Micro-band Boron-doped Diamond Electrode in Capillary Electrophoresis for Simultaneous Detection of AMP, ADP, and ATP

A micro-band boron-doped diamond (BDD) electrode was prepared by sealing a piece of BDD film with an area of 1.11´10^-7 m² between two insulating plates, one Teflon and one silicon rubber, to form sandwich-like layers, so the surface area could be investigated. The micro-band BDD was combined with capillary zone electrophoresis as an electrode for the simultaneous detection of adenosine monophosphate (AMP), adenosine diphosphate (ADP), and adenosine triphosphate (ATP) in a solution. These adenosine phosphates can be separated with a 0.3 m-long fused silica capillary using Britton–Robinson buffers at pH 2.0. Current in the concentration range of 0.1 to 2.0 mM were linear with the limits of detection of 0.004 ?M, 0.006 ?M, and 0.011 ?M for AMP, ADP, and ATP, respectively. A comparison with an unmodified BDD as the detector in the same electrophoresis system showed that the micro-band generated better limits of detection (LODs) than the macroelectrode. This method was successfully applied to human urine samples injected with three adenosine phosphates, as well as adenine and guanine, which can be well-separated with recovery percentages of adenine, guanine, AMP, ADP, and ATP of 99.2%, 102.5%, 107.4%, 107.7%, and 105.4%, respectively.

Adenosine phosphates; Boron-doped diamond; Capillary zone electrophoresis; Electrochemical detection; Micro-band electrode

      Adenosine triphosphate (ATP) is an important small package of energy in a cell. Its hydrolysis reaction to adenosine diphosphate (ADP) and adenosine monophosphate (AMP) is sufficient to promote unfavorable processes required by the cell (Yadav et al., 2017). These adenosine phosphates (APs) are also important as extracellular signaling agents (Giuliani et al., 2019). As signaling agents, the alteration of APs can cause diseases such as epilepsy, Alzheimer’s disease, Parkinson’s disease, and stroke, and it can lead to drug abuse (Effendi et al., 2020). Moreover, the presence of these compounds in urine or blood plasma could be indicators of liver disease (Staufner et al., 2016; Wang et al., 2020). Therefore, detailed and sensitive detection of these compounds is highly important.

        Numerous conventional measurements have been reported on APs detectors using liquid chromatography (Zhu et al., 2017; Andries et al., 2018; Menegollo et al., 2019). However, this method is time-consuming and expensive, and it requires an expert to operate the instruments. Aptamer-based sensors also have been reported (Zhang et al., 2017; Zhou et al., 2020). However, they failed to distinguish ATP from ADP and AMP, as they show a similar affinity (Li and Liu, 2020). Other efforts used carbon fiber in electrophoresis for amperometric detection (Gunawardhana and Lunte, 2018) and electro-analytical detection using a pencil graphite electrode (Krishnan et al., 2020). Previously, electrochemical detection of adenosine phosphates with an unmodified boron-doped diamond (BDD) was successfully reported (Asai et al., 2016). BDD is known as an electrode that exhibits interesting electrochemical properties, such as low background current and wide potential window. These are excellent characteristics of sensor applications (Wahyuni et al., 2015; Hayat et al., 2019). The wide potential window in aqueous media has also been reported and proven to suppress the evolution of H₂ in applications of CO₂ reduction (Natsui et al., 2018; Jiwanti et al., 2020). In addition, high chemical and physical stability also are very useful for applications in extreme conditions (Dettlaff et al., 2019; Muharam et al., 2019; Miao et al., 2020).

    Micro-band BDD electrodes with an effective surface of 1.11´10^-7 m² were successfully fabricated by lamination that used Teflon and silicon as the insulating plates. Cyclic voltametric studies of AMP, ADP, and ATP showed that the optimal parameters for electrochemical detection were at a potential of +0.9 V and a pH of 2.0. Employing the electrode for capillary electrophoresis can be conducted simultaneously on a solution of AMP, ADP, and ATP with linear correlations between the oxidation currents and the concentrations. The results showed that the system shows promise for a real application for simultaneous detection of adenosine phosphates. Hence, further investigations on the fabrication processes of the micro-band BDD electrode, as well as the optimal parameters used in the CE system, are crucial to increase the effectiveness of the modified BDD electrode as an adenosine detector.

    The authors would like to thank the Ministry of Research and Technology, Republic of Indonesia, for the financial support given to this research through Hibah Penelitian Dasar Unggulan Perguruan Tinggi (Grant No.: 1600/UN2.R3.1/HKP.05.00/2019). The authors also declare that there is no conflict of interest regarding the publication of this article.

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