Finnigan surveyor

Prior to analysis, the samples were filtered through a 0.45 μm membrane and injected into the HPLC system. HPLC-PDA analyses were performed using a Finnigan Surveyor equipped with an autosampler, a diode array detector Finnigan Surveyor-PDA Plus (Thermo FisherScientific Inc., Waltham, MA, USA) and ChromQuest 5.0 chromatography software (Thermo FisherScientific Inc., Waltham, MA, USA). The separation conditions were as described by Efenberger-Szmechtyk et al. [36 (link)]. The calibration curves were established using standards for chlorogenic acid, quercetin-glucoside and cyanidin-glucoside to quantify polyphenols at 320, 360 and 520 nm, respectively.

The standards and samples were analyzed on a Finnigan Surveyor high performance liquid chromatography (HPLC) system equipped with a Finnigan linear trap quadrupole (LTQ) mass spectrometer (Thermo Scientific, Germany). The chromatographic separation was performed on a EZ:faast™ AAA-MS HPLC column which is part of EZ:faast™ amino acid analysis kit by Phenomenex. The temperature of the column was set to 35 °C. The mobile phase consisted of a gradient eluting system. Solvent A was methanol with 0.1% formic acid (LC–MS Chromasolv^®, Fluka) and solvent B was water with 0.1% formic acid (LC–MS Chromasolv^®, Fluka). Elution of the standards and samples was achieved with the following solvent gradient: 68% A to 83% A (22 min), 83% A to 68% A (0.01 min) and 68% A isocratic (27.99 min). The flow rate of the mobile phase was set to 0.15 mL/min, and the volume of injection was 2 µL.
The amino acids were identified by ion trap technology and the mass spectrometric ionization was performed with electrospray (Heimer et al. 2014 (link)). MS spectra were recorded consecutively in one segment with one full scan event in the range of m/z 100.00–650.00. The scan event was conducted in a positive ion mode with a skimmer induced dissociation of 35.00. All data was evaluated and interpreted with Xcalibur Software (Thermo Scientific, Dreieich, Germany).

Concentrations of glucose, succinate, acetate, citrate, malate, lactate and formate were quantified by HPLC (Finnigan Surveyor, Thermo Fisher Scientific, Waltham, USA). Detection was performed by refractive index (RI) (RID Agilent 1200, Agilent Technologies, CA, USA) with a 300 mm × 7.8 mm Aminex HPX-87H ion exchange column (Bio-Rad, Hercules, CA, USA). 20 µL sample was injected into the mobile phase of 5 mM H₂SO₄ (aq.), temperature was 60 °C and flow rate 0.35 mL min^− 1. The results from HPLC analysis were used together with DW measurements to calculate yield coefficients. Triplicate datasets are considered using error bars.