Kinetic Rotating Droplet Electrochemistry: A Simple
and Versatile Method for Reaction Progress Kinetic Analysis in Microliter
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Abstract
Here, we demonstrate a new generic,
affordable, simple, versatile,
sensitive, and easy-to-implement electrochemical kinetic method for
monitoring, in real time, the progress of a chemical or biological
reaction in a microdrop of a few tens of microliters, with a kinetic
time resolution of ca. 1 s. The methodology is based on a fast injection
and mixing of a reactant solution (1–10 μL) in a reaction
droplet (15–50 μL) rapidly rotated over the surface of
a nonmoving working electrode and on the recording of the ensuing
transient faradaic current associated with the transformation of one
of the components. Rapid rotation of the droplet was ensured mechanically
by a rotating rod brought in contact atop the droplet. This simple
setup makes it possible to mix reactants efficiently and rotate the
droplet at a high spin rate, hence generating a well-defined hydrodynamic
steady-state convection layer at the underlying stationary electrode.
The features afforded by this new kinetic method were investigated
for three different reaction schemes: (i) the chemical oxidative deprotection
of a boronic ester by H<sub>2</sub>O<sub>2</sub>, (ii) a biomolecular
binding recognition between a small target and an aptamer, and (iii)
the inhibition of the redox-mediated catalytic cycle of horseradish
peroxidase (HRP) by its substrate H<sub>2</sub>O<sub>2</sub>. For
the small target/aptamer binding reaction, the kinetic and thermodynamic
parameters were recovered from rational analysis of the kinetic plots,
whereas for the HRP catalytic/inhibition reaction, the experimental
amperometric kinetic plots were reproduced from numerical simulations.
From the best fits of simulations to the experimental data, the kinetics
rate constants primarily associated with the inactivation/reactivation
pathways of the enzyme were retrieved. The ability to perform kinetics
in microliter-size samples makes this methodology particularly attractive
for reactions involving low-abundance or expensive reagents