104 Lipid Mediator Metabololipidomics LC-MS-MS Spectra ...serhanlab.bwh.harvard.edu/.../UPDATED-2019-Spectra... · Lipid Mediator Metabololipidomics LC-MS-MS Spectra Book 2019 Ian

LipidMediatorMetabololipidomicsLC-MS-MSSpectraBook2019

IanRILEY,PaulNORRIS,KathyLUONG,andCharlesSERHAN

CenterforExperimentalTherapeuticsandReperfusionInjury,HarvardInstituteofMedicine,BrighamandWomen’sHospitalandHarvardMedicalSchool,Boston,Massachusetts02115U.S.A.

Correspondenceshouldbeaddressedto:Prof.CharlesN.SerhanBuildingforTransformativeMedicine60FenwoodRoadBoston,MA02115

Email:[email protected];Tel:617-525-5001;Fax:617-525-5017

HARVARD MEDICAL SCHOOL

BRIGHAM AND WOMEN’S HOSPITAL

Version:02142019

*The2019updateofthe2016SpectraBookWewouldliketoacknowledgethesupportoftheNIHGrantP01GM095467

104

1

CommerciallyAvailableStandards

SpecializedPro-ResolvingMediatorsLipoxinA4LipoxinB45S,15S-diHETE15epi-LipoxinA4ResolvinD117epi-ResolvinD1ResolvinD2ResolvinD3ResolvinD510(S),17(S)-DiHDHA(PDX)Maresin1LipoxinA5

BiosynthesisPathwayMarkersAA5-HETE15(S)-HpETE5(S)-HpETE12-HETE15-HETEDHA17(S)-HpDHA14(S)-HDHA4-HDHA7-HDHA17-HDHAEPA5-HEPE12-HEPE15-HEPE18-HEPEAAbioactiveMetabolomePGE2PGD2PGF2α20-OH-LTB4LTB4LTC4LTD4LTE4TXB2

Prof.CharlesN.SERHANLaboratoryBrigham&Women’sHospitalCenterforExperimentalTherapeuticsandReperfusionInjuryHarvardMedicalSchool


©2019Serhanetal.

CaymanCatalogNumbers

(90410)(90420)(35280)(90415)

(10012554)

(13060)(10007279)

(13834)(10007280)

(10008128)

(10878)

(10011453)

(90010)(34210)(44720)(44230)(34550)(34700)

(90310)(13185)(15253)(33200)(33300)(33650

(90110)(32200)(32540)(32700)(32840)

(14010)(12010)(16010)(20190)(20110)(20210)(20310)(20410)(19030)

2

TableofContents

LipidMediatorStructures&Stereochemistry 6ArachidonicAcidMetabolome 7EicosapentaenoicAcidMetabolome 12DocosahexaenoicAcidMetabolome 16Aspirin-TriggeredSpecializedPro-resolvingMediators 21DeuteriumLabeledInternalStandards 25

Methods 28Operationalizationoflipidmediatormetabololipidomics 29HighPressureLiquidChromatography 30MassSpectrometry 30Methodsvalidation 33

Linearityandattributedinternalstandard 33Recovery,matrixinterferenceandlimitofdetection 34Repeatabilityandreproducibility 35Chemicalstabilityofselectedendogenouslipidmediatorsidentifiedinhumanserum

39

MultipleReactionMonitoringTransitions 40LipidMediators 41Aspirin-TriggeredLipidMediators 41FurtherMetabolitesofLipidMediators 42

LipidMediatorBiosyntheticPathways 43Prostaglandins,LeukotrienesandLipoxins 44Leukotrienes 45E-SeriesResolvins 46D-SeriesResolvins 47Aspirin-TriggeredD-SeriesResolvins 49Protectins 50Maresins 51

LC-MS/MSSpectra 52ArachidonicAcidMetabolome 53

Prostaglandins&Thromboxane 53Leukotrienes 58Lipoxins 62MonoHydroxyETEProducts 66



©2019Serhanetal.

3

EicosapentaenoicAcidMetabolome 70Lipoxins 70E-SeriesResolvins 73MonoHydroxyEPEProducts 77

DocosahexaenoicAcidMetabolome 82D-SeriesResolvins 82Maresins 89Protectins 92MonoHydroxyDHAProducts 94

DocosapentaenoicAcidMetabolome 98D-SeriesResolvins 98Maresins 101Protectins 10313-seriesResolvins 105MonoHydroxyDPAProducts 109Aspirin-TriggeredSpecializedPro-resolvingMediators 112CystenielLeukotrienes 118MaresinConjugateinTissueRepair 122ProtectinConjugateinTissueRepair 126ResolvinConjugateinTissueRepair 130DeuteriumLabeledInternalStandards 134



©2019Serhanetal.

TableofContents

4

LipidMediatorStructures&Stereochemistry



©2019Serhanetal.

5

ArachidonicAcidMetabolome



©2019Serhanetal.

6

Complete

Stereo

chem

istry

5Z,8Z,11

Z,14

Z-Eicosatetraeno

icacid

11-oxo-

9α,15S-

dihydroxy-

5Z,13E-

prostadien

oicacid

9-oxo-

11α,15S-

dihydroxy-

5Z,13E-

prostadien

oicacid

9α,11α

,15S-

trihydroxy-

5Z,13E-

prostadien

oicacid

TrivialN

ame

Arachido

nicacid

ProstaglandinD 2

ProstaglandinE 2

ProstaglandinF 2

α

Chem

icalNam

e

Eicosatetraeno

ic

acid

11-deh

ydro-

prostaglandinF 2

α

9-de

hydro-

prostaglandinF 2

α

9α,11α

,15S-

trihydroxy-

prostaglandinF 2

α

Form

ula

C 20H

32O2

C 20H

32O5

C 20H

32O5

C 20H

34O5

Structure

Abbreviatio

n

AA

PGD 2

PGE 2

PGF 2

α

7

COOH

COOH

OH

O

HO

COOH

OH

HOO

COOH

OH

HO

HO

Complete

Stereo

chem

istry

9α,�11,�15

S-�

trihydroxy-

5Z,�13E-

thromba�dien

�oicacid

5S,�12R

-�dihydroxy-�

6Z,�8E,�10

E,�14

Z-�

eicosatetraeno

icacid

5S,�12R

,20-�

trihydroxy-�

6Z,�8E,�10

E,�14

Z-�

eicosatetraeno

icacid

5S,�12R

-�dihydroxy-

�6Z,�8E,�10

E,�14

Z-�

20-carbo

xy-

eicosatetraene

dioicacid

TrivialN

ame

Thrombo

xane

B2

Leukotrie

neB

4

20-hydroxy-

Leukotrie

neB

4

20-carbo

xy-

Leukotrie

neB

4

Chem

icalNam

e

9α,�11,�15

S-�

trihydroxy-

thrombo

xane

B2

5S,12R

-dihydroxy-�

LTB 4

5S,12R

,20-

trihydroxy-�

LTB 4

5S,12R

-dihydroxy-�

20-carbo

xy-

LTB 4

Form

ula

C 20H

34O6

C 20H

32O4

C 20H

32O5

C 20H

30O6

Structure

Abbreviatio

n

TxB 2

LTB 4

20-OH-LTB 4

20-COOH-LTB 4

COOH

OH

OHO

OH

COOH

OH

OH

COOH

OH

OH

OH

COOH

COOH

OH

OH

8

Complete

Stereo

chem

istry

5S,6R,15

S-

trihydroxy-

7E,9E,11

Z,13

E-eicosatetraeno

icacid

5S,14R

,15S-

trihydroxy-

6E,8Z,10

E,12

E-

eicosatetraeno

icacid

5S,�15S-

�dihydroxy-

�6E,�8Z,�11

Z,�13

E-�

eicosatetraeno

icacid

5-�

hydroxy-�

6E,�8Z,�11

Z,�14

Z-�

eicosatetraeno

icacid

TrivialN

ame

Lipo

xinA 4

Lipo

xinB 4

5S,15S-

dihydroxy-

eicosatetraeno

ic

acid

5-

hydroxy-

eicosatetraeno

ic

acid

Chem

icalNam

e

5S,6R,15

S-

trihydroxy-

LXA 4

5S,14R

,15S-

trihydroxy-

LXB 4

5S,15S-

dihydroxy-

ETE

5-

hydroxy-

ETE

Form

ula

C 20H

32O5

C 20H

32O5

C 20H

32O4

C 20H

32O3

Structure

Abbreviatio

n

LXA 4

LXB 4

5S,15S-diHETE

5-HE

TE

COOH

OH

HO

OH

COOH

OH

HO

OH

COOH

OH

9

COOH

OH

OH

Complete

Stereo

chem

istry

12-�

hydroxy-�

5Z,�8Z,�10

E,�14

Z-�

eicosatetraeno

icacid

15-�

hydroxy-�

5Z,�8Z,�11

Z,�13

E-�

eicosatetraeno

icacid

TrivialN

ame

12-

hydroxy-

eicosatetraeno

ic

acid

15-

hydroxy-

eicosatetraeno

ic

acid

Chem

icalNam

e

12-

hydroxy-

ETE

15-

hydroxy-

ETE

Form

ula

C 20H

32O3

C 20H

32O3

Structure

Abbreviatio

n

12-HETE

15-HETE

10

COOH

OH

COOH

OH

EicosapentaenoicAcidMetabolome



©2019Serhanetal.

11

Complete

Stereo

chem

istry

5Z,8Z,11

Z,14

Z,17

Z-eicosape

ntaeno

icacid

5S,�6R,�15

S-�

trihydroxy-

�7E,�9E,�11

Z,�13

E,�17

Z-�

eicosape

ntaeno

icacid

5S,14R

,15S-

trihydroxy-

6E,8Z,10

E,12

E,17

Z-

eicosape

ntaeno

icacid

5S,�15S-�

dihydroxy-

�6E,�8Z,�11

Z,�13

E,17

Z-�

eicosape

ntaeno

icacid

TrivialN

ame

Eicosape

ntaeno

ic

acid

Lipo

xinA 5

Lipo

xinB 5

5S,15S-

dihydroxy-

eicosape

ntaeno

ic

acid

Chem

icalNam

e

Eicosape

ntaeno

ic

acid

5S,6R,15

S-

trihydroxy-

LXA 5

5S,14R

,15S-

trihydroxy-

LXB 5

5S,15S-

dihydroxy-

EPE

Form

ula

C 20H

30O2

C 20H

30O5

C 20H

30O5

C 20H

30O4

Structure

Abbreviatio

n

EPA

LXA 5

LXB 5

5S,15S-diHEP

E

COOH

OH

HO

OH

COOH

OH

HO

OH

COOH

OH

OH

COOH

12

Complete

Stereo

chem

istry

5S,12R

,18R

-trihydroxy-

6Z,8E,10

E,14

Z,16

E-eicosape

ntaeno

icacid

5S,18R

-dihydroxy-

6E,8Z,11

Z,14

Z,16

E-

eicosape

ntaeno

icacid

17R,18

R-

dihydroxy-

5Z,8Z,11

Z,13

E,15

E-eicosape

ntaeno

icacid

5-

�hydroxy-�

6E,�8Z,�11

Z,�14

Z,�17

Z-�

eicosape

ntaeno

icacid

TrivialN

ame

ResolvinE1

ResolvinE2

ResolvinE3

5-

hydroxy-

eicosape

ntaeno

ic

acid

Chem

icalNam

e

5S,12R

,18R

-trihydroxy-

EPE

5S,18R

-dihydroxy-

EPE

17R,18

R-

dihydroxy-

EPE

5-

hydroxy-

EPE

Form

ula

C 20H

30O5

C 20H

30O4

C 20H

30O4

C 20H

30O3

Structure

Abbreviatio

n

RvE1

RvE2

RvE3

5-HE

PE

COOH

OH

HO

COOH

OH

COOH

HO

OH

COOH

HO

OH

OH

13

Complete

Stereo

chem

istry

12-�

hydroxy-

�5Z,�8Z,�10

E,�14

Z,�17

Z-�

eicosape

ntaeno

icacid

15-

�hydroxy-�

5Z,�8Z,�11

Z,�13

E,�17

Z-�

eicosape

ntaeno

icacid

18-�

hydroxy-�

5Z,�8Z,�11

Z,�14

Z,�16

E-�

eicosape

ntaeno

icacid

TrivialN

ame

12-

hydroxy-

eicosape

ntaeno

ic

acid

15-

hydroxy-

eicosape

ntaeno

ic

acid

18-

hydroxy-

eicosape

ntaeno

ic

acid

Chem

icalNam

e

12-

hydroxy-

EPE

15-

hydroxy-

EPE

18-

hydroxy-

EPE

Form

ula

C 20H

30O3

C 20H

30O3

C 20H

30O3

Structure

Abbreviatio

n

12-HEP

E

15-HEP

E

18-HEP

E

COOH

OH

COOH

OH

COOH

OH

14

DocosahexaenoicAcidMetabolome



©2019Serhanetal.

15

Complete

Stereo

chem

istry

4Z,7Z,10

Z,13

Z,16

Z,19

Z-do

cosahe

xaen

oicacid

7S,8R,17

S-

trihydroxy-

4Z,9E,11

E,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

7S,16R

,17S-

trihydroxy-

4Z,8E,10

Z,12

E,14

E,19

Z-

docosahe

xaen

oicacid

4S,11R

,17S-

trihydroxy-

5Z,7E,9E,13Z,15E,19Z-

docosahe

xaen

oicacid

TrivialN

ame

Docosahe

xaen

oic

acid

ResolvinD1

ResolvinD2

ResolvinD3

Chem

icalNam

e

Docosahe

xaen

oic

acid

7S,8R,17

S-trihydroxy-

DHA

7S,16R

,17S-

trihydroxy-

DHA

4S,11R

,17S-

trihydroxy-

DHA

Form

ula

C 22H

32O2

C 22H

32O5

C 22H

32O5

C 22H

32O5

Structure

Abbreviatio

n

DHA

RvD1

RvD2

RvD3

COOH

OH

HO

HO

COOH

COOH

HO

OH

OH

OH

HO

COOH

HO

16

Complete

Stereo

chem

istry

4S,5,17S-

trihydroxy-

6E,8E,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

7S,17S-

dihydroxy-

4Z,8E,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

4S,17S-

dihydroxy-

5E,7Z,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

7R,14S-

dihydroxy-

4Z,8E,10

E,12

Z,16

Z,19

Z-do

cosahe

xaen

oicacid

TrivialN

ame

ResolvinD4

ResolvinD5

ResolvinD6

Maresin1

Chem

icalNam

e

4S,5R,17

S-

trihydroxy-

DHA

7S,17S-

dihydroxy-

DHA

4S,17S-

dihydroxy-

DHA

7R,14S-

dihydroxy-

DHA

Form

ula

C 22H

32O5

C 22H

32O4

C 22H

32O4

C 22H

32O4

Structure

Abbreviatio

n

RvD4

RvD5

RvD6

MaR

1

COOH

HO

HO

OH

OH

COOH

OH

OH

OH

HOOC

OH

OH

HOOC

17

Complete

Stereo

chem

istry

4S,14S-

dihydroxy-

5E,7Z,10

Z,12

E,16

Z,19

Z-do

cosahe

xaen

oicacid

10R,17

S-

dihydroxy-

4Z,7Z,11

E,13

E,15

Z,19

Z-

docosahe

xaen

oicacid

4-�hydroxy-�

5E,�7Z,�10

Z,�13

Z,�16

Z,�19

Z-�

docosahe

xaen

oicacid

7-�hydroxy-�

4Z,�8E,�10

Z,�13

Z,�16

Z,�19

Z-�

docosahe

xaen

oicacid

TrivialN

ame

4,14-diHDH

A

[Neu

ro]

Protectin

D1

4-hydroxy-

DHA

7-hydroxy-

DHA

Chem

icalNam

e

4S,14S-

dihydroxy-

DHA

10R,17

S-

dihydroxy-

DHA

4-hydroxy-

DHA

7-hydroxy-

DHA

Form

ula

C 22H

32O4

C 22H

32O4

C 22H

32O3

C 22H

32O3

Structure

Abbreviatio

n

4S,14S-diHDH

A

PD1

4-HD

HA

7-HD

HA

COOH

OH

OH

COOH

OH

OH

COOH

OH

COOH

OH

18

Complete

Stereo

chem

istry

14-�hydroxy-�

4Z,�7Z,�10

Z,�12

E,�16

Z,�19

Z-�

docosahe

xaen

oicacid

17-�hydroxy-�

4Z,�7Z,�10

Z,�13

Z,�15

E,�19

Z-�

docosahe

xaen

oicacid

TrivialN

ame

14-hydroxy-

DHA

17-hydroxy-

DHA

Chem

icalNam

e

14-hydroxy-

DHA

17-hydroxy-

DHA

Form

ula

C 22H

32O3

C 22H

32O3

Structure

Abbreviatio

n

14-HDH

A

17-HDH

A

OH

COOH

COOH

OH

19

Aspirin-TriggeredSpecializedPro-resolvingMediators



©2019Serhanetal.

20

Complete

Stereo

chem

istry

5S,6R,15

R-

trihydroxy-

7E,9E,11

Z,13

E-eicosatetraeno

icacid

5S,14R

,15R

-trihydroxy-

6E,8Z,10

E,12

E-

eicosatetraeno

icacid

7S,8R,17

R-

trihydroxy-

4Z,9E,11

E,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

7S,16R

,17R

-trihydroxy-

4Z,8E,10

Z,12

E,14

E,19

Z-do

cosahe

xaen

oicacid

TrivialN

ame

15-epi-LipoxinA

4

15-epi-LipoxinB

4

Aspirin

-trig

gered

ResolvinD1

Aspirin

-trig

gered

ResolvinD2

Chem

icalNam

e

5S,6R,15

R-

trihydroxy-

LXA 4

5S,14R

,15R

-trihydroxy-

LXB 4

7S,8R,17

R-trihydroxy-

DHA

7S,16R

,17R

-trihydroxy-

DHA

Form

ula

C 20H

32O5

C 20H

32O5

C 22H

32O5

C 22H

32O5

Structure

Abbreviatio

n

AT-LXA

4

AT-LXB

4

AT-RvD

1

AT-RvD

2

COOH

OH

HO

OH

COOH

OH

HO

OH

COOH

OH

HO

HO

COOH

OH

HO

OH

21

Complete

Stereo

chem

istry

4S,11R

,17R

-trihydroxy-

5Z,7E,9E,13Z,15E,19Z-

docosahe

xaen

oicacid

4S,5,17R

-trihydroxy-

6E,8E,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

7S,17R

-dihydroxy-

4Z,8E,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

4S,17R

-dihydroxy-

5E,7Z,10

Z,13

Z,15

E,19

Z-

docosahe

xaen

oicacid

TrivialN

ame

Aspirin

-trig

gered

ResolvinD3

Aspirin

-trig

gered

ResolvinD4

Aspirin

-trig

gered

ResolvinD5

Aspirin

-trig

gered

ResolvinD6

Chem

icalNam

e

4S,11R

,17R

-trihydroxy-

DHA

4S,5,17R

-trihydroxy-

DHA

7S,17R

-dihydroxy-

DHA

4S,17R

-dihydroxy-

DHA

Form

ula

C 22H

32O5

C 22H

32O5

C 22H

32O4

C 22H

32O4

Structure

Abbreviatio

n

AT-RvD

3

AT-RvD

4

AT-RvD

5

AT-RvD

6

HOOC

OH

OH

OH

COOH

HO

HO

OH

HO

COOH

HO

OH

OH

HOOC

22

Complete

Stereo

chem

istry

10R,17

R-

dihydroxy-

4Z,7Z,11

E,13E,15

Z,19

Z-

docosahe

xaen

oicacid

TrivialN

ame

Aspirin

-trig

gered

[Neu

ro]

Protectin

D1

Chem

icalNam

e

10R,17

R-

dihydroxy-

DHA

Form

ula

C 22H

32O4

Structure

Abbreviatio

n

AT-PD1

COOH

OH

OH

23

DeuteriumLabeledInternalStandards



©2019Serhanetal.

24

Complete

Stereo

chem

istry

9-oxo-

11α,15S-

dihydroxy-

5Z,13E-

3,3,4,4-d 4-

prostadien

oicacid

5S,6R,15

S-

trihydroxy-

7E,9E,11

Z,13

E-

19,�19,�20,�20,�20-�d

5-

eicosatetraeno

icacid

5S,�12R

-�dihydroxy-�

6Z,�8E,�10

E,�14

Z-�

6,�7,�14,�15-�d 4-

eicosatetraeno

icacid

5S-�

hydroxy-�

6E,�8Z,�11

Z,�14

Z-�

5,6,8,9,11,12,14,15-�d 8-

eicosatetraeno

icacid

TrivialN

ame

d 4-Prostagland

inE

2

d 5-LipoxinA

4

d 4-Leu

kotriene

B4

d 8-5S-

hydroxy-

eicosatetraeno

ic

acid

Chem

icalNam

e

9-de

hydro-

prostaglandinF 2

α-d 4

5S,6R,15

S-

trihydroxy-

LXA 4-d

5

5S,12R

-dihydroxy-�

LTB 4-d

4

5S-

hydroxy-

ETE-d 8

Form

ula

C 20H

28D 4O5

C 20H

27D 5O5

C 20H

28D 4O4

C 20H

24D 8O3

Structure

Abbreviatio

n

d 4-PGE 2

d 5-LXA

4

d 4-LTB

4

d 8-5S-HE

TE

COOH

OH

OH

DD

D

D

COOH

OH

O

HO

DD

DD

COOH

OH

HO

OH

D

D

DD

D

COOH

OH

DD

DD

DD

DD

25

Complete

Stereo

chem

istry

7S,16R

,17S-

trihydroxy-

4Z,8E,10

Z,12

E,14

E,19

Z-19

,19,20

,20,20

,20-d 5-

docosahe

xaen

oicacid

TrivialN

ame

d 5-ResolvinD2

Chem

icalNam

e

7S,16R

,17S-

trihydroxy-

DHA-d

5

Form

ula

C 22H

27D 5O5

Structure

Abbreviatio

n

d 5-RvD

2

COOH

OH

HO

OH D D

D

DD

26

Methods



©2019Serhanetal.

27



©2019Serhanetal.

Auto-extraction(SolidPhaseExtraction)

Samples

LC-MS-MS

TargetedMRMandEPI

Datadependentscan/EMS/MSn

Identification

MatchingtoLMdatabase(RT+≥6diagnosticions)

Quantitation

Deuteratedinternalstandards

Synthesisofauthenticandsynthetic

deuteratedstandards

ConstructionofauthenticandsyntheticSPMandeicosanoids

database

MSn

Tissuevalidationandreferencehumansamples

(i.e.SRM1950)

Computationaldataanalysis

Clusteranalysis

Principalcomponentanalysis

Cells,tissues,organsandindividualprofiles

LMprofiles(>55)Targetedbioactivemetabolomes

Operationalizationoflipidmediatormetabololipidomics

Identificationandsignatureprofilesforpro-resolvingandinflammatorylipidmediatorsinhumantissueColasRA,ShinoharaM,DalliJ,ChiangN,SerhanCN.AmJPhysiolCellPhysiol.2014 28

Table1:HighPerformanceLiquidChromatographysettings.

Table 2:Mass Spectrometer settings forMultiple ReactionMonitoring and EnhanceProductIonforABSciex5500QTRAP.

ShimadzuLC20ADHPLCSettings

SolventA H2O(0.01%aceticacid)

SolventB Methanol(0.01%aceticacid)

Flowrate 0.4mL/min

Column EclipseplusC18,1.8µm,4.6x100mm,AGILENT

ColumnHeater ThermaSphereTS-130,50°C,PHENOMENEX

Gradient

Time(min) Solvent(%B)0.0 552.0 5510.0 8518.0 9820.0 9820.1 5524.5 End

ABSciex5500QTRAPMSsettingforMRMandEPI

MRM EPI CurtainGas 25 25 CollisionGas Medium Medium IonSprayVoltage -4000 -4000 Temperature 500°C 500°C IonSourceGas1 40 40 IonSourceGas2 40 40



©2019Serhanetal.

Table 3:Mass Spectrometer settings forMultiple ReactionMonitoring and EnhanceProductIonforABSciex6500QTRAP.

ABSciex6500QTRAPMSsettingforMRMandEPI

MRM EPICurtainGas 30 30CollisionGas Medium MediumIonSprayVoltage -4500 -4500Temperature 580°C 580°CIonSourceGas1 80 80IonSourceGas2 70 70

29

Table4:MassSpectrometersettingsforMultipleReactionMonitoringforABSciex5500QTRAP. DP=Declustering Potential, EP=Entrance potential, CE=Collision Energy,CXP=CollisionCellExitPotential



©2019Serhanetal.

ABSciex5500QTRAPIndividuallipidmediator,pathwaymarker,andfurthermetabolitesettingforMRM(V)

Compound DP EP CE CXPAA -90 -10 -41 -22PGD2 -80 -10 -30 -15PGE2 -80 -10 -30 -15PGF2α -80 -10 -30 -15TxB2 -80 -10 -30 -15LTB4 -110 -10 -22 -1720-OH-LTB4 -100 -10 -26 -1720-COOH-LTB4 -90 -10 -26 -17LXA4 -95 -10 -22 -11LXB4 -90 -10 -30 -215,15-diHETE -110 -10 -22 -175-HETE -90 -10 -21 -2212-HETE -70 -10 -21 -1515-HETE -65 -10 -19 -18 EPA -90 -10 -41 -22LXA5 -95 -10 -22 -11LXB5 -90 -10 -30 -215,15-diHEPE -110 -10 -22 -17RvE1 -80 -10 -24 -16RvE2 -40 -10 -22 -17RvE3 -40 -10 -22 -175-HEPE -60 -10 -22 -1112-HEPE -70 -10 -21 -1515-HEPE -65 -10 -19 -818-HEPE -65 -10 -18 -23 DHA -90 -10 -41 -22RvD1 -90 -10 -22 -13RvD2 -90 -10 -24 -13RvD3 -80 -10 -28 -19RvD5 -40 -10 -22 -17RvD6 -40 -10 -22 -17MaR1 -40 -10 -22 -174,14-diHDHA -40 -10 -22 -17PD1 -40 -10 -22 -174-HDHA -40 -10 -18 -137-HDHA -40 -10 -18 -1314-HDHA -70 -10 -21 -1517-HDHA -80 -10 -20 -18 d4-PGE2 -80 -10 -30 -11d4-LXA4 -135 -10 -24 -17d4-LTB4 -135 -10 -24 -17d8-5S-HETE -100 -10 -22 -11

d5-RvD2 -90 -10 -24 -13

30

ABSciex6500QTRAPIndividuallipidmediator,pathwaymarker,andfurthermetabolitesettingforMRM(V)

Compound DP EP CE CXPAA -10.0 -10.0 -10.0 -12.0PGD2 -30.0 -10.0 -22.0 -8.0PGE2 -30.0 -10.0 -22.0 -8.0PGF2a -60.0 -10.0 -33.3 -12.0TxB2 -70.0 -10.0 -25.0 -22.0LTB4 -45.0 -10.0 -21.0 -10.820-OH-LTB4 -60.0 -10.0 -22.0 -8.020-COOH-LTB4 -60.0 -10.0 -22.0 -8.0LXA4 -30.0 -10.0 -18.2 -10.0LXB4 -45.0 -10.0 -21.5 -12.05,15-diHETE -35.0 -10.0 -18.0 -12.05-HETE -30.0 -10.0 -18.0 -9.512-HETE -30.0 -10.0 -18.0 -9.515-HETE -30.0 -10.0 -18.0 -9.5EPA -10.0 -10.0 -10.0 -12.0LXA5 -30.0 -10.0 -18.6 -10.0LXB5 -30.0 -10.0 -21.2 -10.05,15-diHEPE -30.0 -12.0 -17.0 -12.0RvE1 -50.0 -10.0 -21.1 -10.0RvE2 -30.0 -12.0 -18.6 -10.0RvE3 -35.0 -10.0 -20.5 -14.05-HEPE -30.0 -10.0 -18.0 -9.512-HEPE -30.0 -10.0 -18.0 -9.515-HEPE -30.0 -10.0 -18.0 -9.518-HEPE -30.0 -10.0 -18.0 -9.5DHA -10.0 -10.0 -10.0 -12.0RvD1 -40.0 -10.0 -19.0 -9.0RvD2 -40.0 -10.0 -27.8 -10.0RvD3 -55.0 -10.0 -24.6 -9.5RvD5 -42.0 -10.0 -18.0 -11.0RvD6 -50.0 -10.0 -20.0 -11.0MaR1 -38.0 -10.0 -19.6 -11.04,14-diHDHA -50.0 -10.0 -20.0 -11.0PD1 -45.0 -10.0 -20.8 -9.54-HDHA -30.0 -10.0 -18.0 -9.57-HDHA -30.0 -10.0 -18.0 -9.514-HDHA -30.0 -10.0 -18.0 -9.517-HDHA -30.0 -10.0 -18.0 -9.5d4-PGE2 -30.0 -10.0 -26.0 -14.0d4-LXA4 -40.0 -10.0 -17.5 -7.5d4-LTB4 -55.0 -10.0 -21.7 -10.0d8-5S-HETE -40.0 -10.0 -24.0 -12.0d5-RvD2 -40.0 -10.0 -20.3 -13.0



©2019Serhanetal.

Table5:MassSpectrometersettingsforMultipleReactionMonitoringforABSciex5500QTRAP. DP=Declustering Potential, EP=Entrance potential, CE=Collision Energy,CXP=CollisionCellExitPotential 31



©2019Serhanetal.

Compoundname Linearequation r2 InternalstandardRvD1 y=0.00002081x 0.999 d5-RvD2RvD2 y=0.00009118x 0.997 d5-RvD2RvD3 y=0.000008163x 0.990 d5-RvD2RvD5 y=0.00001137x 0.993 d4-LTB4PD1 y=0.00001425x 0.995 d4-LTB4MaR1 y=0.00007534x 0.999 d4-LTB4

17-HDHA y=0.00001132x 0.996 d8-5HETE14-HDHA y=0.00001916x 0.999 d8-5HETE7-HDHA y=0.00001096x 0.996 d8-5HETE4-HDHA y=0.00001040x 0.996 d8-5HETELXA5 y=0.00002043x 0.999 d5-LXA4RvE1 y=0.00004358x 0.999 d5-LXA4RvE2 y=0.0001647x 0.996 d4-LTB4RvE3 y=0.0001047x 0.998 d4-LTB4

18-HEPE y=0.00002131x 0.998 d8-5HETE15-HEPE y=0.00002002x 0.999 d8-5HETE12-HEPE y=0.00001899x 0.997 d8-5HETE5-HEPE y=0.00001301x 0.998 d8-5HETELXA4 y=0.00001607x 0.996 d5-LXA4LXB4 y=0.00007325x 0.995 d5-LXA4

5S,15S-diHETE y=0.00008011x 0.997 d4-LTB4LTB4 y=0.00001074x 0.991 d4-LTB4

20-OH-LTB4 y=0.00001524x 0.999 d4-LTB420-COOH-LTB4 y=0.00007931x 0.999 d4-LTB4

PGD2 y=0.00001019x 0.993 d4-PGE2PGE2 y=0.00001698x 0.998 d4-PGE2PGF2α y=0.00001643x 0.997 d4-PGE2TXB2 y=0.00001054x 0.997 d4-PGE2

15-HETE y=0.00001377x 0.999 d8-5HETE12-HETE y=0.00001142x 0.994 d8-5HETE5-HETE y=0.00001010x 0.990 d8-5HETE

Table 6: Linear equation, coefficient of determination (r2) andattributedinternalstandard

32



©2019Serhanetal.

Recovery(%)inplasma

Interference(%signalremaining)

LOD5500(pg)in

LOD6500(pg)in

Internalstandards phase plasma phase plasmad5-RvD2 73 ± 16 28 ± 9 0.17 0.43 0.04 0.09d5-LXA4 75 ± 2 7 ± 2 0.10 0.25 0.02 0.05d4-LTB4 80 ± 3 18 ± 2 0.10 0.51 0.05 0.22d4-PGE2 63 ± 13 21 ± 6 0.19 0.47 0.09 0.23d8-5S-HETE 68 ± 3 28 ± 5 0.10 0.26 0.07 0.15d5-authenticlipidmediatorsd5-10S,17S-diHDHA 88 ± 4 8 ± 2 0.06 0.45 0.03 0.15d5-5S,15S-diHEPE 94 ± 6 9 ± 5 0.13 0.95 0.03 0.15d5-15-HEPE 63 ± 12 26 ± 4 0.09 0.23 0.10 0.19

Table7:Determinationofrecovery, interferenceandLOD.Plasmarecoveries, interferenceandlimit of detection (LOD) of the internal standards and authentic d5-lipid mediators used asrepresentativeofspecificchromatographicregionsandphysicalpropertiesofcompounds.LODwasassessedinphaseandinthematrix.Mean±SEM,n=6forrecoveriesandinterference,d=3forLOD.

33



©2019Serhanetal.

Compoundname

Nominalconcentration

(pg)

Intraday(repeatability) Interday(reproducibility)

Measured(pg) %RSD Accuracy Measured(pg) %RSD AccuracyRvD1 5 5.0 ± 0.1 1% 99% 4.6 ± 0.1 2% 92%

20 17.2 ± 0.7 7% 86% 23.2 ± 0.7 5% 84%100 109.8 ± 2.2 4% 90% 118.5 ± 1.8 3% 82%200 227.8 ± 2.5 2% 86% 233.5 ± 1.8 2% 83%

RvD2 5 5.2 ± 0.1 4% 96% 4.6 ± 0.1 4% 93%20 16.9 ± 0.7 7% 85% 22.6 ± 0.5 4% 87%100 113.0 ± 2.3 4% 87% 116.5 ± 1.6 3% 84%200 227.8 ± 3.1 3% 86% 235.5 ± 2.5 2% 82%

RvD3 5 5.7 ± 0.1 3% 86% 5.4 ± 0.1 4% 91%20 19.8 ± 0.5 4% 99% 26.0 ± 0.3 2% 70%100 118.3 ± 1.4 2% 82% 125.3 ± 0.9 2% 75%200 226.8 ± 2.2 2% 87% 226.8 ± 1.7 1% 87%

RvD5 5 4.8 ± 0.1 5% 97% 4.6 ± 0.04 2% 93%20 16.7 ± 0.6 6% 83% 24.0 ± 0.3 2% 80%100 114.3 ± 0.8 1% 86% 117.0 ± 0.0 0% 83%200 217.0 ± 2.6 2% 92% 217.0 ± 2.4 2% 92%

PD1 5 4.6 ± 0.1 5% 93% 4.4 ± 0.05 2% 88%20 16.7 ± 0.3 3% 84% 23.7 ± 0.5 4% 82%100 114.0 ± 0.7 1% 86% 118.5 ± 1.0 2% 82%200 218.5 ± 1.3 1% 91% 220.0 ± 1.3 1% 90%

MaR1 5 4.6 ± 0.2 9% 91% 4.2 ± 0.02 1% 85%20 16.1 ± 0.1 1% 81% 22.3 ± 0.5 4% 89%100 107.0 ± 3.5 7% 93% 110.8 ± 1.4 3% 89%200 221.3 ± 2.3 2% 89% 229.0 ± 1.8 2% 86%

17-HDHA 5 3.5 ± 0.1 4% 69% 4.4 ± 0.05 2% 87%20 17.8 ± 0.3 3% 89% 23.7 ± 0.8 6% 82%100 115.0 ± 1.1 2% 85% 120.0 ± 0.6 1% 80%200 222.0 ± 1.2 1% 89% 227.5 ± 1.6 1% 86%

14-HDHA 5 3.2 ± 0.1 5% 64% 4.3 ± 0.02 1% 86%20 17.3 ± 0.4 4% 86% 22.2 ± 0.4 3% 89%100 114.8 ± 2.4 4% 85% 109.5 ± 2.3 4% 91%200 212.3 ± 2.3 2% 94% 233.5 ± 0.9 1% 83%

7-HDHA 5 3.5 ± 0.1 4% 70% 4.5 ± 0.1 3% 90%20 19.7 ± 0.1 1% 99% 25.8 ± 0.4 3% 71%100 123.8 ± 2.3 4% 76% 128.0 ± 2.5 4% 72%200 236.5 ± 1.0 1% 82% 240.5 ± 2.4 2% 80%

4-HDHA 5 3.6 ± 0.1 6% 73% 4.6 ± 0.02 1% 91%20 19.8 ± 0.3 2% 99% 26.0 ± 0.3 2% 70%100 125.8 ± 2.0 3% 74% 128.5 ± 1.6 2% 72%200 236.3 ± 2.1 2% 82% 240.3 ± 1.4 1% 80%

Table8 (part 1/4): Precision andaccuracyof LMmetabololipidomics.%RSD, relative standarddeviation;%RSD=(SEM/mean)x100;mean±SEM;n=4

34



©2019Serhanetal.


Compoundname


(pg)


Measured(pg) %RSD Accuracy Measured(pg) %RSD AccuracyLXA5 5 5.6 ± 0.1 2% 87% 5.0 ± 0.1 4% 99%

20 18.0 ± 0.3 3% 90% 21.6 ± 0.7 6% 92%100 111.0 ± 2.6 5% 89% 110.3 ± 1.9 3% 90%200 212.3 ± 2.8 3% 94% 240.3 ± 3.3 3% 80%

RvE1 5 5.8 ± 0.1 4% 85% 5.1 ± 0.1 3% 99%20 17.4 ± 0.1 1% 87% 24.3 ± 0.3 2% 78%100 113.3 ± 2.3 4% 87% 121.8 ± 1.7 3% 78%200 242.0 ± 3.7 3% 79% 241.8 ± 1.7 1% 79%

RvE2 5 5.5 ± 0.4 14% 91% 4.7 ± 0.1 3% 93%20 16.4 ± 0.7 8% 82% 20.7 ± 0.7 6% 97%100 108.8 ± 2.8 5% 91% 101.2 ± 2.9 6% 99%200 199.3 ± 3.3 3% 100% 240.0 ± 1.1 1% 80%

RvE3 5 6.1 ± 0.3 8% 78% 5.0 ± 0.4 16% 99%20 17.8 ± 1.7 17% 89% 23.7 ± 1.6 12% 81%100 105.0 ± 3.0 6% 95% 116.5 ± 1.0 2% 84%200 218.5 ± 3.4 3% 91% 227.8 ± 3.1 3% 86%

18-HEPE 5 4.1 ± 0.1 3% 82% 4.3 ± 0.03 2% 87%20 17.6 ± 0.2 2% 88% 23.3 ± 0.5 4% 83%100 117.5 ± 2.0 3% 83% 121.3 ± 1.5 2% 79%200 239.0 ± 2.3 2% 81% 241.8 ± 1.5 1% 79%

15-HEPE 5 4.3 ± 0.1 3% 87% 4.4 ± 0.04 2% 88%20 17.6 ± 0.2 2% 88% 22.8 ± 0.5 4% 86%100 115.5 ± 2.5 4% 85% 122.3 ± 1.1 2% 78%200 231.3 ± 1.7 1% 84% 239.3 ± 1.7 1% 80%

12-HEPE 5 4.6 ± 0.04 2% 93% 4.4 ± 0.1 3% 88%20 18.1 ± 0.2 1% 91% 22.6 ± 0.4 3% 87%100 122.5 ± 1.3 2% 78% 117.3 ± 1.7 3% 83%200 231.3 ± 2.3 2% 84% 227.8 ± 1.3 1% 86%

5-HEPE 5 4.6 ± 0.05 2% 91% 4.6 ± 0.04 2% 92%20 18.1 ± 0.2 2% 91% 23.4 ± 0.5 4% 83%100 115.3 ± 1.3 2% 85% 119.5 ± 0.5 1% 81%200 223.5 ± 2.0 2% 88% 233.3 ± 1.0 1% 83%

35



©2019Serhanetal.

Compoundname


(pg)


Measured(pg) %RSD Accuracy Measured(pg) %RSD AccuracyLXA4 5 5.5 ± 0.1 2% 90% 4.9 ± 0.1 4% 98%

20 17.7 ± 0.8 8% 88% 23.5 ± 0.1 1% 82%100 114.0 ± 2.1 4% 86% 119.0 ± 1.1 2% 81%200 226.3 ± 2.1 2% 87% 233.5 ± 1.5 1% 83%

LXB4 5 6.2 ± 0.1 4% 76% 5.4 ± 0.1 3% 92%20 18.4 ± 0.6 6% 92% 22.9 ± 0.4 3% 85%100 116.8 ± 1.7 3% 83% 114.5 ± 1.2 2% 86%200 220.0 ± 4.5 4% 90% 231.8 ± 1.6 1% 84%

5S,15S-diHETE 5 5.1 ± 0.1 4% 98% 5.4 ± 0.1 2% 93%20 20.3 ± 0.5 4% 98% 26.4 ± 0.2 1% 68%100 123.3 ± 1.4 2% 77% 129.0 ± 2.0 3% 71%200 246.5 ± 3.7 3% 77% 251.0 ± 6.1 5% 75%

LTB4 5 5.0 ± 0.03 1% 99% 5.8 ± 0.1 2% 84%20 22.3 ± 0.3 2% 89% 27.5 ± 0.4 3% 63%100 125.0 ± 0.6 1% 75% 130.0 ± 1.8 3% 70%200 231.0 ± 3.2 3% 85% 226.8 ± 2.9 3% 87%

20-OH-LTB4 5 5.4 ± 0.1 4% 93% 4.9 ± 0.03 1% 98%20 17.1 ± 0.5 6% 86% 22.6 ± 0.4 3% 87%100 111.3 ± 2.7 5% 89% 116.0 ± 0.8 1% 84%200 223.3 ± 1.1 1% 88% 229.0 ± 3.0 3% 86%

20-COOH-LTB4 5 5.1 ± 0.4 16% 99% 5.2 ± 0.1 2% 97%20 18.0 ± 0.9 9% 90% 21.4 ± 1.6 13% 93%100 117.5 ± 1.3 2% 83% 114.0 ± 2.2 4% 86%200 227.3 ± 2.3 2% 86% 253.8 ± 3.4 3% 73%

PGD2 5 5.7 ± 0.1 2% 87% 5.2 ± 0.1 4% 96%20 17.1 ± 0.2 2% 86% 22.8 ± 0.3 3% 86%100 105.5 ± 2.3 4% 95% 115.5 ± 1.0 2% 85%200 213.3 ± 3.1 3% 93% 218.3 ± 3.7 3% 91%

PGE2 5 5.5 ± 0.1 4% 90% 4.9 ± 0.2 6% 97%20 18.0 ± 0.5 4% 90% 22.9 ± 0.9 7% 86%100 116.8 ± 4.0 7% 83% 113.8 ± 2.3 4% 86%200 206.5 ± 1.2 1% 97% 209.5 ± 4.6 4% 95%

PGF2α 5 5.5 ± 0.1 2% 90% 5.2 ± 0.1 3% 97%20 19.2 ± 0.5 4% 96% 25.6 ± 0.4 3% 72%100 120.3 ± 1.7 3% 80% 123.8 ± 1.1 2% 76%200 233.3 ± 3.1 3% 83% 220.8 ± 0.9 1% 90%

TXB2 5 4.7 ± 0.2 7% 94% 5.0 ± 0.01 0.3% 100%20 16.8 ± 0.8 8% 84% 22.2 ± 1.3 10% 89%100 111.2 ± 4.1 7% 89% 118.0 ± 4.5 8% 82%200 219.0 ± 3.6 3% 91% 189.5 ± 6.9 7% 95%


36



©2019Serhanetal.

Compoundname


(pg)


Measured(pg) %RSD Accuracy Measured(pg) %RSD Accuracy15-HETE 5 4.1 ± 0.1 2% 82% 4.2 ± 0.05 2% 84%

20 16.6 ± 0.2 2% 83% 21.8 ± 0.4 3% 91%100 109.3 ± 2.6 5% 91% 110.8 ± 1.3 2% 89%200 212.8 ± 2.3 2% 94% 219.0 ± 0.8 1% 91%

12-HETE 5 4.5 ± 0.04 2% 89% 4.8 ± 0.04 2% 95%20 17.7 ± 0.3 2% 89% 23.1 ± 0.6 4% 85%100 113.8 ± 1.8 3% 86% 117.0 ± 1.1 2% 83%200 216.5 ± 1.8 2% 92% 220.0 ± 1.7 2% 90%

5-HETE 5 4.9 ± 0.05 2% 97% 5.0 ± 0.1 4% 100%20 20.3 ± 0.3 3% 98% 25.7 ± 0.3 2% 72%100 125.0 ± 3.0 5% 75% 127.8 ± 1.4 2% 72%200 232.5 ± 1.4 1% 84% 239.5 ± 4.7 4% 80%

Table 8 (part 4/4): Precision and accuracy of LMmetabololipidomics. %RSD, relative standarddeviation;%RSD=(SEM/mean)x100;mean±SEM;n=4

37



©2019Serhanetal.Day-0 Day-1 Day-2 Day-3 Day-4 Day-5 2-weeks 4-weeks

DHAbioactivemetabolome Δchange(%)RvD1 49.5±1.3

with -1.0 -0.2 +3.2 -3.2 -1.1without +0.8 -2.0 +2.4 -13.2 -15.5 -33.4 -56.8

RvD2 13.4±0.8with -1.3 +1.1 +6.4 +2.3 +1.1

without -9.2 +2.5 +2.2 -1.2 -19.2 -39.5 -59.0RvD3 68.5±7.5

with -3.1 +8.0 +6.6 -4.4 +6.0without -6.8 -8.5 -3.3 -11.3 -12.6 -18.9 -25.4

RvD5 24.9±2.2with +6.5 -10.5 -18.0 -8.4 -16.8

without -16.4 -14.8 -15.1 -20.2 -17.0 -38.7 -44.0RvD6 21.6±2.0

with +1.7 +2.2 -1.1 +0.4 -11.8without +4.6 -9.8 -12.5 -17.2 -14.3 -19.5 -85.3

PD1 6.1±0.1with -3.0 +5.5 -8.3 -11.9 -10.4

without +6.7 +1.7 -18.3 -26.8 -23.7 -33.0 -36.7MaR1 9.0±0.8

with +2.3 +0.9 -8.0 -6.5 -17.8without -6.5 -2.5 -2.6 -15.7 -24.7 -67.9 -73.1

EPAbioactivemetabolomeRvE1 13.1±0.9

with -3.3 -9.0 -13.9 -9.4 -13.1without +7.6 -4.2 -22.0 -23.7 -24.6 -50.7 -57.1

RvE3 686.3±51.0with -6.4 -7.6 -5.1 -7.2 -4.4

without -1.3 -5.2 -4.5 -3.5 -18.7 -40.9 -42.2AAbioactivemetabolomeLXA4 11.2±1.0

with 1.8 +0.5 -8.6 -8.3 -5.9without 4.4 -3.8 -5.2 -5.7 -3.8 -14.4 -39.9

LXB4 117.6±2.6with -1.7 -3.6 -1.9 +5.2 -5.5

without -1.3 -5.8 -3.6 -5.8 -11.3 -17.7 -57.1PGD2 55.1±1.9

with +9.8 -6.8 +3.3 +3.7 -15.5without +7.6 -2.8 +5.8 +2.1 -16.9 -10.9 -26.7

PGE2 199.2±10.2with +3.7 -1.2 +0.1 -11.8 -15.6

without -6.2 -12.2 -14.3 -20.8 -15.3 -11.9 -29.8PGF2a 143.3±2.5

with -0.4 -0.2 +1.2 +8.9 -13.6without +8.0 +1.1 +1.6 +4.8 -9.0 -15.6 -34.1

TxB2 6067.0±128.4with -5.2 -7.8 +7.4 -5.5 -12.5

without +9.2 -3.2 +7.3 -16.6 -19.2 -17.8 -34.2

Table 9: Human serum (250µl) was stored under -30°Cwith or without 1mlmethanol. On thedesignated day (Day-0 to 4-weeks), LC-MS-MS based LM metabololipidomics were carried out.Resultsareexpressedaspg/mlserum;mean±SEMofd=3determinationsandΔchange(%)fromDay-0. 38

MultipleReactionMonitoringTransitions



©2019Serhanetal.

39

Table1:MRMtransitionforLC-MS-MSbasedmetabololipidomicsofAA,EPAandDHAderivedlipidmediators.

Table2:MRMtransitionforLC-MS-MSbasedmetabololipidomicsofAAandDHAderivedaspirin-triggeredlipidmediators.

Compound Q1 Q3 Compound Q1 Q3

AA 303.3 259.1 15-HEPE 317.2 219.1PGD2 351.3 233.2 18-HEPE 317.2 259.1PGE2 351.3 175.1 PGF2α 353.3 193.1 DHA 327.3 283.1TXB2 369.3 169.1 RvD1 375.2 215.2LTB4 335.3 195.2 RvD2 375.2 141.2LXA4 351.2 235.1 RvD3 375.2 147.1LXB4 351.2 221.1 RvD5 359.2 199.15,15-diHETE 335.2 115.2 RvD6 359.2 159.15-HETE 319.2 115.1 MaR1 359.2 250.112-HETE 319.2 179.1 4,14-diHDHA 359.2 221.115-HETE 319.2 219.1 PD1 359.2 153.1

4-HDHA 343.2 101.1EPA 301.3 257.1 7-HDHA 343.2 141.1LXA5 349.2 215.1 14-HDHA 343.2 205.1LXB5 349.2 221.2 17-HDHA 343.2 245.15,15-diHEPE 333.2 235.2 RvE1 349.2 195.1 d4-PGE2 355.3 193.2RvE2 333.3 253.2 d5-LXA4 356.3 115.2RvE3 333.3 201.1 d4-LTB4 339.3 197.25-HEPE 317.2 115.1 d8-5S-HETE 327.3 116.112-HEPE 317.2 179.1 d5-RvD2 380.2 141.2


AT-LXA4 351.2 235.1 AT-RvD3 375.2 147.1AT-LXB4 351.2 221.1 AT-RvD5 359.2 199.1

AT-RvD6 359.2 159.1AT-RvD1 375.2 215.2AT-RvD2 375.2 141.2 AT-PD1 359.2 153.1



©2019Serhanetal.

40

Additionalreferences:Specific lipid mediator signatures of human phagocytes: microparticles stimulatemacrophageefferocytosisandpro-resolvingmediators.DalliJ,SerhanCN.Blood.2012120(15)Metabolomics-lipidomicsofeicosanoidsanddocosanoidsgeneratedbyphagocytes.YangR,ChiangN,OhSF,SerhanCN.CurrProtocImmunol.2011Chapter14:Unit14.26.

Table3:MRMtransitionforLC-MS-MSbasedmetabololipidomicsoffurthermetabolitesofAA,EPAandDHAderivedlipidmediators.


20-OH-LTB4 351.2 195.1 Dihydro-RvD1 377.2 279.1 20-COOH-LTB4 365.2 195.1 7-oxo-RvD2 373.2 260.1 15-oxo-LXA4 349.2 233.1 16-oxo-RvD2 373.2 260.1 13,14-dihydro-LXA4 353.2 253.1 22-OH-RvD2 391.2 249.1 5-oxo-LXB4 349.2 219.1 Dihydro-RvD2 377.2 279.1

14-oxo-MaR1 357.2 221.1 18-oxo-RvE1 347.2 213.1 7-oxo-MaR1 357.2 219.1 19-OH-RvE1 365.2 205.1 22-OH-MaR1 375.2 177.1 20-OH-RvE1 365.2 291.1 22-COOH-MaR1 389.2 177.1 20-COOH-RvE1 379.2 237.1 Dihydro-MaR1 361.2 223.1 Dihydro-RvE1 351.2 225.1 22-OH-PD1 375.2 153.1

22-COOH-PD1 389.2 153.1 8-oxo-RvD1 373.2 231.1 Dihydro-PD1 361.2 263.1 17-oxo-RvD1 373.2 231.1



©2019Serhanetal.

41

LipidMediatorBiosyntheticPathways



©2019Serhanetal.

42

Keyeicosanoidsthatplaypivotalrolesininitiatinginflammationanditsresolution.

Anti-Inflammatory and Pro-Resolving LipidMediators.Charles N. Serhan, Stephanie Yacoubian and RongYangAnnuRevPathol.20083:279–312.PMCID:PMC2739396

BiosyntheticschemefortheProstaglandins,LeukotrienesandLipoxins



©2019Serhanetal.

Circledenoteshumanlipoxygenases,inmice12/15LOXisasingleenzymeNOTfoundinhumantissue.

43

BiosyntheticschemefortheLeukotrienes



©2019Serhanetal.

Leukotrienes and lipoxins: structures, biosynthesis, and biological effects. Samuelsson B, Dahlén SE,LindgrenJA,RouzerCA,SerhanCNScience.1987237(4819):1171-6PMID:2820055

44

ResolvinsandProtectinsinInflammation-ResolutionCharlesN.Serhan,NicosA.PetasisChemRev.201112111(10):5922–5943.PMCID:PMC3192290

E-seriesresolvinbiosynthesisandmetabolicinactivation.

BiosyntheticschemefortheE-seriesResolvins



©2019Serhanetal.

45


BiosyntheticschemefortheD-seriesResolvins

PathwaysandenzymesinthebiosynthesisofD-seriesresolvins.



©2019Serhanetal.

46



©2019Serhanetal.

ResolvinD3andaspirin-triggeredresolvinD3arepotentimmunoresolventsDalliJ,WinklerJW,ColasRA,ArnardottirH,ChengCY,ChiangN,PetasisNA,SerhanCNChemBiol.201320(2):188-201.PMCID:23438748ResolvinD4stereoassignmentanditsnovelactionsinhostprotectionandbacterialclearanceWinklerJW,OrrSK,DalliJ,ChengCY,SangerJM,ChiangN,PetasisNA,SerhanCNSciRep.20166:18972.PMCID:26743932ResolvinD3multi-levelproresolvingactionsarehostprotectiveduringinfectionPaulC.Norris,HildurArnardottir,JuliaM.Sanger,DavidFichtner,GregoryS.Keyes,CharlesN.SerhanPLEFA.2016InPress,AcceptedManuscript

BiosyntheticschemefortheD-seriesResolvins

47


BiosyntheticschemefortheaspirintriggeredD-seriesResolvins

Pathwaysandenzymesinvolvedinthebiosynthesisofaspirin-triggeredD-seriesresolvins.



©2019Serhanetal.

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©2019Serhanetal.


Novelproresolvingandtissue-regenerativeresolvinandprotectinsulfido-conjugatedpathwaysDalliJ,RamonS,NorrisPC,ColasRA,SerhanCNFASEBJ.201529(5):2120-36PMCID:25713027

IdentificationandActionsofANovelThirdMaresinConjugateinTissueRegeneration:MCTR3.DalliJ,SangerJM,RodriguezAR,ChiangN,SpurBW,SerhanCN.PLOSONE2016;inpress.

BiosyntheticschemefortheProtectins

49



©2019Serhanetal.

BiosyntheticschemefortheMaresins

Maresins:novelmacrophagemediatorswithpotentantiinflammatoryandproresolvingactions.SerhanCN,YangR,MartinodK,KasugaK,PillaiPS,PorterTF,OhSF,SpiteM.JExpMed.2009206(1):15-23.PMCID:PMC2626672

Novelproresolvingandtissue-regenerativeresolvinandprotectinsulfido-conjugatedpathwaysDalliJ,RamonS,NorrisPC,ColasRA,SerhanCNFASEBJ.201529(5):2120-36PMCID:25713027

TheProtectinPCTR1enhancesresolutionofinfectiousinflammationandisproducedbyhumanM2macrophages.RamonS,DalliJ,SangerJM,WinklerJW,AursnesM,TungenJE,HansenTV,SerhanCN.AmJPathol.2016;inpress. 50

LC-MS-MSSpectra



©2019Serhanetal.

51


Prostaglandins&Thromboxane



©2019Serhanetal.

52

PGD2

10.6

351=M-H333=M-H-H2O315=M-H-2H2O289=M-H-H2O-CO2271=M-H-2H2O-CO2235=279-CO2233=251-H2O217=279-H2O-CO2189=251-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

271

189

315

233

333

351289217 251235

100 140 180 220 260 300 340 380m/z,Da

+H251 -H

279

HO

O

COO-

OH



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PGE2

10.5

351=M-H333=M-H-H2O315=M-H-2H2O289=M-H-H2O-CO2271=M-H-2H2O-CO2243=279-2H2O235=279-CO2233=251-H2O217=279-H2O-CO2207=251-CO2189=251-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

271

315 333

189

233 289217 351243

235

207 279

100 140 180 220 260 300 340 380m/z,Da

+H251 -H

279

O

HO

COO-

OH



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PGF2α

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

10.9

353=M-H335=M-H-H2O317=M-H-2H2O299=M-H-3H2O291=309-H2O273=309-2H2O263=281-H2O235=253-H2O219=281-H2O-CO2217=253-2H2O209=253-CO2193=253-H-CH3-CO2191=253-H2O-CO2173=253-2H2O-CO2

253209 281

353

309

335291

317273 299191 219 235173

217

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

HO

HO

COO-

OH

253281

+H-H

193

309

263



©2019Serhanetal.

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TxB2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

10.1

369=M-H333=M-H-2H2O325=M-H-CO2315=M-H-3H2O307=M-H-H2O-CO2289=M-H-2H2O-CO2279=297-H2O233=269-2H2O195=213-H2O189=269-2H2O-CO2177=213-2H2O151=169-H2O133=169-2H2O125=169-CO2

169

195

177

125 289

151369307

315 333279 325189133 233

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

269297

+H-H

213

169

O

OH

HO

COO-

OH

297



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56


Leukotrienes



©2019Serhanetal.

57

COO-

OHOH

LTB4

13.3

335=M-H317=M-H-H2O299=M-H-2H2O291=M-H-CO2273=M-H-H2O-CO2255=M-H-2H2O-CO2205=223-H2O179=223-CO2177=195-H2O161=223-H2O-CO2151=195-CO2133=195-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

335

195

317

151273129

111 177 299255205133

291115

179

219161

100 140 180 220 260 300 340 380m/z,Da

115

219-H

223

111-H

+H195

270

261 281λmax

MeOH=270nmε≈40,000abs.�ℓ/mol.

Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 330310

Wavelength,nm



©2019Serhanetal.

129

58

20-OH-LTB4

8.8

351=M-H333=M-H-H2O315=M-H-2H2O289=M-H-H2O-CO2271=M-H-2H2O-CO2179=223-CO2177=195-H2O151=195-CO2133=195-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

351

195

333

151 315289179177

271133115

100 140 180 220 260 300 340 380m/z,Da

COO-

OHOH

OH

270

261 281λmax


Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 330310

115195

223-H

+H-H

Wavelength,nm



©2019Serhanetal.

129

129

59

20-COOH-LTB4

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

7.75

365=M-H347=M-H-H2O329=M-H-2H2O321=M-H-CO2303=M-H-H2O-CO2285=M-H-2H2O-CO2205=223-H2O177=195-H2O161=223-H2O-CO2151=195-CO2

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

365

347

151

329303

285141 161 205 249177 321115

195

100 140 180 220 260 300 340 380m/z,Da

270

261281

λmaxMeOH=270nm

ε≈40,000abs.�ℓ/mol.

Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 330310

115195

223141 -H

+H-H 249

Wavelength,nm



©2019Serhanetal.

129

129

COO-

COOH

OH OH

60


Lipoxins



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LXA4

11.1

351=M-H333=M-H-H2O315=M-H-2H2O307=M-H-CO2289=M-H-H2O-CO2271=M-H-2H2O-CO2261=279-H2O233=251-H2O217=235-H2O215=251-2H2O207=251-CO2199=235-2H2O189=251-H2O-CO2127=145-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

115 351

217

235

289189271

307 333315251207127 145 233 261215199

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

115

+H

235-H-H

145

251

279-H

COO-

HO OH

OH

301

288

315

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

Wavelength,nm



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62

LXB4

10.5

351=M-H333=M-H-H2O315=M-H-2H2O307=M-H-CO2297=M-H-3H2O289=M-H-H2O-CO2271=M-H-2H2O-CO2261=279-H2O243=279-2H2O233=251-H2O217=235-H2O215=251-2H2O207=251-CO2203=221-H2O189=251-H2O-CO2177=221-CO2159=221-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

351

221

315

233

333

189 251207271203177

159 289297215

243 307261217115

100 140 180 220 260 300 340 380m/z,Da

279251

-H221

115

235

+H

-H

+H

COO-

OH

HO OH

301

288

314

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

Wavelength,nm



©2019Serhanetal.

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13.0

5S,15S-diHETE

335=M-H317=M-H-H2O299=M-H-2H2O291=M-H-CO2273=M-H-H2O-CO2255=M-H-2H2O-CO2227=263-2H2O217=235-H2O201=219-H2O191=235-CO2173=235-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

255

317

335

173

201

115

273235

299217

191291

227219

100 140 180 220 260 300 340 380m/z,Da

COO-

OH

OH

219

235

115

-H+H263

-H

244

λmaxMeOH=244nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

64


MonoHydroxyETEProducts



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65

5-HETE

16.4

319=M-H301=M-H-H2O275=M-H-CO2257=M-H-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

257

319

301

115

203

275

100 140 180 220 260 300 340 380m/z,Da

115232-H

203

-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

232

Wavelength,nm



©2019Serhanetal.

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12-HETE

15.9

319=M-H301=M-H-H2O275=M-H-CO2257=M-H-H2O-CO2163=207-CO2145=207-H2O-CO2135=179-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

319

179257

301

135

163275

145

207

100 140 180 220 260 300 340 380m/z,Da

-H207

+H

179

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



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15-HETE

15.6

319=M-H301=M-H-H2O275=M-H-CO2257=M-H-H2O-CO2203=247-CO2175=219-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

319

301

257

175

219

203 275

247

100 140 180 220 260 300 340 380m/z,Da

219+H

247-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



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68


Lipoxins



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69

LXA5

10.2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

349=M-H331=M-H-H2O313=M-H-2H2O305=M-H-CO2287=M-H-H2O-CO2269=M-H-2H2O-CO2217=279-H2O-CO2215=233-H2O207=251-CO2189=251-H2O-CO2171=251-2H2O-CO2

100 140 180 220 260 300 340 380m/z,Da

301

288

315λmax


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

349

COO-

OH

HO OH

115

+H

233-H-H

145

251279

-H

215 233

115

269 287 305313

331145 251

189

207217171

Wavelength,nm



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-H

5S,15S-diHEPE

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

333

100 140 180 220 260 300 340 380m/z,Da

COO-

OH

OH

246115217-H

235

+H -H263

246

115

217

235

315

297

289

271

253199

333=M-H315=M-H-H2O297=M-H-2H2O289=M-H-CO2271=M-H-H2O-CO2253=M-H-2H2O-CO2245=263-H2O219=263-CO2201=263-H2O-CO2199=217-H2O191=235-CO2173=235-H2O-CO2

191

173

219

201

245

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

244

λmaxMeOH=244nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

12.2

Wavelength,nm



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71


E-SeriesResolvins



©2019Serhanetal.

72

RvE1

349=M-H331=M-H-H2O313=M-H-2H2O305=M-H-CO2295=M-H-3H2O287=M-H-H2O-CO2273=291-H2O269=M-H-2H2O-CO2255=291-2H2O229=291-H2O-CO2205=223-H2O187=223-2H2O179=223-CO2177=195-H2O161=223-H2O-CO2151=195-CO2133=195-H2O-CO2

291

195

223

115

+H

+H

-H

-H

8.3

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

349

195

331

161

205291

313179 287273187151 295115 255229177 305269

100 140 180 220 260 300 340 380m/z,Da

271

262 282λmax


Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 330310

237

Wavelength,nm



©2019Serhanetal.

COO-

HO

OH

OH

73

11.9

RvE2

333=M-H315=M-H-H2O297=M-H-2H2O289=M-H-CO2271=M-H-H2O-CO2257=275-H2O253=M-H-2H2O-CO2231=275-CO2213=275-H2O-CO2199=217-H2O173=217-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

333

271

213 253

115315

199

275297

257 289231217173

100 140 180 220 260 300 340 380m/z,Da

115217-H

275

+H

COO-

HO

OH

243

λmaxMeOH=243nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

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RvE3

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

303275

-H245+H

+H

333

315

297

289

271

253

245

201

275

257

231

213

259

333=M-H315=M-H-H2O297=M-H-2H2O289=M-H-CO2271=M-H-H2O-CO2259=303-CO2257=275-H2O253=M-H-2H2O-CO2231=275-CO2213=275-H2O-CO2201=245-CO2

270

260 281λmax


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

13.2

COO-

HO OH

Wavelength,nm



©2019Serhanetal.

75


MonoHydroxyEPEProducts



©2019Serhanetal.

76

5-HEPE

15.4


Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

255

317115

201299

273

100 140 180 220 260 300 340 380m/z,Da

115201-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

77

12-HEPE

15.0

317=M-H299=M-H-H2O273=M-H-CO2255=M-H-H2O-CO2189=207-H2O163=207-CO2145=207-H2O-CO2135=179-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

317

273145109

179

255

135

299163

207

189

100 140 180 220 260 300 340 380m/z,Da

179+H

207-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

78

15-HEPE

14.8

COO-

OH -H

247219

+H


Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

317

255

219175

299

247

203 273

185229

100 140 180 220 260 300 340 380m/z,Da

237

λmaxMeOH=237nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

79

18-HEPE

14.7

317=M-H299=M-H-H2O273=M-H-CO2255=M-H-H2O-CO2243=287-CO2215=259-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

317

255

215

299

259

273

243

100 140 180 220 260 300 340 380m/z,Da

259

+H

287-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

80


D-SeriesResolvins



©2019Serhanetal.

81

RvD1

11.1

375=M-H357=M-H-H2O339=M-H-2H2O331=M-H-CO2313=M-H-H2O-CO2295=M-H-2H2O-CO2261=305-CO2259=277-H2O243=305-H2O-CO2241=277-2H2O215=233-H2O197=233-2H2O185=203-H2O153=171-H2O135=171-2H2O127=171-CO2123=141-H2O

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

215

375

233

141

135

277357171 295 331

313123 197 259 339241153127 185 243

113

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

COO-

HO

OH

HO

141

233-H

+H277

171-H

113+H

-H305

203

301

288

316

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

261

Wavelength,nm



©2019Serhanetal.

82

COO-

HO

OH

OH

RvD2

10.6

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

375

128

277

295

233 357339197203

313 331185

215

110321

375=M-H357=M-H-H2O339=M-H-2H2O331=M-H-CO2321=M-H-3H2O313=M-H-H2O-CO2295=M-H-2H2O-CO2287=305-H2O259=277-H2O241=277-2H2O215=233-H2O203=247-CO2197=233-2H2O185=247-H2O-CO2110=128-H2O

233

+H247

277+H

305

-H

-H262

128-H

301

288

315

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

287

259

241

141-H

141

247

Wavelength,nm



©2019Serhanetal.

83

RvD3

277+H

195

181165

209-H

+H

101-H

375

147

137357

165 215181 277 313295259159 177 339101 191

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 120 140 160 180 200 220 240 260 280 300 320 340 360 380m/z,Da

375=M-H357=M-H-H2O339=M-H-2H2O313=M-H-H2O-CO2295=M-H-2H2O-CO2259=277-H2O215=277-H2O-CO2191=209-H2O177=195-H2O159=195-2H2O147=165-H2O137=181-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

10.8

220 240 260 280 300 320200

Rel.Ab

s.(%

)

100%

0%

271

λmaxMeOH=271nm


281260

234

Wavelength,nm



©2019Serhanetal.

OH

HO

COO-HO

84

100 120 140 160 180 200 220 240 260 280 300 320 340 360 380 400

375

357

255 331

241313

295 339

225

277

259

306

375=M-H357=M-H2O339=M-H-2H2O331=M+H-CO2313=M-H-H2O-CO2295=M-H-2H2O-CO2259=277-H2O255=273-H2O241=277-2H2O237=273-2H2O233=277+H-CO2225=243-H2O215=277-H2O-CO2113=131-H2O

273

+H

101

101

113

131

243

215

RvD4

12.14275

226

266λmax


0%

Rel.Ab

s.(%

)

50%

100%

230 250 270 290215 310

Wavelength,nm



©2019Serhanetal.

285

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

131

243

306

277

101

-H

-H

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

273

85

COO-

HO

HO

141

-H289

+H

+H246

113

261

-H

-H217

RvD5

13.0

359=M-H341=M-H-H2O323=M-H-2H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2245=289-CO2243=261-H2O228=246-H2O227=289-H2O-CO2199=217-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

199

279 359

141 341297

246 261217113 228

245315323

227

243

100 140 180 220 260 300 340 380m/z,Da

242

λmaxMeOH=242nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

86

RvD6

359=M-H341=M-H-H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2245=289-CO2243=261-H2O239=257-H2O227=289-H2O-CO2217=261-CO2209=289-2H2O-CO2199=261-H2O-CO2

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

359

341

315

297

279

289

261

101

257

239

243

217

199

245

227

101

257

-H

+H261

-H 289

209

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

240

λmaxMeOH=240nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

13.4

Wavelength,nm



©2019Serhanetal.

OH

OH

-OOC

87


Maresins



©2019Serhanetal.

88

OHOH

COO-

MaR1

250 113

246-H

221

+H

+H 141-H

13.1

359=M-H341=M-H-H2O323=M-H-2H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2232=250-H2O228=246-H2O203=221-H2O177=221-CO2159=221-H2O-CO2123=141-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

359

177

250297

123 341

228113 246221 315232 279

141 323

203159

100 140 180 220 260 300 340 380m/z,Da

270

261281

λmaxMeOH=270nm


Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 330310

Wavelength,nm



©2019Serhanetal.

89

13.6

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

244

λmaxMeOH=244nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

4S,14S-diHDHA

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

359

341

100 140 180 220 260 300 340 380m/z,Da

297

279

101

221

138109 323315

239

213203

187

159

177

359=M-H341=M-H-H2O323=M-H-2H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2239=257-H2O231=249-H2O213=249-2H2O203=221-H2O187=249-H2O-CO2177=221-CO2159=221-H2O-CO2

231

COO-

OH

OH

249-H

221+H

101

-H

257

-H138

109

Wavelength,nm



©2019Serhanetal.

90


Protectins



©2019Serhanetal.

91

PD1

13.0

COO-

OH

OH

153

206-H

+H

-H

+H

177 181

261

359=M-H341=M-H-H2O323=M-H-2H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2243=261-H2O217=261-CO2199=261-H2O-CO2188=206-H2O159=177-H2O137=181-CO2119=181-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

153

359

137206

297188

341261217159 323315

199119177

181 279243

100 140 180 220 260 300 340 380m/z,Da

270

261 281λmax


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

92


MonoHydroxyDHAProducts



©2019Serhanetal.

93

4-HDHA

16.7


Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

343

281

101

299

325

241

100 140 180 220 260 300 340 380m/z,Da

101241-H

COO-

OH

235

λmaxMeOH=235nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

94

7-HDHA

16.1

343=M-H325=M-H-H2O299=M-H-CO2281=M-H-H2O-CO2123=141-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

281

343

201

141

325

299113

123 230

100 140 180 220 260 300 340 380m/z,Da

141201

113113

230-H

+H

-H

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

95

14-HDHA

15.9

343=M-H325=M-H-H2O299=M-H-CO2281=M-H-H2O-CO2215=233-H2O189=233-CO2161=205-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

343

281

161

325205

299

233189

215

100 140 180 220 260 300 340 380m/z,Da

233-H

205+H

COO-

OH

237

λmaxMeOH=237nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

96

17-HDHA

15.7


Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

343

255211

281

201

325245

299

229 273

100 140 180 220 260 300 340 380m/z,Da

-H273

+H245

COO-

OH

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

Wavelength,nm



©2019Serhanetal.

97



©2019Serhanetal.

98

DocosapentaenoicAcidMetabolome

n3-DPAResolvins

RvD1n3-DPA

11.8

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

215

377

297143

233359

341315333

279

377=M-H359=M-H-H2O341=M-H-2H2O333=M-H-CO2315=M-H-H2O-CO2297=M-H-2H2O-CO2263=307-CO2261=279-H2O215=233-H2O

301

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

OH

OHHO

COO-279+H

143

233 -H

99

RvD5n3-DPA

14.0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

361=M-H343=M-H-H2O325=M-H-2H2O299=M-H-H2O-CO2281=M-H-2H2O-CO2245=263-H2O199=217-H2O

281

343

361

143

325199

263299

OH

OH

COO-263

+H

143217

-H

λmaxMeOH=244nm


Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

244

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

100



©2019Serhanetal.

101


n3-DPAMaresins

MaR1n3-DPA

14.1

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

361=M-H343=M-H-H2O325=M-H-2H2O317=M-H-CO2299=M-H-H2O-CO2233=252-H2O125=143-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

343

223

361

143 299233

325125

317

244

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

271

λmaxMeOH=271nm


+H

OHOH

COO-

223

252-H

143

102



©2019Serhanetal.

103


n3-DPAProtectins

PD1n3-DPA

14.2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

361=M-H343=M-H-H2O325=M-H-2H2O317=M-H-CO2299=M-H-H2O-CO2281=M-H-2H2O-CO2245=263-H2O219=263-CO2

361

343263

183299 325

155

245219177

317281

OH

OH

COO-

-H183263

177

155

+H

+H

271

244

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

λmaxMeOH=271nm


104



©2019Serhanetal.

105


n3-DPA13-seriesResolvins

RvT1

10.7

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

377=M-H359=M-H-H2O341=M-H-2H2O315=M-H-H2O-CO2301=319-H2O297=M-H-2H2O-CO2215=233-H2O193=211-H2O149=211-H2O-CO2125=143-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

211

143

193

359 377

297 341

215 301

233

315 239 125

149

269

λmaxMeOH=269nm


COO-

OH OH

OH

-H

233

239

211

319

143

+H

+H

+H

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

106

RvT2

12.04

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

377=M-H359=M-H-H2O341=M-H-2H2O333=M-H-CO2315=M-H-H2O-CO2297=M-H-2H2O-CO2227=263-2H2O209=263-3H2O207=225-H2O215=233-H2O163=181-H2O157=255-3H2O-CO2125=143-H2O

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

143

227

255

377

207

341359181

157

315297333163

209

215

125

COO-

OH

OHOH

+H

225

197

255

233 143263

181

-H

-H -H

λmaxMeOH=240nm


Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

240

λmaxMeOH=240nm


107

RvT4

13.8

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

361=M-H343=M-H-H2O325=M-H-2H2O299=M-H-H2O-CO2281=M-H-2H2O-CO2221=239-H2O211=247-2H2O199=217-H2O193=211-H2O177=239-H2O-CO2149=211-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

143

211

281

193

361

199

343

325217

221299

COO-

OHOH-H

143

+H

-H

217

211

239 247

240

λmaxMeOH=240nm


108



©2019Serhanetal.

109


n3-DPAMonoHydroxyDPAProducts

17-HDPA

17.6

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm



©2019Serhanetal.

345=M-H327=M-H-H2O283=M-H-H2O-CO2231=275-CO2213=275-H2O-CO2203=247-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

283

345

327

247

203

275

236

Rel.Ab

s.(%

)

0%

50%

100%

240 260 280 300 320220 340 360

COO-

OH247

235

275

+H

-H

110

λmaxMeOH=236nm


Aspirin-TriggeredSpecialPro-resolvingMediators



©2019Serhanetal.

111

AT-LXA4

11.3

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

351

115

217

235

189 289307

271 333315

233

127

251199145

171279215

207243 261

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

351=M-H333=M-H-H2O315=M-H-2H2O307=M-H-CO2289=M-H-H2O-CO2271=M-H-2H2O-CO2261=279-H2O243=279-2H2O233=251-H2O217=235-H2O215=251-2H2O207=251-CO2199=235-2H2O189=251-H2O-CO2171=251-2H2O-CO2127=145-H2O

100 140 180 220 260 300 340 380m/z,Da

COO-

HO OH

OH

115

+H

235-H-H

145

251

279-H

301

288

315

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

Wavelength,nm



©2019Serhanetal.

112

AT-LXB4

11.1

351=M-H333=M-H-H2O315=M-H-2H2O307=M-H-CO2297=M-H-3H2O289=M-H-H2O-CO2271=M-H-2H2O-CO2261=279-H2O243=279-2H2O233=251-H2O217=235-H2O215=251-2H2O207=251-CO2203=221-H2O189=251-H2O-CO2177=221-CO2159=221-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

351

221

315

233

333

189 251207271203177

159 289297215

243 307261217115

100 140 180 220 260 300 340 380m/z,Da

301

288

314

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

Wavelength,nm

279251

-H221

115

235

+H

-H

+H

COO-

OH

HO OH



©2019Serhanetal.

113

AT-RvD1

11.2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

375=M-H357=M-H-H2O339=M-H-2H2O331=M-H-CO2313=M-H-H2O-CO2295=M-H-2H2O-CO2261=305-CO2259=277-H2O243=305-H2O-CO2241=277-2H2O215=233-H2O197=233-2H2O185=203-H2O153=171-H2O135=171-2H2O127=171-CO2123=141-H2O

215

375

233

141

135

277

185127357171 295 331113 313123 197

153259 339241

2430%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

COO-

HO

OH

HO

141

233-H

+H277

171-H

113+H

-H305

203

301

288316

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

Wavelength,nm



©2019Serhanetal.

114

AT-RvD3

375=M-H357=M-H-H2O339=M-H-2H2O313=M-H-H2O-CO2295=M-H-2H2O-CO2259=277-H2O215=277-H2O-CO2191=209-H2O177=195-H2O159=195-2H2O147=165-H2O137=181-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

10.5

220 240 260 280 300 320200

Rel.Ab

s.(%

)

100%

0%

271

λmaxMeOH=271nm


281260

234

375

147

137357

165 215181 277 313295259159 177 339101 191

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 120 140 160 180 200 220 240 260 280 300 320 340 360 380m/z,Da

Wavelength,nm

277+H

195

181165

209-H

+H

101-H

HO

COO-

OH

HO



©2019Serhanetal.

115

COO-

OH

OH

AT-PD1

12.4

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

270

261 281λmax


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

359=M-H341=M-H-H2O323=M-H-2H2O315=M-H-CO2297=M-H-H2O-CO2279=M-H-2H2O-CO2243=261-H2O217=261-CO2199=261-H2O-CO2188=206-H2O159=177-H2O137=181-CO2119=181-H2O-CO2109=153-CO2

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

153

359

137206

297188

341261217159 323315

199119177

181 279243

100 140 180 220 260 300 340 380m/z,Da

Wavelength,nm

153

206-H

+H

-H

+H

177 181

261



©2019Serhanetal.

116

CysteinylLeukotrienes



©2019Serhanetal.

117

LTC4

10.32

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

270 293

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

626=M+H608=M+H-H2O582=M+H-CO2479=497-H2O301=319-H2O171=189-H2O

Wavelength,nm

319

189

+H

+H

497



©2019Serhanetal.

608

308

Rel.Int.(%

)

100 150 200 250 300 350 400 450 500 550 600 650m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%626

497

301

319

189

171

308

394

+2H

131

131

+H

582479

118

+H

LTD4

9.24

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

497=M+H479=M+H-H2O301=319-H2O275=319-CO2135=179-CO2

Wavelength,nm

-H

+H 179



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

100 150 200 250 300 350 400 450 500m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

479

497

189

116179

301

135

319

189116

131

131

275

119

LTE4

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

10.59

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

440=M+H301=319-H2O283=319-2H2O

116

100 150 200 250 300 350 400 450m/z,Da

440116

189

319

189

301

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

131

131

+H-H

283

120

MaresinConjugatesinTissueRepair



©2019Serhanetal.

121

MCTR1

9.72

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

650=M+H632=M+H-H2O503=521-H2O325=343-H2O217=235-H2O187=205-H2O173=191-H2O161=205-CO2147=191-CO2

Wavelength,nm



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

+H

191

100 150 200 250 300 350 400 450 500 550 600 650m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

109

191

205235

308

343

418 521

173

187 217

325

503 632

650

109

205

235+H

343

308 +H418

521+2H

+H

147

161

122

MCTR2

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

8.81

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

-H

100 150 200 250 300 350 400 450 500 550m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

521

147

109343179

145131115

521=M+H503=M+H-H2O325=343-H2O217=235-H2O187=205-H2O173=191-H2O161=205-CO2147=191-CO2

191

173

325

217

503

191

235109

179343

+H

+H 205

131

145

115

+H

-H

187161

123

MCTR3

10.12

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

464=M+H446=M+H-H2O325=343-H2O217=235-H2O187=205-H2O173=191-H2O161=205-CO2147=191-CO2

100 150 200 250 300 350 400 450 500m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

109

235

205

191

464

325

343 446

217

173

147

191

205

235109

343+H

+H

161

187

124

ProtectinConjugatesinTissueRepair



©2019Serhanetal.

125

PCTR1

9.72

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

650=M+H632=M+H-H2O503=521-H2O325=343-H2O290=308-H2O264=308-CO2257=275-H2O227=245-H2O201=245-CO2

+H

231

245275

308

343

521

+H

+H

+2H

100 150 200 250 300 350 400 450 500 550 600 650m/z,Da

650

503

231

245

275

308

343

521

325

632

264

290

257

227

201

100%

90%

80%

70%

60%

50%

40%

30%

20%

10%

0%

Rel.Int.(%

)

126

8.81

293270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

PCTR2

521=M+H503=M+H-H2O325=343-H2O257=275-H2O239=275-2H2O231=275-CO2213=231-H2O187=231-CO2161=179-H2O135=179-CO2

100 150 200 250 300 350 400 450 500 550m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

521

325

245

231

179343

503

257239

213

187

161

135

343

179

275

231

+H

+H

127

PCTR3

10.12

293

270

281

λmaxMeOH=281nm


Rel.Ab

s.(%

)

0%

50%

100%

250 270 290 310230 330

Wavelength,nm



©2019Serhanetal.

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16

Time,min

0

464=M+H446=M+H-H2O325=343-H2O227=245-H2O213=231-H2O201=245-CO2187=231-CO2

100 150 200 250 300 350 400 450 500m/z,Da

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

464

343446

121

245

231

325

227

213

201

187

275

245343

121

231

+H

+H

+H

128



©2019Serhanetal.

ResolvinConjugatesinTissueRepair

129


130



©2019Serhanetal.

RCTR1

100 m/z,Da

150 200 250 300 350 400 450 500 550 600 650

666=M+H648=666-H2O630=666-2H2O519=537-H2O501=537-2H2O416=434-H2O398=434-2H2O341=359-H2O323=359-2H2O199=217-H2O185=203-H2OCOOH

S

HO

OHO

HNHOOC

HN O

COOHH3N

308

203

359

247

217

434

609+2H

+H

+H

537

+H537

630

341323

179308 416

666

648

203247

185217 519

501

398

434

537

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

Wavelength(nm)240 260 280 300 320 340 360

λmaxEtOH=308nm308

322

296

2 4 6 8 10 12 14 160Time(min)

Rel.Int.(%

)

0%

50%

100%

7.01

Rel.Ab

s.(%

)

0%

50%

100%

131



©2019Serhanetal.

m/z,Da100

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

RCTR2

537=M+H519=537-H20501=537-2H20341=359-H20323=359-2H20211=247-2H20185=203-H20

150 200 250 300 350 400 450 500 550 600 650

537

179

341

323

247

519

162 438

203143

501

185

211

393

COOHS

HO

OHO

HNHOOC

NH3

203

359

393247

438 +H

179

+H

+H

Wavelength(nm)240 260 280 300 320 340 360

λmaxEtOH=308nm308

322296

Time(min)2 4 6 8 10 12 14 160

Rel.Int.(%

)

0%

50%

100%

6.24

Rel.Ab

s.(%

)

0%

50%

100%


132



©2019Serhanetal.

RCTR3

480=M+H462=480-H20444=480-2H20341=359-H20323=359-2H20211=247-2H20199=217-H20185=203-H20

m/z,Da

444323

382341 412247121

480

462

359217203

199 211

100 150 200 250 300 350 400 450 500 550 600 650

185

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

) COOHS

HO

OH

HOOC

NH3

203

359

217247

121

382

412

+H

+H

+H

Wavelength(nm)240 260 280 300 320 340 360

λmaxEtOH=308nm

308

322296

Time(min)2 4 6 8 10 12 14 160

Rel.Int.(%

)

0%

50%

100%

Rel.Ab

s.(%

)

0%

50%

100%


7.42

DeuteriumLabeledInternalStandards



©2019Serhanetal.

133

d4-PGE2

+H255 -H

283

O

HO

COO-

OH

D D

D D

275

319 337

193

239211 355293

237

283

355=M-H337=M-H-H2O319=M-H-2H2O293=M-H-H2O-CO2275=M-H-2H2O-CO2239=283-CO2237=255-H2O211=255-CO2193=255-H2O-CO2

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

10.4

Rel.Int.(%

)Prof.CharlesN.SERHANLaboratoryBrigham&Women’sHospitalCenterforExperimentalTherapeuticsandReperfusionInjuryHarvardMedicalSchool


©2019Serhanetal.

134

d5-LXA4

10.1

356=M-H320=M-H-2H2O312=M-H-CO2294=M-H-H2O-CO2276=M-H-2H2O-CO2233=251-H2O222=240-H2O189=251-H2O-CO2

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

356

115

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

115

+H

240-H

251

301

288

315

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

320312

294

276251

240

233

222

189

Wavelength,nm

-H145

145



©2019Serhanetal.

135

COO-

HO OH

OH

D D

D

D

D

d4-LTB4

COO-

OHOH

D

D

D D

142

197113

225-H

+H223

115-H-H

339=M-H321=M-H-H2O303=M-H-2H2O295=M-H-CO2277=M-H-H2O-CO2259=M-H-2H2O-CO2207=225-H2O205=223-H2O181=225-CO2179=197-H2O163=225-H2O-CO2153=197-CO2135=197-H2O-CO2124=142-H2O

339

197

321

153277207181

115 303259163124135 295142

113 2051790%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

270

261 281λmax


Rel.Ab

s.(%

)

0%

50%

100%

270230 250 290210 3303100%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

13.1

Rel.Int.(%

)

Wavelength,nm



©2019Serhanetal.

136

d8-5S-HETE

COO-

OH

D D D D

D D D D

116210

-H327

309116 265

210

283


0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

236

λmaxMeOH=236nm


Rel.Ab

s.(%

)

0%

50%

100%

220 230 240 250 270210 280 290260

16.2

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

Rel.Int.(%

)

Wavelength,nm



©2019Serhanetal.

137

d5-RvD2

10.6

Rel.Int.(%

)

0%

50%

100%

2 4 6 8 10 12 14 16 18 20 22

Time,min

0

380

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Rel.Int.(%

)

100 140 180 220 260 300 340 380m/z,Da

301

288

315

λmaxMeOH=301nm


Rel.Ab

s.(%

)

0%

50%

100%

260 280 300 320 340240 360 380

362

Wavelength,nm



©2019Serhanetal.

COO-

OH

HO

OH

D

D D

DD

380=M-H362=M-H-H2O344=M-H-2H2O336=M-H-CO2318=M-H-H2O-CO2300=M-H-2H2O-CO2259=277-H2O241=277-2H2O215=277-CO2-H2O185=247-H2O-CO2

238

+H247

277+H

133-H

141-H

344336318300

141 277

133113

113+H

247238

259241

215

185

138

Documents

104 Lipid Mediator Metabololipidomics LC-MS-MS Spectra ...serhanlab.bwh.harvard.edu/.../UPDATED-2019-Spectra... · Lipid Mediator Metabololipidomics LC-MS-MS Spectra Book 2019 Ian