On the preparation of some phospholipid analogues

Article abstract of DOI:10.1039/a910266n

Several structural analogues of the biocompatible diacyl glycerol phosphatidylcholine family have been prepared. Considerable variation in structure is allowed without impairing the desirable biocompatible properties. In particular, derivatives in which the fatty ester link is replaced by ether links and in which the central glycerol group is replaced by simple variants such as the symmetrical tris(hydroxymethyl)ethane group retain biocompatible properties. The Royal Society of Chemistry 2000.

Full text of DOI:10.1039/a910266n

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On the preparation of some phospholipid analogues
Judith E. Browne,a Richard T. Freeman,b Jeremy C. Russellb and Peter G. Sammes,a
a Department of Chemistry, School of Physical Sciences, University of Surrey, Guildford,
Surrey, UK GU2 5XH
1
b Biocompatibles Ltd, Frensham House, Farnham Business Park, Weydon Lane, Farnham,
Surrey, UK GU9 8QL
Received (in Cambridge, UK) 21st December 1999, Accepted 7th January 2000
Several structural analogues of the biocompatible diacyl glycerol phosphatidylcholine family have been prepared.
Considerable variation in structure is allowed without impairing the desirable biocompatible properties. In particular,
derivatives in which the fatty ester link is replaced by ether links and in which the central glycerol group is replaced
by simple variants such as the symmetrical tris(hydroxymethyl)ethane group retain biocompatible properties.
Phosphatidylcholine-containing compounds and polymers are
of importance since they have been shown to render surfaces
‘biocompatible’.1 Biocompatibility, in this context, refers to the
property of surfaces not to interact with body tissues or fluids,
such as blood, in order to avoid adhesion of cells and proteins,2
platelet activation3 and activation of the blood-clotting
cascade. Materials that possess this biocompatibility property
are of importance in a variety of medical dressings and devices
in order to avoid the potentially fatal formation of thrombi
or infection.4
The outer surfaces of erythrocytes and platelet cells are
dominated (>90%) by the presence of phosphorylcholine head
groups where the phosphate and trimethylammonium entities
are separated by two methylene groups. The biocompatibility
properties of typical phosphorylcholine esters, such as 1,
have been linked to its ability to bind water in a hydration shell
that mimics bulk water but through which proteins are unable
to penetrate.1f Water binds tightly to the phosphatidylcholine
head group, with up to twelve molecules of water being tightly
bound, as determined by various techniques.5 Little work, how-
ever, has been carried out to explore the extent to which the
chemical structure of the zwitterion determines its ability to
render surfaces biocompatible. Previous studies have concen-
trated on the pharmacological effects of synthetic phosphoryl-
choline analogues.6
compounds with increased resistance to such enzyme-mediated
hydrolysis. In the first instance, in order to limit the possible
number of structural variations, modifications to the organic
groups were made rather than trying to prepare analogues of
the phosphate ester group.
Ester variations
Our initial approach was to retain the phosphatidylcholine
group 3 and explore structural variations to the ester portion,
normally represented by the di-O-acylglycerol group, e.g. 4.
The latter group poses synthetic and stability problems in
that optically active 1,2-di-O-acylglycerol esters readily equili-
brate with the 1,3-di-O-acyl isomers, thus losing their chirality.
The ester groups are also subject to enzymic attack. Nature
has recognised this ‘stability’ problem in that certain Archae-
bacteria utilise fatty ether derivatives of glycerol in place of
fatty esters, in order to withstand some of the extreme condi-
tions experienced, e.g. thermophilic bacteria.7
In this paper we describe some recent attempts to modify
the phosphorylcholine structure, by the preparation of a series
of analogues, in order to investigate biocompatible structure–
efficacy relationships, using some standard in vitro tests for
biocompatibility.
Results and discussion
In selecting possible structural variations, cognisance was
taken of the ability of the phospholipase enzymes to hydrolyse
phospholipids (see arrows in Fig. 1) and the preference for
The first analogue studied was the known octadecyl ester of
phosphatidylcholine, 7,8 produced by reaction of octadecan-1-
ol with the dioxaphospholane 59 to give the phosphate ester
6 and then ring opening of the dioxophospholane group by
reaction with trimethylamine. However, the product 7 proved to
be an oversimplification and surfaces coated with this derivative
gave only a very poor biological performance (see Table 1).
Fig. 1 Ester-cleavage points for the phospholipases.
DOI: 10.1039/a910266n
J. Chem. Soc., Perkin Trans. 1, 2000, 645–652
This journal is © The Royal Society of Chemistry 2000
645

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Table 1 Biological resultsa
% Reduction,
platelet
% Reduction,
platelet
Concn./
% Fibrinogen
reductionb
Compound
mmol dmϪ3
adhesionc
activationd
7
11
12
17
18
19
20
20
21
21
26
27
28
29
30
30
23
14
14
14
14
14
4.2
14
4.2
14
14
14
4.2
14
4.2
14
10
90
92
94
31
88
91
100
105
96
98
66
77
Ϫ42
28
80
31
86
89
77
77
94
107
71
a Carried out at Biocompatibles Ltd. Average values given. Standard errors within the range 10%. See Experimental section for details. b Decrease
indicates reduced fibrinogen binding to coated polyethylene ribbons. c % Reduction in platelet adhesion to coated poly(vinyl chloride) ribbon quoted;
Ϫve figure indicates an increase in platelet adhesion. d % Reduction in platelet activation quoted, using coated poly(vinyl chloride) ribbon.
Effort was then concentrated on analogues bearing at least two
alkyl side chains.
length variation on biocompatibility had been done.12 Com-
pound 12 was shown to possess similar biocompatible efficacy
to the parent system 1.†
These analogues, 11 and 12, were prepared as racemates.
In order to avoid the use of the glycerol ether backbone and
problems with selective primary vs. secondary alcohol reactivity
and racemate formation, use of the novel glycerol analogue
tris(hydroxymethyl)ethane 13 was next explored. The bis-
hexadecyl ether 14 was produced by direct alkylation of the
triol (Scheme 2) and this was converted into the bromoalkyl
The racemic diether derivative 11 was next prepared (Scheme
1) in high overall yield from 1-O-tritylglycerol,10 using the
Scheme 1 Reagents: i, C16H33Br; ii, toluene-4-sulfonic acid, MeOH;
iii, 8, then H2O; iv, Me3N.
ester 8 via amination of the intermediate 10. Compound 11,
which is similar to the platelet activation factor 9,11 per-
formed in a similar manner to the corresponding ester
control 1 in coated surface tests.†,1 Thus replacement of
the ester functions by the more stable ether groups does not
appear to affect the biocompatible properties of this series of
derivatives.
Scheme 2 Reagents: i, C16H33Br; ii, either 8 or hexyl congener; iii,
nucleophiles, sealed-tube conditions.
phosphate derivatives 15 and 16, the halogeno group of which
could be replaced by reaction with various nucleophiles.
Spacer variations
Variation in the cationic group
Alongside the choline derivative 11, the homologue 12 was
prepared, in which the spacer group between the zwitterionic
groups is increased to six atoms, using the bromohexyl ana-
logue of 8. Earlier work had shown that such a variation in
structure in the parent 2 had only marginally affected its ability
to form liposomes but little work on the effect of the chain-
The intermediate 15 was aminated to form the series of
ammonium derivatives 17–20; the homologue 16 was used to
prepare the derivatives 21 and 22.
The intermediate 15 proved to be unstable to storage and
difficult to obtain pure, even after repeated chromatographic
separations. It was postulated that this instability might be due
to intramolecular bromine displacement to form the cyclic
† Tests carried out in house at Biocompatibles Ltd.
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J. Chem. Soc., Perkin Trans. 1, 2000, 645–652

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Table 2 Activities against phospholipasesa
intermediate 23, which would be expected to readily undergo
hydrolysis. In order to find out if such neighbouring group
participation was of importance in the quaternisation reac-
tions, the 13C-labelled material 25 was prepared (Scheme 3).
Phospholipase C.
Phospholipase D.
% Extent of cleavage
Compound
% Extent of cleavage
1
11
12
17
19
20
22
26
29
30
100
0
0
0
0
0
0
0
0
100
100
90
100
50
50
95
5
30
95
0
Reaction of this with trimethylamine under the standard condi-
tions afforded 13C-labelled 17 in which no scrambling of the
label was observed, indicating that, in this case, intermediate
formation of the cyclic ester 23 was not occurring. Possibly, a
general neighbouring group effect, due to the hydrated phos-
phate function, explains the lability of the bromoethyl group.
Reaction of more hindered nucleophiles with the inter-
mediate 15 often proved low yielding or abortive and, there-
fore, as an alternative method, the dichlorophosphate 24
was initially prepared, and reacted with a range of substituted
alcohols under conditions favouring monoester formation.
In this manner the derivatives 26–28 were prepared.
a See Experimental section for details. Results are averages of multiple
runs; standard errors within 10%.
in fibrinogen adsorption and platelet activation but a very
large platelet-adhesion effect. This result suggests that these
coated surfaces are preferentially adsorbed without the normal
activation, a property that is of potential utility in certain
applications.
Scheme 3
Some initial studies on the stability of these analogues to
enzymic degradation were also carried out; specifically,
resistance to phospholipases C and D. These are known to
hydrolyse at the phosphate ester junctions indicated in Fig. 1
and the results are included in Table 2. Dipalmitoylphos-
phatidylcholine (DPPC) 1 was used as the reference compound
and a literature assay method was used for the lipase C.13 Lipase
D was indirectly estimated by oxidation of the liberated choline
using a commercially available assay kit (see Experimental
section).
As can be seen from the results listed in Table 2, all the
analogues tested were resistant to hydrolysis by phospholipase
C, indicating that the diether linkage was unable to interact
with the enzyme. Phospholipase D was much less specific
and was capable of hydrolysing a wide variety of substrates. Of
these, the triethylammonium analogue 26 was the least affected,
indicating that the cationic site on the enzyme is not able to
accept such a large entity.
Non-alkyl zwitterionic spacers
Simple extension of the ethyl spacer, in the zwitterionic
phosphatidylcholine group, from C2 to C6 had only a marginal
effect on the behaviour of these analogues in in vitro tests.
Some examples of non-alkyl spacers were therefore prepared,
including compounds 29 and 30. The former was obtained
by reaction of 3-(dimethylamino)phenyl dichlorophosphate
with the alcohol 14, followed by hydrolysis to the phosphate
and then methylation of the amine with methyl iodide.
The 3-hydroxypyridine derivative 30 was prepared by initial
preparation of 1-methyl-3-oxidopyridinium 31 and reaction of
this with the dichlorophosphate 8.
Biological results
Table 1 lists representative biocompatibility results obtained
with some of these tris(hydroxymethyl)ethane derivatives.
The surface properties of the prepared compounds were deter-
mined after coating from an ethanolic solution onto either
polyethylene tubing, for fibrinogen adsorption, or onto poly-
(vinyl chloride) ribbon for determination of the adherence of
activated platelets. All results are presented as % reductions
when compared with untreated controls.
The quinuclidinium 18 and dimethylsulfonium 27 derivatives
both gave reductions in the fibrinogen assay of 31% and the
latter surprisingly large reductions on both platelet adhesion
(77%) and activation (77%) assays. The hexyl-spaced triphenyl-
phosphonium compound 21 showed only a moderate reduction
Conclusions
The above results show that considerable latitude is possible in
the design of effective biocompatible analogues of normal
phosphatidylcholine esters, such as DPPC 1. Thus, (i) the
zwitterionic groups can be separated by a spacer length of up
to six atoms without undue loss of biocompatibility; (ii) two
long-chain alkyl groups are required but these may be linked
by an ether link rather than an ester group; (iii) the central
glyceryl group may be replaced by simple variants such as the
tris(hydroxymethyl)ethane structure. The modified structures,
J. Chem. Soc., Perkin Trans. 1, 2000, 645–652
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such as compounds 17 and 26, retain biocompatibility but are
much more resistant to enzymic degradation than the natural
phosphatidylcholine system.
1-O-(Cholinephosphoryl)-2,3-di-O-hexadecylglycerol 11
1,2-Di-O-hexadecyl-3-O-tritylglycerol. 1-O-Tritylglycerol
(5.0 g, 15 mmol prepared as per the literature10) was dissolved
in dry DMF (50 cm3), sodium hydride (1.32 g as a 60% w/w
dispersion in paraffin oil, prewashed with hexane; 33 mmol)
was added and the mixture was stirred at room temperature for
15 min before addition of 1-bromohexadecane (9.13 g, 30
mmol). The mixture was stirred at 40 ЊC for 16 h, cooled, and
partitioned between chloroform (100 cm3) and water (100 cm3).
The organic phase was washed successively with saturated
aq. NaHCO3 (100 cm3) and brine (100 cm3) before drying,
filtering, and removal of solvent under reduced pressure. The
product was purified by chromatography through silica and
eluted with 2:23 ethyl acetate–light petroleum, to give pure 1,2-
di-O-hexadecyl-3-O-tritylglycerol (8.0 g, 68%) as a waxy solid
[Found (FAB MS): MHϩ, 747. C51H87O3 requires m/z 747].
Experimental
Microanalyses were carried out by MEDAC Ltd, Brunel
Science Park, Egham, Surrey. TLC used silica gel 60 F254-coated
glass plates, 0.25 mm, from BDH or, for preparative TLC
(PLC), glass plates precoated with 1 mm of the same material.
NMR spectroscopy was carried out on either a Bruker AC300
spectrometer (1H at 300 MHz, 13C at 75 MHz) or a JEOL
FX200 instrument (1H at 200 MHz); FAB mass spectra
were obtained on a Kratos Concept 1S instrument, and EI
and CI spectra using a VG Trio 2000 machine. Accurate
mass measurements were carried out at the EPSRC National
Mass Spectrometry Service Centre, University of Wales,
Swansea. Where necessary, solvents were dried and distilled
before use.14 Light petroleum refers to the fraction of boiling
range 40–60 ЊC; for chromatography analytical grade solvents
were used as supplied from Romil Chemicals (Cambridge, UK).
Solutions were generally dried over anhydrous sodium sulfate
before removal under reduced pressure using a rotary evapor-
ator. Bromoethyl dichlorophosphate 8 was prepared by the
method of Hirt and Berchtold.15 Where relevant, derivatives are
generally named as esters of the standard phosphatidylcholine
head group, 4.
1,2-Di-O-hexadecylglycerol. 1,2-Di-O-hexadecyl-3-O-trityl-
glycerol (8.0 g, 10.2 mmol) was dissolved in methanol (100 cm3)
with toluene-4-sulfonic acid (0.6 g) and the solution was stirred
at room temperature for 16 h. The bulk of the solvent was
removed under reduced pressure and the residue was par-
titioned between chloroform (100 cm3) and water (100 cm3).
The organic phase was washed successively with water (2 × 100
cm3), and brine (100 cm3) before drying, filtering, and evaporat-
ing to dryness. The residue was chromatographed through silica
and eluted with light petroleum and light petroleum–ethyl
acetate (23:2) to give 1,2-di-O-hexadecylglycerol (3.0 g, 54%) as
a white solid (Found: MHϩ, 541. C35H73O3 requires m/z 541);
δH (300 MHz) 0.88 (6 H, t, J 7 Hz, 2 × CH3), 1.26 [52 H, m,
2 × (CH2)13], 1.56 (4 H, m, 2 × CH2CH2O), 3.4–3.75 (9 H, m,
CHO and CH2O).
Biological assays
The % fibrinogen-reduction, platelet-adhesion and platelet-
activation studies were carried out in house by Biocompatibles
Ltd, using standard protocols. Materials were dissolved in
ethanol before coating onto either polyethylene tubing for the
fibrinogen-adsorption studies or, for the platelet-adhesion and
-activation assays, onto poly(vinyl chloride) ribbon at a con-
centration of 14 mmol dmϪ3 for the former and 4.2 mmol dmϪ3
for the latter.
The activity of phospholipase C (from Clostridium perfrin-
gens) followed the method of Zwaal and Roelofsen,13 using
DPPC 1 as the control, and employing either a colorimetric
determination of the liberated phosphate monoester or TLC to
follow the formation of the liberated alcohol.
Phospholipase D from Streptomyces chromofuscus was
employed using a modification of the method described by
Yang.16 This involves a colorimetric determination (estimation)
of hydrogen peroxide formed during the oxidation of the
released choline. The method was also monitored by TLC
detection of the liberated hydrolysis products.
1,2-Di-O-hexadecylglycerol (0.50 g, 0.924 mmol) was dis-
solved in dry diethyl ether (10 cm3) under an atmosphere of
dry nitrogen and treated with triethylamine (0.22 cm3, 1.57
mmol) followed by the dropwise addition of a solution of 2-
bromoethyl dichlorophosphate 8 (0.33 g, 1.57 mmol) in diethyl
ether (5 cm
3) and the mixture was stirred at room temperature
for 5 h. Further portions of triethylamine (0.22 cm3) and the
dichlorophosphate (0.66 g) were added and the mixture was
heated to reflux for 16 h. A final addition of triethylamine
(0.22 cm3) and the dichlorophosphate (0.33 g) was made and
the mixture was heated for a final 1 h. The reaction mixture was
cooled, triethylamine (1.8 cm3) and water (1 cm3) were added,
and the mixture was stirred for a further 2 h. The bulk of
the ether was removed under reduced pressure and the residue
was dissolved in dichloromethane (100 cm
3) and washed suc-
cessively with water (2 × 100 cm3) and brine (100 cm3) before
drying and filtering. Removal of the solvent afforded com-
pound 10 as an oil, which was immediately dissolved in
acetonitrile (28 cm3) and chloroform (40 cm3). The solution
was heated with trimethylamine (1.5 g, 25 mmol) in a sealed
vessel at 50 ЊC for 14 h. The solvents were then removed and
the residue chromatographed through silica, and eluted with
chloroform–methanol–25% ammonia in the proportions
690:270:64 to afford one major product. Fractions containing
the product were combined, evaporated to dryness and azeo-
troped with benzene before drying over P2O5 in vacuo for 3 h to
afford the title compound 11 (393 mg, 61%) as a hygroscopic
solid (Found: MHϩ, 706. C40H85NO6P requires, m/z 706);
δH (1:20 CD3OD–CDCl3) 0.88 (6 H, t, J 7 Hz, 2 × CH3), 1.26
(52 H, m), 1.54 (4 H, m), 3.42 [9 H, s, (CH3)3Nϩ], 3.57 (6 H, m),
3.88 (5 H, m), 4.33 (2 H, m); δC (1:20 CD3OD–CDCl3) 14.1,
22.7, 26.0, 29.9, 32.2, 54.6 [(CH3)3Nϩ], 58.8, 66.3, 68.6, 70.2,
71.9, 78.9.
Octadecyl phosphatidylcholine8 7
To a mixture of octadecan-1-ol (10 g, 36.9 mmol) and triethyl-
amine (3.74 g, 1 equiv.) in diethyl ether (100 cm3), at room
temperature, was added dropwise a solution of 2-chloro-2-oxo-
1,3,2-dioxaphospholane9 5 (5.27 g, 36.9 mmol) in diethyl
ether (10 cm3). The reaction mixture was stirred for 2 h, then
filtered and the solvent removed. The residue was dissolved
in 1:1 v/v acetonitrile–chloroform (150 cm3) before addition
of trimethylamine (6 g, 0.101 mol) in acetonitrile (15 cm3)
and the mixture was stirred at 50 ЊC for 16 h in a pressure
bottle. On cooling of the reaction mixture to room tem-
perature, a solid precipitated, which was filtered off and
recrystallised from dichloromethane to yield the title
compound (3.13 g, 19%); mp >220 ЊC (decomp.) (lit.8 235 ЊC
decomp.) (Found: C, 59.1; H, 11.4; N, 2.9, P, 6.7. Calc. for
C23H50NO4Pؒ2H2O: C, 58.7; H, 11.5; N, 3.0; P, 6.6%); δH (200
MHz; CDCl3–CD3OD) 0.87 (3 H, m, MeCH2), 1.25 (30 H, br s,
aliphatic H), 1.6 (2 H, m, CH2CH2 OP), 3.25 (9 H, s, Me3Nϩ),
3.65 (2 H, m, CH2Nϩ), 3.8 (2 H, q, CH2OP), 4.25 (2 H, m,
POCH2CH2N); m/z (FAB) 436 (M ϩ Hϩ; 100%).
2,3-Bis-O-(hexadecyloxy)propyl 6-(trimethylammonio)hexyl
phosphate 12
This was prepared in a similar manner to the choline derivative
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J. Chem. Soc., Perkin Trans. 1, 2000, 645–652

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11, using 6-bromohexyl dichlorophosphate in place of the ethyl
homologue. The title compound was obtained in 51% yield
(Found: MHϩ, 763. C44H93NO6P requires m/z, 763); δH (1:20
CD3OD–CDCl3) 0.88 (6 H, t J 7 Hz) 1.25 (52 H, m), 1.50 (8 H,
m), 1.60 (2 H, m), 1.80 (2 H, m), 3.25 [9 H, s, (CH3)3Nϩ],
3.39–3.60 (9 H, m), 3.63 (4 H, m).
J 6 Hz), 3.42 [9 H, s, (CH3)3Nϩ], 3.67 (2 H, d, J 5 Hz), 3.84 (2 H,
s), 4.3 (2 H, s); δC 14.1, 17.1, 22.6, 26.2, 29.3, 29.7, 31.9, 40.7,
54.4, 59.0, 66.4, 68.6, 71.6, 72.9 (Found: MHϩ, 735.6507.
C42H89NO6P requires m/z, 735.6506).
Quinuclidinium derivative 18. Elution from the column with
chloroform–methanol–water (70:25:5, v/v) afforded the
product (23%); δH (CDCl3–CD3OD), 0.88 (6 H, t, MeCH2), 0.95
(3 H, s, MeC), 1.25 (53 H, m, aliphatic H), 1.49 (4 H, m,
OCH2CH2), 2.02 (6 H, m, CH2CH2Nϩ), 3.29 (4 H, s, OCH2C),
3.36 (4 H, t, OCH2CH2), 3.46 (2 H, m, NϩCH2CH2), 3.59 (6 H,
t, CH2Nϩ), 3.71 (2 H, d, POCH2C), 4.21 (2 H, m, NϩCH2-
CH2OP); δC (CDCl3) 14.09 (CH3CH2), 17.13 (CH3C), 19.65
(HCCH2CH2Nϩ), 22.66 (CH2CH3), 24.05 (HCCH2CH2Nϩ),
26.19 (OCH2CH2CH2), 26.94 [(CH2)10 ϩ OCH2CH2], 31.90
(CH2CH2CH3), 40.7 (C), 55.50 (HCCH2CH2Nϩ), 58.29 (CH2-
CH2OP), 64.63 (POCH2C), 68.46 (NϩCH2CH2OP), 71.60
(OCH2CH2) and 73.03 (CCH2O) (Found: C, 66.9; H, 11.9; N.
1.7; P, 4.2. C46H92NO6Pؒ2H2O requires C, 67.1; H, 11.8; N, 1.7;
P, 3.8%).
2,2-Bis(hexadecyloxymethyl)propan-1-ol 14
1,1,1-Tris(hydroxymethyl)ethane 13 (30 g, 0.25 mol) was added
in portions to a stirred suspension of fresh sodium hydride
(12 g, from 60% w/w suspension in mineral oil washed with
hexane and dried under nitrogen; 0.5 mol) in DMF (200 cm3).
The mixture was stirred at room temperature for 1 h before the
addition of bromohexadecane (167.7 g, 0.55 mol). The mixture
was then heated, with stirring, at 50 ЊC for 44 h. After the
mixture had cooled, water was added to destroy the residual
sodium hydride and the mixture was then partitioned between
dichloromethane (200 cm3) and water (200 cm3). The organic
layer was washed successively with more water, dil. HCl (0.1 M)
and brine before drying, filtering, and removal of solvent under
reduced pressure to produce the product as a waxy solid (150 g,
98%). Portions of the product were purified as required by
column chromatography through silica gel, and elution with
9:1 light petroleum–ethyl acetate. The title product showed
δH (CDCl3) 0.85 (3 H, s, MeC), 0.9 (6 H, t, MeCH2), 1.26 (52 H,
aliphatic H), 1.53 (4 H, m, OCH2CH2), 2.06 (1 H, s, OH), 3.41
(8H, OCH2), 3.55 (2 H, s, HOCH2); δC 13.8, 17.2, 22.3, 25.8,
29.0, 29.3, 31.5, 40.0, 69.5, 71.4, 75.1 (Found: C, 78.4; H, 14.0.
C37H76O3 requires C, 78.2; H, 13.5%).
Dimethylammonium derivative 19. The product was chroma-
tographed twice, eluting from the column with 9:1 chloroform–
methanol, to give the dimethylammonium derivative (22%);
δH (CDCl3–CD3OD) 0.88 (6 H, t, MeCH2), 0.95 (3 H, s, MeC),
1.25 (52 H, m, aliphatic H), 1.49 (4 H, OCH2CH2), 2.89 (6 H,
s, Me2NHϩ), 3.25 (2 H, m, CH2Nϩ), 3.28 (4 H, s, OCH2C), 3.35
(4 H, t, OCH2CH2), 3.75 (2 H, POCH2C), 4.2 (2 H, m,
CH2CH2OP) (Found: Mϩ, 719.6192. C41H86NO6P requires M,
719.6208).
2,2-Bis(hexadecyloxymethyl)propyl 2-bromoethyl phosphate 15
Pyridinium derivative 20. The product was chromatographed
using 70:25:5 chloroform–methanol–water as eluent, to give
the product (9%); δH (CDCl3) 0.88 (6 H, t, MeCH2), 0.93 (3 H, s,
Me), 1.25 (52 H, m, aliphatic H), 1.48 (4 H, t, OCH2CCH2),
3.27 (4 H, s, OCH2C), 3.35 (4 H, t, OCH2CH2), 3.67 (2 H, d,
POCH2C), 4.26 (2 H, m, NϩCH2CH2C), 4.82 (2 H, t, Nϩ-
CH2CH2OP), 8.04 (2 H, t, CHCHNϩ), 8.44 (1 H, t, CHCH-
CHNϩ), 9.06 (2 H, d, CHNϩ); δC (CDCl3–CD3OD) 14.05
(CH2CH3), 16.97 (CH3C), 22.61 (CH2CH3), 26.11 (OCH2-
CH2CH2), 29.64 [(CH2)10 ϩ OCH2CH2), 31.85 (CH2CH2CH3),
40.6 (C), 62.18 (NϩCH2CH2OP), 68.5 (POCH2C), 71.6
(OCH2CH2), 72.8 (CCH2O), 128.06 (CH:CHNϩ) and 145.5
(CHNϩ ϩ HCCH:CHNϩ); m/z (FAB) 754 (M ϩ Hϩ, 56%)
(Found: C, 67.1; H, 11.4; N, 1.8; P, 3.8. C4H84NO6Pؒ2H2O
requires C, 66.9; H, 11.2; N, 1.8; P, 3.9%).
To a solution of the alcohol 14 (1 g, 1.75 mmol) in diethyl ether
(20 cm3), kept under an atmosphere of dry nitrogen, was added
triethylamine (0.60 g, 6 mmol) and then, dropwise, bromoethyl
dichlorophosphate 8 (1.26 g, 6 mmol). The mixture was heated
to 50 ЊC for 16 h before cooling and the addition of triethyl-
amine (0.6 g, 6 mmol) and water (4.09 g, 227 mmol); the
mixture was then heated, with stirring, for a further 2 h at 50 ЊC.
After cooling, the water layer was removed and the residue was
dried by azeotroping with benzene (3 × 20 cm3). The residue
was dissolved in diethyl ether (20 cm3) and the solution was
filtered, and evaporated under reduced pressure to afford the
crude product, which was further purified by column chroma-
tography, and elution with chloroform and chloroform–
methanol (5:1) mixtures to give the desired product 15 (0.32 g,
34%) as a waxy solid, δH (200 MHz, CDCl3) 0.9 (9 H, m,
3 × Me), 1.25 (52 H, m, aliphatic H), 1.50 (4 H, m, OCH2CH2),
3.25 (4 H, s, OCH2C), 3.38 (4 H, t, OCH2CH2), 3.51 (2 H, t,
BrCH2), 3.75 (2 H, s, POCH2), 4.2 (2 H, m, BrCH2CH2OP). The
bromoethyl ester was used, without further characterisation,
to prepare the derivatives 17–20.
6-(Triphenylphosphonio)hexyl phosphate analogue 21
6-Bromohexyl dichlorophosphate 8 (n ؍ 6). Distilled phos-
phoryl trichloride (0.85 g, 5.5 mmol) in dichloromethane
(10 cm3) was added dropwise to a solution of 6-bromohexan-1-
ol (1 g, 5.5 mmol) in dichloromethane (10 cm3) under nitrogen.
The mixture was left for 16 h at room temperature before
removal of the solvent and excess of phosphoryl trichloride
under reduced pressure. The crude product was used in
subsequent steps.
General method for reaction of 15 with nucleophiles
The nucleophile (20–30 mmol) was added to a solution of
the bromoethyl ester 15 (1 mmol) in dry chloroform (10 cm3)
and the solution was stirred at 60 ЊC for 16 h, following the
disappearance of the starting halide by TLC. The reaction
mixture was cooled, the solvent and excess of nucleophile
removed by evaporation under reduced pressure, and the
residue chromatographed through silica gel using chloroform–
methanol–water mixtures, generally in the range 65:25:4
respectively.
6-Bromohexyl 2,2-bis(hexadecyloxymethyl)propyl hydrogen
phosphate 16. 2,2-Bis(hexadecyloxymethyl)propan-1-ol 14
(1.00 g, 1.76 mmol) was dissolved in dry diethyl ether (50 cm3)
and to this was added a mixture of triethylamine (0.35 g, 3.5
mmol) and 6-bromohexyl dichlorophosphate 8 (n = 6) (1.05 g,
3.6 mmol) in dry diethyl ether (20 cm3). The reaction mixture
was stirred for 48 h at room temperature before addition of
more triethylamine (0.35 g, 3.5 mmol) and water (1 cm3) and
the mixture was heated to reflux for 3 h before being cooled to
room temperature. The aqueous layer was removed and the
organic layer evaporated under reduced pressure. The residue
was azeotroped with benzene before removal of the solvent
and drying of the residue under reduced pressure. Trituration
Trimethylammonium derivative 17. Carried out on 0.33 mmol
scale, acetonitrile (10 cm3) was used as a co-solvent with chloro-
form (2.5 cm3) for the trimethylamine (0.52 g, 9.9 mmol). The
product (240 mg, 38%) showed δH (CDCl3–CD3OD) 0.88 (6 H,
t, CH2Me), 0.95 (3 H, s, MeC), 1.25 (52 H, m, aliphatic H),
1.42–1.52 (4 H, m, OCH2CH2), 3.26 (4 H, s), 3.34 (4 H, t,
J. Chem. Soc., Perkin Trans. 1, 2000, 645–652
649

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of the crude product with diethyl ether afforded, as a precipi-
tate, the title phosphate (1.40 g, 98%). This was used directly in
the next step. The obtained material showed δH (200 MHz;
CDCl3) 0.9 (9 H, m, Me), 1.25 (52 H, m, aliphatic H), 1.4–1.6
(8 H, m, CH2), 1.9 (4 H, m), 3.28 (4 H, s, CCH2O), 3.3–3.6 (8 H,
m, CH2), 4.05 (2 H, m, CH2OP).
required ester 25 (0.50 g, 93%), δC (CDCl3) 14.12 (H2CMe),
17.62 (CMe), 22.71 (MeCH2), 26.18, 29.71 and 31.93 (CH2),
45.72 (CH2Br), 65.71 and 66.81 (CH2OP), 71.84 and 75.56
(CH2O).
The labelled bromide 25 (0.25 g, 0.33 mmol) as a solution in
chloroform (2.5 cm3) was treated with a solution of trimethyl-
amine (0.59 g, 9.9 mmol) in acetonitrile (10 cm3) in a pressure
bottle and heated to 60 ЊC for 16 h. After cooling to room
temperature some of the solvent was removed under reduced
pressure until a precipitate just started to form. The mixture
was then cooled to Ϫ20 ЊC for 20 min and the precipitate col-
lected (0.43 g) before purification by column chromatography
through silica gel, and elution with chloroform and then
690:270:64 chloroform–methanol–ammonia to afford the
labelled ester 17 (32 mg, 13%; 24% isotopic abundance). The
13C NMR spectrum indicated all the label was in the starting
position, δC (CDCl3) 14.0 (MeCH2), 17.0 (MeC), 22.6 (Me-
CH2), 26.1, 29.6 and 31.9 (CH2), 40.7 (C), 54.3 (Me3Nϩ),
The bromide 16 (1.66 g, 1.9 mmol) was dissolved in dry
chloroform (40 cm3) and a solution of triphenylphosphine (2.32
g, 8.8 mmol) in acetonitrile (20 cm3) was added. The solution
was sealed in a pressure vessel under nitrogen and heated
at 70 ЊC for 16 h. More triphenylphosphine (2.32 g, 8.8 mmol)
was added and the solution reheated for a further 16 h.
The crude material obtained after removal of solvent was chrom-
atographed through silica gel, and eluted with chloroform and
then 690:270:64 chloroform–methanol–conc. ammonia to
afford the crude major product. The material was further puri-
fied by PLC to afford the pure triphenylphosphonium analogue
21 (17 mg, 3.1%) (Found: MHϩ, 993.7229. C61H103O6P2 requires
m/z, 993.7199); δH (CDCl3–CD3OD) 0.88 (9 H, 3 × Me), 1.25
[52 H, br s, 2 × (CH2)13], 1.36–1.48 [8 H, m, 2 × OCH2-
CH2 ϩ (CH2)2CH2CH2OP], 1.61 [4 H, m, CH2(CH2)2CH2CH2-
OP], 3.20 (4 H, s, 2 × CCH2O), 3.29 (4 H, t, 2 × OCH2CH2),
3.47 (2 H, m, PϩCH2), 3.67 (2 H, d, POCH2), 3.80 (2 H, m,
CH2CH2OP), 7.7–7.8 (15 H, ArH); δC (CDCl3–CD3OD) 13.34
(CH2CH3), 16.48 (CH3C), 18.21 (CH2Pϩ), 21.20 (CH2CH2Pϩ),
22.0 (CH2CH3), 24.46 [(CH2)2(CH2)2OP], 25.69 (OCH2CH2-
CH2), 29.18 [(CH2)10 ϩ OCH2CH2 ϩ CH2CH2OP], 31.42 (CH2-
CH2CH3), 40.2 (C), 64.4 (CH2OP), 67.83 (POCH2C), 71.19
(OCH2CH2), 72.48 (CCH2O), 117 (CPϩ), 130.0 (CHCHCPϩ),
132.9 (CHCPϩ) and 134.9 [CH(CH2)2CPϩ]; δP (121 MHz,
CDCl3–CD3OD) Ϫ0.99 (1 P, s), 25.01 (1 P, s).
59 (CH2OP), 66.5 (CCH2OP), 68.5 (Me3NϩCH2), 71.6 and
12
72.8 (CCH2O) [Found: Mϩ(13C), 736.6524.
requires M, 736.6539].
C
13CH90NO6P
41
2,2-Bis(hexadecyloxymethyl)propyl dichlorophosphate 24
To a solution of freshly distilled phosphorus oxychloride (0.4 g,
2.64 mmol) dissolved in ether (10 cm3) was added, at room
temperature, a solution of triethylamine (0.18 g, 1.76 mmol)
and 2,2-bis(hexadecyloxymethyl)propanol (1.00 g, 1.76 mmol)
in ether (10 cm3). The mixture was then stirred at 40 ЊC for 16 h
in a sealed vessel before cooling to room temperature, filtering
off the precipitated triethylammonium chloride and concen-
trating the filtrate under reduced pressure. The labile acid halide
24 (0.85 g, 70%) was used without further purification. The
viscous oil showed δH (200 MHz, CDCl3) 0.85 (6 H, t, MeCH2),
1.0 (3 H, s, Me), 1.25 (52 H, m, CH2), 1.5 (4 H, m, OCH2CH2),
3.25 (4 H, s, CCH2O), 3.35 (4 H, t, OCH2CH2), 4.26 (2 H, d,
POCH2).
Preparation of the parent 2-(triphenylphosphonio)ethyl
derivative was accomplished in very small quantity and the
material was not fully characterised.
13C-Labelled trimethylammonium derivative 17
2-Bromo[1-13C]ethanol. To a stirred solution of bromoacetic-
[13C] acid (0.5 g, 3.6 mmol; 99% isotopic abundance) in dry
tetrahydrofuran (5 cm3), at 0 ЊC under nitrogen, was added
borane–tetrahydrofuran complex (1 M; 4.3 cm3, 4.3 mmol)
dropwise during 10 min. After stirring for a further 20 min at
0 ЊC the solution was warmed to room temperature and stirred
for a further 64 h. Aq. HCl (3 M; 1.5 cm3) was added and the
solution stirred for a further 90 min before addition of more
water (10 cm3) and extraction into diethyl ether (4 × 10 cm3);
the extract was dried, filtered, and the volume reduced by
distillation under atmospheric pressure to ≈10 cm3. The solu-
tion was diluted with unlabelled bromoethanol (1.35 g) and
the rest of the solvent was removed before microdistillation
of the product to give labelled bromoethanol (1.03 g; 24%
isotopic abundance); δC (50 MHz; CDCl3) 35.5 (H2CBr), 62.5
(H2COH).
Triethylammonioethyl phosphate analogue 26
2-Bromoethanol (2.0 g, 16 mmol) and triethylamine (1.62 g,
16 mmol) were heated in dichloromethane (40 cm3) at reflux
for 16 h. The resultant precipitate was filtered off and dried
over phosphorus pentaoxide in vacuo to afford 2-triethyl-
ammonioethanol bromide (1.47 g, 41%), δH (D2O) 1.3 (9 H, t,
Me), 3.4 (8 H, m, CH2Nϩ), 4.0 (2 H, m, CH2OH).
The salt (0.18 g, 1.23 mmol) and triethylamine (0.12 g,
1.23 mmol) in diethyl ether (10 cm3) were added to a solution
of the dichlorophosphate 24 (0.843 g, 1.23 mmol) in diethyl
ether (5 cm3). The solvent was removed and replaced with
acetonitrile (20 cm3), and the solution heated to reflux for 16 h.
Water (1.33 g, 74 mmol) and triethylamine (0.12 g, 1.23 mmol)
were added and the mixture was heated to reflux for a further
2 h, before being evaporated to dryness, with azeotroping with
benzene to remove the water. The residue was chromatographed
through silica gel, and eluted with 690:270:64 chloroform–
methanol–ammonia, to give the required title product (14 mg,
1.5%); δH (CDCl3) 0.88 (6 H, t, MeCH2), 0.93 (3 H, s, Me), 1.26
(52 H, m, aliphatic H), 1.36 (9 H, m, MeCH2Nϩ), 1.50 (4 H,
t, OCH2CH2), 3.26 (4 H, s, CCH2O), 3.35 (4 H, t, OCH2CH2),
3.48 (6 H, m, CH2Nϩ), 3.66 (4 H, m, CH2Nϩ, CCH2OP), 4.29
(2 H, m, NϩCH2CH2OP) (Found: MHϩ, 776.6865. C45H95NO6P
requires m/z, 776.6897).
2-Bromo[1-13C]ethyl dichlorophosphate 13C-8. A solution of
the labelled bromoethanol (1.03 g, 8.2 mmol) in dichloro-
methane (5 cm3) was added, dropwise, to a solution of freshly
distilled phosphoryl trichloride in dichloromethane (5 cm3)
and the solution was stirred for 16 h under nitrogen. The
solvent was removed and the residue distilled to give the
dichlorophosphate as a pale yellow liquid (0.551 g; 26%
isotopic abundance).
Preparation of 13C-labelled ester 25. To the labelled chloride
13C-8 (0.27 g, 1.2 mmol) was added dropwise, over a period
of 1 min, a solution of the alcohol 14 (0.40 g, 0.71 mmol)
and triethylamine (0.12 g, 1.2 mmol) in diethyl ether (10 cm3).
The mixture was stirred, whilst warming to 40 ЊC, for 2 h.
After cooling, the solvent was removed and the residue
azeotroped with benzene. The dry residue was triturated
with diethyl ether and filtered, when evaporation afforded the
2-(Dimethylsulfonio)ethyl phosphate analogue 27
2-(Dimethylsulfonio)ethyl toluene-4-sulfonate (0.49 g, 1.75
mmol), the dichlorophosphate 24 (1.20 g, 1.75 mmol) and
triethylamine (0.18 g, 1.75 mmol) were heated in acetonitrile
(10 cm3) at 50 ЊC for 16 h. The reaction mixture was cooled, and
concentrated in a stream of nitrogen, before chromatography
through silica gel, and elution with chloroform and then
650
J. Chem. Soc., Perkin Trans. 1, 2000, 645–652

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690:270:64 chloroform–methanol–ammonia. The fractions
containing the product were evaporated and the solid residue
was triturated with hexane (5 × 10 cm3) and acetone (10 ×
10 cm3) before the solid precipitate was collected by centri-
fugation. The solid was dried over phosphorus pentaoxide to
afford the sulfonium product (40 mg, 3%), δH (CDCl3–CD3OD)
0.88 (6 H, t, Me), 0.95 (3 H, s, Me), 1.25 (52 H, m, aliphatic H),
1.49 (4 H, m, OCH2CH2), 3.04 (6 H, s, Me2Sϩ), 3.28 (4 H, s,
C CH2O), 3.36 (4 H, t, OCH2CH2), 3.58 (2 H, m, SϩCH2), 3.71
(2 H, d, POCH2C), 4.26 (2 H, m, POCH2CH2); δC (CD3OD–
CDCl3) 14.0, 17.0, 22.9, 26.5, 27.0, 30.0, 32.0, 41.2, 46.4, 59.5,
69.0, 72.3, 73.5 [Found: (M ϩ Hϩ), 737.5904; C, 66.8; H, 11.6;
S, 4.3. C41H85O6SP requires M ϩ H, 737.5882; C, 66.8; H, 11.6;
S, 4.35%].
before drying (under reduced pressure) to afford a waxy residue
(1.86 g). This was chromatographed through silica gel with 5:1
chloroform–methanol as eluent to give 2,2-bis(hexadecyloxy-
methyl)propyl 3-(dimethylamino)phenyl hydrogen phosphate
(350 mg, 13%); δH 0.85 (9 H, m), 1.25 (52 H, m), 1.4 (4 H, m),
2.8 (6 H, s), 2.9–3.35 (8 H, m), 3.75 (2 H, m), 6.35 (1 H, m), 6.65
(2 H, m), 7.0 (1 H, s). This material was used in the next step
without further purification.
Potassium carbonate (0.08 g, 0.64 mmol) and 18-crown-
6 (0.17 g, 0.64 mmol) were stirred at room temperature for
20 min in dichloromethane (4 cm3) and to this mixture was
added a solution of the above dimethylaminophenyl phosphate
(0.35 g, 0.45 mmol) in dichloromethane (4 cm3), followed
by iodomethane (8 cm3). The reaction mixture was sealed
under nitrogen and heated at room temperature for 40 h. The
solvent and excess of iodomethane were removed under
reduced pressure; the residue was dissolved in acetonitrile
(10 cm3) and the solution was filtered and then dried. The
solid product was purified by PLC on silica gel with chloro-
form–methanol–water (65:25:4) as eluent. The major band
was collected by extraction with chloroform–methanol and
evaporated to give a solid (50 mg), which was re-dissolved in
chloroform (1 cm3), and acetone was added dropwise to precipi-
tate the solid. The solid was collected, and dried over phos-
phorus pentaoxide as the title compound (14.9 mg, 4.2%);
δH 0.88 (9 H, m), 1.25 (52 H, m), 1.48 (4 H, m), 3.25 (4 H, s),
3.31 (4 H, t, J 7 Hz), 3.71 (9 H, s), 3.86 ( 2 H, d, J 3 Hz), 7.25–
7.5 (3 H, m) and 7.92 (1 H, s); δC 14.10, 17.10, 22.66, 26.16m,
29.69, 31.90, 40.70, 57.22, 69.01, 71.55, 72.89, 112.40, 122.63,
130.56, 147.30, 155.40 (Found: MHϩ, 782.6421. C46H89NO6P
requires m/z, 782.6427).
2-(Hexyldimethylammonio)ethyl phosphate analogue 28
2-(Hexyldimethylammonio)ethyl toluene-4-sulfonate (0.302 g,
0.44 mmol), prepared from 2-(4-tolylsulfonyloxy)ethanol and
hexyldimethylamine, the chlorophosphate ester 24 (0.302 g,
0.44 mmol) and triethylamine (0.044 g, 0.44 mmol) were heated
in dichloromethane (5 cm3) in a pressure bottle at 50 ЊC for
16 h. Water (1 cm3) was then added together with triethylamine
(0.044 g, 0.44 mmol) and the mixture was heated at 50 ЊC for a
further 6 days. The reaction mixture was evaporated and the
residue chromatographed through silica gel, eluting with, first,
chloroform and then with 690:270:64 chloroform–methanol–
ammonia. The fractions containing the phosphatidylcholine
analogue were combined, dried and the residual solid triturated
with acetone to give the title product (7 mg, 2%), δH (CDCl3–
CD3OD) 0.80–0.95 (12 H, m), 1.25 (52 H, aliphatic H), 1.35
(6 H, m, CH2), 1.51 (4 H, OCH2CH2), 1.65 (2 H, m CH2), 3.15
(6 H, s, Me2Nϩ), 3.22 (4 H, s, CH2O), 3.35 (6 H, m, CH2O and
CH2Nϩ), 3.60 (2 H, m, NϩCH2), 3.70 (2 H, m, POCH2OP),
4.23 (2 H, m, CH2OP) (Found: MHϩ, 804.7203. C47H99NO6P
requires m/z, 804.7210).
N-Methylpyridinium-3-yl phosphate analogue 30
3-Hydroxypyridine (0.5 g, 5.2 mmol) and methyl toluene-4-
sulfonate (1.47 g, 7.9 mmol) were heated in acetonitrile (15 cm3)
at 50 ЊC for 16 h in a sealed vessel. The resulting precipitate was
collected, and dried over phosphorus pentaoxide in vacuo for
48 h. The solid was suspended in acetonitrile (20 cm3) and the
suspension shaken with Amberlite IRA 401 ion-exchange
resin [6 g, preactivated and washed with 1 M aq. sodium
hydroxide (1 dm3), distilled water (1 dm3) and acetone (1 dm3)]
for 4 h. The mixture was filtered and the filtrate evaporated
to give N-methyl-3-oxidopyridinium betaine 31 as a solid. This
was stored in vacuo over phosphorus pentaoxide and used as
soon as possible after preparation.
Triethylamine (0.18 g, 1.73 mmol) and the betaine 31 (0.29 g,
2.6 mmol) were dissolved in a mixture of dry acetonitrile
(10 cm3) and diethyl ether (10 cm3) and the solution was added
dropwise to a solution of the dichlorophosphate 24 (1.19 g,
1.78 mmol) in acetonitrile (10 cm3) at room temperature, pro-
ducing an immediate precipitate. The reaction mixture was
heated to 40 ЊC in a sealed vessel for 16 h before addition of
water (5 ml) and triethylamine (0.18 g, 1.73 mmol) and re-
heating the mixture to 40 ЊC for a further 2 h. The solvents were
removed under reduced pressure and the residue azeotroped
with benzene, before removal of the solvent and drying the
residue in vacuo to give crude product (1.8 g). This was chrom-
atographed through silica and eluted with chloroform–
methanol–water (65:25:4). The initially enriched fractions
containing the product were rechromatographed, using
chloroform–methanol–0.88 ammonia (690:270:64) as eluent,
to yield the title compound (242 mg, 19%) δH (CD3OD–CDCl3)
0.88 (6 H, t, J 7 Hz), 0.94 (3 H, s), 1.25 (52 H, m), 1.49 (4 H, m),
3.28 (4 H, s), 3.34 (4 H, t, J 7 Hz), 3.86 ( 2 H, d J 3 Hz), 4.41
(3 H, s), 7.80 (1 H, m), 8.20 (1 H, m), 8.27 (1 H, m), 8.87 (1 H,
s); δC (CD3OD–CDCl3) 14.0, 16.99, 22.58, 26.08, 29.27–29.6,
31.85, 40.7, 48.63, 69.51, 71.57, 72.76, 127.94, 135.75, 137.96,
154.11 (Found: C, 67.9; H, 11.1; N, 2.0; P, 3.9. C43H82NO6Pؒ
H2O requires C, 68.1; H, 11.2; N, 1.85; P, 4.0%).
3-(Trimethylammonio)phenyl phosphate analogue 29
3-(Dimethylamino)phenol was purified by recrystallisation
from boiling water several times before use; activated charcoal
was used to decolourise the material.
A solution of the phenol (2 g, 14.6 mmol) in dichloro-
methane (10 cm3) was added to a stirred solution of phosphoryl
trichloride (2.24 g, 14.6 mmol) in dichloromethane (20 cm3)
over a period of 30 min and the mixture stirred under nitrogen
for 16 h. A further portion of phosphoryl trichloride was then
added (2.24 g, 14.6 mmol) and stirring was continued for a
further 2 h before removal of the solvent and excess of the
reagent under reduced pressure. The residue was dried at a
high vacuum pump before triturating with light petroleum and
drying (under reduced pressure) to produce 3-(dimethylamino)-
phenyl dichlorophosphate as the hydrochloride salt (2.37 g,
56%), δH (CDCl3) 2.9 (6 H, s, Me2N), 6.2, 6.35 and 7.1 (4 H,
ArH). This material was unstable to storage in air and so was
used immediately after preparation.
To a solution of the ether alcohol 14 (2.00 g, 3.5 mmol)
in dichloromethane (10 cm3) containing triethylamine (0.71 g,
7.0 mmol) was added 3-(dimethylamino)phenyl dichloro-
phosphate (3.55 g, 12.2 mmol), described above. The reaction
mixture was heated under nitrogen at 50 ЊC for 20 h. Water
(3.78 g, 210 mmol) and triethylamine (0.71 g, 7 mmol) were
added and the mixture reheated to 50 ЊC for a further 5 h. The
organic layer was collected, the solvent removed under reduced
pressure, and the residue azeotroped with benzene to give a
gelatinous product. The solvent was removed, the residue was
dissolved in chloroform (100 cm3), and the solution washed
with water (2 × 10 cm3); emulsions formed during this step and
were left to settle for 16 h. The organic layer was dried, the
solvent removed, and the residue again azeotroped with benzene
J. Chem. Soc., Perkin Trans. 1, 2000, 645–652
651

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5 H. Hauser, in Proceedings of the International Symposium, Water
Related to Foods, ed. R. B. Duckworth, 1975, pp. 37–71.
6 K. Cooper and M. J. Parry, Annu. Rep. Med. Chem., 1989, 24, 81;
Phospholipids, ed. I. Hanin and G. Pepeu, Plenum Press, New York,
1990; cf. P. Klotz, L. A. Cuccia, N. Mohamed, G. Just and R. B.
Lennox, J. Chem. Soc., Chem. Commun., 1994, 2043.
7 F. M. Menger, X. Y. Chen, S. Brocchini, H. P. Hopkins and
D. Hamilton, J. Am. Chem. Soc., 1993, 115, 6600.
8 F. Ramirez, H. Okazaki and J. F. Marecek, J. Org. Chem., 1978, 43,
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Acknowledgements
We thank Mr Trevor Boyd, Mr Terry Vick, and Dr Alistair
Taylor for coating the samples and Mr Ewan Campbell,
Dr Brenda Hall and Miss Monica Parmar for the fibrinogen
and activated platelet assays. Thanks are also expressed to
Mr Jim Bloxsidge, University of Surrey, for help with the high-
field NMR spectra.
9 N. T. Thuong, M. Chassignol, U. Asseline and P. Chalvier, Bull. Soc.
Chim. Fr., 1981, (part 2) 51.
10 P. E. Verkade and J. van der Lee, Recl. Trav. Chim. Pays-Bas, 1938,
57, 417.
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