Lipid Mass Spectrometry in the

1
Lipid Mass Spectrometry in the Cardiovascular
System Plasmalogen Targeting by Reactive
Chlorinating Species Derived from Phagocytes in
Infarcted Myocardium David Ford, Ph.D.
Department of Biochemistry and Molecular Biology
St. Louis University School of Medicine
2
MOLECULAR SUBCLASSES
OF ETHANOLAMINE GLYCEROPHOSPHOLIPIDS
O
H
H
C
O
C
H
CH
O
C
H
C
O
C
C
H
15
31
2
15
31
14
29
O
O
O
O
C
C
H
O
C
C
H
O
C
C
H
19
31
19
31
19
31
O
O
O



P
O
CH
NH
O
CH
P
O
O
CH
NH
CH
P
O
O
CH
NH
CH
3
2
2
-
3
3
2
2
2
2
-
-
O
O
O
ALKYL ETHER
DIACYL
PLASMALOGEN
(Plasmanylethanolamine)
(Phosphatidylethanolamine)
(Plasmenylethanolamine)
3
Plasmalogen Targeting by Reactive Chlorinating
Species Derived from Phagocytes
Cl
Schiff base adducts
C
C
C
O
H
14
29
H
H
2-Chlorohexadecanal
(
-Chloro-fatty aldehyde)
a

H
H
H
O
C
C
C
14
29
H
, Cl
O
O
2
2
OH
O
C
R
1
Myeloperoxidase
O
CH
O
LPC chlorohydrins
3
O
C
R
1
O
O
P
(CH
)
N
CH
3
2
2

CH
O
3
CH
O
3
Plasmenylcholine
O
O
P
(CH
)
N
CH
3
2
2

(Plasmalogen)
CH
O
3
Lysophosphatidylcholine
4
GC-MS analysis by negative ion electron capture
of the PFB oxime of 2-chlorohexadecanal produced
from RCS attack of plasmalogens in infarcted
myocardium
5
2-Chlorohexadecanal in infarcted myocardium
1 day LAD Occlusion
D
Mid-Ventricle
1 day sham surgery
E
Mid-Ventricle
6
Concomitant accumulation of neutrophils and
2-chlorohexadecanal
7
Schiff Base Formation
O
O
O
R
R
2
1
O
O
O
O
O
O
P
O
-
R
R
O
2
N
H
1
2
O
O
Phosphatidylethanolamine (PE)
O
P
O

-
O
N
H2O
O
Cl
C
H
C
H
Cl
29
14
14
29
N-2-ClHDA-PE
2-Chlorohexadecanal (2-ClHDA)
8
2-ClHDA Forms Schiff Bases with di160-PE
9
N-2-ClHDA-PE Schiff Base Adducts in HCAEC
Incubate HCAEC with 25 mM 2-ClHDA Stabilize
Schiff bases with NaCNBH3 Detect and quantify
Schiff base adducts by LC-MS/MS (SRM)
SRM M-H-
m/z 362
10
N-2-ClHDA-PE Schiff Base Adducts in Human
Coronary Artery Endothelial Cells
11
Secondary RCS Attack of Plasmalogens Production
of LPC Chlorohydrins
12
CID of m/z 596 (18 carbon LPC Chlorohydrin)
13
Detection of LPC chlorohydrins in HCAEC treated
with HOCl Neutral Loss Scanning of m/z 95
No Treatment
500 mM HOCl
17C (int.std.)
19C (int.std.)
17C (int.std.)
19C (int.std.)
14
Summary of reactive chlorinating species attack
of plasmalogens
Phagocyte Chemoattractant, Schiff base
adducts Cell Death
Cl
C
C
C
O
H
14
29
H
H
2-Chlorohexadecanal
(
-Chloro-fatty aldehyde)
a

H
H
O
C
C
C
H
29
14
H
, Cl
O
LPC Chlorohydrin PKA activation Gene
activation? Effects on Endothelial Cells Cell
Death?
O
2
2
OH
O
C
R
1
Myeloperoxidase
O
O
CH
3
O
C
R
1
CH
O
O
P
(CH
)
N
2
2
3

O
CH
3
O
CH
3
Plasmenylcholine
CH
O
O
P
(CH
)
N
2
2
3

(Plasmalogen)
O
CH
3
Lysophosphatidylcholine
15
ACKNOWLEDGEMENTS
NIH and American Heart Association
Saint Louis University Carolyn Albert Arun
Thukkani Kristin Wildsmith Maria Messner Viral
Brahmbhatt Dhanam Anbukumar George Vogler Brad
Martinson Washington University Mass
Spectrometry Resource John Turk Fong Hsu Jan
Crowley