Note: Descriptions are shown in the official language in which they were submitted.
7 1 ~
Derivati~res of g~ycerophosphQcho~ine and
g~ycerophosphoethanolamine, their preparation
a nd th ei r us e
Th~ present invention rel.ates to new triphenylL--
methy~ de~ivatives of sn-g~ycero-3-phosphocho~ine and sn-
g~ycer o-3 phosphoe~h~no~am; ne, processes ~or thei r pre-
para~ionO and their use for the synthesis of chemîea~y
de~ined, pure en~ntiomer~c 1~2-diacyl sn~ycero ;3-phospho-
eho~ine~ and 1~2 diacyl-sn-g-ycero-3 phosphoethano~amines
10 l~hieh are substituted in positions 1 and ;2 of the g-ysero-,.
~ndependent~y of one another, by different acy~ radicais.
In the current state of th~ artD che~ica~y d~fin~d
phosphatidy~cho~nes and phosphatidylethano~amin~s ~hich
carry diflFerent acy~ radisa~s in positions 1 and 2 of the
g~ycero~ are obtainab~e only via a mu~tistage tota- syn-
thesis or expensive semisynthetie processes~ uhich require
the use of enzymes and a ~arge excess of carboxy~1c acids~
The relevant pr~or art is described 1n two review articles
b~ H~ E1b~ 1n Chem. Phys. L~pids~ Vo~ume Z6 ~1980~ pages
Z0 405-429~ and in Zeitschritt fur Ange~andte Chemie CJournaL
of App~ied Chemistry~ , Volume 96 tl984), pages 2$7-262~
The industria~-sca~e synthesis of these phospha-
tidylcho~ines and phosphatidy~eth3nolamines, each of Jh;eh
ca rri es di f f erent substi tuents,. i s not f easib~e by means
o~ the processes known to date and the starting materia~s
and intermediates used there;n.
I~ uas therefore the object of the present inven-
tion to start from simple and eas~-y obtainabLe startin~
materia-s and provide new intermediates for a process ~hich
30 p~rmits pure enantiomeric 1,2-diacy~-sn-g~ycero-3-phospho~
cho~ i nes a nd 1 ,2-dia cy~-sn-g~yc ero-3-phosphoetha no~ami nes~
eaeh of ~hich carries different substituents and has a
d~lFined chemica~ structureO to b~ prepared in an eGonon~ca~
and time-saving manner on an industria- scal.ea This ~bject
35 ~das aehieYed by the present invention in an unexpected~y
s i mp~ e a nd ef f ect i ve ma nn~r .
The present invent;on aceording~y re~ates to lthe
.
L4t;;~
-- 2 --
1 new triphenylmethyl derivatives of sn-glycero-3-
phosphocholine and sn-glycero-3-phosphoethanolamine of the
formula
H2C - O - T
HO - C - H o~ R
H2C - O - P - O - CH2 - CH2 - N --R2
O R3
in which
T denotes an unsubstituted or substituted triphenylmethyl
group and the radicals
Rl, R2 and R3 either are identical and each represent a
methyl group or are different, in which case two of the
radicals Rl, R2 and R3 always denote hydrogen and the
third radical represents an unsubstituted or substituted
triphenylmethyl group. T preferably denotes a
triphenylmethyl group which is unsubstituted or
monosubstituted or polysubstituted by Cl to C6-alkyl, C
to C6-alkoxy or halogen.
The compounds of the formula I in which Rl, R2
and R3 each denote methyl are derivatives of
glycerophosphocholine, while the compounds of the formula
I in which two of the radicals Rl, R2 and R3 denote
hydrogen and the third represents a substituted or
unsubstituted triphenylmethyl group constitute derivatives
of glycerophosphoethanolamine.
In the general formula I, the radical denoted by
T in the compounds according to the invention, and one of
the radicals Rl, R2 and R3, where the other two radicals
denote hydrogen~ are each preferably an unsubstituted
.~
- 2a -
l triphenylmethyl group. However, the general formula I
also embraces those compounds in which the l-0-triphenyl-
methyl group, and/or any N-triphenylmethyl group which may
be present in the molecule, independently of one another
are monosubstituted or polysubstituted by alkyl radicals,
such as methyl, ethyl, propyl and the like, alkoxy
radicals, such as methoxy, ethoxy, propoxy and the like,i
or halogen, such as fluorine, chlorine or bromine. Among
; the compounds of the formula I containing substituted
triphenylmethyl groups, preferred compounds are in turn
those in which one or more phenyl radicals of the
triphenylmethyl groups are substituted in the para-
position by alkoxy radicals having l to 6 carbons atoms.
In the compounds of the formula I, the l-0-
triphenylmethyl group and any N-triphenylmethyl group
which may be present
~LZr~
-- 3 ~
1 in the molecule, are protecting groups which are
eliminated during the preparation of l,2-diacyl-sn-
glycero-3-phosphocholines and 1,2-diacyl-sn-glycero-3-
phosphoethanolamines. Therefore, the meaning and the
number of the substituents on the triphenylmethyl groups
in the compounds of formula I are of minor interest, as
long as the triphenylmethyl protecting groups can be
easily removed by the action of acids.
The nomenclature and designation of positions
used in the present description for glycerophosphocholine,
glycerophosphoethanolamine and their derivatives follow
the rules stated in Biochem. J. 1~1, 29-35 (1978). The
abbreviation sn in the systematic chemical names of the
compounds mentioned denotes stereospecifically
numbered . All designations oE positions in the present
description which relate to the position of the
substituents on the glycerol radical are based on this
stereospecific numbering.
The compounds of the general formula I are
prepared by a method in which sn-glycero-3-phosphocholine
or sn-glycero-3-phospho-(N-triphenylmethyl)-ethanolamine
of the general formula
H2C - OH II
HO - C - H 1- /Rl
H2C - O - I - O - CH2 - CH2 - N - R2
R3
in which
Rl, R2 and R3 have the meanings given above, their salts
.~
,~
46;~
1 with inorganic or organic acids or bases, or their
complexes with metal salts are tritylated at the oxygen in
position l of the glycerol by reaction with a reactive
triphenylmethyl derivative of the general formula
T-X III
in which
T is defined as in formula I and
x denotes a reactive leaving group, such as chlorine,
bromine or iodine,
in an inert organic solvent or solvent mixture at
.~ "~. .
~ _ f
temperatures from room temperature to the boi~ing po1n~
of the solvent or o~ the lowest-boilin~ solvent componen~ -
of the solvent m;xtureO
~he tritylat1ng agent used in the process accord-
5 ing to ~he invention ;s a reactive triphenylmethyl deriva-
tive~ preferabLy tripheny~meehy~ chloride or tripheny~-
~ethy~ bromide, particu~ar~y pre~erab-y ~ripheny~methy~
chloride. For the preparation o~ the 1-0-tripheny~methy~
derivati~e of the general formula X~ one mole e~uiva~nt
o~ the trity~ating agent is consumed. ALthough advan-
tageous in many cases for accelerating the eourse o~ the
reaction and for comp~eting the reaction, it is not
absoluteLy necessary ~o emp~oy an excess o~ the trity~atin~
agent. The amount of tritylatin~ agent emp~oyed can be
varied ~ith;n wide limits~ as desired. The us~ of a smal~
excess to a several-~o~d molar excess of tritylat1n~ agent
per ~o~e of the start;ng material of the formula II has
proved part~cular~y useful, a 1.5-foLd to 3-~old ~o~ar
excess being particularly preferred.
The reac~ion of the compound of the formula ~I
~i~h that o~ the formula II~ is advantageousLy carried out
in the presence af an excess of a suita~le proton acceptorlO
Suitable proton acceptors are inorganic or organie bases.
The reaction is particularLy preferabLy carried out in the
presence of organic bases, such as tertiary a~ines, for
example trimethylam;ne, triethylamine, N-methyLpiperidine~
N,~J dimethylaniline or N,N-diethy~aniline, Hunig' base or
heterocycLic bases, for example pyridine, 4-N,N-dimethy~
am;nopyr;dine9 4-pyrrolid;nopyridine, picolines, co~
dine, quinol;ne, ;soquinoLine and the ~ike.
The react;on is carried out in organic so~vents or
solvent mixtures wh;ch are inert to the particuLar rea~
tantsr Particularly suitab~e so~vents are aprotic, po~ar
solvents. These preferably include dimethyl sulfoxide,
carboxamides~ such as dimethylformamide, dimethy~aceta-
mide, hexamethylphosphQrotriamide or N-methy~pyrro~idone,
nitriles, such as acetonitrile or propionitri-e~ hetero-
eyciic bases, such as pyridine~ quino~ine or pico~ines,
Ot mixtures of such so~vents. In many cases, it may be
advantageous to carry out the reaction in a solvent mix-
ture consisting ~of an inert aprotic polar solvent and an
~nert apro~ic solvent o~ Lo~ polarity. xamples of so~-
vent components of ~o~ polarity ~hich are suitable for
S such so~vent mixtures are a~iphatic or aromatic hydrocar-
bons, such as Low-boil;ng or high-boiling petro~eum ether,
h~xane, heptane, benzene, toLuene and tl1e ~ike, ha~o~enated
a~ipha~ic or aromatic hydroçarbons~ such as methy~ene
chlorid~, ch~oroform, 1,2~dichloroeth3neO carbon tetra-
- 10 ch~or;de, eh~orobenzene, p-ch~orotoluene and the ~ike.
The reaction temperatures can be varied ~Sthin a
~;de range. ~n genera~, the reaction is carried out at
temperatures bet~een room temperature and ~h~ boi~;ng
point sf the part~cu~ar so~vent. emp~oyed or of the lo~est-
boi~ing so~vent eomponent of.the so~vent ~ixture used,prefer~b~y at temperatures between 20C and 200~C~ in par-
t~cu~ar at temperatures between 20 and 80co
To prepare the compounds of the genera- for~ula I,
~t is possil~Le to start from the comp,ounds of the ~eneraL
Z0 formu~a ~I as such, from the1r ~a~ts ~ith inorganic or
organ~c acids or bases, or from their complexes uith metal
sa~ts. Particularly in the preparation of 1-~-trity~-sn-
~ycero-3-phosphocholine, it may be advantageous in many
ca-~es to em~loy the more stable meta~ salt complexes h~v-
ing a ~on~er shelf ~ife~ for exampLe the cadmium ch~orideadduct of sn-g~ycero-3-phosphocholine.
Working up is advantageously carried out by
evaporating do~n the reaction so-ution or precipitating
the products from the reaction so~ution by di~ution ~ith.
so~vents in ~hich the compounds of the genera~ formu~a ~
are poor~y saluble. The crude products obtained by son-
vent~ona~ chemical work~ng-up methods are very suitable,
~ithout further purification operations, as intermediate
compounds for the preparation of phosphat;dylcho~ines and
phosphatidy~ethano~amines, each of ~hich carries different
substituents. Part;cu~arly suitable ~ethods for purifying
the compounds of the general formula I are the conventiona~
chromatographic methods~ such as preparative thin-~ayer
chromatography, column chromatography, adsorption
t.~
., ` !~
- 6 -
chromatographyO medium pressure liquid chromatography
or high pres~ure lîqu;d chromatography.
The invention furthermore re~ates to the use of
the new compounds of the general formula I for the prepara-
t;on of pure en~nt;omer;c phosphat;dy~chol;nes and phos-
phatidy~e~hano~amines ~h;ch carry dif~erent acy~.rad~ca~s
~n positions 1 and 2 o~ the glycerol, independently o~ one
another~
~he compounds of ~h~ f~xmula 1 ca~ preerably be
~sed in a method ~or preparing 1,2-diacyl-sn-glycero-3~
phosphocho1ines and 1,2-diacy1-sn-g~ycero~3~phosphoethanol-
ami~es o~ the formula
.
0 ~ C -~ - C - R5 IY
~t
R~ C - O CH o ~ ~, R l
H2C - 0-P~0 - CH2-cH2-N - R2'
i~ ~hich
R~', R2~ and R3' are identical and in each case denote either
3 ~ydrogen atoms or 3 methyl groups, and
and R5 are ~i~Pere~t and independently o one another de-
~ote a stxaight chai~ or branched C1 to C24- alkyl ra~ical~
a straight chain or branched Cl to C24-al~yl radical substi--
20 tuted by one or more halogen atoms or alkoxy g:roups, a straig~t
chain or bra~ched monourlsaturated or pol~nsaturated C3 to
C24-alkenyl radical or a straight chain or branched mono~--
saturated or polyunsaturated C3 to C24-al}cenyl radical sub--
stituted by one or more halogen atoms or alkoxy groups" ~rhich
method comprises
a) acylating a compound o~ the ~ormula I by reaction with an
acylatlng derivative o a carboxylic acid o~ the o~mula
~ COOH to give a compound o~ the formula
~;~514~ 1
o H2C-O~l'
Il
R4 ~ C ~ O - C - H O_ V
H2C - O - 1l - O - CH~ ~H2 - ~Y ~ R2
O ~3
~he rei n
T represents a~ unsubstituted or su~s~ ed triphenylme~
group and
5 Rl, R2 a~d R3 either are identical a~d each represent a met:hy
group or are diferer~, i~ ~rhich case ~ro o~ the radicals Rl~
R2 and R3 always denote hydroge~ and t~e third radieal repre-
sents an unsubsti~uted or substituted triphenylmethyl group,
b) eliminati~g the 1-0-triphe~ylmethyl group from the resulting
compou~d of the formula V by the action o acids9 with the
proviso that, i~ the ca.se o~ compound~ o~ ormula V in ~hich
one o~ the radicals Rl ~2 and R3 is a triphenylmethyl group,
~he acid in a Lewls acid, with ~ormulation o~ a 2-acyl-sn-
glycero-3-phosphocholine or a 2-acyl-sn-glycero-3-phospho-
(N~triphenylmethyl)-ethanolamine o~ the ~ormula
O H2C-OH
R4-C- O~ C- H ~ ~ ~ R
W2C- ~ -CH2_ CH2_ N -R
0 3
in ~hich
:æl, R~, R3 and R4 have the meaning given abo~e,
c:) a~rlating the compound of the fo~nula VI by react;on ~nth
an acylating derivative o~ a car~oxylic acid o~ the formula
R5-COOH to give a 1,2-diacyl-sn-glycero-3-phosphocholine
or 1, 2-diacyl-sn-glycero--3-phospho- (N-triphen~rlmet~yl )
ethanolamine o~ the ormula
5~4t;;~ ~
.. -- S --
o
o lH2C~Q-C_R5
R4 - C - V - ~ H c~
HzC - 9 - P - O - CH2 C:H;2 - N R2
O R3
tn ~h;ch
R1 R2 and R3 a:re defined as in fo:rmula ~r, and R~l a~d R5
have ~he meani3lg given in ~ormula IV" a:nd
5 d) elimirLati~g any N-triphe~ylmethyl group prese~t irl the
compou~d oE the ~ormula VII by the actio~n o~ a~ acid i~ a~L
ap~otic ~olventO
In the defin~t;or,s given in the pr~sent descripo
t~on for R4 and Rs, a "straight-chain or branched
aLkyL radical" ~s understood as meaning a saturated a~i-
phatic hydrocarbon radicaL ~hich contains 1 to 24 c~rbon
at~ms and can be branched as often as desired. The
- expression "straight-chaln or branched, monounsaturated or
poLyunsaturated aLkeny~ radicaL" ~epresents an unsatura~ed
aliphatic hydroca~bon rad;caL which has 3 to 24 carbon
ato~s and one or more olefinic doub~e bonds and can ~1keo
~ise be branched as often as desired. Both ln the a~ky~
radicaLs and in the a~keny~ radicaLs, one or more hydrogen
atoms can be replaced ~ith halogen, such as fLuor;ne,
chlorineD bromine or i~dine, or with a~ko~y radicaLs~
such as methoxy, ethoxy, propoxy, isopropoxy~ butoxy and
the l~keO
The acyLation of co~pounds of the generaL form~la
I to give the 2-acyL derivatives of the generaL for~u~a
is carried out using reactive derlYatives of carboxy~ic
acids of the general formuLa R~cooH~ ExampLes of suitable
acyLat;ng agents of this type are ha~ides, anhydridesO
acti~e ~sters and azolides of these carboxy~ic acids, car-
boxy~ic acid imidazo~ides being part1cu~arly preferred.
Th~ reaction is carried out under the conventional
~s~
_ 9 _
acylat~on conditions~ for example in anhydrous~ poLar apro-
tic soLvents or solvent mixtures ~hich are ;nert to the
partisu~ar reactants~ These preferably include eehers,
such as tetrahydrofuran, dioxane or ethy~ene gLyco~ dime-
thy~ ether, haLogenated aliphatic or aromat;c hydrocarbons,such as me~h~ene ch~or;de, ch~oroform~ 1,2-dich~oroethane,
ch~orobenzene or p-ch~oroto~uene~ carboxamides, such as
d;~ethylfor~amide or dimethylacetamide, hexamethyLphosph~ro-
triamide, N-methy~pyrroL;done, dimethy~ su~foxide,
heterocycLic bases which at the same time act as protQn
acc~ptors, such as pyridine~ aikylated pyridines9 N~N-
dia~kylaminopyridines and the ~ike, or mixtures of th~se
so~vents. When carboxyLic acid i~;dazo~ides are used as
~cy~ating agents, mixtures of di~ethyl sulfox;de ~nd
tetrahydrofuran in a ratio of t:4 to 4:t have proved par
ticu~ar-y usefu~. The acyiation is advantag~ous~y carried
out us~ng an acid acceptor ~hich simultaneous~y acts as a
catalyst and i5 of inorganic nature, such as so~ium
hydrideO metalLic sodium and the like, or of organic nature,
2~ such as triethyla~;ne~ N-meth~lp;peridine, N,N-dimethyL-
aniLine or N,N diethyLani~;ne, or in the presence o~
heterocyc~ic bases, such as pyr;dine, N,N-dimethyLamino-
prrid;ne, pico~ines, coL~;d;ne, quinoLine, isoquino~ine
and the ~ike. The acyLation reaction ean be carr;ed out
2S at temperatures bet~een 0C and the boiling point o~ the
so~vent, but is preferably effected at temperatures bet~een
2~C and 30C.
The unconverted carboxyiic ac;ds obtained in th~
reaction9 and any unreacted residues o~ acylating agents
3~ stiLL present, must be separated off be~ore the subsequent
reaction-step, for example by chromatography over siLica
ge~ or by other purification methods conventiona~y used
in preparative chemistry~ such as by extrac~ion of a
soLut;on of the crude product in ~ater-;mmiscib~e so~vents
~ith dilu~e aqueous bases, for example aqueous ammonia~
or by recrystal~;zation or reprecipitation o~ the crude
product f rom suitabLe solvents.
The 1-0-tripheny~methyL group is then eL;minated
from the 2-acyl derivat;ves o~ the generaL formula V by
5~4~2
-- 10 --
1 treatment with an acid. In eliminating the l-0-triphenyl-
methyl group, the use of a Lewis acid is necessary in the
case of the glycerophosphoethanolamine derivatives of the
general formula V in which one of the radicals Rl, R2 and
R3 denotes a triphenylmethyl radical in order to retain
the N-triphenylmethyl group, and is advantageous in the
case of the glycerophosphocholine derivatives of the
formula V in which the radicals Rl, R2 and R3 each denote
methyl. Elimination of the l-0-teiphenylmethyl group in
the case of the glycerophosphocholine derivatives can,
however, also be effected with inorganic acids, for
example mineral acids, such as hydrochloric acid,
hydrobromic acid, per-chloric acid and the like, or with
organic acids, such as trifluoroacetic acid or
trichloroacetic acid and the like. To prevent migration
of an acyl group, it is advantageous to maintain
temperatures below 20C. The boron trifluoride/methanol
complex in methylene chloride is employed at 0C, this
being a particularly effective detritylation agent. The
compounds of the general formula VI are obtained in this
procedure in such good purity that they do not need to be
subjected to any further purification operations before
further processing. The embodiment of the synthesis in
which the compounds of the general formula VI which are
obtained after elimination of the l-0-triphenylmethyl
group are fed directly for further acylation is
particularly advantageous since any acyl migrations which
may occur during the purification operations are
decisively reduced at this stage of the synthesis.
Further acylation of compounds of the general
formula VI to give the 1,2-diacyl derivatives of the
general formula IVa is again carried out using one of the
abovementioned reactive carboxylic acid derivatives of the
~-
`"` ~L~S~6;~
- 10a -
1 acids of the general formula R5COOH. To introduce the
acyl radical into position 1 of the glycerol, carboxylic
anhydrides have proved particularly useful in this case.
The reaction is carried out under the conventional
acylation conditions, advantageously in anhydrous, polar
aprotic solvents or solvent mixtures, suitable solvents being not
only the solvents stated above for the acylation of the
,
~Z51~6f~
-- 11 --
1 2-position but also solvent mixtures containing halogenated
hydrocarbons, such as methylene chloride, chloroform, carbon
tetrachloride, cholorobenzene and the like. Mixtures which
have proved particularly useful are mixtures of methylene
chloride and 4-N,N-dimethylaminopyridine, which at the same
time acts as an acid acceptor. The temperature range from
room temperature to the boiling point of the solvent or of
the lowest-boiling solvent component can be chosen for the
reaction. Preferably~ the reaction is carried out at room
temperature or slightly elevated temperatures of 20 to 40C.
The diacyl derivatives of the general formula VII
which are formed can be obtained in pure form by means of
the purification operations usually employed in preparative
chemistry. Thus, oc example, the crude products can be
dissolved in water-immiscible solvents and freed from excess
acylating agents by extraction with dilute aqueous bases,
for example with aqueous ammonia, or can be purified by
recrystallization or reprecipitation.
The conventional chromatographic methods or
multiplicative partition between two phases are also
suitable for the purification of the compounds of the
formula VII.
~he 1,2-diacyl-sn-glycero-3-phosphoethanolamines
of the formula IV, wherein Rl~,R2' and R3' each denote
hydrogen, can be obtained from compounds of the formula VII
in which one of the radicals Rl~ R2 or R3 denotes a
triphenylmethyl group, by elimination of the N-
triphenylmethyl radical. The elimination reaction can be
carried out using inorgainic acids, for example mineralacids, such as hydrochloric acid or hydrobromic acid, or
preferably organic acids, for example halogenated carboxylic
acid, such as trichloro or trifluoroacetic acid, the
..~
~'~S~
- lla -
1 reaction being carried out in aprotic solvents in order to
avoid deacylation in position 1 and 2 of the glycerol.
Preferably, the N-triphenylmethyl radical is removed using
trifluoroacetic acid in methylene chloride at temperatures
of about 0C.
~ he compounds described here are characterized,
and their purity is checked, by means of thin-Layer
s~
- 12 -
1 chromatography and spectroscopic methods, in particular
lH_NMR spectroscopy. The spectroscopic data determined are
in agreement with the chemical structures described for the
compounds,
The compounds of the general formula I are new,
central intermediates for the universal chemical synthesis
of 1,2-diacyl-sn-glycero-3-phosphocholines and 1,2-diacyl-
sn-glycero-3-phosphoethanolamines of the general formula IV,
each of which carries different substituents. when
intermediates of the general formula 1 are employed, it is
no longer necessary to use enzymes, which previously had to
be employed in the semi-synthetic preparation of such
phosphatidylcholines and phosphatidylethanolamines, each of
which carries different substituents; hence, the compounds
o~ the general formula IV are also easily obtainable on an
industrial scale. Other advantages of this invention are
that the carboxylic acid derivatives used for the acylation
in position 1 or in position 2 are virtually completely
utilized. In the enzymatic methods used to date, the
positions 1 and 2 in the glycerophosphocholine or in N-
protected glycerophosphoethanolamines are first substitutedby two identical acyl radicals, after which the acyl radical
in position 2 is eliminated by the action of phospholipase
A2, and the desired acyl radical is introduced into this
intermediate by means of a further acylation reaction, In
this procedure, the carboxylic acid used for the first
acylation in position 2 is lost. Furthermore, the rate and
efficiency of the enzymatic hydrolysis depend on the type of
carboxylic acid, whereas the preparation, according to the
invention, of the compounds of the formula IV is completely
independent of this.
~ ,-.
- 12a -
l The phosphatidylcholines and phosphatidylethanol-
amines of the general ~ormula IV are, for example, valuable
emulsifiers in the preparation of medicaments and crop
protection agents or can be used in the photographic
industry. These compounds can also be used as chemically
defined starting materials for the preparation of liposomes,
liposome solutions and liposome gels.
''i
13
sn-~lycero-3-phosphocho~;ne or sn-g~y~e~o-3-
phospho-tN-tripheny~m2thy~ thanolamine is used as a
start;ng material for the preparation of the compounds of
the gen~raL formuLa ~. sn-Glycero 3~phosphochoL;ne can
; 5 b~ prepared in a simp~e m3nner in virtuaL~y ~n~i~ited
amounts by con~entiona~ ~ethods, ~or exampLe by alkal;ne
hydro~ysis of naturaL ~ecithin~ Preferab~y, co~merciaL~y
- avai~ab~e products are used.
~n-6iycero-3-phospho-(N-tr;pheny~methyl,~; ethano~-
10 a~ne ~s readi~y obtainab~e by N-Srity~at;on of sn-sLycero-
3-phosphoethano~amine from any source. sn-6~ycero-3-
phospho-CN-tripheny~ethy~)-ethano~amine is prepared in
a particu~ar~y advantageous and simp~e manner by trity~a-
ti on of phospha t i dyl e~ha no~a m; ne i n a mi xture of phospho-
15 lipids of vegetab~e, animal. or microbia~ origin }Jith areae~ive tripheny~methy~ derivativ~, preferab~y triphenyL
~ethy~ bromide, fo~-o~ed by hydroLysis of the tota~
~ipids. The sn-3~ycero-3-phospho-~N-triphenyL-methy~)-
qthanoLa~ine can be separated from water-soLuble c~ea~a0e
products by extracting lt from this mixture with water-
~m~iscib~e so~ventsO pre~erably ~ith ch~oroform in the
presence of methanoL, and can then be obtained in pure
1;or~ by ~ashing out the fatty acids ~ith an a~ka~ine
aqueous phase, if appropriate mixed with methanoL. sn
61ycero-3-phospho-(N-tripheny~methyL)-ethano~a-~ine can be
prepared, ~or exampLe, by the fo~o~ing procedure:
1~5 9 o~ soya phosphoLipids and 1.4 g of tripheny~-
~ethy~ bromide are dissoLved in 50 ~L of chLoro~orm~
A~ter the add;tion of 1.1 9 of triethyLamine, the reaction
30 Ini%t~re is stirred for 12 hours at 20C,, 50 m~ of chLoro-
form and lU0 mL of a 0.5 N sodium hrdroxide soLution in
methano~ are then added, and stirring is con~;nued for
30 ~inutes at 20C. 100 m~ of ch~oroform are then added,
and ~he organic phase is separa~ed off and washed three
times with an a~kaLine aqueous phase. Fina~lyO the or~anic
soLYerlt is stripped of ~ in ~acuo. The residue, ~Ihich cun-
ta1ns the sn-g~ycero-3-phospho-~N tripheny~methy~)-
ethanoLamine can be used d;rectly for the subsequent 0-
trit~ation. The pure compound~ which is obtained by
S~ 6~
- 14 -
1 ch~omatographing the crude product over silica gel with a
chloroform/methanol gradient, shows a single spot with Rf
0.3 in thin-layer chromatography over silica gel (mobile
phase: 50:25:6 (v/v/v) chloroform-methanol-25% NH3 mixture).
The examples below serve to illustrate the
invention in more detail without restricting it.
a) Preparation of compounds of the general formula I
Example 1:
l-0-Triphenylmethyl-sn-glycero-3-phosphocholine
5 g of sn-glycero-3-phosphocholine/CdC12 complex and 4.6 g
of triphenylmethyl chloride are dissolved in anhydrous
dimethylformamide (50 ml) at 70C. After the addition of
2,3 ml of triethylamine, the mixture is stirred for 30
minutes at 70C in the absence oE moisture. When the
reaction mixture has cooled, S g oP powdered NaHC03 are
added, and the mixture is stirred for 20 minutes at room
temperature. The reaction solution is then filtered, and
300 ml of diethyl ether are added. The oil formed is
separated off by centrifuging, washed once with diethyl
ether and finally dissolved in 150 ml of methanol. After
the addition of 300 ml of CHC13, washing is carried out
using 90 ml of a solvent mixture tuPPer phase) consisting of
3:48:47 (v/v/v/) CHC13/CH30H/H20, and the lower phase is
diluted with lS0 ml of 2:1 (v/v) CHC13/CH30H. 3 ml of 25
strength agueous NH3 solution are then added, and the
mixture is left to stand for 15 minutes at room temperature
and finally separated off from the resulting colorless
precipitate by centrifuging. When the solvent has been
stripped off in vacuo, the oily residue is digested with
three times 50 ml of diethyl ether, This procedure gives a
yellowish solid crude product (5.5g) which, when subjected
to thin-layer chromatography over silica gel (mobile phase:
65:35:5 (v/v/v) CHC13/CH30H/25~ NH3 mixture), shows an Rf
:~2~ 6;~
,
- 14a -
1 value of 0.15 and is found to contain only very small
amounts of impurities. The pure compound is obtained by
medium pressure chromatography over silica gel. A
CHC13/CH30H gradient is used for the elution, the eluant
containing 0,5~ by volume of 25% strength aqueous NH3
solution, When
~ 15 -
1 subjected to thin-layer chromatography over silica gel
(mobile phase as for the analysis of the crude product), the
pure substance exhibits a single spot at Rf 0.15.
lH_NMR spectrum: ~ ppm
3.1 (N-CH3); 3.3-4.3 (choline and glycerol); 7,25
(triphenylmethyl C-H).
Example 2:
l-0-Triphenylmethyl-sn-glycero-3-phospho-(N-triphenyl-
methyl)-ethanolamine
946 mg of sn-glycero-3-phospho-(N-triphenylmethyl)-
ethanolamine and l.735 g of triphenylmethyl chloride in 32ml
of anhydrous pyridine are stirred for 48 hours at room
temperature in the absence of moisture. The mixture is then
poured onto ice water, The product is extracted with three
portions of diethyl ether. The combined ether phases are
washed twice with water and then dried over Na2S04. When
the solvent has been stripped off in vacuo, the remaining
pyridine is removed by evaporating the residue in the
presence of toluene and in vacuo. The residue is taken up in
30 ml of a l:l (v/v) chloroform/methanol mixture, and the
solution is left to stand overnight at 4C. After the
precipitated triphenylcarbinol has been filtered off and the
solvent mixture then stripped off, 1.7 g of crude product
are obtained, which is used directly for the subsequent
~5 acylation. Chromatography over silica gel with a
chloroform/methanol gradient gives the pure compound which
shows a single spot at R~ 0.6 when subjected to thin layer
chromatography over silica gel (mobile phase: 6:4 (v/v)
chloroform/methanol mixture).
lH_NMR spectrum : ~ ppm (DMS0-D6)
3.5-4.3 (glycerol and ethanolamine C-H, diffuse); 7.25
(triphenylmethyl C-H, m).
Example 3:
~J
. ~
S~l46~
- 15a -
1 ~sing the method stated in Example 1, the
following compound was obtained:
1-0 (4,4'-dimethoxytriphenylmethyl)-sn-glycero-3-phospho-
choline
5 The thin-layer chromatogram over silica gel ~mobile phase:
65:35:5 (v/v/v) CHC13/CH30H/25% strength aqueous
NH3 mi xture~
shous a s;ngle spot, R~ vaLue 0.15.
1H-NMR spectrum~ ~ ppm CCD30D)
3.1 tN-tll3~; 3.8 ~pheny~-0-SH3~; 3.3-4.,3 gcho~ine and
5 g~yceroL~; 7023 ~aromatic3.
Exa~np~ e 4O
1-0-~4~4' I)imethoxytripheny~methy~-sn-g~ycero-3-phospho-
~iN-tripheny~methy~-ethano~a~ne
A sol~lon of 2.4 g of sn-g~ycero-3-phospho-~N ~
10 tripheny~methy~3 ethano~am; ne and 4..5 9 of 4,4'-dimethoxy-
~ripheny~methy~ ch~oride in 80 m~ of anhydrous pyridine
~s s~i rred for 72 hours at room temperature. Af'ter the
pyr~dtne has been stripped off in vacuo, the residue is
disso~ved in 200 Inl. of Z:l CV/V3 C)lÇ~3tCH30H. This so~u--
15 t;on is ~ashed ~ith ttice 40 n~L of 3:48:47 (v~Y~v) CHel3/
CH30HtHzO, and the soLvent is then disti~cd of ~ ~in vacuo.
This procedure gives a brown oi~ which, when digested
~ith 80 m~ and 30 m~ portions of diethy- ether, g~ves
2.5 9 of Sh~ crude prnduct as a color~ess amorphous pu~der.
Z0 Th~n-~ayer chromatography over si~ica ge~ Cmob;Le phase:
~:4 Cvlv) CHCl3/CH30H mixture) shows that the crude product
conta i ns more tha n 90X of the desi red produc~ tRf 0.,6),
~hich can be used, ~ithout further purificationO for the
preparation of the pure enantiomeric 1,2-diacy~-sn-g~ycero-
Z5 3-phosphoethano~amines~ each of ~hich carries differenC
substi tuents ,,
The pure compound is obtained by m~dium pressure
chromatography over si~;ca ge( with a CHC~3/CH30H grad~ent
~n the presence of 0.5 X by vo~ume o~ aqueous NH3) and sho~s
a sing~e spot at R~ 0.6 ~n the thin-~ayer chroma~ogra~
~conditisns as in Examp~e 3).
lHUNMR spectrumu ~ ppm
C2:1 ~v~v) CDcl3/cD3oD)
3~1 CN CH3); 3.8 tphenyl-0-CH3); 3.3-4.3 Scholine and
giycero~); 7~23 ~aromatic9.
b) Use of the compounds of the genera~ formu~a I, ob~ined
acc~rding to the above examp~es, for the preparation of
compound~ of the general for0u~a IY
:1;2S~6
17
Example S~
A) 1-0-Tripheny~methy~-2-oleoyl-sn~g~ycero-3-phosph~-
choline
1~Z~ g of oleic acid are allo~ed to react ~ith
794 mg of carbony~diimidazole in 25 m~ of tetrahydrofuran
f :7r 45 winutes at room temperature, after which the solven~
is removed in Yacuo. A soLution of 1.1 9 of the crude
produGt obtained from 1-0-tr;pheny--methy~-sn-g~ycero-3-
phospho~ho~ine taceording to Example 1) in 28 m~ of
di~ethy~ sulfoxide ;s then adde~ to the res;due, ~hich
contains the tatty acid im;dazoiide formed. After the
addi.tion of a cataLyst, ~hich is prepared by disso~ving
148 mg of metal~ic sodiu~ in 11.5 ~1 of dimethy~ su~-
~oxide, the reaction mixture is allo~ed to stand for
20 ~inutes at Z0C~ ~ith occasionaL shaking. F~na~yg
~4 m~ of a 0.1 N aqueous acetic a~id are added a~l at
once. The ~ixture is extracted S~ice ~ith a 2:1 ~v~v)
ch~oroformtmethano~ mixture, and the co~;ned organic
phases are washed tu~c~ w1th a 1:1 ~v/v) methanol/H20
mixture. After the solvent ha5 been evaporated of~ in
vacuo~ the 6rude product is dissolved in chLoroform, and
the solution is introduced onto a s;~ica gel co~umn. The
pure compound ~1.1 g~ is eluted with a chloroform/
methanol ~radient, and thin-layer chromatography over
si~ica ge- tmobile phase: 65:35:5 ~v/v/v) ch~oroform/
~ethano~/25% NH3) gives a single spot ~R~ 0.4)~
Ba Z-O~eoyl-sn-glycero-3-phosphocho~ine
O.S ml of a 20% strength solution of boron tri-
fLuor~de in methanol is added to a solution o~ 500 mg o~
1-O-tr;phenylmethy~-2~oleoyl-sn-~lYcero-3-phosphocho~in~
in 30 ml of methyLene chloride, after ~hich the m;xture
is stirred for 30 minutes at 0C~ 15 ml of methano~ and
9 ~ of ~ater are then added, the ~ixture is shaken to
effect extraction, and the organ;c phase is iso~ated. The
solvent i5 removed by stripping it off in a high vacuum
at room temperatureD
C~ 1-Pa~mitoy~-2-oleoy~sn-glycero-3-pho~phocholine
The residue, ~hich contains the resu~ting Z-o~eoy~-
sn-glycero-3-phosphocholine, is d;ssolYed in 30 ml o~ dry
s~
chL~rofor~ 820 mg ~f palm;t1c anhydr;de and 200 m~ o~
dimethy'dminopyridine are then added to this so~ution~
and the reaction mixture is stirred for 6 hours at 205~
Thereafter, 15 ml of methanol are added, and the m;xture
is extracted by shaking, ~irst ~ith 0~1 N HC~ and then
with H20~ After the organ;c so~vent has been stripped off
;n vacuoO the substance is purif;ed by chromatography over
silica ge~9 usin~ a ch~oroformJmethano~ gradient. The
purified compound C400 mg~ gives a sing7~e spot tRf ~ 4)
~hen subjected to thin-layer chromatography oYer siLica
ge~ ~mobiLe phase: 65:35:5 (v/vJv~ ch~oro~orm/m~thano~/Z5X
strength NH3>~
Examp~e 6:
1-Stearoy~ o~eoy~-sn-g~ycero-3-phosphocho~ine
A so~ution ot 2-o~eoy--sn-g-ycero-3-phosphochoLine
~obtained as desc~ibed in E~ample 5, stages A and B~ is
reacted uith 1.2 9 of stear;c anhydride according to the
method stated in E~ample 5, stage C, and the product is
purified chromatographically and anaLyzed. 55Q mg t76X
yield, relative to 1-0-triphenylmethyl-sn-glycero-3-
phosphoGholine) of 1-stearoyL-Z-oleoyl-~n-glycero-3- -
phosphocholine are obta;ned, this product ~iving a sing~e
spot ~n the th;n-layer chromatogram over silica ge~ Cmobi~e
phase: 6S:35:5 Cv~v/v) chlorofocm/me~hano~/25Z strength
aqueous NH3 solution); R~ value 0.4.
Examp~e 7.
A) 1-0-Tripheny~methyL~2-lino~eoy--sn-g~ycero-3-phospho-
cho~ne
600 mg of 1~0-tripheny~methyL-sn-~lycero-3-phos-
phocholine taccording to Example 1) are acyLated according
to the method stated in ExampLe 5, stage A~ with linolecrl-
imidazo~ide, ~hich ~as prepared from ~90 mg of linoleic
acid and ~40 mg o~ carbonyldiim;dazole~ After ~edium
pressure chromatography over si~ica gel, the p~re compound
~690 mg~ y;eLd 75X of theory) sho~s a single spot tR~
va~ue 0~3) in the thin-~ayer chromatogram over 5i ~ica ge~
tmobile phase. 65~35:5 (v/v/v) ch~oroformJmethano~/25X
strength aqueous NH3 solution)O
~;~5~6;~
- 19 -
1 B) 2-Linoleoyl-sn-glycero-3-phosphocholine
In order to eliminate the l-0-triphenylmethyl
radical, a solution of 1 g of 1-0-triphenylmethyl-2-
linoleoyl-sn-glycero-3-phosphocholine in 60ml of methylene
chloride is treated with a 20~ strength solution of boron
trifluoride in methanol, as stated in Example 5, stage B.
C) l-Stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine
The 2-linoleoyl-sn-glycero-3-phosphocholine
obtained in stage B is reacted with 1.6 q of stearic
anhydride by the method stated in Example 5, stage C, and
the product is purified by chromatography. 820 mg (80
yield, relative to 1-~-triphenylmethyl-sn-glycero-3
phosphocholine) of pure product are obtained, which gives a
single spot in the thin-layer ch~omatogram over silica gel
(mobile phase: 65:35:5 (v/v/v) chloroform/methanol/25
strength aqueous NH3 solution); Rf value 0.4.
Example fl:
A) l-0-Triphenylmethyl-2-acetyl-sn-glycero-3-phosphocholine
320 mg of 1-0-triphenylmethyl-sn-glycero-3-
phosphocholine (according to Example 1) are reacted with 0.7
me of acetic anhydride in 4 ml of anhydrous pyridine
overnight at room temperature. The reaction mixture is then
poured onto ice water and extracted twice with
chloroform/methanol (2:1, v/v). The combined organic phases
are washed twice with chloroform/methanol/water (3:48:47
v/v/v), dried over sodium bicarbonate and finally brought to
dryness. The crude product is purified by medium pressure
chromatography over silica gel, using a chloroform/methanol
gradient. The pure product (180 mg, 51~ yield) gives a
single spot in the thin-layer chromatogram over silica gel
(mobile phase: 65 35:5 (v/v/v) chloroform/methanol/25%
strength aqueous NH3 solution); Rf 0.16.
`~'
"
S1~6;~
- l9a -
1 lH_NMR spectrum ~ ppm (CDC13-CD3+0D, 2:1 v/v)
2,13 (H3C-C0, S, 3H); 3.16 (H3C-N, s, 9H); 3.3-4.4 (glycerol
and choline-CH2); 5.23 (glycerol C-H, lH3; 7.33 (aromatic,
15H)
B) 2-Acetyl-sn-glycero-3-phosphocholine
15 `'
t
-- 2~ --
C) 1-Pa~mito~l-2-acetyl-sn-glycero 3-phosphocholine
The 1-0-triphenylmethyl ~roup uas e~imi~ated from
1-0-triphenylmethyl-2-acet~l-sn-glycero-3-phosphocholine
by ~he method stated in Ex~mp~e 5, stage 2, and the
; 5 result;ng 2-acety~-sn-glycero-3-phosphochoL;ne was reac-
ted with pa~mitîc anhydride by the method stated in
Examp~e 5, stage CO ~o give 1-pa~mitoyL-Z-acety~-sn-
giycero~3-phosphocholine.
Examp~e 9:
~0 A~ 1~0 Tripheny~methy~-2-methoxyacetyl-sn-g~ycero 3-
phosphochol;ne
316 ~g of 1-0-tripheny~methyL-sn-s-ycero-3-phos;
phochoLine according to Example 1 is reacted by the method
- stated in Examp~ 5, stage A, ~ith ~ethoxyacety~i~idazo-
~ide, ~h~ch ~as prepared from 90 mg of methoxyacet;c acidand 178 mg o~ carbonyld;imidazole, and the r~ixture was
worked UPD Z50 mg ~69X yield) of pure l-0-triphenylmethy~-
2-methoxyaGety~-sn-glycero-3-phosphocho~ine is obtained,
~hich g;ve~ a 3ingle spot in the thin-layer chromatogra~
Z0 (mobile ph~e: 65:35:5 ~Y/V~) chlorofo~m/methano~/25Z
strength a~ueous NH3 solution); R~ value 0.25.
B~ 2-Methoxyacetyl-sn-glycero-3-phosphocholine
C) 1-Stear3y~-2-metboxyacety~-sn-glycero-3-phosphocho~ine
The 1-0-tripheny~methy~ group was e~iminated by
Z5 the method stated in Examp~e S, stage ~, from 1-0-tri-
phenylmeth~l-2-methoxyacety~-sn-gLycero-3-phosphocholine~
and the resu~ting 2-methoxyacety~-sn-g~rcer~-3-phospho-
choline is reacted ~ith stearic anhydride as in Exa~p~e 6
to give 1-stearoyl-2-methoxyacetyL-sn~g~ycero-3-phospbo-
cho~ine, and the product is pur~fied by chromatography~Examp~e 10:
A) 1-O~Tripheny~methy~-2-~2'-ethy~hexanoy~)-sn-glycero-3-
phosphocho~ine
A soLut;on of 250 mg of 1-0-triphenyl~ethyl-sn-
~lycero-3-phQsphochol~ne ~according to Examp~e 1) is re
3cted by the method stated i-n ExampLe 5, stage A, ~ith Z-
ethy~hexanoy~;midazolide, ~hich was prepared from 144 ~
of 2 ethy~hexanoic acid and 178 mg of carbonrldiimidazoLe,
and the mixture ~as ~orked up. 200 mg ~9X yie~d) o~ pure
l'~ g
. ~ Z1
1-0-triphenylmethyL-2-~2'-ethy~hexanoyL~-sn-g~ycero-3-
phosphocho~ine are obtained, the product gi~;ns a ~in~le
spot in the ~hin-~ayer chromatogram tmobi~e phas2:
65:35:5 cY/vr~ ch~oro~orm/methanol~35X stren9th WH3~;
S ~f value 0.2~.
8) 2~ Ethy~hexanoy~) sn-~ycero-3-phosphochn~in~
C) l~=Oleoy~-2 t2'-ethy~hexanoy~)-sn-g-ycero-3-phosPho-
cho~ine
The l-0-tripheny-methr- group was eLiminated by
the method stated in Examp~e 5, stage ~0 from 1-0-tr~-
pheny~ethy~-2-t2~-ethy~hexanoy-~-5n-~ycero-3-phosphochoo
~iRe~ and the res~tin~ 20t2'-ethy~hexanoy~-sn-g~ycero-
3-ph~sphocho~n~ uas reacted ~ith o~eic anb~drid~ by the
~ethod sta~ed in Exa~p-e 5, sta~e C, to give ~-o~eoy~-2-
~5 t~'-ethy~hexanoy-~-sn-~y~ero-3-phosphochG-ine~ and the
product ~as purified b~ chromatography.
Example 11:. . ......... . . . .
A) 1-0-Tr;pheny-methy~2~ 4C)~dOdeCanOY~-sn~g~ycero-
3-p;llosphocholine
2n ~ 100 mg of.C1'-~C)-dodecanoic acid t1 IJci/m~ol~
and 90 ~9 of carbony~dîimidazo~e in 3 m- of anhydrous
tetrahydrofuran are reacted for 45 minut~s at ;2~~, The
soaut;on of the result;ng acy~im;dazo~ide is added t~
127 mg o~ 1 OoSripheny~ethy--sn-g-ycero-~ophosphoch31in~
After the soLvent has been stripped off in ~acuo, the
residue ts d;sso-ved ~n 1 ~ of d~methy~ su~foxide, and
the PeaCtion is initiat~d by addin~ the cata~ytic ac1d
: acceptor, ~hich has been obtained by d1sso-Ying Z3 mg o~
~eta~-~c sodiu~ in 18 ml of dimethy- su~foxide. The
reaction mixture is kept at Z0C ~or 10 minutes~ ~Sth
occasional shaking, after ~hlch it is neutra~;z~d by ?dd-
~ng 10 ~ of aqueous acetic acid a-~ at once~ Thereaft~r~
the ~ixture ls extracted ~sith t~ice 15 m~ of chlsro~oro~
m~thano~ t2:1, v~v).. Tb~ combined organic phases are
35 uashed successive~y uith 6 ml of chloro~orm~'methar)o~r
aqueo~ls NH3 solut;on C3~ 8:47,~ v~v/v) and ~ mt o~
chlloroform/methano-tualter t3:~8:47, vrYJv~ str;pp~nSl off
the so-vent in vaeuo and evaporat;on in the presence of
benzene gives the crude productO ~Ihich is purified by
~25~4~i~
- 22 -
1 medium pressure chromatography over silica gel using a
chloroform/methanol gradient, 110 mg of pure substance (82
yield) being eluted at a chloroform/methanol ratio of 6:4
( v/v ) .
The pure substance gives a single spot in the
thin-layer chromatogram over silica gel (mobile phase:
65:35:5 (v/v/v) chloroform/methanol/25~ strength aqueous NH3
solution); RfO.25. The radioacti~ity is 1 uci/mmol).
1H-NMR spectrum ~ in ppm (CDC13/CD30D, 2:1, v/v) 0.90
(acyl-Ch3, 3H) 1.26 ((CH2)n, s) 16H); 1.66 (H2C-C-CO, m,
2H); 3 16 (H3C-N, s, 9H); 3.3-4.4 (glycerol and choline-CH2,
8H); 5.23 (glycerol-CH, m, lH); 7.33 (aromatic, 15 H).
B) 2-(1'-14C)-Dodecanoyl-sn-glycero-3-phosphocholine
C) l-Oleoyl-2-(1~-14C)- dodecanoyl-sn-glycero-3-phospho-
choline
The l-O-triphenylmethyl group was eliminated by
the method stated in Example 5, stage B, from l-O-tri-
phenylmethyl-2-(2 -14C)-dodecanoyl-sn-glycero-3-
phosphocholine, and the resulting 2-(1 -14C)-dodecanoyl-sn-
glycero-3-phosphocholine was reacted with oleic anhydride as
in Example 5, stage C, to give l-oleoyl-2-(2~-14C)-
dodecanoyl-sn-glycero-3-phosphocholine, and the product was
purified by chromatography.
Example 12:
A) l-O-Triphenylmethyl-2-(9', 10 -dibromostearoyl)-sn-
glycero-3-phosphocholine
300 mg of 1-0-triphenylmethyl-sn-glycero-3-phos-
phocholine (according to Example 1) is reacted with 1.3 g of
9,10-dibromostearic anhydride in the presence of 300 mg of
4-N,N-dimethylaminopyridine in 20 ml of chloro~orm for 5
hours at 20C by the method stated in Example 8, stage A, and
the mixture is worked up. 460 mg (82% yield) of pure 1-0-
- ~2S~
- 22a -
1 triphenylmethyl-2-(9 ,10 -dibromostearoyl)-sn-glycero-3-
phosphocholine are obtained, which gives a single spot in
the thin layer chromatogram ~mobile phase: 65:35:5 (v/v/v)
chloroform/methanol 25% strength aqueous NH3 solution); Rf
5 value 0.3.
~`?
12S~
23 --
~ 2-~9tolol-Dibromostearoyl)-sn-g~ycero-3-phosphocholine
C) 1-Stearoyl-2~~Q~,lO'-dibromostearoY~)-sn-g~ycero-3
pho 5p hocho~ine
The 1-0-tripheny~methyl group uas e~iminated fro~
5 1-0-tripheny~ethy~-2-$9D~10'-dibromostearoyl~-sn~g~ycero-
3-phosphocholine by ~he method stated in Exa~p~e 5~ stage
B~ and the resulting 2-~9',10'-dibromos~earoy-3-sn-
g~ycero-3-phosphocho~in~ was reacted ~ith stearic anhy~
dride as in Examp~e 6 ~o give 1-stearoyl-2-C9'~10'-dibromo~
stearoy~-sn-gLycero 3-phosphoch~ine, and the produc~
~as puri~ied by chromatography.
Exa~p~e 13:
A) 1-O~TriphenyLmethyl-Z-arachidQnoyL-sn g-ycero-3-phos~
phochoLine
~S 550 ~g of 1-O~tripheny~ethyLrsn-g~ycero-3 phospho-
cho~ine taccording to Example 1) is r~acted ~ith 1.50 ~
of arach~donic anhydride under an argon atmosphere by the
method stated in Examp~e 12~ A~ter the add1tion o~ lS m~
o~ methanol~ the mixture is washed with 9 mL of chloro-
form/methanol/uater ~3:48:47, v/v~v), after ~hich the
~o~er phase is brought to dryness ~n vacuo, and the pro-
duct is purified by medium pressure chromatography~
750 mg ~80X yieLd) of pure 1-0-tripheny~methy~-2-
arach;donoy~ sn-g~ycero-3-phosphochoLine are obtaîned,
~hich g;ves a sing~c spot in the thin-Layer chromato~ram
(mobi~e phase: 65:35:5 Cv/v/v~ chloroform/methano~/Z5%
strength aqueous NH3 so~uti4n); Rf vaLue 0.3.
8) 2-Arachidonoy~-sn-g~ycero-3-phosphocho~ine
C~ 1-Stearoy~ 2-arachidonoy~-sn-g~ycero-3-phosphocho~ine
The 1-~-triphenylmethyl group ~as eLiminated fro~
l-O-triphenyLmethyi-2-arach;donoy~-sn-gLycero-3-phospho-
cho~ine by the method stated in Exampie 5, stage ~O and
the resu~t;ng 2-arachidonoyl-sn-g~ycero-3 phosphocholine
~as reac~ed with stearic anhydride as in Examp~e ~ to giv~
1-stearoy~ 2-arachidonoy~-sn-g~ycero-3-phosphocho~ne,
and the product ~as purified by chromatography~
ExampLe 14:
A~ 1-O TriphenylmethyL-2-tetracosanoy~-sn-g~ycero-3
phosphocho~;ne
~ZS3~46;~
- 24 -
1 316 mg of 1-0-triphenylmethyl-sn-glycero-3-phos-
phocholine (according to Example 1) is reacted by the method
stated in Example 5, stage A, with tetracosanoyl-
imidazolide, which has been prepared from 1,7 g of tetra-
cosanoic acid and 356 mg of carbonyldiimidazole, and the
mixture is worked up. 850 mg (62% yield) of pure 1-0-
triphenylmethy-2-tetracosanoyl-sn-glycero-3-phosphocholine
are obtained, which gives a single spot in the thin-layer
chromatogram (moblie phase: 65:35:5 (v/v/v)
chloroform/methanol/25% strength agueous NH3 solution); Rf
value 0.3.
B) 2-Tetracosanoyl-sn-glycerO-3-phosphocholine
C) l-Oleoyl-2-tetracosanoyl-sn-glycero-3-phosphocholine
The 1-0-triphenylmethyl group was elimlnated from
1-0-triphenylmethyl-2-tetracosanoyl-sn-glycero-3-
phosphocholine by the method stated in Example 5, stage B,and the resulting 2-tetracosanoyl-sn-glycero-3-
phosphocholine was reacted with oleic anhydride to give 1-
oleoyl-2-tetra-cosanoyl-sn-glycero-3-phosphocholine, and the
product WaS purified by chromatography, these steps being
carried out as in Example 5, stage C.
Example 15:
A) 1-0-Triphenylmethyl-2-oleoyl-sn-glycero-3-phospho-(N-
triphenylmethyl)-ethanolamine
386 mg of oleic acid and 255 mg of carbonyldi-
imidazole in 10 ml of tetraphydrofuran are stirred for 45
minutes at room temperature in the absence of moisture, 480
mg of l-0-triphenylmethyl-sn-glycero-3-phospho-(N-
triphenylmethyl)-ethanolamine (according to Bxample 2) are
taken up in the solution of the oleoylimidazolide, after
which 3,1 ml of a solution of 41 mg of sodium metal in
dimethyl sulfoxide are added. The mixture is allowed to
=
~25~
- 24a -
1 react for 20 minutes at room temperature and is then
neutralized with 17.8 ml of 0.1 N aqueous acetic acid. it
is then taken up in ice water and extracted three times with
diethyl ether, the combined organic phases are washed twice
with water and dried over Na2S04, and the solvent is then
stripped off in vacuo. After purification of the crude
product by chromatography over silica gel using a
~.
;~5, ~S ~6;~ ~
petro~eum ether/chloroform gradient,~ 633 mg of the pure
target compound are obta;ned. Thin-Layer chromatography
of ~he product over si Li ca ge~ Cmobi Le phase: 9:1 tYtY~
ch~oroformlmethanol~ giYes a s;ngle spot t~ 0.6).
S 1H-NPlR spectrum: S ppn~ (CDCL33
0.88 t-CH3),, 1.2~ t-eH2~); 1.8-2.3 ~-CH2-CH2-~=C,,
-CH2~cH2~ C=0); 3,1-4.3 ~g~ycero~ and lethano~amine C-H);
5.1 ~g~ycero~ C2`c-Ho o~eoyl HC-CH~; 7.25 Ctripheny~-
methy~ C-H~.,
B) 2-Oleoyl~sn-g~ycero-3-phospho-(N-~riphenylme~hy~-
ethano~amtne
A ~o~ution o~ 406 mg of ~ 0-tripheny~methy~-.2-
o~eoy~-~n-g~ycero-3~phospho-CN~tripheny~methyL~-ethanoL-
amine in a mixture of 30 m~ of methy~en~ ch~oride and 1 m~
of a ZOX strength soLution of boron trifLuorite in
~ethanol i 5 st~ rred for 30 m;nutes at 0C. 30 m~ of
~ethy~ene l:hloride are then added, the mixture is ~ashed
three times with water, and the organic phase is dried
over Na2S0,~.
ZO C) 1-Palm;~:oyl-2-ol~oyl-sn-g~Ycero-3-phospho-tN-tripheny~-
methyl)-ethanolamine
1.S g of palmitic anhydride and Z30 mg of dimethylo-
aminopyrid;ne are added to the so~ution of the r~su~ting
2-o~eoy~-sn-gLycero-3-phospho-~N~tripheny~methyl~-ethano~-
amine~ The reaction mixture is ~tirred for 6 hours at20C~ The so~vent is then stripped off in vacuo. Aft2r
the residu~ has been chromatographed over si~ica ~e~ using
a~petroLeuln ether/chloroform gradient~ 3~1 mg of the pure
acylation product are obta;ned. When s~bjected to th~n-
layer chromatography over si~ica gel Cmob;~e phase: 9:1
SY/V) ch~oroform~methanol)~ the compound gives a sing~e
spot ~Rf o~
~H-NMR spectrumo S ppm ~CDCl3)
OD88 t-CH3); 1.22 t-CHz~ 55 t-CH2 C-C0); 108-2~3
~-CH2~CH2-C~ ~CH2-C=0), 3.1-4,3 tg~ycero~ and
ethanolamine C~H~; S.l ~glycerol = C-H, o~eoyl CH-CH);
7.25 ~tr;phenylmethyl C-H).
D~ a~m;toy~ 2-o~eoy~-sn-g~ycero-3-phosphoethano~amine
A solution of 190 mg of 1-pall~itoyl-Z-o~eoy~-sn-3-
' 26 - ~5~4~
glycero-~N-tr;pherly~methyL~-ethanolamine in 12 ml of a
~ixture of ~ m~ of methylene ch~oride and 6 m~ of
trifluoroacetic ac;d i5 (eft to stand in the absence o~
moist~re for 5 m;nutes at O~C~ It is then neutralized
by addin~ 23 ~ of a six per cen~ strength aqueous
ammonia solut;on aLl at once. The aqueous phase is sepa-
rated off and extracted ~ith 2~1 Cvrv) ch~oro~ormJ
methano~. The combined OrganiG phases are ~ashed ~ith
~ater. A~ter the solYent has been stripped o~f in vacuo~
'che crude product is purified by chromatography over
s~ica ge~ using 3 ch~oro~orm/methano~ gradien~. ~hen
sub jested to thin-layer chromatography over si ~ica ge~
~obi~e phase: 50~25:b Cvrv/v) chLorofor~/methano~125%
- strength aqueous NH3 sclution), the co~pound gives a
sing~e spot ~R~ C.
Examp~e 16:
A) 1-0-Tripheny-methyl-2-lino~eoy--sn-glycero-~-phospho-
CN-triphenylmethyl)-ethanola~ne
464 mg o~ l1nole~c ac~d and 365 mg o~ carbonyl-
d;;midazo~e in 15 m~ of tetrahydrofuran are reacted for
45 minutes at 20C. The solution of the resuLting acy~
;midazolide ;s added to 530 ~9 of 1-0-triphenylmethyi-sn-
gl.ycero-3-phospho-~N-triphenylmethy~)-ethanolamine
taccording to Example 2~. After the solvent has been
stripped off ;n vacuo, the residue is d;sso~ved in t5 m~
- of dimeth~l su~foxide, and the reaction is initiate~ by
addiny the catalyt;c ac;d acceptor, ~hich has been obt~fned
by dissolving 49 mg of metallic sodium in 3.7 m~ of
dimethy~ suLfoxide. The reaction mixture is kept at 20C
for 20 minutes with occas;onal shaking, after ~h;ch it is
neutra~ized with 10~4 m~ of aqueou~ 0.1 N acetic acid,
added a~ at once. The m;xture is then extracted ~ith
t~ice 20 m~ of 2:1 ~v/v3 chloroform/metha~o~. The combined
organic ph3ses are washed successive~y ~ith 8 m~ of
3S chloroform/~ethano~/aqueous NH3 solution ~3:~8:47,
v/vlv9 and 8 ~ of ch~oroform/methano~/hater ~3 48 47O
v1v/v). Str;pping off the solvent in vacuo and evaporatin~
the residue in the presence o~ benzene gave a crude pro-
duct ~hich, ~hen purified by ~edium pressure chromatography
. . .
. - ~ z7 - :~l2~4~
O~Jer si ~ica ge~ us;n~ a ch~oroformJtnethano~ gradient~
~ves 670 mg (9ZX y;e~d~ of the pure compound. In th~
th~ layer chromatogr3m over si ~ica ge~ Cmob; ~e ~hase:
8:3 ~v~v~ rrlethano~/chloroform)~ the pure compound gives
S a sin~e 5pot; Rf 0.61
B~ 2-Lino~eoy~-sn~glycero-3-phospho-~N-triphenyl~ethy~
~thano~amine
The 1 0-tripheny-me~hyl gro.up 1s e-iminated fro~
435 ~ of ~ tr~pheny~meth~l-2-linoleoyl-~n-~lycero-3-phospbo-tN-tri
tO pheny~ethy~)-ethano~a~ine by the method stated in
Exa~le 15, sta~ ~, using O.S ~ o~ a 2~X stren~th 50~u-
.tion o~ boron trif~uorlde ln ~eth~noL~
eoy~-2~ o~eoyl-sn-g-ycero-3-phospho-tN-tr~-
phenyl~ethy~)-ethano~am~ne .
D~ 1-Oleoyl-2--ino~eoy~-sn-g~ycero 3-phosphoethano~am~ne
2-Lino~eoylL-sn-g~ycefo-3-phospho-tN-tripheny~--
~ethy~-ethano~anine is ~urther acylated 3na-0gous~ to
the ~ethod in Example 150 ~tage C, by reaction ~ith ole1c
anh~dr~d~, to give 1-o~eoyl-2~1inoleoyl-sn-glycero-3--
phospho tN-triphenylmethyL)-ethanolamine, and the ~-tri-
pheny~methy~ radical is e-imlnated frQm this ana~o~ous~y
to the ~ethod stated ;n ~ample 15, stase D.
Examp~e ~7:
A) 1-0-TriphenyLmethyL-2 stearoy~-sn-gLycero-3-phospho-
~N-tripheny-~ethy~)-ethano-amine was prepared anaLogous~y
to th~ ~e~hod ~n Examp~e 16, stage A, by reaction o~ 380 ~9
of 1-0-~ripheny-m~thy~sn-g~ycero-3-ph5spho-C~-tr~phe~y~
~ethy~-ethanola~;ne ~lth 350 ~ o~ stearo~imidazo~ideO
83 Z-Steal~oy--sn-glycero-3-phospho-CN-tripherly-metby~)-
ethano~2mine ~as obta;ned ~rom the product obtained instage A by e~i~inating the 1-0-tr~pheny~meth~- group b~
the ~thod stated in Examp-e 150 stage 8.
C) .'I-O~eoyl-2-ctearoyL-sn-g-ycero-3-phospho-~N-triph~ny~
~ethy~) et~ano~amine
D) 1-O~eoy~-2-stearo~sn~g-ycero-3-phosphoethanolam~ne
The 2-stearoyl-snoglycero~3-phospho-~N-tr;pheny~-
~ethyl~-ethanolamine obtained in stagè B is reacted ~ith
105 ~ of o~eic anhydride to gi~e 340 mg of 1-o~eoyl-2-
- 28 - ~ ~ S~
stearoy~-sn-glycero-3-phospho-~N-tripheny~-methyL~-
ethanolamine ana~ogous~y to the method in Examp~e 150
stage C. 3etr;tyla~ion analogous to the method in
ExampLe 15, stage D, ~i~es 250 mg of 1-o~eoy~-2-stearoy~-
S sn-g~ycero-3-phosphoethanolamine~ chromatographing this
over s;lgsa ge~ using a chloroform/methano~ grad;ent
gives 140 mg of pure product, ~hich sho~s a sing~e spot
in the th;n-layer ch~omatogram ~mobi~e phas~: 50:25:~
~Y~V/V) ch~orofor~tmethanol/25X strength aqueous NH3
solution); Rf ~a~ue 0,5
Examp~e 18:
A~ 1 O Tripheny~methy~ Z-acety~ sn-glycero-3-phospho-CN-
tr~pheny~methy~9-ethanolamine
300 mg o~ 1-0-triphenylmethyL-sn-g~ycero-3-phospho-
~N-triphenylmethy~)-ethano~amine Caccording to Examp~e 2)
are reacted with 500 m~ of acetic anhydride in 5 m~ of
anhydrous pyridine for 20 hours at 20C. The m;xture is
then pa~red onto 50 m~ of ice ~ater and extracted twice
w~th 2:1 ~v/v) chloroform/~ethanol~ The combin~d organic
2~ phases are ~ashed t~ice ~ith 3:48:47 ~v/v/v) chloro~orm/
methanol/~aterO dried over sodium bicarbonate and brough~
to dryness in vacuo. Residua~ pyridine is removed by
evaporation in the presence of ~o~uene. The crude product
~s purified by medium pressure chromatography over si~ica
gel using a ch~oroform/methanol gradient. The pure sub-
stance obtained C128 mg, 40X y;eld) g;ves a singLe spot
in the thin--ayer chromatogram over silica ge~ ~mobi~e
phaser 8:2 tv/v) ch~oro~ormlmethano-); R~ va~ue 0~50.
1H-NMR spectrum S ppm tcDcl3/cD3oD 2:1 v~v);
2~0 ~H3C-Co, s~ 3H); 2.8-4.2 Sg~ycerol and cho~ine-CHz3;
5016 ~g~ycero~-CH, m, 1H~; 7~23 ~aromatic, 30H)
1-n-Tripheny~methy~-2-acety~-sn-g~ycero-3-pho~pho
t~-tripheny~methy~amine)-ethano~amine ~a5 converted ana~o-
~ous~y to the methods ln stages 8, C and D of Examp~e 15
to give the fo~owing further products:
B) 2-Acety~-sn-g~ycero-3-phospho-tN-tripheny~me~hyl)-
ethanolam;ne
C) 1-Palmitoyl-2~ acety~-sn-glycero 3-phospho-~N-tri-
phenylmethrl)-ethanolanline
- ` ~ 29 ~ ~ ~ 5~
D~ 1-palmitoyL-2-acety~-sn-s~ycero-3-phosphoeth3noLa~ine
Examp~e 19:
A) 1-0-Triphenylmethy~ 2-propionyl-sn-~lycsro-3-phospho-
CN-tr;pheny~methy~-ethano~amine
450 m~ o~ 1 0-triphenr~methrL-sn-~(ycero-3-phosph~-
CN-tripheny~methyl~ethano~a~ine taccording to Examp~e 2a
were reacted ~;th 900 ~9 of propionir anhydride in 10
of pyridine at 30C to give 1-0-tripheny-methy~-Z-pro-
piony~-sn-gLycero-3-phospho-~N-tripheny~methy-) ethano~-
a~ineD the ~ixture ~as ~orked up and the product ~as puri-
..
_ f;ed, these s~eps being carr;ed out an3~0gous~y t~ the
~ethod in Exahp-e 18, stage A.
YieLd~ 220 mg ~5X of theory); Rf va~ue 0.5.
- The product obtained ~as converted ana~ogously to the
~ethods in Examp~e 15, sta~es 8~ C and D, to give the
7n~owing compounds:
~) 2-Propionyl-sn-glycero-3-phospho-tN-tripheny~meehyl~-
ethanolamlne
C~ 1-Palm~toyl-2-prop~onyl-sn-glycero-3-phospho-~N-tri
phenylmethyl)-ethanola~;ne
D~ 1~Palmitoy~-2-propionyl-sn-glycero-3-phosphoethanoL-
amine
Examp~e 20:
A3 1-0-Triphenylmethy~-2-butyry~ sn-g~ycero-3-phospho-CN-
triphenylmethy~)-ethano~amine
500 mg of 1-0-triphenylmethy~-sn g~ycero-3-
phospho-CN-triphenyLmethyl)-ethano~amine ~accordin~ to
Examp~e 2) ~as acylated ~ith ~utyry~imida20lide, ~hich was
obtained from 132 mg of but~ric acid and 267 mg of car-
bony~diimidazo~e, the mixture was worked up and the proo
duct ~as purified, these steps being carried out as ~n
Example 16, stage A. The product abtained ~300 mg~ 56%
- yie~d; ~f va~ue 0.5Z, over silica ge~, ~obi~e phase:
8:2 ~v/v~ ch~orsformtmethano~) ~as reacted further ana~o-
~ous~y to the methods in Examp~e 15, stages 8, C and D~
to give the ~o~lowing compo~nds:
~) 2-8utyry~-sn-g~ycero-3-phospho-~N-triphenylmeth~
ethano~amine
~ 2~4~
- 30 -
C) 1-PaLmitoy~-2-butyry--sn-g~ycero-3-phospho-CN-tri-
phenylmethyl~ethanolam;ne
D~ 1-Palm;toyl-Z-butyryl-sn~ Cero-3-Phosphoethano~a~ine
ExampLe Z1:
A~ 1-0-Tripheny~methy- 2-;so~utyry~-sn-g-ycero-~-phospho-
~N-triphenyL~ethy~)-ethanolamine
500 mg of 1-0-tripheny~methy~-sn-g~ycero-3-
phospho-tN-tripheny~methy~)-ethano~amine taccording to
Examp-e 2) was acylated ~ith isobutyryLimidazo~ide~ which
~as obta;ned from 732 mg of ;sobutyr;c ac;d and Z67 mg of
carbonyldiimidazo~e, the mixture ~as ~orked ~p and the
product was purified, these steps be~ng carr;ed out as in
Examp~e 16, stage A, The product obta;ned ~27Z ~9~ 50X
yield; R~ va~ue 0.53~ over si~ica ge~, mobiLe phase~
8:2 Cv/v) chloroformJmethanol) g;ves the folLouing
spectroscopic data:
~H-NMR spectrum: ~ ppm ~cDcl3/cD3oD~ 2:1, v/v)
1.2 tCH3, 6H); 2.5-3.9 ~glycerol and cho~ine CHz, ~H);
5~Z6 ~glycerol-C~I~ m, 1H~; 7.23 ~aromatic, 3UH?.
1-0-Tr;phenylmethyL-Z-isobutyryl-sn-g~ycero-3-
phospho-~N-tripheny~methy~)-ethano~am;ne was converted
anaiogous~y to Examp~e 16, stages B, C and D, to gi~e the
fo~owin~ compo~nds:
B) 2-Isobutyry~-sn-g~ycero-3-phospho-tN-triphenylmet~y~
ethanolamine
C) ~-Oleoyl~Z-isobutyry~-sn-g~yeero-3-phospho-~N-tri-
phenylmethy~)-ethano~amine
D) 1-Oleoyl-2-isob~tyryl-sn-g~ycero-3~PhosPhoethano~amîne
Examp~e 22:
A) 1-0-Tr;phenylmethy~ 3'-trifLuoromethylbutyryl~-sn-
~lycero-3-phospho-CN-triphenyLmethy~)-ethano-amine
900 mg of 1-0-tripheny~methy--sn-g~ycero-3-phos-
pho-~N-triphenylmethyl)-ethanolamine Caccording to
Exa0p~e 2~ and 500 mg of 3-trifluoromethy~butyryl anhy-
dride in S0 m~ of methylene ch~oride ~ere stirred for~ hours at Z0C after 300 mg of 4~N,N-dimethy~amino-
pyridin~ had been added. After the addition of ~ater,
the mixture is extracted by shaking, and the organic phas~
is dried cver NazS04 and evaporated down in Yacuo. After
~25~6~ ~ I
- 31 -
the crude product has been purified by medium pressure
chromatography oYer silica ~el ~s;ng a chloroform/methano~
gradient in the presence of 005X by voiume of aqueous NH3
so~ution, $Z0 m~ ~76% yi~d; R~ Yalue 0~500 over si~ica
S gel, mob;~e phase: 8:2 ~ chloroformtmethanoL) o~ 1-
0-triphenylmethyL 2-~3'-tri~uoromethy~butyry~3-sn-g~yc~ro-
3-phospho-tN-tripheny~methy~)-ethanoLamlne are obtained,
This product ~as conYerted analogously to Example 150
stages a~ C and D, to g;ve the follo~;ng compounds:
B) Z-~3'-Trif~uoromethylkutyry~3~sn-g~ycero-3-phospho-
tN-triphe~ylmethyl) ethanolamine
C) 1-Pa~m;toy~-2 (3~-trif~uoromethy~butyry~-sn-g~ycero-
3-phospho-~N-tripheny~methyl)-ethano~amin2
D) 1-Pa~mitoy~-Z-C3'-trifluoromethy~butyryl)-sn-g~ycero~
3-phosphoethanolam;ne
Examp~e 23:
A~ 1-0-Triphenylmethyl-2-t2'-butylhexanoyl)-sn~glycero-
3-phospho-CN-triphenylmethy~)-ethanolamine
430 mg of 1-0-~ripheny~methy--sn-glycelo-3-phospho-
CN-triphenylmethyi)-ethanola~nine ~accord~ng to Example 2)
~as acy~ated w;th 2-butylhexanoy~imidazo~ideg ~hich ~as
obtained from 200 mg of Z-butylhexanoic acid and Z1~0 mg
of carbonyldiimidazoleO the mixture was worked up and tbe
product ~as purified, these steps being carried out as in
Examp~e 16, s~age A. The product obtained C37~ ~9~ 70X
y;eld; R~ va~ue 0.55, over silica gel, mobile phase: 8:2
~v/v) chloroform/methanol) ~as reacted further ana~ogous~y
to the methods ;n Example 15, stages ~ C and D, to give
the following compounds:
~ 2-~2'-Butylhexanoyl)-sn-glycero-3-phospho-~N-tri-
phenyLmethyl)-ethanolamine -
C) l-Pa-mitoy~-Z-~2~-buty~hexanoyl)-sn-glyGero-3-phosphQ-
tN-tripheny~methyl)-ethanolamine
DS l-PalmitQyL-Z-~Z'-butylhexanoy~)-sn-gLycero-3-phospho-
3S ethanolamine
Examp~e 24:
A) 1-0-Triphenylmethy~ ~-arachidonoyl-sn-glycero-3-
phospho-~N-tripheny~methyl)-ethanolamine
32 - ~ Z S~
500 ~9 of 1~0-triphenyLmeth~--sn-sLyc~ro-3-ph~s-
pho-~N-tr;phenylmethyl)-ethanoLamine taccording to
ExampLe 2) and 89Q mg of arach;donic anhydride in 50 m~
o~ methyLene chLoride ~ere st;rred for 4 hours at 20C
; 5 after 300 m~ of 4-NrN-dimethyLa~inopyridine had been added.
After the addition of ~ater, the mixture is extracted by
shaking, and the organic phase ;s dried over Na2so4 and
evaporated do~n in vacuo. After purification of the crude
product by medium pressure chromatography over siLica ~el
10 us;ng a ch~oroform/methanoL gradient in the presence of
O.SX by vo~ume of aqueous NH3, 520 mg (82X yieLd; R~
vaLue 0.~1, over si-;ca ge-, mobile phase: 8:2 tvfv~
chLoroform/methano-) of ~-0-tripheny-methy~-2-araGh~dony~-
sn gLyrero-3~phospho CN-tr;phenyLmethy~)-ethano-amine are
t5 obtained. This product ~as converted anaLo~ousLy to
Example 15, stages 8, C and D, to give the folLowing
compounds:
B) 2-Arachidonoy~-sn-gl,ycero-3-phaspho-~N-triphenyl
methyL)-ethano~amin6!
C) 1-PaLmito~L~2-arachldonoyl-sn-gLycero-3-phospho-~N-
triphenylmethyl)-ethano~amine
D) 1-Pa~mitoy~-2-arachldonoyL-sn-~ycero-3-phosphoethan
am;ne
Examp~e 25:
A) 1-0-TriphenyLmethyL-2-tetraGosanoyL-sn~gLycero-3-
phospho-CN-triphenyLmethyl)-ethanoLamine
S00 mg o~ 1-U-~ripheny-methy--sn-gLycero-3-
phospho-tN-triphenylmethyL3-ethanoLamine ~according to
Examp-e 2) was acyLated with tetracosanoyLimidazoLide,
which was obtained from 1~57 9 of tetracosano;c acid and
Z67 mg of carbonyLdiimidazoLe~ the mixture was worked up
and the product was p~rif;ed, these steps bein~ carried
out as in ExampLe 16, stage A. The product obtained
t640 mgO 90X yieLd; Rf vaLue 0.6, over siLica ge~,
3S mobi~e phase: 8:Z ~v/v~,ch-oroform/methano-) was reac~ed
further anaLogous~y to the methods in ExampLe 15, stages
~9 e and D~ to give the fo~Lowing co~pounds:
9) 2-TetracosanoyL-sn gLycero-3-phospho-(N-tr;phenyL-
methyL3-ethanoiamine
- 33 ~ 4~ ~
C~ 1-Pa~itoy~~2-tetracosanoy~sn-~ycero-3-pho~pho~
tr;phenyl~ethy~)-ethanolamine
Dl 1-Palmitoy~-2-tetracosanoyi-sn-glycero-3-phospho^
ethanolamine
; 5 Examp~e ~6:
C) 1-StearoyL-Z-oLeoy!L-sn-g~ycero-3-phospho ~N~tripheny~
~e~hy ~) -ethano ~ami ne
3.5 9 o~ steario anhy~ride and 6Z5 mg o~ 4-N,N-
dimethy~a~;nopyr;dine are add~d to a solut;on~ in ~50 m~
of meth~Lene ch~oride, of 2-o~eoy~-sn-gLycero-3-phospho
tN-tripheny~methy~)-ethano~am~ne~ ~hieh ~as obtained ~ro~
1~1 9 of 1-0-tripheny~methyl-2-o~eoy--sn-g-ycero-3 phos-
pho-~N-triphen~l.methy~-etha~o~amirle tsee Examp~e 15,
stages A and B). The reaction mixture was stirred for
3 hours at 2~to after ~hich 45 m~ of chlorofor~Jmethano~/
~ater (3:48:47, Y/V/~) and 75 ml of methano~ ~ere added.
After extract~on by shaking, the lower phase is separated
nff, and ~ashed with t~ice 45 ml o~ chloroformtmethanol/
aqueous ammon~a ~3:48:47, v~v/Y) and once ~ith chloroform/
~0 methanol/~ater C3.48:47, v/v/v). A~ter the so~vent has
been s~ripped off in vacuo, the crude product is purified
by medium pressure chromatography over silica gel using
a ch~oroform/methano~ gradient, 990 mg C~9X yie~d, Rf
va~ue 0;~ over siLica ge~, mobiLe phase: 9:1 ~v/v)
Z5 chloroform/methanol) of pure substanc~ being obtained.
D) 1-Stearoy~-2-oleoy~-sn-~lycero-3-phosphoethanolamine
1 9 o~ 1-stearoy~-2-oleoy~-sn-glycero-3-phospho-
C~-triphenylmethy~)-ethano~amine in 30 m~ o~ trif~uoro-
acetlc acid ;s ~eft to stand for 5 minutes at 0C.
Working up ;s carried out as in Examp~e 15, stage ~.
550 mg of pure substance S72X yield) are obtained, ~hieh
gives a sing~e spot ;n the thin-~ayer chromatogram over
si~ica ge~ tmobi~e phase: 50:25 6 CYIV/V1 chlorofor~t
methano~Z5% strength NH3); Rf va~ue 0.5.
xamp~e 27:
Cl 1-Stearoy~-2-~;no~eoyl-sn-~ycero-3-phospho-tN-tri-
phenylme~hyl)-ethano~amine
D) 1-Stearoy~-2-l;noleoyl-sn-glycero-3-phosphoeehanola~ine
2.2 9 of stearic anhydrlde and 390 mg of
~ 34 _ 1 ~ S~ ~6
4~N,N-dimethylaminopyridine are added to Z-~ino~eoy~-sn-
g~ycero-3-phospho ~N-tr;pheny~methyl)-ethano~amine~
obtained from 70D mg of 1-0-triphenylmethyl-Z-~inoLeoy~-
sn-g~ycero-3-phospho-~N-tripheny~methyl)-ethanolam;ne
S ~see Examp~e 16~ stages A and B), in methylene ch~oride,
and ~he mixture ;s stirred for 3 hours at room tempera~
ture~ The product obtained is reacted ~ith trifluoroacetic
acîd, and the product is purified, these steps being
car~ied out as described in Examp~e 15, stage D.
This procedure giYeS 250 mg of pure 1-stearoy~-2-
~inoleoy~ sn gLycero-3-phosphoethano~amine, ~hich ShDW5
sing~e spot in the thin-Layer chromatogram over ~iLica
~e~ ~mobi~e phase: ch~oroformJmethano~ as in ExampLe Z~);
Rf vaLue 0.5.
Thls application is a divisional of the applicant's
Canadian patent application, serial number 48l,204, filed
May 9, 19~5.
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