of this invention are maintained in a nonreactive state by string the components in a powdered or pilled form and the activated by audiometer in a suitable liquid carrier for use The components of this material are compatible with common discounts such as silica gel which might be used to prolong the shelf-life Do a powered mixture or a pill. hermetically Lulled container could a;ternGt2iy be used to prolong the she.f-life when) extended product lifetime is necessary.

DESCRIPTION OF THE PREFEk~ED E~RODIMENT
The material used as a bactericide in accordance with the method of this invention is a combination of preselected donor molecules a peroxide and a peroxides stored in a nonreactive state sulk as in pill or powder form which if, activated Jo generate bactericidal free radicals when combined to permit a catalyzed reaction by said peroxides preferably by admixture in a suitable liquid carrier such as a buffered normal saline 501 union.
The components may be rendered nonreactive by Losing a lyophilized peroxides, a solid donor molecule and a salt us other dry source of a suitable peroxide. Any solid entity which liberates a peroxide Loon dissolution which is usable by perDxidase can serve as the source of peroxide. Sodium perorate page 8 HOD FOR DISINFECTING CONTACT LENS

or aft organic derivative of hydrogen peroxide could serve a the source o; peroxide in this invention. When this is done all components can be mixed together and activated by introduction into a liquid carrier such as a mildly buffered aqueous saline solution. This system it computable n-lmeroLIs different types of buffers which allows for a formulation organoleptically compatible with the eye.
The system of this invention incorporates a peroxide as an acceptor molecule. The enzyme periods catalyzes transfer ox electrons from donor molecules to acceptor molecules. When an electron is removed from the donor molecule this molecule is transformed into a bactericidal free radical A cycle of the enzyme mechanism i 5 illustrated below:

Enzyme + HOO~pero~ide) -------------- OR I HO
Enamel Ok Ho ------------ OR I-Enzyme Enzyme OR + huh ------- Enwomb HO
Enzyme where R = hydrogen methyl or ethyl AH = donor molecule I- = free radical of donor molecule The increased rate of formation of free radicals produced by this chemical system allows for rapid generation of high concentrations of bactericidal free radicals relative to the non-catalyzed decomposition of hydrogen purred.
The purred in this invention it a material or materials page Rudy FOR DISIhlFECTING CONTACT LENS

which form hydrogen peroxide upon dissolution in a suitable carrier. Sodium purred dilemma perbDrate or other salts ox hydrogen peroxide can readily serve as the source of peroxide for this invention since these compounds form hydrogen peroxide upon dissolution. Other periods live methyl peroxide and ethyl peroxide can serve as substrate for peroxides. When compounds which generate peroxides other than hydrogen purred are used the cost is increased and no added advantage is obtained. any compound which generates a peroxide that peroxides can use to oxidize donor molecules is an acceptable source of purred for this invention this includes a large number of compounds as one skilled in the art isle recogni~c. Likewise the org~;~ic ~lol~cule 1,1-bis-lp4-dia~abicycloC~ octane purred generates hydrogen peroxide upon dissolution in water and it an acceptable source ox purred for this invention. any compound or combination of compounds that generate a peroxide upon dissolution in water which peroxides can use for its en2ymnatic reaction with donor molecules is a suitable source of peroxide dry this invention The preferred periods is horse radish periods identified ho the IT and IUP~CI Enzyme commission identification No. EKE. 1.11~1.7. Peroxides can be obtained from a wide variety of sources although other paradises p live myelopero~:idase and lactoperoxidasep can be used the cost is increased and the stability of the final product it reduced.
Commercially obtained peroxides comes lyophili~ed as a dry powder. Periods, which uses a purred as an acceptor molecule imparts to the bactericidal composition ox this invention an enormous catalytic advantage in generating active page 10 aye SHOD FOR DISINFECTING A CONTACT LENS

constituents capable ox killing selected bacteria in defined areas relative to using only hydrogen pro ire. high concentration of free radicals are produced in short time period.
for employ the reaction rate to form free radicals occurs essentially instantaneously and proceeds at a rate determined by the initial concentration of each of the three critical components of the system and the environment in which the reaction Occurs. The periods Jan come from a variety of sources and can be isolated by any of the well-~.nown prior art procedures as Llsed by the many companies which offer a periods for sale. The use of horseradish peroxides is preferred since it is easily isolated has low cost and has very high stability giving it a long lifetime however, other sources of periods can be used. Paradises have variable substrate specificities depending upon the source from which they are isolated. Hydrogen peroxide it often the most effective substrate.
The donor molecules are molecules which can be acted upon to aid in formation ox bactericidal free radials zany donor molecules can be use a will be recognized by those skilled in the art. The general substrate specificity of paradises it such that they can use phenols, aureole and allele amine, hydroquinones~ NED N~DPH, palpitate halogens, glutathione~
ferrocytochrome c and ascorbate as donor molecules. This broad enzymatic substrate specificity allows for a large choice of a single donor molecule or a combination of several Different donor molecules have different abilities and reactivities and can be selected to focus bactericidal selectivity on any given page 11 ~-~26~
OX FOR DISINFECTING CONTACT LENS

preparation by careful selection of donor molecules or by designing specific donor molecules with high selectivity for specific bacteria. We have observed that certain donor molecules are effective at high levels of bacteria (l million/ml) while others are only effective at very low levels of bacteria. The following compounds have been found to be effective as donor molecules: ascorbic acid Tarzan phenylalanine~ bunk acid, salicylic acid hydroquinone~ dehydrophenylalanine7 vanilla and iodide salts like sodium iodide.
Typically the number of bacteria on a contact lens after one day of wear is between I and lC1C~; regulatory requirements can necessitate a formulation of this invention that is b~cte.-icidal to an environment which contains many orders of magnitude more bacteria than that found on a contact lens after a typical day ox use. When formulating the invention to kill large numbers of bacteria there are certain donor molecules which are more effective than others. If an environment contains a high concentration of a bacteria which secretes an active kettles then this enzyme will compete for the hydrogen purred present in the formLIlation and reduce the effectiveness of the invention.
It is therefore necessary to low something about the environment of the contact lens which it to be sterilized in order to formLIlate the invention properly.
prerequisite for the storage of any preparation it not allowing all three components donor molecLIles~ acceptor molecules) and periods) of the system to combine under conditions where the catalytic process can occur. That is it is imperative that the storage of the components will not allow the pave lo Mr--rHOD FOR DISINFECTING A CONTACT LENS I

depletion of the component parts of the system until the reaction is initiated immediately prior to its use If the components are allowed to react before intended for use the combination of these components under such conditions will precipitate the depletion of the enemies substrate molecules and thereby attenuate the effectiveness of the preparation. Any combination of the components of this system donor molecules, acceptor molecules or periods) which precludes the catalytic reaction from occurring is acceptable for storage prior to use. That is if it it practical to separate any one of the three components from the other two prior to administration, this would serve the purpose ox preserving the integrity of the system. alternately it is possible to have two separate mysterious which contain any two of the components of the system in any combination and to combine these two mixtures prior to use. The present invention accomplishes this by combining the three components of the invention in a dry form.
Toe presort invent on can utilize a concentration of purred preferably hydrogen pursued, ranging from 1 millimolar to to 1 micro molar with a preferred concentration range between Owl millimolar and Oily millimolar. The present invention can utilize concentrations of donor molecule ranging from 100 millimolar to 10 micromDlar with a preferred range of I
micro molar to 10 millimolar. The present invention can utilize a concentration range of horseradish periods from O.OOOOlmg~ml to 1 mg~ml with a preferred range of 0.5 to O.Olmg~ml.

page I

F HOD FOR DISINFECTING CONTACT LENS 12X61i4 EXPELS

Example l - Four patients used vows containing 0,C19 my sodium peroxide I my Nikolai ~.12 my L-tyrosine and 5 units of HOP. To sterilize their contacts patients placed their contact lens in the vial containing these components and added 10 ml of distilled water The contents were gently mixed and exposed to the contact lens for to 5 minutes The lens was then removed from the vial and rinsed in distilled water. There was no clinical discomfort or danger to any of the patients in this study.

Example Al - contact lens was exposed to ten ml of normal saline containing Okay my sodium peroxide 20 my Nikolai 0.1~ my L-Tarzan and 5 units of HOP for several lolls minute sterilization cycles at room temperature To determine if there were any deleterioLIs effects to the lens electron micro graphs were Tony and the surface morphology of this lens was compared to a lens a patient had sterilized in saline with a commercially allele Sheldon me_hndolcgy~ there wan no difference between the two in terms of the polymers structLlre.

Example - after a day offer a Wesley-Jensen Direst lens was divided into your equal sections. One section was plated I

sheep-blaod agree out direct One section was immersed in 5 x lo molar peroxide for 5 minutes and then plated out One section was sterilized using a standard Bullish I< Lomb heat sterilizer and then plated out. One section was treated with S x molar peroxide 1 x 10 molar Tarzan and 2.5 x 10 molar HOP Hall components in 0.010 molar sodium phosphate 0~15 molar page 14 THEA I DISINFECTING CONTACT Lucy .~2Z61~

Nikolai pi 6.8) for 5 minutes and then plated out. ill lens sections were sterilized at room temperature with the exception of the Bausch Lomb heat sterilizer. The plates were grown our 5 days at 37 C and then counted. The only plates which did not have growth were prom the contacts sterilized with heat and the contact sterilized in accordance with this invention.

Employ 4 - contact way contaminated with a pure hospital culture o-f S. Ayers! episode to a ten ml of normal saline containing 0.0~ my sodium peroxide, 20 my Nikolai 0.l2 my L-Tarzan and 5 ur1its of HOP for five minutes at room temperature and the: plated out by the -..bDve procedure; no growth we-observed.

.

Example 5 - lamely of buffer, sodium phosphate pi 7.0~ Oslo molar was pipette into two lo x 75mm test tubes. loop was flamed and inserted into the buffer of one o-f the two test tubes a colony us staph. Ayers cells was removed from a pure slant and placed no the l quad of on ox those two Tut Tokyo. Tess sty tube was shaken to evenly suspend the cells and then placed in a clinical centrifuge to spin at to 4 thousand rum.
I ~icroliters of potassium iodide, lo milli~olar, Wow pipette into each of the test tubes to be Swede 70 micro liters of Oslo molar sodium phosphate, pi 7~0 was pipette into each us the test tubes to be used. The centrifuge containing the Staph.
cells way stopped and the supernatant was poured off; the 5t~ph.
cells were resuspended in phosphate buffer lamely 7 and spun in the centrifuge again at to 4 thousand rum.

page 15 HOD FOR DISINFECTING CUNT~CT LENS i2261~

Hydrogen purred lo millimolar was diluted serially by factors of ten to a L~ncentratiorl of lo nanomolar; JO
micro liters of the serial dilution ox hydrogen peroxide were added to individual test tubes containing both 7C~ micro liters of phosphate buffer and potassium iodide.
The centrifuge containing the Staph. cells was stopped and the supernatant was poured off; the Staph. cells were resuspended in phosphate buffer lamely, and spun in the centrifuge again at 3 to 4 thousand rum.
Horse radish periods was weighed out and dissolved in phosphate buffer to a concentration of OHS my per my The synergy was stopped and tube supernatant of the jest tube containing the Staph. cells was poured of. The cells were suspended in lamely ox phosphate buffer and their optical density way measured in a Go 1 f or spectrophotometer at 60~ no. The optical density was recorded and the cell suspension was diluted until the optical density was 0.05 ODE after this was done ZOO
micro liters of this suspension of cells was added to each test tube.
O Micro liters of the solution of horse radish periods was adder to each test tube and the test tubes were gently Chilean. The test tubes were allowed is incubate for 45 minutes at I degrees centigrade. After the incubation time lamely Do phosphate buffer was pipette into a test tube. calibrated loop was flamed placed into the test tube. plate of 5%-sheep blood Gore was streaked or each test tube in the experiment. The colonies on each plate were counted and compared to control plates which had not received periods, positive control, or page I

MF-lHUD O'ER DISlNFECrING A CONTACT LENS I

cells negative control.
The positive control had over lC)C~ colonies on its surface the negative control had no colonies on its carafes, all plates with a final Concentration of hydrogen peroxide in the test 501 UtiDn greater than or equal to 1 micro molar had no colonies or, their surface.

Employ 6 - The protocol of example 5 was followed except that serial dilutions of horse radish peroxides and hydrogen peroxide were made the enzymatic reaction was alloyed to proceed only for five minutes at room temperature before being plated out and 1 million bacteria per ml of S. a -eye were used in the assay. The minimum concentration of hydrogen purred that was necessary to I
-Jill all the bacteria was 10 M in the final reaction. the minimum concentration of enzyme sigma 150 unitsJmg) necessary to kill all bacteria present was O.Ol~mg~ml.

Example 7- The protocol of experiment 5 was followed except that the hyclr~gen peroxide concentration and horse ravish peroxide concentration were kept at 0~10 millimolar and 0.~5mg~ml (I50units/mg). The concentration of potassium iodide was serially diluted by factors of ten. The minimum concentration of potassium iodide necessary to insure complete bactericidal action was 35 micro molar.

page 17

Claims (11)

???HOD FOR DISINFECTING A CONTACT LENS

1. method for disinfecting a contact lens comprising forming bactericide having a limited period of bacteriological activity, said bactericide comprising three components including a peroxide, a peroxidase and a source of predetermined donor molecules adapted to act as a substrate for said peroxidase, storing said three components in a nonreacting state such that said batericide is rendered inactive; admixing the three components in a liquid carrier to cause a catalyzed reaction by said peroxidase for generating free radicals from said source of donor molecules; selecting the concentration level of said three components such that a substantial percentage of said peroxide is consumed during said catalyzed reaction; and immersing the contact lens into said solution substantially simultaneous with the admixture of all three components whereby bacteria present on said contact lens will be killed during said limited period of bacteriological activity.

2. A method as defined in claim 1 wherein said components are stored in a dry form such as a powder or in a pill needing only to be dissolved in a suitable liquid carrier to be activated.

3. A method as defined in claim 1 wherein said source of donor molecule is selected from the class consisting of iodide salts, tyrosine, phenylalanine, benzoic acid dehydrophenylalanine and vanillan.

4. A method as defined in claim 3 wherein the concentration of said donor molecule is in a concentration range from 10 micromolar to 100 millimolar.

page 18 METHOD FOR DISINFECTING A CONTACT LENS

5. A method as defined in claim 4 wherein said source of peroxide is selected from the class consisting of hydrogen peroxide, sodium perborate, methyl peroxide, ethyl peroxide or any compound which generates a peroxide that peroxidase can use to oxidize donor molecules.

6. A method as defined in claim 5 wherein said peroxide concentration is in a range of between 1 millimolar to 1 micromolar.

7. A method as defined in claim 6 wherein said source of peroxidase is horseradish peroxidase.

8. A method as defined in claim 7 wherein the concentration of said peroxidase is in a range between 0.00001 and 1.0 mg/ml.

9. A method as defined in claim 8 wherein said liquid carrier comprises water or a saline solution.

10. A method as defined in claim 2, 4 or 9 wherein said source of donor molecules is a salt of iodide.

11. A method as defined in claim 5 wherein said source of peroxide is sodium perborate.

page 19