Figure 4.

Chromatographic conditions for monoamines measurement: Focus on NA. The purpose of these chromatograms is to illustrate that the measurement of NA using classical procedures, which is not an easy task, because several species can be eluted at the same time. (A) Experimental procedures. We used classical procedures for the chromatographic conditions, as recently published [94], with some modifications as regards to the electrochemical detection. A conditioning cell (5020, ESA) was placed before the manual injector and set at +450 mV. Then, after the column, two coulometric cells (5011, ESA; two electrodes each) were placed in series, each cell being connected to one coulometric detector (CoulochemII, ESA). The potential at the four electrodes was as follows: +125, +350, +500, and −300 mV (reduction). The detectors were connected to the CED 1401 interface (Cambridge Electronic Design Ltd.), and signals were acquired using the spike 2 software (Cambridge Electronic Design Ltd.). B. Chromatograms (mV as a function of time, in minutes) obtained for each electrode for different groups of molecules: First column (B1,B2): L-DOPA, NA, adrenaline, DA; second column (C1,C2): NA and octopamine; third column (D1,D2): VMA, DHPG, DOPAC, HVA, 3-MT, and 5-hydroxytryptamine (5-HT). It is possible to act on the time of elution of the compounds by modifying the pH (here by adding 20 µL NaOH, the below panels B2, C2, and D2), the concentration of pairing ions, and the initial concentration of methanol (here 7%). Note that L-DOPA and NA are confounded in the initial condition and can be separated by basifying the mobile phase. Nonetheless, it is convenient to have different potentials here because some compounds do not generate signals at low potentials (E1), including octopamine, DHPG, and 3-MT. Other compounds would have given signals in these conditions: MHPG, 5-hydroxyindole acetic acid, 5-hydroxytryptophan, tryptophan, and most likely the various metabolites of L-DOPA.