Naslov
Effect-directed analysis reveals inhibition of zebrafish uptake transporter Oatp1d1 by caulerpenyne, a major secondary metabolite from the invasive marine alga Caulerpa taxifolia

Autori
Marić, Petra ; Ahel, Marijan ; Senta, Ivan ; Terzić, Senka ; Mikac, Iva ; Žuljević, Ante ; Smital, Tvrtko

Izvornik
Chemosphere (0045-6535) 174 (2017); 643-654

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
Caulerpa taxifolia ; secondary metabolites ; cellular detoxification mechanism ; uptake transporters ; Oatp1d1 ; Oct1 ; zebrafish ; caulerpenyne ; effect-directed analysis (EDA)

Sažetak
Caulerpa taxifolia is a tropical marine algae that generates secondary metabolites that contribute to its invasive behavior. Some of these metabolites are toxic or repulsive substances that may represent a significant threat to the environment and human health. Currently there is no knowledge about the interaction of these compounds with the uptake membrane transporters that are crucial mediators of the phase 0 cellular detoxification in aquatic organisms. In this study we investigated possible inhibitory effects of secondary metabolites from C. taxifolia on the activity of specific uptake membrane transporters in zebrafish (Danio rerio). We have focused on two uptake transporters that transport organic anions (Oatp1d1) and cations (Oct1), respectively. Both transporters, cloned previously from zebrafish liver, are transiently transfected and overexpressed in HEK293 cells. Transport activity assays using lucifer yellow (LY) and 4-(4- (dimethylamino)styryl)-N-methylpyridinium iodide (ASP+) as a model substrate were applied for determination of Oatp1d1 and Oct1 interactors, respectively. A two-step Effect-Directed Analysis (EDA) procedure based on the advanced fractionation, followed by liquid chromatography coupled to high-resolution mass spectrometry (LC- MS) detection was applied for the separation and identification of compounds. Using the described methodology we were able to identify caulerpenyne (CYN) as the major metabolite in C. taxifolia and reveal its inhibitory effect on the transport activity of zebrafish Oatp1d1. The observed effect was confirmed by testing CYN purified from C. taxifolia, resulting in the determined CYN IC50 of 17.97 µM. A weak interaction of pure CYN was also determined with zebrafish Oct1 transporter. By following Michaelis-Menten kinetics we identified CYN as a non-competitive inhibitor of zebrafish Oatp1d1 transporter. In conclusion, in this study we described a novel mechanism of toxic action of C. taxifolia, showed that CYN is a potent non- competitive inhibitor of zebrafish Oatp1d1, and demonstrated that EDA procedure can be reliably used for both characterization of environmentally relevant complex biological samples and potential identification of new biologically active substances.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

POVEZANOST RADA