Palladium and platinum ions interfere with the measurement of erythrocyte vesiculation by inhibiting the acetylcholinesterase activity of the released spectrin-depleted microvesicles

Palladium (Pd(2+)) and platinum (Pt(2+)) ions were found to inhibit erythrocyte membrane-bound acetylcholinesterase (AChE) with Ki values of 6.0 and 6.5 microg/ml, respectively. Lineweaver-Burke plots revealed that the inhibition of erythrocyte AChE by both metal ions was competitive in nature. Binding studies using alkaline phosphatase as a reporting enzyme confirmed that both metal ions indeed did bind to the enzyme molecules. In the process of red cell vesiculation, membrane-bound AChE is shed along with vesicles. The measurement of AChE activities in the medium containing vesiculated RBC could potentially be served as an index of vesiculation. Inhibition of AChE activities by both metal ions can thus constitute a potential source of error in vesiculation measurement. To illustrate these effects, a simulated vesiculation system, using green tea polyphenol in the presence (25 microg/ml) or absence of Pd(2+) ion was simultaneously examined by the electronmicrography and the AChE method. We observed vesiculation under the experimental condition in Pd(2+)-free controls that was associated with a time-dependent increase in AChE activity were barely detected in the Pd(2+)-spiked specimen because of the masking effect exerted by the metal ions themselves.