LC-MS/MS Raw Files: Anthocyanin identification from mixed flavonoid populations in flowers by positive and negative mode LC-MS/MS
Description
This data set includes the raw files for methanolic extracts of Clitoria ternatea and Viola sororia run in LC-MS/MS which we obtained for the purpose of qualitatively identifying anthocyanin and flavonol molecules present in such flowering plants. Anthocyanins are the primary pigments found in flowers and fruits and we aim to develop a rapid and sensitive mass spectrometry-based workflow to identify anthocyanins and their sugar an acyl substituents and differentiate them from a mixed flavonoid population. Targeting anthocyanins and flavonols, we first searched the MS1 data for high-mass (>300 Da) chromatographic peaks in the 5-12.5 minute region of our gradient where we expected such molecules to elute. After identifying potential peaks, we then searched the MS2 spectra associated with these peaks, based on the apparent parent mass in the MS1 domain, looking for obvious signs of anthocyanidin or flavonol aglycone fragments, which appear prominently under our fragmentation scheme below ∼350 Da. Once parent adducts and associated MS2 spectra were assigned, we then manually annotated the fragmentation spectra based on observed losses associated with various sugar or acid moieties being cleaved from the central aglycone structure.
Files
Steps to reproduce
Extracted pigment samples were analyzed using a Waters (Milford, Massachusetts) Xevo TQ Absolute UPLC-MS/MS system with ACQUITY (Milford, Massachusetts) UPLC Premier. Compound separation was performed using an ACQUITY PREMIER CSH Phenyl-Hexyl column (1.7 µM VanGuard Fl 2.1 x 50 mm Column) with a column temperature set to 50 ◦C. Mobile phase A was composed of 0.1% formic acid from Macron (Radnor, Pennsylvania) and mobile phase B was 100% acetonitrile, LC/MS grade from Fisher Scientific (Fair Lawn, New Jersey). The flow rate was set to 0.4 mL/min and the gradient was non linear with composition as follows: 0-2 min, 0-10% B; 3-23 min, 10-40% B; 23-26 min, 40-100% B; 28-30 min, 100-0% B. Electrospray ionization was employed in both positive and negative modes. Time of- flight mass spectra in the m/z range 100- 1200 and MS/MS scans in the m/z range 50- 2000 were obtained with fast data-dependent acquisition in centroid mode. Other mass spectrometry conditions were all as follows: nitrogen gas temperature, 350 ◦C; drying gas flow rate, 800 L/hr; cone gas, 50 L/hr; cone voltage, 75 V; capillary voltage, 3 kV (positive mode) and 2 kV (negative mode); and collision energy, 10- 100 V. The mass axis was calibrated using a solution of sodium formate clusters ranging from 50-2000 Da. The instrument was calibrated real time with internal calibrant (LockSpray Leucine-Enkephalin Single Point MS) every 10 seconds.
Institutions
- University of California Irvine