`In Therapy®
`
`Volume 18 No. 5
`September/October 2001
`
`Effects of Common
`Ophthalmic Preservatives
`on Ocular Health
`
`Robert Noecker, M.D.
`Tucson, Arizona
`
`ABSTRACT
`Preservatives are an important component of ophthalmic preparations, providing
`antimicrobial activity in the bottle and preventing decomposition of active drug.
`Often underrecognized, however, are the significant cytotoxic effects of preserva-
`tives associated with long-term therapy and especially use of multiple preserved
`drugs. The most common preservatives in ophthalmic preparations for glaucoma
`and surface eye disease—benzalkonium chloride (BAK), chlorobutanol, sodium
`perborate, and stabilized oxychloro complex (SOC)—were reviewed. Compared
`with other preservatives, SOC caused the least amount of damage to rabbit
`corneal epithelial cells. BAK has demonstrated cytotoxic effects in cell culture, as
`well as in animal and human studies. Physicians should consider treatment with
`new-generation preparations containing low-risk preservatives such as SOC, espe-
`cially in patients receiving multiple ophthalmic medications.
`
`Keywords: ophthalmic preservatives; benzalkonium chloride;
`chlorobutanol; SOC; sodium perborate
`
`INTRODUCTION
`The contents of multidose medication containers used twice daily
`often undergo bacterial contamination within 1 or 2 weeks.1 As a result,
`the US Food and Drug Administration and the US Pharmacopoeia man-
`date that all multidose ophthalmic preparations contain a preservative to
`ensure a nonhazardous degree of contamination. By providing a level of
`antimicrobial activity in the bottle, preservatives limit bacterial, mycotic,
`and amoebal ocular infections caused by contaminated solutions and
`prolong shelf life by preventing biodegradation and maintaining drug
`potency. The primary concern with many preservatives is not their effica-
`cy but, rather, their recognized cytotoxic side effects.2
`
`©2001 Health Communications Inc.
`Transmission and reproduction of this material in whole
`or part without prior written approval are prohibited.
`
`0639
`
`Address reprint requests to
`Robert Noecker, M.D.
`655 North Alveron Way, Suite 108
`Tucson, AZ 85711
`
`205
`
`Page 1 of 11
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`SENJU EXHIBIT 2089
`LUPIN v. SENJU
`IPR2015-01097
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`High concentrations of some preservatives can damage and irritate ocular tissue.
`Preservative-free products may prevent the toxic side effects, but they are expensive
`and the small unit-dose containers can be difficult to use, hindering compliance.3
`Nonetheless, some patients require preservative-free products because of sensitivi-
`ties or allergies. The goal of the physician should be to prescribe effective agents that
`contain preservatives with minimal effects on ocular tissues.
`
`CLINICAL RELEVANCE
`In chronic diseases, such as glaucoma or dry eye syndrome, high concentrations
`of preservatives or repeated exposure to preserved medications increases the likeli-
`hood of adverse effects. For example, high incidences of endothelial damage, epithe-
`lial edema, and bulbous keratopathy characterize patients with glaucoma, dry eyes,
`infections, or iritis whose use of preservative-containing ophthalmic solutions is fre-
`quent and prolonged.4 Even with infrequent administration, preserved solutions
`may be contraindicated in the presence of trophic ulcers or other states of severely
`compromised corneal epithelial integrity.5 Patients with defective epithelia or
`corneal ulcers may be most at risk because of increased penetration of the medica-
`tion and preservatives.4
`
`Glaucoma
`Long-term use of antiglaucoma drugs has been linked to toxic and inflammatory
`changes of the ocular surface.6-8 Conjunctival biopsies from glaucoma patients show
`a significant increase in immune cells and fibroblasts possibly related to prolonged
`treatment.9,10 Repeated doses of preserved eyedrops can have a cumulative effect,
`and extended contact with the epithelium may lead to chronic irritation and sub-
`conjunctival fibrosis, increasing the risk that trabeculectomy will fail.7,11 Multidrug
`treatment of glaucoma may also raise the risk for the ocular surface effects of preser-
`vatives. Less frequent daily administration, lower preservative concentrations (cur-
`rently ≤0.01% for most antiglaucoma drugs), and new formulations may help to
`minimize this ocular surface damage.
`
`Keratoconjunctivitis sicca (KCS)
`Patients with severe KCS may need to instill tear substitutes as often as every
`20 minutes. Preservatives may worsen the condition by disrupting the precorneal
`tear film and damaging the epithelial surface.5 Many corneal specialists believe that
`KCS may be aggravated by frequent use of preservative-containing artificial tears,
`especially because these patients may not produce enough natural tears to dilute a
`harmful preservative.2,5 Overuse of nonprescription eyedrops can also contribute to
`adverse effects. When patients with glaucoma or KCS discontinue use of preserva-
`tive-containing medications, allergic complaints or chronic irritation of the conjunc-
`tiva and eyelids also ceases.4
`
`Managing Preservative-Induced Ocular Damage
`Damage due to ophthalmic preservatives often goes unnoticed because it is diffi-
`cult to differentiate side effects of an active ingredient from those of the preservative.
`
`206
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`R. Noecker
`Ophthalmic Preservatives
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`Page 2 of 11
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`The following sections review the mechanism of action and results of tissue culture
`and animal studies to compare toxic effects of four preservatives.
`
`COMMONLY USED PRESERVATIVES
`A spectrum of preservatives are found in nearly every type of ophthalmic solu-
`tion. Benzalkonium chloride (BAK) is one of the most commonly used preservatives.
`Less common are benzododecinium bromide (BDD), cetrimonium chloride, thiom-
`ersal, methyl parahydroxybenzoate, sorbic acid, polyquarternium ammonium chlo-
`ride (PQAC), polyaminopropyl biguanide, and hydrogen peroxide. Tables 1 and 2
`list commonly used products and their preservative concentrations.
`
`Table 1. Preservative Composition of Antiglaucoma Medications
`
`Trade Name
`
`Alphagan®
`Alphagan P®
`Azopt®
`Betagan®
`Betoptic S®
`Cosopt®
`Lumigan™
`Propine®
`Rescula®
`Timoptic®
`Timoptic-XE®
`Trusopt®
`Xalatan®
`
`Manufacturer
`
`Allergan, Inc.
`Allergan, Inc.
`Alcon
`Allergan, Inc.
`Alcon
`Merck & Co., Inc.
`Allergan, Inc.
`Allergan, Inc.
`CIBA Vision
`Merck & Co., Inc.
`Merck & Co., Inc.
`Merck & Co., Inc.
`Pharmacia & Upjohn
`
`Preservative
`
`BAK 0.005%
`SOC 50 ppm
`BAK 0.01%
`BAK 0.005%
`BAK 0.01%
`BAK 0.0075%
`BAK 0.005%
`BAK 0.005%
`BAK 0.015%
`BAK 0.01%
`BDD 0.012%
`BAK 0.0075%
`BAK 0.02%
`
`SOC = stabilized oxychloro complex.
`
`Mechanism of Action
`Preservatives interfere with microbial organisms by causing lysis of plasma mem-
`branes, inhibiting cellular metabolism, oxidizing or coagulating cellular con-
`stituents, or promoting hydrolysis.2 Preservatives can be classified in two main cat-
`egories as oxidants or detergents.12 Oxidative preservatives, such as stabilized
`oxychloro complex (SOC) and sodium perborate, are usually small molecules that
`penetrate cell membranes and disrupt cellular function by modifying lipids, pro-
`teins, and DNA.13 Their membrane-destabilizing activity is less potent than that of
`
`Advances In Therapy®
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`Table 2. Over-the-Counter Products for Ocular Surface Disease and Their
`Preservative Concentrations
`
`Product Name
`
`Manufacturer
`
`Preservative
`
`Use
`
`GenTeal®
`Hypotears®
`Naphcon-A®
`Refresh Tears®
`Tears Naturale II®
`Vasocon-A®
`Visine®
`
`CIBA Vision
`CIBA Vision
`Alcon
`Allergan, Inc.
`Alcon
`CIBA Vision
`Pfizer
`
`Sodium perborate
`BAK 0.01%
`BAK 0.01%
`SOC 50 ppm
`Polyquad 0.001%
`BAK 0.01%
`BAK 0.01%
`
`Artificial tears
`Artificial tears
`Vasoconstrictor
`Artificial tears
`Artificial tears
`Vasoconstrictor
`Vasoconstrictor
`
`detergent preservatives. At low levels, oxidative preservatives have an advantage
`over detergent preservatives by providing enough activity against microorganisms
`while exerting only negligible toxic effects on eukaryotic cells. This occurs because
`many microorganisms cannot cope with oxidative stress. In comparison, mam-
`malian cells are equipped with antioxidants, oxidases, and catalases to neutralize
`the effect of a low-level oxidant.
`Detergent preservatives, such as BAK, are usually monomeric or polymeric com-
`pounds that have surfactant effects and alter cell membrane permeability by caus-
`ing lipid dispersion and lysis of cytoplasmic contents.14 Some may have a similar
`action on eukaryotic cells and cause cytotoxic effects. Mammalian cells cannot neu-
`tralize detergent preservatives, which can be incorporated into the cell by liposomes
`or other intracellular vacuoles and cause cellular damage.15
`
`Purite®* (SOC)
`SOC destroys many types of bacteria as well as the fungus Aspergillus niger.16
`Introduced in 1996, it consists of an equilibrium mixture of oxychloro species—
`99.5% chlorite (ClO2), 0.5% chlorate (ClO3), and trace amounts of chlorine dioxide
`(ClO2)—that have bactericidal and viricidal activity.17 In saline solution, SOC gener-
`ates chlorine dioxide free radicals in the presence of microbial contamination.
`However, SOC is an oxidizing, not a chlorinating, agent.12 The free radicals provide
`the antimicrobial activity by oxidizing unsaturated lipids and glutathione in the cell.12
`When SOC is administered in the eye, it is converted into natural tear components,
`such as sodium and chloride ions, oxygen, and water. This conversion occurs by
`way of cascade-type reactions between SOC and tear-film components and pho-
`tolytic reactions between SOC and light.16,17
`Mild cytotoxic effects and an excellent safety record have earned SOC a US Envir-
`onmental Protection Agency category II rating as a mild eye irritant on the basis
`of rabbit studies. Although exposure to 2% SOC can produce slight irritation of
`
`*Registered trademark of Allergan, Inc., Irvine, Calif, USA.
`
`208
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`R. Noecker
`Ophthalmic Preservatives
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`the conjunctiva, cornea, and eyelid, this concentration is higher than that used in most
`commercial products. Efficacy at low concentrations (0.005% w/v) that are benign to
`the eye makes SOC an ideal ophthalmic preservative. Safety and tolerability were
`established in a study of 62 patients with mild to moderate dry eye who were treated
`with an SOC-containing product four to eight times per day for 4 weeks.18 No evi-
`dence of in vivo or in vitro mutagenicity or carcinogenicity has been found.
`Chlorine dioxide has been used since 1944 to purify water, and conventional
`doses appear to be safe for that indication.17 Mild cytotoxic effects make it a common
`ingredient in toothpaste, mouthwash, and antacids. Chlorine dioxide destroys
`microorganisms in fish, fruit, and vegetables without altering the food’s nutritive
`and organoleptic qualities.17
`
`Sodium Perborate
`One of the first oxidative preservatives, sodium perborate is converted to hydro-
`gen peroxide when combined with water. Sodium perborate oxidizes cell walls
`or membranes, affects membrane-bound enzymes, and disrupts protein synthesis.
`On entering the eye, it is rapidly decomposed to water and oxygen by catalase and
`other enzymes in the conjunctival sac.16 Sodium perborate is bactericidal and can kill
`A. niger.16 Low levels retain antimicrobial activity and are comfortable in the eye.
`However, hydrogen peroxide levels between 30 and 100 ppm, normally produced
`by ophthalmic preparations containing sodium perborate, can cause ocular sting-
`ing.19,20 Limited testing has also identified sodium perborate as a direct-acting in
`vitro mutagen.21 It can also destabilize cell walls and membranes, albeit to a lesser
`degree than other types of preservatives.
`
`BAK
`The quaternary ammonium compound BAK is the most common antimicrobial
`preservative,3,5 found at an average concentration of 0.01% (range, 0.004%–0.02%)3
`in topical multiuse ophthalmic preparations. Nearly all antiglaucoma medications
`contain BAK. Highly efficacious against numerous microbes, BAK denatures pro-
`teins and causes lysis of cytoplasmic membranes. The surfactant effect of quaternary
`ammonium compounds, including BAK, can solubilize the intercellular cement of
`the corneal epithelium, thereby increasing the compound’s penetration.4,14
`Moreover, because BAK can accumulate and remain in ocular tissue for relatively
`lengthy periods,4,15 it can induce different types of cell death in a dose-dependent
`manner: growth arrest at low concentrations, apoptosis at 0.01%, and necrosis at
`higher concentrations.21
`In an antiglaucoma preparation, BAK does not alter the drug’s ability to lower
`intraocular pressure but can modify the ocular surface with long-term use.22 For exam-
`ple, timolol maleate, which contains 0.01% BAK, rapidly decreased cell viability and
`numbers in a human conjunctival cell line.23 Similarly, patients treated with timolol
`maleate 0.5% containing 0.01 g/100 mL of BAK exhibited ocular surface damage
`attributed to a reduced rate of aqueous layer production and impairment of the tear-
`film mucus layer.24 In another study, 127 patients instilling various antiglaucoma
`drugs containing BAK had significant conjunctival metaplasia compared with
`patients not using topical treatment.25 There was no significant difference on cytolog-
`ic examination between those using any of the medications for less or more than
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`Advances In Therapy®
`Volume 18 No. 5, September/October 2001
`
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`1 year, suggesting that the damage occurred relatively quickly. Long-term use of
`antiglaucoma medications containing BAK changes the conjunctival surface and tear-
`film function,26 which may increase the risks attendant on future glaucoma surgery.
`Patients with dry eye syndrome may have increased vulnerability to BAK-
`induced effects. Artificial tears containing BAK enhance corneal epithelial perme-
`ability, contributing to ocular surface disease.27 In these patients, the cornea is par-
`ticularly susceptible to the effects of preservatives because the epithelium is exposed
`to the full strength of topical preparations.2 Further, severely affected patients do not
`produce enough tears to dilute the compound in the eyedrop. Ophthalmic prepara-
`tions that contain high concentrations of BAK interfere with the integrity of the
`superficial lipid layer and reduce tear breakup time, causing the duplex tear film to
`become unstable.28,29 This compromised stability may not be physiologically harm-
`ful, as the lipid layer of the tear film is re-formed every 15 to 30 seconds, but should
`be taken into account for patients with a compromised tear film.28 Ideally, these indi-
`viduals should use products with preservatives that do not break up the tear film.
`At low BAK concentrations or with infrequent use, BAK-preserved preparations
`may pose little risk.
`
`Chlorobutanol
`As an alcohol-based preservative, chlorobutanol lacks surfactant activity14; there-
`fore, unlike BAK, it does not increase penetration of additional chlorobutanol mole-
`cules into the cell.4 Instead, chlorobutanol disorganizes the lipid structure of the mem-
`brane, which increases permeability and leads to cell lysis. Chlorobutanol has
`broad-spectrum antimicrobial action. In vivo, even at concentrations 100 times that of
`commercial products, it did not damage rabbit cornea, including the endothelium.4
`Chlorobutanol 0.5% did not affect the stability of the tear-film lipid layer in non–
`contact lens users30 and enhanced in vitro transcorneal permeation of ibuprofen.31
`Despite its relative safety, chlorobutanol 0.5% m/v in artificial tears caused irrita-
`tion in more than 50% of patients in a double-blind crossover study,32 most likely as
`a result of cellular retraction and cessation of normal cytokinesis, cell movement,
`and mitotic activity.33 Degeneration of human corneal epithelial cells and generation
`of conspicuous membranous blebs have also been observed.34 In normal intact rab-
`bit corneas, chlorobutanol 0.5% caused cytoplasmic swelling and occasional breaks
`in the external cell membrane. In keratectomized rabbit corneas, some mitochondria
`showed swollen and distorted cristae.35 A 0.1% concentration caused near-complete
`loss of the squamous cell layer in isolated rabbit cornea, suggesting disruption of the
`barrier and transport properties of the corneal epithelium.35 Chlorobutanol also
`inhibits oxygen use by the cornea, which increases susceptibility to infection.36
`Another study,37 however, found minimal cytotoxic effects of chlorobutanol 0.5%
`in artificial tears given twice a day for 12 days to pigmented rabbits. Scanning elec-
`tron microscopy of the corneal epithelial surface detected only occasional cell exfo-
`liation, which peaked after 2 to 3 days and returned close to zero at day 12.37 Also,
`compared with BAK 0.004% to 0.02%, chlorobutanol 0.2% to 0.5% is less toxic to rab-
`bit corneal epithelial cells in vitro.38 In human corneal epithelial cells, the cytotoxic
`effects of chlorobutanol occur less rapidly and are less severe than those of BAK.33
`
`210
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`R. Noecker
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`SOC Compared With Other Preservatives
`Relative to other preservatives, SOC is minimally toxic to the eye, as determined
`by scanning electron microscopy. In one study,39 rabbits were treated four times
`a day for 7 days with artificial tears containing polyquad 0.001%, sodium perborate,
`or SOC 50 ppm. On completion of the study, the corneas were removed, fixed in
`2.6% glutaraldehyde, osmicated, serially dehydrated, and critical point dried. The
`specimens were mounted on aluminum stubs, gold sputtered-coated, and evaluat-
`ed. The untreated rabbit corneal epithelium had extensive microvilli and tight inter-
`cellular junctions (Fig 1), as did the eye treated with SOC (Fig 2), and was nearly
`identical to the eye treated with sodium perborate (Fig 3). The eye exposed to
`polyquad 0.001% showed extensive superficial epithelial erosion and lack of pro-
`truding microvilli (Fig 4). Compared with the untreated eye, the extent of corneal
`epithelial damage was polyquad > sodium perborate > SOC.38
`Attempts to improve drug tolerability by minimizing the toxic effects of preserva-
`tives are underway. One aim is to decrease cumulative exposure to the preservative.
`A once-daily form of timolol with 0.012% BDD (a preservative similar to BAK) is avail-
`able; however, BDD also damages the corneal epithelium and the gel-forming prepa-
`ration may prolong contact of the preservative with the corneal surface.3 Another
`approach is to reformulate existing products with better-tolerated preservatives. One
`such product, a brimonidine compound approved by the FDA in March 2001, has
`replaced BAK with SOC in the current formulation. A 12-month clinical comparison
`in patients with glaucoma or ocular hypertension40 showed that brimonidine-SOC
`was well tolerated and produced a significantly lower incidence of allergic conjunc-
`tivitis than brimonidine, as well as equivalent IOP-lowering efficacy.
`
`Fig 1.
`
`Scanning electron microscopy of untreated rabbit corneal epithelium. The tissue is
`normal, with extensive microvilli and tight intercellular junctions (×14,000).
`
`Advances In Therapy®
`Volume 18 No. 5, September/October 2001
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`Fig 2.
`
`Scanning electron microscopy of rabbit corneal epithelium treated with SOC. The epithelium
`is normal with extensive microvilli and tight intercellular cell-to-cell junctions (×14,000).
`
`Fig 3.
`
`Scanning electron microscopy of rabbit corneal epithelium treated with sodium perborate.
`The mostly normal epithelium has extensive microvilli and tight epithelial cell-to-cell
`junctions (×14,000).
`
`212
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`Ophthalmic Preservatives
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`Fig 4.
`
`Scanning electron microscopy of rabbit corneal epithelium treated with polyquad 0.001%.
`Extensive superficial epithelial erosion and lack of protruding microvilli are evident
`(×14,000).
`
`CONCLUSIONS
`Many preservatives in eyedrops induce histopathologic, inflammatory, and toxic
`changes on the ocular surface. Choosing eyedrops with a less harmful preservative
`or with lower concentrations of preservatives can be beneficial to the patient. SOC
`represents a new generation of ophthalmic preservative that breaks down into nat-
`ural tear components on instillation and thus has low potential to cause toxic effects.
`Preservative-induced toxic reactions can escalate with the use of multiple oph-
`thalmic drops. Therefore, physicians should consider drugs containing low-risk
`preservatives or preservatives at concentrations least likely to cause damage.
`
`REFERENCES
`
`1. Schein OD, Hibberd PL, Starck T, Baker AS, Kenyon KR. Microbial contamination of in-use ocular
`medications. Arch Ophthalmol. 1992;110:82-85.
`2. Tripathi BJ, Tripathi RC, Kolli SP. Cytotoxicity of ophthalmic preservatives on human corneal
`epithelium. Lens Eye Toxicol Res. 1992;9:361-375.
`3. Pisella PJ, Fillacier K, Elena PP, Debbasch C, Baudouin C. Comparison of the effects of pre-
`served and unpreserved formulations of timolol on the ocular surface of albino rabbits.
`Ophthalmic Res. 2000;32:3-8.
`4. Gasset AR, Ishii Y, Kaufman HE, Miller T. Cytotoxicity of ophthalmic preservatives. Am J
`Ophthalmol. 1974;78:98-105.
`5. Berdy GJ, Abelson MB, Smith LM, George MA. Preservative-free artificial tear preparations.
`Assessment of corneal epithelial toxic effects. Arch Ophthalmol. 1992;110:528-532.
`
`Advances In Therapy®
`Volume 18 No. 5, September/October 2001
`
`213
`
`Page 9 of 11
`
`
`
`6. Debbasch C, Rat P, Warnet J-M, De Saint Jean M, Baudouin C, Pisella P-J. Evaluation of the toxici-
`ty of benzalkonium chloride on the ocular surface. J Toxicol Cutan Ocul Toxicol. 2000;19:105-115.
`7. Baudouin C. Side effects of antiglaucomatous drugs on the ocular surface. Curr Opin Ophthalmol.
`1996;7:80-86.
`8. Baudouin C. Mechanisms of failure in glaucoma filtering surgery: a consequence of antiglauco-
`matous drugs? Int J Clin Pharmacol Res. 1996;16:29-41.
`9. Broadway DC, Grierson I, O’Brien C, Hitchings RA. Adverse effects of topical antiglaucoma
`medication, I: the conjunctival cell profile. Arch Ophthalmol. 1994;112:1437-1445.
`10. Sherwood MB, Grierson I, Millar L, Hitchings RA. Long-term morphologic effects of antiglauco-
`ma drugs on the conjunctiva and Tenon’s capsule in glaucomatous patients. Ophthalmology.
`1989;96:327-335.
`11. Wilson WS, Duncan AJ, Jay JL. Effect of benzalkonium chloride on the stability of the precorneal
`tear film in rabbit and man. Br J Ophthalmol. 1975;59:667-669.
`12. Spickett C. Studies of Cellular Responses to Purite and Other Preservatives. Strathclyde, UK:
`University of Strathclyde; 2001.
`13. Block SS. Disinfection, Sterilization, and Preservation. Malvern, Pa: Lea & Febiger; 1991.
`14. Kreiner C. Biochemical aspects of ophthalmic preservatives. Contacto. 1979;Nov:10-14.
`15. Grant WM, Schuman JS. Toxicology of the Eye. Springfield, Ill: Charles C Thomas; 1990.
`16. Grant R, Ajello M, Vlass E. Salt water or high tech? A look at two new rinsing solutions for
`contact lenses. Optician. 1996;212:38-41.
`17. Masschelein WJ, Rice RG. Chlorine Dioxide. Chemistry and Environmental Impact of Oxychlorine
`Compounds. Ann Arbor, Mich: Ann Arbor Science Publishers; 1979.
`18. Rozen S, Abelson M, Giovanoni A, Welch D. Assessment of the comfort and tolerance of
`0.5% carboxymethylcellulose preserved with Purite (Refresh Tears) in dry eye sufferers. Invest
`Ophthalmol Vis Sci. 1998;39(4):S451.
`19. Vaughan JS, Porter DA. A new in vitro method for assessing the potential toxicity of soft contact
`lens care solutions. CLAO J. 1993;19:54-57.
`20. Paugh JR, Brennan NA, Efron N. Ocular response to hydrogen peroxide. Am J Optom Physiol
`Opt. 1988;65:91-98.
`21. Seiler JP. The mutagenic activity of sodium perborate. Mutat Res. 1989;224:219-227.
`22. Campagna P, Macri A, Rolando M, Calabria G. Chronic topical eye preservative-free beta-blocker
`therapy effect on the ocular surface in glaucomatous patients. Acta Ophthalmol Scand Suppl.
`1997;224:53.
`23. De Saint Jean M, Debbasch C, Brignole F, Rat P, Warnet JM, Baudouin C. Toxicity of preserved
`and unpreserved antiglaucoma topical drugs in an in vitro model of conjunctival cells. Curr
`Eye Res. 2000;20:85-94.
`24. Yalvac IS, Gedikoglu G, Karagoz Y, et al. Effects of antiglaucoma drugs on ocular surface. Acta
`Ophthalmol Scand. 1995;73:246-248.
`25. Turacli E, Budak K, Kaur A, Mizrak B, Ekinci C. The effects of long-term topical glaucoma
`medication on conjunctival impression cytology. Int Ophthalmol. 1997;21:27-33.
`26. Arici MK, Arici DS, Topalkara A, Guler C. Adverse effects of topical antiglaucoma drugs on
`the ocular surface. Clin Exp Ophthalmol. 2000;28:113-117.
`27. Gobbels M, Spitznas M. Corneal epithelial permeability of dry eyes before and after treatment
`with artificial tears. Ophthalmology. 1992;99:873-878.
`28. Holly FJ. Surface chemical evaluation of artificial tears and their ingredients, II: interaction with
`a superficial lipid layer. Contact Lens. 1978;4:52-65.
`
`214
`
`R. Noecker
`Ophthalmic Preservatives
`
`Page 10 of 11
`
`
`
`29. Baudouin C, de Lunardo C. Short-term comparative study of topical 2% carteolol with and
`without benzalkonium chloride in healthy volunteers. Br J Ophthalmol. 1998;82:39-42.
`30. Tomlinson A, Trees GR. Effect of preservatives in artificial tear solutions on tear film evaporation.
`Ophthalmic Physiol Opt. 1991;11:48-52.
`31. Gupta M, Majumdar DK. Effect of concentration, pH, and preservative on in vitro transcorneal
`permeation of ibuprofen and flurbiprofen from non-buffered aqueous drops. Indian J Exp Biol.
`1997;35:844-849.
`32. Fassihi AR, Naidoo NT. Irritation associated with tear-replacement ophthalmic drops.
`A pharmaceutical and subjective investigation. S Afr Med J. 1989;75:233-235.
`33. Tripathi BJ, Tripathi RC. Cytotoxic effects of benzalkonium chloride and chlorobutanol on
`human corneal epithelial cells in vitro. Lens Eye Toxicol Res. 1989;6:395-403.
`34. Collin HB. Ultrastructural changes to corneal stromal cells due to ophthalmic preservatives.
`Acta Ophthalmol (Copenh). 1986;64:72-78.
`35. Burstein NL, Klyce SD. Electrophysiologic and morphologic effects of ophthalmic preparations
`on rabbit cornea epithelium. Invest Ophthalmol Vis Sci. 1977;16:899-911.
`36. Burstein N. Corneal cytotoxicity of topically applied drugs, vehicles, and preservatives. Surv
`Ophthalmol. 1980;25:15.
`37. Doughty MJ. Twice-daily use of a chlorobutanol-preserved artificial tear on rabbit corneal
`epithelium assessed by scanning electron microscopy. Ophthalmic Physiol Opt. 1992;12:457-466.
`38. Lazarus HM, Imperia PS, Botti RE, Mack RJ, Lass JH. An in vitro method which assesses corneal
`epithelial toxicity due to antineoplastic, preservative and antimicrobial agents. Lens Eye Toxicol
`Res. 1989;6:59-85.
`39. Way WA, Matsumoto S, Apel LJ, Wiese A, Tarlo K, Vehige J. Purite™ as a non-disruptive preser-
`vative for lubricating eye drop solutions in comparison to alternative preservatives.
`Invest Ophthalmol Vis Sci. 2001;42:S39.
`40. Walters TR, Batoosingh AL. 12-Month evaluation of brimonidine-Purite compared with
`brimonidine in patients with glaucoma or ocular hypertension. Invest Ophthalmol Vis Sci. 2001;
`42:S558.
`
`Advances In Therapy®
`Volume 18 No. 5, September/October 2001
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`215
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`Page 11 of 11