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Table 2 Characteristics for the putatively identified marker metabolites in liquid chromatography-mass spectrometry analysis

From: Non-targeted metabolite profiling reveals changes in oxidative stress, tryptophan and lipid metabolisms in fearful dogs

 

Column

Ionization mode

MW

m/z

RT (min)

Putative annotation

p-valuea

FDR corrected p-valueb

Fold change (FC)c

CID (eV)

MS/MS fragmentation

Identification referenced

VIP

Cluster 1

RP

ESI+

821.577

822.584

10.67

PC(16:0/23:5)

0.0017

0.0226

−2.06

20

822.584, 184.074; ESI(−) 40 eV: 806.556, 343.249, 255.233

MS/MS

2.24

RP

ESI+

801.587

802.595

11.35

PC(18:0/19:1)

0.0376

0.1316

−1.98

20

184.072, 784.5833, 802.594; ESI(−) 40 eV: 295.229, 283.261, 786.565

MS/MS

1.30

RP

ESI+

204.09

205.097

2.30

Tryptophan

4.22E−04

0.0087

−1.58

10

188.0698, 146.0599, 144.0806, 130.0613, 132.0788, 159. 0881, 205.0947

Standard

0.84

RP

ESI+

537.295

538.309

10.25

LysoPC(19:0)

0.0488

0.1461

−1.53

20

104.106, 501.236, 560.310 [M + Na]+; ESI(−) 20 eV: 522.323, 297.245

MS/MS

1.68

Cluster 2

RP

ESI−

579.319

578.312

8.79

Unknown LysoPC

0.0103

0.0884

−2.79

20

293.209, 578.310, 518.291; ESI(+) 20 eV: 104.107, 534.319, 184.074

MS/MS

2.08

HILIC

ESI−

88.016

87.009

1.77

Unknown metabolite

0.0427

0.1108

−2.58

10

 44.999, 73.857

MS/MS

1.34

RP

ESI+

809.592

810.599

12.64

PC(18:0/20:4)

0.0200

0.0965

−2.00

40

184.073, 86.095; ESI(−) 40 eV: 303.234, 283.265, 794.567

MS/MS

1.82

RP

ESI+

517.316

518.323

8.81

LysoPC(18:3)

0.0472

0.1443

−1.86

40

184.072, 104.104, 86.094, 60.082; ESI(−) 20 eV: 502.2945, 277.2162

MS/MS

1.47

RP

ESI+

499.270

500.277

9.17

LysoPE(20:5)

0.0416

0.1376

−1.64

10

500.2786, 359.2548; ESI(-) 20 eV: 498.2860, 169.1368, 301.2172

MS/MS

1.28

Cluster 3

RP

ESI+

311.319

312.326

11.01

Unknown metabolite

0.0240

0.1055

2.85

20

312.326, 57.071, 102.095, 100.075, 214.214, 81.068

 MS/MS

1.88

HILIC

ESI−

189.994

188.986

0.69

Pyrocatechol sulfate

0.0150

0.0734

2.36

40

108.024, 79.957, 53.042, 80.965, 109.027

Pyrocatechol standard

1.55

HILIC

ESI+

246.137

247.144

1.42

Hypaphorine

0.0485

0.1719

2.17

10

188.071, 60.081, 146.061, 55.017, 247.206, 144.079, 85.0245, 118.928

Keller et al. [34]

1.29

RP

ESI−

213.009

212.002

2.43

Indoxylsulfate

0.0480

0.1870

1.78

10

212.007, 80.966, 132.043

MID 253

1.85

RP

ESI+

370.308

371.315

10.96

Unknown fatty acyl, either di-(2-ethylhexyl)adipate or dioctyl hexanedioate

0.0335

0.1253

1.55

10

129.0557, 111.0459, 147.0635, 101.0612, 57.0694, 241.1772

MS/MS

1.60

RP

ESI+

315.277

316.285

8.7

Unknown sphingosine, either dehydrophytosphingosine, 6-hydroxysphingosine, or (4OH,8Z,t18:1) sphingosine

0.0087

0.0619

1.50

40

93.071, 43.055, 57.069, 81.070, 69,069, 67.055, 95.048, 77.040

MID 392

0.68

RP

ESI+

283.287

284.294

10.59

Stearamide

0.0266

0.1108

1.28

20

284.295, 57.070, 102.091, 88.076, 71.085, 43.054

MID 34494

1.73

Cluster 4

HILIC

ESI+

136.039

137.046

1.43

Hypoxanthine

0.0250

0.1225

1.87

20

137.046, 119.035, 94.040, 110.035, 55.029, 82.038

MID 83

1.60

RP

ESI−

749.54

748.531

12.51

PE(P-18:1/20:4)

0.0447

0.1866

1.81

20

748.526, 303.234; ESI(+) 20 eV: 361.275, 390.2773, 609.529, 750.551

MS/MS

1.61

  1. Also the most significant non-identified marker metabolites are included. The characteristics include both uncorrected and FDR corrected p values, fold changes, Variable influence on projection (VIP) –values, and identification references, together with parameters for the LC–MS analysis, including the chromatography (Column), ionization mode in the mass spectrometry (Ionization mode), molecular weight (MW), identified ion (m/z), retention time (RT), collision induced dissociation energy (CID), and fragment ions in the tandem mass spectrometry (MS/MS fragments). n = 20 dogs (10 fearful and 10 non-fearful dogs). Note that two metabolic features, tryptophan and unknown sphingosine, are included in the table despite their low VIP values, since they otherwise show statistical significance
  2. LysoPC lysophosphatidylcholine, LysoPE lysophosphatidylethanolamine, PC phosphatidylcholine, PE phosphatidylethanolamine
  3. aStudent’s t-test comparing the fold change against the Control group. P values <0.05 were considered as statistically significant
  4. bBenjamini–Hochberg false discovery rate (FDR) corrected p value
  5. cAverage fold change when compared against the Control group, with p values. Fold changes ≥±1.2 were considered as statistically significant. Positive values indicate increased whole blood levels in case dogs vs. control dogs, whereas negative values indicate decreased whole blood levels in case dogs vs. control dogs
  6. dIdentification of metabolites is based on manual MS/MS spectral interpretation, METLIN ID when MS/MS spectrum available, commercial standard compound, or some earlier published fragmentation patterns. Keller et al. [34]