An in Vitro Comparison of the Effects of Tetrabromobisphenol A and Tetrabromobisphenol S on Threshold-Like Hemolysis in Human Erythrocytes

BACKGROUND/AIMS: The use of tetrabromobisphenol A (TBBPA) as a brominated flame retardant has raised growing concerns of its potential toxicity due to its widespread use and persistence. The present study compares the threshold-like mechanism of hemolysis induced by TBBPA with that of its sulfone analog, tetrabromobisphenol S (TBBPS), using an in vitro model of human erythrocytes. METHODS: Isolated red blood cells (5% hematocrit) obtained from healthy donors were exposed to TBBPA (10-50 µg/mL) or TBBPS (10-100 µg/mL) for 24 hours at 37°C. Hemolysis, methemoglobin formation, lipid peroxidation, glutathione levels, ATPase activity, membrane protein integrity, and interactions with human serum albumin were evaluated using spectrophotometric, fluorometric, flow cytometric, and electrophoretic (SDS-PAGE) analyses. Interactions of both compounds with human serum albumin (HSA) were investigated by fluorescence quenching and fluorescence lifetime measurements. RESULTS: TBBPA induced a distinct threshold-like hemolytic response, with hemolysis increasing sharply above 12.5 µg/mL and reaching approximately 43.7% at 30 µg/mL, whereas TBBPS did not induce hemolysis within the tested concentration range. TBBPA-induced hemolysis was preceded by increased MetHb formation and accompanied by depletion of glutathione, impaired ATPase activity, and alterations in membrane proteins, including aggregation and loss of cytoskeletal components, but not by detectable lipid peroxidation. TBBPS produced substantially weaker effects on erythrocyte membrane integrity and antioxidant status. Fluorescence studies demonstrated that both TBBPs interacted with HSA, with predominantly static quenching accompanied by a contribution from dynamic quenching. The two compounds exhibited similar effects on HSA fluorescence under the experimental conditions used. CONCLUSION: TBBPA and TBBPS exhibit markedly different effects on erythrocytes despite their structural similarity, with TBBPA showing a distinct threshold-like hemolytic response associated with protein and antioxidant system damage rather than lipid peroxidation. The results suggest that differences in the chemical structure of TBBPs influence their interactions with proteins and may contribute to their distinct biological effects.