Photochemical degradation of selected pharmaceuticals under light conditions relevant to natural waters and study of toxicity of photoproducts

M. Poncarová1, Š. Klementová1, M. Šorf2

1Faculty of Science, University of South Bohemia, Branišovská 1760, 370 05 České Budějovice

2Department of Agronomy, MENDEL University, ÚZRHV AF, Zemědělská 1, 613 00 Brno

poncam00@prf.jcu.cz


Xenobiotics in the environment include a wide variety of compounds, e.g. pesticides, drugs, textile dyes, personal care products, stabilisers, and many others. Among xenobiotics, pharmaceuticals have recently acquired increasing attention [1, 2]. Pharmacological products enter natural waters mainly via wastewater either from manufacturing facilities or from municipal wastewater (excretion of unmetabolised drugs, disposal of unused drugs). The contamination of natural aquatic systems results in adverse negative effects on aquatic organisms.

In surface waters, physical, chemical, and biological processes contribute to the transformations of polluting substances. Photoinitiated processes may represent important degradation pathways in surface waters for compounds resistant to both biological degradation and chemical reactions such
as hydrolysis
[3]. Photochemical degradation may lead to a decrease in contaminant concentration, and, in some cases, generate photoproducts with even higher harmful effects than that of the parent compound [4].

Ecotoxicology represents a framework enabling to test a given compound and to reveal or at least estimate its potential harmful effect. This study is focused on aquatic organisms. In autotrophs, algae Chlorella sp. and Desmodesmus sp. are often used due to their simple laboratory maintenance [5, 6].
The flagship of heterotrophs toxicity testing in surface water is the planktonic microcrustacean Daphnia magna
[7]. It has several characteristics that in toxicological tests, especially those targeted at acute toxicity estimation – it can be relatively easily maintained in the laboratory and, when under suitable conditions, D. magna reproduces parthenogenetically. A common model of vertebrates in ecotoxicology is the zebrafish Danio rerio. Although the extrapolation of the obtained results to higher vertebrates is not straightforward and should be done with care, the response of fish to xenobiotics is a significant indicator of how a particular compound (or products of its photodegradation) affect fish assemblages in surface waters.

In this study, toxicity of atorvastatin, a widely prescribed hypolipidemic drug, and the mixture of its photoproducts were investigated. The photoproduct mixture was produced by irradiation of the solution of atorvastatin (c = 50 mg/l) by the radiation in the range between 300 – 350 nm (to imitate the
short-wavelength solar radiation that reaches the Earth´s surface) for 15 minutes. Then, two toxicity assays based on OECD 202
[8] and 236 [9] guidelines were performed.

In the case of acute toxicity test on the model organism Daphnia magna (OECD 202), 2 juveniles not older than 24 h were introduced into 5 ml of pure media, other juveniles in pairs in the media with atorvastatin in the concentration range from 1 to 10 000 µg/l; photoproducts solutions were tested
at concentration range
of remaining atorvastatin in the irradiated solution up to 1000 µg/l. During this test (48 hrs) constant temperature was held at 18.7 ± 0.2 °C; photoperiod was 16 hrs light and 6 hrs dark and the juvenils were not fed. Atorvastatin did not cause any mortality, photoproducts caused
20 % mortality at the highest concentration used. The LC50 value could not be evaluated from this experiment
. The data show that lethal concentration for 50 % of daphnids is higher than the highest used concentration. Photoproducts seem to be more toxic than atorvastatin itself since in addition to
the observed mortality each daphind showed odd swimming at the highest concentration of photoproducts.

Toxicity assay based on OECD 236 guideline was done on embryos of Danio rerio. One fertilized egg was introduced into 2 ml of ISO water (control) or into 2 ml of atorvastatin or 2 ml of photoproducts (concentration ranges as in the tests with D.magna). Tested embryos were kept in the incubator Climacell EVO line, for 96 hours at the temperature 25 – 26 °C and the photoperiod 14 hrs light/ 10 hrs darkness. The evolution of the embryos was monitored visually every 24 hours. Four key parameters indicating the lethality were sought for: coagulated embryos, lack of somite formation, non – detachment of the tail and lack of heartbeat. The value of LC50 for atorvastatin was determined by software Prism 6, its value is 1976 µg/l. Regarding photoproducts, 40 % mortality was observed at the photoproduct mixture
with remaining 500 
µg/l of atorvastatin. In the higher concentration there was also retardation in
the development and the drug had adverse effect on blood formation – lack of cell flow, transparent cells without pigmentation.

1. M. Mezzelani, S. Gorbi, F. Regoli, Marine Environmental Research, 140, (2018), 41.

2. M. de Oliviera, B. E. F. Frihling, J. Velasques et al., Sci. Tot. Env., 705, (2020), 135568.

3. A. Nikolaou, S. Meric, S. Fatta, Anal. Bioanal. Chem., 387, (2007), 1225.

4. Š. Klementová, M. Poncarová, D. Kahoun, M. Šorf, E. Dokoupilová, P. Fojtíková, Environ. Sci. Pollut. Res., 27, (2020), 35650.

5. OECD, Guidelines for testing chemicals 201, Freshwater Alga and Cyanobacteria, Growth Inhibition Test (2011).

6. E. Geiger, R. Hornek-Gausterer, M. T. Sacan, Ecotox. Environ. Safe, 129, (2016), 189.

7. J. Seďa & A. Petrusek, Journal of Limnology, 70, (2011), 337.

8. OECD, Guidelines for the testing of chemicals 202, Daphnia sp., Acute Immobilisation Test (2004).

9. OECD, Guidelines for testing of chemicals 236, Fish Embryo Acute Toxicity (FET) Test (2013).