現代人使用染髮劑愈趨普遍來改變外表，增加生活的品質。在歐美國家，最受歡迎的染髮劑為氧化性染髮劑。由於染髮劑的普遍，及接觸於人類皮膚，其成份必須是安全的。染髮劑由許多成分組成，包括芳香胺類成分，某些芳香胺成分已確認為致癌物。雖在動物試驗及流病學研究中，目前證據不足以判斷染髮劑之使用與癌症發生率的相關性，但某些研究結果仍是不樂觀的。由於人類皮膚對於某些外用物質仍具穿透性，因此評估染髮劑的經皮吸收是有必要的，歐盟即建議可以體外試驗作為評估模式。評估染髮劑的經頭皮吸收有其特殊的考量，包括頭髮具相當大的吸收表面積、頭皮毛髮密度大及皮脂含量高等。
本研究目的為建立高效液相層析方法，分離及定量染髮劑裡的多種成分，並應用於分析自製模擬處方及市售產品。接著評估不同皮膚部位(背部及腹部)、皮膚模式(人皮、豬皮及鼠皮)、處方溶劑(乳劑及溶液)、毛髮(保留毛髮及除去毛髮之豬皮)、暴露皮膚時間(30分鐘、60分鐘)、皮膚障壁功能(膠帶撕取角質層皮膚及正常皮膚)及皮脂分泌(添加人類皮脂及正常皮膚)對於含對苯二胺或對氨基酚氧化性染髮劑經皮吸收之影響。
研究結果首先建立以高效液相層析定量分析方法，可同時分析染髮劑多種成分，為快速方便、高選擇性且具相當程度之專一性的方法，並可應用於分析模擬處方。應用於含三種染髮成分之市售品時，其定量回收率可達91%以上。
以取皮刀切割之豬皮作為染髮劑之體外皮膚模式，研究結果顯示：含PPD和PAP氧化性染髮劑之豬背皮膚的經皮吸收分別為豬腹部皮膚之1.48和1.24倍；含PPD染髮劑溶液劑型的經皮吸收量為乳劑劑型的3.27倍；染髮劑暴露時間由30分鐘延長至60分鐘使其經皮吸收量分別增為3.47與3.03倍。破壞皮膚障壁功能使電阻值自10.84 ± 0.67 × 103Ω降為7.30 ± 0.29 × 103Ω時，PPD與PAP之經皮吸收量分別增至2.06與1.37倍；而添加皮脂於皮膚上1小時使得其經皮吸收分別增至1.31~1.93與1.15~1.93倍；至於保留毛髮於背部皮膚上，則PPD與PAP之經皮吸收量分別略減為0.92與0.89倍。以上各組比較之差異，除了毛髮之影響及部分皮脂之影響組別外，均具有統計上之顯著意義。PPD與PAP染髮劑的經皮吸收量分別在人皮>鼠皮>豬皮及鼠皮≧人皮>豬皮。建議未來可嚐試以分離表皮之豬背皮作為評估染髮劑經人類頭皮吸收之體外皮膚模式。

Personal use of hair dyes is getting more popular to improve people’s appearance. Oxidative hair dyes are the most important group and the largest market share in EU or the US. Taking into account the extent and frequency of human contact with hair coloring products, their ingredients must be safe. Hair dyes contain a variety of chemical agents, such as aromatic amines, some of which are considered carcinogens. Although there is inadequate evidence (toxicological and epidemiological studies) that personal exposures to hair dyes are carcinogenic, some results appear less favorable. The recognition that human skin is not an impermeable barrier for some topically applied substances initiated the investigation of percutaneous absorption of hair dyes and their ingredients. In vitro methodologies are recommended for ethical reason and feasibility in EU. Percutaneous absorption of hair dyes from scalp under actual-use conditions requires some special considerations. For example, hairs themselves provide a large surface for dye permeation. There are also high follicular density and sebum content in the scalp.
The purpose of this study was to develop a HPLC method which enables the identification and quantification of the dye ingredients in simulated and commercial oxidative hair dyes in the first place. Secondly, the percutaneous absorption of PPD or PAP containing oxidative hair dyes was evaluated in vitro. The effects of skin sites (abdominal and back skin), skin models (human, pig and mouse skin), formulation types (cream and solution), presence of hairs (hair-clipped skin or non hair- clipped skin), exposure time (30 min and 60 min), barrier function (tape-stripped skin and intact skin), and sebum supplement (sebum-treated skin and normal skin) were investigated.
We have developed a fast, selective and specific HPLC method which enabled simultaneous separation of six ingredients of oxidative hair dyes. We have also successfully applied the method to content analysis of one commercial product containing three dye components with recovery > 91%. In vitro studies using dermatomed pig skin demonstrated that the percutaneous absorption of PPD- and PAP- containing oxidative hair dyes through back skin was 1.48 and 1.24 fold of abdominal skin, respectively. Solution formulation delivered 3.27 fold amount of PPD as compared to cream formulation. When exposure time was increased from 30 to 60 min, 3.47 and 3.30 fold of increase in the absorption were found with PPD and PAP creams, respectively. When the transdermal electrical resistance of pig skin was decreased from 10.84 ± 0.67 ×103 Ω to 7.30 ± 0.29 ×103 Ω by 3 tape stripping, the absorption of two hair dyes were increased by 2.06 and 1.37 fold. Sebum treatment of the skin for 1 hour resulted in 1.31-1.93 and 1.15-1.93 fold percutaneous absorption of these hair dyes than the control skin. Absorption of two hair dyes through non-hair-clipped skin slightly decreased to 0.92 and 0.89 fold of hair-clipped skin. All the above changes were statistically significant (p < 0.05) with the exception of the hair and sebum effects. Finally, the percutaneous absorption of PPD- and PAP- containing hair dyes was in the order of human cadaver skin > mouse skin > pig skin and mouse skin ≧ human cadaver skin > pig skin, respectively. It was suggested that the isolated pig epidermis from back skin may be appropriate in vitro model for the evaluation of the percutaneous absorption of hair dyes in human scalps in the future.

Abrams K, Harvell JD, et al. Effect of organic solvents on in vitro human skin water barrier function. J Invest Dermatol 101(4): 609-13.1993.
Andrew AS, Schned AR, et al. Bladder cancer risk and personal hair dye use. Int J Cancer 109(4): 581-6.2004.
Aungst BJ. Structure/effect studies of fatty acid isomers as skin penetration enhancers and skin irritants. Pharm Res 6(3): 244-7.1989.
Barry BW. Novel mechanisms and devices to enable successful transdermal drug delivery. Eur J Pharm Sci 14(2): 101-14.2001.
Bi W, Hayes RB, et al. Mortality and incidence of bladder cancer in benzidine-exposed workers in China. Am J Ind Med 21(4): 481-9.1992.
Blume U, Verschoore M, et al. The vellus hair follicle in acne: hair growth and sebum excretion. Br J Dermatol 129(1): 23-7.1993.
Bommannan D, Potts RO, et al. Examination of stratum corneum barrier function in vivo by infrared spectroscopy. J Invest Dermatol 95(4): 403-8.1990.
Bronaugh RL and Maibach HI. Percutaneous absorption of nitroarmatic compounds: in vivo and in vitro studies in the human and monkey. J Invest Dermatol 84(3): 180-183.1985.
Budavari S. The merck Index-An Encyclopedia of chemicals, drugs and biologicals. Whitehouse Station, NJ, Merck and Co., Inc.1996.
Case RA, Hosker ME, et al. Tumours of the urinary bladder in workmen engaged in the manufacture and use of certain dyestuff intermediates in the British chemical industry. I. The role of aniline, benzidine, alpha-naphthylamine, and beta-naphthylamine. Br J Ind Med 11(2): 75-104.1954.
Cottingto EM KR, Tolgyesi WS. Observation of female scalp hair population: distribution and diameter. j Soc Cosmet Chem 28: 219-229.1977.
Diembeck W, Beck H, et al. Test guidelines for in vitro assessment of dermal absorption and percutaneous penetration of cosmetic ingredients. European Cosmetic, Toiletry and Perfumery Association. Food Chem Toxicol 37(2-3): 191-205.1999.
Downing DT, Strauss JS, et al. Variability in the chemical composition of human skin surface lipids. J Invest Dermatol 53(5): 322-7.1969.
Dressler WE. Percutaneous Absorption of Hair Dyes. In Dermal Absorption and Toxicity Assessment. Roberts MS and Walters KA MD. New York: 489-536.1998.
Gago-Dominguez M, Castelao JE, et al. Use of permanent hair dyes and bladder-cancer risk. Int J Cancer 91(4): 575-9.2001b.
Gago-Dominguez M, Chan KK, Ross RK, Yu MC. Permanent hair dyes and bladder cancer risk. Int J Cancer 94: 905-906.2001a.
Grodstein F, Hennekens CH, et al. A prospective study of permanent hair dye use and hematopoietic cancer. J Natl Cancer Inst 86(19): 1466-70.1994.
Haley TJ. Benzidine revisited: a review of the literature and problems associated with the use of benzidine and its congeners. Clin Toxicol 8(1): 13-42.1975.
Holly EA, Lele C, et al. Hair-color products and risk for non-Hodgkin's lymphoma: a population-based study in the San Francisco bay area. Am J Public Health 88(12): 1767-73.1998.
Howes D and Black JG. Percutaneous absorption of 2-nitro-p-phenylenediamine. Int J Cosmet Sci 5: 215-226.1983.
Hruby R. The absorption of p-toluenediamine by the skin of rats and dogs. Food Cosmet Toxicol 15(6): 595-9.1977.
Hueber-Becker F, Nohynek GJ, et al. Human systemic exposure to a [14C]-para-phenylenediamine-containing oxidative hair dye and correlation with in vitro percutaneous absorption in human or pig skin. Food Chem Toxicol 42(8): 1227-36.2004.
Kogevinas M, Marcos R, et al. Hairdye use and genetic cariation in relation to micronuclei in urothelial cells and bladder cancer risk in Spanish women. American Association for Cancer Research.2005.
La VC and Tavani A. Epidemiological evidence on hair dyes and the risk of cancer in humans. Eur J Cancer Prev 4(1): 31-43.1995.
Lyon, Ed. IARC monographs on the evaluation of carcinogenic risks to humans, International Agency for Research on Cancer (IARC).1993.
Nohynek GJ, Fautz R, et al. Toxicity and human health risk of hair dyes. Food Chem Toxicol 42(4): 517-43.2004.
O'goshi K, Iguchi M, et al. Functional analysis of the stratum corneum of scalp skin: studies in patients with alopecia areata and androgenetic alopecia. Arch Dermatol Res 292(12): 605-11.2000.
Ogiso T, Shiraki T, et al. Transfollicular drug delivery: penetration of drugs through human scalp skin and comparison of penetration between scalp and abdominal skins in vitro. J Drug Target 10(5): 369-78.2002.
Otberg N, Richter H, et al. Variations of hair follicle size and distribution in different body sites. J Invest Dermatol 122(1): 14-9.2004.
Pagnoni A, Kligman AM, et al. Determination of density of follicles on various regions of the face by cyanoacrylate biopsy: correlation with sebum output. Br J Dermatol 131(6): 862-5.1994.
Piacquadio D and Kligman A. The critical role of the vehicle to therapeutic efficacy and patient compliance. J Am Acad Dermatol 39(2 Pt 3): S67-73.1998.
Qiao GL, Chang SK, et al. Effects of anatomical site and occlusion on the percutaneous absorption and residue pattern of 2,6-[ring-14C]parathion in vivo in pigs. Toxicol Appl Pharmacol 122(1): 131-8.1993.
Rehn. Blasengeschwulste bei Fuchsinarbeitern. Arch Klin Chir 52: 588-600.1985.
Ritschel WA, Sabouni A, et al. Percutaneous absorption of coumarin, griseofulvin and propranolol across human scalp and abdominal skin. Meth Find Exp Clin Pharmacol 11(10): 643-646.1989.
Ross JH, Dong MH, et al. Conservatism in pesticide exposure assessment. Regul Toxicol Pharmacol 31(1): 53-8.2000.
SCCNFP/0129/99 F. Opinion concerning p-Phenylenediamine (Colipa n° A7) adopted by the SCCNFP during the 19th plenary meeting of 27 February 2002.
SCCNFP/0690/03 F. Notes of Guidance for Testing of Cosmetic Ingredients for Their Safety Evaluation, 5th revision, adopted by SCCNFP during the plenary meeting of 20 October 2003.
SCCP/0867/05. para-Aminophenol COLIPA n° A16. Adopted by the SCCP during the 3rd plenary meeting of 15 March 2005.
SCCP/0930/05. Personal Use of Hair Dyes and Cancer Hazard. Adopted by the SCCP during the 5th plenary meeting of 20 September 2005.
Schaefer H and TE R. Principles of Percutaneous Absorption. In Skin Barrier. London: 18.1996.
Scott RC, Corrigan MA, et al. The influence of skin structure on permeability: an intersite and interspecies comparison with hydrophilic penetrants. J Invest Dermatol 96(6): 921-5.1991.
Seago SV and Ebling FJ. The hair cycle on the human thigh and upper arm. Br J Dermatol 113(1): 9-16.1985.
Skoog DA and Leary JJ. Principles of Industrumental Analysis. New York.1992.
Stewart ME and Downing DT. Chemistry and function of mammalian sebaceous lipids. Adv Lipid Res 24: 263-301.1991.
Thun MJ, SF A, et al. Hair dye use and risk of fatl cancers in U.S. women. J Natl Cancer Inst 86: 941-944.1994.
Tsai JC, Hung PL, et al. Molecular weight dependence of polyethylene glycol penetration across acetone-disrupted permeability barrier. Arch Dermatol Res 293(6): 302-7.2001.
Verschueren K. Handbook of enviromental Data on organic Chemicals. 3rd ed. New York, NY: Van Nostrand Reinfold Co.1996.
Vincent U, Bordin G, et al. Validation of an analytical procedure for the determination of oxidative hair dyes in cosmetic formulations. J Cosmet Sci 53(1): 43-58.2002.
Wilhelm KP, Cua AB, et al. Skin aging. Effect on transepidermal water loss, stratum corneum hydration, skin surface pH, and casual sebum content. Arch Dermatol 127(12): 1806-9.1991.
Wolfram LJ and Maibach HI. Percutaneous penetration of hair dyes. Arch Dermatol Res 277(3): 235-41.1985.
Yalkowsky SH and Dannenfelser RM. The aquasol database of aqueous solubility Ver 5.1992.
Zenser TV, Lakshmi VM, et al. Metabolism of N-acetylbenzidine and initiation of bladder cancer. Mutat Res 506-507: 29-40.2002.
Zhang Y HT, Leaderer B, Boyle P, Zahm SH, Flynn S, Tallini G, Owens PH, Zhang T. Hair-coloring product use and risk of non-Hodgkin's lymphoma: a population-base case-control study in Connecticut. Am J Epidem 159: 148-154.2004.
林銘楷論文 年. 人體皮脂對藥物角質層滲透性之影響. 國立成功大學. 台南市.2004.