Analytical Sciences

Electrochemical Determination of Oxidative Damaged DNA with High Sensitivity and Stability Using a Nanocarbon Film

Dai KATO,* Mayuri KOMORIYA,** Kohei NAKAMOTO,** Ryoji KURITA,* Shigeru HIRONO,*** and Osamu NIWA*,**

*National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan
**University of Tsukuba, 1-1-1 Tenno-dai, Tsukuba, Ibaraki 305-8571, Japan
***MES-Afty Corporation, 2-35-2 Hyoe, Hachioji, Tokyo 192-0918, Japan

We describe the electrochemical determination of oxidative damaged DNA by using a nanocarbon film electrode combined with a high performance liquid chromatography (HPLC) system. The nanocarbon film was formed by employing the electron cyclotron resonance sputtering method, and has a nano-crystalline sp² and sp³ mixed bond structure with an atomically flat surface. This film electrode provided the high electrode activity and stability needed to quantitatively detect oxidative damaged DNA, 8-hydroxy-2′-deoxyguanosine (8-OHdG), by direct electrochemical oxidation. The coefficient of variation (C.V.) value of 1 μM 8-OHdG at our film electrode was 0.75% (n = 12), which constitutes superior reproducibility to that of a conventional glassy carbon (GC) electrode (9.28%, n = 12) in flow-injection analysis. This was because the nanocarbon film suppressed fouling for the oxidized product of 8-OHdG owing to its hydrophilically ultraflat and chemically stable surface. We also investigated the performance of HPLC with an electrochemical detection (HPLC-ECD) system using our nanocarbon film electrode. The detection limit for 8-OHdG at the nanocarbon film electrode was 3 nM, which was superior to the detection limit of the GC electrode (7.2 nM). Furthermore, this electrode was more suitable for use in a urinary 8-OHdG experiment than the GC electrode. The concentration of urinary 8-OHdG in the urinary sample was 8.30 nM. These results indicate that this HPLC-ECD system with our nanocarbon film electrode enables us to realize an accurate, sensitive, reproducible and easy to use analysis technique.

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