2.2. Electrochemical Impedance Spectroscopy System

The measurement technique used was two-electrode impedance spectroscopy (Figure 1a). This technique consists of applying a potential difference between two electrodes and measuring the current through them in order to find the electrode–sample–electrode impedance. The electrodes used are stainless steel needles. The material has a high corrosion resistance and is widely used in food contact applications [44]. In addition, the resistance of the electrodes is very small (~ 3 mΩ), in the order of about one million times smaller than the sample resistance, so the resistive part of measured impedance is practically the sample resistance. The electrodes were connected to two wires housed in an epoxy resin frame (Figure 1b).

Schemes of (a) the applied two-electrode measurement technique and (b) the stainless-steel electrodes used in the assays.

The system consisted of an electronic equipment and a software application that ran on a PC (Figure 2). Sinusoidal alternating electrical signals with different frequencies were applied to the sample, and the current response for each one of the frequencies was measured. Then, the system calculated the impedance spectrum of the sample by means of the Discrete Fourier Transform (DFT) and displayed it on the screen. The parameters of the system (signal amplitude, frequency range, current scale, etc.) were configured by the user through the graphical user interface. The EIS system was divided into two clearly differentiated parts.

Electrochemical impedance spectroscopy system.

A software application ran on a PC. It carried out a frequency sweep to obtain the impedance modulus and the phase of the sample. The user established the frequency range and the amplitude of the sinusoidal signals applied to the electrodes. For each one of the frequencies, the application calculated the signal temporal evolution and sent this information (along with the rest of the data needed to generate the signal) to the electronic equipment through a USB port. Then, with the data response of the electronic equipment, the software application determined the amplitude and the phase of the voltage and current signals through a DFT. From these data, the software application calculated the modulus and phase of the sample impedance for the current frequency. Then, the application stored the result of the measurement in a file and repeated the same process for the rest of the frequencies. The specifications of the EIS measurement system are shown in Table 1.

Specifications of the electrochemical impedance spectroscopy (EIS) measurement system.

The electronic equipment received the information sent by the computer, generated the corresponding sinusoidal waveform and applied it to the sample. Then, the current and voltage signal responses were sampled and sent to the PC. For the receiving process of the data sent by the PC and the signal generation, the equipment used a complex programmable logic device (CPLD, Altera EPM7160SLC84), a 10-bit digital analogue converter (DAC) and a static 2 KB (2048 bytes) random access memory (RAM). A second CPLD, two 8-bit digital analogue converter and a configurable current sensor sampled the signals corresponding to the voltage applied to the electrode and the current flowing through it. The samples were stored in others two static RAM memories. Once a complete cycle of the signal was sampled, the values are transmitted to the PC.

The electronic measurement system was designed by the Group of Electronic Development and Printed Sensors (GED and PS) of the Interuniversity Institute for Molecular Recognition and Technological Development (IDM) at the Universitat Politècnica de València (UPV) [45].