Lack of significant dermal penetration of titanium dioxide from sunscreen formulations containing nano- and submicron-size TiO2 particles

Abstract

Titanium dioxide (TiO(2)) is included in some sunscreen formulations to physically block ultraviolet radiation. A dermal penetration study was conducted in minipigs with three TiO(2) particles (uncoated submicron sized, uncoated nano-sized, and dimethicone/methicone copolymer-coated nanosized) applied 5% by weight in a sunscreen. These and control formulations were topically applied to minipigs at 2 mg cream/cm(2) skin (4 applications/day, 5 days/week, 4 weeks). Skin (multiple sites), lymph nodes, liver, spleen, and kidneys were removed, and the TiO(2) content was determined (as titanium) using inductively coupled plasma mass spectroscopy. Titanium levels in lymph nodes and liver from treated animals were not increased over the values in control animals. The epidermis from minipigs treated with sunscreens containing TiO(2) showed elevated titanium. Increased titanium was detected in abdominal and neck dermis of minipigs treated with uncoated and coated nanoscale TiO(2). Using electron microscopy-energy dispersive x-ray analysis, all three types of TiO(2) particles were found in the stratum corneum and upper follicular lumens in all treated skin samples (more particles visible with coated nanoscale TiO(2)). Isolated titanium particles were also present at various locations in the dermis of animals treated with all three types of TiO(2)-containing sunscreens; however, there was no pattern of distribution or pathology suggesting the particles could be the result of contamination. At most, the few isolated particles represent a tiny fraction of the total amount of applied TiO(2). These findings indicate that there is no significant penetration of TiO(2) nanoparticles through the intact normal epidermis.

FIG. 1.

Levels of titanium in left, right, and combined (Comb) inguinal, prescapular, and submandibular lymph nodes taken from minipigs treated with sunscreen formulations evaluated via ICP-MS: control, no TiO₂ (open bar); submicron-sized TiO₂ (gray bar); uncoated nanosized TiO₂ (hashed bar); and coated nanosized TiO₂ (filled bar). The “all combined” group represents the average of all lymph nodes for animals in the indicated treatment groups. Data are presented as mean ± SE for each group (three minipigs per group). *statistically significant difference from the control value for each group (p ≤ 0.05).

FIG. 2.

Levels of titanium in left, right, and middle lobes of the liver taken from minipigs treated with sunscreen formulations containing the following TiO₂ (from left to right): control, no TiO₂ (open bar); submicron (grey bar); uncoated nano (hashed bar); and coated nano (filled bar). The “combined” group represents the average value for liver samples for each animal in the indicated treatment group. Data are presented as mean ± SE for three minipigs per group.

FIG. 3.

Level of titanium in epidermis isolated from skin taken from minipigs treated with sunscreen formulations containing the following TiO₂ (from left to right): control, no TiO₂ (open bar); submicron (grey bar); uncoated nano (hashed bar); and coated nano (filled bar). The skin was taken from the abdomen, right and left axillary (axil) areas, neck, and back, and the dermis separated from the epidermis with a dry heat method. The “combined” group represents the average value for all epidermal samples for each animal. Data are presented as mean ± SE for three minipigs per group. *statistical significance (p ≤ 0.05) from the control values for each grouping.

FIG. 4.

Level of titanium in dermis isolated from skin taken from minipigs treated with sunscreen formulations containing the following TiO₂ (from left to right): control, no TiO₂ (open bar); submicron (grey bar); uncoated nano (hashed bar); and coated nano (filled bar). The skin was taken from the abdomen, right and left axillary (axil) and inguinal (Ing) areas, neck, and back, and the dermis separated from the epidermis with a dry heat method. The “combined” group represents the average value for all epidermal samples for each animal. Data are presented as mean ± SE for three minipigs per group. *statistical significance (p ≤ 0.05) from the control values for each grouping.

FIG. 5.

Photomicrograph of a section of minipig skin before (left panel) and after (right panel) separation of the epidermis from the dermis using the dry heat method. Sections were stained with hematoxylin and eosin. Fragments of the epidermis that were not removed with the dry heat method are identified in right panel.

FIG. 6.

Representative TEM micrographs and corresponding SEM-EDX spectra (below micrograph) of minipig skin stratum corneum. (upper left) Control (no TiO₂); (upper right) Sub-micron-sized TiO₂; (lower left) Uncoated nanosized TiO₂; (lower right) Coated nanosized TiO₂. The areas of stratum corneum of skin were surveyed and EDX spectra were acquired at the circled area (lower right micrograph, dotted circle) in the TEM micrographs, respectively. The acquisition time for the EDX spectrum associated with lower right image is much shorter than the acquisition times for the other EDX spectra, reflecting the higher density of particles in the coated nano TiO₂ treated sample. A higher magnification TEM image of the circled area in lower right panel is shown in Figure 8.

FIG. 7.

Representative TEM micrographs of minipig skin in areas containing hair follicles. (left column of photomicrographs) Sub-micron-sized TiO₂; (middle column of photomicrographs) Uncoated nano-sized TiO₂; (right column of photomicrographs) Coated nano-sized TiO₂ in hair follicle areas. The images in the lower photomicrographs are higher magnification images of the corresponding upper panels.

FIG. 8.

×15,000 magnification TEM image of tissue in Figure 6d (minipig skin treated with coated nanosized TiO₂). Hundreds of particles are visible in this small region of stratum corneum tissue, illustrating the higher density of particles in the coated nano-TiO₂–treated skin samples.

FIG. 9.

Representation of TiO₂ particle distribution in different layers of minipig abdominal skin (longitudinal slice of skin) exposed to submicron TiO₂ showing the results from the cross section analysis of each skin layer. Numbers in parentheses are estimates of the numbers of TiO₂ particles observed in each layer. Abbreviations are: SC, stratum corneum; SS, stratum spinosum; SG, stratum granulosum; and SB, stratum basale.