On-Chip Detection of Hydrogen Peroxide with Bimetallic Nanoparticles in Electrochemical Microfluidic Sensor
Authors: Ko Euna , Tran Van-Khue , Geng Yangfang , Kim Min Ki , Jin Ga Hyun , Seong Gi Hun 
Published: February 2020

A simple, accurate and inexpensive quantitative determination of hydrogen peroxide (H2O2) is an active area of research due to potential biological, industrial, and environmental applications. Biologically, H2O2 plays a significant role as a signaling and regulating molecule within cells and between cells and H2O2 is a reactive oxygen species (ROS) that is a product of metabolism in the human body, resulting in oxidative stress that may damage DNA and contribute to cancer. Accordingly, an efficient detection method for simple, exact, and expeditious sensing of H2O2 has been sought.
Microfluidic platforms provide a convenient and immediate sensing to detect specific biomolecules. Making use of properties of microfluidics, such as small sample volume, low cost, fast sample analysis, and improved reaction reliability and reproducibility, numerous biosensors combined with microfluidics have been developed to improve the overall performance of the sensing system. Particularly, electrochemical biosensors integrated in microfluidics have been widely applied for on-chip and real–time monitoring systems due to their inexpensive analysis, simple manipulations, high sensitivity and selectivity, and ease of portability for miniaturized healthcare sensors. To improve the sensitivity of the electrochemical sensors for quantification of H2O2, the modification of noble metal nanoparticles (NPs) onto the electrode could be used to decrease overpotential and increase electron transfer rate. Moreover, the electrochemical deposition of metal NPs into the microchannel can accurately control the position of nanostructures, maximize the stability of the electrode, and ensure that miniaturized electrochemical sensors with NPs are more specific and highly sensitive for biomolecular diagnostics.
We developed microfluidic devices for real-time detection of H2O2 with the electrochemically deposited Au covered AgNPs (Au-AgNPs) modified on single-walled carbon nanotube (SWCNT) electrodes (Figure. A & B). Decoration of Au-AgNPs on the SWCNT working electrode was performed by electrochemical deposition of AgNPs and galvanic replacement of AuNPs that results in an increase in Au seed formation on AgNP surfaces (Figure. C). The size of the Au seed increase with applied electrical potential (10 μC at -0.4 V) and thus the increased roughness of the nanoparticles associated with newly created pits augments active surface energy and reactivity. As shown in Figure. D, the Au-AgNPs were electrochemically deposited on the SWCNT working electrode and had very good adhesion and stability after washing.
In the electrochemical analysis by the microfluidic device, 1 mM H2O2 in 0.1 M PBS buffer (pH 7.2) was investigated using on-chip CVs measurement on Au-AgNPs modified SWCNT electrodes. The larger reduction current toward H2O2 comes from the synergistic enhancement of the electro-catalytic activity between Au and Ag. AgNPs were used to improve the reduction of H2O2, and AuNPs could support the enhanced catalytic reaction of AgNPs toward H2O2 detection, resulting from the effective electron transfer ability of Au. Under optimized conditions, the cyclic voltammograms of electro-reduction at various concentrations of H2O2 were measured in the microfluidic device (Figure. E). The reduction currents at -0.4 V increased gradually with higher concentrations of H2O2. The calibration plot indicates good linearity for the reduction current versus H2O2 concentrations in the range from 0.3 mM to 1.8 mM with a correlation coefficient of r2 = 0.985. The lowest concentration of H2O2 that could be estimated by this microfluidic electrochemical sensor was 26.8 μM (S/N = 3). The long-term stability studies of the device filled with 0.1 M PBS buffer during 7 days exhibited only a small deviation with a relative standard deviation (RSD) of 2.01%. The reproducibility of this sensor showed that RSD was 0.42–3.69% for 24 different sensors in 1 mM H2O2 solution, indicating the electrodes can be reproduced reliably. For real sample analysis, commercial antiseptic solutions containing H2O2 were analyzed to evaluate the performance of the sensors (Table). The recovery values ranged from 97% to 107%, and RSDs were below 5%. In view of the results achieved, these microfluidic sensors combined with electrochemical detection could contribute to the growth of biosensing technologies as an advance in microfluidic applications.

Figure. (A) An optical image of the microfluidic device integrated with microelectrodes. (B) a schematic image of on-chip device. (C) A microscope image of SWCNT electrodes deposited with Au-AgNPs. (D) FE-SEM image of Au-AgNPs on a working electrode. Inset: image of a single Au-Ag nanoparticle. (E) CV responses of H2O2 reduction over different concentrations in a microfluidic sensor at a flow rate of 1 μL·min-1. Inset: calibration curve of responses for H2O2 reduction current toward concentration at -0.4 V.

Table. H2O2 determination in pharmaceutical samples using the electrochemical microfluidic devices.

Figure 1