20
CHAPTER 37 Cosmetic Toxicology JOHN F. CORBETT, RAJ K. SHARMA, WILLIAM E. DRESSLER Bristol-Myers Squibb Worldwide Beauty Care, Stamford, Connecticut INTRODUCTION COSMETIC SAFETY REGULATION Decorative cosmetics and fragrances have been part of almost every culture for many thousands of years but a formal approach to the toxicological eval- uation of cosmetic products has only evolved during the 20th century. Initially; the evaluation involved only dermal, ocular, and, in some cases, mucous membrane irritation toxicity and allergenicity via the Draize eye and skin tests in rabbits and guinea pigs. This was sometimes followed by clinical studies in humans. The recognition that the skin was not an imperme- able barrier to topically applied materials required that systemic toxicity, including both acute and chronic effects, also be taken into account. For the most part, and for obvious reasons, ingredients used in products subject to incidental ingestion, such as lipstick, have received particular attention, followed by those used in the eye area and products applied to and left on the skin, while rinse-off products, except for hair dyes, have been of least concern. This chapter will consider the scope of testing that is generally accepted as desirable for cosmetic prod- ucts, together with the chemical nature of the active ingredients used in selected specialized hair products such as dyes, bleaches, permanent waves, and straighteners. For other varied types of cosmetic prod- ucts, use experience and some general principles re- garding safety evaluation and risk assessment will be discussed. In addition, reference will be made to the particular requirements for and methods of reviews of safety data in the European Union, the United States and Japan. The United States In the U.S., the Food, Drug and Cosmetic Act des- ignates the Food and Drug Administration as the agency responsible for cosmetic safety. For this purpose, the FDA has defined cosmetics as "articles to be rubbed, poured, sprinkled, or sprayed on or introduced into, or otherwise applied to the human body or any part thereof for cleansing, beau- tifying, promoting attractiveness, or altering the ap- pearance, and articles intended for use as a compo- nent of any such articles; except that such term shall not include soap." The exemption for soap applies only to a composition consisting of an alkali metal salt of a fatty acid that is intended only for cleansing. Certain cosmetic products that are "intended for use in the diagnosis, cure, mitigation, treatment or prevention of disease" are classified by FDA as drugs. This category includes sunscreen products, anticavity toothpastes, antiperspirants (as opposed to deodor- ants, which are "cosmetics"), antidandruff prepara- tions, "medicated" skin lotions and liquids, skin pro- tectants, and hair restorers. Under the regulations, the active ingredients in cos- metic drug products must be safe and effective ac- cording to the appropriate monograph covering the claimed indication. The only category of cosmetic ingredient subject to FDA approval is colors used for purposes other than dyeing the hair. There are only 36 "certified colors" and 23 "permitted" colors available for general use in cosmetics, and a further 7 that are permitted only for TOXICOLOGY 899 Copyright 9 1999 Academic Press. All rights of reproduction in any form reserved.

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Page 1: Cosmetic Toxi

C H A P T E R

37

Cosmetic Toxicology JOHN F. CORBETT, RAJ K. SHARMA, WILLIAM E. DRESSLER

Bristol-Myers Squibb Worldwide Beauty Care, Stamford, Connecticut

I N T R O D U C T I O N C O S M E T I C SAFETY R E G U L A T I O N

Decorative cosmetics and fragrances have been part of almost every culture for many thousands of years but a formal approach to the toxicological eval- uation of cosmetic products has only evolved during the 20th century. Initially; the evaluation involved only dermal, ocular, and, in some cases, mucous membrane irritation toxicity and allergenicity via the Draize eye and skin tests in rabbits and guinea pigs. This was sometimes followed by clinical studies in humans.

The recognition that the skin was not an imperme- able barrier to topically applied materials required that systemic toxicity, including both acute and chronic effects, also be taken into account. For the most part, and for obvious reasons, ingredients used in products subject to incidental ingestion, such as lipstick, have received particular attention, followed by those used in the eye area and products applied to and left on the skin, while rinse-off products, except for hair dyes, have been of least concern.

This chapter will consider the scope of testing that is generally accepted as desirable for cosmetic prod- ucts, together with the chemical nature of the active ingredients used in selected specialized hair products such as dyes, bleaches, permanent waves, and straighteners. For other varied types of cosmetic prod- ucts, use experience and some general principles re- garding safety evaluation and risk assessment will be discussed. In addition, reference will be made to the particular requirements for and methods of reviews of safety data in the European Union, the United States and Japan.

The United States

In the U.S., the Food, Drug and Cosmetic Act des- ignates the Food and Drug Administration as the agency responsible for cosmetic safety.

For this purpose, the FDA has defined cosmetics as "articles to be rubbed, poured, sprinkled, or sprayed on or introduced into, or otherwise applied to the human body or any part thereof for cleansing, beau- tifying, promoting attractiveness, or altering the ap- pearance, and articles intended for use as a compo- nent of any such articles; except that such term shall not include soap." The exemption for soap applies only to a composition consisting of an alkali metal salt of a fatty acid that is intended only for cleansing.

Certain cosmetic products that are "intended for use in the diagnosis, cure, mitigation, treatment or prevention of disease" are classified by FDA as drugs. This category includes sunscreen products, anticavity toothpastes, antiperspirants (as opposed to deodor- ants, which are "cosmetics"), antidandruff prepara- tions, "medicated" skin lotions and liquids, skin pro- tectants, and hair restorers.

Under the regulations, the active ingredients in cos- metic drug products must be safe and effective ac- cording to the appropriate monograph covering the claimed indication.

The only category of cosmetic ingredient subject to FDA approval is colors used for purposes other than dyeing the hair. There are only 36 "certified colors" and 23 "permitted" colors available for general use in cosmetics, and a further 7 that are permitted only for

TOXICOLOGY 899 Copyright 9 1999 Academic Press.

All rights of reproduction in any form reserved.

Page 2: Cosmetic Toxi

900 Corbett et al.

specific uses. In addition, a number of lakes of the soluble certified colors are available.

For all other cosmetic ingredients, the safety eval- uation is the responsibility of the manufacturer. In order to assist its members in this, the Cosmetic, Toi- letry, and Fragrance Association (CTFA) established in 1976 the Cosmetic Ingredient Review (CIR) to re- view all the available data on an ingredient and to decide whether the ingredient is safe under the con- ditions of use. The CIR expert panel comprises six independent scientists as voting members and a non- voting member representing each of the CTFA, the FDA, and the consumer. By 1996, the CIR had re- viewed over 600 ingredients, finding 64% to be safe as used, 27% to be safe for use under defined condi- tions, 8% to have insufficient data, and five ingredi- ents to be unsafe.

The CIR expert panel currently requests the follow- ing as a minimum data requirement for reviewing the safety of the cosmetic ingredients that are on its pri- ority list:

1. Current concentration of use data. 2. Chemistry data, including method of manufacture

and impurities. 3. UV-absorption data; if absorption occurs in the

UVA or UVB range, photosensitization data are needed.

4. Skin-irritation and -sensitization data at concentra- tions of use in humans.

5. Dermal-absorption data; if significant dermal ab- sorption occurs, 28-day dermal-toxicity and devel- opment-toxicity data are needed.

6. Two genotoxicity studies, one using a mammalian system; if positive, a 2-year dermal carcinogenicity assay performed using NTP methods is needed.

Additional studies may be requested based on the nature and use of the ingredient.

The European Union In the European Union, cosmetics are regulated un-

der the Cosmetics Directive (76/768/EEC) of 1976 and amendments thereto. For the purpose of regulation, cosmetics are more broadly defined than in the United States, viz. "any substance or preparation intended to be placed in contact with various external parts of the human body (epidermis, hair system, nails, lips and external genitalia) and with the teeth or mucous mem- branes of the oral cavity with a view exclusively or mainly to cleaning them, perfuming them, changing their appearance and /o r correcting body odors and / or protecting them or keeping them in good condi-

tion." This definition covers many of the product cat- egories that are considered to be over-the-counter drugs in the U.S.

In common with the United States, cosmetics do not require premarket clearance in Europe. However, certain ingredients, notably colorants (other than hair dyes), preservatives, and sunscreens, do require ap- proval before they can be used in cosmetic products. The list of permitted colorants (Annex IV) is broader than the U.S. list, comprising 157 materials.

The approval process involves the submission of data via COLIPA, the European trade association, to the European Commission. The data is evaluated by a group of independent experts who make up the Sci- entific Committee for Cosmetology and Non-Food Products (SCCNFP) and who decide whether the in- gredient can be accepted for listing in the appropriate Annex. Data requirements are similar to those listed above for the CIR.

The General Toxicological Requirements for Cos- metic Ingredients (1996 revision) state:

When requested, the manufacturer shall provide the Com- mission with the information set out below: 1. Acute toxicity 2. Dermal absorption 3. Dermal irritation 4. Mucous membrane irritation 5. Skin sensitization 6. Sub-chronic toxicity 7. Mutagenicity 8. Phototoxicity and Photomutagenciny (in case of UV-

fight absorbing substances) 9. Human data (if available)

When considerable oral intake can be expected or when the data or dermal absorption do indicate a considerable pene- tration of the ingredients through the skin, taking into ac- count the toxicological profile of the substance and its chem- ical structure, the following further information may be necessary: 10. Toxicokinetics 11. Teratogenicity, Reproduction toxicity, Carcinogenicity,

and additional Genotoxicity.

Additionally, there is now a legal requirement, for- realized in Article 7a of the sixth amendment to the Cosmetics Directive, that cosmetic companies hold a technical dossier of information on each of their prod- ucts. This requirement applies to every cosmetic prod- uct, both retail and professional, including imported products.

This dossier must include information or product composition, specifications, and method of manufac- ture, as well as an assessment of product safety car- fled out by an appropriately qualified expert. The assessment must take account of the general toxico-

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Cosmetic Toxicology 901

logical profile of the ingredients, their chemical struc- tures, and the potential levels of exposure.

The SCC is also responsible for recommending the listing of substances prohibited for use in cosmetics (Annex II) and substances that can be used with cer- tain limitations with regard to product category, con- centration, and/or special labeling requirements (An- nex III).

In addition to these requirements, the EU regula- tion for the Notification of New Substances must be followed by companies introducing new chemicals. These regulations state that "a material is considered to be a new substahce within the EU if it is not listed in EINECS (European Inventory of Existing Chemical Substances)." EINECS is a closed list (closed in 1981), that is, no new substances can now be added.

Substances not in EINECS must be notified accord- ing to Council Directive 92/32/EEC (seventh amend- ment to the EC Dangerous Substances Directive) prior to marketing in the EU. Notified substances are issued in ELINCS (European List of New Chemical Sub- stances). ELINCS is an open list, that is, new sub- stances may be added once notified according to the Directive.

The precise contents of a notification depend on the quantity of the substance to be placed or already placed on the market. The notification system is banded, the information required increasing as the quantity of a substance placed on the market in- creases.

Japan The Japanese regulation of cosmetics is the most

restrictive. While the definition of a cosmetic is similar to that in the U.S., the number of product types con- sidered to be "quasi-drugs" (analogous to cosmetic drugs or O.T.C. products) is much greater. Thus, in addition to the U.S. categories, products to combat bad breath, bath preparations, hair dyes, permanent waves, talcum powder, depilatories, shaving lotions, and skin packs are considered "quasi-drugs."

In Japan, all cosmetic products are subject to pre- market approval by the Ministry of Health and Wel- fare and can contain only those ingredients included in the Comprehensive Licensing Standards of Cosmet- ics by Category (CLS) and these must conform to cer- tain defined specifications. New ingredients can be added to the list on the basis of the submission of appropriate toxicological data.

For quasi-drugs, there are lists of permitted active ingredients for use in hair dyes, permanent waves,

and medicated toothpastes. Lists for other categories are being developed.

For countries other than those discussed here, in- gredients acceptable in the United States and/or Eu- rope would usually be acceptable.

HAIR COSMETICS

Permanent Waves The conformational integrity of the hair fiber is

maintained by various interactions between protein chains making up the cortex. These comprise polar (electrostatic) and nonpolar interactions between the side chains of the amino acids, hydrogen bonding, and covalent bonding-- in particular the interchain disulfide bonds of the amino acid cystine.

A temporary set can be imparted to hair simply by wetting the hair, restraining it in the desired new conformation, and drying it while still constrained. On the release of the constraint, the hair will retain the new conformation until it is again exposed to wa- ter or even to high humidity.

If a more permanent effect is desired, it is necessary to break and reform covalent bonds, as is done in the process of permanent waving. For this purpose, thiols (compounds containing t h e - S H group) have been found to be the most effective at reducing the disul- fide bond.

Essentially the permanent wave process involves about six or seven steps:

1. Rolling, in which a section of hair is wound onto a perm rod. In this step the hair may be first damp- ened with water or with the waving lotion.

2. The application of waving lotion, if the hair was dry-rolled or water-rolled, to the hair on the rods in such a way as to saturate each hair mass.

3. The creep step, in which the reduced hair is left on the rods for about 20 minutes to allow movement of segments within the individual cortex.

4. Water rinsing, which is then carried out with the hair still on the rods.

5. Neutralization, in which the disulfide bonds are rebuilt. This step can be performed on or off the rods depending on the desired end result.

6. Rinsing to remove the excess neutralizer and other chemicals, which is performed after removal of the rods.

7. Styling and drying.

Because of the complexity of the process and the skill required to obtain satisfactory results, the market

Page 4: Cosmetic Toxi

902 Corbett et al.

for home perms is small compared to that for salon products. As a consequence, professional hairdressers have greater exposure to perm chemicals than do the general public.

Although literally thousands of thiol compounds have been investigated as reducing agents for the ker- atin disulfide bonds, only derivatives of thioglycolic acid, cysteine and, to a lesser extent, ammonium bi- sulfite and ammonium thiolacetate have achieved commercial importance.

The thiols work best at pH levels slightly below the pKa of the thiol group. Most "cold wave" or "alkaline" perms are based on ammonium thioglycolate at a pH of about 9.2 (the pKa of thioglycolic acid is 10.4), while the "acid waves" employ glyceryl monothioglycolate (GMT), which has a pK~ of 7.8 and is applied in a composition having a pH of about 6.8, the reduction step generally being performed under a heating de- vice such as a hair dryer of infrared lamp.

The reduction step can be represented as

K-S-S-K + RS---~ K-S-S-R + KS-

K-S-S-R + RS----* K-S- + R-S-S-R

where K represents a keratin protein chain and RS- is the thiolate perming agent.

The rebuilding step, which usually employs hydro- gen peroxide, although the previously widely used potassium bromate neutralizer is still occasionally used, can be represented as

2K-S-H + H202 ----)K-S-S-K + 2H20

In the case of bisulfite perms, which give much weaker results, the reduction step is performed at pH 6 and involves the formation of a Bunte salt.

K-S-S-K + HSO3 ---* K-S-H + K-S-SO;

Neutralization is performed by reversing the above reaction by treatment with a solution buffered at pH 8 and containing a little hydrogen peroxide, which serves to oxidize the liberated sulfite to sulfate.

Apart from occasional cases of skin irritation, der- mal problems among recipients of permanent waves are rare. More common are problems with dryness and brittleness of the hair due to overprocessing in the form of too strong a lotion for the particular hair type, too long a residence time of the waving lotion, or inadequate rinsing or neutralization.

The use of ammonium thioglycolate in the cold wave process has the advantage that evaporation of the ammonia results in lowering of the pH and, with that, the possibility of over-reduction of the disulfide bonds. The use of alkanolamines in place of ammonia

has the advantage of better odor but carries with it the possibility of over-reduction.

There have been few examples of sensitization from exposure to the alkaline thioglycolate waves, even among hairdressers. On the other hand, glyceryl monothioglycolate has been shown to be a sensitizer and allergies to it are reported to be common among hairdressers in some countries, notably Germany and Holland.

Adverse reactions can be minimized by the adher- ence to certain guidelines.

1. Commercial use of single-dose rather than bulk GMT to avoid contamination of work surfaces and measuring equipment.

2. Water rolling rather than lotion rolling of the hair. 3. The use of suitable disposable gloves while han-

dling the hair and rinsing of the gloves before dis- posal.

4. General good work practices to avoid contamina- tion of work surfaces and equipment.

Hair Straighteners Conventional thioglycolate and sulfite permanent

waving products are relatively ineffective when used to straighten African hair because the hair tends to revert to the curly state within a short time.

Treatment with highly alkaline compositions (pH --- 13), known in the trade as relaxers, is the most effective way to achieve a long-lasting result. The most widely used products contain 2-4% sodium hy- droxide in an emulsion system containing petrolatum, fatty alcohols, mineral oil, and 40-50% water.

Products promoted as "no lye relaxers" employ ei- ther guanidinium hydroxide or lithium hydroxide as the alkali.

Under these highly alkaline conditions it is be- lieved that a C-S bond in cystine is broken to give an aminoacrylic residue that reacts with the disulfide to give lanthiomine or with an aminoalkyl side chain to give lysinoalanine. These new interchain linkages are unreactive to chemicals used in hair products, includ- ing alkaline and reducing agents, thus creating a per- manent conformational change.

Due to their high alkalinity, great care must be exercised in the use of relaxers. Every effort must be made to avoid contact with the scalp, which would result in a chemical burn. Sometimes a base coat of petrolatum is applied to the scalp and around the hair line, although this is very difficult given the density of the hair.

In use, the thick composition is applied to the hair, section by section, and pulled through with a comb

Page 5: Cosmetic Toxi

Cosmetic Toxicology 903

vertically away from the scalp. If the hair is combed parallel to the scalp, a right angle will be set into the hair where it emerges from the follicle and, after about two weeks of new hair growth this will become a point of breakage when the hair is combed. Contact time for these products is about 15 minutes and should not exceed 20 minutes.

Hair Dyes There are a wide variety of products marketed for

changing the color of human hair and they can con- veniently be divided into three groups according to the type of coloring materials.

The most important of these groups, representing about 80% of the market in the U.S., is the oxidation dyes, which employ colorless precursors and an oxi- dizing agent (usually hydrogen peroxide) that to- gether are capable of producing colored molecules inside the hair. This class is often referred to as "per- manent" because the dyes formed do not readily dif- fuse out of the hair during subsequent shampooing. A unique feature of this class of product is the ability to formulate them to simultaneously lighten the hair's natural color and deposit the synthetic color. This is because hydrogen peroxide is a good bleaching agent for the melanin, which constitutes the natural hair pigment. Thus, oxidation dyes can be used to change the color of hair to darker or lighter shades that may have warm red, gold, neutral, or cool ash nuances, as well as to cover grey without necessarily producing a darker shade than that of the individual's naturally pigmented hair.

The second group in commercial importance is the direct or preformed dyes. This group can be further subdivided into semipermanent dyes, which with- stand six to twelve shampoos, and temporary dyes, which wash out in one shampoo.

A smaller segment of the market is the group based on natural dyes extracted from plants, which have lasting properties similar to those of the semiperma- nent dyes.

Finally, there is the group for products based on metal salts. In Europe and the United States, these usually contain a solution of lead acetate and are known as progressive dyes, which are mainly used by men for grey coverage. In Asia, there are also prod- ucts that employ polyhydric phenols, particularly py- rogallol, and ferrous salts. These are used by both men and women exclusively for covering grey hair.

Oxidation Dyes The oxidation dyes have been in use for over 100

years and were based on the discovery that a mixture

of p-phenylenediamine and an oxidizing agent could produce color on hair. This color was shown to be due to the formation of Bandrowski's base.

Modern oxidation dyes contain four essential clas- ses of ingredients in a suitably thickened detersive base.

p-Phenylenediamine and other para diamines, and o- and p-aminophenols, referred to as the primary intermediates, are materials that individually are ca- pable of being oxidized to give colored reaction prod- ucts.

The second class contains the couplers, which are aromatic compounds that do not themselves produce colored products on oxidation but, when present dur- ing the oxidation of the primary intermediates, result in the production of colored species.

The couplers are generally meta di-functional ben- zene derivatives such as m-phenylenediamines, m- aminophenols, and resorcinols. Additionally, certain monohydric phenols, such as 1-naphthols and thymol, are useful as couplers.

The color-forming reactions have been shown to involve the initial oxidation of the primary interme- diate to form a p-benzoquinonediimine from p-diam- ines, or monoamine from p-aminophenols. These im- ines are highly reactive toward the nucleophilic couplers and react to form diphenylamine leuco-dyes, which are rapidly oxidized to indo-dyes, (Fig. 1).

H 2 N / ~ NH2 = H N / ~ NH

Primary Intermediate CH 3

+ H 2 N ' ~ O H I, Coupler H

H2N" ~eu~2Nye "~ OH

Oxidation l / ~ N . A CH 3

Indoaniline Dye

Page 6: Cosmetic Toxi

904 Corbett et al.

p-Phenylenediamine 2,5-Diaminotoluene N-Phenyl-p-phenylenediamine N,N'-bis(2-Hydroxyethyl)-phenylenediamine 2-/3-Hydroxyethyl-phenylenediamine N,N'-bis-( ]3- Hydroxyethyl ) N,N'-bis-(4'-aminop henyl )- 1,3-di-

amino-2-hydroxypropane p-Aminophenol 3-Methyl-4-aminophenol N-Methyl-p-aminophenol o-Aminophenol

The oxidant of choice is hydrogen peroxide (gen- erally 3% in the applied mixture), although some products employ solid oxidants, such as urea perox- ide, sodium percarbonate, or perborate. These latter are efficient in initiating the color-forming reactions but have little bleaching power for melanin.

The chemistry involved in the color formation and in melanin bleaching occurs optimally at a pH of 9.0- 9.5 and the preferred alkalizing agent is ammonia. Monoethanolamine can also be used and has a better odor but at the cost of some bleaching efficiency. If no bleaching is required, as in the "no-lift, . . . . demiper- manent," "tone on tone," or "deposit only" colorants, then a higher alkanolamine such as aminomethylpro- panol can be used. Secondary alkanolamines, such as diethanolamine, are now avoided because of the pos- sibility of nitrosamine formation.

The m-difunctional benzene couplers, such as res- orcinol and m-aminophenol, having no substituent para to either of the functional groups, react beyond the initial indo-dye stage to produce trinuclear or even polymeric dyes that are drabber in color than

the binuclear indo-dyes formed with couplers having a blocking group para to one of the functional groups and serve to provide a background brownish or black- ish coloration. In contrast, the blocked couplers pro- duce intense bright colors.

Using the ingredients listed in Tables 1 and 2, it is possible to formulate a full line of natural-looking shades and many "fashion" shades. Occasionally cer- tain red and yellow direct dyes of the nitroamino- phenol or nitrophenylenediamine type are added when a particularly bright shade is desired.

Direct Dyes for Hair

The temporary hair dyes include a variety of acid and /o r basic dyes mainly chosen from among those used in textile dyeing. In the truly temporary dyes, which are removed by a single shampoo treatment, acid dyes are precipitated by the addition of a cationic surfactant to give a fine dispersion of colored parti- cles, which is applied to the hair and left on to dry without rinsing, thus leaving a colored surface coating on the hair.

Semipermanent colorants, which withstand 6 to 12 shampoos, employ low-molecular-weight dyes that were specifically designed for use in hair dyes (Fig. 2). These are generally nitro phenylenediamines or nitroaminophenols and their N-substituted deriva- tives, which cover the color range of yellow to violet, together with a small number of azo or anthraquinone dyes, which provide violet to blue tones. These dyes are small enough to diffuse into the hair under ambi- ent conditions and, consequently, will also diffuse out slowly during subsequent washings of the hair.

Typical dyes used in semipermanent colorants are shown in Table 3. The dyes are presented in a low- foaming surfactant base and are applied to the hair

::~:::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::::: ::::::::::::::::::::::::::::::::~:~:~:~:: ~.-~:,.,~:~;;::::-:::~.'.::~,;;.;:~;;~ ~:~. ,:-,: ~ : -~ , - , a~ : ~ : - ,~..:~;~.~.:..,.;.,.~:~,,~::~..:~: ~ . . . . . . . . . . . ~ . ~ . , ~ , , . . . ~ . . . . . . . . . . . . . . . . . . . . . ~ a , . . . . . . . . . . . . . . . . . . . . . . . . . . . . o: ............................................ 9 ................................................. . o . .....

Color produced with Coupler p-Diamine p-Aminophenol

Resorcinol 2-Methylresorcinol m-Aminophenol m-Phenylenediamine 2,4-Diaminophenoxyethanol 1-Methoxy-2-amino-4-~-hydroxyethylaminobenzene 1,3-Bis-(2,4-diaminophenoxy-propane) 5-Amino-2-methylphenol 5-]3-Hydroxyethylamino-2-methylphenol 1-Naphthol

greenish brown beige yellowish brown beige warm brown pinkish beige blue-black reddish beige blue-violet orange-red blue-violet orange-red blue-violet orange-red magenta red-orange magenta red-orange blue-violet red

Page 7: Cosmetic Toxi

Cosmetic Toxicology 905

NHCH2CH2OH

NH 2

HC Red 3

H

NI) OH

HC Orange 1

NHCH2CH2OH

N(CH2CH2OH) 2

HC Blue 2

OCH2GH2OH ~ NHCH2OH2OH NO2

HC Yellow 4

CH 3

O NHCH2CH2OH

Disperse Blue 3

~ ~ ~ I ~ ~ ~ : ~ ~ ~ i ~ ! ~ ~ 4 . . . ~ | ~

and left for 10 to 30 minutes before the composition is rinsed from the hair.

Natural Dyes for Hair The practice of using plant extracts to dye the hair

goes back over 2500 years when henna was employed for this purpose by the Egyptian nobility.

Henna is made from the dried macerated leaves of shrubs of the Lawsonia family. The major coloring agent is lawsone (2-hydroxy-l,4-naphthoquinone) to- gether with a smaller amount of luteolin, 2-(3',4'-di- hydroxyphenyl)-5,6-dihydroxyflavone, which is ex- tracted when leaves are mixed with hot water. The resulting paste is applied to the hair and left on for up to 1 hour, imparting an orange-red color that is quite resistant to shampooing.

In order to extend the range of shades, henna is sometimes mixed with other plant materials, such as indigo, logwood, chamomile, and hematin.

Natural dyes have a particular advantage in being essentially nonsensitizing. However, the coloring re- sults and, in particular the grey coverage do not en' courage their widespread use.

In some parts of the world, henna is mixed with powdered p-phenylenediamine and used to dye both the hair and the skin. Such a practice should certainly be discouraged on toxicological grounds.

Metallic Dyes for Hair The use of lead acetate solutions containing sulfur

to develop a gradual darkening of the hair derives

from the ancient practice of dipping lead combs in vineger and combing the hair.

The modern products command less than 1% of the market and are mainly used by men for masking grey hair. The product is applied to the hair on a daily basis and, by interaction with the air and hydrogen sulfide, possibly produced by bacterial action on the sulfur, produces a deposit of lead oxide and lead sul- fide on the surface of the hair. With repeat applica- tion, a significant brown coloration develops. Studies have shown that use of these products does not in- crease the body burden of lead, provided that the scalp is intact.

The other metallic dyes are used mainly in Asia and have one of two component systems, one contain- ing a ferrous salt and the other containing pyrogallol.

Hair-Dye Toxicology Sensitization

Because a number of the oxidative dye intermedi- ates are known to be sensitizers, it is an almost uni- versal requirement that these products carry a cau- tionary notice saying that the product may cause a reaction in some individuals. Additionally, instruc- tions for carrying out a preliminary skin-patch test must be included. In the U.S., the mandatory warning reads, "This product contains ingredients which may cause skin irritation on certain individuals and a pre- liminary test according to accompanying directions should first be made. This product must not be used for dyeing the eyelashes or eyebrows: to do so may

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906 Corbett et al.

iiiiiiiiiiii tii ii N-(2-Hydroxyethyl)-o-nitroaniline (HC Yellow 2) 4-Nitro-o-phenylenediamine N~-(2-Hydroxyethyl)-4-nitro-o-phenylenediamine (HC Yellow 5) 2-Amino-3-nitrophenol O,N-Bis(2-hydroxyethyl)-2-amino-5-nitrophenol (HC Yellow 4) N-(2-Hydroxyethyl)-2-amino-5-nitrophenol 4-Amino-3-nitrophenol N-2-Hydroxyethyl)-2-amino-3-nitrophenol N-2-(Hydroxyethyl)-4-amino-3-nitroanisole 1-(3-Methylamino-4-nitrophenoxy)propane-2,3 diol 3-Methylamino-4-nitrophenoxyethanol 2-Nitro-p-phenylenediamine NM2-Hydroxyethyl)-2-nitro-p-phenylenediamine (HC Red 3) N4-(2-Hydroxyethyl)-2-nitro-p-phenylenediamine (HC Red 7) N ~-Methyl-2-nitro-p-phenylenediamine N 1,N4,N4-Tris(2-hydroxyethyl)-2-nitro-p-phenylenediamine (HC Blue 2) N4-(2-Hydroxyethyl)-N ~,N4-dimethyl-2-nitro-p-phenylenediamine 4-Nitro-m-phenylenediamine Picramic acid N-Methyl-iso-picramic acid 4-Amino-2-nitrodiphenylenediamine (HC Red) 2-Hydroxy-2'-nitrodiphenylenediamine (HC Orange) 4-(p-Aminophenylazo)-N,N-bis(2-hydroxyethyl)aniline 4-(p-Aminophenylazo)-3-methyl-N,N-bis(2-hydroxyethyl) aniline (Disperse Red 17) 1,4,5,8-Tetraminoanthraquinone (Disperse Blue 1) 1,4-Diaminoanthraquinone 1-Amino-4-methylaminoanthraquinone 1-(2-Hydroxyethylamino)-4-methyaminoanthraquinone (Disperse Blue 3)

cause blindness". There is no evidence to suggest that the use around the eye would cause blindness. In fact most of the ingredients have been subjected to in vivo animal tests and the incidence of accidental instilla- tion of the product during use is not insignificant; yet no evidence of permanent injury has been docu- mented. In Europe the warning states, "Can cause allergic reactions. Contains phenylenediamines. Do not use to dye eyelashes or eyebrows," and, on prod- ucts for professional use, "wear suitable gloves."

Clinical tests have shown some of the nitro and anthraquinine dyes to be weak allergens and that there is some cross-sensitivity with p-phenylenedi- amine.

Mutagenicity

Much attention has been focused on the mutagenic- ity of hair dyes since the initial report by Ames of a study conducted as part of a biochemistry class exper- iment that showed positive responses for a number of dyes in Salmonella tymphimurium. While positive activ- ity has also been reported for a number of dye mate- rials in cultured mammalian cells, there have been only infrequent reports of positive results using in

vivo test systems. Accumulated experience has shown that Ames mutagenicity is not a reliable predictor of rodent carcinogenic potential, particularly for aro- matic amines. Such compounds were among the 33/ 301 NTP-tested chemicals, and invariably showed Ames-positive results for both transpecies, as well as for noncarcinogens. Batteries of short-term mutage- nicity tests, despite some initial promise of facilitating the detection of rodent carcinogens, do not appear to improve substantially on the overall performance of the Salmonella assay for chemicals in general, and for hair dyes specifically. As an example, a highly puri- fied sample of HC Blue 1, which was negative in a battery of short-term tests, produced hepatocellular carcinomas in mice, as did an unpurified mutagenic commercial material. However, other investigators subsequently found very small amounts of mutagenic impurities in ethanol-recrystallized HC Blue 1 using an Ames-directed HPLC fractionation assay. They speculated that these impurities may have been present in insufficient quantities to express an effect at a 5 mg maximum dose in the Ames test.

The consideration of the chemistry of the oxidative hair color process and in vitro assay conditions also

Page 9: Cosmetic Toxi

Cosmetic Toxicology 907

have implications for the interpretation of hair dye mutagenicity data. For example, it was found that aqueous and freshly prepared DMSO solutions of p- phenylenediamine were nonmutagenic in Ames tests (TA98 and TA1538), while DMSO solutions allowed to stand at room temperature for 4 hours were active. The mutagenicity of p-phenylenediamine in the Ames test, the mouse lymphoma assay, and in human lym- phocytes in v i t r o has been attributed to the formation of Bandrowski's base, a trimer of p-phenylenedi- amine. The addition of adequate amounts of couplers and restricted incubation times prevented the muta- genic activity in these assays. This is especially rele- vant because Bandrowski's base is not formed under conditions of actual hair dyeing in formulations that contain materials that preferentially couple with p- phenylenediamine to produce the desired colored dye polymer.

In view of these observations and complexities, the interpretation of the in v i t r o or in v i v o mutagenic ef- fects of individual hair dye materials and the possible relevance of such findings to the carcinogenic or mu- tagenic potential of dye materials under actual condi- tions of use needs to be carefully considered in a risk- assessment process.

C a r c i n o g e n i c i t y

A substantial number of hair-dye ingredients have been subjected to subchronic toxicity, chronic toxicity, and life-time or multigeneration carcinogenicity stud- ies, as well as to evaluations of teratologic and muta-

genic potential. A number of these studies have in- volved composite oxidative and semipermanent (direct) hair-dye formulations given by topical appli- cation, dietary administration, or oral gavage (Table 4). These composite formulations reflected complete palettes of individual dyes used to produce various hair color shades.

Other than local irritant-type effects at the sites of topical application, none of these formulation studies have found any evidence of tumorigenicity of skin or internal organs, or clear evidence of other systemic nontumor pathology.

In addition to carcinogenicity tests involving topi- cal application of composite hair dye formulations, the U.S. National Toxicology Program (NTP) has eval- uated a number of individual dye materials by feed- ing or gavage routes using maximally tolerated doses (MTDs) determined from preliminary 14- and 90-day subchronic studies.

The results from these studies are summarized in Table 5 (direct dyes) and Table 6 (oxidative dyes). A few of these dyes, none of which remain in use, have elicited positive responses or "clear evidence" of car- cinogenicity in one or more species or sex (e.g., 2,4- diaminoanisole, 2,4-toluenediamine, and HC Blue 1). Other materials have produced lesser degrees of evi- dence ("some" or "equivocal" responses) or "clear ev- idence" isolated to an individual sex or species. A number of dyes have been prohibited in Europe on the basis of such oral studies or because of a close structural relationship to the tested dyes (Table 7).

Number of formulations

Type of study--species Permanent Semipermanent Reference

Subchronic 13 wk topical application--rabbits

Chronic 2 year dietary administration--dogs

Carcinogenicity 18 month topical application--mice Life-time topical application--mice Life-time topical application--mice Multigeneration topical application--rats

Teratology Oral gavage--rabbits Dietary administration-- rats Topical application-- rats

Mutagenicity Heritable translocation, topical--rats

9 3 Burnett et al. (1976)

1 Wernick et al. (1975)

1 Wernick et al. (1975) 1 Wernick et al. (1975)

9 3 Burnett et al. (1976)

1 1 Burnett et al. (1981)

3 m Burnett et al. (1975) 6 3 Burnett et al. (1980) 2 12 Jacobs et al. (1984) 7 3 Burnett et al. (1988)

Page 10: Cosmetic Toxi

908 Corbett et al.

F344/N rats B6C3F1 mice Source Overall

Dye Route call Male Female Male Female NTP TR, year

HC Blue 2 Gav NEG NE NE NE NE 293, 1985 HC Yellow 4 Feed NEG EE-PIT AD NE NE NE 419, 1992 HC Red 3 Gav NEG NE NE EE-LIV A D / C A IS 281, 1986 Acid Orange 3 Gav POS NE CE-KID TC CA NE NE 335, 1988 Disperse Blue 1 Feed POS CE-UBLAD CA CE-UBLAD CA EE-LIV A D / C A NE 299, 1986 HC Blue I c Gav POS EE-LW CA CE-LUNG CA CE-LW/THY C A / A D CE-LW CA 271, 1985

aSummary data can be found in compendium of abstracts from long-term cancer studies reported by The National Toxicology Program from 1976 to 1992 (Environ. Health Perspectives 101 (Supplement 1), April 1993). Overall calls were given in 1993 correspondence from NTP Position paper on "Hair dyes, NTP research findings" (January 22, 1993).

b Abbreviations: AD = ademoma; CA = carcinoma; CE = clear evidence; EE = equivocal evidence; IS = inadequate study; IT = pituitary gland; KID = kidney; LIV = liver; NEG = negative; NE = no evidence; POS = positive; SE = some evidence; TC = transitional cell; THY = thyroid gland; UBLAD = urinary bladder.

CNot in current use.

However, in some instances the positive findings in such oral studies are of unlikely relevance to human health, in view of mechanistic and exposure consid- erations, as well as in light of potential confounding influences that may have affected the study outcomes.

For example, urinary bladder tumors produced in rats by Disperse Blue 1 (squamous- and transitional- cell tumors, and leiomyosarcomas) were highly asso- ciated with urinary bladder calculi and were not found at the low dose in the three-dose level NTP bioassay. This was suggestive of a secondary mecha- nism for tumor induction. Quantitative risk assess- ments, assuming a threshold effect, indicated the safe level was 20-fold higher than the maximum average daily-applied dose associated with product use.

For HC Red No. 3, the NTP concluded that there was "equivocal evidence of carcinogenicity" in male B6C3F1 mice due to an increased incidence (35/50 mice) of combined liver tumors (adenomas and carci- nomas) in the high-dose group (250 m g / k g / d a y ) as compared to the control group (25/50 mice). How- ever, there was also a significant reduction in these tumors in the low-dose group (15/50 mice at 125 mg / kg/day). The possible importance of these findings is reduced by the known variability of hepatic cell neo- plasms in this strain. Moreover, a perfect dose-re- sponse (15/50; 25/50; 35/50) association was found between cage shelf heights and tumor incidence on reanalysis of these data (and other NTP bioassay re- sults) for "systematic" effects unrelated to chemical treatment.

For 2-amino-5-nitrophenol, the NTP call of "some evidence of carcinogenicity" for male rats was based on an increased incidence of acinar cell adenomas of

the pancreas. The dye was administered by gavage in corn oil. Subsequently, NTP found increases in the incidence of pancreatic exocrine hyperplasia and ade- noma with either corn oil, safflower oil, tricapyrilin alone, and with dichloromethane given in corn oil. NTP concluded that the use of corn oil as a gavage vehicle may have a confounding effect on the inter- pretation of chemical-induced proliferative lesions of the exocrine pancreas (as well as on mononuclear cell leukemias) in male F344/N rats.

Exposure is an important consideration with re- spect to risk assessment for hair dyes, which may display a carcinogenic hazard in rodent studies but are used on an infrequent basis and display a rela- tively low potential for percutaneous penetration. For 2,4-diaminoanisole, an oxidative hair dye that in- duced thyroid tumors in mice and rats, as well as tumors of the integumentary system in rats, exposures based on percutaneous penetration data obtained in vivo in rhesus monkeys (about 0.032% of the applied dose) and risks estimated by conservative mathemat- ical approaches (Mantel-Bryan and linear models) yielded cancer risks in the range of 6.1 • 10 -8 to 2.8 • 10 -11 for dark shades and 4.1 • 10 -11 to 4.9 • 10 -19 for light shades of hair color containing this material. Subsequent human data on 2,4-diaminoani- sole obtained under actual-use conditions showed a lower absorption of about 0.02% of the applied dose. Such risks are clearly in the range of being de minimus. In vivo human studies with other dye materials have shown percutaneous absorption values to generally less than 1% of the applied dose. For representative oxidative and direct hair dyes, this translated to ex- posures in the range of about 2 to 7 mg per applica-

Page 11: Cosmetic Toxi

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910 Corbett et al.

0-Phenylenediamine ~ 2,4-Diaminotoluen~ 2,4-Diaminoanisole ~ 2,4-Diaminophenetole b 2,4-Diaminophenylethanol b 2,5-Diaminoanisole b 2-Amino-4-nitrophenol a 2-Amino-5-nitrophenol a 4-Amino-2-nitrophenol a Catechol c Pyrogallol c

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oxide. Ammonium and potassium persulfates are very effective in this role. While the persulfates alone have no effect on the pigment granules, when used in admixture with ammonium hydroxide and hydrogen peroxide in the form of a paste, they are very effective in completely destroying the natural pigments. Appli- cation time varies from 20 minutes to more than 1 hour, depending on the original color of the hair.

Unfortunately, the strong oxidizing mixture also results in the oxidation of the hair keratin, particularly at the cystine moiety. Injudicious use of bleaches can thus result in brittle hair that requires careful han- dling and the extensive use of conditioners. This phe- nomenon is often referred to as "drying," which is a misnomer resulting from the feel of hair with a rough- ened cuticle. Indeed, damaged hair has a higher water content than virgin hair under the same ambient con- ditions.

The persulfates have been reported to be sensitiz- ers, but such reactions appear to be rare.

tion for relatively dark shades or about 1 - 4 / ~ g / k g / day, taking into account a 4-week use frequency.

Relevant to the extrapolation of risks to low-level human exposures from high-dose rodent tumorgenic- ity studies is an observation based on an evaluation of 139 NTP studies. It was found that the MTD/ 740,000 provides an esimate of the 10 -6 life-time risk, which was generally within a factor of 10 of that dose estimated by applying linear multistage models to the 2-year bioassay data. Thus, for materials exhibiting MTDs in the range of 1 g m / k g / d a y (106/~g/kg/day) in 2-year rodent bioassays, life-time average daily ex- posures of about 1 .4 /~g/kg/day (i.e., 106/(0.74 X 106) would generally be expected not to pose significant carcinogenic risk. Extended use of hair dyes contain- ing the materials would pose a risk of considerably less than the de min imus one in a million.

Hair Bleaches

Lightening of the natural hair color requires de- struction of the brown-black eumelanin and /o r the yellow to red phaeomelanin pigments that occur as granules in the cortex of the hair.

Hydrogen peroxide, particularly under alkaline conditions, is effective in disintegrating the eumelanin granules and in bleaching some of the dispersed pig- ment. However, the end result is a reddish brown that requires modulating with oxidation dyes if a natural shade is desired.

Complete decolorization of the natural pigment re- quires the use of "boosters" with the hydrogen per-

O T H E R C O S M E T I C P R O D U C T C A T E G O R I E S

It is beyond the scope of this chapter to detail the toxicological profiles and issues associated with indi- vidual cosmetic ingredients used in the wide variety of available product types. However, it is relevant to put the associated risks into an appropriate perspec- tive based on actual-use history and experience.

Accidental and Emergency-Related Human Exposures

In 1995, cosmetic and other personal-care products accounted for 8.5% (n = 171,426) of the human expo- sures reported to poison control centers in the United States, ranking third, below cleaning substances and analgesics. It is important to acknowledge that mate- rials with a high frequency of involvement may not necessarily be the most toxic but may be merely the most accessible, particularly to children. Indeed, it is reassuring that only a small proportion (9.5%) of such exposures involved treatment in a health-care facility, while that the majority resulted in no significant out- come (33.2%) or a minor outcome (21.4%) associated with minimally bothersome and generally resolvable signs and symptoms. Major effects were relatively in- frequent (n = 69) and largely associated with alcohol- containing mouthwashes. Deaths were associated with exceptional circumstances, such as intentional abuse of a hair spray (n = 1) or suicides involving mixed chemical exposures (n = 2).

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Cosmetic Toxicology 911

Other A dverse React ions

U.S. FDA Reports

Industry Data Until recently, the U.S. cosmetic industry reported

to the FDA the incidence of adverse reactions received via toll-free 800 telephone numbers or other corre- spondence, together with the numbers of units dis- tributed as part of the Voluntary Cosmetic Registra- tion Program. In turn, the FDA would calculate the number of such cosmetic experience reports (CERs) per million units sold, fit the data to a linear regres- sion model and report back to the individual manu- facturers which of their products had reaction rates that were more than one standard deviation above the calculated mean rate. It was then the responsibility of the manufacturer to determine the relevance, if any, of the reported observations and take any appropriate action. Representative rates (CERs per 106 units sold) for various product types are shown for the years 1989-1992 in Table 8.

While such data provide a general idea of the pro- pensity of various cosmetic product types to produce undesired reactions in consumers, the lack of detailed information and standardized medical follow-up lim- its more meaningful analysis of the causative ingredi-

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Number of reactions a Product types

262

111 70-89 50-69

30-49

20-29

10-19

-<10

Skin moisturizers, cleansers, night creams, and other skin care

Hair dyes Shampoos; Nail products Permanent waves; eye products; hair

conditioners / sprays Suntan products; facial foundations; hair

straightenersb; deodorants; depilatories Fragrances; toothpastes and oral-hygiene

products; lipsticks; hair tonics Bath soaps and detergents; bath products;

baby products Hair bleaches and lighteners; other hair colors;

shaving products

a n - 1305. bExcludes 157 injury reports that were related to one partic-

ular product.

ent(s) and the relative severity or importance of the reported adverse experiences.

CER Product types (10 6 units)

20-30 Baby shampoo

10-19

5-9

<5

Other baby products Permanent waves Hair rinses

Noncoloring shampoos Deodorants Other skin-care

preparations Other suntan

preparations

Baby lotions, oils Bath oils, tablets Eyeliners Eye shadows Eye-makeup removers Powders All other categories

Bath soaps and detergents

Skin cleansers Skin moisturizers Night creams Indoor-tanning

preparations Hair sprays Hair straighteners Hair dyes and colors

Hair lighteners Hair bleaches Foundations Skin fresheners Suntan gels, creams

Consumer Complaints Reported Directly to the FDA The FDA continues to report cosmetics injuries re-

ported directly to them organized by the same prod- uct categories as the industry data. Reports provide details of the brand name, injury complaints (e.g., der- matitis, pain, tissue damage, and respiratory effects), and affected body part or area. A n analysis of the incidence of the complaints by cosmetic product type for the period July 1987 through 1995 is shown in Table 9. Skin-care products (e.g., moisturizers, cleans- ers, and night creams), hair dyes, shampoos, nail products, permanent waves, eye products, and hair sprays and conditioners showed the highest incidence of adverse reactions. A high incidence of reports as- sociated with hair straighteners for this period was primarily attributable to a single product and, while not representative of the product category in general, points to ability of the system to detect (and react to) problematical materials.

The availability of such data through Freedom of Information Act requests to the FDA affords individ- ual manufacturers the opportunity of examining the type and frequency of adverse reactions associated with their material compared to those for related or competitive products. It is likely that there is overlap

Page 14: Cosmetic Toxi

912 Corbett et al.

in these databases, in that those consumers reporting adverse reactions directly to the FDA would also in- form the manufacturer, particularly if medical costs were incurred.

Swedish MPA Reports

Adverse reactions to cosmetics and toiletries are reported to the Swedish Medical Products Agency (MPA) in a voluntary system introduced in 1989. In the period from 1989-1994, 191 cases involving 253 products were evaluated and causality was estab- lished by dermatologists using World Health Organi- zation (WHO) criteria for suspected adverse drug re- actions. The large majority of adverse reactions (90%) involved moisturizers, followed by hair-care and nail products, sunscreens and tanning products, eye makeup, and perfume. Most of the effects were lim- ited to the skin and eczematous reactions were often (7%) classified as contact allergic in nature, as con- firmed by patch-test reactions to the product or its individual components. The most frequent compo- nents implicated were fragrances (22/79), toluene sul- fonamide-formaldehyde resin (19/79), various preser- vatives (19/79), and glyceryl monothioglycolate (7/79).

Dermatological Reactions U.S. Experience

In a 5-year prospective study of cosmetic reactions conducted by the North American Contact Dermatitis Group, 713 cosmetic-related cases of contact dermati- tis were identified among 13,216 of their patients. Of these, 89% of the cosmetic reactions were confirmed by patch testing, with the remainder based on case history and medical examination. The majority of the reactions were related to skin-care products (28%), hair preparations including hair color (24%), and fa- cial makeup (11%), with lesser contributions associ- ated with other product types. Fragrance (usually un- specified) accounted for the majority of reactions, followed by preservatives, (Quaternium 15, imidazo- lidinyl urea, parabens, 2-bromo-2-nitropropane-l,3 diol, formaldehyde); p-phenylenediamine; lanolin and its derivatives; glyceryl monothioglycolate; propylene glycol; toluene sufonamide-formaldehyde resin; sun- screens and other UV absorbers; and acrylate or meth- acrylate. The group acknowledged that their data might not have been representative of the country at large because of the specialized interest of the derma- tologists involved.

Belgium Experience

The European experience with respect to dermato- logical reactions appears to be quite consistent. Among 3970 patients evaluated in a contact allergy unit from 1985-1990, 11.9% (462) represented cos- metic contact dermatitis. The majority of the reactions were associated with face- and body-care products (29%), followed by perfumes and toilet waters (22%), sun products (7%), and a variety of other cosmetic products with lower reaction incidences (1-5%). In patients who had positive patch tests, the most fre- quent allergens among ingredient types were fra- grance mix, oak moss, balsam of Peru, preservatives (e.g., [chloro] methylisothiazoline, formaldehyde, and parabens), vehicles and emulsifiers (e.g., wood alco- hol, cocamidopropylbetaine, and propylene glycol), active ingredients and hair dyes (e.g., p-toluene sul- fonamide formaldehyde resin, glyceryl, or ammo- nium thioglycolate, and paraphenylenediamine), and sunscreens (e.g., benzophenone-3, Eusolex 8021, and p-aminobenzoic acid).

C O S M E T I C I N G R E D I E N T S

Fragrances Fragrances are used widely in cosmetic products,

either to provide a direct end benefit as in the case of perfumes, toilet waters, and deodorants, or to increase the acceptability of other cosmetic product types by masking undesired odors or imparting a pleasant scent to enhance the consumer's product-use experi- ence.

The potential for fragrance materials to elicit aller- gic contact dermatitis and, in some cases, contact ur- ticaria of immunologic or nonimmunologic origin is well recognized and extensively studied (see the re- view by Scheinman, 1996). Self-regulatory initiatives by the Research Institute for Fragrance Materials (RIFM) and the International Fragrance Research As- sociation (IFRA) has led to the systematic review of fragrance components for toxicological, ecotoxicologi- cal, and sensitization potential. As a result, certain components have been prohibited for use or have had limits imposed based on their concentrations.

North American, European, and Japanese derma- tologists have routinely included either fragrance mixtures or a screening series of selected individual fragrance components in their diagnosis and surveil- lance of cosmetic allergic reactions. The use concentra- tion and use circumstances (i.e., leave on or rinse off), extent and location of product application, use fre- quency, integrity of the skin barrier, and underlying

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Cosmetic Toxicology 913

skin disease can all impact the propensity of potential sensitizers to elicit dermatological reactions. There- fore, after exclusion of known strong sensitizers, the routine safety evaluation for finished cosmetic prod- ucts invariably incorporates a human prophetic patch test to ascertain the potential of the material to induce an allergic response under exaggerated (e.g., occlusive patches) conditions.

Based on the chemical analysis of fragrance com- ponents from 17 common cosmetic products (8 leave- on and 9 wash-off types) in Denmark, from one to five constituents of the European fragrance mix mate- rials were present in each of the products. However, patch testing performed in 500 consecutive eczema patients revealed that not all cases of perfume allergy were detected by the mix, as reflected by positive concordance values of 52.4% and 81.3% for wash-off and leave-on products, respectively. Conversely, 70% of the subjects that were fragrance-mix positive did not react to the product perfumes. This emphasizes the importance of establishing the clinical relevance of observed patch-test reactions based on actual product testing.

In a worldwide multicenter investigation of fra- grance contact dermatitis involving the diagnostic patch testing of 67 subjects at seven centers, atopic individuals were over-represented as compared to the general population. The face and hands were the most common sites of eczematous dermatitis. Racial differ- ences included a higher likelihood of reactions to fra- grance mix in white persons and a higher prevalence of reactions to benzyl salicylate among Asians.

In addition to the clinically related issues associ- ated with fragrance materials, the chemical reactivity of certain fragrances may predispose them to more toxicologically significant findings in animal studies, such as reproductive and carcinogenic effects. These are discussed with regards to an important class of fragrance materials, the nitromusks and the polycyclic musks.

Nitromusks

The terms "musk" and "nitromusk" are nonchemi- cal classifications that are used in the perfume indus- try to describe substances that have a typical "musky" odor. There are currently four nitroaromatic com- pounds of interest to the fragrance industry, musk xylene, musk ketone, musk tibetine, and moskene. Musk xylene and musk ketone are the most important of these, while musk tibetine and moskene are used in much smaller amounts. These compounds have had a safe history of use as perfume ingredients for over 100 years and there are no reports of adverse

effects in humans. A fifth nitromusk, musk ambrette, was found to cause neurotoxicity and testicular atro- phy in rats via both oral and topical routes. In addi- tion, musk ambrette was also found to be phototoxic in a few individuals. As a result, musk ambrette is banned from all fragrances and was placed on Annex II of the European Cosmetics Directive in 1995. The RIFM has evaluated the other nitromusks and con- cluded that they did not produce the same toxicity as musk ambrette.

In studies for potential mutagenicity and carcino- genicity, musk xylene was found to produce hepato- cellular carcinomas and adenomas when fed to mice for 2 years. Musk xylene was subsequently catego- rized as a nongenotoxic carcinogen causing hepatic tumors in mice by mechanisms similar to those ob- served for the barbiturate phenobarbital, and thus it is not considered to represent a carcinogenic risk for humans. Through mechanistic studies, musk ketone and musk tibetine were shown to be metabolized dif- ferently and hence unlikely to show the same toxicity as musk xylene. As a result of the musk xylene obser- vations, the RIFM proceeded to evaluate more exten- sively the genotoxic potential of the nitromusks. Musk xylene was tested in the mouse lymphoma assay, in an in vitro cytogenetics assay in Chinese hamster ovary (CHO) cells, in an in vitro UDS assay, and in an in vivo unscheduled DNA synthesis (UDS) assay. When these were combined with the negative Ames assay performed earlier, the RIFM concluded that musk xylene showed no evidence of genotoxic poten- tial.

The RIFM also evaluated the genotoxicity of musk ketone in a battery of short-term tests and found no evidence of genotoxic potential based on results from the mouse lymphoma, in vitro cytogenetics, and in vitro UDS assays. In addition, musk ketone was found to be negative in an in vivo mouse micronucleus assay.

Recent studies indicate that trace amounts of nitro- musks have been detected in mother's milk and in environmental samples. Although nitromusks have been the subject of an extensive testing program ex- amining both human health and environmental safety, due to these relatively recent findings the SCCNFP has requested specific toxicological data from the industry on these compounds in order to evaluate their significance for human health. Because these four ingredients are, in fact, four different chem- ical substances with different toxicological profiles, each substance is being evaluated separately by the SCCNFP.

The RIFM has undertaken research to provide ad- ditional data to help in the risk assessment. Studies have been conducted to determine the effects of musk

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914 Corbett et al.

ketone and musk xylene on developmental toxicity. These studies demonstrated that developmental tox- icity was not a biologically significant factor in the risk assessment for musk ketone or musk xylene when assessed in relation to expected human expo- sure through the use of cosmetics.

Using dermal-absorption figures obtained from rat studies and NOELs (no observed effect levels) deter- mined from subchronic dermal studies also in rats, large safety factors can be calculated for musk xylene and musk ketone, 2000 and 3000 respectively, under simulated conditions of human use. Large safety fac- tors have also been demonstrated for indirect human exposure through the environment.

The SCCNFP is currently evaluating the toxicolog- ical data submitted by industry on musk ketone and musk xylene. For the other two nitromusks, musk tibetine and moskene, the SCCNFP considered the available data to be insufficient to enable a safety eval- uation and classified these as 2b and not to be used in cosmetic products. The SCCNFP has indicated that there are no adverse findings or identified health haz- ards on musk tibetine and moskene and is prepared to reassess its opinion subject to additional data sub- missions.

Polycyclic M u s k s

These ingredients have been widely used in fra- grance formulations for a wide variety of consumer products, including perfumes, soaps, shampoos, and detergents, for over 40 years. Extensive safety dossiers have also been submitted to the SCCNFP for both HHCB (1,3,4,6,7,8-hexahydro-4,6,6,7,8,8,-hexamethyl- cyclopenta-y-2-benzopyran) and AHTN (6-acetyl- 1,1,2,4,4,7-hexamethyltetraline). As in the case of the nitromusks the SCCNFP will review an extensive bat- tery of mutagenicity assays, subchronic studies, skin absorption, and teratology and developmental toxic- ity studies investigating exposure via maternal milk.

As part of the environmental risk assessment for AHTN, a chronic fish study was conducted on fathead minnows. An absent tail fin was observed in the ma- jority of fish at the lowest observed-effect concentra- tion level of 67/~g. All fish at the no-observed-effect concentration level of 35/~g/liter and below had nor- mal tail fins. This finding was reported to the U.S. EPA (Environmental Protection Agency); however, it is not considered to be of environmental concern or to be relevant to human health. The SCCNFP is yet to complete its evaluation of HHCB and AHTN.

Preservatives As with fragrances, cosmetic use experience has

demonstrated a relatively high propensity for allergic

reactions to this widely used ingredient category. Ex- amples of potential contact sensitizers used in cosmet- ics have included formaldehyde, paraben esters, sorbic acid, isothiazoles, and organic mercurial com- pounds. Skin irritation is also a potential limiting fac- tor with preservatives, and cosmetic-formulation goals strive to achieve adequate microbial preserva- tion using the lowest feasible preservative levels. Hu- man repeat-insult patch tests and cumulative irrita- tion studies on prototype or finished cosmetic products can usefully serve in identifying such poten- tial issues prior to marketing.

In a study among 19,546 unselected dermatitis pa- tients in Italy, positive patch-test incidences of 1.9% with parabens and 0.3% with formaldehyde were re- ported. In subsets of this population, comprising from 980 to 14,897 patients, incidences ranged between 0.1% (Quaternium 15) to 1.6% (chloromethylisothia- zolone and methylisothiazolone). Except for sodium metabisulfite and chloracetamide, with incidences of 1.4% and 1.0%, respectively, all of the other preserva- tives tested (triclosan, butylated hydroxyanisole, imi- dazolidinyl urea, bronopol, and butylated hydroxyto- luene) showed reaction rates of less than 0.3%. Temporal analysis showed that the incidence of con- tact allergy to parabens declined from about 2.5-2.7% in the period between 1968 and 1982 to about 1.0- 1.3% in the period between 1983 and 1991. Paraben allergy appeared to be associated more with local remedies applied to diseased or compromised skin than with cosmetic products per se. For parabens, the hands and face were considered to be moderate and low risk areas, respectively, while for chloromethyli- sothiazolone and methylisothiazolone, the hands and face were the predominant cutaneous sites of contact allergy.

As is the case for fragrances, the presence of positive patch-test results for preservatives may not be indicative of intolerance to the material under use conditions. For example, 24/27 subjects with positive patch-test reactions to 100 ppm chloromethylisothiazolone and methylisothiazolone tolerated a 2-week use period with a shampoo formulated at 15 ppm. The authors emphasize that it still may be prudent to suggest alternative products to patients with known sensitivities. They also point to the paucity of controlled clinical trials evaluating positive patch-test allergens under use conditions.

In the European Union, cosmetic preservatives are systematically reviewed and the maximum autho- rized concentrations and other limitations and re- quirements are listed in Annex VI, Part 1 (allowed) and Part 2 (provisionally allowed).

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Cosmetic Toxicology 915

Surfactants Surfactants are another broadly used cosmetic in-

gredient class employed at varied concentrations in diverse product types, such as shampoos, body washes, and other types of skin cleansing products, and at relatively lower concentrations as emulsifiers and wetting agents in many other product categories. The toxicology of surfactants used in cosmetics has been reviewed recently by Sterzel (1997). Their bio- chemical properties include interactions with mem- branes involving adsorption to surfaces and altera- tions in membrane permeability and, at higher concentrations, cell lysis and solubilization of integral membrane proteins. Cationic and anionic surfactants also interact with proteins, with consequent swelling of the stratum corneum, and may also inactivate en- zymes due to conformational changes or binding to active enzyme sites.

The skin compatibility of anionic surfactants ap- pears to vary by chain length and, for sodium soaps, alkyl sulfates, and alkylbenzene or ~-olefin sulfonates, compounds with a saturated side chain of 10-12 car- bon atoms are usually the least compatible. It is gen- erally acknowledged that for mucous membranes, the rank order of surfactant compatibility is nonionic anionic > cationic. While surfactants as a class have a concentration-dependent potential of inducing skin and mucous membrane irritation, they are not gener- ally regarded as having a significant sensitization lia- bility because of their inability to act as haptens by forming strong bonds to endogenous proteins.

Because of their relatively low potential for percu- taneous absorption under normal cosmetic use condi- tions, the likelihood of systemic adverse effects from surfactants are low, and feeding or drinking water toxicology studies with these materials have generally shown substantial safety margins.

Other Ingredients In addition to the fragrances, preservatives, and

surfactants that are used widely in a variety of cos- metic products, there is a diversity of other product and ingredient categories with more specialized toxi- cological issues, which presents a significant challenge to summarize adequately. Some examples include UV absorbers with potential for photoxicity, photosensiti- zation and, possibly, photomutagenicity; hair sprays; nail-polish solvents; powdered materials with inhala- tion potential; lipstick components for which oral ex- posure may be an expected exposure route; and nail- polish resins with sensitization issues. Nater and DeGroot (1988) have compiled comprehensive lists of

representative cosmetic formulations, their ingredi- ents, and their reported side effects, including contact allergy, urticaria, irritant dermatitis, and systemic ef- fects such as respiratory symptoms associated with an immediate-type allergy. Included are recommended skin patch-test concentrations and vehicles for diag- nostic patch testing. The reader is referred to this text for detailed information.

EXPOSURE A N D RISK A S S E S S M E N T C O N S I D E R A T I O N S FOR C O S M E T I C

P R O D U C T S A N D I N G R E D I E N T S

The toxicological evaluation of cosmetic ingredi- ents serves usefully to identify potential hazards, which then need to be put into appropriate perspec- tive via a risk assessment that incorporates an esti- mation of exposure and a calculation of the margin of safety by relating the human exposure dose to the no- observed-adverse-effect level obtained from animal tests. Key exposure parameters include the amount of product and the concentration of ingredient applied, the route and site of application, and the frequency and duration (e.g., rinse off or leave on) of applica- tion. Although, there are no good quantitative data that adequately reflect the variations in consumer habits and practices, particularly for several of the newer cosmetic types and forms, some conventions that are used for cosmetic exposure assessment are given in Table 10. Because demographics and cultural differences would be expected to have a significant influence, particularly on use frequency or on amounts, there may be considerable variations in the actual exposures, which often may be remarkably overestimated using conservative exposure assump- tions.

While the intended route of exposure for most cos- metic products, by definition, is topical, inadvertent exposure may also occur via the mucous membranes, orally, or via inhalation for aerosol or powdered cos- metic forms. Inhalation may also be a potential expo- sure route for ingredients, including contaminants, that may be volatilized under use conditions. In some cases, such as for hair cosmetics, skin (scalp) exposure is an inadvertent but unavoidable consequence of product use. While it is important to consider the relative contribution of various exposure routes to an ingredient on a case-by-case basis, oral and inhalation exposures would generally be considered to poten- tially engender higher risks than would topical appli- cation.

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916 Corbett et al.

Amount Exposure type Use frequency (gm/application) Product types

Rinse off

Leave on

Mucous membrane contact

3-6/day 0.8 Soap bar 1/day 2.0 Shave cream 1-7/week 12.0 Shampoo 1-4/week 10.0 Shower gel 1-3/week 14.0 Hair conditioner 1-2/week 17.0 Foam bath (undiluted) 8-18/year 30.0 Semipermanent hair dye 8-12/year 50.0 Permanent hair dye (ready-to-use) 2/day 0.80 Face cream 1-5/day 0.75 Toilet water 1- 3/day 3.0 Antiperspirant/deodorant (spray) 1 - 3 / day 0.5 Antiperspirant / deodorant (others) 1-2/day 1 (mg/cm 2) General-purpose cream 1-2/day 1.2 After shave 1-2/day 2.5 Make-up remover 1-2/day 2.5 Talcum powder 1-2/day 7.5 Body lotion 1-2/day 10.0 Hair spray 2- 3/week 0.25 Nail product 1-2/week 12.0 Setting product 1-2/week 12.0 Temporary hair dye 2-3 day for 2 wk/year 8.0 Sun cream

1-6/day 0.01 Lipstick 1- 3 / day 12.0 Mouthwash (ready-to-use) 1-3/day 0.01 Eye make-up: powder 1-2/day 0.5 Eye make-up remover (wipe-off) 1 - 2 / day 1.5 Toothpaste 1/day 0.025 Eye make-up: mascara 1/day 0.005 Eye make-up: liner

aAdapted from ECETOC (1993).

I N G R E D I E N T , T O X I C O L O G Y A N D P R O D U C T S A F E T Y

While the toxicologic potential of cosmetic ingredi- ents, some of which have a broader use in other con- sumer, industrial, and pharmaceutical products or in food applications, has traditionally been characterized in conventional animal studies such as those listed earlier in this chapter, the European Union sixth amendmen t to the Cosmetics Directive has brought increased emphasis on the use of nonanimal s tudy alternatives for both ingredients and finished prod- ucts. The use of selected individual in vitro tests or test batteries shows good promise for cosmetic mate- rials in certain areas, such as skin irritation, percuta- neous absorption, and photoirri tation or photosensi- tization. While considerable progress has been made in unders tanding their utility and limitations, there is less of a consensus on alternative methods for testing eye irritation and, particularly, skin sensitization. It is

generally acknowledged that for the evaluation of sys- temic effects, such as studies of chronic toxicity, repro- ductive toxicity, and immunotoxic potential as cur- rently required or requested by international cosmetic regulations and guidelines, there appear to be no near-term alternatives to animal testing.

In many circumstances, finished product testing may be accomplished without or with minimal use of animals by a stepwise approach involving the thor- ough evaluation of the available toxicological data on the ingredients and consideration of prior experience with related ingredients and formulations ("paper" toxicology); the conservative application of controlled clinical studies with appropria te ethical considera- tions and close medical supervision; and the use of in vitro methods that have been validated for a specific purpose with the appropria te known "benchmark" materials. This scheme, incorporating the option of conducting limited animal tests under selected cir- cumstances to resolve outs tanding issues or, alterna-

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Cosmetic Toxicology 917

"Paper"Toxicology

Assessment

Clinical Tests , ,

Assessment

Consumer Use Tests

Assessment

Marketing

Post-Market Survey

IN VITRO BATTERY

LIMITED ANIMAL TESTS ] . . . . . .

~ Y

> Data-based Confidence

Question/Issue

Concern

! ~ 4 ~ i ~ i ~ ~i~!~,'!!~ ~ i l ~ ' ~ ; ~ ~ i ~ ~ - ' ~ ~X~,~i~j~ '~, '~;~,~ '~. '~:~;~:~: ' ,~~i~i .... ~ ~ ' , ' . ! ~ ~.~;~~@~/~a~i~, 9 .~!~ ~`~;~:~;~`~.~;~;~.:~`~`.~``r

tively, to suspend development, is shown in Fig. 3. An important element in such a process is the inclu- sion of feedback loops to address issues that may develop during the various stages of product devel- opment, including the exposure of a broader numbers of individuals under less controlled circumstances, such as consumer-use tests or test marketing. Post- market surveillance is an important element of cos- metic product safety in that low-frequency events may only be ascertained after broad consumer use.

The viability of maintaining the option to include animal tests of finished products, even on a selected basis, remains to be determined by future interna- tional regulatory activities and by the policy and prac- tices of individual cosmetic companies.

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