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The MS data was acquired in selected-reaction monitoring
(SRM) mode on a Thermo Scientific
™
TSQ Quantiva
™
triple-stage quadrupole MS equipped with a HESI
interface.
MS Conditions
Ion mode
Positive HESI
Cycle time (s)
0.75
CID gas pressure (mTorr)
1.5
Spray voltage (V)
3500
Sheath gas (arb units)
60
Aux gas (arb units)
20
Sweep gas (arb units)
2
Ion transfer tube temp (˚C)
350
Vaporizer temp (˚C)
350
RF lens
Used calibrated RF lens values
Data processing, calibration, and quality control were
performed using Thermo Scientific
™
TraceFinder
™
software
version 3.1.
2
Broad-Spectrum HPLC-HRAM MS Screening
To begin analysis, broad-spectrum MS screening was
performed on a Thermo Scientific
™
LTQ Orbitrap Velos
™
hybrid ion trap-Orbitrap MS using heated electrospray
ionization (HESI). The instrument was operated in
positive full-scan (
m/z
100-1000) mode at a resolving
power of 60,000 (FWHM) at
m/z
400. Data-Dependent
Top 3 HRAM MS/MS experiments were performed with
dynamic exclusion and peak apex detection.
Non-Targeted Compound Identification
After broad-spectrum data acquisition, Thermo Scientific
™
ExactFinder
™
software version 2.5 was used for non-
targeted compound identification. The HRAM data was
screened for approximately 1000 known contaminants
using the environmental and food safety (EFS) compound
database and HRAM MS/MS spectral library. Automated
feature scoring and filtering was based on chromatographic
peak shape, mass error (ppm), and isotope pattern.
Structures were tentatively assigned by library searching
and later confirmed by analysis of authentic standards.
Targeted Quantitation
Targeted quantitation was performed with the
Thermo Scientific
™
EQuan MAX Plus
™
online SPE and
HPLC system. A 1 mL injection was loaded onto a
Thermo Scientific
™
Hypersil GOLD aQ
™
column (20 x
2.1 mm, 12 µm particle size) and separated on a
Thermo Scientific
™
Accucore
™
aQ analytical column
(50 x 2.1 mm, 2.6 µm particle size) by gradient elution
with methanol/water mobile phase.
LC Conditions
Loading pump
Thermo Scientific
™
Dionex
™
UltiMate
™
3000
Quaternary Analytical Pump LPG-3400SD
Flow rate
Isocratic 1 mL/min
Solvent A (water)
98%
Solvent B (methanol)
2%
Total run time 1
8.4 min
Analytical pump
UltiMate 3000 Binary Rapid Separation
Pump
HPG-3200RS
Solvent A (water)
98%
Solvent B (methanol)
2%
Gradient elution
0.3 mL/min
Gradient
Time
%A
0
98
1.5
98
12.0
2
15.0
2
15.1
98
Total run time
18.4 min
Autosampler
Thermo Scientific
™
Open Accela
™
autosampler
Valve switching
At 1.5 min and 16.6 min