Chromolith rp 18e 100 x 4.6 mm column

Brain extracts from all 44 individuals included in the analysis were homogenized (1:2 v/v) in ice-cold 0.25 M perchloric acid containing 0.1 M NaS2O5 and 0.25 M ethylenediaminetetraacetate (EDTA) and kept frozen at -80°C until use. After centrifugation at 12000 x g for 10 min at 4 °C, the concentration of catecholamines (NA, DA, DOPAC and HVA) and indoleamines (5-HT and 5-HIAA) were determined in 20 μL aliquots using HPLC (Elite LaCHrom, Merck, Hitachi, Japan) equipped with a Chromolith Rp-18e 100 x 4.6 mm column (Merck KgaA, Darmstadt, Germany) with electrochemical detection (ESA Coulochem II 5200, Bedford, MA). The mobile phase consisted of 0.5 M citrate buffer pH 2.8, 0.05 mM EDTA, 1.2 mM sodium octyl sulphate (SOS) and 1 % acetonitrile. The applied voltage was set at 400 mV and the flow rate was 1 mL/min [48] (link). Validation of the methodology is described in Arroyo et al. [43] (link). The internal control DHBA allowed the comparison between runs. Dopaminergic total system (DA-system) and serotonergic total system (5-HT-system) are calculated as the sum of all metabolites in the pathway (DA, DOPAC and DA; and 5-HT and 5-HIAA; respectively).