Analysis of biothiols by HPLC-DAD
Biothiols in shoots and roots were analysed by HPLC-UV-diode array detector (DAD) (Sobrino-Plata et al., 2009). Extracts (100 µL) were injected in a Mediterranea SEA18 column (5 μm, 250 x 4.6 mm; Teknokroma, San Cugat del Vallés, Spain), using a 1200 HPLC system (Agilent), and biothiols were detected after post-column derivatization with Ellman reagent. Quantification was carried out adding N-acetyl cysteine (N-AcCys; final concentration 250 μM) as internal standard prior to sample homogenization.