CE coupled with amperometric detection using a boron-doped diamond microelectrode: validation of a method for endogenous norepinephrine analysis in tissue

Abstract

The level of endogenous norepinephrine (NE) in several tissue types was determined by CE with amperometric detection. We report herein on the method validation by HPLC using both amperometric and coulometric detection (CD). Keys to the method were the use of a diamond microelectrode for detection and off-line SPE for sample preparation. The run buffer was a 250 mM borate solution adjusted to pH 8.8 with potassium hydroxide. The diamond microelectrode exhibited a low and stable background current, and a low peak-to-peak noise < or =0.65 pA at the detection potential of +0.86 V versus Ag/AgCl. For standard solutions, the detector signal (i.e., oxidation current) changed linearly with the NE concentration (r(2) = 0.999) between 60 and 1000 nmol/L with an estimated LOD of 51 nmol/L (S/N = 3) and a response variability of 4.5% (RSD, n = 5). An Oasis MCX sorbent was used for SPE and the procedure produced an NE recovery of 95.1 +/- 5.6% (n = 6) from tissue homogenates. NE levels in the spleen, small intestine, and heart of a normotensive rat were found to be in the range of 0.77-0.97, 0.22-0.32, and 0.29-0.45 microg/g tissue (n = 3), respectively.