Note: Descriptions are shown in the official language in which they were submitted.
CA 02307664 2000-04-28
WO 99/24542 PCTIUS98/23818
DISINFECTING CONTACT LENSES WITH BIS(BIGUANIDES) AND POLYMERIC BIGUANIDES
Field of the Invention
This invention relates to new and improved solutions for the treatment of
contact
lenses and to methods for treating contact lenses with such solutions. In
particular, the
present invention is directed to disinfecting systems comprising the novel
combination of
two disinfecting agents, namely a bis{biguanide) and a polymeric biguanide.
Background of the Invention
Generally, contact lenses in wide use fall into three categories: (1) hard
lenses
formed from materials prepared by polymerization of acrylic esters, such as
polymethyl
methacrylate (PMMA), (2) rigid gas permeable (RGP) lenses formed from silicone
acrylates and fluorosilicone methacrylates, and (3) gel, hydrogel or soft type
lenses made
of polymerized hydrophilic or hydrophobic monomers, such as 2-hydroxyethyl
methacrylate (HEMA). The hard acrylic type contact lenses are characterized by
low
water vapor diffusion constants, resistance to the effects of light, oxygen
and hydrolysis,
and absorb only minor amounts of aqueous fluids. Because of the durability of
hard
contact lenses, coupled with their tendency not to absorb appreciable amounts
of water,
the selection of suitable disinfecting agents, cleaning agents or other lens
care
compounds is relatively non-critical.
However, unlike hard lenses, soft-type contact lenses have a tendency to hind
and
concentrate significantly more fluids, environmental pollutants, water
impurities, as well
as antimicrobial agents and other active ingredients commonly found in lens-
care
solutions. In most instances, the low levels of the ingredients in lens-care
solutions do
not lead to eye tissue irritation when used properly. Nevertheless, especially
due to the
inherent binding action of protein deposits to soft-lens materials, some
disinfecting
agents and preservatives tend to build up on lens surfaces and may become
concentrated
to potentially hazardous levels, such that when released could cause corneal
inflammation and other eye tissue irntation.
CA 02307664 2000-04-28
WO 99/24542 PCT/US98123818
Certain antibacterial agents were found to be more compatible with contact
lenses
and exhibit less binding on lens surfaces. In one case, it was found that
chlorhexidine, a
biguanide, binds to soft lens material seven times less than benzalkonium
chloride. The
presence of proteinaceous oily tear-film deposits on a lens, however, can
double the
amount of chlorhexidine absorbed on the lens compared to a clean lens. U.S.
patent
4,354,952 discloses very dilute disinfecting and cleaning solutions containing
chlorhexidine or its salt in combination with certain amphoteric and non-ionic
surfactants. These solutions were found to reduce the amount of binding of
chlorhexidine on hydrophilic soft contact lenses. Notwithstanding the
reduction in
binding achieved by this invention, the use of chlorhexidine did result in
certain
tradeoffs. The antimicrobial activity of the chlorhexidine may be diminished
when used
with certain amphoteric surfactants. Furthermore, it was reported that if not
used in
proper ratio, the surfactant and disinfectant will precipitate unless a non-
ionic type
surfactant is also employed.
British Patent 1,432,345 discloses contact lens disinfecting compositions
containing a polymeric biguanide and a mixed phosphate buffer. Compositions as
disclosed by this patent, however, have corneal staining values of 17% or
more, far
above that which is desirable for patient acceptability.
U.S. Patent 4,758,595 to Ogunbiyi et al. disclosed that a contact-lens
solution
containing a polyaminopropyl biguanide (PAPB), also known as polyhexamethylene
biguanide (PHMB), has enhanced efficacy when combined with a borate buffer.
These
disinfecting and preservative solutions are especially noteworthy for their
broad
spectrum of bactericidal and fungicidai activity at low concentrations coupled
with very
low toxicity when used with soft-type contact lenses.
U.S. Patent No. 5,453,435 to Raheja et al., disclosed a preservative system
that
comprises a combination of chlorhexidine and polyhexamethylene biguanide. This
preservative system, used in commercial products for rigid-gas-permeable
lenses, was
found to exhibit an improved combination of efficacy and low eye irritation.
Compositions containing PHMB and borate have been commercialized in various
products, but at levels of about 1 ppm or less for use with soft contact
lenses. It is
generally desirable to provide the lowest level of a bactericide possible,
while
-2-
CA 02307664 2000-04-28
WO 99124542 PCT/US98/23818
maintaining the desirable level of disinfection efficacy, in order to provide
a generous
margin for safety and comfort.
Some of the most popular products for disinfecting lenses are multipurpose
solutions that can be used to clean, disinfect and wet contact lenses,
followed by direct
insertion (placement on the eye) without rinsing. Obviously, the ability to
use a single
solution for contact-lens care is an advantage. Such a solution, however, must
be
particularly gentle to the eye, since, as indicated above, some of the
solution will be on
the lens when inserted and will come into contact with the eye.
With conventional contact-lens cleaners or disinfectants, including multi-
purpose
solutions, lens wearers typically need to digitally or manually rub the
contact lenses
(typically between a finger and palm or between fingers) during treatment of
the contact
lenses. The necessity for the daily "rubbing" of contact lenses adds to the
time and effort
involved in the daily care of contact lenses. Many contact-lens wearers
dislike having to
perform such a regimen or consider it to be an inconvenience. Some wearers may
be
negligent in the proper "rubbing" regimen, which may result in contact-lens
discomfort
and other problems. Sometimes rubbing, if performed too rigorously, which is
particularly apt to occur with beginning lens wearers, may damage the lenses.
This can
be problematic when a replacement lens is not immediately available.
Contact lens solutions that qualify as a "Chemical Disinfecting Solution" do
not
require rubbing to meet biocidal performance criteria (for destroying
representative
bacteria and fungi) set by the US Food and Drug Administration (FDA) under the
Premarket Notification (S l Ok) Guidance Document For Contact Lens Care
Products,
May l, 1997. In contrast, a contact-lens solution, referred to as a "Chemical
Disinfecting System," that does not qualify as a Chemical Disinfecting
Solution, requires
a rubbing regimen to pass biocidal performance criteria. Traditionally, mufti-
purpose
solutions (used for disinfecting and wetting or for disinfecting, cleaning,
and wetting)
have qualified as a Chemical Disinfecting System, but not as a Chemical
Disinfecting
Solution.
A Chemical Disinfecting Solution would generally require a more efficacious or
stronger disinfectant than a Chemical Disinfecting System. The stronger the
biocidal
-3-
CA 02307664 2000-04-28
WO 99124542 PCT/US98123818
effect of a solution, however, the more likely that it may exhibit toxic
effects or
adversely effect lens-wearer comfort. For example, many very efficacious
bactericides
used in other contexts, such as mouthwashes, cosmetics, or shampoos, while
being
sufficiently safe for use in such products, would be too toxic for ophthalmic
use,
especially for use with soft lenses because of the above-mentioned tendency of
soft
lenses to bind chemicals and the sensitivity of eye tissues. ~~milarly, the
concentrations
of certain bactericides may need to be within lower limits in solutions for
use with soft
contact lenses than in other products or in solutions for other types of
lenses, especially
when such solutions are not rinsed from the contact lens before placing the
lens in the
eye.
It would be desirable to obtain a contact-lens solution that would
simultaneously
provide both (1) an increased level and/or broader spectrum of biocidal
activity, and (2) a
low order of toxicity to eye tissue, such that the solution can be used to
treat a contact
lens such that the lens can subsequently be placed on the eye without rinsing
the solution
from the lens. While challenging to develop, it would be especially desirable
to obtain a
Chemical Disinfecting Solution that could be used for soft contact lenses and
that would
allow direct placement of a contact lens on an eye following soaking in the
solution
and/or rinsing and rewetting with the solution. Such a product may provide
increased
efficacy, resulting in greater protection to the lens wearer against infection
caused by
microorganisms, while providing maximum convenience. Finally, it would be
desirable
for the biocidal efficacy of the disinfecting solution to be sufficiently high
to achieve
efficacious disinfection, or at least not inherently inefficacious
disinfection, of a contact
lens with respect to bacteria and fungi in the event, for whatever reason,
that the contact
lens wearer does not carry out a regimen involving mechanical rubbing or the
like using
the contact-lens solution.
-4-
CA 02307664 2000-04-28
WO 99124542 PCT/US98/23818
Brief Description of the Invention
The present invention is directed to an ophthalmically safe disinfecting
solution
for contact lenses comprising:
(a) about 0.10 to about 4.0 ppm of a bis(biguanide) in the form of a
dihydrochloride salt, or a corresponding concentration of the same
bis(biguanide) in the
form of the free base or a different water-soluble salt, which bis(biguanide)
has the
following general formula:
Rl-NH- i -NH- i -NH-(CH2)n NH- i -NH-C-NH-R2
NH NH NH NH (I)
or their water-soluble salt form, wherein R' and RZ are independently selected
from the
group consisting of branched or unbranched alkyl, alkoxyalkyl or alkylsulfide
radical,
and n is 4 to 16; and
(b) about 0.1 to about 3.0 ppm of a polymeric biguanide having the formula:
Xl~-Z-NH- i -NH-C-NH~-Z-X'-
NH NH (II)
wherein Z is an organic divalent bridging group which may be the same or
different
throughout the polymer, n on average is at least 3, and X~ and X2 are
independently selected from the groups -NHZ and -NH - i - NH - CN.
NH
(c) an effective amount of a buffering agent; and
(d) water in an amount of at least about 80% by weight.
Preferably, the compositions of the present invention also include one or more
surfactants. In one embodiment of the invention, the surfactant is a neutral
or non-ionic
surfactant.
The invention is also directed to a method of disinfecting, or cleaning and
disinfecting, a contact lens comprising soaking the lens for a given period of
time in the
aqueous solution described above, and subsequently directly placing the
treated lens on
-5-
CA 02307664 2000-04-28
WO 99124542 PCT/US98I23818
an eye of the wearer. In one embodiment of this method, a contact lens does
not require
rubbing with the solution to achieve the necessary disinfection.
Bri f Description of the Drawings
FIG. 1 is a bar graph showing the biocidal efficacy, after 15 minutes, of a
solution containing the combination of two disinfecting agents, namely a
bis(biguanide)
and a biguanide polymer, versus the theoretical sum of the biocidal efficacy
of separate
solutions of each disinfecting agent. The increased efficacy of the
combination
compared to the theoretical sum is a measure of the synergy of the
combination. I n
particular, Fig. 1 shows the theoretical log reduction compared to the actual
log reduction
for alexidine and PHMB, with respect to C. albicans microorganisms, after 15
minutes
exposure.
FIG. 2 is a bar graph showing the biocidal efficacy, after 30 minutes, of a
solution containing the combination of a bis(biguanide) and a biguanide
polymer versus
the theoretical sum of the biocidal efficacy of separate solutions of each
disinfecting
agent. In particular, Fig. 2. shows the theoretical log reduction compared to
the actual
log reduction for alexidine and PHMB, with respect to C. albicans
microorganisms, after
30 minutes exposure.
Detailed Description of the Invention
As indicated above, the present invention is directed to a composition
involving
the combined used of a biguanide polymer and a bis(biguanide), and a method of
using
the composition, in the form of an aqueous solution, for disinfecting and/or
preserving
contact lenses, especially soft contact lenses. This synergistic combination
offers
maximum convenience while providing increased efficacy and hence better
protection
against microorganisms compared to traditional disinfecting products for
contact lenses.
The solution according to the present invention provides a broader, more
potent and
faster antimicrobial activity overall, when considering the entire range of
microorganisms, based on representative bacteria and fungi commonly tested. In
particular, the disinfecting solutions of the present invention are effective
at low
-6-
CA 02307664 2000-04-28
WO 99/24542 PCT/US98/Z3818
concentrations against a wide spectrum of microorganisms, including but not
limited to
Staphylococcus aureus, Pseudomonas aeruginosa, Serratia marcescens, Candida
albicans, and Ftrsarium solani.
A disinfecting solution is generally defined as a contact-lens care product
containing one or more active ingredients (for example, anti-microbial agents
and/or
preservatives) in sufficient concentrations to destroy harmful microorganisms
on the
surface of a contact lens within the recommended minimum soaking time. The
recommended minimum soaking time is included in the package instructions for
use of
the disinfecting solution. The term "disinfecting solution" does not exclude
the
possibility that the solution may also be useful as a preserving solution, or
that the
disinfecting solution may also be useful for other purposes such as daily
cleaning, rinsing
and storage of contact lenses, depending on the particular formulation. The
present
solution, in combination with its container or bottle and packaging, including
instructions
for use in accordance with a specified regimen, may be considered a novel and
improved
kit, package, or system for the care of contact lenses.
By the term "soft lens" is meant a lens having a proportion of hydrophilic
repeat
units such that the water content of the lens during use is at least 20% by
weight. The
term "soft contact lens" as used herein generally refers to those contact
lenses which
readily flex under small amounts of force. Typically, soft contact lenses are
formulated
from polymers having a certain proportion of repeat units derived from
hydroxyethyl
methacrylate andlor other hydrophilic monomers, typically crosslinked with a
crosslinking agent. However, newer soft lenses, are being made from high-Dk
silicone-
containing materials.
By the term "ophthalmically safe" with respect to a contact-lens solution is
meant
that a contact lens treated with the solution is safe for direct placement on
the eye
without rinsing, that is, the solution is safe and comfortable for daily
contact with the eye
via a contact lens that has been wetted with the solution. An ophthalmically
safe
solution has a tonicity and pH that is compatible with the eye and comprises
materials,
and amounts thereof, that are non-cytotoxic according to IS4 standards and
U.S. FDA
(Food & Drug Administration) regulations.
CA 02307664 2000-04-28
WO 99!24542 PCTJITS98/Z3818
A solution that is useful for cleaning, chemical disinfection, storing, and
rinsing a
soft contact lens is referred to herein as a "rnulti-purpose solution." Multi-
purpose
solutions do not exclude the possibility that some wearers, for example,
wearers
particularly sensitive to chemical disinfectants or other chemical agents, may
prefer to
rinse or wet a contact lens with another solution, for example, a sterile
saline solution
prior to insertion of the lens. The term "mufti-purpose solution" also does
nit exclude
the possibility of periodic cleaners not used on a daily basis or supplemental
cleaners for
removing proteins, for example enzyme cleaners, which are typically used on a
weekly
basis. By the term "cleaning" is meant that the solution contains one or more
cleaning
agents in sufficient concentrations to loosen and remove loosely held lens
deposits and
other contaminants on the surface of a contact lens, especially if used in
conjunction with
digital manipulation (for example, manual rubbing of the lens with a solution)
or with an
accessory device that agitates the solution in contact with the lens, for
example, a
mechanical cleaning aid. The critical micelle concentration of a surfactant-
containing
solution is one way to evaluate its cleaning effectiveness.
The term "effective mufti-purpose solution" analogously refers to a solution
useful for daily chemical disinfection, storing, and rinsing a contact lens,
which solution
does not claim to clean a contact lens, but which solution still obviates the
need for any
other solution for daily cleaning, that is, no other solution must necessarily
be used in
conjunction or combination with the solution on a daily basis. Although such
solutions
may comprise a surfactant or other agent that may inherently loosen or
preventing lens
deposits to some extent, such solutions are not necessarily capable of
cleaning a contact
lens. Effective mufti-purpose solutions are therefore only applicable for
lenses used for
limited period of time; either disposable or frequent replacement lenses.
Traditionally, mufti-purpose solutions on the market require a regimen
involving
mechanical rubbing of the contact lens with the mufti-purpose solution, in
order to
provide the required disinfection. That is, such a regimen is required under
governmental regulatory authorities (for example, the FDA or Food & Drug
Administration in the USA) for a Chemical Disinfection System that does not
qualify as
a Chemical Disinfecting Solution. The invention according to the present
invention has
the advantage that it is possible to formulate a product that, on the one
hand, is gentle
-g_
CA 02307664 2000-04-28
WO 99124542 PCTNS9$/23818
enough to be used as both a disinfecting solution and a wetting agent and, on
the other
hand, is able to meet the biocidal performance disinfection for a Chemical
Disinfecting
Solution under criteria established by the US FDA for Contact Lens Care
Products (May
1, 1997) that does not require a regimen involving rubbing of the lenses (even
though
rubbing of the lens may provide further removal of microorganisms). In other
words, the
compositions according to the present invention may optionally be formulate.i
to meet
the requirements of the FDA or ISO Stand-Alone Procedure for contact lens
disinfecting
products. Accordingly, it is possible to make formulations that offer higher
patient
compliance and greater universal appeal than traditional disinfecting or
disinfecting and
cleaning products.
It is noted that the combination of the biguanide polymer and the
bis(biguanide)
provides enhanced efficacy while not causing irritation or discomfort to the
eyes, always
an important and challenging concern in the art of contact-lens care. Thus,
increased
amounts of the biguanide polymer, by itself, to achieve the same efficacy as
the
combination would result in greater eye irritation. Specifically, it has been
found that
increased amounts of the biguanide polymer, by itself, to achieve the
necessary
disinfection for a Chemical Disinfecting Solution would result in unacceptable
eye
irritation. Even if a rubbing regimen is recommended when using solutions of
the
present invention, the enhanced biocidal activity may provide greater
protection against
infection, especially if the rubbing by the contact-lens wearer is inadequate
or omitted
through negligence or disregard of the product instructions.
According to the present invention, the bis(biguanide) germicides employed in
the present solutions include compounds, and their water-soluble salts, having
following
formula:
R~-NH-C-NH-C-NH-(CH2)n NH- i -NH- i -NH-R2
NH NH NH NH (I)
wherein R1 and R2 are independently selected (i.e., the same or different)
from the group
consisting of branched or unbranched alkyl having 4-12, preferably 6-10,
carbon atoms,
alkoxyalkyl (i.e., ether) or alkylsulfide (thioether or dialkylsulfide)
radical having 4-12,
preferably 6-10, carbon atoms, or cycloalkyl or cycloalkyl-alkyl radical
having 5-12,
_9_
CA 02307664 2000-04-28
WO 99/24542 PCT/US98I23818
preferably 7-10, carbon atoms; and n is 4 to 16, preferably 6 to 10. By the
term
"cycloalkyl," either in cycloalkyl or cycloalkyl-alkyl, is meant unsubstituted
or
substituted cycloalkyl, where the substituents are one or more alkyl, alkoxy (-
OR) , or
alkylthio (-SR) groups having 1-6 carbon atoms.
In the present disinfecting solution, the biguanides of Formula (I) are
suitably
used in the total amount of 0.1 to 4.0 ppm, preferably 1.0 to_3.0 ppm based on
the total
aqueous solution. More preferably, the bis(biguanides) are used in the amount
of 1.5 to
2.5, most preferably about 2.0 ppm. The concentration of the bis(biguanide) in
solution is
directly related to its bactericidal efficacy. The term "ppm" refers to "parts
per million"
and 1.0 ppm corresponds to 0.0001 percent by weight. It is based on the total
weight of
the composition or, in this case, the total weight of the aqueous disinfecting
solution.
In the present application, the amount of the bis(biguanide) or other
components
in a solution according to the present invention refers to the amount
formulated and
introduced into the solution at the time the solution is made. Over time (for
example,
over a storage period of 18 months), the assayed amount of a bis(biguanide) in
solution
may decrease somewhat.
Preferably, the bis(biguanide) compounds have the above Formula (I) wherein R~
and RZ are independently selected from the group consisting of branched or
unbranched
alkyl, alkoxyalkyl (i.e., ether) or alkylsulfde (thioether) radical, and n is
5 to 7.
Each of R' and Rz in Formulas (I) above may be, for example, an n-butyl,
isobutyl, sec-butyl, tent-butyl, pentyl, neopentyl, octyl, 2-ethylhexyl,
dodecyl, cyclobutyl,
cyclopentyl, cyclohexyl, cycloheptyl, cyclopentylmethyl or cyclohexylmethyl
radical.
Preferred are 2-ethylhexyl (alexidine), 1,5-dimethylhexyl, 1-methylhexyl, 1,3-
dimethylpentyl, 1,4-dimethylpentyl, cyclohexylmethyl, 2-norbornyl,
propyloxyoctyl, and
propyloxybutyl.
The acid-addition salts of the invention may be derived from an inorganic or
organic acid. In mast circumstances it is preferable that the salts be derived
from an acid
which is readily water soluble and which affords an anion which is suitable
for human
usage, for example a pharmaceutically-acceptable anion. Examples of such acids
are
hydrochloric, hydrobromic, phosphoric, sulphuric, acetic, D-gluconic, 2-
pyrrolidino-S-
-10-
CA 02307664 2000-04-28
wo mzasaz Pcrius9sn3sis
carboxylic, methanesulphonic, carbonic, lactic and glutamic acids. The
hydrochloride
salt is preferred.
The bis(biguanides} of Formula (I) have relatively hydrophobic end groups.
Preferably, the Log P of the compounds is 5 to 10, more preferably 6 to 8,
wherein P is
the partition coefficient of the free base, using the following equation,
wherein C is the
molar concentration of the bis(biguanide) in each phase and_ a is the degree
of ionization
of the bis(biguanide):
P - nl
Cbuffer( 1-af,)
To obtain the partition coefficient of a bis(biguanide), the compound is
partitioned
between a 0.05 M phosphate buffer (pH 11 ) saturated with octanol and octanol
saturated
with phosphate buffer after gentle shaking at room temperature (26 °C).
The volume
ratio of these two phases and the amount of sample are chosen so that the
absorbence of
the sample from the buffered layer after partitioning has a value between 0.2
and 0.9,
using a 1-cm cell and buffer solution as a blank. By working at a fixed pH and
knowing
or calculating the pKa, the P value can be determined using the above formula.
See
"Quantitative Structure-Activity Relationships for Biguanides, Carbamidates,
and
Bisbiguanides as Inhibitors of Streptococcus mutans No. 6715", Warner, V. and
Lynch,
D., J. Med. Chem, 1979, Vol. 22, no. 4 at 359, 365; and Albert, and Serjeant,
E.,
"Determination of Ionization and Stability Constants," Butler and Tanner Ltd.,
London,
England, 1962, both references hereby incorporated by reference.
Particularly preferred bis(biguanide) compounds of this invention are 2-
(decylthiomethyl)-pentane-1,5-bis(5-isopropylbiguanide), 2-(decylthio-
methyl)pentane-
1,5-bis(5,5-diethylbiguanide), and hexane-1,6-bis(2-ethylhexylbiguanide), the
latter also
known as alexidine or l,1'-hexamethyienebis(5-(2-ethylhexyl)-biguanide)
dihydrochloride. Other preferred bis(biguanides) include 1,1'-
hexamethylenebis(5-
heptyl-biguanide) dihydrochloride, 1,1'-hexamethylenebis(5-octyl-biguanide)
dihydrochloride, and 1,1'-hexamethylenebis(5-hexyl-biguanide) dihydrochloride.
The biguanide compounds of Formula {I) may be made by reacting a bis-
cyanoguanidine of the formula:
-11-
CA 02307664 2000-04-28
WO 99/?,4542 PGT/US98IZ3818
NC-NH-C-NH-A-NH-C-NH-CN
(I II
NH NH
(VI)
with an amine R'NH2, or with two different amines R'NH2 and R~NH2, in the form
of an
acid addition salt thereof, wherein R' and R2 have the meanings stated above,
at a
temperature of 100°C to 170°C and A is an alkylene group having
the required number
of carbon atoms. A preferred amine salt is the hydrochloride. Most diamines
are
commercially available from a variety of sources.
The reactants are heated together until the reaction is complete. The reaction
proceeds fastest at higher temperatures, but if thermal stability is a
problem, the reaction
should be carried out at lower temperature for a longer period. The reactants
are most
conveniently melted together in the absence of a solvent, but if desired an
inert solvent
such as DMSO, 2-methoxyethanol, 2-ethoxyethanol, nitrobenzene, sulpholane,
isopropanol, n-butanol, ethylene glycol dimethyl ether or water, or a mixture
of such
solvents, may be used.
The bis-cyanoguanidine of the Formula (VI) may be manufactured from known
starting materials such as hexamethylenedinitrile which is reduced, for
example, with
hydrogen and Raney nickel or with borane in dimethyl sulphide to the
corresponding
diamine (VIII), and the diamine in the form of an acid-addition salt,
conveniently the
dihydrochloride, is reacted with sodium dicyanamide or other suitable salt to
form the
required starting material (VI), as depicted below.
NC-A-CN -~~ NHS A-NH,,
(VII) (VIII)
VIII -~ 2-MN(CN).,--ANC-NH- j -NH-A-NH-C-NH-CN
NH NH
-12-
CA 02307664 2000-04-28
WO 99124542 PCT/US98I23818
wherein M is a sodium, potassium, zinc or other suitable salt. The sodium salt
is
commercially available.
The compounds of the present invention can also be made by reacting a diamine
of the Formula (VIII) in the form of an acid addition salt, with a
cyanoguanidine of the
formula:
R~HN-C-NH-CN
II
NH
(
or with a cyanoguanidine of the Formula (IX) and a cyanoguanidine of the
formula:
R2HN-C-NH-CN
II
NH
(X)
wherein R' and R2 have the meanings stated above, at a temperature of
100° to 170°C.
A suitable salt of the diamine is, for example, the dihydrochloride. The
reactants
are heated together until the reaction is complete. The reaction proceeds
fastest at higher
temperature, but if thermal stability is a problem, the reaction should be
carried out at
lower temperature over a longer period. If a melt can be formed at those
temperatures
the reactants are conveniently melted together in the absence of a solvent. If
not, or
alternatively, the reactants are heated together in a suitable inert solvent,
for example
those mentioned above. The acid-addition salts of the invention are obtained
by
conventional means.
The cyanoguanidines of the Formulae (IX) and (X), which may be used as
starting materials in the above process, may be obtained by reacting sodium
dicyanamide
with an appropriate amine R~NHz or RZNH2, in the form of an acid-addition
salt,
conveniently the dihydrochioride, in a suitable inert solvent.
-13-
CA 02307664 2000-04-28
_ WO 99124542 PCT/US98123818
For example, the bis(biguanide) known as alexidine is produced from the
following sequence of reactions.
a
. 2 Cl
Na~
~ ~ N
H3N-(CH2)6-NH3 + 2 NC CN
(~) (~)
NC-NH- i -NH-(CH2)~-NH- i -NH-CN
NH NH
(XIII)
C~
XIII -- 2 H3N~
(XIV)
NH- i-NH- i-NH-(CH2)6-NH-i -NH-i -NH ~ 2 HC1
NH NH NH NH
(XV)
Compound (XI) is hexamethylenediamine dihydrochloride (MW 189), Compound (XII)
is sodium dicyanamide, Compound XIII is HMBDA, hexamethylene
bis(cyanoguanido),
Compound (XIV) is 2-ethyl-hexylamine hydrochloride (MW 165.?), and Compound
(XV) is alexidine dihydrochloride a.k.a. [1,6-bis-(2-
ethylhexylbiguanido)hexane
dihydrochloride a.k.a. hexane-I,6-bis(2-ethylhexyl biguanide) dihydrochloride.
This
compound has a molecular weight in g/mole (MW) of 581.7 and empirical formula
Cz6Hs6Nio-2HC1. The Compound (XV) is commercially available from various
sources,
including Sigma Chemical Co. (St. Louis, Missouri).
-14-
CA 02307664 2000-04-28
WO 99124542 PCT/US98123818
The methods for synthesized compounds of the present invention are also
disclosed in European Patent Application Publication No. 0 I25 092 (published
14.1 I.84); Rose, F.L. and Swain, G., "Bisdiguanide Having Antibacterial
Activity," J.
Chem. Soc., p. 4422-4425 (1956); and Wamer, Victor D. and Lynch, Donald,
"Quantitative Structure-Activity Relationships of Biguanide, Carbamimidates,
and
Bisdiguanides as Inhibitors of Streptococcus Mutans No. 67I 5, " J. Med.
Chem., Vol. 22,
No. 6, p. 359-366 (1979).
The bis(biguanides) of the present invention (Formula I) may be used in
combination with one or more polymeric biguanides, and water-soluble salts
thereof,
having the following formula:
X~~Z-NH-C-NH- i-NH~-Z-X2
NH NH
(IV)
wherein Z is an organic divalent bridging group which may be the same or
different
throughout the polymer, n is on average at least 3, preferably on average 5 to
20, and X'
and Xz are independently selected from the groups -NH2 and -NH - i - NH - CN.
NH
One preferred group of water-soluble polymeric biguanides will have number
average
molecular weights of at least 1,000 and more preferably will have number
average
molecular weights from 1,000 to 50,000. Suitable water-soluble salts of the
free bases
include, but are not limited to hydrochloride, borate. acetate, gluconate,
sulfonate,
:artrate and citrate salts.
The above-disclosed biguanides and methods of preparation are described in the
literature. For example, U.S. patent 3,428,576 describes the preparation of
polymeric
biguanides from a diamine and salts thereof and a diamine salt of dicyanimide.
The polymeric biguanides, in combination with the bisbiguanides of the present
invention, are effective in concentrations as low as 0.00001 weight percent
(0.1 ppm). It
has also been found that the bactericidal activity of the solutions may be
enhanced or the
spectrum of activity broadened through the use of a combination of such
polymeric
biguanides with the compounds of Formula (I) above. The effective amount of
the
-15-
CA 02307664 2000-04-28
WO 99/24542 PCT/US98123818
polymeric biguanides {irrespective of the particular salt form or whether the
free base is
used ) may in total be as low as about 0.000010 weight percent (0.10 ppm) and
up to
about 0.00030 weight percent {3.0 ppm) in the present invention, whether in
the form of
a water-soluble salt or the free base. Preferably, the total amount of
polymeric
biguanide, in combination with the total amount of compounds of Formula (I)
above is
about 0.3 to 2.0 ppm, more preferably about 0.4 to 1.0, most preferably aboui
0.5 to 0.8
ppm.
Most preferred are the polymeric hexamethylene biguanides, commercially
available, for example, as the hydrochloride salt from Zeneca (Wilmington, DE)
under
the trademark CosmocilTM CQ. Such polymers and water-soluble salts are
referred to as
polyhexamethylene (PHMB) or polyaminopropyl biguanide (PAPB). The term PHMB
or PAPB, as used herein, is meant to encompass one or more biguanides have the
following formula:
X~-(CH~)3 {CH2)3-NH- i-NH-i -NH-(CH~)3 {CH2)3-X2
NH NH n
(V)
wherein X' and X2 are as defined above and n is from 1 to 500.
Depending on the manner in which the biguanides are prepared, the predominant
compound falling within the above formula may have different X' and XZ groups
or the
same groups, with lesser amounts of other compounds within the formula. Such
compounds are known and are disclosed in US Patent No. 4,758,595 and British
Patent
1,432,345, which patents are hereby incorporated herein by reference.
Preferably, the
water-soluble salts are compounds where n has an average value of 2 to 15,
most
preferably 3 to 12.
Optional additional disinfectant/germicide components may be employed in the
present invention to further potentiate, compliment or broaden the spectrum of
microbiocidal activity of the invention. This includes microbiocidally
effective amounts
of germicides which are compatible with and do not precipitate in the
solution, in
concentrations ranging from about 0.000001 to about 0.5 weight percent,
depending on
the particular disinfecting agent as will be appreciated by the skilled
artisan. Suitable
complementary germicidal agents include, but are not limited to thimerosol,
sorbic acid,
-16-
CA 02307664 2000-04-28
WO 99lZ4542 PCTIUS98/23818
alkyl triethanolamines, phenylmercuric salts, quaternary ammonium compounds,
and
polyquaternium copolymers, and mixtures thereof. Suitable salts are soluble in
water at
ambient temperature to the extent of at least 0.5 weight percent. These salts
include the
gluconate, isethionate, (2-hydroxyethanesulfonate), fonmate, acetate,
glutamate,
succinanate, monodiglycollate, methanesulfonate, lactate, isobutyrate and
glucoheptonate. Representative examples of the quaternary ammonium corr~pounds
are
compositions comprised of balanced mixtures of n-alkyl dimethyl benzyl
ammonium
chlorides. An example of a polyquaternium polymer used in ophthalmic
applications
include Polyquaternium 1~ (chemical registry number 75345-27-6) available from
Onyx
corporation.
The present solution optionally comprises at least one surfactant. Suitable
surfactants can be either amphoteric, cationic, anionic, or nonionic which may
be present
(individually or in combination) in amounts up to 15 percent, preferably up to
5 percent
by weight of the composition or solution. Preferred surfactants are amphoteric
or
nonionic surfactants, which when used impart cleaning and conditioning
properties. The
surfactant should be soluble in the lens care solution and non-irritating to
eye tissues.
Many nonionic surfactants comprise one or more chains or polymeric components
having oxyalkylene (-O-R-) repeats units wherein R has 2 to 6 carbon atoms.
Preferred
non-ionic surfactants comprise block polymers of two or more different kinds
of
oxyalkylene repeat units, which ratio of different repeat units determines the
HLB of the
surfactant. Satisfactory non-ionic surfactants include polyethylene glycol
esters of fatty
acids, e.g. coconut, polysorbate, polyoxyethylene or polyoxypropylene ethers
of higher
alkanes (C12-Clg). Examples of the preferred class include polysorbate 20
(available
under the trademark Tween~ 20), polyoxyethylene (23) lauryl ether (Brij~ 35),
polyoxyethyene (40) stearate (Myrj~ 52), polyoxyethylene (25) propylene glycol
stearate (Atlas~ G 26I2). One non-ionic surfactant in particular consisting of
a
poly(oxypropylene)-poly(oxyethylene) adduct of ethylene diamine having a
molecular
weight from about 7,500 to about 27,000 wherein at least 40 weight percent of
said
adduct is poly(oxyethylene) has been found to be particularly advantageous for
use in
cleaning and conditioning both soft and hard contact lenses when used in
amounts from
-17-
CA 02307664 2000-04-28
WO 99124542 PCTIUS98I23818
about 0.01 to about 15 weight percent. The CTFA Cosmetic Ingredient
Dictionary's
adopted name for this group of surfactants is poloxamine. Such surfactants are
available
from BASF Wyandotte Corp., Wyandotte, Michigan, under the registered trademark
"Tetronic". An analogous of series of surfactants, suitable for use in the
present
invention, is the poloxamer series which is a poly(oxyethylene)
poly(oxypropylene)
block polymers available under the trademark "Pluronic" (commercially.
available form
BASF).
Various other ionic as well as amphoteric and anionic surfactants suitable for
in
the invention can be readily ascertained, in view of the foregoing
description, from
McCutcheon's Detergents and Emulsifiers, North American Edition, McCutcheon
Division, MC Publishing Co., Glen Rock, NJ 07452 and the CTFA International
Cosmetic Ineredient Handbook, Published by The Cosmetic, Toiletry, and
Fragrance
Association, Washington, D.C.
Amphoteric surfactants suitable for use in a composition according to the
present
invention include materials of the type are offered commercially under the
trade name
"Miranol." Another useful class of amphoteric surfactants is exemplified by
cocoamidopropyl betaine, commercially available from various sources.
The foregoing surfactants when employed with a buffer enhancer will generally
be present in an amount from 0.01 to 5.0 percent (w/w), preferably 0.1 to 5.0
percent
Typically, the aqueous solutions of the present invention for treating contact
lenses are also adjusted with tonicity agents, to approximate the osmotic
pressure of
normal lacrimal fluids which is equivalent to a 0.9 percent solution of sodium
chloride or
2.5 percent of glycerol solution. The solutions are made substantially
isotonic with
physiological saline used alone or in combination, otherwise if simply blended
with
sterile water and made hypotonic or made hypertonic the lenses will lose their
desirable
optical parameters. Correspondingly, excess saline may result in the formation
of a
hypertonic solution which will cause stinging and eye irritation.
The pH of the present solutions should be maintained within the range of 5.0
to
8.0, more preferably about 6.0 to 8.0, most preferably about 6.5 to 7.8,
suitable buffers
may be added, such as boric acid, sodium borate, potassium citrate, citric
acid, sodium
bicarbonate, TRIS, and various mixed phosphate buffers (including combinations
of
-18-
CA 02307664 2000-04-28
WO 99/24542 PCTIUS98/23818
Na2HP04, NaHZP04 and KH2P04) and mixtures thereof. Borate buffers are
preferred,
particularly for enhancing the efficacy of biguanides. Generally, buffers will
be used in
amounts ranging from about 0.05 to 2.5 percent by weight, and preferably, from
0.1 to
1.5 percent. The disinfecting/preserving solutions of this invention
preferably contain a
borate buffer system, containing one or mare of boric acid, sodium borate,
potassium
tetraborate, potassium metaborate, or mixtures of the same. _ .
In addition to buffering agents, in some instances it may be desirable to
include
sequestering agents in the present solutions in order to bind metal ions which
might
otherwise react with the lens and/or protein deposits and collect on the lens.
Ethylene-
diaminetetraacetic acid (EDTA) and its salts (disodium) are preferred
examples. They
are usually added in amounts ranging from about 0.01 to about 0.2 weight
percent.
Other suitable sequestering agents include gluconic acid, citric acid,
tartaric acid and
their salts, e.g. sodium salts. Preferred sequestering agents, which are also
effective for
removing protein deposits, are the phosphonate compounds disclosed in WO
97/31659.
The aqueous solutions of the present invention are especially useful for soft
contact lenses, with or without further additives. Nevertheless, the solutions
of the
present invention may be formulated into specific contact lens care products,
such as
wetting solutions, soaking solutions, cleaning and conditioning solutions, as
well as
mufti-purpose type lens care solutions, etc. and mixtures thereof. Finally,
such solutions
can be applied to the lenses outside the eye or while on the eye, for example,
in the form
of droplets.
It may also be desirable to include water-soluble viscosity builders in the
solutions of the present invention. Because of their demulcent effect,
viscosity builders
have a tendency to enhance the lens wearer's comfort by means of a film on the
lens
surface cushioning impact against the eye. Included among the water-soluble
viscosity
builders are the cellulose polymers like hydroxyethyl or hydroxypropyl
cellulose,
carboxymethyl cellulose, povidone, polyvinyl alcohol, and the like. Such
viscosity
builders may be employed in amounts ranging from about 0.01 to about 4.0
weight
percent or less. The present solutions may also include optional demulcents.
-19-
CA 02307664 2000-04-28
WO 99124542 PCT/US98123818
The aqueous solutions according to the present invention can be effectively
used
in disinfecting contact lenses by any of the well recognized methods. The
lenses may be
treated by the "cold" soaking method at room temperature for a period ranging
from
about 5 minutes to about 12 hours. The lenses are then removed from the
solution,
rinsed with the same or a different solution, for example a preserved isotonic
saline
solution and then replaced on the eye.
As indicated above, contact-lens wearers are commonly required to digitally or
manually rub the contact lenses (typically between a finger and palm or
between fingers)
during daily cleaning andlor disinfecting of contact lenses. In one embodiment
of the
present invention, a method is provided in which rubbing is not required
during
treatment with the claimed specified solution, between removal from the eye
and
replacement of the lens following lens care. In a preferred embodiment of such
a
method, a soft lens is disinfected or both disinfected and cleaned with a
multipurpose
solution or an effective multipurpose solution that is the only daily solution
needed for
treating the lens outside the eye. Thus, in one embodiment of a method
according to the
invention, the described solution is used to treat a contact lens without
rubbing, by a
method comprising:
(a) soaking the contact lens that has not been rubbed with the solution
for a specified time period, and
(b) direct placement of the treated contact lens on the eye of the
wearer.
Typically, step (a) may involve immersing the contact lens in the solution.
Soaking may optionally comprise shaking or similarly agitating a container of
the
solution by manual means. Preferably, step (a) involves a period of soaking
the contact
lens in a container wherein the contact lens is completely immersed in the
solution. By
the term "direct placement" is herein meant that the solution is not diluted
or rinsed off
the lens with a different contact-lens solution prior to "insertion" or
placement on the
eye. In a particularly preferred embodiment, the method uses a product that is
formulated as a mufti-purpose or effective mufti-purpose solution, wherein no
other
solution or product is required for daily cleaning of the lens, with the
possible exception
of an enzyme cleaner.
-20-
CA 02307664 2000-04-28
WO 99124542 PCT/US98/23818
In yet another embodiment of a method according to the present invention, the
claimed solution is used to clean a frequent replacement lens (FRL) that is
planned for
replacement after not more than about three months of use in the eye, or that
is planned
for replacement after not more than about 30 days of use in the eye, or that
is planned for
replacement after not more than about two weeks in the eye. Preferably, the
lens is made
from a polymer comprising about 0.0 to 5 mole percent repeat units derived
from
methacrylic acid (MAA), 10 to 99 mole percent of repeat units derived from
hydroxyethyl methacrylate, and about 0.5 to 5 mole percent of cross-linking
repeat units.
Cross-linking repeat units may be derived, for example, from such monomers as
ethyleneglycol dimethacrylate, divinylbenzene, and trimethylpropane
trimethacryiate.
The following Examples illustrate the compositions and methods of the instant
invention.
EXAMPLE 1
This Example illustrates the preparation of 1,6 bis(cyanoguanidino) hexane,
used
as a starting material for bis(biguanides) of the present invention. In the
amount of 35.80
g (0.402 mole), sodium dicyanamide (NaC2N3) is suspended in 400 mL of 1-
butanol.
Then, 23.60 g (0.204 mole) of 1,6-hexanediamine were added as well as 33.0 mL
of
cone. aqueous hydrochloric acid (0.400 mole). A milky white precipitate
appeared
immediately which was probably the amine hydrochloride. The mixture was then
refluxed for 3.5 hr. The suspension was then cooled to room temperature and
filtered.
The white solid was then washed well with distilled water before drying under
vacuum.
Yield 46.38 g; 93.1%. C 10 H 18 N 8 calc'd: C 48.0%; H 7.20%; N 44.80%; found:
C
47.7%; H 7.40%; N 45.12%. 300 MHz ~H NMR {d~-DMSO) 6.60 ppm (6p, br m); 2.93
ppm (4p, m); 1.34 ppm (4p, br s); 1.15 ppm (4p, br s). IR (KBr pellet, cm')
3142 (m);
2943; 2912; 2862 (w); 2179 (s); 1658; 1609 {s).
EXAMPLE 2
This Example illustrates the preparation of the bis(biguanide) known as
alexidine
for use in the present invention. Hexamethylene bis{cyanoguanido) in the
amount of
1.003g (0.004498 moles) was placed into a flask. To this was added 1.474 mL
(1.163g;
-21-
CA 02307664 2000-04-28
wo ~nasaz rc~rms9sn3sis
0.008996 moles) of 2-ethylhexylamine. Then, 0.74 mL (0.008996 moles) of
concentrated HCl was added. The mixture was heated in a flask to boil away the
H20.
After the H20 was gone, the temperature of the melt had risen to
195°C. The
temperature was decreased to 150-160°C and maintained for 1 hour. The
material was
cooled to room temperature. The solid can be dissolved in hot water and
allowed to
crystallize. - . -
EXAMPLE 3
This Example illustrates the preparation of poly(hexamethylene biguanide},
also
referred to as PAPB or PHMB, for use in combination with bis(biguanides) in
the present
invention. In 500 mL of distilled water was suspended 25.08 g (0.100 mole) of
1,6-
bis(cyanoguanidino)hexane and 18.99 g (O.I00 mole) of 1,6-hexanediamine
dihydrochloride. The pH of this mixture was then brought down to 6.8 with
dilute
hydrochloric acid. The water was then removed by distillation under reduced
pressure.
The white solid was then transferred to a three-necked flask fitted with a
mechanical
stirrer and heating mantle. The intimate mixture of solids was then placed
under
nitrogen and the temperature of the mixture was raised to 150-55°C. The
molten
reaction mixture possessed the consistency of honey. The mixture was stirred
at 150-
55°C for 1 - 1.5 hr. before cooling to room temperature. The resulting
poly{hexamethylene biguanide) is obtained as a glassy solid. The yield is
essentially
quantitative. Melting range 105-125°C. 300 MHz ~H 1~TMR (Dz0) 3.13 ppm
(2l.lp, br
t); 2.93 ppm (Zp, t); 1.49 ppm (2l.lp, br s); 1.28 ppm (2l.lp, br s). IR {KBr
pellet, cm-1)
3325; 3201 (s); 2931; 2858 (m); 2175 (m-w); 1631; 1589; 1550(s).
EXAMPLE 4
This example illustrates the preparation of an aqueous disinfecting solution
according to the present invention comprising a combination of alexidine and
polyhexamtheylene biguanide (also referred to as PHMB). The following
components
are used, in the indicated percent weight per total volume of the solution:
-22-
CA 02307664 2000-04-28
. WO 99/24542 PCT/US98I23818
Percent (w/v)
PHMB 0.00008
Alexidine2HC1 0.0002
Poloxamine 1107** 1.0
Na2EDTA 0.11
Boric Acid 0.66
Sodium Borate 0.10
Sodium Chloride 0.54
Distilled Water (qs) 100.0
** molecular weight 14,500, Tetronic~ 1107, a poly(oxypropylene)
poly(oxyethylene) block copolymer
adduct of ethylene diamine, a trademark of BASF Wyandotte Corp., Wyandotte,
MI.
The solution is prepared by gradually heating 80 percent of the water to
80°C
while dissolving the disodium EDTA therein. The boric acid and sodium borate
are
added to the heated solution of disodium EDTA and dissolved. The sodium
chloride is
then added to the solution and dissolved, followed by the addition of the
surfactant. The
solution is sterilized by autoclaving to 120°C for 45 minute. After the
solution is cooled
to room temperature, the alexidine bis(biguanide) and the PHMB are added as a
solution
through a sterile filter, followed by the balance of distilled water. The
solution is
packaged in sterilized plastic containers.
EXAMPLE 5
This Example illustrates the improved antimicrobial efficacy of the
combination
of alexidine with polyhexamethylene biguanide (PHMB) in an aqueous
disinfecting
solution for contact lenses. The antimicrobial efficacy of each of various
compositions
for the chemical disinfection of contact lenses was evaluated.
Microbial challenge inoculums were prepared using Pseacdomonas aeruginosa
(ATCC 9027), Staphylococcus aureus (ATCC 6538), Serratia marcescens (ATCC
13880), Candida albicans (ATCC 10231), and Fusarium solani (ATCC 36031). The
test
organisms were cultured on appropriate agar and the cultures were harvested
using sterile
DPBST (Dulbecco's Phosphate Buffered Saline plus 0.05% wlv polysorbate 80) or
a
suitable diluent and transferred to a suitable vessel. Spore suspensions were
filtered
-23-
CA 02307664 2000-04-28
WO 99124542 PCT/US98l23818
through sterile glass wool to remove hyphal fragments. Serratia murcesceris,
as
appropriate, was filtered (eg., through a 1.2p, filter) to clarify the
suspension. After
harvesting, the suspension was centrifuged at no more than 5000 x g for a
maximum of
30 minutes at 20-25°C. The supernatant was poured off and resuspended
in DPBST or
other suitable diluent. The suspension was centrifuged a second time, and
resuspended
in DPBST or other suitable diluent. All challenge bacterial and fungal.cell
suspensions
were adjusted with DPBST or other suitable diluent to 1 x 10~-10g cfu/mL. The
appropriate cell concentration may be estimated by measuring the turbidity of
the
suspension, for example using a spectrophotometer at a preselected wavelength,
for
example 490 nm. One tube was prepared containing a minimum of 10 mL of test
solution per challenge organism. Each tube of the solution to be tested was
inoculated
with a suspension of the test organism sufficient to provide a final count of
1.0 x 105-10G
cfulmL, the volume of the inocuium not exceeding 1 % of the sample volume.
Dispersion of the inoculum was ensured by vortexing the sample for at least 15
seconds.
The inoculated product was stored at 10-25°C. Aliquots in the amount of
1.0 mL were
taken of the inoculated product for determination of viable counts after
certain time
periods of disinfection. The time points for the bacteria were, for example,
1, 2, 3, and 4
hours when the proposed regimen soaking time was 4 hours. Yeast and mold were
tested
at an additional time point of >_ 16 hours (4 times the regimen time). The
suspension was
mixed well by vortexing vigorously for at least S second. The 1.0 mL aliquots
removed
at the specified time intervals were subjected to a suitable series of decimal
dilutions in
validated neutralizing media. The suspensions were mixed vigoroaslv and
incubated for
a suitable period of time to allow for neutralization of the microbial agent.
The viable
count of organisms was determined in appropriate dilutions by preparation of
triplicate
plates of trypticase soy (TSA) agar for bacteria and Sabouraud dextrose agar
(SDA) for
mold and yeast. The bacterial recovery plates were incubated at 30-35°C
for 2-4 days.
The yeast was incubated at 20-30°C for 2-4 days and mold recovery
plates at 20-25°C
for 3-7 days. The average number of colony forming units was determined on
countable
plates. Countable plates refer to 30-300 cfu/plates for bacteria and yeast,
and 8 to 80
cfu/plate for mold except when colonies are observed only for the 10°
or 10-' dilution
plates. The microbial reduction was then calculated at the specified time
points. In order
-24-
CA 02307664 2000-04-28
WO 99124542 PCTIUS98/23818
to demonstrate the suitability of the medium used for growth of the test
organisms and to
provide an estimation of the initial inoculum concentration, inoculum controls
were
made by dispersing an identical aliquot of the inoculum into a suitable
diluent, for
example DPBST, using the same volume of diluent used to suspend the organism
as
listed above. Following inoculation in a validated neutralizing broth and
incubation for
an appropriate period of time, the inoculum control must be between 1.0_x ltis
- 1.0 x 106
cfu/mL
The solutions were evaluated based on the performance requirement referred to
as the "Stand-Alone Procedure for Disinfecting Products" (hereafter the "stand-
alone
test") and is based on the Disinfection Efficacy Testing for contact lens care
products
under the Premarket Notification (510(k)) Guidance Document For Contact Lens
Care
Products dated May l, 1997, prepared by the U.S. Food and Drug Administration,
Division of Ophthalmic Devices. This performance requirement is comparable to
current ISO standards for disinfection of contact lenses (revised 1995). The
stand-alone
test challenges a disinfecting product with a standard inoculum of a
representative range
of microorganisms and establishes the extent of viability loss at pre-
determined time
intervals comparable with those during which the product may be used. The
primary
criteria for a given disinfection period (corresponding to a potential minimum
recommended disinfection period) is that the number of bacteria recovered per
mL must
be reduced by a mean value of not less than 3.0 logs within the given
disinfection period.
The number of mold and yeast recovered per mL must be reduced by a mean value
of not
less than 1.0 log within the minimum recommended disinfection time with no
increase at
four times the minimum recommended disinfection time.
The testing procedures described above were followed to determine whether the
primary performance criteria would be passed at time intervals of 5 minutes,
15 minutes,
30 minutes, and 4 hours. The concentration of alexidine in this set of tests
was
formulated in amounts ranging from 0.0 to 4.0 ppm in combination with either
0.0 or 0.8
ppm PAPB, the latter the amount of PAPB currently used in commercial multi-
purpose
solutions for soft contact lenses. The results are shown in Table 1 below.
-25-
CA 02307664 2000-04-28
WO 99124542 PCTNS98l23818
TABLE 1
Alexidiae
2HC1 PAPB Log ReductionLog Reduction
{ppm) (ppm) Soak Period S. marcescensC. albicans
5 min - 0.6
0.5 0.8 15 min 2.0 1.1
30 min 3:2 - 1.8
60 min 4.3 3.5
5 min - 0.5
1.0 0.8 15 min 2.6 1.1
30 min 3.7 2.2
60 min >4.3 4.2
5 min - 0.7
2.6 0.8 15 min 2.7 2.2
30 rnin >4.3 3.3
60 min >4,3 >4.2
5 min - 1.1
4.0 0.8 15 min >4.3 3.0
30 min >4.3 >4.2
60 min >4.3 >4.2
5 min - 0.3
0.5 0.0 15 min 0.6 1.0
30 min 1.6 -0.2
60 min 2.5 -0.1
5 min - 0.4
2.6 0.0 15 min 2.2 0.5
30 min >4.3 0.5
60 min >4,3 1.4
15 min - 0.6
0.0 0.8 30 min 1.3 0.8
45 min 2.0 1.4
60 min 3.1 2.9
The results show that the addition of alexidine to the polyhexametheylene
biguanide improved antimicrobial efficacy, with the improved efficacy reaching
a trade-
off, for practical purposes, at about 4.0 ppm, such that any improved
antimicrobial
efficacy would be unlikely to be justified by the increased potential for
toxicity at higher
concentrations of the antimicrobial agent. With respect to G albicans, there
appears to
be a synergistic effect at time periods of 15 minutes and 30 minutes, with the
combination of 2.6 ppm alexidine and 0.8 ppm PA.PB showing greater efficacy
than the
sum of 2.6 ppm alexidine by itself and 0.8 ppm PAPB by itself.
-26-
CA 02307664 2000-04-28
_ WO 99124542 PCTIITS98/23818
EXAMPLE 6
This Example illustrates the microbiocidal efficacy of solutions according to
the
present invention. The above testing procedures were used for evaluating the
antimicrobial efficacy against C. albicans of disinfecting solutions such as
prepared in
Example 4, but which contain the bis(biguanide) alexidine at various
concemrations
extending from 1 ppm to 4 ppm and the biguanide polymer at various
concentrations
extending from 0.3 to 1.5 ppm. The results are shown in Table 2 after a 5
minute soak,
Table 3 after a 15 minute soak, Table 4 after a 30 minute soak, and Table 5
after a 45
minute soak. Tables 6 to 9, corresponding respectively to Tables 2 to 5,
compares the
theoretical kill, based on the sum of individual disinfecting agents versus
actual kill
using the combination of disinfecting agents. For this calculation, the
following equation
was employed: log 10(exp 10(PHMB) + exp 10(alexidine)). In other words, to
calculate
the theoretical log kill, the log reduction kill values for each separate
disinfecting agent
were converted to the numbers of organisms killed, and then these values were
added
and a new log value derived for the sum.
TABLE 2
(5 Minutes)
PHMB
0 ppm 0.3 ppm 0.8 ppm 1.5 ppm
I
0 ppm 0.4 0.4 0.7 1.0
1 ppm 0.5 0.6 0.9 1.1
Alexidine 2 ppm 0.7 0.8 0.9 1.5
3 ppm 0.8 1.1 1.6 2.6
4 ppm 1.1 1.8 I 2.8 3.4
-27-
CA 02307664 2000-04-28
WO 99I24S42 PG"T/ITS98/23818
TABLE 3
(15 Minutes)
PHMB
0 ppm 0.3 ppm 0.8 ppm 1.5 ppm
0 ppm 0.4 0.7 1.5 I 1.9
1 ppm 0.4 1.1 2.2 2.9
Alexidine 2 ppm 0.8 1.7 2.8 3.5
3 ppm 1.6 2.7 3.7 4.5
4 ppm 2.6 3.8 4.8 >4.8
TABLE 4
(30 Minutes)
PHMB
0 ppm 0.3 ppm 0.8 ppm 1.5 ppm
0 ppm 0.4 I .0 2.1 3.1
1 ppm 0.5 1.8 3.3 4.4
Alexidine 2 ppm 1.2 2.1 3.7 4.5
3 ppm I .9 3.5 4.8 >4.8
4 ppm 3.4 >4.8 I >4.8 >4.8
-28-
CA 02307664 2000-04-28
WO 99124542 PCT/US98123818
TABLE 5
(45 Minutes)
PHMB
0 ppm 0.3 ppm 0.8 ppm 1.5 ppm
0 ppm 0.4 1.1 3.0 I 4.1
1 ppm 0.4 2.1 3.7 >4.8
Alexidine 2 ppm 0.9 2.6 4.5 >4.8
3 ppm 2.1 4. S >4. 8 >4. 8
4 ppm 3.7 4.8 >4.8 >4.8
TABLE b
(5 Minutes)
Log Kill
Formulation Theoretical Actual
1 ppm Alex/0.3 ppm PHMB 0.8 0.6
2 ppm Alex/0.3 ppm PHMB 0.9 0.8
3 ppm Alex/0.3 ppm PHMB 0.9 1. I
4 ppm Alex/0.3 ppm PHMB 1.2 1.8
1 ppm Alex/0.8 ppm PHMB 0.9 0.9
2 ppm Alex/0.8 ppm PHMB I .0 0.9
3 ppm Alex/0.8 ppm PHMB I .I 1.6
4 ppm Alex/0.8 ppm PHMB 1.2 2.8
1 ppm Alex/1.5 ppm PHMB 1.1 1.1
2 ppm Alex11.5 ppm PHMB 1.2 1.5
3 ppm AlexIl.S ppm PHMB 1.2 2.6
4 ppm Alex/1.5 ppm PHMB 1.4 3.4
-29-
CA 02307664 2000-04-28
WO 99/24542 PCTIUS98I23818
TABLE 7
(15 Minutes)
Log Kill
Formulation Theoretical Actual
I ppm Alex/0.3 ppm PHMB 0.9 -_. ~.i -
2. ppm Alex/0.3 ppm PHMB 1.1 _ 1.7
3 ppm Alex/0.3 ppm PHMB I .7 2.7
4 ppm Alex/0.3 ppm PHMB 2.6 3.8
I ppm Alex/0.8 ppm PHMB 1.5 2.2
2 ppm Alex/0.8 ppm PHMB 1.6 2.8
3 ppm Alex/0.8 ppm PHMB 1.9 3.7
4 ppm Alex/0.8 ppm PHMB 2.6 4.8
1 ppm Alex/1.5 ppm PHMB 1.9 2.9
2 ppm Aiexll .5 ppm PHMB 1.9 3.5
3 ppm Alexll .5 ppm PHMB 2.1 4.5
4 ppm Alex/1.5 ppm PHMB 2.7 4.8
-3 0-
CA 02307664 2000-04-28
WO 99124542 PCTIUS98/23818
TABLE 8
(30 Minutes)
Log Kill
Formulation Theoretical Actual
1 ppm Alexl0.3 ppm PHMB 1.1 1.8
2 ppm AlexI0.3 ppm PHMB 1.4 _ 2.1
3 ppm Alex/0.3 ppm PHMB 2.0 3.5
4 ppm Alex/0.3 ppm PHMB 3.4 4.8
1 ppm AlexI0.8 ppm PHMB 2.1 3.3
2 ppm Alex/0.8 ppm PHMB 2.2 3.7
3 ppm Alex/0.8 ppm PHMB 2.3 4.8
4 ppm Alex/0.8 ppm PHMB 3.4 4.8
1 ppm Alex/1.5 ppm PHMB 3.1 4.4
2 ppm Alex/1.5 ppm PHMB 3.1 4.5
3 ppm Alex/1.5 ppm PHMB 3.1 4.8
4 ppm Alex/1.5 ppm PHMB 3.6 4.8
-31-
CA 02307664 2000-04-28
WO 99124542 PCTIUS98/23818
TABLE 9
(4S Minutes)
Log Kill
Formulation Theoretical Actual
1 ppm Alex/0.3 ppm PHMB 1.2 2.1
_ -
-
2 ppm Alex/0.3 ppm PHMB 1.3 _ - '.6
3 ppm Alex10.3 ppm PHMB 2.1 4.5
4 ppm Alex/0.3 ppm PHMB 3.7 4.8
1 ppm Alex/0.8 ppm PHMB 3.0 3.7
2 ppm Alex/0.8 ppm PHMB 3.0 4.5
3 ppm Alex/0.8 ppm PHMB 3.1 4.8
4 ppm Alex/0.8 ppm PHMB 38 4.8
1 ppm Alex/1.5 ppm PHMB 4.1 4.8
2 ppm Alex/1.5 ppm PHMB 4.1 4.8
3 ppm Alexll .5 ppm PHMB 4.1 4.8
4 ppm Alex/1.5 ppm PHMB 4.2 4.8
The above results show synergistic microbicidal effects against C. albicans,
in
which the log kill from the combination of both alexidine and PHMB, in a good
proportion of cases, is higher than the sum of the individual disinfecting
agents, which
synergistic effects are evident, beginning with the results after 1 S minutes.
At 45
minutes, these synergistic effects may become less evident, when a higher
proportion of
the microorganisms have already been killed. Based on the above data, Figure 1
shows
the biocidal efficacy against C. albicans of the test solutions containing the
combination
of alexidine and PHMB, after 15 minutes, compared to the theoretical efficacy,
based on
the sum of individual solutions containing, respectively, alexidine alone and
PHMB
alone. Figure 2 shows the biocidal eff cacy against C. albicans of the test
solutions after
a period of 30 minutes compared to the theoretical efficacy.
-32-
CA 02307664 2000-04-28
WO 99124542 PCT/US98/23818
While the invention has been described in conjunction with specific examples
thereof, this is illustrative only. Accordingly, many alternatives,
modifications, and
variations will be apparent to those skilled in the art in light of the
foregoing description
and it is, therefore, intended to embrace all such alternatives,
modifications, and
variations as to fall within the spirit and scope of the appended claims.
-33-
