Development of Electrochemical Sensor for Uric Acid Detection in Intestinal Closed Loop

Abstract

Uric acid is a ulitimate metabolite of purine nucleotides in humans, because the uricase had been lost completely. So, the serum uric acid level is higher than other mammalians. The advantage of the higher serum uric acid levels has been suggested to be antioxidant activity of uric acid. On the other hand, hyperuricemia is an abnormally high level of uric acid in the serum that leads to uric acid precipitation in tissues and the main risk factors for the onset of gout, hypertension, chronic kidney disease, atherosclerosis, diabetes and so on. There are two important physiological mechanisms caused hyperuricemia: (a) increase production of the uric acid from dietary and endogenous substrates that raise purine levels and (b) reduced renal and extra-renal excretion of uric acid. It is commonly accepted that two thirds of the urate is excreted from the kidney into urine. The remaining one third is excreted via extra-renal excretion, mainly intestine. Recently, importance of the uric acid excretion from intestine has been gathering attention because the considerable inpact of urate transporter function in the intestine on the serum uric acid level has been revealed gradually. A novel concept in hyperuricemia was proposed as “extra renal underexcretion type”.[1] Intestinal closed loop method is one of the basic techniques to investigate the excretion of uric acid via trasporters in intestin. However, it needs troublesome procedure and time consuming process. If it is possible to detect the concentration of excreted uric acid with electrochemical sensor, dynamic analysis of urate excretion via the urate transporters will easily be available. In this study, we developed an electrode to detect uric acid sensitively. Gold beads electrode was prepared according to the literature.[2] 6-Ferrocenyl-1-hexanethiol which works as a mediator for the uric acid oxidation was used to make self -assenbled monolayers (SAMs) on gold surface. 6-Hydroxyl-1-hexananethiol, 5-Carboxyl-1-pentanethiol and 6-Amino-1-hexanethiol were also used to construct mixed SAMs. Gold beads electrodes were soaked in the ethanol solution containing these alkanethiols. The SAM-coated electrode was used as working electrode (C.E: Pt wire, R.E.: Ag/AgCl). Uric acid dissolved in phosphate buffer (pH 7.4) was added to the measurement system. Cyclic voltammograms were measeured in buffer by addition of uric acid (-0.2 to +0.5 V). Oxdation current based on the uric acid oxidation was observed with 6-Ferrocenyl-1-hexanethiol modified electrode at around +0.45 V. The oxidation current was increased with increasing of uric acid concentration. When the mixed SAMs-coated electrodes were examined, similar responses were observed regardress the component of SAMs. However, no response was observed from the SAM-coated electrode without 6-Ferrocenyl-1-hexanethiol. Responsible change was observed at oxidation current by 0.1 mg/mL change of uric acid. Furthermore, ammperometric measurement showed the step-like current change according to the added uric acid concentration.