Faculty of Food Technology and Biotechnology, University of Zagreb
Abstract
The presence of endocrine-disrupting compounds in wastewater, surface water, groundwater and even drinking water has become a major concern worldwide, since they negatively affect wildlife and humans. Therefore, these substances should be effectively removed from effluents before they are discharged into surface water to prevent pollution of groundwater, which can be a source of drinking water. Furthermore, an efficient control of endocrine-disrupting compounds in wastewater based on biological and analytical techniques is required. In this study, a yeast estrogen screen (YES) bioassay has been introduced and optimized with the aim to assess potential estrogenic activity of waters. First, assay duration, concentration of added substrate to the assay medium and wavelength used to measure the absorbance of the substrate were estimated. Several compounds, such as 17-β-estradiol, 17-α-ethinylestradiol, bisphenol A, nonylphenol, genisteine, hydrocortisone, dieldrin, atrazine, methoxychlor, testosterone and progesterone were used to verify its specificity and sensitivity. The optimized YES assay was sensitive and responded specifically to the selected estrogenic and nonestrogenic compounds in aqueous samples. Potential estrogenicity of influent and effluent samples of two wastewater treatment plants was assessed after the samples had been concentrated by solid-phase extraction (SPE) procedure using Oasis® HLB cartridges and methanol as eluting solvent. Up to 90 % of relative estrogenic activity was detected in concentrated samples of influents to wastewater treatment plants and estrogenic activity was still present in the concentrated effluent samples. We found that the introduced YES assay is a suitable screening tool for monitoring the potential estrogenicity of effluents that are discharged into surface water.Prisutnost spojeva koji imaju estrogeni učinak u otpadnim, površinskim, podzemnim, a čak i pitkim vodama sve više zabrinjava stručnjake diljem svijeta zbog negativnog učinka na zdravlje ljudi i životinja. Stoga takve spojeve treba odstraniti iz otpadnih voda prije njihovog ispuštanja u površinske vode, te na taj način spriječiti zagađivanje podzemnih voda, što se koriste kao izvor pitke vode. Kemijskom analizom i biološkim testovima potrebno je učinkovito pratiti prisutnost spojeva s estrogenim učinkom u otpadnim vodama. U ovom smo radu razvili i optimirali metodu ispitivanja estrogenog učinka spojeva pomoću kvasca (test YES) radi procjene potencijalne estrogene aktivnosti u vodama. Optimirali smo trajanje testa, koncentraciju supstrata dodanog podlozi i valnu duljinu za mjerenje apsorbancije razgrađenog supstrata. Specifičnost i osjetljivost testa YES provjerili smo pomoću sljedećih spojeva: 17-β-estradiol, 17-α-etinilestradiol, bisfenol A, nonilfenol, genistein, hidrokortizon, dieldrin, atrazin, metoksiklor, testosteron i progesteron. Optimiranim testom YES mogu se u uzorcima voda detektirati odabrani spojevi s estrogenim učinkom ili bez njega. Moguća estrogenost pritoka i istoka vode iz dva postrojenja za obradu otpadnih voda određena je prethodnom ekstrakcijom uzoraka vode na čvrstoj fazi (SPE ekstrakcija) pomoću kolone Oasis® HLB i metanola kao otapala. U koncentriranim je uzorcima pritoka u postrojenjima određeno 90 % relativne estrogene aktivnosti. Isto tako je estrogena aktivnost pronađena i u koncentriranim uzorcima istoka iz postrojenja. Iz dobivenih se rezultata može zaključiti da je test YES prikladan za ispitivanje estrogene aktivnosti otpadnih voda, što naknadno dospijevaju u površinske vode
