Identification of Darmstoff analogs as selective agonists and antagonists of lysophosphatidic acid receptors

Article abstract of DOI:10.1016/j.bmcl.2005.08.096

Darmstoff describes a family of gut smooth muscle-stimulating acetal phosphatidic acids initially isolated and characterized from the bath fluid of stimulated gut over 50 years ago. Despite similar structural and biological profiles, Darmstoff analogs have not previously been examined as potential LPA mimetics. Here, we report a facile method for the synthesis of potassium salts of Darmstoff analogs. To understand the effect of stereochemistry on lysophosphatidic acid mimetic activity, synthesis of optically pure stereoisomers of selected Darmstoff analogs was achieved starting with chiral methyl glycerates. Each Darmstoff analog was evaluated for subtype-specific LPA receptor agonist/antagonist activity, PPARγ activation, and autotaxin inhibition. From this study we identified compound 12 as a pan-antagonist and several pan-agonists for the LPA_1-3 receptors. Introduction of an aromatic ring in the lipid chain such as analog 22 produced a subtype-specific LPA₃ agonist with an EC₅₀ of 692 nM. Interestingly, regardless of their LPA_1/2/3 ligand properties all of the Darmstoff analogs tested activated PPARγ. However, these compounds are weak inhibitors of autotaxin. The results indicate that Darmstoff analogs constitute a novel class of lysophosphatidic acid mimetics.

Full text of DOI:10.1016/j.bmcl.2005.08.096

Bioorganic & Medicinal Chemistry Letters 16 (2006) 451–456
Identification of Darmstoff analogs as selective agonists
and antagonists of lysophosphatidic acid receptors
Veeresa Gududuru,a Kui Zeng,a Ryoko Tsukahara,b Natalia Makarova,b Yuko Fujiwara,b
Kathryn R. Pigg,c Daniel L. Baker,c Gabor Tigyib and Duane D. Millera,*
aDepartment of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, TN 38163, USA
bDepartment of Physiology, and University of Tennessee Health Science Center, Memphis, TN 38163, USA
cDepartment of Medicine and Vascular Biology and Genomics and Bioinformatics Centers of Excellence,
University of Tennessee Health Science Center, Memphis, TN 38163, USA
Received 10 August 2005; revised 29 August 2005; accepted 29 August 2005
Available online 14 November 2005
Abstract—Darmstoff describes a family of gut smooth muscle-stimulating acetal phosphatidic acids initially isolated and character-
ized from the bath fluid of stimulated gut over 50 years ago. Despite similar structural and biological profiles, Darmstoff analogs
have not previously been examined as potential LPA mimetics. Here, we report a facile method for the synthesis of potassium salts
of Darmstoff analogs. To understand the effect of stereochemistry on lysophosphatidic acid mimetic activity, synthesis of optically
pure stereoisomers of selected Darmstoff analogs was achieved starting with chiral methyl glycerates. Each Darmstoff analog was
evaluated for subtype-specific LPA receptor agonist/antagonist activity, PPARc activation, and autotaxin inhibition. From this
study we identified compound 12 as a pan-antagonist and several pan-agonists for the LPA1–3 receptors. Introduction of an aro-
matic ring in the lipid chain such as analog 22 produced a subtype-specific LPA3 agonist with an EC50 of 692 nM. Interestingly,
regardless of their LPA1/2/3 ligand properties all of the Darmstoff analogs tested activated PPARc. However, these compounds
are weak inhibitors of autotaxin. The results indicate that Darmstoff analogs constitute a novel class of lysophosphatidic acid
mimetics.
ꢀ 2005 Elsevier Ltd. All rights reserved.
Lysophosphatidic acid (LPA) is a growth factor-like
lysophospholipid1 involved in many physiological and
pathological processes including neurogenesis,2 myelina-
tion, angiogenesis,3 wound healing,4 and cancer progres-
sion.5 LPA is produced by a number of cell types
including platelets, adipocytes, fibroblasts, and ovarian
cancer cells.6,7 LPA is a product of lysophospholipase
D, also known as autotaxin, an esterase that cleaves
lysophosphatidylcholine to LPA and choline.8,9 LPA
acts through three G protein coupled receptors, LPA1
(EDG-2), LPA2 (EDG-4), and LPA3 (EDG-7).10,11
Recently, two non-EDG LPA receptors, LPA4/GPR23
and the nuclear transcription factor/receptor PPARc,
have been reported.12,13 Most cells express a combina-
tion of these receptors, making it difficult to dissect
the biological effects mediated by an individual receptor
subtype. The need to understand the biological function
of LPA receptors and the desire to pharmacologically
exploit the differences in their ligand recognition require
the development of receptor subtype-selective agonists
and antagonists.14,15
Based on Weilandꢀs observations16 in 1949, Vogt was
first to report the isolation of an acidic phospholipid
from the bath fluid of horse intestine that stimulated
smooth muscle contraction, which he named Darmst-
off.17 Darmstoff, present not only in the bath but also
in the gut extract, was subsequently shown to be a mix-
ture of acetal phosphatidic acids primarily composed of
1a, 1b, and 1c (Fig. 1). Interestingly, all the intestinal
smooth muscle contracting activity possessed by
Darmstoff resides in the oleyl acetal 1b.18,19 These com-
pounds were also shown to display hypotensive and
cardiodepressant properties20 and stimulate platelet
aggregation. Vogt was also the first to describe an
LPA-induced physiological response. Surprisingly, nei-
ther Vogt nor anyone else has published work in which
Darmstoff or its analogs were examined as LPA mimics.
In our continuing efforts to develop subtype-selective
Keywords: Darmstoff; LPA mimetics; PPARg; ATX.
*
Corresponding author. Tel.: +1 (901) 4486026; fax: +1 (901)
4483446; e-mail: dmiller@utmem.edu
0960-894X/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.08.096

452
V. Gududuru et al. / Bioorg. Med. Chem. Lett. 16 (2006) 451–456
O
O
O
R
H
O
HO P O
OH
O
O
(CH2)nCH3
O
P
OH
OH
OH
Darmstoff
LPA (2)
1a, R = (CH2)14CH3
1b, R = (CH2)7CH=CH(CH2)7CH3
1c, R = (CH2)7CH=CHCH2CH=CH(CH2)4CH3
Figure 1. Chemical structures of Darmstoff and LPA.
LPA agonists and antagonists, we revisited the possibil-
ity that acetal phosphatidates would mimic the effects of
LPA and for the first time we show here that Darmstoff
analogs constitute a new class of LPA-like ligands that
act on multiple LPA targets.21 The details of synthesis
and pharmacological characterization of a series of
Darmstoff analogs are discussed herein.
conditions formed the dioxolane 20. Debenzylation of
20 followed by phosphorylation and removal of the pro-
tecting groups gave target compounds 21 and 22.
To investigate the effect of stereochemistry on biological
activity, we decided to synthesize all stereoisomers of
Darmstoff analogs 13 and 14. From the chemical struc-
tures it is clear that 2,4-disubstituted-1,3-dioxolanes of
this type exist as a mixture of four stereoisomers. Previ-
ously it was difficult to separate optically pure dioxol-
anes, although there have been reports that utilize
chiral stationary phases24or enzymatic methods.25
Recently, Lin et al. showed the separation of 2,4-disub-
stituted-1,3-dioxolane stereoisomers utilizing a semi-
preparative HPLC method.26
Previously reported methods19,22 for the synthesis of this
compound class have limitations and are particularly
unsuitable for the preparation of Darmstoff analogs
containing thiophosphate head groups. To facilitate
the synthesis of various Darmstoff analogs in an effort
to develop a structure–activity relationship, we devel-
oped a general method for the preparation of these
compounds utilizing phosphoramidite chemistry.
Accordingly, PCC mediated oxidation of fatty alcohols
produced the corresponding aldehydes, which were con-
densed with glycerol in the presence of PTSA under
reported conditions23 to give dioxolanes 6a–c (Scheme
1). Phosphorylation of 6a–c using bis(cyanoethyl)-
N,N-diisopropylphosphoramidite (7) in the presence of
1H-tetrazole formed phosphorous acid esters that were
converted in situ to phosphate or thiophosphate esters
using hydrogen peroxide or sulfur, respectively. Finally,
treatment of the phosphate or thiophosphate esters with
methanolic KOH at ambient temperature provided
potassium salts of Darmstoff analogs.
Our synthetic approach for the preparation of the four
possible stereoisomers of 13 and 14 is outlined in
Scheme 3. Accordingly, acid mediated removal of the
isopropylidene group from commercially available 23
(R-isomer) gave methyl glycerate (24) in a quantitative
yield. The acid-catalyzed condensation of cis-9-octadec-
enal with 24 afforded a mixture of dioxolanes 25 and 26,
which were readily separated by column chromato-
graphy.27,28 LiBH4 mediated reduction of the ester
functionality of 25 gave alcohol 27 that was phosphory-
lated using 7 to form phosphate 28 and thiophosphate
29. Finally, treatment of these esters with methanolic
KOH gave the corresponding Darmstoff stereoisomers
30 and 31 as shown in Scheme 3. Similarly, dioxolane
intermediate 26 was converted to target compounds 34
and 35 using the same chemistry. Synthesis of the other
four Darmstoff stereoisomers (37–40) utilized the same
procedure but used 36 (S-isomer of 23) as the starting
material. To examine the purity of these stereoisomers,
The synthesis of compounds 21 and 22 containing a
phenyl ring in the lipid chain is shown in Scheme 2. Fri-
edel–Crafts acylation of n-octyl benzene with pimelic
anhydride gave arylketo acid 18 that was converted to
the required aldehyde 19 in three steps. Condensation
of 19 with 3-benzyloxy-propane-1,2-diol under standard
O
R
HO
b
a
R CHO +
4a-4c
R
3a-3c
CH2OH
HO
OH
O
OH
6a-6c
5
O
O
O
c
d
X
X
R
R
O P O-K+
O P O(CH2)2CN
O(CH2)2CN
O
OH
12-15
8-11
R
X
O
O
S
C
12H25 (13:0)
12
C17H33 (18:1, 9)
C17H33 (18:1, 9)
13
14
15
C17H29 (18:3, 9,12,15)
O
Scheme 1. Reagents and conditions: (a) PCC, CH2Cl2, 90–95%; (b) PTSA, C6H6, Dean–Stark, reflux, 70–78%; (c) (1) bis(cyanoethyl)-N,N-
diisopropylphosphoramidite (7), 1H-tetrazole, CH2Cl2, (2) 30% H2O2, rt or sulfur, reflux, 70–75%; (d) KOH, MeOH, 80–85%.

V. Gududuru et al. / Bioorg. Med. Chem. Lett. 16 (2006) 451–456
453
O
O
O
O
OH
H
b, c, d
a
+
O
O
19
C8H17
C8H17
C8H17
18
16
17
H
O
H
O
O
X
f, g, h
e
P O-K+
O
O
OBn
OH
C8H17
C8H17
21, X = O
22, X = S
20
Scheme 2. Reagents and conditions: (a) AlCl3, CH2Cl2, 60%; (b) N2H4ÆH2O, KOH, ethylene glycol, MW, 87%; (c) BH3ÆTHF, 78%; (d) PCC, CH2Cl2,
95%; (e) 3-benzyloxy-propane-1,2-diol, PTSA, C6H6, Dean–Stark, reflux, 83%; (f) H2/Pd(OH)2, EtOAc, 100%; (g) (1) bis(cyanoethyl)-N,N-
diisopropylphosphoramidite, 1H-tetrazole, CH2Cl2, (2) 30% H2O2, rt or sulfur, reflux, 70%; (h) KOH, MeOH, 80%.
HO
HO
O
O
O
C17H33
C17H33
a
OMe
b
OMe
OMe
OMe
+
e
O
O
O
H
H
H
H
O
O
O
26
25
23
O
24
X
X
O
O
O
C17H33
C17H33
d
c
C17H33
O P O-K+
OH
O P O(CH2)2CN
O(CH2)2CN
25
O
O
O
H
H
H
OH
H
H
H
27
32
28, X = O
29, X = S
30, X = O
31, X = S
f
O
C17H33
OBn
O
H
H
X
O
C17H33
O P O-K+
c,d,e
O
26
H
OH
H
34, X = O
35, X = S
X
X
O
O
C17H33
C17H33
O P O-K+
O
O P O-K+
+
OMe
O
O
H
H
OH
OH
H
H
O
39, X = O
40, X = S
37, X = O
38, X = S
O
36
O
O
f
C17H33
C17H33
OBn
OH
O
O
33
H
H
H
H
41
Scheme 3. Reagents and conditions: (a) HCl, MeOH, 88%; (b) C17H33CHO, PTSA, C6H6, Dean–Stark, reflux, 75%; (c) LiBH4, Et2O, 83%;
(d) (1) bis(cyanoethyl)-N,N-diisopropylphosphoramidite, 1H-tetrazole, CH2Cl2, (2) 30% H2O2, rt or sulfur, reflux, 73%; (e) KOH, MeOH, 81%;
(f) BnBr, NaH, THF, reflux, 75%.
HPLC profiles of compounds 32 and 41 were analyzed.
Benzyl ethers (32 and 41, Scheme 3) were prepared to in-
crease their detection by UV. The HPLC analysis (Chir-
alpak AS-RH 150 · 4.6 mm, 1:1 water–acetonitrile) of
benzyl ethers confirmed the purity of 32 and 41. All
compounds were fully characterized by spectroscopic
methods.29
Darmstoff analogs. Second, PPARc activation was
examined in CV1 cells, transfected with an acyl-coen-
zyme A oxidase-luciferase (PPRE-Acox-Rluc) reporter
gene construct as previously reported.30 Third, inhibi-
tion of the lysophospholipase D autotaxin was deter-
mined using a previously described procedure.31
Compound 12 containing a C13 alkyl chain and no dou-
ble bond inhibited Ca2+ mobilization in cells expressing
all three LPA GPCRs there by providing a pan-antago-
nist of LPA1–3 (Table 1). An increase in chain length to
C18 and introduction of the C9@C10 double bond result-
ed in analog 13, which produced LPA1/3 antagonist
activity. It is interesting to note that oleoyl-LPA is an
agonist of LPA1–3 while Darmstoff analog 13 containing
The biological effects of all synthesized compounds were
tested using three high-throughput assays. First, intra-
cellular calcium transients in rat hepatoma (RH7777,
an LPA receptor null cell line) cell lines individually
expressing either LPA1, LPA2, and LPA3 receptors were
analyzed to examine agonism and antagonism.30 Wild
type RH7777 cells did not respond to any of the

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V. Gududuru et al. / Bioorg. Med. Chem. Lett. 16 (2006) 451–456
Table 1. Ligand properties of Darmstoff analogs at LPA1/2/3, PPARc, and ATX
Compound
LPA1
IC50
(Emax)a(nM) (Ki)(nM)
LPA2
IC50
(Emax) (nM) (Ki) (nM)
LPA3
IC50
(Emax) (nM) (Ki) (nM)
7430 (745) NE 2870 (681) Agonist 232 (26)
PPARc ATX IC50 (inhibitionmax %)
(nM)
EC50
EC50
EC50
12
13
14
15
21
22
30
31
34
35
37
38
39
40
NEb
NE
1110 (652)
915 (497)
NE
NE
>10,000
1170 (87)
1710 (51)
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
527 (548)
NE
NE
Agonist 141 (30)
Agonist 415 (51)
Agonist 803 (54)
Agonist 106 (10)
Agonist 449 (55)
Agonist 120 (30)
Agonist 252 (74)
Agonist 97 (28)
Agonist 344 (66)
Agonist 238 (46)
Agonist 363 (64)
Agonist 178 (25)
Agonist 403 (60)
981 (45)
3600 (55)
NE
639 (73)
7590 (29)
NE
NE
4660 (1930) NE
504 (171)
NE
WA
NE
NE
WAc
WA
NE
NE
NE
NE
NE
WA
NE
NE
692 (87)
NE
1170 (39)
1300 (77)
1710 (42)
1090 (85)
>10,000
1540 (72)
NE
1580 (89)
NE
127 (127)
NE
NE
136 (83)
NE
1410 (71)
>10,000
2260 (68)
NE
194 (113)
NE
484 (241)
NE
204 (102)
NE
265 (78)
209 (77)
NE
1560 (65)
1320 (87)
a Emax, maximal efficacy of drug/maximal efficacy of LPA 18:1, expressed as the percentage.
b NE, no effect was shown at the highest concentration (30 lM) tested.
c WA, weak antagonist.
an oleoyl chain at C-2 position of the 1,3-dioxolane
inhibited LPA1–3 receptors indicating the significant role
played by the acetal moiety in ligand recognition. The
replacement of the phosphate head group with a thio-
phosphate in our previously reported fatty alcohol phos-
phate analogs had a significant effect on their ligand
properties with regard to the LPA1–3 receptors.30 To
examine the effect of this modification with Darmstoff
series compound 14 was synthesized. This analog was
an agonist at all three LPA receptor subtypes and was
most potent at LPA3 (EC50 of 639 nM). The phosphate
analog 15 with conjugated double bonds at C9@C10,
C12@C13, and C15@C16 positions was an agonist for all
three LPA receptors. Though analog 15 was less potent
than 14, these compounds were identified as two new
LPA GPCR pan-agonists. We are currently using these
leads to optimize the receptor potency and introduce
specificity.
phosphate head groups were LPA3 antagonists,
whereas analogs with thiophosphate groups (31, 35,
38, and 40) were pan-agonists. Among the phosphate
stereoisomers, analog 34 was identified as the most po-
tent LPA3 antagonist with an IC50 of 136 nM
(Ki = 83 nM). Interestingly, compound 30 weakly acti-
vated LPA1/3 and was a partial LPA2 agonist with an
EC50 of 1.17 lM (Emax = 39%). It is noteworthy that
stereoisomers with a thiophosphate head group were
found to be more potent at LPA3 receptor than parent
compound 14. In this series all other stereoisomers (31,
35, and 38), with the exception of 40, were full agonists
of LPA3 receptor, with the most potent being 31 (EC50
of 127 nM, Emax = 127%) there by indicating the
importance of stereochemistry on biological activity
of Darmstoff analogs.
PPARc is a lipid-activated transcription factor that be-
longs to the nuclear hormone receptor superfamily.
Recently, McIntyre et al. showed that LPA is a direct ago-
nist of PPARc.13 They demonstrated that LPA competed
against the synthetic PPARc agonist Rosiglitazone and
the natural agonist azaleylphosphatidylcholine. LPA
binding to PPARc has a nanomolar Kd and LPA does
not bind to PPARa or PPARb.13 LPA-induced activation
of PPARc appears to be an important step in the regula-
tion of adipogenesis, macrophage function, neointima
formation, and atherogenesis.33 Zhang et al. showed that
unsaturated and alkyl ether analogs of LPA induce neoin-
tima formation, an early step that leads to the develop-
ment of atherogenic plaques, through PPARc
activation.33 We examined the activity of all synthesized
Darmstoff analogs as PPARc activators in vitro in CV1
cells using a PPRE-Acox-Rluc reporter gene assay. For
comparison purposes we also included Rosiglitazone, a
known PPARc agonist, as a positive control in all analy-
ses. The results (Fig. 2) indicate that all tested Darmstoff
analogs, regardless of whether they were agonist or antag-
onist of LPA GPCR, activated the PPARc reporter con-
struct. These results are consistent with previous
A multitude of aldehydes are produced via oxidative
cleavage of unsaturated fatty acids and their phospho-
lipid derivatives.32 The cis-olefinic bond of analogs
13–15 is susceptible to oxidative cleavage. In order
to avoid this problem and to examine the effect of
structural rigidity on biological activity, we replaced
the double bond with an aromatic ring and screened
against LPA GPCR, PPARc, and autotaxin. Incorpo-
ration of an aromatic ring in the alkyl chain led to
compounds 21 and 22. Analog 21 was an antagonist
of LPA1/3 receptors but had no effect against LPA2.
The thiophosphate analog, compound 22, was a weak
LPA1 antagonist, without any effect on LPA2 but
stimulated LPA3 with an EC50 of 692 nM
(Emax = 87%).
Next, we examined the importance of stereochemistry
on biological activity. Analysis of pure stereoisomers
(Table 1) with respect to LPA GPCR activation sug-
gests that regardless of their stereochemistry at C-2
and C-4, Darmstoff analogs (30, 34, 37, and 39) with

V. Gududuru et al. / Bioorg. Med. Chem. Lett. 16 (2006) 451–456
455
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3624.
28. Dioxolanes 25 (49%) and 26 (26%) were separated on a
silica gel column (eluted with a mixture of diethyl ether–
hexanes, 5:95, v/v). Configurational assignment for the C2
acetal proton of 25 and 26 was based on 1H NMR studies.
The C2 acetal proton appeared as a triplet at d 5.09 ppm
for the trans-stereoisomer (25) and at 4.99 ppm for the cis-
stereoisomer (26). These observations were in complete
agreement with the earlier reports26,27 wherein the C2
proton for the trans-stereoisomer of 2,4-disubstituted-1,
Acknowledgments
This research was supported by a grant from the
Department of Defense (DAMD17-01-1-0830) and a
grant from National Institutes of Health (CA
92160).

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V. Gududuru et al. / Bioorg. Med. Chem. Lett. 16 (2006) 451–456
3-dioxolanes always appeared in the downfield of 1H
NMR spectra.
J = 6 Hz, 2H), 4.08–4.13 (m, 1H), 4.30–4.38 (m, 1H), 4.99
(t, J = 4.8 Hz, 1H), 5.33 (m, 2H); 13C NMR (CD3OD) d
13.04, 21.93, 23.25, 26.44, 28.51, 28.56, 28.78, 28.89, 29.0,
31.20, 31.83, 33.27, 64.24, 66.26, 74.74, 74.85, 103.65,
104.03, 129.05, 129.19; MS (ESI) m/z 435 [MÀH]; 31P
NMR (CD3OD) d 63.82.
29. Characteristic data for some compounds: 13: 1H NMR
(300 MHz, CD3OD) d 0.86 (m, 3H), 1.27 (m, 24H), 2.0
(m, 4H), 3.77–4.0 (m, 3H), 4.14–4.21 (m, 1H), 4.26–4.35
(m, 1H), 4.89 (t, J = 4.8 Hz, 0.5H), 5.0 (t, J = 4.8 Hz,
0.5H), 5.35 (m, 2H); 13C NMR (CD3OD) d 12.68, 21.68,
22.91, 23.12, 25.99, 26.09, 28.06, 28.25, 28.33, 28.51,
28.60, 28.69, 30.92, 31.43, 32.76, 33.22, 33.72, 63.89,
64.44, 66.17, 69.66, 69.87, 74.54, 74.75, 101.55, 103.80,
104.37, 129.20, 129.25; MS (ESI) m/z 419 [MÀH]; 31P
NMR (CD3OD) d 19.68. Compound 14: 1H NMR
(300 MHz, CD3OD) d 0.78 (m, 3H), 1.19 (br s, 24H),
1.93 (m, 4H), 3.76–3.89 (m, 3H), 4.05–4.25 (m, 2H), 4.4–
4.47 (m, 0.4H), 4.75 (m, 0.75H), 4.97 (m, 0.3H), 5.25 (m,
2H); 13C NMR (CD3OD) d 13.36, 22.09, 26.62, 28.81,
29.21, 31.39, 33.09, 64.24, 66.18, 74.86, 74.90, 103.87,
104.50, 129.19, 129.42; MS (ESI) m/z 435 [MÀH]; 31P
NMR (CD3OD) d 62.85.
Compound 34: 1H NMR (300 MHz, CD3OD) d 0.84
(t, J = 6.3 Hz, 3H), 1.25 (m, 24H), 1.97 (m, 4H), 3.71–
3.80 (m, 2H), 3.85–3.91 (m, 1H), 4.17–4.26 (m, 2H),
4.84 (t, J = 5.1 Hz, 1H), 5.32 (m, 2H); 13C NMR
(CD3OD) d 12.84, 21.86, 23.33, 26.35, 28.45, 28.54,
28.71, 28.81, 28.88, 28.98, 31.16, 31.76, 32.72, 33.16,
38.22, 64.46, 64.52, 66.37, 67.49, 75.04, 75.16, 104.39,
128.97, 129.07; MS (ESI) m/z 419 [MÀH]; 31P
NMR(CD3OD) d 20.28.
30. Durgam, G. G.; Virag, T.; Walker, M. D.; Tsukahara, R.;
Yasuda, S.; Liliom, K.; van Meeteren, L. A.; Moolenaar,
W. H.; Tigyi, G.; Miller, D. D. J. Med. Chem. 2005, 48,
4919.
31. van Meeteren, L. A.; Ruurs, P.; Christodoulou, E.;
Goding, J. W.; Takakusa, H.; Kikuchi, K.; Perrakis, A.;
Nagano, T.; Moolenaar, W. H. J. Biol. Chem. 2005, 280,
21155.
32. Esterbauer, H.; Schaur, R. J.; Zollner, H. Free Radical
Biol. Med. 1991, 11, 81.
33. Zhang, C.; Baker, D. L.; Yasuda, S.; Makarova, N.;
Balazs, L.; Johnson, L. R.; Marathe, G. K.; McIntyre, T.
M.; Xu, Y.; Prestwich, G. D.; Byun, H. S.; Bittman, R.;
Tigyi, G. J. Exp. Med. 2004, 199, 763.
Compound 21: 1H NMR (300 MHz, CD3OD) d 0.90 (t,
J = 6.3 Hz, 3H), 1.27 (m, 24H), 2.56 (m, 4H), 3.77–3.78
(m, 1H), 3.87–3.95 (m, 3H), 4.22–4.25 (m, 1H), 4.98 (m,
1H), 7.06 (m, 4H); 13C NMR (CD3OD) d 12.62, 21.66,
22.86, 23.0, 28.05, 28.10, 28.24, 28.41, 30.54, 30.60, 30.90,
32.78, 34.41, 64.17, 64.69, 64.76, 66.07, 66.18, 74.15,
74.27, 74.44, 103.90, 104.41, 127.44; MS (ESI) m/z 455
1
[MÀH]; 31P NMR (CD3OD) d 19.26. Compound 31: H
NMR (300 MHz, CD3OD) d 0.86 (t, J = 6.3 Hz, 3H),
1.25 (br s, 24H), 1.98 (m, 4H), 3.64 (m, 1H), 3.89 (t,