Draize sensitization test

There are 4 basic predictive human sensitization tests in current use (1) single induction/ single challenge patch tests (2) repeated insult patch tests (RIPT) (3) RIPT with continuous exposure (modified Draize) and (4) the maximization test. Principal features of human sensitization assays are summarized in Table 2. 

RIPT has four major variations that are commonly used (1) the Draize human sensitization test (1955, 1959) (2) the Shelanski/Shelanski test (1953) (3) the Voss/Griffith test (1969, 1976) and (4) Marzill/Maibach modification. 

In the Draize human sensitization test, an occlusive patch containing the test material is applied to the upper armor upper back of 200 volunteers. The patch remains in place for 24 h and is then removed. The test site is evaluated for erythema and edema at patch removal. Twenty-four hours after the removal of the 1st patch, a 2nd patch test is applied. This... 

The Draize method has generally erred on the side of safety in that it over predicts the severity of skin damage produced by chemicals, thus producing a safety factor for those exposed. Some investigators report repeatedly that the test is not sensitive enough to separate mild from moderate irritants. Although Draize-type tests will be replaced by in vitro assays some time in the future, we have no validated in vitro substitute at present. 

There are four basic predictive human sensitization tests in current use (1) a single-induction/single-challenge patch test (2) repeated-insult patch test (RIPT) (3) RIPT with continuous exposure (modified Draize) and (4) the maximization test all of these use similar customized patches (Frosch and Kligman 1979 Kaminsky et al. 1986). Principal features of human sensitization assays are summarized in Table 1, and further details can be found in MarzuUi and Maibach (1996). For assays other than maximization, 150-200 subjects are usually tested. Henderson and Riley (1945) statistically showed that if no positive reactions are observed in 200 randomly selected subjects, as many as 15/1000 of the general population may react (95% confidence). As sample size is reduced, the likelihood of unpredicted adverse reactions in the general population increases. 

Perrot S, DutertreCatella H, Martin C et aL (2003) Rezasurin metabolism assay is a new sensitive alternative test in isolated pig cornea. Toxicol Sci 72 122-129 Prinsen MK, Koeter HB (1993) Justification of the enucleated eye test with eyes of slaughterhouse animals as an alternative to the Draize eye irritation test with rabbits. Food Chem Toxicol 31 69-76... 

The eyes of the rabbit differ in certain aspects from the eyes of humans. They are more sensitive, have a lower tear production and blink frequency and posses a nictitating membrane. Nevertheless, the Draize test predicts human ocular toxicity correctly in 85 % cases but overestimates in 10% and underestimates in 5 % (Gad and Chengelis 1991). In addition, ethical concerns have been raised in the use of animals and benefit vs risk of these tests for the protection of the human eye must be carefully evaluated. 

Results of RIPT, modified Draize test, and human maximization tests have been accepted as valid by regulatory agencies however, some sponsors routinely use one of the methods described and defend its use as the standard of the industry . FDA reviewed details of sensitization procedures and developed a guidance document (1999) for evaluating skin sensitization to chemicals in natural rubber products. 

There are several types of irritancy testing protocols that are used to comply with federal and international safety regulations. The classic Draize test was developed in 1944 to measure acute primary irritation. The test compound is applied in an occluded fashion to a clipped area of abraded and intact skin of at least six albino rabbits and evaluated 24 hr and 72 hr after patch removal. The degree of erythema and edema, ranging from one to four, is recorded to reflect severity of the irritation. Because these tests are occluded, irritancy is potentiated due to hydration, which reduces the skin barrier. The Draize test may be modified to assess sensitization by preexposing animals to a sensitizing dose of the study chemical and then rechallenging the animals at a later date to illicit the immune-mediated response. 

B. Recommendations for a Skin Sensitization Study (Modified Draize Test)... 

Iodine solution in its earlier form without povidone (tincture of iodine) proved suboptimum for chronic treatment of open wounds because of staining of the skin, local irritation, and sensitization of skin. Experimentally, such problems are avoided with PVP-I as solubility increases and chemical reactivity decreases due to binding with povidone, A review of the safety literature evaluating PVP-I concludes there is a lack of irritation as demonstrated by skin patch tests, sensitization assays, phototoxicity and allergenicity assays in humans, and eye and dermal Draize irritation assays in rabbits [17],... 

The sensitizing potential of this preservative has been established in a modified Draize test (17% of 205 volunteers) (Maibach 1971). The industrial exposure is widespread (e. g., wood, leather, and tanning industries, wallpaper glue, nylon spin finish, cutting oils). Chloracetamide in cosmetics and pharmaceutical preparations has caused contact allergy (Nater 1971 Calnan 1971) of 27 contact allergic reactions to Hirudoid ointment 7 were caused by chloracetamide (Smeenk and Prins 1972),... 

German 115 is used as an antimicrobial agent in cosmetics and has caused allergic contact dermatitis (Fisher 1975 b). In a modified Draize test, Germall 115 sensitized 2 of 150 subjects (Jordan and King 1977). It appears as a common allergen in cosmetics (FDA 1979). 

In a survey of cosmetic allergy Schorr (1971) mentioned other cosmetic preservatives and antimicrobial agents betaines, miranols (amphoteric), dimethoxane (Dioxin), and dehydroacetic acid. How often they produce allergic reactions is unknown only a few have been reported. Dimethoxane (1.25%) sensitized 50 of 205 subjects in a Draize test (Maibach 1971). 

Captan (Dangard, Vancide 89) is used as a fungicide on fruits and vegetables. Used as an antiseborrhoeic agent in a shampoo, it caused one case of photoallergic contact dermatitis (Epstein 1968). In a human Draize test 1 % Captan sensitized 4.4% of the subjects (Marzulli and Maibach 1973). 

Maibach HI (1971) Allergic sensitization potential (Draize test) in man of several preservatives. Contact Dermatitis Newslett 9 213 Maibach HI (1975a) Acute laryngeal obstruction presumed secondary to thiomersal (mer-thiolate) delayed hypersensitivity. Contact Dermatitis 1 221-222 Maibach HI (1975 b) Glutaraldehyde cross reactions to formaldehyde. Contact Dermatitis 1 326-327... 

Maibach HI, MarzuUi FN (1971) Allergic sensitization potential of riot control lacrimants. Human draize test. Contact Dermatits Newsl 5 209... 

The microbicide is judged to be non-sensitizing according to the guinea pig test (Draize test). - In a series of in-vitro and in-vivo assays it did not demonstrate mutagenic activity. Teratogenicity studies (oral application, 6 to 15 days) no-reproductive-effect level 4mg/kg/day for rabbits and lOOOmg/kg/day for rats. 

In tests for irritant response, the substances can be applied in the eye (Draize test) or on the skin. Skin sensitivity patch tests may be undertaken on man. 

This prompted us to study the pH]-arachidonic acid release test (pH]-AART) (KIdcking et al., 1994b) which is highly sensitive to membrane-toxic agents for its suitability to replace the EIT according to Draize. 

For all substances, we noticed a remarkably high sensitivity of the in-vitro method. Each of the substances found to be irritating by the Draize test was characterized as harmful in the pH]-AART at much lower concentrations. This leads to the conclusion that concentrations below the MTC values found by pH]-AART, will probably not irritate the rabbit eye. 

In case this hypothesis will be confirmed for a larger number of chemical compounds, this in-vitro system could be considered as a good candidate for the replacement of the Draize test because of its high sensitivity, objectivity, automatization degree and ethical acceptance. This test only reflect the inflammatory alterations of the Draize EIT and must therefore completed by other in vivo tests. 

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