Development of novel brush-type chiral stationary phases based on terpenoid selectors: HPLC evaluation and theoretical investigation of enantioselective binding interactions

Article abstract of DOI:10.1016/j.tetasy.2006.11.036

The terpenoid chiral selectors dehydroabietic acid, 12,14-dinitrodehydroabietic acid and friedelin have been covalently linked to silica gel yielding three chiral stationary phases CSP 1, CSP 2 and CSP 3, respectively. The enantiodiscriminating capability

Full text of DOI:10.1016/j.tetasy.2006.11.036

Tetrahedron: Asymmetry 17 (2006) 3248–3264
Development of novel brush-type chiral stationary phases based
on terpenoid selectors: HPLC evaluation and theoretical
investigation of enantioselective binding interactions
Cristina Moiteiro,a,* Nelson Fonseca,b Maria J. M. Curto,a Regina Tavares,a Ana M. Lobo,c
b
b,
d
*
´
Paulo Ribeiro-Claro, Vitor Felix and Michael G. B. Drew
aINETI, Instituto National de Engenharia, Tecnologia e Inovac¸a˜o I.P., Estrada do Pac¸o do Lumiar, 1649-038 Lisboa, Portugal
bDepartamento de Quı´mica, CICECO, Universidade de Aveiro, 3810-193 Aveiro, Portugal
cDepartamento de Quı´mica, Centro de Quı´mica Fina e Biotecnologia Requinte, Sintor-Uninova,
ˆ
Faculdade de Ciencias e Tecnologia, UNL, 2829-516 Monte de Caparica, Portugal
dSchool of Chemistry, University of Reading, Whiteknights, Reading RG6 6AD, UK
Received 23 October 2006; accepted 23 November 2006
Abstract—The terpenoid chiral selectors dehydroabietic acid, 12,14-dinitrodehydroabietic acid and friedelin have been covalently linked
to silica gel yielding three chiral stationary phases CSP 1, CSP 2 and CSP 3, respectively. The enantiodiscriminating capability of each
one of these phases was evaluated by HPLC with four families of chiral aromatic compounds composed of alcohols, amines, phenyl-
alanine and tryptophan amino acid derivatives and b-lactams. The CSP 3 phase, containing a selector with a large friedelane backbone
is particularly suitable for resolving free alcohols and their derivatives bearing fluorine substituents, while CSP 2 with a dehydroabietic
architecture is the only phase that efficiently discriminates 1,10-binaphthol atropisomers. CSP 3 also gives efficient resolution of the free
amines. All three phases resolve well the racemates of N-trifluoracetyl and N-3,5-dinitrobenzoyl phenylalanine amino acid ester deriv-
atives. Good enantioseparation of b-lactams and N-benzoyl tryptophan amino acid derivatives was achieved on CSP 1.
In order to understand the structural factors that govern the chiral molecular recognition ability of these phases, molecular dynamics
simulations were carried out in the gas phase with binary diastereomeric complexes formed by the selectors of CSP 1 and CSP 2 and
several amino acid derivatives. Decomposition of molecular mechanics energies shows that van der Waals interactions dominate the for-
mation of the diastereomeric transient complexes while the electrostatic binding interactions are primarily responsible for the enantio-
selective binding of the (R)- and (S)-analytes. Analysis of the hydrogen bonds shows that electrostatic interactions are mainly associated
with the formation of N–Hꢀ ꢀ ꢀO@C enantioselective hydrogen bonds between the amide binding sites from the selectors and the carbonyl
groups of the analytes. The role of mobile phase polarity, a mixture of n-hexane and propan-2-ol in different ratios, was also evaluated
through molecular dynamics simulations in explicit solvent.
ꢀ 2006 Elsevier Ltd. All rights reserved.
1. Introduction
the last two decades an intensive search for novel chiral sta-
tionary phases (CSP) for high-performance liquid chroma-
tography (HPLC) has been carried out and CSPs generated
from cyclodextrines,3,4 polysaccharide derivatives,5 pro-
teins,6 macrocyclic antibiotics,7–9 synthetic polymers,10,11
chiral crown ethers12 and low molecular weight optically
active compounds are already available on the market.13,14
The latter ones, denominated brush-type or Pirkle-type, are
very attractive types to be used in the molecular design of
novel phases with enhanced chiral discrimininating ability.
Indeed, chiral selectors can be selectively modified allowing
a prompt evaluation of the structural changes on the
enantioselective recognition process.15,16 On the other
The enantiomers of chiral compounds with recognised bio-
logical activity usually exhibit different physiological effects
following different pathways in biological processes.1,2
Thus, optical resolution of racemic synthetic compounds
is one of the most interesting challenges for a synthetic
chemist and is essential in several research fields such as
pharmaceutical, agrochemical and food chemistry.2 Over
*
Corresponding authors. Tel.: +351 210924751; fax: +351 217168100
(C.M.); e-mail addresses: cristina.moiteiro@ineti.pt; vfelix@dq.ua.pt
0957-4166/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2006.11.036

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3249
hand, these phases can be produced using selectors
obtained directly from the natural chiral pool such as
amino acids,16 alkaloids,14,17–22 sterols,14,23–25 and tartaric
acid derivatives.26,27
characterisation of these novel phases. The enantioselective
chromatographic resolution capability of these phases was
evaluated by HPLC with four sets of aromatic compounds
composed of a representative number of alcohols, amines,
amino acid derivatives and cis-b-lactams (Chart 1).
Herein we report novel brush-type phases containing chiral
selectors derived from natural sources, dehydroabietic acid
1 and friedelin 3, anchored onto a solid support of silica
gel. Both compounds exhibit different functionalities
assembled in a rigid skeleton, which allows the chiral
molecular recognition of a wide range of organic sub-
strates. Dehydroabietic acid 1 is available in large amounts
from the dehydrogenated commercial resin of the pines ex-
tracted from Pinus pinaster.28 Friedelin 3 can be obtained
from the cork smoker wash solids, which are a by-product
generated in large amount by the cork processing industry
associated with Quercus suber L.29 Furthermore, this black
wax has not yet found a practical application and repre-
sents an environmental problem. Both species are wide-
spread in Portuguese forests, one of the most important
renewable sources of biomass available in the country for
industrial proposes including the production of energy.30
In an attempt to find other potential applications for these
inexpensive natural raw materials with a well recognised
high trade value, dehydroabietic acid 1, 12,14-dinitrode-
hydroabietic acid 2 and friedelin 3 were selected as chiral
selectors of three novel brush-type phases shown in Figures
1 and 2. Selector 2 was obtained by the nitration of 1.31
Subsequently, selectors 1, 2 and 3 were covalently linked
to a silica gel support using the larger spacer, 10-undecenyl-
amine, leading to CSP 1, CSP 2 and CSP 3 phases.32 Here-
in we report the synthesis and the structural
In order to understand the accumulated chromatographic
resolution data at a molecular level, the structural features
associated with the chiral recognition modes, molecular
modelling studies were carried out on the diastereomeric
binding associations formed between the chiral selectors
of CSP 1 and CSP 2 phases and the analytes of phenylal-
anine amino acid derivatives 17a and 17d. This theoretical
investigation comprised of molecular mechanics (MM)
energy calculations and molecular dynamics (MD) simula-
tions in the gas phase. The effect of an enhancement of
the mobile phase polarity on the enantioselective separa-
tion of the amino acid 17a on CSP 2 was also evaluated
through MD simulations in solution.
2. Results and discussion
2.1. Synthesis of the chiral stationary phases
12,14-Dinitrodehydroabietic acid 2 was synthesised by the
introduction of two nitro groups at the 12- and 14-posi-
tions of dehydroabietic acid 1 via electrophilic aromatic
substitution. These two chiral selectors contain a rigid
framework incorporating an aromatic ring and a carboxyl
group at the C-4 position, which was used for the covalent
link to the silica gel support.
16
R
R
R
12
17
20
1
4
14
10
R
R
R
SOCl2, reflux
NH2(CH2)9CHCH2
toluene
1´ - 9´
11´
H
H
H
19
CO2H
CONH(CH2)9CHCH2
COCl
5 R= H
1 R= H
4 R= H
8 R= NO2
2 R= NO2
7 R= NO2
(CH3)2SiHCl
H6PtCl6
16
R
R
17
20
1
4
R
10
R
silica gel, toluene
reflux
CH3
1´ - 11´
H
NH(CH2)11Si CH3
H
O
CONH(CH2)11Si(CH3)2Cl
O
OH
HO
6 R= H
OH
HO
9 R= NO2
silica gel
CSP 1 R= H
CSP 2 R= NO2
Figure 1. Syntheses of CSP 1 and CSP 2 diterpenoid brush-type phases.

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C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
29
30
18
20
27
COCl
1.
13
15
11
ClOC
H
H
H
28
1
4-DMAP, toluene, reflux
10
O
25
6
3
26
,
2.NH2(CH2)9CHCH2 pyridine
Na reduction
O
O
H
N
HO
24
23
(H2C)9
3a
3
11
O
(CH3)2SiHCl
H6PtCl6
29
30
18
20
27
H
13
15
11
H
O
28
1
10
O
CH3
O
H
N
3
26
1´
6
(H2C)11
O
Cl Si
CH3
CH3
silica gel,
11´´- 1´´
24
H3C Si (H2C)11HN
23
O
4´
toluene
reflux
12
O
O
OH
HO
OH
silica gel
CSP 3
Figure 2. Synthesis of CSP 3 triterpenoid brush-type phase.
CH3
F3C
OH
OH
OH
OH
CH NHR1
CH
CH3
R1
H
16
15
14
13
16a COCH3
16b COCF3
16c COPh
16d 3,5-DNB
CO2R
CO2R
CH2CH NHR1
(H3C)2N
CH2CH NHR1
O
O
N
H
O
O
N R
R
R1
H
H
R
R1
H
H
17
H
18
H
O
R
17a CH3
18a CH3
17b CH3 COCF3
17c CH3 COPh
17d CH3 3,5-DNB
18b CH3 COCF3
18c CH3 COPh
18d CH3 3,5-DNB
19a CH2Ph
19b o-FPh
Chart 1.
CSP 1 and CSP 2 were prepared from precursors 1 and 2,
respectively, as outlined in Figure 1. The carboxylic groups
of 1 and 2, directly bound to the stereogenic centre, were
previously transformed into their acid chlorides 4 and 7.
Subsequently, intermediates 4 and 7 were reacted with
the 10-undecenylamine spacer to afford the amides 5 and
8 in 93% and 86% yields, respectively. This derivatisation
introduces a double bond, which is well known to be an
appropriate structural moiety for the anchoring of chiral
selectors to a silica gel support. In fact, hydrosilylation of
amides 5 and 8 with dimethylchlorosilane in the presence
of a catalytic amount of chloroplatinic acid quantitatively
afforded monochlorosilane compounds 6 and 9, respec-
tively. Subsequently, these intermediates were covalently
linked to a 5 lm porous silica gel via a straight nucleophilic
reaction leading to CSP 1 and CSP 2, respectively.

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3251
The complete synthesis of CSP 3 is depicted in Figure 2. In
order to prepare this triterpenoid phase, friedelin 3,
isolated from cork smoker wash solids, was selectively
reduced to friedelan-3a-ol 3a.29 The friedelane scaffold does
not contain suitable functionalities for enantioselective dis-
crimination, such as p–p interactions; thus the hydroxyl
group of 3a was reacted with terephthaloyl chloride in
the presence of 4-DMAP leading to the formation of deriv-
ative 10. This intermediate now incorporates one aromatic
moiety with potential binding properties and one free acyl
group, which can be used to bind the spacer. Treatment of
this intermediate with 10-undecenylamine gave the N-1000-
undecenylamide compound 11, which was hydrosilylated
to silyl compound 12, using the reaction conditions de-
scribed for the preparation of the other two CSPs. Finally,
this intermediate was directly linked to 5 lm silica gel
support.
mixtures under normal phase conditions. The two enantio-
meric forms of alcohols 14 and 15 and amino acids 17 and
18 were also eluted separately in order to identify the
elution order. For comparison, all compounds were also
eluted on the (R,R)-Whelk-O1 commercial phase, which
has been widely used for the separation of the enantiomers
of a variety of compounds.34 The chromatographic results
are summarised in Table 1 together with the most adequate
mobile phases compositions found.
CSP 1 is unable to promote the resolution of the racemic
mixtures alcohols 13–15, in spite of the several mobile
phases with different degrees of polarity have been
experimented.
a-methylbenzyl alcohol 13 exhibits a good resolution on
CSP 2 and CSP 3 and (R,R)-Whelk-O1 with a values of
1.26, 1.43 and 1.68, respectively. The enantiomers of 9-
anthryltrifluoromethylcarbinol 15 are separated onto
CSP 3 with an a of 1.23 while on the (R,R)-Whelk-O1
phase they are discriminated with a a value of only 1.16.
The binding interactions between the (S)-15 analyte and
the selectors of CSP 2 and CSP 3 are stronger than those
with the (R)-15 analyte, as the latter is the first to be eluted
from both columns. Whereas, the (S)-15 analyte is the first
to be eluted on the (R,R)-Whelk-O1 phase.
The grafting yield of the selector immobilised on the silica
gel support was determined from the elemental analyses of
either carbon or nitrogen (see Experimental). The results
found are consistent with covalent linkage of the three
selectors to the silica gel support and comparable with
those reported for other brush-type phases. CSP 2 shows
a slightly higher grafting yield (0.31 mmol gꢁ1) than CSP
1 and CSP 3.32
The three phases were then packed into 150 and 100 ·
4.6 mm ID stainless-steel HPLC columns. In order to
increase the enantioselectivity, endcapping of the modified
silicas was carried out in situ throughout the reaction of the
free silanol groups with hexamethyldisilazane.33
An efficient enantioseparation of 1,10-bi-2,20-naphthol 14
was achieved on CSP 2 phase with an a value of 1.14. This
result is noteworthy compared to a resolution of 1.04 ob-
tained with the commercial phase for this atropisomer. In
fact, the separation of binaphthol compounds by (R,R)-
Whelk-O1 phase is achieved only after derivatisation of
the hydroxy groups of the atropisomer with a butane
bridge.35 Separation of 14 by CSP 2 is probably due to
the formation of transient complexes stabilised by enantio-
selective hydrogen bonds between the nitro groups from
the p-acidic selector, and free hydroxy groups of analytes,
which can be complemented concomitantly with p–p inter-
actions between the p-acidic CSP 2 and the p-basic binaph-
thol aromatic sites. These interactions are more effective
with the (S)-analyte. Moreover, it is interesting to note that
the best enantioseparations of alcohols 13–15 on this phase
were achieved using a low polarity mobile phase composed
of n-hexane/propan-2-ol/trifluoracetic acid (99:1:0.09 v/v),
in which the acid was added in order to improve the peak
shapes.
2.2. Evaluation of chiral resolution ability of the stationary
phases
Selectors 1 and 2 have rigid diterpenoid architectures con-
taining an aromatic moiety capable of enantioselective rec-
ognition of the aromatic analytes through p–p interactions.
However, these two chiral molecules display different elec-
tronic binding properties, 2 with two electron-withdrawing
nitro groups has a p acidic character, while 1 exhibits a p-
basic character. In addition, 2 can establish enantioselec-
tive N-Oꢀ ꢀ ꢀH hydrogen bonding interactions between the
NO2 groups and O–H or N–H groups from the analytes
(Fig. 1). Therefore, both compounds seem very attractive
candidates to be chiral selectors of novel brush-type
phases. CSP 3 has an extended pre-organised triterpenoid
backbone coupled with an aromatic moiety via an ester
group, which can be used concomitantly for enantioselec-
tive binding of the analytes. Moreover, the N-undecenyl-
amide fragment of the CSPs provides N–H and CO
binding groups, which extend the enantioselective discri-
mination possibilities via multiple and cooperative inter-
molecular hydrogen bonding interactions.
The chromatographic data shows that the (R,R)-Whelk-O1
is the more appropriate to separate the majority of amine
derivatives than the brush-type phases described here, with
the exception of the underivatised amine 16, which is well
separated by CSP 3 with an a of 1.55. This racemate is
not resolved by a commercial phase. Amine derivative
16b with a trifluoracetyl group is also well resolved upon
CSP 3 with an a of 1.74, but the enantiochromatographic
separation of this compound on (R,R)-Whelk-O1 phase
occurs with a 3.00. The N-3,5-dinitrobenzoyl (N-3,5-DNB)
amine derivative 16d with a p-acidic character is slightly
discriminated on CSP 1 with an a of 1.04 and CSP 2 with
an a of 1.07. This compound finds an efficient resolution on
(R,R)-Whelk-O1 phase with an a of 1.98.
The enantioselective resolution capabilities of CSP 1, CSP
2 and CSP 3 were evaluated with four representative sets of
chiral aromatic analytes composed of alcohols 13–15,
amines 16–d, amino acid derivatives from phenylalanine
17–d and tryptophan 18–d and cis-b-lactams 19a–b, respec-
tively (Chart 1). All compounds were eluted as racemic

3252
C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
Table 1. Chromatographic resolution of racemic compounds on CSP 1, CSP 2, CSP 3 and commercial (R,R)-Whelk-01
Compound
CSP 1
CSP 2
CSP 3
(R,R)-Whelk-01
a
k10
Rs
a
k10
Rs
a
k10
Rs
a
k10
Rs
13
14
1.00
1.00
1.00
1.00
1.00
1.06f
1.00
1.04g
1.00
4.83d
1.30c
1.04e
3.65e
1.00
2.00d
1.00
1.00
1.00
3.94e
2.93e
—
—
—
—
1.26a
1.14a
1.08a
1.00
0.66
10.47n
9.95n
—
1.13
1.54
0.65
—
1.43b
1.00
1.23c
1.55j
1.00
1.74k
1.00
1.00
1.00
2.44c
1.00
1.00
2.94c
1.00
1.00
1.00
1.00
1.00
1.00
1.00
0.45
—
2.00
—
0.80
2.00
—
—
—
—
—
3.40
—
—
1.56
—
—
—
—
—
—
1.68c
1.04d
1.16e
1.00
2.21c
3.00e
3.20c
1.98c
1.00
0.69
6.57o
1.89o
—
3.30
1.64
3.60
7.20
—
—
—
—
1.42o
3.50
—
1.13
—
3.91
8.82
9.40
4.85
—
—
—
—
—
15
16
1.26n
0.20
—
—
16a
16b
16c
16d
17
1.00
—
—
4.55
—
22.36
—
0.70n
0.92o
2.58
0.92o
0.70
—
—
1.06h
1.09h
1.07h
1.00
3.62
14.0
17.8
—
1.62n
1.21n
—
1.03n
—
—
—
8.68o
—
0.28
—
—
0.76
0.82
—
1.13
—
—
9.22
—
—
—
1.69
—
—
—
—
17a
17b
17c
17d
18
3.79
0.89
—
2.87i
1.05c
1.00
5.03e
1.00
0.35n
—
1.00
1.00
—
1.00
6.40
0.32o
—
1.30l
1.00
18a
18b
18c
18d
19a
19b
0.73n
—
—
—
0.16
0.17
1.95
—
—
—
6.78
4.78
1.00
—
—
—
—
—
—
1.00
1.00
1.00
1.15l
1.36m
1.41m
1.00
1.34e
1.00
1.23o
4.00
4.20
—
3.35
3.45
1.00
1.00
—
—
—
—
Mobile phase: a n-hexane/propan-2-ol/trifluoroacetic acid (99:1:0.09) (v/v); b n-hexane/dichloromethane (80:20); c n-hexane/propan-2-ol (90:10); d n-hex-
ane/propan-2-ol (80:20); e n-hexane/propan-2-ol (95:5); f n-hexane/propan-2-ol (99:1); g n-hexane/propan-2-ol/trifluoroacetic acid (97:3:0.09); h n-hexane/
propan-2-ol/methanol (96:3.5:0.5); i n-hexane/propan-2-ol/ethanol (99:0.5:0.5); j n-hexane/dichloromethane (90:10); k n-hexane/dichloromethane (95:5);
l n-hexane/propan-2-ol/ethanol (95:2:3); m n-hexane/propan-2-ol (50:50); n (R) absolute configuration of the first eluted enantiomer; o(S) absolute confi-
guration of the first eluted enantiomer.
The free amino acid 17 is not separated by any of the four
phases either. In contrast, the methyl ester of phenylalanine
17a has a good resolution on the three novel phases with
enantioseparation factors of 4.83, 2.87 and 2.44 for CSP
1, CSP 2 and CSP 3, respectively; while on the commercial
phase, the racemate of this derivative is not separated. This
result suggest that the presence of a methyl ester can im-
prove enantioseparation as reported previously by several
authors.36 However, the attachment of bulky substituents
at the amine group, such as N-trifluoracetyl in 17b and
N-benzoyl (an extra p-donor aromatic binding site) in
17c, decreases the enantiodiscriminating capability of all
the brush-type phases. Indeed, only racemate 17b is sepa-
rated on CSP 1 with an a of 1.30 while 17c is only slightly
resolved with an a of 1.04 when the polarity of the mobile
phase is reduced (see Table 1). These results indicate that
the steric bulk of the N-substituent prevents the formation
of enantioselective N–Hꢀ ꢀ ꢀO hydrogen bonds between the
analytes and the selectors. Efficient enantioselective separa-
tions of the analytes of the N-3,5-dinitrobenzoyl (3,5-DNB)
methyl ester amino acid derivative 17d are obtained on
CSP 1, CSP 2 and CSP 3 with separation factors of
3.65, 5.03 and 2.94, respectively. It is noteworthy that the
best enantioseparation of this p-acidic analyte is achieved
with the p-acidic CSP 2 phase suggesting that the forma-
tion of the transient complexes is mainly governed by enan-
tioselective hydrogen bonding interactions rather than p–p
interactions. We will return to this point later. The race-
mate of 17d is also resolved by the commercial phase, but
with a lower a of 1.30.
of an indolic ring decreases or turns off the enantioselectiv-
ity. Indeed, methyl ester 18a is resolved upon CSP 1 with
an a of 2.00, while the analytes of methyl ester 18c having
a N-benzoyl group are separated by CSP 2 with an a value
of 1.34. The N-(3,5-DNB) methyl ester derivative 18d is not
discriminated by any of the three CSPs. This derivative is
resolved on the (R,R)-Whelk-O1 phase with an a of 1.15.
Furthermore, CSP 3 was shown to be inadequate in resolv-
ing any of the tryptophan amino acid derivatives.
The two cis-b-lactams display low capacity factors, below
0.7, but they are resolved by CSP 1 with a values of 3.94
for 19a and 2.93 for 19b. By contrast, these compounds
are more retained on the (R,R)-Whelk-O1 phase with k10
values of 4.00 for 19a and 4.20 for 19b, nevertheless its sep-
aration on this phase is less efficient with a values of 1.36
and 1.41 for 19a and 19b, respectively.
2.3. Molecular modelling studies
In order to obtain further insights into the enantioselective
binding behaviour of CSP 1 and CSP 2 phases, molecular
mechanics energy calculations were carried out for the dia-
stereomeric complexes between the selectors of these two
chiral stationary phases and the two enantiomers amino
acid derivatives 17a and 17d, which exhibit the higher sep-
aration factors on both phases (see Table 1). The starting
geometries of all the binary complexes were generated
using the docking strategy described in Section 4.
Figure 3 shows the structures of the lowest energy binding
associations found for each diastereomeric complex with
the corresponding differences in molecular mechanics ener-
gies between (R)- and (S)-complexes (DR–S). The binding
The enantioseparation results for tryptophan amino acid
18 and their ester derivatives 18a–d, by comparison with
those obtained for phenylalanine suggest that the presence

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3253
Figure 3. Lowest energy binding scenarios found on conformational analyses on diastereomeric complexes with DR–S energies (see text) in kcal molꢁ1. The
carbon skeleton of selectors and analytes are in yellow and green, respectively. The oxygen and nitrogen atoms are in the conventional colours red and
blue, respectively. The hydrogen bonding interactions between the binding sites are drawn as dotted lines in light blue.
regions of the CSP 1 phase are illustrated in Figure 4
through the isodensity contour map, which was drawn
sampling all (R)-analyte orientations from the minimised
CPS 1Æ17a binding scenarios saved along the molecular
dynamics quenching run. The shell surface almost encases
the selector domain validating the docking strategy
adopted. As would be expected, the most densely popu-
lated region is the hydrogen bonding site of CSP 1, namely
the amide group. Subsequently, MD simulations of 40 ns
were carried out in the gas phase at 300 K using the lowest
energy structures shown in Figure 3 as starting models.
The binding energy (DE) associated with an enantioselec-
tive interaction between the selector and a single enantio-
mer is calculated using the equation:
DE ¼ Ecomplex ꢁ Eselector ꢁ Eenantiomer
ð1Þ
In which Ecomplex is the total energy for the diastereomeric
complex while Eselector and Eenantiomer are the individual
energies for the selector and the enantiomer, respectively.
Usually, these three energetic terms are evaluated from
three independent simulations carried with complex, selec-
tor and the analyte, respectively. However, when the

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C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
The DEelec and DEvdw terms are average energy differences
calculated with snapshots of the complex, selector and ana-
lyte retrieved by post-processing the MD trajectory file of
each diastereomeric complex with the MM-PBSA meth-
od37 as described in Section 4. The results obtained for
the eight complex diastereomers studied are compiled in
Table 2. The evolution of accumulated binding energies
for 40 ns of simulation is plotted in Figure 5 for the tran-
sient complexes between the analytes of 17a and 17d and
the CSP 1 selector and in Figure 6 for the complexes
formed with these analytes and the CSP 2 selector. The
plot for the two CSP 1Æ17a diastereomeric associations
(Fig. 5a) shows that after the first 5 ns of simulation the
(S) enantiomer is more tightly bound to the phase and
the interaction energy of the two diastereomeric complexes
remains almost constant until the end of the simulations.
Thus, the (R) enantiomer is predicted to be the first one
to be eluted from the HPLC column, which is in agreement
with experimental findings. Figure 5b shows that over the
majority of the simulation, the CSP 1 selector has a clear
binding preference for the (R) enantiomer of 17d. The
(R) diastereomeric complex displays a constant energetic
stabilisation from 3 ns until the end of the simulation. By
contrast, the simulation with the (S) complex starts with
a marked increase of the binding energy of simulation until
16 ns of simulation, then the energy shows a progressive
slight decrease for the remaining 24 ns of simulation apart
from a short period of 2 ns around 26 ns.
Figure 4. Isodensity contour map for CSP 1Æ(R)-19a built sampling all
(R)-19a analyte orientations saved over molecular dynamics run in the gas
phase.
formation of the transient complexes occur without a
significant conformational change of selector and analytes,
these three individual energetic contributions can be evalu-
ated from the MD simulation of the complex and then the
binding energy is only determined by electrostatic (DEelec
)
and van der Waals (DEvdw) energy differences being Eq. 1
simplified to
DE ¼ DEelec þ DEvdw
ð2Þ
Table 2. Interaction energies (kcal molꢁ1) for the diastereomeric complexes with CSP 1 and CSP 2 selectors averaged over the 200,000 frames taken from
the 40 ns molecular dynamics simulation in the gas phase
DDEc
a
b
Phase
Analyte
DEele
DEvdw
DE
DDEele
DDEvdw
CSP 1
(R)-17a
(S)-17a
(R)-17d
(S)-17d
ꢁ3.95
ꢁ4.31
ꢁ10.37
ꢁ9.07
ꢁ8.63
ꢁ8.77
ꢁ12.58
ꢁ13.08
ꢁ26.84
ꢁ24.36
0.36
0.14
0.50
ꢁ16.47
ꢁ1.30
ꢁ1.18
ꢁ2.48
ꢁ15.29
CSP 2
(R)-17a
(S)-17a
(R)-17d
(S)-17d
ꢁ4.40
ꢁ4.93
ꢁ7.74
ꢁ8.74
ꢁ10.29
ꢁ10.33
ꢁ17.51
ꢁ17.31
ꢁ14.69
ꢁ15.26
ꢁ25.25
ꢁ26.05
0.53
1.00
0.04
0.57
0.80
ꢁ0.20
a DDEele = DEele(R) ꢁ Eele(S).
b DDEvdw = DEvdw(R) ꢁ Evdw(S).
c DDE = DE(R) ꢁ DE(S). Remaining energy terms are defined in the text.
Figure 5. Evolution of the average binding energies over the MD simulations on diastereomeric binding associations between CSP 1 and analytes 17a (a)
and 17d (b).

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3255
Figure 6. Evolution of the average binding energies over the MD simulations on diastereomeric binding associations between CSP 2 and analytes 17a (a)
and 17d (b).
Figure 6a shows that the simulation between the (R)-17a
and the CSP 2 selector starts with a pronounced increase
of the average binding interaction energy and during the
first 3 ns of the MD the enantioselective binding forces
favour the formation of CSP 2Æ(R)-17a complex. However,
after 5 ns, this binding preference is reversed with the bind-
ing energies for the two diastereomeric complexes remain-
ing almost constant to the end of the MD simulations.
sents the molecular mechanics energy, which is only one
of the three components of the free energy. The remaining
two, the entropic and the enthalpic contributions were not
taken into account in our calculations nor were the solvent
effects. However, the entropy was estimated for all diaste-
reomeric pairs using the NMODE program as implemented
in AMBER 8 package. The values for the (R) and (S) com-
plexes were similar showing that entropy contribution is
negligible on the chromatographic chiral discrimination as
suggested previously by Lipkowitz.38 Finally, the molecular
modelling models are an oversimplification of the chro-
matographic experiments since that neither the spacers
nor the anchor of the spacer onto the silica gel support
are taken into account.
The graphs for CSP 2Æ17d diastereomeric complexes plotted
in Figure 6b show that the binding interaction with the (S)
enantiomer is favoured after the first 5.5 ns until the end of
the simulation, which agrees with the experimental chro-
matographic data. Further insight on the enantioselective
discrimination is derived from the difference between the
intermolecular binding energies of (R) and (S) complexes,
DDE = DER ꢁ DES, which can be considered to be a quan-
titative measure of the chiral discrimination. The DDE val-
ues calculated for all pairs of diastereomeric complexes,
given in Table 2, are in agreement with the experimental
HPLC elution orders found for the (R) and (S) analytes
of 17a and 17d on CSP 1 and CSP 2. The negative values
of DEelec and DEvdw energy differences obtained for each
diastereomeric complex indicate that in all of them both
terms contribute to hold the receptor and the analyte to-
gether. Furthermore, the van der Waals interactions are
the dominant stabilising intermolecular forces while the
electrostatic binding interactions are the discriminating
forces responsible for the DDE energy differences between
the (R) and (S) complexes. The unique exception is the
enantioselective binding of the CSP 1 to the analytes of
17d, where the energy differences between (R) and (S)
complexes are ꢁ1.30 kcal molꢁ1 for DDEelec term and
ꢁ1.18 kcal molꢁ1 for the DDEvdw term leading to a DDE
discriminating energy of ꢁ2.48 kcal molꢁ1. Furthermore,
these enantiodifferentiating energies (DDE) are very small
when compared with the energies associated with the for-
mation of each diastereomeric complex (DE). The jDDEj
values lead to the following resolution orders for (R) over
(S) 17d > 17a on CSP 1 and CSP 2. The order for CSP2
is entirely consistent with experimental a values while the
order for CSP 1 is not. However, this result is not necessar-
ily inconsistent with our results since the DDE term repre-
Further analysis of MD simulations was carried out with
the ANAL program within the AMBER 8 package. The selec-
tors of the CSP 1 and CSP 2 phases were split at chiral car-
bon C-4 into three fragments as shown in Figure 7. The
first fragment is the N-propyl amide group, the second is
the diterpenoid backbone and the methyl group is the third
one. This fragment decomposition is arbitrary, albeit it
provides a better understanding of how the enantiomers
are discriminated. The energies associated with each one
of these three fragments interacting with the analytes are
collected in Table 3.
The selectors and analyte are held together by short-range
dispersion forces and so the diterpenoid fragment is mainly
responsible for the stabilisation of most of the diastereo-
meric complexes. The second most important stabilising
factor comes from the amide group, which interacts with
the analytes via electrostatic forces. As mentioned above,
the unique exception is the complex between CSP 1
and the (R) enantiomer of 17d, where both fragments con-
tribute almost identically towards stabilisation. The methyl
group favours only marginally the formation of the com-
plexes because the approach of the analytes to the chiral
binding site of CSP 1 and CSP 2 from the side of the
methyl group is not favoured because of steric repulsions.
The enantioselective differentiating forces (electrostatic
interactions as reported above) are consistent with the pat-
tern of the hydrogen bonds established by the selector and

3256
C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
atom. The CSP 1 and CSP 2 phases discriminate the ana-
R
lytes of 17a and 17d diastereomers mainly through the
N–Hꢀ ꢀ ꢀO@C hydrogen bonding interactions established
between the N–H and carbonyl binding sites from the
amide group of the phase and analyte, respectively. This
interaction was found to be quite strong in all diastereo-
meric complexes with average Hꢀ ꢀ ꢀO distances within the
˚
narrow range of 2.09–2.26 A. Furthermore, this dis-
R
crimination comes mainly from the different occupancies
(the percentage of time that the hydrogen bond is formed
over the complete MD trajectory) of this hydrogen for
the R and S complexes. For example, the N–Hꢀ ꢀ ꢀO@C
hydrogen participates with an occupancy of 99.3% for
the formation of CSP 1Æ(R)-17d and only with 77% for
CSP 1Æ(S)-17d and the formation of the first complex is
favoured by an electrostatic stabilisation of 1.30 kcal molꢁ1
in the gas phase. Surprisingly, the nitro groups of CSP 2
occasionally establish weak N–Hꢀ ꢀ ꢀO hydrogen bonds with
the N–H binding site of the R analyte but not with the S
isomer. However this will have a marginal impact on
enantioresolution.
Fragment 2
4
H
CONHCH2CH2CH3
Fragment 3
Fragment 1
CSP 1 R = H
CSP 2 R = NO2
Figure 7. Diagram showing the fragment decomposition of CSP 1 and
CSP 2 selectors used on energetic analyses.
the analytes during the molecular simulations. Table 4 lists
the dimensions of the hydrogen bonds found for all tran-
sient complexes with occupancies greater than 10% and
an Hꢀ ꢀ ꢀA distance less than 2.50 A, A being an acceptor
As reported above CSP 1 was found experimentally to be
unable to discriminate between the 17a analytes in n-hex-
ane but could discriminate with a mobile phase composed
of n-hexane and propan-2-ol (ratio 80:20). In order to eval-
˚
Table 3. Energetic contribution of two fragments in the binding interaction between the analytes and CSP 1 and CSP 2 phases
Fragment 1
Fragment 2
Fragment 3a
Analyte
DEele
DEvdw
DEtotal
DEelec
DEvdw
DEtotal
DEelec
DEvdw
DEtotal
CSP 1
CSP 2
(R)-17a
(S)-17a
(R)-17d
(S)-17d
ꢁ3.11
ꢁ3.36
ꢁ10.32
ꢁ7.05
ꢁ2.23
ꢁ2.43
ꢁ3.41
ꢁ3.25
ꢁ2.80
ꢁ2.51
ꢁ3.32
ꢁ3.27
ꢁ5.34
ꢁ5.79
ꢁ0.82
ꢁ0.94
ꢁ0.16
ꢁ2.08
ꢁ7.93
ꢁ9.29
ꢁ0.26
ꢁ0.15
ꢁ6.26
ꢁ6.21
ꢁ7.08
ꢁ7.15
ꢁ0.02
ꢁ0.02
0.12
ꢁ0.13
ꢁ0.12
ꢁ0.29
ꢁ0.28
ꢁ0.08
ꢁ0.09
ꢁ0.27
ꢁ0.32
ꢁ0.15
ꢁ0.14
ꢁ0.17
ꢁ0.22
ꢁ0.24
ꢁ0.26
ꢁ0.24
ꢁ0.21
ꢁ13.73
ꢁ12.77
ꢁ12.93
ꢁ10.30
ꢁ11.74
ꢁ13.82
0.06
(R)-17a
(S)-17a
(R)-17d
(S)-17d
3.69
4.35
0.89
1.84
ꢁ7.41
ꢁ7.71
ꢁ15.34
ꢁ17.00
ꢁ14.18
ꢁ13.85
ꢁ0.16
ꢁ0.17
0.03
ꢁ7.51
ꢁ10.83
ꢁ13.92
ꢁ8.72
ꢁ11.99
ꢁ13.70
0.11
Values are averaged from the 200,000 frames taken from the 40 ns molecular dynamics simulation in the gas phase.
a The fragment definitions are given in the text and in Figure 7.
Table 4. Average dimensions of the hydrogen bonds in transient diastereomeric complexes formed by selectors (S) and enantiomers (E)
˚
Complex
Selector
Enantiomer
% Occupation
Hꢀ ꢀ ꢀA (A)
D–Hꢀ ꢀ ꢀA (ꢁ)
CSP 1Æ(S)-17a
CSP 1Æ(R)-17a
CSP 1Æ(S)-17d
N–H
N–H
O@C
H–N
O@C
N–H
N–H
N–H
NO2
O@C
O@C
N–H
O@C
N–H
O@C
O@C
N
NH2
N–H
O@C
N–H
71.9
64.7
77.0
20.0
99.3
80.4
64.6
14.1
11.6
72.8
22.5
93.6
2.21(26)
2.25(28)
2.11(24)
2.30(27)
2.09(22)
2.22(27)
2.26(28)
2.43(29)
2.47(31)
2.10(23)
2.24(26)
2.11(23)
163(8)
161(9)
154(10)
158(9)
158(10)
165(9)
163(9)
159(12)
151(17)
159(10)
164(8)
156(12)
CSP 1Æ(R)-17d
CSP 2Æ(S)-17a
CSP 2Æ(R)-17a
CSP 2Æ(R)-17d
CSP 2Æ(S)-17d
O@C
N–H
O@C
O@C and N–H denote oxygen and hydrogen atoms from the amide binding site; O and O–H denote oxygen and hydrogen atoms from the alcohol binding
site, N and NH2 denote nitrogen and hydrogen atoms from amine group; NO2 denote oxygen atoms from the NO2 group. A and D represent acceptor and
donor atoms, respectively.

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3257
Figure 8. Molecular dynamics simulations of the enantioselective binding recognition of 19a analytes by CSP 1 in the n-hexane/2-propan-ol (80:20 vv)
mobile phase: (a) evolution of the cumulative average binding energies; (b) variations of the intermolecular distance between the centre of mass of aromatic
rings of the selector and (R)-17a and (S)-17a analytes.
uate the role of the polarity of the mobile phase on the
enantioresolution of 17a enantiomers by CSP 1, molecular
dynamics simulations in this medium were carried out for
4 ns. The binding energy difference DDE between the (R)
and (S) analytes, calculated with the MM-PBSA method37
is 5.43 kcal molꢁ1, showing a significant increase compared
to the value of 0.50 kcal molꢁ1 determined in the gas phase.
The van der Waals and the electrostatic energy components
are both attractive contributing to the stabilisation of the
two diastereomeric complexes. However, the formation of
the (S) complex is favoured by 2.10 and 3.32 kcal molꢁ1
,
respectively. The average accumulated binding intermolec-
ular energies for the two diastereomeric complexes are plot-
ted against time in Figure 8a, while Figure 8b shows the
variation in intermolecular distances between the centres
of mass of the selector and the analytes. During the
Figure 9. Rdfs showing the interaction between the propan-2-ol solvents molecules with (R) and (S) CSP 1Æ17a diastereomeric complexes: top with the
CSP 1 selector; bottom with the 17a analytes. MCN represents the centre of mass of the amide binding group of the selector.

3258
C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
simulation the selector binds tightly with the (S) analyte
atives were achieved on CSP 1. All three phases can be also
used for the efficient resolution of racemates of N-trifluo-
racetyl and N-3,5-DNB ester phenylalanine derivatives.
Experimental results show that CSP 2 containing two nitro
groups can discriminate better than CSP 1. Therefore, the
introduction of the two nitro groups at the C-12 and C-14
positions of the aromatic ring of the dehydroabietic acid
provides new and additional sites for intermolecular inter-
actions with chiral analytes and this can significantly
enhance chiral recognition ability, especially for binaphthols
and methyl ester phenylalanine derivatives. The highest a
value was found for the enantioresolution of 17d upon
CSP 2. In this case, the phase and selector both have p-
acidic character showing that the cooperative binding
interaction between p-acidic and p-basic centres is not nec-
essary for the success of an enantioseparation. Indeed, our
molecular modelling studies shown that the stability of the
transient complexes formed between CSP 1 and CSP 2 and
analytes 17a, 17d and 14 is determined by van der Waals
interactions while the enantioselectivity is mainly dictated
by hydrogen bonds. Moreover, the efficiency of enantiosep-
aration depends largely on the expertise of the analytical
chemist to discover a mobile phase with an appropriate
polarity as shown by our experimental and simulations
studies in an n-hexane/propan-2-ol solvent mixture. The
chromatographic data obtained for the four different fam-
ilies of compounds composed of a representative number
of structurally related molecules also shows that the mole-
cular design of chiral phases for general use, even for similar
compounds, is still a hard task and a challenge for the
chemists as recently noticed.39 The ability of CSP 1, CSP
2 and CSP 3 phases to promote the chiral enantiosepara-
tion of the aminoacid derivatives using reverse polarity mo-
bile phases such as water and methanol are in progress in
our laboratories.
such that this intermolecular distance displays only small
˚
changes around its average value of ca. 4.0 A. By contrast,
for the (R) analyte the binding association is interrupted
during two periods of simulation, between 1.6 and 1.8 ns
and 2.3–3.8 ns, with concomitant solvation of the analyte
with propan-2-ol eventually via weak N–Hꢀ ꢀ ꢀO–H hydro-
gen bonding interactions with Hꢀ ꢀ ꢀO distances ranging
˚
from 2.26 to 2.38 A. At this stage, it is important to note
that in order to evaluate the solvent discriminating role,
that is the competition between the propan-2-ol solvent
molecules and the analytes for the binding sites of phase,
no restraint distance was applied between the enantiomers
and the selector. The binding energy for CSP 1Æ(S)-17a is
lower than that found for CSP 1Æ(R)-17a over the time of
simulation, with the exception for the period between 1.2
and 1.8 ns. The fragment decomposition analysis indicates
that in solution discriminating energies are derived from
the binding interaction between the amide group and the
diterpenoid moiety with the analytes. As found in the gas
phase calculations, CSP 1 binds with the analytes via N–
Hꢀ ꢀ ꢀO@C hydrogen bonds with occupancies of 26% and
61% for (R) and (S) analytes, respectively, which is consis-
tent with an electrostatic stabilisation of the (S) complex by
1.51 kcal molꢁ1. The diterpenoid fragment interacts with
analytes via dispersive van der Waals forces, which also
favour the formation of the latter complex in
2.51 kcal molꢁ1. The radial distribution functions (rdf)
for the intermolecular distances from the oxygen of the
propan-2-ol solvent molecules to the chiral centre of ana-
lyte and the centre of mass of the amide binding group
are presented in Figure 9 for the two CSP 1Æ(S)-17a and
CSP 1Æ(R)-17a diastereomeric complexes and show identi-
cal profiles with two well-defined peaks centred at 1.9
˚
and 3.0 A. The first one has higher intensity characteristic
of O–Hꢀ ꢀ ꢀO@C hydrogen bonding interactions between
propan-2-ol molecules and the carbonyl from the CSP 1.
Indeed, this peak integrates approximately to 3.95 for (R)
complex and 4.53 for (S) complex, suggesting that in this
diastereomeric binding association the selector is more
accessible for solvent hydrogen bonding interactions. On
the other hand, the rdfs for the interaction with analytes
show two broad peaks consistent with the formation of
4. Experimental
4.1. Chemicals and reagents
12,14-Dinitrodehydroabietic acid 2 was synthesised by the
nitration of dehydroabietic acid 1,31 which was isolated
from commercially dehydrogenated rosin.28 Friedelin 3
was isolated from cork smoker wash solids while the frie-
delan-3a-ol derivative 3a was prepared by the selective
reduction of friedelin 3.29
˚
two solvent shells at ca 3.2 and 4.5 A around the analyte
(R) while for the (S) analyte only the first coordination
shell is observed. Therefore, propan-2-ol molecules solvate
better with the (R) analyte than the (S) analyte leading to
the enantioselective separation of 17a analytes on CSP 1.
Toluene and THF were refluxed in the presence of sodium
benzophenone and distilled before use. The remaining
reagents and solvents were used without further purification.
All chiral test compounds are drawn in Chart 1 together
with the numbering scheme adopted. For amines, amino
acids and cis-b-lactams the numbering scheme is composed
of a number indicating the type of compound and a letter
used to distinguish between molecules with different R and
R1 substituents. The chiral test compounds 14–18 were
purchased from Aldrich or Merck. Racemic mixture of
alcohol 13 was prepared by the reduction of the corre-
sponding ketones with LiAlH4. Amine 16a–c, amino acid
17a–d and 18a–d derivatives were synthesised from their
counterparts using standard procedures. Thus, the analytes
3. Conclusions
Three novel brush phases based on two terpenoid skeletons
with different enantioselective capabilities were prepared.
The CSP 3 phase can resolve racemates of alcohols, specif-
ically derivatives containing fluorine substituents, and free
amines. CSP 2 is the only phase to resolve unequivocally
1,10-binaphthol atropisomers. This result is noteworthy
since that with the brush-type phases reported in the liter-
ature, the enantioseparation of these alcohols is achieved
only with their derivatives. Good enantioseparations of
cis-b-lactams and N-benzoyl tryptophan amino acid deriv-

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3259
of methyl esters 17a and 18a were obtained by the treat-
ment of the corresponding analytes of 17 and 18 with thio-
nyl chloride in methanol. Reaction of 16 with acetic and
trifluoroacetic anhydrides yielded the amide derivatives
16a and 16b, respectively, as racemic mixtures. Treatment
of 16, 17 and 18 with benzoyl chloride in the presence of
pyridine in toluene afforded compounds 16c, 17c and 18c,
respectively; while derivatives 16d, 17d and 18d were
obtained using 3,5-dinitrobenzoyl chloride under the same
reaction conditions. The cis-b-lactams 19a and 19b were
synthesised following the synthetic procedure described in
Ref. 40, and spectroscopic data recorded (comprise [a]D
for each analyte) for all derivatives are consistent with their
structures. Silica gel for TLC refers to Merck silica gel
GF254 and for flash chromatography to Merck silica gel
60 230–400 mesh. Organic phases were dried over anhydr-
ous sodium sulfate.
ment (FAB) on a 3-nitrobenzyl-alcohol matrix. High-reso-
lution MS spectra (HRMS) were obtained on a FTICR/
MS Finnigan FT/MS 2001-DT spectrometer at 70 eV by
electron impact or on a Finnigan MAT 900 ST spectro-
meter by ESI. Infrared (IR) spectra were recorded on a
Perkin–Elmer 1725X FTIR spectrometer. Melting points
were taken on a Reichert Thermovar thermal analyser and
are uncorrected. Elemental analyses were performed in a CE
EA-1110 microanalyser. Optical rotations were measured
at the sodium D-line on a Perkin–Elmer 241 polarimeter.
4.3. Synthesis of CSP 1
CSP 1 and CSP 2 phases were prepared by covalent link of
1 and 2 onto the silica gel support following similar syn-
thetic procedures (Fig. 1).
4.3.1. 10-Undecenylamine spacer and its 10-undecenamide
intermediate. 10-Undecenylamine spacer and its 10-
undecenamide intermediate were synthesised using the pro-
cedure described elsewhere.27 The full characterization of
these compounds was not reported previously, being there-
fore presented here.
Porous spherical silica gel Nucleosilꢂ 100 A–5 lm from
˚
Macherey-Nagel (Duren, Germany; Batch 20404134) was
¨
used as the support material of the three CSPs. Mobile
phases were prepared using n-hexane and the organic mod-
ifiers methanol, propan-2-ol, ethanol and dichloromethane
of HPLC-grade Riedel-deHae¨n (Germany).
10-Undecenamide, white crystals (81% yield); mp 85–87 ꢁC
(CHCl3/n-hexane) (mp lit.27 88.5–89 ꢁC); IR (KBr, cmꢁ1):
3359 (NH), 3192 (NH), 3083, 2922, 2851, 1662 (CONH),
Comparative chromatographic studies were carried out on
the commercial column (R,R)-Whelk-O1 (LiChroCARTꢂ
250-4) purchased from the Merck Company.
1
1632 (C@C), 1469, 1426, 1420, 912, 702, 637; H NMR
(CDCl3):
d 1.30 (10 H, s, CH2), 1.63 (2H, m,
CH2CH2C@O), 2.04 (2H, m, CH2CH@CH2), 2.22 (2H, t,
J = 7.5 Hz, CH2C@O), 4.93 (1H, br d, J11,10(cis) = 10.2 Hz,
HCH@CH), 4.99 (1H, dd, J11,10(trans) = 17.1 and
Jgem = 1.5 Hz, HCH@CH), 5.44 (1H, br s, N–H, D2O
exchange), 5.61 (1H, br s, N–H, D2O exchange), 5.82
(1H, dddd, J10,11(trans) = 17.1, J10,11(cis) = 10.2 Hz, J10,9 = 6.9
and 6.9 Hz, CH@CH2); EIMS m/z (relative intensity) 183
[M]+ (24.3), 141 (18.2), 123 (12.2), 111 (15.3), 97 (14.2),
83 (5.7), 59 [H2NC(OH)CH2]+ (84.3), 44 [H2NCO]+
(28.5), 43 (100).
4.2. Instrumentation
The chromatographic separations were performed using a
Spectra-Physics liquid chromatograph system with a UV
spectra Chrom 100 detector, injector equipped with a 20-
lL sample loop, connected to a Chrom Jet register. The
UV detector was operated at 254 nm. All separations were
carried out at room temperature. The dead time (t0) of the
column was determined by the retention time of 1,3,5-tri-
tert-butylbenzene, a presumed unretained analyte. The sep-
aration factor (a) between analytes and resolution factor
(Rs) were defined as
10-Undecenylamine, colourless oil (76.3% yield), bp
106 ꢁC, 10 mmHg (bp lit.27 123–124 ꢁC, 21 mmHg); IR
(NaCl cell, cmꢁ1): 3330 (NH), 3080, 2920, 2850, 1640
(C@C), 1570, 1480, 1470, 1150, 990, 910; 1H NMR
(CDCl3): d 1.28 (12H, s, CH2), 1.44 (2H, t, J = 6.9 Hz,
CH2CH2NH2), 2.04 (2H, m, CH2CH2@CH2), 2.68 (2H, t,
J = 6.9 Hz, CH2NH2), 4.93 (1H, br d, J11,10(cis) = 10.2 Hz,
t2 ꢁ t0 k02
a ¼
¼
ð3Þ
ð4Þ
t1 ꢁ t0 k01
2ðt2 ꢁ t1Þ
Rs ¼
W 1 þ W 2
where t1 and t2 are the retention times, k01 and k20 are the
capacity factors, W1 and W2 are the peak widths at the
base for the first and second analytes to leave the column,
respectively.
HCH@CH), 4.99 (1H, dd, J11,10(trans) = 17.1 and Jgem =
1.5 Hz, HCH@CH), 5.82 (1H, dddd, J10,11(trans) = 17.1,
J10,11(cis) = 10.2 Hz, J10,9 = 6.9 and 6.9 Hz, CH@CH2);
EIMS m/z (relative intensity): 169 [M]+ (30.0), 88 (10.3),
70 (20.4), 61 (40.2), 43 [CH2CHNH2]+ (100).
1H and 13C NMR spectra were recorded on a General
Electric GE Plus-300 spectrometer. Chemical shifts were
reported in parts per million (ppm) relative to
tetramethylsilane as the internal standard. The following
abbreviations were used: s = singlet; br s = broad singlet;
d = doublet; br d = broad doublet; dd = double dou-
blet; t = triplet; m = multiplet; coupling constants J are
expressed in hertz (Hz). MS spectra were obtained on a
Kratos MS-25 RF spectrometer via electron impact using
an ionising energy of 70 eV or through fast atom bombard-
4.3.2. Dehydroabietoyl chloride, 4. Dehydroabietic acid
(0.99 g, 3.29 mmol) dissolved in SOCl2 was heated under
reflux (6 h). The excess of SOCl2 was evaporated under re-
duced pressure and the crude product was washed succes-
sively with toluene (40 mL). A yellow crystalline product
23
4 was obtained, mp 150–152 ꢁC (toluene); ½aꢂD ¼ þ59:0 (c
1, ethanol); IR (KBr, cmꢁ1): 3060, 2900, 2850, 1780
1
(C@OCl), 1530, 1460, 1420, 990, 914; H NMR (CDCl3):
d 1.14 (3H, s, CH3-20), 1.15 (6H, d, J = 6.9 Hz, CH3-16

3260
C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
and 17), 1.30 (3H, s, CH3-19), 1.51 (2H, m, H-1ax and
H-6ax), 1.72 (5H, m, 2H-2, 2H-3 and H-6eq), 2.13 (1H, br
d, J = 12.6 Hz, H-5), 2.27 (1H, m, H-1eq), 2.86 (3H, m,
2H-7 and H-15), 6.83 (1H, s, H-14), 7.01 (1H, dd,
J12,14 = 1.2 and J12,11 = 8.1 Hz, H-12), 7.10 (1H, d,
J11,12 = 8.1 Hz, H-11); EIMS m/z (relative intensity, %):
320 [M+2]+ (2.0), 318 [M]+ (6.6), 303 [MꢁCH3]+, (12.0),
255 [MꢁCOCl]+ (35.0), 43 [CH(CH3)2)]+ (100).
3300 (NH), 3080, 2930, 1630 (C@O), 1467, 1380, 1254 (Si–
C), 1058 (Si–O–C), 830 (Si–O), 800 (Si–C); 1H NMR
(CDCl3): d 0.54 (6H, sl, (CH3)2Si), 0.81 (2H, m, H-110-
Si), 1.21 (6H, d, J = 6.9 Hz, CH3-16 and 17), 1.22 (3H, s,
CH3-20), 1.26 (3H, s, CH3-19), 1.27 (12H, s, H-30–80),
1.54 (4H, m, H-20, H-1ax and H-6ax), 1.75 (4H, m, 2H-2
and 2H-3), 2.13 (1H, br d, J = 12.6 Hz, H-5), 2.32 (1H,
br d, J = 12.6 Hz, H-1eq), 2.84 (3H, m, H-15 and H-7),
3.22 (2H, m, H-10), 5.72 (1H, br s, NH, D2O exchange),
6.86 (1H, d, J14,12 = 1.2 Hz, H-14), 7.00 (1H, dd,
J12,14 = 1.2 and J12,11 = 8.1 Hz, H-12), 7.16 (1H, d,
J11,12 = 8.1 Hz, H-11); 13C NMR (CDCl3): d 39.45 (C-1),
18.36 (C-2), 36.87 (C-3), 46.79 (C-4), 45.14 (C-5), 20.68
(C-6), 26.59 (C-7), 134.30 (C-8), 146.60 (C-9), 37.58 (C-10),
123.65 (C-11), 123.45 (C-12), 145.27 (C-13), 126.46
(C-14), 33.04 (C-15), 23.58 (C-16 and C-17), 177.84 (C-18),
16.13 (C-19), 24.84 (C-20), 36.66 (C-10), 29.61 (C-20),
29.24 (C-30 and C-40), 29.03 (C-50), 28.93 (C-60), 33.09
(C-80), 22.89 (C-100), 18.02 (C-110), 1.60 (CH3–Si); EIMS
m/z (relative intensity, %): 460 [Mꢁ85]+ (9.0), 454 (61.7),
451 [MꢁHSi(CH3)2Cl]+ (7.8), 452 [MꢁSi(CH3)2Cl]+
(17.4), 438 [MꢁCH2Si(CH3)2Cl]+ (45.5), 282 [MꢁNH2-
(CH2)11Si(CH3)2Cl]+ (8.1), 267 [MꢁNH2(CH2)11Si(CH3)2-
ClꢁCH3]+ (3.3), 255 (28.2), 239 [MꢁCONH2(CH2)11-
Si(CH3)2ClꢁCH3]+ (72.4), 199 (15.9), 185 (40.0), 173 (100),
159 (27.9), 131 (24.0), 117 (15.9), 109 (19.9), 97 (17.3), 95
(18.8), 93 [Si(CH3)2Cl]+ (5.8), 83 (15.2), 81 (18.9), 69 (2.5),
55 (43.8), 43 [CH(CH3)2]+ (53.8), 41 (28.2).
4.3.3. N-(100-Undecenyl)dehydroabietamide, 5. Dehydro-
abietoyl chloride (0.04 g, 0.13 mmol) dissolved in dry tolu-
ene (3 mL) was added to a vigorously stirred solution of
10-undecenylamine (0.45 g, 0.66 mmol) in toluene (3 mL).
Subsequently, the mixture was stirred for 48 h at room
temperature. The solvent was evaporated under reduce
pressure and the crude product (1.9 g) was purified by col-
umn chromatography on silica gel using petroleum ether/
dichloromethane (2:1) as an eluent, affording a colourless
oil of N-(100-undecenyl)dehydroabietamide 5 (0.05 g, 93%
23
yield); ½aꢂD ¼ þ57:7 (c 1, ethanol); IR (NaCl cell, cmꢁ1):
3350 (NH), 3060, 2900–2840, 1640 (C@O, amide), 1530,
1
1460, 1380, 1360, 1270, 900, 810; H NMR (CDCl3): d
1.21 (6H, d, J = 6.9 Hz, CH3-16 and 17), 1.22 (3H, s,
CH3-20), 1.26 (3H, s, CH3-19), 1.28 (12H, s, H-30 to H-
80), 1.51 (4H, m, H-20, H-1ax and H-6ax), 1.73 (5H, m,
2H-2, 2H-3 and H-6eq), 2.05 (2H, q, J = 6.9 Hz, H-90),
2.12 (1H, dd, J5ax6eq = 1.8 and J5ax6ax = 12.3 Hz, H-5),
2.30 (1H, br d, J = 12.9 Hz, H-1eq), 2.87 (3H, m, 2H-7
and H-15), 3.24 (2H, m, H-10), 4.88 (1H, br d,
0
0
J11 ;10 ðcisÞ ¼ 10:2 Hz,
CH2@CH),
4.90
(1H,
dd,
4.3.5. Chemically bonded CSP 1 and HPLC column
packing. A solution of 6 (1 g, 1.84 mmol) in dry toluene
(8 mL) was added dropwise to silica gel (2.063 g, Nucleosil
100-5) previously dried at 120 ꢁC under high vacuum for
30 h. The mixture was gently stirred at reflux. The solvent
was removed by evaporation under reduce pressure and the
modified silica was kept in a high vacuum oven at 100 ꢁC
for 24 h. The bonded phase was then washed and filtered
successively with toluene, methanol, acetone and n-hexane.
Elemental analysis of modified silica gel CSP 1 (C, 16.33;
H, 2.60; N, 0.30) showed a loading of 0.22 mmol of the chi-
ral selector (based on C) or 0.24 mmol (based on N) per
gram of stationary phase. The bonded phase was slurried
in methanol and packed in a 100 · 4.6 mm ID stainless-
steel HPLC column using a conventional slurry packing
method. A solution of 2 mL of hexamethyldisilazane in
50 mL of dichloromethane was eluted through the column
to endcapping the remaining free silanol groups of the
bonded phase. Then the unreacted hexamethyldisilazane
was removed out washing the column with 100 mL of
dichloromethane.33
0
J11 ;100ðtransÞ ¼ 17:1 and Jgem = 1.5 Hz, CH2@CH), 5.82
(2H, m, H-100 and N–H, D2O exchange), 6.86 (1H, d,
J14,12 = 1.2 Hz, H-14), 7.00 (1H, dd, J12,14 = 1.2 and
J12,11 = 8.1 Hz, H-12), 7.16 (1H, d, J11,12 = 8.1 Hz, H-11);
13C NMR (CDCl3): d 39.76 (C-1), 18.73 (C-2), 37.26 (C-
3), 47.14 (C-4), 45.54 (C-5), 21.04 (C-6), 26.91 (C-7),
134.58 (C-8), 146.97 (C-9), 37.96 (C-10), 123.98 (C-11),
123.81 (C-12), 145.61 (C-13), 126.81 (C-14), 33.40 (C-15),
23.92 (C-16 and C-17), 178.12 (C-18), 16.48 (C-19), 25.17
(C-20), 37.03 (C-10), 29.96 (C-20), 29.60 (C-30), 29.43 (C-
40), 29.35 (C-50), 29.22 (C-60), 29.05 (C-70), 28.87 (C-80),
33.74 (C-90), 139.10 (C-100), 114.09 (C-110); EIMS m/z
(relative intensity, %): 451 [M]+ (88.1), 437 [Mꢁ14]+
(87.0), 436 [Mꢁ15]+ (6.2%), 282 (8.3), 255 (23.8), 267 [Mꢁ
NH2(CH2)9CHCH2ꢁCH3]+ (5.0), 239 [MꢁCONH2(CH2)9-
CHCH2ꢁCH3]+ (82.3), 225 [MꢁCONH(CH2)9CHCH2]+
(23.8), 199 (15.6), 185 (36.4), 173 (100), 159 (27.5), 131
(25.1), 109 (23.4), 81 (18.4), 69 (33.3), 55 (39.4), 43 (30.0),
41 (20.3); HRMS (ESI) m/z: 452.388 [M+H]+ (calculated
for C31H49NO, 452.389).
4.3.4.
N-[110-(Chlorodimethylsilyl)undecyl]dehydroabiet-
4.4. Synthesis of CSP 2
amide, 6. A solution of 5 (0.12 g, 0.27 mmol) in 5 mL dry
dichloromethane was added to a stirred solution of chloro-
platinic acid in propan-2-ol (0.1 mL, 0.13 mmol/mL) and
the mixture was stirred at room temperature. Then, di-
methylchlorosilane (0.3 mL) was added and the mixture
was heated under reflux for 4 h and 30 min. The solvent
and excess silane were eliminated under reduced pressure
leading to the N-[110-(chlorodimethylsilyl)undecyl]de-
hydroabietamide 6 (0.14 g, 99% yield) as a brownish gum,
used without further purification. IR (NaCl cell, cmꢁ1):
12,14-Dinitrodehydroabietic acid 2 was obtained by nitra-
tion of dehydroabietic acid 131 and CSP 2 was prepared
similarly to the procedure described above for CSP 1.
4.4.1. 12,14-Dinitrodehydroabietoyl chloride, 7. Com-
pound 7, prepared following the procedure described in
Section 4.3.2, was obtained as yellow crystals in an 85%
23
yield; mp 157–160 ꢁC (Et2O); ½aꢂD ¼ þ47:5 (c 1, acetone);
IR (KBr, cmꢁ1): 2955, 2870, 1789 (C@O), 1498, 1460,

C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
3261
1
1419, 994, 914; H NMR (CDCl3): d 1.26 (3H, s, CH3-20),
intensity, %): 637 [M+2]+ (1.2), 635 [M]+ (28.1), 619
(25.9), 617 [MꢁH2O]+ (33.6), 601 (30.0), 541 [Mꢁ
HSi(CH3)2Cl]+ (22.2), 525 [Mꢁ(OH+Si(CH3)2Cl)]+
(89.3), 508 (14.6), 329 (6.6), 328 (13.1), 312 (9.4), 289
(1.5), 263 (19.6), 262 (40.4), 247 (26.0), 93 (100), 83
(23.6), 69 (46.8), 55 (60.9), 43 [CH(CH3)2]+ (40.5), 41
(32.8).
1.32 (6H, d, J = 6.9 Hz, CH3-16 and 17), 1.39 (3H, s, CH3-
19), 1.54 (2H, m, H-1ax and H-6ax), 1.88 (5H, m, 2H-2, 2H-
3, H-6eq), 2.26 (1H, br d, J = 13.6 Hz, H-1eq), 2.30 (1H, dl,
J = 12.1 Hz, H-5), 2.82 (2H, m, 2H-7), 3.03 (1H, hept,
J = 6.9 Hz, H-15), 7.54 (1H, s, H-11); EIMS m/z (relative
intensity, %): 410 [M+2]+ (1.8), 408 [M]+ (4.9), 393
[MꢁCH3]+ (14.7), 391 [MꢁOH]+ (36.5), 329 [Mꢁ(CO+
HCl)]+ (55.7), 263 (45.4), 43 [CH(CH3)2]+ (100).
4.4.4. Chemically bonded CSP 2. The covalent linkage of
chiral selector 9 to silica gel was carried out following a
similar procedure described above for CSP 1. Elemental
analysis of CSP 2 (found: C, 14.26; H, 2.30; N, 1.29) gave
a loading of 0.36 mmol of selector (based on C) or
0.31 mmol (based on N) per gram of stationary phase.
The bonded phase was slurried in methanol and packed
in a 150 · 4.6 mm ID stainless steel HPLC column using
a conventional slurry packing method. Endcapping of
modified silica was carried out using the procedure detailed
in Section 4.3.5.
4.4.2. N-(100-Undecenyl)-12,14-dinitrodehydroabietamide,
8. Compound 8, prepared following the procedure de-
scribed in Section 4.3.3 was obtained as a yellow oil in an
23
86% yield; ½aꢂD ¼ þ36:9 (c 1, acetone); IR (NaCl cell,
cmꢁ1): 3367 (NH), 3075, 2927, 2855, 1631 (C@O), 1536
1
(NO), 1466, 1365 (NO), 961, 750, 735; H NMR (CDCl3):
d 1.24 (3H, s, CH3-20), 1.26 (3H, s, CH3-19), 1.28 (12H, s,
H-30–80), 1.31 (6H, d, J = 6.9 Hz, CH3-16 and 17), 1.53
(5H, m, H-1ax, 2H-6 and 2H-20), 1.76 (4H, m, 2H-2 and
2H-3), 2.03 (2H, q, J = 6.9 Hz, H-90), 2.23 (2 H, m, H-1eq
and H-5), 2.85 (2H, m, 2H-7), 3.02 (1H, hept, J =
6.9 Hz, H-15), 3.26 (2H, q, J = 7.2 Hz, H-10), 4.88
4.5. Synthesis of CSP 3
0
0
ð1H; br d; J11 ;10 ðcisÞ ¼ 10:2 Hz, HCH@CH), 4.90 (1H, dd,
4.5.1. 40-(Chlorocarbonyl)-10-friedelan-3a-yl-benzoate, 10.
Friedelan-3a-ol 3a (0.28 g, 0.65 mmol) dissolved in dry
toluene (25 mL) was added to tereftaloyl chloride
(0.599 g, 2.97 mmol) and 4-dimethylaminepyridine (4-
DMAP) (0.125 g, 1.03 mmol) and then the mixture was
heated under reflux for 8 h. The toluene was removed
under reduced pressure and the residue was used in the
next reaction without purification.
0
0
J11 ;10 ðtransÞ ¼ 17:1 and Jgem = 1.5 Hz, HCH@CH), 5.82
(2H, m, H-100 and NH, D2O exchange), 7.54 (1H, s, H-
11); 13C NMR (CDCl3): d 37.64 (C-1), 18.32 (C-2), 36.74
(C-3), 46.73 (C-4), 43.54 (C-5), 20.54 (C-6), 24.50 (C-7),
130.62 (C-8), 151.03 (C-9), 37.52 (C-10), 121.21 (C-11),
149.24 (C-12), 139.02 (C-13), 151.87 (C-14), 33.77 (C-15),
20.61 (C-16), 20.64 (C-17), 177.20 (C-18, C@O amide),
16.56 (C-19), 24.90 (C-20), 39.92 (C-10), 29.57, 29.46,
29.37, 29.23, 29.06, 28.95, (7C, C-20 to C-80 of aliphatic
chain), 26.94 (C-90), 139.02 (C-100), 113.99 (C-110); EIMS
m/z (relative intensity, %): 541 [M]+ (19.0), 526 [MꢁCH3]+
(54.3), 524 [MꢁOH]+ (37.8), 329 [Mꢁ(CONH2(CH2)9-
CHCH2+CH3)]+ (19.4), 313 (7.6), 289 (1.7), 275 (13.0),
263 (49.1), 249 (4.8), 238 (24.5), 224 (9.6), 55 (100), 43 [CH-
(CH3)2]+ (72.4), 41 (75.7); HRMS(EI) m/z: 540.343898
[MꢁH]ꢁ (calculated for C31H46N3O5 540.344296).
4.5.2. 40-(N-1000-Undecenylamide)-10-friedelan-3a-yl-benzo-
ate, 11. 10-Undecenylamine (0.133 g, 0.79 mmol) in
5 mL ether was added to a solution of 10 in dry pyridine
(0.5 mL) at 0 ꢁC and the mixture was stirred at room
temperature for 2 h. The solvent was evaporated under
reduced pressure and the white crude product obtained
was purified by column chromatography on silica gel using
dichloromethane/methanol (1%) as the eluent affording
0.211 g of 11 in
a
45% yield. Mp 218–220 ꢁC;
23
4.4.3. N-[110-(Chlorodimethylsilyl)undecyl]-12,14-dinitrode-
hydroabietamide, 9. Compound 9, prepared using the
procedure described in Section 4.3.4 was isolated as a yel-
low oil in a quantitative yield; IR (NaCl cell, cmꢁ1): 3300
(NH), 3075, 2926, 2855, 1631 (CO), 1536 (NO), 1467,
1368 (NO), 1255 (Si–C), 1059 (Si–O–C), 909, 840 (Si–O),
½aꢂD ¼ ꢁ6:4 (c 0.94, CH2Cl2); IR (KBr, cmꢁ1): 3402
(NH), 2926, 2855, 2870, 1715 (C@O ester), 1642 (C@C
and C@O amide), 1545, 1437, 1278; H NMR (CDCl3): d
1
0.83 (3H, d, J = 6.3 Hz, CH3-23), 0.85 (3H, s, CH3-24),
0.89 (3H, s, CH3-25), 0.96 (3H, s, CH3-30), 1.01 (6H, s,
CH3-27 and CH3-29), 1.03 (3H, s, CH3-26), 1.17 (3H, s,
CH3-28), 1.28 (14H, s, H-200 to H-800), 2.04 (2H, dt,
J = 6.5 and 7.3 Hz, H-900), 2.23 (1H, m, H-4), 3.47 (2H,
q, J = 6.7 Hz, H-100), 4.96 (3H, m, H-3 and 2H-1100), 5.84
(1H, m, H-1000), 6.29 (1H, t, J = 5.1 Hz, NH, D2O ex-
change), 7.82 (2H, d, J=8.1 Hz, Ar–H-30 and Ar–H-50),
8.07 (2H, d, J = 8.1 Hz, Ar–H-20 and Ar–H-60); 13C
NMR (CDCl3): d 19.31 (C-1), 32.62 (C-2), 76.40 (C-3),
50.10 (C-4), 38.25 (C-5), 41.28 (C-6), 17.8 (C-7), 52.96
(C-8), 36.97 (C-9), 59.85 (C-10), 35.51 (C-11), 30.51
(C-12), 39.62 (C-13), 38.45 (C-14), 32.75 (C-15), 35.98
(C-16), 29.95, (C-17), 42.75 (C-18), 35.27 (C-19), 28.12
(C-20), 32.29 (C-21), 39.19 (C-22), 10.10 (C-23), 14.50
(C-24), 18.1 (C-25), 18.60 (C-26), 20.10 (C-27), 32.05 (C-28),
31.74 (C-29), 34.96 (C-30), 133.32 (C-10), 126.77 (C-20),
129.65 (C-30), 138.47 (C-40), 129.65 (C-50), 126.77 (C-60),
40.20 (C-100), 29.56, 29.42, 29.33, 29.23, 29.03, 28.84 (C-
1
799 (Si–C), 735; H NMR (CDCl3): d 0.40 (6H, br s, Si–
(CH3)2), 0.83 (2H, m, H-110), 1.29 (3H, s, CH3-19), 1.27
(12H, s, H-30–80), 1.24 (3H, s, CH3-20), 1.30 (6H, d,
J = 7.2 Hz, CH3-16 and 17), 1.56 (5H, m, H-1ax, 2H-6
and H-20), 1.78 (4H, m, 2H-2 and 2H-3), 2.24 (2 H, m,
H-1eq and H-5), 2.75 (2H, m, 2H-7), 3.01 (1H, hept,
J = 6.9 Hz, H-15), 3.22 (2H, q, J = 7.2 Hz, H-10), 6.04
(1H, br s, NH, D2O exchange), 7.55 (1H, s, H-11); 13C
NMR (CDCl3): d 37.63 (C-1), 18.28 (C-2), 36.59 (C-3),
46.76 (C-4), 43.47 (C-5), 19.53 (C-6), 24.47 (C-7), 130.63
(C-8), 151.04 (C-9), 37.42 (C-10), 121.30 (C-11), 149.35
(C-12), 128.33 (C-13), 151.96 (C-14), 32.91 (C-15), 20.62
(C-16), 20.53 (C-17), 177.60 (C-18, C@O amide), 16.58
(C-19), 24.88 (C-20), 40.14 (C-10), 29.50, 29.43, 29.22,
28.94, 26.91 (7C of aliphatic chain, C-20 to C-80), 22.93,
(C-100), 18.94 (C-110), 1.65 (CH3–Si); EIMS m/z (relative

3262
C. Moiteiro et al. / Tetrahedron: Asymmetry 17 (2006) 3248–3264
200–C-700), 26.94 (C-800), 33.74 (C-900), 139.12 (C-1000), 114.09
(C-1100), 166.59 (C@O amide), 165.50 (C@O ester); FAB-
MS (NBA) m/z (relative, intensity %): 729 [M+1]+ (30.9),
559 [MꢁNH(CH2)9CHCH2]+ (9.2), 485 (5.8), 411 (11.0),
374 (6.4), 318 (100), 300 (38.9), 297 (8.9). Elemental analy-
sis calculated for C49H77NO3: C, 80.89; H, 10.69; N, 2.01.
Found: C, 80.83; H, 10.66; N, 1.93.
mised by MM using an appropriated Python in-house
script. The lowest energy structures found for each com-
plex were used as starting geometries in further MD runs
carried out for 40 ns in the gas phase. Frames were saved
every 0.1 ps of simulation giving arise to a trajectory files
containing 400,000 structures. The kinetic energy of the
systems was kept constant coupling the system to a Lange-
vin thermostat with a collision frequency of 1.0 psꢁ1. All
nonbonded interactions were evaluated applying a virtually
4.5.3. N-[110-(Chlorodimethylsilyl)undecyl]-10-friedelan-3a-
yl-benzoate, 12. Compound 12 was prepared using the
procedure reported in Section 4.3.4 and isolated as a yellow
˚
infinite cut-off of 99 A. In order to keep the analyte and the
selector together a weak restraint distance using a force
constant of 5.0 kcal molꢁ1 was applied when the distance
between the chiral centre of the analytes and C-4 chiral car-
1
oil in a quantitative yield. H NMR (CDCl3): d 0.39 (6H,
br s, Si–(CH3)2), 0.82 (3H, d, J = 6.6 Hz, Me-23), 0.84
(3H, s, Me-24), 0.88 (3H, s, Me-25), 0.94 (3H, s, Me-29),
0.99 (6H, s, Me-27 and Me-30), 1.02 (3H, s, Me-26), 1.17
˚
bon from stationary phases was higher than 10.0 A using a
parabolic function as defined in AMBER 8. Otherwise the
selector and the analyte showed a tendency to separate.
(3H, s, Me-28), 1.27 (12H, s, ðCH2Þ00), 3.46 (2H, m, H-
6
100), 4.89 (1H, m, H-3), 6.13 (1H, br s, NH, D2O exchange),
7.82 (2H, d, J = 8.4 Hz, H-20 and H-60), 8.07 (2H, d,
J = 8.4 Hz, H-30 and H-50).
The MD simulations in explicit solvent were carried in
equilibrated cubic boxes composed of n-hexane and
propan-2-ol molecules in a ratio consistent with the
n-hexane/propan-2-ol solvent mixture (80:20 v/v) used in
HPLC separations. The solvent molecules were simulated
using all atom models with parameters taken from the
GAFF force field and ESP atomic charges calculated at
the RHF/6-31G* level.45 An individual cubic box com-
posed of 64 propan-2-ol molecules was built replicating a
single molecular mechanics minimized propan-2-ol mole-
cule with the Vega software.46 The same procedure was
adopted to generate a cubic box having 155 n-hexane sol-
vent molecules. Subsequently, the two boxes were merged
yielding the n-hexane/propan-2-ol mobile phase. This new
cubic box was input into AMBER 8 and minimised by
MM in order to remove bad intermolecular contacts. Then,
the system was heated at 300 K with a NVT ensemble for
200 ps, followed of a NPT molecular dynamics run of
400 ps at an average pressure of 1 atm. At the end of this
4.5.4. Chemically bonded CSP 3. Covalent linkage of chi-
ral selector 12 to silica gel was undertaken using the proce-
dure described in Section 4.3.5. Elemental analysis of CSP
3 (Found: C, 14.71; H, 2.67; N, 0.32) showed a loading of
0.24 mmol of selector (based on C) or 0.23 mmol (based on
N) per gram of stationary phase. The modified silica was
packed into a 100 · 4.6 mm ID stainless-steel HPLC col-
umn and the endcapping of modified silica was carried
out following the procedure described in Section 4.3.5.
4.6. Molecular modelling
The molecular modelling studies were performed through
molecular mechanics (MM) and MD simulations using
the AMBER 8 software suite41 with parameters taken from
the GAFF force field, of general use for organic mole-
cules.42 Partial atomic charges were calculated using the
AM1-BCC bond charge correction model43 as imple-
mented in AMBER 8.
simulation, the cubic box displayed
a density of
0.607 g cmꢁ3, which is within the range of expected values
for the experimental density of n-hexane/propan-2-ol
solvent mixture (80:20 v/v). Furthermore, the majority of
propan-2-ol solvent molecules are self-assembled in
supramolecular aggregates stabilized by O–Hꢀ ꢀ ꢀO hydro-
gen bonds interactions. Indeed, the equilibrated box exhib-
ited four propan-2-ol molecules separated from others as
well as small isolated solvent clusters composed of three
tetramers and five dimers. The remaining propan-2-ol mol-
ecules were involved in the formation of a large solvent
network. Therefore, in order to obtain a more regular
distribution of the alcohol molecules into the solvent box,
the system was subsequently heated at 500 K for 200 ps
using a NVT ensemble. The last frame saved for this simu-
lation displayed an effective dispersion of solvent molecules
containing only nine dimers and one trimer of propan-2-ol
molecules. This frame was then minimized by molecular
mechanics and subsequently used to solvate separately
the two CSP 1. Compound 17a diastereomeric complexes.
The final cubic boxes composed of one complex sur-
rounded by 62 propan-2-ol and 142 n-hexane molecules
were obtained. Each system was then equilibrated using a
multistage protocol. The equilibration process started with
minimisation of the solvent molecules by MM with 500
steps by the steepest descent method followed by 2000 steps
The structures of the two binaphthol atropisomers 14 were
retrieved from their crystal structures available from Cam-
bridge Structural Data Base44 while the structures of the
amino acids as well as the structures of the chiral selectors
of CSP 1 and CSP 2 were built by manipulation of the
atomic coordinates of the structures of related compounds
deposited on this data base. For example, the selectors
of both chiral stationary phases were generated from
dehydroabietic acid 1 structure with the silica gel support
omitted and the –(CH2)11SiMe2 spacer replaced by a termi-
nal n-propyl group. The starting geometries of all binary
selector–analyte binding associations (diastereomeric
complexes) were obtained by quenched molecular dynam-
ics using the following methodology. The (R) and (S) enan-
tiomers of two racemic amino acids, 17a and 17d, were
docked separately to the selectors of CSP 1 and CSP 2
leading to eight independent diastereomeric structures.
Subsequently, all (R) and (S) complexes were minimised
by MM and subject to MD runs of 2 ns at 2000 K using
a relaxation time step of 1 fs. Frames were collected at
0.2 ps intervals of simulation leading to a trajectory file
containing 20,000 structures, which were then fully mini-

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˚
A
500 kcal molꢁ1
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the system was allowed to relax using an identical mole-
cular mechanics protocol. The equilibration then continued
with the heating of the system at 300 K for 400 ps using
a NPT ensemble and a weak positional restraint of
ꢁ2
5 kcal molꢁ1
A
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˚
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Bond lengths involving hydrogen bonded atoms were con-
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