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Use of ophthalmic topical anaesthetics
Fd Chem. Toxic. Vol. 31, No. 2, pp. 95-98, 1993 Printed in Great Britain
0278-6915/93$6.00+ 0.00 PergamonPress Lid
USE OF OPHTHALMIC TOPICAL ANAESTHETICS* V. M. SEABAUGHt~,W. A. CHAMBERS~,S. GREEN§,K. C. GUPTAll,R. N. HILLt, P. M. HURLEYt, L. A. LAMBERT§,C. C. LEEt, J. K. LEE§, P. T. LIU§, D. K. LOWTHER§,C. D. ROBERTS~,J. A. SPRINGER§and N. L. WILCOX§ :~US Environmental Protection Agency, 401 M St, SW, Washington, DC 20460, §US Food and Drug Administration, 200 C St, SW, Washington, DC 20204 and [IUS Consumer Product Safety Commission, 5401 Westbard Ave., Bethesda, MD 20207, USA Abstraet--Pretreatment of the eyes of rabbits with a topical anaesthetic can be viewed as a refinement of the test for eye irritation. It reduces pain at the time of test-material administration, decreases animal distress and permits easier application of the test agent to the eye. In some cases, however, use of an anaesthetic either alone or in combination with the test substance may alter ocular responses or provide little benefit. Although anaesthetic pretreatment may result in decreased pain at the time of test-compound administration, it does not affect possible pain after the effects of the anaesthetic have dissipated. Some anaesthetics are themselvesirritating to eyes. In addition, anaesthetics reduce blinking and tearing, thereby maintaining the test-material concentration at the surface of the eye longer. Corneal permeability may also be increased with pretreatment use of an anaesthetic, and may bring the test agent into contact with more structures of the eye. Some anaesthetics delay healing after ocular injury. All of these varied effects may result in increased irritation to the eye. Overall, pretreatment with anaesthetics has usually resulted in a tendency for slightly higher irritation scores; eye irritancy classification is usually unaffected.
Introduction
After the effects wear off, the animal could experience pain. For example, the peak anaesthetic effect for topical tetracaine is 3-8 rain, and anaesthesia lasts for 30-60rain (Ritchie and Greene, 1990). Maurice (1985) indicated that 0.1% proparacaine gives about 10 min of protection. It appears that the use of anaesthetics could reduce the normal blinking and tearing that dilute and flush materials from the eye. In rabbits, the nictitating gland may be continuous with the harderian gland (present in rabbits, but not in humans and non-human primates). The rabbit's lacrimal gland is very large and nearly encloses the globe near the orbital rim. This gland secretes a serous material. The nictitating and harderian glands secrete lipid materials. Tearing in rabbits is much less than in humans. Each of these structures can affect ocular scores by contributing to exudates and discharges or altering the properties or concentration of test substances. Rabbit eyes normally blink less frequently than do human eyes. However, topical administration of test materials to the rabbit may be followed by frequent rapid blinking and tearing. What responses these effects play in factors such as dilution, changes in pH and ionic concentration of test materials are difficult to evaluate (McDonald et al., 1987). It should be noted that some synthetic surfactants used as cleansing agents, emulsifiers, bacteriostats and wetting agents may themselves have local anaesthetic properties (Becher and Soehring, 1950; Draize and Kelley, 1952; Litt-Janssens, 1956; Martin et al., 1962; Quevauviller and Blanpin, 1955). Harris et al. (1975) reported that some suffactants and soaps
The substance of this paper was presented at the Workshop on Updating Eye Irritation Test Methods (Proposal for Regulatory Consensus) held on 26-27 September 1991, at the Sheraton Washington Hotel, Washington, DC. The workshop was the culmination of many meetings of an a d hoc Interagency Regulatory Alternatives Group (IRAG). The IRAG's proposal for anaesthetics is a consensus of those participating in the committee, and is not an official policy of any US Federal agency. Discussion Pretreatment of rabbit eyes with a topical anaesthetic has the benefit of reducing painful stimuli after administration of a test substance. However, the maximum effect for ocular anaesthetics is short-acting. The duration of anaesthesia depends on the concentration in the volume (drop) of an anaesthetic. *The opinions expressed in this article are of the authors, and do not necessarily reflect the views of any US Federal agency. $To whom correspondence should be addressed. Abbreviations: B -- benzalkonium chloride; BTS = British Toxicology Society; C = chlorhexidine; CPSC= US Consumer Product Safety Commission; ECETOC = European Chemical Industry Ecology and Toxicology Centre; EEC ~- European Economic Commission; EPA = US Environmental Protection Agency; FDA = US Food and Drug Administration; IRAG = Interagency Regulatory Alternatives Group; OECD = Organisation for Economic Cooperation and Development; 0 = oxybuprocaine; OSHA -- US Occupational Safety and Health Administration; SOT-Society of Toxicology; T ~ tetracaine. 95
96
V.M. Sr.ABAUGHet al.
produce prolonged and profound anaesthesia in rabbits. According to Grant (1974), it has been well established that local anaesthetics may affect the corneal epithelium and increase permeability. Heywood and James (1978) used 0.5% proparacaine to test 10% sodium lauryi sulphate in rabbits and monkeys. They found no appreciable differences between anaesthetized and unanaesthetized rabbit corneas. However, the anaesthetic intensified reactions in the monkey. Johnson (1980) reported increased intensity of response in eyes treated with local anaesthetics but no evidence of prolonged effect. Ulsamer et al. (1977) tested various chemicals with and without butacaine sulphate. Measurements included opacity, corneal water content, dry weight and electrophoretic protein patterns. No appreciable differences were noted in dry weight or corneal electrophoretic protein patterns with any irritants tested. With 10% acetic acid and 10% ammonia, the unanaesthetized eyes had lower opacity scores. The anaesthetized cornea had a higher water content after the administration of 5% acetic acid and 1% ammonia. Anaesthesia produced a lower water content with 3% sodium hydroxide. Hoheisei et al. (cited by Falahee et al., 1981) tested a number of irritants with and without proparacaine hydrochloride, butacaine hydrochloride, benoxinate, lidocaine and tetracaine. Both proparacaine and benoxinate were effective in producing anaesthesia; lidocaine was ineffective, and the dosing pattern for tetracaine (single application) was not fully adequate. (Note that two different instillations are required to produce anaesthesia from tetracaine.) Both proparacaine and benoxinate tended to increase the severity of ocular reactions and the time of recovery. No adverse effects were caused by the dose of tetracaine administered. Gunderson and Liebman (1944) studied regeneration of mechanically induced corneal lesions in the guinea pig with and without multiple administrations of various anaesthetics. The anaesthetic agents in order of decreasing inhibitory activity were: 10% cocaine > 4% cocaine > 4% larocaine > 1% butacaine. Both 0.5% tetracaine and 1% phenacaine showed minimal inhibitory effects. Cats and rabbits were also studied, but the anaesthetic-induced effects were equivocal. In another study, 2% cocaine and 0.5% tetracaine delayed epithelial regeneration more than 2% lidocaine (Bykov and Semenova, 1972). Marr et al. (1957) investigated the effect of repeated administration (11 times over 3 hr) of several different anaesthetics on healing of mechanically induced corneal lesions in the rat. They found that the use of amylsine, butyn, cocaine, dorsacaine, holocaine, nupercaine and ophthaine interfered with healing, whereas metycaine, tetracaine and xylocaine allowed for recovery. Pfister and Burstein (1976) reported no observable disruptive effects on rabbit eyes tested with 0.5%
proparacaine or 0.5% tetracaine with the use of an electron microscope. However, 4% cocaine produced appreciable plasma membrane injury and loss of surface cells. Leuenberger (1973) studied the ultrastructural examination of the rat corneal epithelium, using the scanning and transmission electron microscopes. He reported corneal loss of microplicae and microvilli with repeated applications of 0.4 and 1.0% benoximate, 4% cocaine, 2% lidocaine or 0.5% tetracaine. This same finding was reported as 'stippling' when the slit lamp was used. No basic differences in the type of ultrastructural changes or severity of lesions were found for the different anaesthetics tested. Maurice and Singh (1986) tested the penetration of sulphorhodamine B into the cornea of eyes of freshly killed mice, and the ratio of dye concentration in treated and control eyes provided a numerical index of toxicity to the corneal epithelium. With a group of six animals, a 50% rise in the index could be distinguished (P =0.05). Testing included 0.1 and 1% proparacaine freshly dissolved with preservative and commercial 0.5% tetracaine (preservative-free). The data indicated that proparacaine had no appreciable effect on the ratio at either concentration. Tetracaine gave a rise in the ratio of three out of a maximum achievable rise of approximately 30. Ramselaar et al. (1988) studied the effect of local anaesthetics [oxybuprocaine (O) and tetracaine (T)], and preservatives [chlorhexidine (C) and benzalkonium chloride (B)] on corneal epithelial permeability, measured by fluorophotometry in normal human eyes (10-12 people). Five instillations of one drop of an ophthalmic solution of the test compounds were administered to one eye of each volunteer, and the other eye received a control solution. Permeability was not significantly increased after the instillation of the anaesthetics. The preservatives and the combination O + C increased permeability significantly (P < 0.05) as did combinations O + B and T + B. When the number of drops was reduced to fewer than four, permeability did not increase after co-administration of T + B. Arthur et al. (1986) reported a collaborative study in which eight industrial laboratories investigated the pretreatment use of topical ocular anaesthetics, low volume and corneal or conjunctival sac instillation. They found that the pretreatment use of an ocular anaesthetic (proparacaine or tetracaine) had no appreciable effect on the course or intensity of ocular responses from treatment with 20 or 100% shampoo, 80% ethyl alcohol or 100% talc.
The I R A G proposes the following information. Topical anaesthetic pretreatment is a part of the ocular irritation test that should be used because of the benefit in ameliorating pain and distress in treated animals. When test materials are considered to have
Use of ophthalmic topical anaesthetics
97
Table 1. Results of survey question on the use of topical anaesthetics Topical anaesthetics should be employed (%): Case-by-case No Routinely determination Never Other response Respondents
Parameter Overall response summary Affiliation: Academia Government Industry Public interest Other No response Residence: USA Other Experience: Conduct/have conducted Draize test Review eye-test data Knowledge of method through literature Work with in rive systems Little/no direct experience Professional training in ocular disease Review of alternatives No response
8.33
72.92
8.33
9.38
1.04
96.00
0.00 11.10 3.75 100.00 33.33 0.00
71.43 70.37 76.79 0.00 66.67 100.00
14.29 7.41 8.93 0.00 0.00 0.00
14.29 7.41 10.17 0.00 0.00 0.00
0.00 3.70 0.00 0.00 0.00 0.00
7.29 28.13 58.33 2.08 3.13 1.04
7.32 14.29
71.95 78.57
8.54 7.14
10.98 0.00
1.22 0.00
85.42 14.58
7.14
69.05
9.52
11.90
2.38
43.75
7.55
73.58
7.55
11.32
0.00
55.21
20.00
65.00
0.00
15.00
0.00
20.83
8.82
65.65
5.88
17.65
0.00
35.42
15.38
76.92
0.00
7.69
0.00
15.34
0.00
60.00
0.00
40.00
0.00
5.21
0.00 0.00
100.00 100.00
0.00 0.00
0.00 0.00
0.00 0.00
1.04 1.04
pain-inducing potential, analgesia deserves special consideration. Anaesthetic agents should be selected that will minimize differences in test outcomes. Survey summary Participants at the Workshop responded to an anaesthetics question as part of a survey as follows (96 people submitted questionnaires): should topical anaesthetics be employed: routinely? (8.33%); caseby-case determination? (72.92%); never? (8.33%); other? (9.38%); no response (1.04%). Further information is given in Table 1. Status among organizations The Federal Hazardous Substances Act protocol for eye irritation testing is used by the US F o o d and Drug Administration (FDA), the U S Occupational Safety and Health Administration (OSHA) and the U S Consumer Product Safety Commission (CPSC); the protocol is silent about the use of anaesthetics. A CPSC (1984) policy recommends the use of 0.5% tetracaine in ocular testing. The current guideline of the European Economic Community (EEC, 1990) permits the use of anaesthetics. The Organisation for Economic Cooperation and Development ( O E C D , 1987) and the U S Environmental Protection Agency (EPA) both permit anaesthetic use if extreme pain is expected, but they caution that agents should be carefully selected to ensure that no appreciable difference in reactions is caused by the anaesthetic. The European Chemical Industry Ecology and Toxicology Centre ( E C E T O C , 1988) rec-
ommends the use of anaesthetics when unnecessary pain or distress may occur without the qualification mentioned by the O E C D and the EPA. The European Economic Community (EEC, 1990) permits anaesthetics. The Society of Toxicology (SOT, 1989) lists anaesthetic use as a consideration in ocular testing, while the British Toxicology Society (BTS, 1987) did not give unequivocal endorsement to the use of anaesthetics. REFERENCES Arthur B. H., Pennisi S. C., Dipasquale L. C., Re T., Dinardo J., Kennedy G. L., North-Root H., Penny D. A. and Sererke H. J. (1986) Effects of anaesthetic prctreatment and low volume dosage on ocular irritancy potential of cosmetics: a collaborative study. Journal of Toxicology-Cutaneous and Ocular Toxicology 5, 215-227. Becher F. and Soehring K. (1950) Deutsche Zeitschriftfiir Nervenheilkunde 164, 381. BTS (1987) A hierarchical approach to the assessment of dermal and ocular irritancy; a report of the British Toxicology Society Working Party on Irritancy. Human Toxicology 6, 269-278. Bykov N. F. and Semenova D. I. (1972) The effect of local anaesthetics reparative regeneration of corneal epithelium. Oftal'mologicheskii Zhurnal 27, 173-175. Consumer Product Safety Commission (1984) Animal testing policy. Federal Register 49, 22522-22523. Draiz¢ J. H. and Kelley E. A. (1952) Proceedings of the Scientific Section, Toilet Goods Association 17, 1-4. ECETOC (1988) Eye Irritation Testing. Monograph no. 11. European Chemical Industry Ecology and Toxicology Centre, Brussels. EEC (1990) Methods for the determination of toxicity. B5. Acute Toxicity (Eye Irritation). EEC Directive 79/831 Annex V. European Economic Community, Brussels. Falahe¢ K. J., Rose C. S. and Seifried E. (1981) Eye irritation testing: an assessment of methods and guidelines
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V.M. SEABAUGHet al.
for testing materials for eye irritancy (EPA-560/ll-82001). Grant W. M. (1974) Toxicology of the Eye. Thomas, Springfield, IL. Gundersen T. and Liebman S. D. (1944) Effect of local anaesthetics on regeneration of corneal epithelium. Archives of Ophthalmology 31, 29-33. Harris L. S., Kahanowica Y. and Shimmyo M. (1975) Cornea anaesthesia induced by soaps and surfactants, lack of correlation in rabbits and humans. Ophthalmologica 178, 320-325. Heywood R. and James R. W. (1978) Towards objectivity in the assessment of eye irritation. Journal of the Society of Cosmetic Chemists 29, 25. Johnson, A. W. (1980) Use of small dosage and corneal anaesthetic for eye testing in vivo. Proceedings of the CTFA Ocular Safety Testing Workshop: In Vivo and In Vitro Approaches. Cosmetic, Toiletries and Fragrance Association, Washington, DC, October 6-7, 1980. Leuenberger P. M. (1973) Ultrastructural changes of corneal epithelium after topical anaesthesia, Ulbrecht v. Graefas. Archiv fiir Klinische und Experimentelle Ophthalmologie 186, 73-90. Litt-Janssens M. (1956) Bruxelles--Medical 36, 1149. McDonald T. O., Seabaugh V., Shadduck J. A. and Edelhauser H. F. (1987) Eye irritation. In Advances In Modern Toxicology. pp. 641~96. Edited by F. N. Marzulli and H. I. Maibach. Hemisphere, Washington, DC. Marr W. G., Wood R., Senterfit L. and Sigelman S. (1957) Effect of topical anaesthetics. American Journal of Ophthalmology 43, 606--610. Martin G., Draize J. H. and Kelley E. A. (1962) Proceedings of the Scientific Section. Toilet Goods Association 37, 2-3. Maurice D. (1985) Chairman's introduction: pain and acute
toxicity testing in the eye. In In Vitro Toxicology: A Progress Report from the Johns Hopkins Center for Alternatives to Animal Testing. pp. 342-354. Edited by A. M. Goldberg. Mary Ann Liebert, New York. Maurice D. and Singh T. (1986) A permeability test for acute corneal toxicity. Toxicology Letters 321, 125-130. OECD (1987) Guidelines for testing chemicals. Acute eye irritation/corrosion. Publication 405. Organisation for Economic Cooperation and Development, Paris. Pfister R. R. and Burstein N. (1976) The effects of ophthalmic drugs, vehicles, and preservatives on corneal epithelium: a scanning electron microscope study. Investigative Ophthalmology 15, 246-258. Quevauviller A. and Blanpin O. (1955) Comptes Rendus des Sbances de la Socibtd de Biologic et de ses Filiales 149, 1113. Ramselaar J. A. M., Boot J. P., van Haeringen N. J. and van Best Oosterhuis J. A, (1988) Corneal epithelial permeability after instillation of ophthalmic solutions conmining local anaesthetics and preservatives. Current Eye Research 7, 947-950. Ritchie J. M. and Greene N. M. (1990) Local anaesthetics. In The Pharmacological Basis of Therapeutics. 8th Ed. p. 322. Edited by A. G. Gilman, T. W. Rail, A. S. Niew, and Taylor. Pergamon Press, New York. SOT (1989) Society of Toxicology position paper: comments on the LD50 and acute eye and skin irritation tests. Prepared by the Animals In Research Committee of the Society of Toxicology and the SOT Council. Fundamental and Applied Toxicology 13, 621~23. Ulsamer A. G., Wright P, L. and Osterberg R. E. (1977). A comparison of the effects of model irritants on anesthetized and non-anesthetized rabbit eyes (Abstract). Toxicology and Applied Pharmacology 23, 175-178.
0278-6915/93$6.00+ 0.00 PergamonPress Lid
USE OF OPHTHALMIC TOPICAL ANAESTHETICS* V. M. SEABAUGHt~,W. A. CHAMBERS~,S. GREEN§,K. C. GUPTAll,R. N. HILLt, P. M. HURLEYt, L. A. LAMBERT§,C. C. LEEt, J. K. LEE§, P. T. LIU§, D. K. LOWTHER§,C. D. ROBERTS~,J. A. SPRINGER§and N. L. WILCOX§ :~US Environmental Protection Agency, 401 M St, SW, Washington, DC 20460, §US Food and Drug Administration, 200 C St, SW, Washington, DC 20204 and [IUS Consumer Product Safety Commission, 5401 Westbard Ave., Bethesda, MD 20207, USA Abstraet--Pretreatment of the eyes of rabbits with a topical anaesthetic can be viewed as a refinement of the test for eye irritation. It reduces pain at the time of test-material administration, decreases animal distress and permits easier application of the test agent to the eye. In some cases, however, use of an anaesthetic either alone or in combination with the test substance may alter ocular responses or provide little benefit. Although anaesthetic pretreatment may result in decreased pain at the time of test-compound administration, it does not affect possible pain after the effects of the anaesthetic have dissipated. Some anaesthetics are themselvesirritating to eyes. In addition, anaesthetics reduce blinking and tearing, thereby maintaining the test-material concentration at the surface of the eye longer. Corneal permeability may also be increased with pretreatment use of an anaesthetic, and may bring the test agent into contact with more structures of the eye. Some anaesthetics delay healing after ocular injury. All of these varied effects may result in increased irritation to the eye. Overall, pretreatment with anaesthetics has usually resulted in a tendency for slightly higher irritation scores; eye irritancy classification is usually unaffected.
Introduction
After the effects wear off, the animal could experience pain. For example, the peak anaesthetic effect for topical tetracaine is 3-8 rain, and anaesthesia lasts for 30-60rain (Ritchie and Greene, 1990). Maurice (1985) indicated that 0.1% proparacaine gives about 10 min of protection. It appears that the use of anaesthetics could reduce the normal blinking and tearing that dilute and flush materials from the eye. In rabbits, the nictitating gland may be continuous with the harderian gland (present in rabbits, but not in humans and non-human primates). The rabbit's lacrimal gland is very large and nearly encloses the globe near the orbital rim. This gland secretes a serous material. The nictitating and harderian glands secrete lipid materials. Tearing in rabbits is much less than in humans. Each of these structures can affect ocular scores by contributing to exudates and discharges or altering the properties or concentration of test substances. Rabbit eyes normally blink less frequently than do human eyes. However, topical administration of test materials to the rabbit may be followed by frequent rapid blinking and tearing. What responses these effects play in factors such as dilution, changes in pH and ionic concentration of test materials are difficult to evaluate (McDonald et al., 1987). It should be noted that some synthetic surfactants used as cleansing agents, emulsifiers, bacteriostats and wetting agents may themselves have local anaesthetic properties (Becher and Soehring, 1950; Draize and Kelley, 1952; Litt-Janssens, 1956; Martin et al., 1962; Quevauviller and Blanpin, 1955). Harris et al. (1975) reported that some suffactants and soaps
The substance of this paper was presented at the Workshop on Updating Eye Irritation Test Methods (Proposal for Regulatory Consensus) held on 26-27 September 1991, at the Sheraton Washington Hotel, Washington, DC. The workshop was the culmination of many meetings of an a d hoc Interagency Regulatory Alternatives Group (IRAG). The IRAG's proposal for anaesthetics is a consensus of those participating in the committee, and is not an official policy of any US Federal agency. Discussion Pretreatment of rabbit eyes with a topical anaesthetic has the benefit of reducing painful stimuli after administration of a test substance. However, the maximum effect for ocular anaesthetics is short-acting. The duration of anaesthesia depends on the concentration in the volume (drop) of an anaesthetic. *The opinions expressed in this article are of the authors, and do not necessarily reflect the views of any US Federal agency. $To whom correspondence should be addressed. Abbreviations: B -- benzalkonium chloride; BTS = British Toxicology Society; C = chlorhexidine; CPSC= US Consumer Product Safety Commission; ECETOC = European Chemical Industry Ecology and Toxicology Centre; EEC ~- European Economic Commission; EPA = US Environmental Protection Agency; FDA = US Food and Drug Administration; IRAG = Interagency Regulatory Alternatives Group; OECD = Organisation for Economic Cooperation and Development; 0 = oxybuprocaine; OSHA -- US Occupational Safety and Health Administration; SOT-Society of Toxicology; T ~ tetracaine. 95
96
V.M. Sr.ABAUGHet al.
produce prolonged and profound anaesthesia in rabbits. According to Grant (1974), it has been well established that local anaesthetics may affect the corneal epithelium and increase permeability. Heywood and James (1978) used 0.5% proparacaine to test 10% sodium lauryi sulphate in rabbits and monkeys. They found no appreciable differences between anaesthetized and unanaesthetized rabbit corneas. However, the anaesthetic intensified reactions in the monkey. Johnson (1980) reported increased intensity of response in eyes treated with local anaesthetics but no evidence of prolonged effect. Ulsamer et al. (1977) tested various chemicals with and without butacaine sulphate. Measurements included opacity, corneal water content, dry weight and electrophoretic protein patterns. No appreciable differences were noted in dry weight or corneal electrophoretic protein patterns with any irritants tested. With 10% acetic acid and 10% ammonia, the unanaesthetized eyes had lower opacity scores. The anaesthetized cornea had a higher water content after the administration of 5% acetic acid and 1% ammonia. Anaesthesia produced a lower water content with 3% sodium hydroxide. Hoheisei et al. (cited by Falahee et al., 1981) tested a number of irritants with and without proparacaine hydrochloride, butacaine hydrochloride, benoxinate, lidocaine and tetracaine. Both proparacaine and benoxinate were effective in producing anaesthesia; lidocaine was ineffective, and the dosing pattern for tetracaine (single application) was not fully adequate. (Note that two different instillations are required to produce anaesthesia from tetracaine.) Both proparacaine and benoxinate tended to increase the severity of ocular reactions and the time of recovery. No adverse effects were caused by the dose of tetracaine administered. Gunderson and Liebman (1944) studied regeneration of mechanically induced corneal lesions in the guinea pig with and without multiple administrations of various anaesthetics. The anaesthetic agents in order of decreasing inhibitory activity were: 10% cocaine > 4% cocaine > 4% larocaine > 1% butacaine. Both 0.5% tetracaine and 1% phenacaine showed minimal inhibitory effects. Cats and rabbits were also studied, but the anaesthetic-induced effects were equivocal. In another study, 2% cocaine and 0.5% tetracaine delayed epithelial regeneration more than 2% lidocaine (Bykov and Semenova, 1972). Marr et al. (1957) investigated the effect of repeated administration (11 times over 3 hr) of several different anaesthetics on healing of mechanically induced corneal lesions in the rat. They found that the use of amylsine, butyn, cocaine, dorsacaine, holocaine, nupercaine and ophthaine interfered with healing, whereas metycaine, tetracaine and xylocaine allowed for recovery. Pfister and Burstein (1976) reported no observable disruptive effects on rabbit eyes tested with 0.5%
proparacaine or 0.5% tetracaine with the use of an electron microscope. However, 4% cocaine produced appreciable plasma membrane injury and loss of surface cells. Leuenberger (1973) studied the ultrastructural examination of the rat corneal epithelium, using the scanning and transmission electron microscopes. He reported corneal loss of microplicae and microvilli with repeated applications of 0.4 and 1.0% benoximate, 4% cocaine, 2% lidocaine or 0.5% tetracaine. This same finding was reported as 'stippling' when the slit lamp was used. No basic differences in the type of ultrastructural changes or severity of lesions were found for the different anaesthetics tested. Maurice and Singh (1986) tested the penetration of sulphorhodamine B into the cornea of eyes of freshly killed mice, and the ratio of dye concentration in treated and control eyes provided a numerical index of toxicity to the corneal epithelium. With a group of six animals, a 50% rise in the index could be distinguished (P =0.05). Testing included 0.1 and 1% proparacaine freshly dissolved with preservative and commercial 0.5% tetracaine (preservative-free). The data indicated that proparacaine had no appreciable effect on the ratio at either concentration. Tetracaine gave a rise in the ratio of three out of a maximum achievable rise of approximately 30. Ramselaar et al. (1988) studied the effect of local anaesthetics [oxybuprocaine (O) and tetracaine (T)], and preservatives [chlorhexidine (C) and benzalkonium chloride (B)] on corneal epithelial permeability, measured by fluorophotometry in normal human eyes (10-12 people). Five instillations of one drop of an ophthalmic solution of the test compounds were administered to one eye of each volunteer, and the other eye received a control solution. Permeability was not significantly increased after the instillation of the anaesthetics. The preservatives and the combination O + C increased permeability significantly (P < 0.05) as did combinations O + B and T + B. When the number of drops was reduced to fewer than four, permeability did not increase after co-administration of T + B. Arthur et al. (1986) reported a collaborative study in which eight industrial laboratories investigated the pretreatment use of topical ocular anaesthetics, low volume and corneal or conjunctival sac instillation. They found that the pretreatment use of an ocular anaesthetic (proparacaine or tetracaine) had no appreciable effect on the course or intensity of ocular responses from treatment with 20 or 100% shampoo, 80% ethyl alcohol or 100% talc.
The I R A G proposes the following information. Topical anaesthetic pretreatment is a part of the ocular irritation test that should be used because of the benefit in ameliorating pain and distress in treated animals. When test materials are considered to have
Use of ophthalmic topical anaesthetics
97
Table 1. Results of survey question on the use of topical anaesthetics Topical anaesthetics should be employed (%): Case-by-case No Routinely determination Never Other response Respondents
Parameter Overall response summary Affiliation: Academia Government Industry Public interest Other No response Residence: USA Other Experience: Conduct/have conducted Draize test Review eye-test data Knowledge of method through literature Work with in rive systems Little/no direct experience Professional training in ocular disease Review of alternatives No response
8.33
72.92
8.33
9.38
1.04
96.00
0.00 11.10 3.75 100.00 33.33 0.00
71.43 70.37 76.79 0.00 66.67 100.00
14.29 7.41 8.93 0.00 0.00 0.00
14.29 7.41 10.17 0.00 0.00 0.00
0.00 3.70 0.00 0.00 0.00 0.00
7.29 28.13 58.33 2.08 3.13 1.04
7.32 14.29
71.95 78.57
8.54 7.14
10.98 0.00
1.22 0.00
85.42 14.58
7.14
69.05
9.52
11.90
2.38
43.75
7.55
73.58
7.55
11.32
0.00
55.21
20.00
65.00
0.00
15.00
0.00
20.83
8.82
65.65
5.88
17.65
0.00
35.42
15.38
76.92
0.00
7.69
0.00
15.34
0.00
60.00
0.00
40.00
0.00
5.21
0.00 0.00
100.00 100.00
0.00 0.00
0.00 0.00
0.00 0.00
1.04 1.04
pain-inducing potential, analgesia deserves special consideration. Anaesthetic agents should be selected that will minimize differences in test outcomes. Survey summary Participants at the Workshop responded to an anaesthetics question as part of a survey as follows (96 people submitted questionnaires): should topical anaesthetics be employed: routinely? (8.33%); caseby-case determination? (72.92%); never? (8.33%); other? (9.38%); no response (1.04%). Further information is given in Table 1. Status among organizations The Federal Hazardous Substances Act protocol for eye irritation testing is used by the US F o o d and Drug Administration (FDA), the U S Occupational Safety and Health Administration (OSHA) and the U S Consumer Product Safety Commission (CPSC); the protocol is silent about the use of anaesthetics. A CPSC (1984) policy recommends the use of 0.5% tetracaine in ocular testing. The current guideline of the European Economic Community (EEC, 1990) permits the use of anaesthetics. The Organisation for Economic Cooperation and Development ( O E C D , 1987) and the U S Environmental Protection Agency (EPA) both permit anaesthetic use if extreme pain is expected, but they caution that agents should be carefully selected to ensure that no appreciable difference in reactions is caused by the anaesthetic. The European Chemical Industry Ecology and Toxicology Centre ( E C E T O C , 1988) rec-
ommends the use of anaesthetics when unnecessary pain or distress may occur without the qualification mentioned by the O E C D and the EPA. The European Economic Community (EEC, 1990) permits anaesthetics. The Society of Toxicology (SOT, 1989) lists anaesthetic use as a consideration in ocular testing, while the British Toxicology Society (BTS, 1987) did not give unequivocal endorsement to the use of anaesthetics. REFERENCES Arthur B. H., Pennisi S. C., Dipasquale L. C., Re T., Dinardo J., Kennedy G. L., North-Root H., Penny D. A. and Sererke H. J. (1986) Effects of anaesthetic prctreatment and low volume dosage on ocular irritancy potential of cosmetics: a collaborative study. Journal of Toxicology-Cutaneous and Ocular Toxicology 5, 215-227. Becher F. and Soehring K. (1950) Deutsche Zeitschriftfiir Nervenheilkunde 164, 381. BTS (1987) A hierarchical approach to the assessment of dermal and ocular irritancy; a report of the British Toxicology Society Working Party on Irritancy. Human Toxicology 6, 269-278. Bykov N. F. and Semenova D. I. (1972) The effect of local anaesthetics reparative regeneration of corneal epithelium. Oftal'mologicheskii Zhurnal 27, 173-175. Consumer Product Safety Commission (1984) Animal testing policy. Federal Register 49, 22522-22523. Draiz¢ J. H. and Kelley E. A. (1952) Proceedings of the Scientific Section, Toilet Goods Association 17, 1-4. ECETOC (1988) Eye Irritation Testing. Monograph no. 11. European Chemical Industry Ecology and Toxicology Centre, Brussels. EEC (1990) Methods for the determination of toxicity. B5. Acute Toxicity (Eye Irritation). EEC Directive 79/831 Annex V. European Economic Community, Brussels. Falahe¢ K. J., Rose C. S. and Seifried E. (1981) Eye irritation testing: an assessment of methods and guidelines
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