The use of Co-crystals for the determination of absolute stereochemistry: An alternative to salt formation

Article abstract of DOI:10.1021/jo102148p

Absolute stereochemistry of oils and viscous liquids can be difficult to determine. Co-crystallization involves generating a crystalline material consisting of more than one neutral compound. The combination of co-crystallization with both X-ray diffraction and chiral HPLC was particularly powerful in overcoming these difficulties for a series of chiral 3-arylbutanoic acids. Co-crystallization offers advantages over salt formation because co-crystals dissociate in solution, meaning identical HPLC conditions can be used for both the materials of interest and their co-crystals.

Full text of DOI:10.1021/jo102148p

pubs.acs.org/joc
the material, via either additional chemical transformations
The Use of Co-crystals for the Determination of
Absolute Stereochemistry: An Alternative to
Salt Formation
or salt formation.4 Both of these strategies, however, have
limitations. The use of chemical transformations can be
limited by the availability of only small quantities of enantio-
pure material, and furthermore, additional reaction steps may
affect the stereochemical integrity of the required compound.
For salt formation, in addition to the limitations mentioned
above, the compound also requires ionizable sites. For both
strategies, chiral HPLC analysis has to be developed not only
for the pure material but also for the derivatives, since in many
cases the conditions are nontransferrable.4
†
Kevin S. Eccles, Rebecca E. Deasy, Laszlo Fabian,
†
†
ꢀ
ꢀ ꢀ ꢀ
Anita R. Maguire,§ and Simon E. Lawrence*,†
†Department of Chemistry, Analytical and Biological
Chemistry Research Facility and §Department of Chemistry
and School of Pharmacy, Analytical and Biological Chemistry
Research Facility, University College Cork, Cork, Ireland
As part of an ongoing program of research into enantio-
selective biotransformations, we have been interested in
determining the absolute stereochemistry of a series of sub-
stituted 3-arylbutanoic acids 1a-d, obtained via enzymatic
hydrolysis of the corresponding ethyl ester (Figure 1).5 The
enantiopure products were isolated as liquids at room tem-
perature. The enantioselectivity of the enzyme-catalyzed
reactions was investigated by chiral HPLC analysis,5 and
appropriate conditions to separate and quantify the enan-
tiomers of 1a-d were identified. However, knowledge of
absolute stereochemistry was necessary to determine the
sense of enantioselection with each of the biocatalysts em-
ployed. Chemical transformations were not attempted, as
the enantiopure samples were available in limited quantities.
s.lawrence@ucc.ie
Received November 1, 2010
Absolute stereochemistry of oils and viscous liquids can
be difficult to determine. Co-crystallization involves gen-
erating a crystalline material consisting of more than one
neutral compound. The combination of co-crystallization
with both X-ray diffraction and chiral HPLC was parti-
cularly powerful in overcoming these difficulties for a
series of chiral 3-arylbutanoic acids. Co-crystallization
offers advantages over salt formation because co-crystals
dissociate in solution, meaning identical HPLC condi-
tions can be used for both the materials of interest and
their co-crystals.
FIGURE 1. The enzymatic biotransformation employed in this
work.5
Co-crystallization, involving crystallizing two (or more)
neutral molecules together in one crystalline material, has
recently garnered great interest as an alternative to salt
formation for improving the physical properties of active
Despite major advances in asymmetric synthesis over the
past 30 years,1 one of the major challenges that remains is
definitively assigning absolute stereochemistry,2 especially
with materials which are not readily crystalline. This is
increasingly important as new synthetic products are often
difficult to crystallize,3 and many can only be isolated as
viscous oils.
(4) McConnell, O.; Bach, A.; Balibar, C.; Byrne, N.; Cai, Y. X.; Carter,
G.; Chlenov, M.; Di, L.; Fan, K.; Goljer, I.; He, Y. N.; Herold, D.; Kagan,
M.; Kerns, E.; Koehn, F.; Kraml, C.; Marathias, V.; Marquez, B.;
McDonald, L.; Nogle, L.; Petucci, C.; Schlingmann, G.; Tawa, G.; Tischler,
M.; Williamson, T.; Sutherland, A.; Watts, W.; Young, M.; Zhang, M.-Y.;
Zhang, Y. R.; Zhou, D. H.; Ho, D. Chirality 2007, 19, 658–682.
(5) Deasy, R. E.; Brossat, M.; Moody, T. S.; Maguire, A. R. Tetrahedron
Asymmetry. Accepted.
The two main synthetic strategies which have been em-
ployed to circumvent these problems involve modification of
(1) (a) Bornscheuer, U. T.; Kazlauskas, R. J. Hydrolases in Organic
Synthesis: Regio- or Stereoselective Biotransformations, 2nd ed.; John Wiley
& Sons: Chichester, UK, 2006. (b) Koeller, K. M.; Wong, C. H. Nature 2001,
409, 232–240. (c) Theil, F. Chem. Rev. 1995, 95, 2203–2227.
(2) Allenmark, S.; Gawronski, J. Chirality 2008, 20, 606–608.
(3) Hursthouse, M. B.; Huth, L. S.; Threlfall, T. L. Org. Process Res. Dev.
2009, 13, 1231–1240.
(6) (a) Schultheiss, N.; Newman, A. Cryst. Growth Des. 2009, 9, 2950–
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2967. (b) Karki, S.; Friscic, T.; Fabian, L.; Laity, P. R.; Day, G. M.; Jones,
W. Adv. Mater. 2009, 21, 3905–3909. (c) Shan, N.; Zaworotko, M. J. Drug
Discovery Today 2008, 13, 440–446. (d) Blagden, N.; de Matas, M.; Gavan,
P. T.; York, P. Adv. Drug Delivery Rev. 2007, 59, 617–630. (e) Remenar, J. F.;
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Morissette, S. L.; Peterson, M. L.; Moulton, B.; MacPhee, J. M.; Guzman,
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H. R.; Almarsson, O. J. Am. Chem. Soc. 2003, 125, 8456–8457.
DOI: 10.1021/jo102148p
r
Published on Web 01/18/2011
J. Org. Chem. 2011, 76, 1159–1162 1159
2011 American Chemical Society

Eccles et al.
JOCNote
FIGURE 2. Co-crystallization of the 3-arylbutanoic acids with
isonicotinamide.
FIGURE 3. Hydrogen bonding interactions in (S)-2a. The different
colors indicate unique, symmetry-independent (S)-2a (green and
blue) and isonicotinamide (red and orange) molecules. The same
motif is formed in (S)-2c, (S)-2d, (()-2b, and (()-2c, see the
Supporting Information.
pharmaceutical ingredients, without detrimental effects on
the chemical properties.6 Therefore, we decided to investi-
gate if co-crystals involving the carboxylic acids 1a-d would
be suitable materials for determining their absolute stereo-
chemistry.
A search of the Cambridge Structural Database7 revealed
that monocarboxylic acids often co-crystallize with nicotin-
amide and isonicotinamide.8-10 Initial co-crystal screening
was performed by neat grinding with the racemic acid, and
isonicotinamide was identified by powder X-ray diffraction
(see the Supporting Information) as a suitable co-crystal
former for each of the acids (Figure 2).
Suitable conditions for growing single crystals from solu-
tion were identified by using the racemic acids and similar
conditions were applied to the enantiopure analogues. The
advantage of this approach was that once the conditions
were developed, a small sample of the enantiopure material
(<8 mg) was sufficient. All co-crystals, both enantiopure
and racemic, were grown by slow evaporation of acetonitrile:
acetone (70:30) solutions.
FIGURE 4. Hydrogen bond motif in (S)-2b. The same color
scheme is applied as for Figure 1.
ensure this was the case, powder X-ray diffraction, PXRD,
was performed on each of the ground materials 2a-d and
compared to the theoretical PXRD patterns calculated from
the single crystal data.
We anticipated that acid molecules, once dissolved, would
have the same properties irrespective of whether they are
obtained by dissolving the pure acid or the co-crystal, as co-
crystallization only affects the properties of the solid phase.
Thus, co-crystallization has the clear advantage that the
conditions developed to separate the enantiomers of the pure
compound by chiral HPLC can be applied directly for anal-
ysis of the co-crystal employed for X-ray crystallography,
enabling direct correlation with the chiral HPLC peak for the
enantiomer. The only potential concern is the impact of the
co-crystal former. To check for possible interference, chiral
HPLC of the racemic acids, in the presence of isonicotin-
amide, was recorded, clearly showing that chromatographic
behavior of 1a-d was unaffected, confirming the validity of
the methodology employed. Thus, chiral HPLC were re-
corded on the individual crystals used for the X-ray diffrac-
tion experiments. This enabled the use of chiral HPLC to
directly monitor the direction of enantioselection in the
enzyme-mediated resolutions, Figure 5.5 Thus, while (S)-1a
has been previously described in the literature,12 this is the
first time the absolute stereochemistry of this compound
has been definitively determined by X-ray diffraction. This
procedure was successfully employed to determine the abso-
lute stereochemistry of the novel enantiopure acids (S)-
1b-d.
Single crystal diffraction data, in combination with the
chromatographic experiments, allowed the unambiguous
assignment of absolute configuration.11 Each enantiopure
co-crystal was formed with the (S)-enantiomer of the acid.
Each of the co-crystals display a common set of intermole-
cular interactions (Figures 3 and 4) with amide-amide [N;
H
OdC] and acid-pyridine [COOH N] hydrogen
3 3 3
3 3 3
bonds linking the co-crystal components.
Interestingly, irrespective of the chiral or enantiopure
nature of the acid molecules, similar infinite one-dimensional
hydrogen-bonded ribbons are formed in all the co-crystals
except (()-2d, which exhibits two-dimensional puckered
sheets (see the Supporting Information). Thus, the overall
core hydrogen-bonded motif is retained across the struc-
tures, with the conformation of the acids varying. Presum-
ably, this conformational variation is necessary to maintain
both a close-packed arrangement of molecules with asym-
metric shapes and a (topologically) symmetrical hydrogen
bond network. Further details of the structural features
present in these co-crystals are in the Supporting Infor-
mation.
An important consideration was whether the diffraction
experiment was a true representation of the bulk sample. To
(7) Allen, F. H. Acta Crystallogr., Sect. B: Struct. Sci. 2002, 58, 380–388.
ꢀ
(8) Lemmerer, A.; Bathori, N. B.; Bourne, S. A. Acta Crystallogr., Sect.
B: Struct. Sci. 2008, 64, 780–790.
Co-crystallization involving achiral compounds with en-
antiopure and racemic partners is known; in particular
interest has centered on the different physical properties that
are obtained for the racemic and enantiopure co-crystals,
€
(9) (a) Aakeroy, C. B.; Beatty, A. M.; Helfrich, B. A. Angew. Chem., Int.
Ed. 2001, 40, 3240–3242. (b) Bhogala, B. R.; Basavoju, S.; Nangia, A.
CrystEngComm 2005, 7, 551–562.
€
(10) Aakeroy, C. B.; Beatty, A. M.; Helfrich, B. A. J. Am. Chem. Soc.
2002, 124, 14425–14432.
(11) (a) Flack, H. D.; Bernardinelli, G. Chirality 2008, 20, 681–690. (b)
Hooft, R. W. W.; Straver, L. H.; Spek, A. L. J. Appl. Crystallogr. 2008, 41,
96–103. (c) Hooft, R. W. W.; Straver, L. H.; Spek, A. L. J. Appl. Crystallogr.
2010, 41, 665–668.
(12) (a) Kanemasa, S.; Suenaga, H.; Onimura, K. J. Org. Chem. 1994, 59,
6949–6954. (b) Sun, X. F.; Zhou, L.; Wang, C.-J.; Zhang, X. M. Angew.
Chem., Int. Ed. 2007, 46, 2623–2626.
1160 J. Org. Chem. Vol. 76, No. 4, 2011

Eccles et al.
JOCNote
such as density and stability.8,13 It has been used to introduce
heavy atoms into a chiral structure, enabling absolute ste-
reochemistry determination of known materials with Mo KR
radiation.14 To date the stereochemistry of the starting
material has always been known.
Our results demonstrate for the first time that the applica-
tion of co-crystallization methodology can be utilized suc-
cessfully for determination of the absolute stereochemistry
of compounds that are oils or viscous liquids at ambient
conditions. The advantage of co-crystals over salt formation
is that the chromatographic conditions developed to sepa-
rate the enantiomers do not change on co-crystallization,
unless the co-crystal former interferes with the separation of
the enantiomers. Furthermore, co-crystals do not require
ionizable sites, which in principle means that co-crystalliza-
tion should be more universally applicable than salt forma-
tion. In conclusion, co-crystallization and X-ray diffraction,
in combination with chiral HPLC, provides an effective
method in synthetic research to identify absolute stereo-
chemistry in novel compounds.
Experimental Section
General Experimental. Compounds (()-1a, (()-1b, (()-1c,
(()-1d, (S)-1a, (S)-1b, (S)-1c, and (S)-1d were prepared accord-
ing to the literature methods.5 Grinding was undertaken with a
mixer mill equipped with stainless steel 5 mL grinding jars and
one 2.5 mm stainless steel grinding ball per jar. Powder
diffraction experiments were performed with graphite mono-
chromatized Cu KR radiation at room temperature. Single
crystal diffraction experiments were performed with either
graphite monochromatized Mo KR or doubly curved silicon
crystal monochromatized Cu KR radiation at 100 K. Analysis
was undertaken with the SHELX suite of programs15 and
diagrams prepared with Mercury 2.3.16 The Hooft y param-
eter11b,c was used to check the correct assignment of the
absolute stereochemistry of the enantiopure co-crystals. This
was done using PLATON.17 Differential Scanning Calorimetry
was undertaken over the temperature range 40-140 °C and the
sample was heated at a rate of 4 deg min-1. Chiral HPLC
analysis was undertaken with a chiral column (Chiralcel OJ-H
for 2a, 2b, and 2c; Chiralcel AS-H for 2d) at 0 °C.
General Procedure for the Preparation of the Racemic Acid:
Isonicotinamide Co-crystals. The corresponding racemic acid
(1 equiv) and isonicotinamide (1 equiv) were placed in the
grinding jars, and the mill was operated at 30 Hz for 30 min.
The material obtained was analyzed by PXRD. It was then
dissolved in a minimum amount of acetonitrile and acetone
was added until the solvent ratio was 70:30, respectively. The
solution was allowed to stand at ambient conditions and
crystals suitable for single crystal diffraction were obtained
by slow evaporation over 3-4 d. After single crystal analysis,
ꢁꢁ ꢀ
(13) (a) Friscic, T.; Jones, W. Faraday Discuss. 2007, 136, 167–178. (b)
ꢁꢁ ꢀ
ꢀ ꢀ
Friscic, T.; Fabian, L.; Burley, J. C.; Reid, D. G.; Duer, M. J.; Jones, W.
Chem. Commun. 2008, 1644–1646. (c) Chen, S.; Xi, H.; Henry, R. F.;
Marsden, I.; Zhang, G. G. Z. CrystEngComm 2010, 12, 1485–1493.
(14) Bhatt, P. M.; Desiraju, G. R. CrystEngComm 2008, 10, 1747–1749.
(15) Sheldrick, G. M. Acta Crystallogr. 2008, A64, 112–122.
(16) Macrae, C. F.; Bruno, I. J.; Chisholm, J. A.; Edgington, P. R.;
McCabe, P.; Pidcock, E.; Rodriguez-Monge, L.; Taylor, R.; van de Streek, J.;
Wood, P. A. J. Appl. Crystallogr. 2008, 41, 466–470.
FIGURE 5. Chiral HPLC trace for (()-1b, top; (()-2b, middle; and
(S)-2b, bottom. For (()-2b and (S)-2b, the data are obtained by
using the individual crystal from the single crystal analysis.
(17) Spek, A. L. Acta Crystallogr., Sect. D: Biol. Crystallogr. 2009, D65,
148–155.
J. Org. Chem. Vol. 76, No. 4, 2011 1161

Eccles et al.
JOCNote
3
˚
chiral HPLC analysis was performed on the actual crystal
used in the diffraction experiment.
92.659(3)°, γ = 95.430(3)°, V = 800.90(8) A , T = 100.(2)
K, space group P1, Z = 2, 18797 reflections measured, 5028
unique (Rint = 0.0238). The final R1 values were 0.0278 (I >
2σ(I)) and 0.0285 (all data). The final wR(F2) values were 0.0773
(I > 2σ(I)) and 0.0781 (all data). Flack parameter = -0.10(9),
Hooft y parameter = 0.05(6). HPLC data: Chiralcel OH-J,
eluent hexane:i-PrOH (3% trifluoroacetic acid), 94:6, flow rate
0.25 mL min-1, λ = 209.8 nm, RT (min): S = 32.7.
Synthesis of (()-3-o-Tolylbutanoic Acid:Isonicotinamide Co-
crystal, (()-2b. Isonicotinamide (27 mg, 0.22 mmol) and (()-1b
(40 mg, 0.22 mmol) were used; mp 84 - 86 °C. Crystal data:
˚
C17H20N2O3, M = 300.35, triclinic, a = 9.6495(16) A, b =
˚
˚
12.915(2) A, c= 13.390(2) A, R= 102.007(3)°,β= 103.616(4)°,
3
˚
γ = 92.047(4)°, V = 1580.0(4) A , T = 100.(2) K, space group
P1, Z = 4, 43120 reflections measured, 7818 unique (Rint =
0.0647). The final R1 values were 0.0402 (I > 2σ(I)) and 0.0570
(all data). The final wR(F2) values were 0.1009 (I > 2σ(I)) and
0.1070 (all data). HPLC data: Chiralcel OH-J, eluent hexane:
Acknowledgment. This publication has emanated from
research conducted with the financial support of Science
Foundation Ireland, under Grant nos. 08/RFP/MTR1664,
07/SRC/B1158, 05/PICA/B802 TIDA 09, and 05/PICA/
B802/EC07, IRCSET, and Eli Lilly.
i-PrOH (3% trifluoroacetic acid), 94:6, flow rate 0.25 mL min-1
,
λ = 209.8 nm, RT (min): S = 33.1 and R = 62.7.
General Procedure for the Preparation of the Enantiopure
Acid:Isonicotinamide Co-crystals. The corresponding enan-
tiopure acid (1 equiv) and isonicotinamide (1 equiv) were
treated in an identical fashion to the racemic compounds,
except the initial grinding and subsequent PXRD analysis
steps were omitted.
Supporting Information Available: Additional figures, HP-
LC traces, DSC data, PXRD data and the cif files for (()-
2a-d and (S)-2a-d. This material is available free of charge
via the Internet at http://pubs.acs.org. The crystallographic
coordinates have been deposited with the Cambridge Crys-
tallographic Data Centre; deposition nos. 790893 to 790899.
These data can be obtained free of charge from the
Cambridge Crystallographic Data Centre, 12 Union Rd.,
Cambridge CB2 1EZ, UK or via www.ccdc.cam.ac.uk/
data_request/cif.
Synthesis of (S)-3-o-Tolylbutanoic Acid:Isonicotinamide
Co-crystal, (S)-2b. Isonicotinamide (5 mg, 0.06 mmol) and
(S)-1b (7 mg, 0.05 mmol) were used; mp 89-91 °C. Crystal
˚
data: C17H20N2O3, M = 300.35, triclinic, a = 7.1279(4) A,
˚
˚
b = 8.8202(5) A, c = 13.1405(8) A, R = 102.542(3)°, β =
1162 J. Org. Chem. Vol. 76, No. 4, 2011