New functionalised silicas for highly selective cation exchange SPE purification in medicinal chemistry

Article abstract of DOI:10.1016/j.tetlet.2008.05.087

Functionalised silicas 2 and 3 are effective for the cation exchange SPE purification of basic molecules containing acid-sensitive functionalities, the selective separations of mixtures of basic compounds and accelerated reaction work-ups/product isolatio

Full text of DOI:10.1016/j.tetlet.2008.05.087

Tetrahedron Letters 49 (2008) 4968–4971
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
New functionalised silicas for highly selective cation exchange SPE
purification in medicinal chemistry
Jane Brown b, Alessandra Chighine a, Marie A. Colucci b, Nicola Galaffu a, Simon C. Hirst b,
b
a
Helen M. Seymour a, Jason J. Shiers b, Robin D. Wilkes a, , Jonathan G. Williams , John R. H. Wilson
*
a PhosphonicSTM Ltd, 114 Milton Park, Abingdon, Oxfordshire OX14 4SA, United Kingdom
b Sygnature Chemical Services Ltd, BioCity, Pennyfoot Street, Nottingham NG1 1GF, United Kingdom
a r t i c l e i n f o
a b s t r a c t
Article history:
Functionalised silicas 2 and 3 are effective for the cation exchange SPE purification of basic molecules
containing acid-sensitive functionalities, the selective separations of mixtures of basic compounds and
accelerated reaction work-ups/product isolations.
Received 22 April 2008
Revised 9 May 2008
Accepted 19 May 2008
Available online 23 May 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Solid phase extraction (SPE)
Cation exchange
SCX
Amine separation
Purification
Functionalised silica
Immobilised acid
Recent years have seen a continuing drive for increased produc-
tivity from Medicinal Chemistry groups as one of many initiatives
to optimise the chances of new medicines emerging from pharma-
ceutical company pipelines. New technologies need to be adopted
to overcome bottlenecks within the drug discovery process, the
most significant of which for medicinal chemists is compound
purification, which can be tackled by specialist purification groups
or more directly by the use of a number of continually emerging
easy-to-use laboratory purification techniques.
Solid phase extraction (SPE) has rapidly become an essential
tool in Discovery Medicinal Chemistry,1 offering considerable
operational benefits and time savings over traditional extractive
work-up and chromatographic methods. Whilst utilised initially
for high-throughput applications in lead discovery, this simple
technique has now become widely-used for the purification of both
high value single compounds and small focused compound arrays
at both hit-to-lead and lead optimisation project stages. It is a tech-
nique ideally and equally suited to purifying milligrams through to
multiple grams of product.
away, before the desired basic product is eluted (‘catch and
release’). Product purification can also be performed by the reten-
tion of basic impurities by the SCX material, with the non-retained,
non-basic reaction product washed off and collected.
Products can be typically isolated in purities of >90%, generally
rendering additional purification unnecessary before further syn-
thetic steps. For screening compounds, SCX can deliver compounds
of sufficiently high purity (97–>99%) for direct submission to
screens; however, if the compound purity specification is not
reached, the technique will generally have cleaned up the reaction
mixture sufficiently to greatly facilitate additional purification by
other techniques such as preparative HPLC.
Cation exchange materials based on both polystyrene2 and
silica are commercially available in easy-to-use cartridge formats.
Silica adsorbents show excellent stability and flexibility to a wide
range of both organic and aqueous solvents and do not swell; as
such silicas have generally become the materials of choice for
Medicinal Chemistry.
Commercially available cation exchange materials typically fea-
ture sulfonic acid functionalities attached to the support. Aryl sul-
fonic acids (pKa <1) are slightly more acidic than the alkyl variants
(pKa ꢀ 1). Alkyl carboxylic acids (pKa 4–5) are described as weak
cation exchangers, though these materials have received little
attention. Materials of this latter type add a degree of selectivity
to the existing SCX technology, and our paper elaborates this
Strong cation exchange (SCX) is the most widely used method
of SPE purification. Basic reaction products are typically retained
by the SCX material, allowing non-basic impurities to be washed
* Corresponding author. Tel.: +44 01235 834466; fax: +44 01235 835566.
E-mail address: robin.wilkes@phosphonics.com (R. D. Wilkes).
0040-4039/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2008.05.087

J. Brown et al. / Tetrahedron Letters 49 (2008) 4968–4971
4969
aspect of cation exchange methodology with the introduction and
application of new immobilised acids (Fig. 1).
variety of key industrial problems, including scavenging of metal
and organic impurities from active pharmaceutical ingredients
(APIs)9 and as heterogeneous catalysts for chemical processes
including oxidations and metal-mediated cross couplings.10
Figure 1 shows a selection of immobilised acids which were
prepared using this approach, and which feature sulfonic acid
1,11 phosphonic acid 2 and carboxylic acid functionalities, leading
to coverage of a pKa range from approximately 1–6. For this Letter,
our efforts were focused on the use of immobilised phosphonic 2
and mercaptosuccinic 3 acids, for cationic exchange purifications.
Commonly used protecting groups include TBDMS and tert-
butyl (tBu) for alcohols and phenols, and Boc for amines. Attempts
to purify basic molecules containing these groups with immo-
bilised sulfonic acids can result in partial or complete removal of
the protecting group. Phosphonic acid 2 and mercaptosuccinic acid
3, are however, effective for the catch and release of molecules
such as those shown in Figure 2, all of which were caught and then
released without any protecting group loss. Molecules containing
tetrahydropyran (THP) groups have also been eluted through 3
without protecting group removal, allowing non-retained purifica-
tion operations to be considered.
Limitations of existing cation exchange technology include the
strongly acidic nature of the sulfonic acids, which although respon-
sible for the key ionic exchange with the basic products or reaction
components to be retained, also have the effect of limiting the
range of molecules which can be purified effectively—the presence
of any acid-sensitive functionalities renders the technology unsuit-
able. In fact, SCX adsorbents are effective for the removal of tert-
butoxycarbonyl (Boc) groups from a diverse range of protected
amines3 and alkyl/aryl sulfamides,4 and tert-butyldimethylsilyl
(TBDMS) groups from protected 1° and 2° alkyl ethers.5
Used in catalytic amounts, alkyl sulfonic acid functionalised sil-
ica has also been reported as a general method for the preparation
of Boc-protected derivatives of amines, amino-esters and sulfona-
mides.6 Sulfonic acid functionalised materials are also capable of
catalysing additional reactions, such as the formation of pyrazoles
from enol ketones and substituted hydrazines.7 The potential for
the sulfonic acid functionality present on SCX adsorbents to cleave
acid-sensitive moieties and catalyse additional reactions has led to
small compound arrays, involving purification by this method,
being relatively limited in terms of additional functionalities.7,8
Existing SCX materials can also fail to differentiate adequately
between different basicities of reaction components, for example,
molecules containing a 2° amine and a 3° amine. Such mixtures
are commonplace after a variety of standard synthetic steps
including amine deprotections and reductive alkylation reactions.
We have developed the addition of free radicals to vinyl trialk-
oxysilanes to yield highly functionalised trialkoxysilanes, which
are grafted onto existing silica frameworks, as depicted in Scheme
1, to provide novel functionalised materials of use for solving a
t
Amide formation, featuring Boc and Bu protecting groups on
the amino and phenolic groups, respectively, proceeded smoothly
with N-methylpiperazine as shown in Scheme 2. Use of excess ami-
no acid 4 ensured complete consumption of the N-methylpipera-
zine and cation exchange treatment of the reaction mixture, in
this case with 2, resulted in only the desired amide product 5 being
retained, with the excess amino acid, HOBt and N,N0-diisopropyl-
urea by-product from the DIC coupling agent all being washed
off with methanol. Piperazine amide 5 was eluted from the immo-
bilised acid cleanly, in high yield and without loss of either of the
O
O
O
S
P
S
S
OH
OH
OH
OH
OH
OH
S
O
O
O
2
3
1
= silica
Increased Acidity
Lower pKa
Figure 1.
RO
RO
Si
FG
OR
FG
RO
X
Silica
Initiator
X
(
)
Si
m
(
)
m
+
RO
toluene
OR
FG
HX
(
)
m
For 2, X(CH2)mFG = P(O)(OH)2
= silica
(prepared from HP(O)(OMe)2 with subsequent ester hydrolysis)
For 3, X(CH2)mFG = SCH(CO2H)CH2CO2H
FG = single or multiple functional group(s)
Scheme 1.
O
HN
O
HN
N
H
N
HO
O
NH2
Si
O
O
O
Si
H2N
CO2H
NH2
O
O
2
2
3
3
3
Purified using...
Figure 2.

4970
J. Brown et al. / Tetrahedron Letters 49 (2008) 4968–4971
O
4
O
O
P
O
O
OH
N
H
OH
OH
(
xs.)
O
Caught
N
N
2
O
O
NH
N
DIC, HOBt (1.5 eq.)
DCM
0.7 M NH3
in MeOH
Released
N
H
O
5
Washed off
96% yield
98% purity
xs. Acid
DCU Urea By-Product
HOBt
Scheme 2.
protecting groups, by use of ca. 0.7 M ammonia in methanol solu-
tion.12 Treatment of the amide product with immobilised sulfonic
acids resulted in partial loss of both protecting groups almost
immediately and complete removal of both protecting groups
upon shaking overnight.
amounts of the unwanted phenethyl pyridine 7. Further treatment
of the mixture was resisted due to the potential for reduction of the
phenyl ring. Sulfonic acid-based SCX resulted in both products
co-eluting, and no separation could be achieved.
Treatment of the R = H product mixture (from 6b) with 2
resulted in both basic compounds being retained; however, the
less basic pyridine product 7 was eluted selectively with triethyl-
amine in methanol, allowing the more basic piperidine product 8
to be eluted cleanly with ammonia in methanol. With acid 6a,
use of 3 resulted solely in the more basic piperidine product 8
being retained, with the pyridine product 7 not retained but
instead washed through with methanol. Piperidine 8 was eluted
with ammonia in methanol solution. A 1:1 mixture of the two reac-
tion products was used in these studies, and compound recoveries
and purities were >95%.
Preliminary screening of a range of basic compounds, including
heterocycles and 1° and 2° amines, covering a pKa range of 5–11,
revealed that 2 retained all compounds in the range, including pyr-
idines and anilines. At lower pKa, basic compound release from the
cation exchange material was possible with ca. 0.35 M triethyl-
amine in methanol solution,13 whereas ca. 0.7 M ammonia in
methanol solution was required as the pKa of the retained basic
compound increased beyond 9. Mercaptosuccinic acid 3 failed to
catch basic compounds including anilines and pyridines of lower
pKa (5–6), but basic compounds above pKa 7–8 were retained. As
with 2, as the pKa of the caught basic component moved from low-
er to higher pKa, the use of ammonia in methanol12 was required
for adequate compound elution rather than triethylamine in meth-
anol.13 We hoped to exploit these observations to perform selec-
tive separations of basic compounds and report here some of our
initial results. A full investigation of these functionalised silicas
for the separation of basic molecules will be reported in due
course.
The separation of some 2° and 3° amine mixtures is possible
using these cationic exchange materials of reduced acidity. N-Benz-
ylaniline was readily separated from N-benzylethanolamine using
3. N-Benzylaniline was not retained by 3 but was washed off
within the methanolic loading solution, whereas N-benzylethanol-
amine was retained and only subsequently eluted with triethyl-
amine in methanol solution. A weight recovery of >95% for the
caught and separated compound was achieved. Acids 2 and 3 can
also be used for the separation of basic product mixtures including
cyclic 2° and 3° amines, and saturated and unsaturated
heterocycles.
Deprotection of cyclic amines is another widely used and
generally reliable hydrogenative synthetic step. Access to the
synthetically-attractive 3-hydroxy-3-aryl azetidine building block
10 shown in Scheme 4 is best achieved through formation of the
corresponding benzhydryl-protected azetidine
9 followed by
hydrogenation of the amino protecting group. Unfortunately, the
deprotection step sometimes failed to go to completion resulting
in a mixture of 2° and 3° amines, which are extremely difficult to
separate by conventional column chromatography or with SCX
sulfonic acids.
Treatment of the mixture with either 2 or 3 allowed clean and
high-yielding separation of the 2° and 3° amines. Both basic
materials were caught by 2, from which selective elution could
again be achieved, using first triethylamine in methanol to release
exclusively the starting material 9 followed by ammonia in meth-
anol solution to release the desired 2° amine reaction product 10.
Both compounds were isolated in >95% purity.
These functionalised silicas are also useful to simplify reaction
work-ups and allow quicker access to purified products. For exam-
ple, the synthesis of 2-phenyl-3-aminopyridine 12 is reported to
proceed most efficiently from 2-chloro-3-aminopyridine by the
use of an imine protecting group, which facilitates the desired
Suzuki coupling step before being removed.14 Product work-up
Hydrogenation of the central olefin along with the pyridyl moi-
ety of the phenyl vinyl pyridine substrates 6a,b shown in Scheme 3
did not always proceed to completion to give the desired phen-
ethyl piperidine 8, and this was often accompanied by significant
N
N
NH
H2, Pd/C
+
EtOH
7
8
6a,b
R
R
R
Silica 3 :
Not retained
MeOH
Retained
~1% NH3/MeOH
R = CO2H
6a
-
eluted with
Silica 2 :
eluted with
Retained
5% Et3N/MeOH
Retained
~1% NH3/MeOH
6b R = H
-
Scheme 3.

J. Brown et al. / Tetrahedron Letters 49 (2008) 4968–4971
4971
F
F
HO
Ph
HO
H2, Pd/C
9
+
N
N
H
Ph
9
9
10
Silica 3 :
eluted with
Not retained
MeOH
Retained
5% Et3N/MeOH
-
Silica 2 :
eluted with
Retained
5% Et3N/MeOH
Retained
~1% NH3/MeOH
-
Scheme 4.
Ph
N
O
P
OH
OH
NH2
NH2
Cl
PhCHO (xs), PhB(OH)2 (xs
)
2
Caught
PdCl2(PPh3)2
0.7 M NH3
in MeOH
Released
N
N
N
Na2CO3, toluene
11
12
Washed off
impurities
60% yield
98% purity
Scheme 5.
3. Liu, Y-S.; Zhao, C.; Bergbreiter, D. E.; Romo, D. J. Org. Chem. 1998, 63, 3471.
4. Ghassemi, S.; Fuchs, K. Mol. Divers. 2005, 9, 295.
5. Karimi, B.; Zareyee, D. Tetrahedron Lett. 2005, 46, 4661.
6. Das, B.; Venkateswarlu, K.; Krishnaiah, M.; Holla, H. Tetrahedron Lett. 2006, 47,
7551.
7. Humphries, P. S.; Finefield, J. M. Tetrahedron Lett. 2006, 47, 2443.
8. Siegel, M. G.; Hahn, P. J.; Dressman, B. A.; Fritz, J. E.; Grunwell, J. R.; Kaldor, S. W.
Tetrahedron Lett. 1997, 38, 3357; Wang, Y.; Sarris, K.; Sauer, D. R.; Djuric, S. W.
Tetrahedron Lett. 2007, 48, 5181.
9. Galaffu, N.; Man, S.; Wilkes, R. D.; Wilson, J. R. H. Org. Process Res. Dev. 2007, 11,
406.
10. Al-Hashimi, M.; Fisset, E.; Sullivan, A. C.; Wilson, J. R. H. Tetrahedron Lett. 2006,
47, 8017 and references cited therein; Al-Hashimi, M.; Sullivan, A. C.; Wilson, J.
R. H. J. Mol. Catal. A: Chem. 2007, 273, 298; Al-Hashimi, M.; Qazi, A.; Sullivan, A.
C.; Wilson, J. R. H. J. Mol. Catal. A: Chem. 2007, 278, 160.
was achieved by extractive aqueous work-up, involving initial
imine 11 cleavage with aqueous hydrochloric acid, followed in
some cases by a crystallisation step. We have found that this inten-
sive work-up protocol can be replaced by simply loading the crude
Suzuki reaction product onto 2, as shown in Scheme 5, resulting in
the imine 11 being retained, purified, deprotected and finally
released as the desired 1° aminopyridine reaction product 12 using
ammonia in methanol solution as eluant. The product was isolated
in a promising, unoptimised yield of 60% and in high purity, with
no additional purification steps required.
A typical protocol for reaction purification using these function-
alised cation exchange silicas is provided.15
11. Bugrayev, A.; Al-Haq, N.; Okopie, R. A.; Qazi, A.; Suggate, M.; Sullivan, A. C.;
Wilson, J. R. H. J. Mol. Catal. A: Chem. 2008, 280, 96.
In conclusion, we have shown the potential of functionalised
silicas with modified acidities,16 in comparison to the tradition-
ally-employed sulfonic acid materials, for catch and release SPE
purifications. These materials allow acid-sensitive functionalities
such as Boc, TBDMS and OtBu to be retained during the purification
of basic compounds. Use of the materials for the separation of basic
products such as 2° and 3° amines and saturated and unsaturated
nitrogen heterocycles has also been demonstrated, and further
elaboration of this work is ongoing.
12. Commercially available 7 N ammonia in methanol solution (Aldrich) was
diluted ꢁ10 with methanol (ꢀ1% v/v solution).
13. Triethylamine (5 mL) was added to methanol (95 mL) to give a 5% v/v solution
(ꢀ0.35 M).
14. Caron, S.; Massett, S. S.; Bogle, D. E.; Castaldi, M. J.; Braish, T. F. Org. Process Res.
Dev. 2001, 5, 254.
15. Typical purification procedure using functionalised silicas for cation exchange: The
appropriate SPE cartridge containing 1 g of functionalised silica was
conditioned by passing 10 mL of MeOH through it, either under gravity or
with positive pressure or under controlled vacuum. The reaction mixture was
next loaded onto the top of the cartridge in the solvent of choice (ꢀ1–2 mL
solvent/0.050 g product). Loading onto the cartridge should typically be 5% by
weight (reaction mixture: functionalised silica), though higher loadings are
sometimes possible. Once the reaction mixture has been eluted through, a
further 5 mL of methanol was passed through the cartridge. This ‘load’ fraction
may contain unretained reaction components. The cartridge was washed with
2 ꢁ 10 mL of MeOH and fractions were collected. These ‘wash’ fractions should
contain any further traces of unretained reaction components. Basic product
elution is dependent on the functionalised silica used. The cartridge was eluted
with either triethylamine in MeOH solution13 (5% v/v; ꢀ0.35 M, 10 mL) or
ammonia in MeOH solution12 (ꢀ1.2% v/v; ꢀ0.7 M, 10 mL), or both, and the
fractions collected. Fractions from these elution steps should contain any
retained reaction components. The cartridge was finally rinsed using 10 mL of
MeOH, which was also collected. Fractions were analysed to confirm that the
desired purification had been achieved. The desired fractions were
concentrated in vacuo to afford the purified product(s). In some cases,
solvent quantities and wash steps may be reduced as the user deems most
suitable for their particular purification process.
Supplementary data
Example HPLC traces for the separations of the basic mixtures
described are available. Supplementary data associated with this
article can be found, in the online version, at doi:10.1016/
j.tetlet.2008.05.087.
References and notes
1. Cork, D.; Hird, N. Drug Discovery Today 2002, 7, 56; Ripka, W. C.; Barker, G.;
Krakover, J. Drug Discovery Today 2001, 6, 471; Weinbrenner, S.; Tzschucke, C.
C. In Combinatorial Chemistry; Bannwarth, W., Hinzen, B., Eds.; Wiley-VCH:
Weinheim, 2006; Chapter 1; pp 1–30.
2. Salimi, H.; Rahimi, A.; Pourjavadi, A. Monatsh. Chem. 2007, 138, 363 and
references cited therein.
16. PhosphonicSTM cation exchange SPE materials POH1d
2 and STMAd 3 are
available loose as well as in easy-to-use, pre-packed polypropylene 1 g or 10 g
cartridges. Cartridges are also available for metal SPE purifications and organic
scavenging—see www.phosphonics.com for further information.