Phototoxic substances

Some essential oils, such as Citrus bergamia essential oil, contain photoactive molecules that can be activated under light exposure, and consequently these oils have potential phototoxicity [53]. The mechanism of phototoxicity differs from that of cytotoxicity because phototoxic essential oils penetrate the cell without destroying membranes or biomolecules, and radical reactions only occur when the cells are exposed to light, resulting in production of phototoxic substances. However, cytotoxicity and phototoxicity are both dependent on the molecules in the essential oils and their compartmentalization in cells, with production of different types of radicals with or without light exposure [10]. 

The test is based on an in vitro assay of the uptake of the dye, neutral red (NR), in Balb/c 3T3 fibroblasts. It was developed to detect the phototoxicity induced by the combined interaction of the test substance and light of the wavelength range from 315 to 400 nm, the so-called UVA. The cytotoxicity is evaluated in the presence (+UVA) or absence (-UVA) of UVA light exposure, after application of a nontoxic dose of the compound. The cytotoxicological impact is assessed via the inhibition of the fibroblasts to take up the vital dye NR (NR is a weak cationic dye, penetrating easily into the cell membrane by a nonionic diffusion and accumulates in the lysosomes) one day after the initial treatment. Normally, healthy cells may incorporate and bind NR. Alterations of the cell surface or the lysosomal membranes, however, lead to a decreased uptake and binding of the dye. 

A further option to investigate the phototoxic potential of substances is the use of reconstructed human skin models. The evaluation of the cell viability is based on the MTT-assay that is sensitive for the mitochondria activity in cells. Currently, these in vivo substitutes are still under validation and are not approved as full standard test methods for the investigation of the phototoxicity potency of a test chemical. However, several existing models are in use for prevalidation studies and are described elsewhere in more detail [92],... 

F. Netzlaff, C.-M. Lehr, P. W. Wertz, and U. F. Schaefer. The human epidermis models EpiSkin , SkinEthic and EpiDerm An evaluation of morphology and their suitability for testing phototoxicity, irritancy, corrosivity, and substance transport. Eur. J. Pharm. Biopharm. 60 167-178 (2005). 

As mentioned in the introduction, the main process leading to phototoxicity is the production of ROS such as 02 or by a photoactivated substance after energy or electron transfer to oxygen. 

The red blood cell phototoxicity test (photo-RBC test) is based on the ability of a light-activated substance to produce lysis of freshly isolated erythrocytes and to... 

The phototoxicity test 3T3 NRU was proposed in 1994 and is so far the only in vitro method that has been validated by European regulatory authorities for predicting the photoirritant potential of substances [5,40,41]. In this test, the mouse fibroblasts cell line Balb/c 3T3 is exposed to simulated solar UV (or, more frequently, solar UVA) in the presence of the test compound after an incubation of 1 h in the dark. Evaluation of cytotoxicity is performed 24h post-exposure using the neutral red uptake (NRU) method. N RU permits to distinguish live and dead cells, since intact cells retain this dye (detailed method in INVITOX protocol 78). The validation was performed with substances selected on the basis of their in vivo photoirritant or phototoxic properties. Some of these structures are shown in Table 19.1. 

Analytical studies on the prediction of photosensitive/phototoxic potential of pharmaceutical substances. Pharmaceutical Research, 23, 156-164. 

Despite the hypothesis that the evolutionary significance of phototoxic secondary substances may be linked to their ability to discourage Insect herbivores, most research has been directed toward their effects on human skin and range animals (42). In an attempt to extend our knowledge of insect photosensitizers we have screened a number of plant secondary substances (TABLE TI) for their photosensitizing activity to 4th instar mosquito larvae Aedes atropalpus under solar simulating lamps. 

Furocoumarins, such as xanthotoxin (20), apparently increase the sensitivity of Ehrlich tumor cells to y-rays.80 The use of phototoxic furocoumarins as anticancer substances has been investigated, but does not seem to give satisfactory results.81... 

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