Abstract
.We describe the contribution of our in vivo multiphoton microscopy (MPM) studies over the last ten years with DermaInspect® (JenLab, Germany), a CE-certified medical tomograph based on detection of fluorescent biomolecules, to the assessment of possible penetration of nanoparticulate zinc oxide in sunscreen through human skin. At the time we started our work, there was a strong movement for the precautionary principle to be applied to the use of nanoparticles in consumer products due to a lack of knowledge. The combined application of different MPM modalities, including spectral imaging, fluorescence lifetime imaging, second harmonic fluorescence generation, and phosphorescence microscopy, has provided overwhelming evidence that nanoparticle zinc oxide particles do not penetrate human skin when applied to various skin types with a range of methods of topical sunscreen application. MPM has also been used to study the viable epidermal morphology and redox state in supporting the safe use of topical zinc oxide nanoparticles. The impact of this work is emphasized by the recent proposed rule by the United States FDA on Sunscreen Drug Products for Over-the-Counter Human Use, which listed only zinc oxide and titanium dioxide of the currently marketed products to be generally recognized as safe and effective.
Highlights
Researchers at the forefront of advances in technology have always had the flow-on benefits of being able to apply that technology in addressing previously unresolvable key concerns in biology and other disciplines. These findings may, in turn, stimulate the wave of technological advances or refinements, resulting in further research and development in resolving particular ill-defined issues of interest. One such example of a vital juxtaposition is the role played by the commercial development of multiphoton microscopy (MPM) for human studies in the assessment and, the regulatory approval of the nano-sized and transparent zinc oxide sunscreens, which we present here as a case study
We describe the imaging of zinc oxide nanoparticles (ZnO NPs) via different methods:[9,10,11,12,13,14] (1) fluorescence imaging using spectral filters, (2) fluorescence lifetime imaging microscopy (FLIM) analysis of fluorescence lifetimes τ1 and τ2, (3) second harmonic generation (SHG) and hyper-Rayleigh scatter (HRS), (4) multiphoton excited photoluminescence (MEP) defined by the FLIM fluorescence lifetime amplitude a1%, and (5) photoluminescence offset
It is important to recognize that the spectral imaging method, as we introduced it, has limitations in sensitivity and selectivity at low ZnO NP concentrations when the overlapping signal from autofluorescence is overwhelming, especially from keratin, which spectrally overlaps with the ZnO NP
Summary
Researchers at the forefront of advances in technology have always had the flow-on benefits of being able to apply that technology in addressing previously unresolvable key concerns in biology and other disciplines. These findings may, in turn, stimulate the wave of technological advances or refinements, resulting in further research and development in resolving particular ill-defined issues of interest One such example of a vital juxtaposition is the role played by the commercial development of multiphoton microscopy (MPM) for human studies in the assessment and, the regulatory approval of the nano-sized and transparent zinc oxide sunscreens, which we present here as a case study. We emphasize that this is only one of many examples of how this commercial MPM has been used. We describe our use of MPM to assess possible morphological changes in the morphology and redox state of the viable epidermis in the assessment of the possible direct viable epidermal toxicity of nanoparticulate zinc oxide
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