Establishment and intra‐/inter‐laboratory validation of a standard protocol of reactive oxygen species assay for chemical photosafety evaluation

Abstract

A reactive oxygen species (ROS) assay was previously developed for photosafety evaluation of pharmaceuticals, and the present multi-center study aimed to establish and validate a standard protocol for ROS assay. In three participating laboratories, two standards and 42 coded chemicals, including 23 phototoxins and 19 nonphototoxic drugs/chemicals, were assessed by the ROS assay according to the standardized protocol. Most phototoxins tended to generate singlet oxygen and/or superoxide under UV-vis exposure, but nonphototoxic chemicals were less photoreactive. In the ROS assay on quinine (200 µm), a typical phototoxic drug, the intra- and inter-day precisions (coefficient of variation; CV) were found to be 1.5-7.4% and 1.7-9.3%, respectively. The inter-laboratory CV for quinine averaged 15.4% for singlet oxygen and 17.0% for superoxide. The ROS assay on 42 coded chemicals (200 µm) provided no false negative predictions upon previously defined criteria as compared with the in vitro/in vivo phototoxicity, although several false positives appeared. Outcomes from the validation study were indicative of satisfactory transferability, intra- and inter-laboratory variability, and predictive capacity of the ROS assay.

Establishment of an <i>in silico</i> phototoxicity prediction method by combining descriptors related to photo-absorption and photo-reaction

“…ROS assay can detect ROS generations both of results of photo-reactions and photo-decomposition of chemicals, but phototoxicity was related to the ROS generation in the result photo-reactions (Onoue et al, 2008(Onoue et al, , 2012. Therefore, the false positive of ROS assay to 3T3 NRU-PT might be related to not photo-reaction but photo-decomposition.…”

“…Therefore, the false positive of ROS assay to 3T3 NRU-PT might be related to not photo-reaction but photo-decomposition. We removed photo-decomposable compounds from the dataset for ROS assay prediction which were corresponded with 3T3 NRU-PT (true positive and negative compounds) including 33 compounds (22 positives and 11 negatives) (Onoue et al, 2012). The dataset for ROS assay prediction included some kinds of compound structures such as amino acid, tetracyclines, non-steroidal anti-inflammatory drugs, polycyclic-aromatic hydrocarbons, camphor derivatives and artificial pigments.…”

Photostability evaluation of five UV-filters, trans-resveratrol and beta-carotene in sunscreens

“…This study was based on ROS Assay Protocol Version 1.0 (JaCVAM, 2013a) recommended by the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) and the Japanese Center for the Validation of Alternative Methods (JaCVAM), which was based on previous validation studies conducted by JaCVAM (JaCVAM, 2013b(JaCVAM, , 2013cOnoue et al, 2013).…”

“…Results such as these are of increasing concern to the scientific community involved in the study of active ingredients to be used in sunscreen products. Some assays employing probes to indirectly evaluate ROS quenching ability as well as their generation are also being reported (Onoue et al, 2014(Onoue et al, , 2013.…”

Phototoxic Risk Assessments on Benzophenone Derivatives: Photobiochemical Assessments and Dermal Cassette-Dosing Pharmacokinetic Study

“…For the detection of both singlet oxygen and superoxide generation from irradiated compounds, the ROS assay was carried out according to the validated protocol with minor modification (Onoue et al, 2013(Onoue et al, , 2014b). Briefly, each tested compound was dissolved in DMSO at 10 mM for stock solution.…”