Improved detection limit for a direct determination of 8-hydroxy-2′-deoxyguanosine in untreated urine samples by capillary electrophoresis with optical detection

Method for a direct determination of 8-hydroxy-2′-deoxyguanosine (8OHdG) in untreated urine samples by capillary electrophoresis with optical detection was developed. Optimisation of conditions resulted in a significant lowering of the limit of detection (LOD) by a factor of 400 as compared to our previous study. Optimum separation of 8OHdG from other urine components was achieved using the separation electrolyte containing 80 mM 2-(cyclohexylamino)ethanesulfonic acid, 9 mM LiOH (pH 8.6), and 0.1 mM cetyltrimethylammonium bromide ensuring the electro-osmotic flow inversion. In the model aqueous samples, these conditions allow separating 8OHdG and 2′-deoxyguanosine (dG) from other nucleosides/nucleotides including 2′-deoxycitidine 5′-monophosphate (dCMP), thymidine 5′-monophosphate (TMP), adenosine (A), and thymidine (T). On the other hand, 2′-deoxyadenosine 5′-monophosphate (dAMP) and 2′-deoxyguanosine 5′-monophosphate (dGMP) migrate together, and guanosine (G), 2′-deoxyadenosine (dA), 2′-deoxycytidine (dC) are transported as neutral species with the electro-osmotic flow. In the spiked urine samples, 8OHdG and dG are well separated from each other and from other urine components and exhibit a linear calibration over the concentration range of 0.1–2.0 μM for 8OHdG (LOD = 42 nM) and 0.2–5.0 μM for dG (LOD = 86 nM), but urine metabolites interfere with the determination of dCMP, TMP, A and T. Method is applicable to untreated urine samples with slightly enhanced levels of 8OHdG compared to that found in healthy individuals.