3.6.1. MALDI MS

MALDI analysis was carried out using a Bruker Autoflex III Smartbeam (Bruker Daltonik, Bremen, Germany). All MALDI MS equipment, software and consumables were from Bruker Daltonics. The laser (355 nm) had a repetition rate of 200 Hz and operated in the positive reflectron ion mode for MS data over the mass range of 400 to 2200 Da under the control of the Flexcontrol and FlexAnalysis software (V 3.3 build 108). External calibration was performed using the sodium-attached ions from a Polyethylene Glycol of average molecular weight 1000. MS spectra were processed in FlexAnalysis (version 3.3, Bruker Daltonik, Bremen, Germany). MALDI MS<sup>2</sup> spectra were obtained using the LIFT mode of the Bruker Autoflex III with the aid of CID. The isolated ions were subjected to collision against argon in the collision cell to be fragmented, affording intense product ion signals. For MALDI a laser was used to provide both good signal levels and mass resolution with the laser energy for MS2 analysis being generally 25% higher than for MS analysis.

The samples were spotted onto a MALDI stainless steel MPT Anchorchip TM 600/384 target plate. Alpha-cyano-4-hydroxycinnamic acid (CHCA) in acetone/iso-propanol in ratio of 2:1 (15 mg mL<sup>−</sup>1) was used as a matrix to produce gas-phase ions. The matrix solution (1 μL) was placed onto the MALDI target plate and air-dried. Subsequently 1 μL of sample was added to the matrix crystals and air-dried [3,12]. Finally, 1 μL of NaI (Sigma-Aldrich # 383112, St Louis, MI, USA) solution (2 mg/mL in acetonitrile) was applied onto the sample spots. The samples were mixed on the probe surface and dried prior to analysis.
