The hidden toxicity of solid phase extraction blanks in water analysis

Nontoxic blank samples are a prerequisite in effect-directed analysis to ensure that any measured bioassay activity stems from chemicals in the environmental sample, and not from chemicals added through the sampling and sample preparation procedures. In a study on wastewater, solid phase extraction (SPE) blank samples, prepared by extracting clean water (such as tap water, demineralized water, etc.) using the same methodology as real wastewater, showed toxic effects in algae, daphnia, and in vitro bioassays. The aim of this study was to pinpoint the origin of the chemicals responsible for the observed toxicity, and to provide guidance on how to remediate their occurrence. Analysis revealed that the elution solvents optimized for SPE, composed of methanol (MeOH), NH4OH, and formic acid, constituted a significant source of toxicity, even when evaporated to dryness. Ammonium was the primary source of toxicity in the algae assay. Formic acid released toxic metal ions from the system and SPE materials, particularly Cu and Zn, which strongly affected daphnia. Switching to pure MeOH as the elution solvent alleviated most of the toxicity, although some metals remained. Further reductions in toxicity were achieved by replacing ultra-pure water with an ion-rich artificial freshwater medium for preparing blank samples. Three tested SPE materials (graphitized carbon, hydrophilic-lipophilic balance, and ENV+) released unidentified toxic chemicals correlating with algae toxicity and estrogen receptor activity, but these could be eliminated through extensive MeOH washing before packing the cartridges. Nontoxic levels of plasticizers were detected from system components. In conclusion, altering SPE elution solvents, washing SPE materials, and using artificial freshwater as blanks minimized the release of chemicals and ensured nontoxic blank samples.