Selective synthesis of chlorophosphoramidites using ionic liquids

Article abstract of DOI:10.1039/b905000k

A range of chlorophosphoramidites have been prepared in ionic liquids and compared with material synthesised in molecular solvents. Through the use of ionic liquids as reaction media the moisture sensitivity and impurity issues hampering existing traditio

Full text of DOI:10.1039/b905000k

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Selective synthesis of chlorophosphoramidites using ionic liquids†
Eric J. Amigues,a Christopher Hardacre,*a Gillian Keane,a Marie E. Migaud,*a Sarah E. Normana and
William R. Pitnerb
Received 11th March 2009, Accepted 27th May 2009
First published as an Advance Article on the web 16th June 2009
DOI: 10.1039/b905000k
A range of chlorophosphoramidites have been prepared in ionic liquids and compared with
material synthesised in molecular solvents. Through the use of ionic liquids as reaction media the
moisture sensitivity and impurity issues hampering existing traditional synthetic routes have been
eased. Not only can stock chemicals be used without purification, but the reactions may be
conducted at room temperature and at high concentrations. Furthermore, reaction times are
reduced and rapid addition of reagents is possible whilst retaining tight control over product
selectivity. Beyond their role as reaction media, ionic liquids also present a unique storage medium
for these highly moisture sensitive chlorophosphoramidites.
there is still significant room for improvement.7 For example,
3-hydroxypropionitrile must be purified to remove the ethylene
Introduction
Nucleoside chemistry represents an important area of research
glycol contaminant via an expensive and complex synthetic
for drug discovery, with oligonucleotide-based therapeutics be-
purification sequence.
ing evaluated in preclinical and clinical studies for the treatment
Ionic liquids are known to be viable alternatives to molecular
of a variety of disease states such as cancer, cardiovascular
disease, diabetes and inflammatory disease disorders.1–3
solvents as they have negligible vapour pressure, unprecedented
ability to dissolve a broad range of compounds both organic
In order to prepare oligonucleotide-based therapeutics, nu-
and inorganic in nature, and have a wide liquid range.8 As such,
cleotides need to be synthesised by phosphorylation of partially
ionic liquids are finding applications in synthesis owing to their
protected nucleosides. Phosphorylation can be achieved in
utility in easing the preparation and recovery of targets in a num-
two ways; either by the introduction of a PV reagent or by
ber of cases.9 In particular, bis{(trifluoromethyl)sulfonyl}imide
phosphitylation with a PIII reagent followed by oxidation.4 Phos-
([NTf2]-) based ionic liquids represent a hydrophobic subsection
phitylation procedures with phosphoramidites and chlorophos-
of this class of solvents and, even when ‘wet’, by organic solvent
phoramidites have been used most extensively owing to the
standards, provide an environment which prevents hydrolysis
increased reactivity compared with the PV reagents and, with
of a range of hydrolytically unstable species, such as PCl3
the development of phosphoramidite chemistry, oligonucleotide
and PN(R1R2)Cl2.10,11 Herein, we report the facile preparation
synthesis has been transformed into an efficient and automated
of a range of chlorophosphoramidite reagents using ionic
process.5,6 However, phosphoramidites and chlorophospho-
liquids showing enhanced yields and selectivities over molecular
ramidites, in particular, are very sensitive to moisture and these
solvents (Scheme 1).
compounds often require fresh preparation before use since
they cannot be reliably stored over significant time periods.
In general, the preparation of these derivatives requires the
treatment of an excess of PCl3 with a highly purified alcohol,
with 3-hydroxypropionitrile being the most commonly used
protecting group in oligonucleotide synthesis, producing an
alkoxydichlorophosphine, which requires purification by distil-
lation. Subsequent reaction with an appropriate amine, again
requires purification by distillation before use. Although this
route has been extensively optimised in molecular solvents,
aQUILL and School of Chemistry and Chemical Engineering,
Queen’s University Belfast, Belfast, Northern Ireland BT9 5AG.
E-mail: c.hardacre@qub.ac.uk, m.migaud@qub.ac.uk;
Fax: +44 28 9097 4687
bMerck KGaA, PC R & D Ionic Liquids, Frankfurter Str. 250, 64271,
Darmstadt, Germany
Scheme 1 Chlorophosphoramidite preparation: i) Hu¨nigs base (1 eq),
3-hydroxypropionitrile (1 eq), IL, 30 min. ii) Either Hu¨nigs base (1 eq),
R¢2NH (1 eq), 40 mins or R¢2NH (2 eq), 40 min.
† Electronic supplementary information (ESI) available: Reaction mix-
ture compositions for the systems illustrated in Figs. 1–3 and 31P NMR
chemical shifts. See DOI: 10.1039/b905000k
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Table 1 Relative product yields of 1 and 2 as a function of the
ionic liquids determined by 31P NMR integration and compared with
the reaction performed in diethyl ether for the reaction of PCl3 and
3-hydroxypropionitrile in the presence of 1 eq. Hu¨nigs base at room
temperature for 30 min
Results and discussion
Reaction of PCl3 and 3-hydroxypropionitrile
In order to maximise atom efficiency with respect to both the
conversion and yield of the product, stoichiometric ratios were
maintained throughout the experiments with respect to PCl3,
alcohol, amine, and ionic liquid. The results in ionic liquid
were compared with previously reported data for the prepa-
ration of chlorophosphoramidites in acetonitrile and diethyl
ether.12,13 [NTf2]- based ionic liquids were employed as the
reaction medium as they are known to provide a hydrolytically
stable environment for the PCl3 whilst maintaining a good
balance between solubility and an inertness of the anion.10
In many other ionic liquids, the anion is nucleophilic and
has been found to react with the PIII reagents leading to side
product formation thus limiting their use for this application.
Tris(perfluoroethyl)trifluorophosphate ([FAP]-) based ionic liq-
uids provide an alternative which does not lead to nucleophilic
displacement of chloride; however, PCl3 has a low solubility in
these ionic liquids again limiting their applicability. Therefore,
in the majority of this work, only the effect of the cation was
examined in detail.
The yields were determined from the 31P NMR peak integra-
tion ratio, where the percentage yield corresponds to the ratio
of the peak area of a given compound to the total of peak areas
of all phosphorus containing species in a given NMR spectrum.
This ratio corresponds to the molar speciation of the phosphorus
containing compounds in that sample.
As previously reported, the reaction between PCl3 and
an amine generally gives quantitative yields of the desired
PCl2NR2 product in short reaction times. The reaction of the
aminochlorophosphine with an alcohol provides a possible
route to the synthesis of chlorophosphoramidites. However,
on reaction with the alcohol, significant hydrolysis and side
product formation was observed. For example, the reaction
of 3-hydroxypropionitrile with PCl3 resulted in quantitative
conversion of the aminochlorophosphine but only ~5% of the
desired materials were formed. Therefore, in all subsequent
reactions reported, herein, PCl3 was first reacted with the alcohol
followed by the amine.
The initial step in the chlorophosphoramidite synthesis was
the reaction of PCl3 and 3-hydroxypropionitrile in the presence
of Hu¨nigs base (Scheme 1). In molecular solvents, this reaction
takes place at high dilution with external cooling of the
reaction mixture. In addition to the extensive treatment of
3-hydroxypropionitrile, PCl3 has to be distilled before use and
used in a large excess to encourage the reaction to go to com-
pletion. The Hu¨nigs base and the solvent of choice must also be
dried or distilled before use. In contrast, the ionic liquid mediated
reactions only require the ionic liquid to be dried under high
vacuum for two hours before use. In the ionic liquid reactions
the commercial reagents were used without pretreatment.
In the ionic liquids, PCl3 was first solubilised with the
subsequent addition of the Hu¨nigs base. The mixture is then
stirred vigorously for 5 min before the 3-hydroxypropionitrile
is added. While the reaction is exothermic, no cooling of the
system is required, and after 30 min an aliquot was removed
for 31P NMR analysis. Table 1 details the percentage yield
Composition (%)
Ionic liquida or
molecular solvent
PCl3 P(OR)Cl2 (1) P(OR)2Cl (2) H-phos
[C4mim][NTf2]
[C4dmim][NTf2]
[C4mpyrr][NTf2]
[C4mpip][NTf2]
[C4py][NTf2]
3
4
3
5
3
85
89
83
89
90
59
5
7
8
3
5
7
6
2
2
Diethyl etherb
a [C4mim]+ = 1-butyl-3-methylimidazolium, [C4dmim]+ = 1-butyl-2,3-
dimethylimidazolium, [C4mpyrr]+ = 1-butyl-1-methylpyrrolidinium,
[C4mpip]+ = 1-butyl-1-methylpiperidinium, [C4py]+ = 1-butylpyri-
dinium, [NTf2]- = bis{(trifluoromethyl)sulfonyl}imide. b Other products
not reported and the reaction was performed at 0 ◦C with an 6 mol%
excess of PCl3.13
distribution for all the ionic liquids used in this reaction. As
can be seen from the data, in all cases the P(OR)2Cl derivative
2 was formed before all the PCl3 had reacted. In addition, an
H-phosphonate derivative was also formed during the alcohol
addition. Interestingly, this formation was found to be reversible
for some systems as shown by the absence of the 31P NMR peaks
corresponding to derivatives of these side-products in the final
product composition after the subsequent amine addition.
In comparison with commonly utilised methods for the prepa-
ration of 1, the reactions in ionic liquids give excellent yields and
with good selectivity. For example using [C4dmim][NTf2] 89% of
1 was formed with no H-phosphonate contamination. Although
it is possible to isolate 1 from the ionic liquid at this stage by
distillation, addition of the nucleophilic amines was undertaken
directly into the ionic liquid reaction mixture without workup
to minimise the number of stages and to maximise yield. In
contrast, the optimised reactions in molecular solvents, such as
diethyl ether, require filtration of the amine hydrochloride salts
that are formed during the reaction from the reaction mixture
and isolation of the crude material, 1, by evaporation of the
solvent before further reaction with the amine. High reagent
dilution is required in the molecular solvent to limit the effect of
the salt waste on the conversion of PCl3. Owing to the sensitive
nature of 1, careful work-up is also needed to reduce product
hydrolysis.
The formation of chlorophosphoramidites in molecular sol-
vents requires 1 to be dissolved in a large volume of an-
hydrous acetonitrile (or hexane) followed by the addition of
freshly distilled nucleophilic amine in anhydrous acetonitrile
(or hexane). In general, for most reactions diisopropylamine
is used as the product formed displays a good balance between
stability and reactivity. For example, using acetonitrile as the
molecular solvent, the reaction of 1 with the nucleophilic
amines is performed at -20 ◦C for 1.5 h and then at room
temperature for a further 20 h resulting in the respective
chlorophosphoramidites being formed in yields ranging from
50–85%.12 Again the amine hydrochloride salt is filtered from
the reaction mixture and the solvent concentrated to give the
product 3, with purification generally performed by distillation.
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Careful storage of the material at low temperatures and under
an argon atmosphere is required to prevent hydrolysis. For
some materials, in particular the morpholino and diethylamino
derivatives, storage is not a viable option and they must be used
directly. This may be contrasted with the reactions performed
in the ionic liquid wherein amine (1 eq) and Hu¨nigs base
(1 eq) or amine (2 eq) was added to the ionic liquid reaction
mixture without any workup following the formation of 1. It
should be noted that the amine hydrochloride salt is soluble
in the ionic liquid allowing high concentrations of reagents
to be used. In contrast, in the molecular solvents the slurry
formed limits the concentration able to be utilised due to mass
transfer limitations which reduces the reaction rate. Figs. 1–3
show the percentage yield distribution for the reaction mixtures
as determined by 31P NMR analysis of samples taken 40 min
after amine addition to the ionic liquid reaction mixture without
any workup following the formation of 1 for diisopropylamine,
morpholine and diethylamine, respectively.†
Fig. 3 Percentage yield for the reation of 1 with diethylamine in [NTf2]-
based ionic liquids as a function of the cation with either a reaction ratio
of 1:1:1 or 1:2:0 PCl3:Amine:Hu¨nigs Base. H corresponds to hydrolysis.
The factors determining the selectivities observed are a
complex combination of the nucleophilicity of the amine and the
alcohol as well as the pKa of the amine and the base in the ionic
liquid solvent and these determine the equilibrium constants
for the reaction matrix shown in Scheme 1. The underlying
reason for the enhanced chemoselectivity in the ionic liquids
compared with the molecular solvents is not clear. It is possible
that the ionic liquid provides a medium whereby the nucleophilic
molecules and the PCl3 become well dispersed from other like
molecules, as found in the case of the water, and, therefore, this
limits the chance of multiple nucleophilic attack on the PCl3 by
amine, for example. This is aided by the higher viscosity of the
ionic liquid which reduces the diffusion coefficient and slows
intermixing of reagents. Although the mechanism is not well
understood to date, it is clear that the predominant product
formed in all cases is the desired P(OR)(NR¢2)Cl material
irrespective of the ionic liquid used, the nature of the amine
and whether or not Hu¨nigs base is present. However, the order
of addition of reagents was found to be critical in optimising the
formation of 3.
Fig. 1 Percentage yield for the reaction 1 with diispropylamine in
[NTf2]- based ionic liquids as a function of the cation with either a
reaction ratio of 1:1:1 or 1:2:0 PCl3:Amine:Hu¨nigs Base. H corresponds
to hydrolysis.
In general, reactions using 2 equivalents of amine give higher
yields of the desired products 3a-c than the corresponding
reaction involving 1 equivalent of both Hu¨nigs base and amine.
Interestingly, in some cases the dialkoxy product (P(OR)2(NR¢2),
4a-c) is formed but in a much higher yield than would be expected
based on the product from the first reaction step (Table 1)
indicating that the entire system is under equilibrium. For the
diethylamine set of products, the 1:1:0 conditions generally
formed P(NR¢2)2Cl derivatives in the product mixture, whereas
the 1:2:0 reagent ratio lead to P(NR¢2)Cl2 type species. This
trend was not observed for the other amine systems with the
P(NR¢2)2Cl derivative not being observed for diisopropylamine.
This difference may be associated with the increased steric
hindrance of the amine used. In addition, P(OR)2(NR¢2) type
species was not observed using morpholine, and the formation
of side products for this amine show no significant trend between
reaction conditions or ionic liquid employed.
Fig. 2 Percentage yield for the reaction of 1 with morpholine in [NTf2]-
based ionic liquids as a function of the cation with either a reaction ratio
of 1:1:1 or 1:2:0 PCl3:Amine:Hu¨nigs Base. H corresponds to hydrolysis.
Of the three products formed, diisopropylamino derivative is
commercially available, but the quality of the product can vary.
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The other two derivatives are too unstable to store, and as such
are not available to purchase.
an inert atmosphere. Using the stability provided by the ionic
liquid, a simple phosphitylation reaction was undertaken with
the phosphoramidite and a protected (Scheme 2).
Isolation and characterisation
Two main methods were employed for the isolation of 3 from the
ionic liquid mixture, namely extraction and distillation. Solvent
extraction proved to be a useful method of isolating 3a from
the reaction mixture. Using diethyl ether the product could be
isolated; however, some hydrolysis did occur on removal from
the ionic liquid. Unfortunately it was not possible to isolate the
derivatives 3b and 3c using this methodology. Both compounds
are highly sensitive to hydrolysis, and once removed from the
ionic liquid, significant hydrolysis occurred and little product
remained intact.
Distillation proved to be a more useful method for the isola-
tion of these sensitive materials with both the diisopropylamino
and diethylamino compounds successfully removed from the
reaction mixture cleanly without decomposition of the ionic
liquid. The diisopropyl derivative 3a was isolated; however, as
with solvent extraction, the diethyl derivative 3c decomposed
immediately due to its high sensitivity. This could be overcome
by distillation of the product material directly into the reaction
flask, or into fresh ionic liquid for storage purposes until
required. Using distillation, it was also possible to distil the
chlorophosphoramidites into alternative ionic liquids, namely
the tris(perfluoroethyl)trifluorophosphate ([FAP]-) based ionic
liquids. Ionic liquids based on this anion are even more
hydrophobic than those based on the [NTf2]- anion and this
media provided a more stable environment for these sensitive
reagents. As described above, due to the insolubility of PCl3 in
[FAP]- based ionic liquids, it was not possible to undertake the
full synthetic procedure in this media. Due to the high thermal
instability of the morpholino derivative, 3b, distillation was not
possible. However, preliminary investigations into the use of
supercritical CO2 extraction have shown promise for this and the
other systems and are currently being optimised. In addition, the
use of thin film distillation could also improve upon the isolated
yields of the distillable materials.
Scheme 2 Phosphitylation reaction of protected 2-deoxyadenosine in
DCM with 3a in [C6mim][FAP] in the presence of Hu¨nigs base.
Before use the nucleoside was azeotroped and dried under
high vacuum to reduce the potential for the co-crystallised
water to hinder the reaction; however, the Hu¨nigs base was
not distilled before use. The reaction was performed in anhy-
drous dichloromethane, as reactions in anhydrous acetonitrile
only resulted in deprotection of the trityl protecting group.
After 1.5 h at room temperature, quantitative conversion of
the chlorophosphoramidite to a diastereotopic mixture of
phosphitylated nucleosides with chemical shifts at 149 and
150 ppm by 31P NMR was found. After purification by column
chromatography the phosphitylated nucleotide was isolated in
92% yield, illustrating that the ionic liquid stabilised chlorophos-
phoramidite could be successfully employed for the synthesis of
nucleotides.
Conclusions
[NTf2]- based ionic liquids provide a media in which chlorophos-
phoramidites can be prepared without extensive purification
of the starting materials and external cooling of the reaction
mixture. The products were formed with high selectivity without
significant hydrolysis of the product. These selectivities are
difficult to achieve in molecular solvents and normally require
the use of excess reagents, derivatisation of the starting materials
and careful control of the reaction temperature and concentra-
tion. The ionic liquid reactions were also carried out at high
concentration, reducing the amount of solvent required when
compared with the analogous reaction in molecular solvent.
The chlorophosphoramidites were found to be readily dis-
tilled from the reaction mixture as isolated compounds or
into a stabilising clean ionic liquid. In particular, distillation
into [FAP]- based ionic liquids provided a medium in which
the reagents could be added directly to molecular solvents,
without isolation, providing a stable environment to protect
these sensitive materials before use. This has been utilised to
prepare phosphitylated nucleosides in quantitative yield without
isolation of the intermediate molecules.
Stability and phosphitylation
All reaction mixtures were monitored over an extended period
of time to establish the stability of the chlorophosphoramidites,
not only when distilled into fresh ionic liquids, but also within
the reaction mixtures. In agreement with the previous findings
for aminochlorophosphines,11 the chlorophosphoramidites once
formed were found to be stable with respect to hydrolysis
within the ionic liquid, illustrating that this class of solvent not
only represents an excellent medium for the formation of these
sensitive reagents but also offers exceptional storage qualities.
Furthermore, no decomposition of the chlorophosphoramidite
reagents was observed after being stored in the ionic liquid at
room temperature for several weeks even in the absence of an
inert atmosphere.
Nucleoside chemistry is an extremely important area of
research, but one in which the solvent choice is severely
compromised by poor solubility of the reagents. The solubility
of nucleoside and nucleic bases in ionic liquids have already been
reported,14–16 Using the methodology developed herein, there is
scope for the complete preparation of a wide range nucleotides
in ionic liquids to be realised with high selectivity.
Further studies showed that the chlorophosphoramidite,
3a, dissolved in [C6mim][FAP] could be added to anhydrous
acetonitrile and 31P NMR indicated no degradation of material
over 72 h in acetonitrile at room temperature in the absence of
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added or Hu¨nigs base (1 eq) and nucleophilic amine (1 eq).
Reactions were generally complete after a further 40 min.
Experimental
General procedures
(2 - Cyanoethoxy) - N,N - diisopropylamino - chlorophosphora -
midite 3a. Distilled from [C4dmim][NTf2] at 88–90 ◦C at
0.01 mmHg, or extracted with diethyl ether to give a colourless
liquid (46%). 1H NMR (300 MHz, CDCl3) 1.26 (12H, d, J 6.8 Hz,
4 CH3), 2.73 (2H, t, J 6.25 Hz, OCH2CH2CN), 3.68–3.86 (2H,
m, 2 CH), 4.05 (2H, dt, J 8.17, 6.25 Hz, OCH2CH2CN). 13C
NMR (75 MHz, CDCl3) 19.7 (CH3), 46.5 (d, CH), 47.8 (CH2),
60.8 (d, CH2), 117.3 (CN). 31P NMR (121 MHz, CDCl3) 181
(P(OR)(NiPr2)Cl).
The product distribution of the reactions of PCl3 for
the synthesis of chlorophosphoramidites was examined
in situ by 31P NMR and 1H-31P coupled NMR. Large
scale reactions were carried out in order to establish a
protocol for product isolation and to examine the product
stability in ionic liquids. Five sets of parallel experiments
were performed using 1-butyl-3-methylimidazolium bis-
{(trifluoromethyl)sulfonyl}imide ([C4mim][NTf2]), 1-butyl-2,3-
dimethylimidazolium bis{(trifluoromethyl)sulfonyl}imide ([C4-
dmim][NTf2]), 1-butyl-3-methylpyrrolidinium bis{(trifluoro-
methyl)sulfonyl}imide ([C4mpyrr][NTf2]), 1-butyl-3-methyl-
piperidinium bis{(trifluoromethyl)sulfonyl}imide ([C4mpip]-
[NTf2]), and 1-butyl-pyridinium bis{(trifluoromethyl)sul-
fonyl}imide ([C4py][NTf2]). The ionic liquids used were
prepared in house using standard literature methods17 from
the appropiate halide salt. 1-hexyl-1-methyl-imidazolium
tris(pentafluoroethyl)trifluorophosphate ([C6mim][FAP]) was
supplied by Merck KGaA. All ionic liquids were dried under
high vacuum for 2 h prior to use. The water content and
bromide content were measured for each ionic liquid using Karl
Fischer titration and ion chromatography, respectively. In each
case the bromide levels were below 5 ppm and the water content
for the dried ionic liquids were < 0.04 wt%.
(2-Cyanoethoxy)-N-morpholino-chlorophosphoramidite 3b.
It was not possible to distil the morpholino derivative due
to thermal instability or extract with molecular solvent due
high hydrolytic instability. Selected data from crude reaction
mixture: 13C NMR (75 MHz, CDCl3) 16.7 (CH2), 58.6 (d, CH2),
66.1 (N(CH)2), 71.2 (CH2), 118.1 (CN). 31P NMR (121 MHz,
CDCl3) 168.6 (P(OR)(NiPr2)Cl).
(2-Cyanoethoxy)-N,N -diethylamino-chlorophosphoramidite
3c. Distilled from [C4dmim][NTf2] at 90–92 ◦C at 0.01 mmHg,
into fresh [C4dmim][NTf2] (44%). Data from mixture of ionic
liquid and product. 1H NMR (300 MHz, CDCl3) 1.12 (6H, t, J
9.0 Hz, 2 CH3), 2.72 (2H, t, J 6.0 Hz, OCH2CH2CN), 3.09–3.16
(4H, m, 2 NCH2), 4.03 (2H, dt, J 8.0, 6.0 Hz, OCH2CH2CN). 13
C
NMR (75 MHz, CDCl3) 18.2 (CH3), 39.8 (d, J 18.5 Hz, CH2),
42.5 (CH2), 61.5 (d, J 17.5 Hz, NCH2), 118.3 (CN). 31P NMR
(121 MHz, CDCl3) 176.9 (P(OR)(NEt2)Cl).
For each reaction, the mole ratio of PCl3 (Aldrich, 98%), the
nucleophilic amine and the base, diisopropylethylamine (Hu¨nigs
base) (Aldrich, 99%) were varied. The mole ratio of PCl3 and
3-hydroxypropionitrile (Aldrich, 99%) was kept constant. The
amines investigated were diisopropylamine, morpholine and
diethylamine and were obtained from Aldrich. Diisopropy-
lamine was distilled over calcium hydride prior to use. PCl3,
3-hydroxypropionitrile, Hu¨nigs base and the remaining amines
were used as supplied.
General procedure for nucleoside phosphitylation18
Protected 2-deoxyadenosine, 8, (1 eq) was azeotroped with
toluene (3 times) and dried under high vacuum for 3 h before
use. The nucleoside was taken up in anhydrous DCM and 2 eq of
Hu¨nigs base was added. The solution was stirred for 10 min then
chlorophosphoramidite, 3a, in [C6mim][FAP] (2.5 eq) was added
to the reaction mixture. After 45 min, the 31P NMR showed
peaks at 149 and 150 ppm. After a further 1 h the mixture was
concentrated in vacuo and purified by column chromatography
(1:1–0:1 hexane:ethyl acetate, 1% NEt3) to yield the protected
nucleoside, 8, as a white solid (92%, 2:1 diastereotopic mixture).
Spectroscopic details
All the 31P, 1H-31P nuclear magnetic resonance spectra were
◦
recorded on a Bruker Bruker Avance 300 or 500 at 25 C. For
ionic liquid samples an aliquot was transferred directly into
the NMR tube with no addition of deuterated solvents. The
31P-NMR chemical shifts were recorded in parts per million
(ppm) relative to an external probe (sealed capillary inside
the NMR tube sample) of triethylphosphonate (PO(OEt)3) in
CDCl3 (solvent used for locking/shimming optimisation). The
PO(OEt)3 probe was referenced to 0.2 ppm. For the nucleotide
the NMR was recorded in CDCl3 referenced to 0.00 ppm using
N6-Benzoyl-5¢-O-(4,4¢-dimethoxytrityl)-2¢-deoxyadenosine-
3¢-O-[O-(2-cyanoethyl)-N,N¢-diisopropylphosphoramidite
9.
1H NMR (500 MHz, CDCl3) 1.12–1.22 (12H, m, 4 CH3),
2.48 (2H, t, J 6.4 Hz, OCH2CH2CN-a), 2.63 (2H, t, J 6.3 Hz,
OCH2CH2CN-b), 2.75–2.78 (1H, m, H-2¢), 3.31–3.3.63 (7H, m,
H-5¢, NCH, OCH2CH2CN), 3.76 (3H, s, OCH3-a), 3.78 (3H, s,
OCH3-b), 4.28–4.35 (1H, m, H-4¢), 4.76–4.82 (1H, m, H-3¢),
6.49–6.55 (1H, m, H-1¢), 6.76–6.82 (4H, m, ArCH), 7.17–7.30
(6H, m, ArCH), 7.38–7.7.41 (2H, m, ArCH), 7.51–7.62 (4H, m,
ArCH), 8.02 (2H, d, J 12.0 Hz, ArCH), 8.20 (1H, s, H-2-a), 8.22
(1H, s, H-2-b), 8.75 (1H, s, H-8-a), 8.76 (1H, s, H-8-b), 8.97
(1H, br s, NH). 13C NMR (125 MHz, CDCl3) 22.9–23.1 (CH3),
24.66–24.68 (CH2), 39.6 (NCH), 43.3–43.3 (C-2¢), 50.37 (CH2),
55.28 (OCH3), 58.1 (C-4¢), 63.4 (C-5¢), 77.2 (C-3¢), 84.7 (C-1¢),
86.5 (C), 113.1 (ArCH), 117.5 (CN), 122.0–132.8 (ArCH),
135.6, 135.7 (ArC), 141.8 (C-8), 144.4, 149.4 (ArC), 152.6
TMS for the 1H NMR and 77.00 ppm using CDCl3 for the 13
NMR.
C
General experimental conditions
To a stirred solution of PCl3 (1 eq) in dried ionic liquid
(1 eq) under an atmosphere of argon was added Hu¨nigs base
(1 eq). The solution was stirred vigorously for 5 min and
3-hydroxypropionitrile (1 eq) was added. The reaction mixture
was stirred for 30 min then either nucleophilic amine (2 eq) was
(C-2), 158.5 (C O). 31P NMR (121 MHz, CDCl3) 149.9, 150.0.
=
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HRMS (ES, M + H+) calculated for C47H53N7O7P 858.3744,
found 858.3756.
7 R. Gukathasan, M. Massoudipour, I. Gupta, A. Chowdhury, S.
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1396 | Green Chem., 2009, 11, 1391–1396
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