Ultra-sensitive conductometric detection of heavy metals based on inhibition of alkaline phosphatase activity from Arthrospira platensis.
Résumé
This study is based on the conductometric measurement of alkaline phosphatase activity (APA) from the cyanobacterium, Arthrospira platensis, called Spirulina. Cyanobacterium cells were directly immobilized, by physical adsorption, on the ceramic part of gold interdigitated transducers. This activity was inhibited in the presence of heavy metals and a variation of the local conductivity was measured after addition of the substrate. The Michaelis-Menten constant (Km) was evaluated to be 0.75 mM through a calibration curve of the substrate, disodium 4-nitrophenylphosphate p-nitrophenyl phosphate (pNPP). Inhibition of APA was observed with cadmium and mercury with a detection limit of 10(-20) M. The half maximal inhibitory concentration (IC50) was determined at 10(-19) M for Cd(2+) and 10(-17) M for Hg(2+), and the binding affinity of heavy metal (Ki) was equal to the IC50. On the sensor surface, scanning electron microscopy (SEM) images revealed a remarkable evolution of the cyanobacterium's external surface that was attributable to the first defense mechanism against toxic heavy metals in trace. This effect was also confirmed through the important increase of response time τ(90%) recorded for APA response towards the substrate pNPP after cell exposure to metallic cations. Lifetime of the Spirulina-based biosensor was estimated to be more than 25 days.