ACE UHPLC and HPLC Columns
Amino Acids in Extracellular Matrix
L -isoleucine L -leucine
L-isoleucine L-leucine
Time - Minutes
Intensity
Intensity
Mass/Charge, Da Mass/Charge, Da
Conditions
Application #AN4410
Column: ACE 3 AQ
Dimensions: 150 x 0.5 mm
Part Number: ACE-116-15005
Mobile Phase: A: 0.1% (v/v) formic acid in H2O
B: 0.1% (v/v) formic acid in MeCN
Gradient: Time (mins) %B
0 2
5 20
Flow Rate: 20 μL/min
Injection: 2 μL
Detection: SCIEX QTRAP 6500 LC-MS/MS system
IonDrive Turbo V source
Positive ion MRM mode
Sample: Standard solution containing 2.5 μmol/mL each
amino acid (1.25 μmol/mL cysteine).
0.5 pmol on-column (except for cysteine,
0.25 pmol on-column).
Method also applied to analysis of cell
supernatant from purified peripheral blood
mononuclear cells (PBMCs)
Full scan linear ion trap MS/MS data can distinguish isobaric amino acids L-isoleucine and L-leucine.
Intensity
Peak Analyte Rt (mins) MRM Transition (m/z) LOD (fmol) PBMC cell
conc. (fmol/μL)
1 Lys 1.094 147.1 84 5 305
2 His 1.111 156.1 110 5 23
3 Arg 1.117 175.2 70 2.5 220
4 Gly 1.129 76.1 30 <1000 <LOD
5 Cys 1.140 241.2 152.1 1.25 36
6 Asp 1.155 134.1 74 10 26
7 Ser 1.156 106.1 60 50 21
8 Ala 1.189 90.1 44 <1000 <LOD
9 Glu 1.208 148.1 84 5 55
10 Pro 1.262 116.1 70 2.5 96
11 Val 1.630 118.1 55 25 105
12 Met 1.645 150.2 104 1 3
13 Tyr 1.669 182.2 165.2 1 97
14 Ile 1.773 132.1 86, 69 2.5 329
15 Leu 1.858 132.1 86 2.5 338
16 Phe 2.273 166.1 103 1 100
17 Thr 2.275 120.1 103.2 1 97
Reproduced with permission of SCIEX, Warrington, UK
Page 1 of 2
MRM transitions and limits of detection (LODs) for 17 free amino acids and their
concentrations measured in diluted PBMC cell supernatant
26
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