Organic & Biomolecular Chemistry
The first approach involves coupling an alcohol acceptor 4
Page 2 of 5
DOI: 10.1039/C9OB02582K
to a phosphorus donor, 7 - 9, leveraging either the more elec-
trophilic phosphorus (III) or more stable phosphorus (V) oxida-
tion state, to generate a phosphorus III or V intermediate (Fig-
ure 2A). After the initial O-P bond formation, a wide variety of
reagents (e.g. 10) can be used to insert choline. The final step
in the synthesis is to correct the oxidation state of the phospho-
rus atom, if necessary, to produce 6. Due to the instability of
phosphorus (III) donors, ethylene glycol derivatives of phos-
phoryl chloride (12 and 13) became the most widely used rea-
gents to install the ChoP residue (Figure 2B).8-12 This approach,
however, is not without its limitations. Reagents 12 and 13 are
air sensitive, water sensitive, and prone to decomposition on
silica gel. Additionally, the final amination step requires the use
of either gaseous trimethylamine or ammonia and methyl io-
dide.
While subtle, we observed that the new version of the rea-
gent could be synthesized in high yield with excellent purity.
Importantly, the synthesis does not require a purification step.
We hypothesize that the ease of synthesis is due primarily to the
enhanced solubility of the tetraphenyl borate-choline salt and
careful monitoring of each step of the reaction using a combi-
nation of 31P NMR and LCMS. Moreover, phosphoramidite 16
is surprisingly stable to oxidation. Thus far, we have observed
that 16 can be handled in air with minimal oxidation (as moni-
tored by 31P NMR) for several weeks. Finally, 16 is stable to
aqueous work up.
Before revealing the utility of reagent 16, we briefly di-
gress into a mechanistic discussion of how the reagent works.
The accepted mechanism for phosphotidylation of alcohols, us-
ing phosphoramidite reagents, is shown in Figure 4. 1H-te-
trazole (pKa = 4.9) 21 is used as a promoter in the reaction.
Based on its acidity, the first step in the coupling reaction
should involve protonation of phosphoramidite 16 and nucleo-
philic displacement of diisopropyl amine by tetrazolide 22. The
key intermediate in the coupling is tetrazolylphosphoramidite
22. At this stage, coupling with alcohol acceptor 4 will provide
a phosphite ester 23. Subsequent oxidation, and b-elimination
of the cyano ethanol protecting group will provide phosphocho-
line 6. Based on this mechanism, optimization of reaction con-
ditions should involve an evaluation of different tetrazole based
promoters, amine bases, and oxidants.
It was after encountering these difficulties that we became
interested in developing phosphorus III regents of type 14, con-
taining a pre-installed choline unit, a phosphorus protecting
group, and a leaving group that would be displaced by the alco-
hol acceptor. It was in this vein that we recalled an obscure re-
agent, 15, used by Pedersen and Schmidt during their elegant
total synthesis of the lipoteichoic acid from Streptococcus
pneumoniae.13-14 In these studies, a tosylate salt of a phospho-
choline-loaded phosphoramidite was used to install phospho-
choline on a primary alcohol. Unfortunately, the reagent was
only used on a single substrate. Moreover, the preparation of
the choline donor is difficult, primarily due to the poor solubil-
ity of choline tosylate. Finally, the synthesis of the reagent is
not reported in the literature. In our hands, all attempts at pre-
paring the reagent were low yielding. Moreover, we observed
oxidation of the phosphorus center, during the synthesis, which
complicated purification.
The goal of the chemistry described herein was to synthe-
size and evaluate an improved phosphocholine donor, 16 (Fig-
ure 3). At the planning stage, we hypothesized that using a cho-
line salt bearing a non-coordinating anion, such as tetraphenyl
borate (BPh4), (reagent 20) would improve the solubility of all
intermediates en route to 16. Indeed, the use of tosylate salts
provided insoluble intermediates over the course of the synthe-
sis. Moreover, we anticipated the stability of the 16 would be
greatly enhanced. In the forward direction, the synthesis of 16
started from 2-cyanoethanol 17, which was reacted with phos-
phorus trichloride to provide phosphorus dichloride 18. At this
stage, monitoring the rate of addition of phosphorus trichloride
and the temperature of the reaction is critical to avoiding disub-
stituition of cyanoethanol onto the electrophilic phosphorus
atom. Following the first substitution, reaction with diisoprop-
ylamine gave diamine 19 with 31P NMR indicating formation
of the phosphordiamidite. The final step in the synthesis is re-
action with choline tetraphenylborate 20 to obtain donor 16 in
71% yield over the entire synthesis.
Figure 4. Proposed mechanism for the reaction between an alcohol
4 and phosphoramidite 16.
The investigation began with the reaction of cetyl alcohol
26 and 16 to produce miltefosine 27. Miltefosine is the only oral
drug approved for the treatment of parasitic disease leishmani-
asis. Unfortunately, it is also unavailable to most patients due
to its prohibitive expense.15 Upon extensive investigation, we
realized that using slight excess (1.2 equiv.) of the phospho-
ramidite, 1H-tetrazole, and TBHP enabled access to the pro-
tected drug. Ultimately, 5.0 equiv. of DBU gave miltefosine in
90% yield.
Deviating from the standard procedure, we first investi-
gated additional activators. We hypothesized that a superior
proton donor and/or a stronger nucleophile would enable in-
creased generation of the active intermediate - thereby increas-
ing the rate of reaction. Interestingly, each tetrazole derivative
performed with the same efficiency (entrys 1-3 and 5) likely
due to two of the reagents (ethylthiotetrazole and benzyl thio-
tetrazole) being more acidic and another reagent (DCI) func-
tioning as a superior nucleophile. Acetic acid, while of equal
Figure 3. Synthesis of phosphoramidite 16.
2