The Development of a Direct Electron Transfer Type Open Circuit Potential Based Glucose Sensor Utilizing Microneedle Type Gold Electrode
Authors: Lee Inyoung , Loew Noya , Okuda-Shimazaki Junko , Tsugawa Wakako , Ikebukuro Kazunori , Sode Koji 
Published: February 2020

Introduction

To control the blood glucose level of Type I diabetes patients, the continuous monitoring of the blood glucose concentration is the inevitable information for the subsequent infusion of insulin. Recently, the U.S. Food and Drug Administration (FDA) approved a medical device comprising a continuous glucose monitoring (CGM) system, a continuous subcutaneous insulin infusion pump and a control algorithm that can supply an appropriate amount of basal insulin based on the CGM results as an artificial pancreas. Consequently, it became possible to automatically and continuously control the blood glucose level.
CGM is the core component in the artificial pancreas system because the amount of insulin being supplied is calculated based on the CGM results. The incorrect amount of insulin infusion due to incorrect CGM results might make diabetes patient iatrogenic-hypoglycemia. To stably and accurately control the blood glucose level, CGM should be stable and accurate for a long period. Currently, commercial available CGM sensors are based on amperometric methods employing either 1st or 2nd generation enzyme electrochemical sensing principles; using either oxygen as the electron acceptor for 1st generation, or artificial electron acceptors for 2nd generation. Besides, the 3rd generation principle, which is the direct electron transfer principle, is the ideal electrochemical method where no external electron acceptors are required, since the enzyme itself can transfer electron directly with electrode. The direct electron transfer (DET) principle can minimize the electrochemical reactions and the sensor components, therefore this principle is considered as the ideal system for sensor used for CGM.
We have been engaged in the development of DET principle based glucose sensor employing DET-type FAD dependent glucose dehydrogenase (FADGDH) (Kakehi et al., 2007; Sode et al., 2016; Lee et al., 2018). FADGDH was originally discovered by our group, and molecular science as well as its mechanisms of electron transfer have been studied (Inose et al., 2003; Tsuya et al., 2006; Yamashita et al., 2018). We recently reported on the open circuit potential (OCP) type sensor for CGM based on DET principle. OCP is the potential of an electrode in the open circuit state. When the circuit is opened, current cannot flow in the circuit; therefore, there is no current. In other words, the turn-over of enzyme can be minimized during open circuit state. The OCP based glucose sensor can measure glucose concentration accurately and stably during 9 days without being affected by interference substances (I. Lee et al., 2019).
Here, we attempted to develop the miniaturized OCP based glucose sensor by utilizing microneedle type gold electrode which is sufficiently small for implanting. We demonstrated measurement of glucose concentration and continuous operation using the miniaturized OCP based glucose sensor.

Method

Recombinantly prepared bacterial FADGDH, capable of direct electron transfer, was used in this study, and the enzymes were immobilized onto microneedle type gold electrode. OCP of the miniaturized OCP based glucose sensor was monitored continuously toward the sample containing various concentration of glucose, against Ag/AgCl reference electrode. Calibration curve was prepared based on plotting ∆OCP versus glucose concentration.

Result and discussion

OCP decreased with increasing on glucose concentration, and ∆OCP of the miniaturized OCP based glucose sensor increased depending on glucose concentration (1 mM~20 mM). Even though the electrode surface area became extremely smaller by employing microneedle disk electrode, we could not observe the significant difference on both sensor response and sensitivity between microneedle gold electrode and gold disk electrode (I. Lee et al., 2019). In other words, by employing OCP for glucose measurement, we could obtain sufficiently high sensor signal without being affected by the area of electrode surface. This characteristic of OCP is suitable for CGM which requires miniaturization of sensor chip for implanting. Furthermore, this miniaturized OCP based glucose sensor could generate stable OCP signal for more than 43 hours of continuous operation. From these results, it is indicated that this miniaturized glucose sensor utilizing microneedle gold electrode can be applied for CGM by employing OCP.

References

Inose et al., Biochimica et Biophysica Acta, 1645, 133-138 (2003)

Tsuya et al., Journal of Biotechnology, 123, 127-136 (2006)

Kakehi et al., Biosensors and Bioelectronics, 22, 2250-2255 (2007)

Sode et al., Biosensors and Bioelectronics, 76, 20-28 (2016)

Lee et al., Bioelectrochemistry, 121, 1-6 (2018)

Yamashita et al., Current Opinion in Electrochemistry, 12, 92-100 (2018)

Lee et al., Biosensors and Bioelectronics, 124-125, 216-223 (2019)