Human percutaneous absorption of a direct hair dye comparing in vitro and in vivo results: Implications for safety assessment and animal testing

Although in vitro skin absorption studies often detect small residues of applied test material in the epidermis/dermis, it is uncertain whether the residue is within the living skin. We studied the dermal absorption of a hair dye hydroxyanthraquinone–aminopropyl methyl morpholinium methosulphate (HAM) in human skin in vivo and in vitro. In vivo, skin (back and scalp) received 0.5% HAM in a commercial formulation at 20 μg/cm2 After 0.5 or 48 h, skin was tape stripped, followed by cyanoacrylate biopsies (CAB). Sebum from scalp sites was collected for 48 h. In vitro, skin was treated with 20 mg/cm2 dye for 0.5 h, penetration determined after 24 h. In vivo, at 0.5 h, total recovery (back) was 0.67 μg/cm2 (tape strips + CAB). Fluorescence microscopy showed HAM in the hair follicle openings (HFO). At 0.5 h, scalp tape strips contained 1.80 μg/cm2, HFO 0.82 μg/cm2. At 48 h, HFO contained 0.21 μg/cm2, sebum 0.80 μg/cm2. In vivo, skin residues were in the non-living skin and eliminated via desquamation and sebum secretion. In vitro, the SC contained 1.50 μg/cm2, epidermis/dermis 0.86 μg/cm2, receptor fluid < 0.04 μg/cm2, a total of 0.90 μg/cm2 was considered to be bioavailable. In vitro epidermis/dermis residues were nearly identical to those located in non-living skin in vivo. In conclusion, in vitro percutaneous penetration studies may produce seemingly bioavailable material , which raises the need for a Threshold of Skin Absorption (TSA) addressing a negligible dermal absorption in order to avoid unnecessary in vivo toxicity studies on substances that produce no significant human systemic exposure.