Phe-ala-based diazaspirocyclic lactam as nucleator of type II0 β-Turn

Article abstract of DOI:10.1021/jo1019927

The synthesis of a novel Phe-Ala dipeptide mimic, built up on a diazaspirocyclic lactam core, is presented. This new scaffold was evaluated for conformational mimicry of reverse turn by combining molecular modeling, IR, NMR, and X-ray diffraction experime

Full text of DOI:10.1021/jo1019927

pubs.acs.org/joc
Phe-Ala-Based Diazaspirocyclic Lactam as Nucleator of Type II0 β-Turn
Alessandro Sacchetti,*,† Alessandra Silvani,*,‡ Giordano Lesma,‡ and Tullio Pilati§
†Politecnico di Milano, Dipartimento di Chimica, Materiali ed Ingegneria Chimica ‘Giulio Natta’, via
‡
Mancinelli 7, 20131 Milano, Italy, Dipartimento di Chimica Organica e Industriale, Universita degli Studi di
ꢀ
Milano, via G. Venezian 21, 20133 Milano, Italy, and §Istituto di Scienze e Tecnologie Molecolari - CNR,
Via Golgi 19, 20133 Milano, Italy
alessandro.sacchetti@polimi.it
Received October 13, 2010
The synthesis of a novel Phe-Ala dipeptide mimic, built up on a diazaspirocyclic lactam core, is
presented. This new scaffold was evaluated for conformational mimicry of reverse turn by combining
molecular modeling, IR, NMR, and X-ray diffraction experiments. All these tools agree on the
presence of a strong intramolecular hydrogen bond, thus demonstrating the ability of this spiro
compound to act as a type II0 β-turn inducer.
Introduction
spirocyclic scaffolds4 have attracted attention as privileged
structures, able to provide, upon the attachment of appro-
priate functional groups, useful high-affinity ligands, rele-
vant to the field of drug discovery. The polysubstituted
central atom common to the rings of spiro compounds
confers on the overall molecular framework unique 3D
properties that were early on recognized by medicinal chem-
ists. For instance, in the simplest two-ring system, the
almost mutually orthogonal position of the rings can be
used to mimic structural motifs found in proteins, thus
increasing the probability of interactions with biological
systems. In particular, making reference to β-turns, proline-
based spirocyclic lactams have proven to be very effec-
tive nucleators of type II-II0 β-turn conformations.5
The importance of reverse turn for the biological proper-
ties of peptides has in recent years spurred the search for new
scaffolds able to influence the secondary structure when
placed into a peptide chain.1 One of the most studied reverse
motifs is the β-turn, a four amino acid fragment capable of
inverting the chain direction, which is often stabilized by an
intramolecular hydrogen bond between the carbonyl oxygen
of the first residue (i) and the amide proton of the fourth
one (i þ 3).2 Among the many proposed β-turn mimics,3
(1) For reviews, see: (a) Ohfune, Y.; Shinada, T. Eur. J. Org. Chem. 2005,
24, 5127–5143. (b) Cheng, R. P.; Gellman, S. H.; DeGrado, W. F. Chem. Rev.
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Abbenante, G.; Fairlie, D. P. Chem. Rev. 2005, 105, 793–826. (c) Ball,
J. B.; Hughes, R. A.; Alewood, P. F.; Andrews, P. R. Tetrahedron 1993,
49, 3467–3478.
(3) For selected references about reverse-turn surrogates, see: (a) Beierle,
J. M.; Horne, W. S.; van Maarseveen, J. H.; Waser, B.; Reubi, J. C.; Ghadiri,
M. R. Angew. Chem., Int. Ed. 2009, 48, 4725–4729. (b) Molteni, M.; Bellucci,
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2005, 1031–1048.
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R. K. Tetrahedron 2006, 62, 779–828. (b) Van Rompaey, K.; Ballet, S.;
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DOI: 10.1021/jo1019927
r
Published on Web 01/11/2011
J. Org. Chem. 2011, 76, 833–839 833
2011 American Chemical Society

Sacchetti et al.
JOCArticle
FIGURE 1. Proposed β-turn inducer 1 and the derived tetrapeptide
mimic 2.
Recently, type II β-turn mimics based upon L-Pro-containing
spirocyclic scaffolds have served for the synthesis of new Pro-
Leu-Gly (PLG) peptidomimetics with efficient dopamine
receptor modulating activities.6 Among benzo-condensed
compounds, benzazepinones-based spirocyclic derivatives,
bearing mainly β-turn conformations, were recently reported
as potent and selective μ-opioid agonists.7
FIGURE 2. Structures submitted to computer-aided conforma-
tional analysis.
Following our ongoing interest in the design and synthesis
of new β-turn mimics,8 we report here on the preparation of
an unprecedented diazaspirocyclic peptidomimetic capable
of efficiently stabilizing a type II0 β-turn. The stereogenic
spiro carbon atom is common to a tetrahydroisoquinoline
and to a piperidin-2-one ring and enables the construction of
a benzo-condensed 1,8-diazaspiro[5.5]undecan-7-one ring
system. The tetrahydroisoquinoline-3-carboxylic acid (Tic),
a conformationally constrained mimic of Phe, is a key
element in several peptide-based drugs, since its incorpora-
tion itself not only restricts the conformational freedom of
the aromatic ring but also places constraints on the peptide
backbone.9 We reasoned that elaborating the Tic into a
diazaspyrocyclic structure would lead to an efficient β-turn
nucleator. For investigation purposes, we selected Ala as the
(i þ 2) amino acid residue. The target β-turn inducer would
consist of the Phe-Ala mimic 1 (Figure 1). The derived 2
could be considered as a simplified tetrapeptide model, in
which the N-terminal amino acid (i) has been substituted by
an acetyl group and the C-terminal residue (i þ 3) by an N-
methyl moiety.
TABLE 1. Results from Computer-Aided Conformational Analysisa
no. of conf
compd <6 kcal/mol
% dR
< 7 A
% |β|
< 30°
% H bond
C1dO---HN4
2a
2b
3a
3b
6
16
23
43
66 (5.37 A) 66 (-15.9°)
44 (6.30 A) 25 (-35.6°)
52 (7.58 A) 69 (-34.7°)
16 (9.93 A) 12 (-178.4°)
33 (present)
0 (absent)
8 (absent)
0 (absent)
aResults are reported as percentage of conformers which meet the
indicated requirement. Values for the global minimum are reported in
parentheses.
the interatomic distance dR were evaluated. In addition, the
presence of the intramolecular hydrogen bond C1dO---HN4,
involved in a 10-membered ring, was assumed as a critical
requirement.10 To support the assumption that the spirocy-
clic moiety would be crucial for the β-turn stabilization, we
also investigated compound 3a,b in which the precursor Phe
amino acid is not constrained into a Tic ring system. Struc-
tures 2a,b and 3a,b were submitted to a computational
procedure11 consisting of an unconstrained Monte-Carlo/
Energy Minimization conformational search using the mo-
lecular mechanics MMFF94 force field12 in vacuo. For each
compound only conformations within 6 kcal/mol of the
global minimum were kept. Results are reported in Table 1
as the percentage of conformers which meet the requirements
for a generic β-turn.
Results and Discussion
The tetrapeptide mimic 2 was first investigated by compu-
tational methods to determine the correct stereochemistry
required to maximize the β-turn propensity.
The best candidate for our purpose was compound 2a,
embodying D-Phe and L-Ala and exhibiting both a low
number of stable conformations and good percentages of
conformers with the desired geometrical requirements. In
particular, with respect to 2b, the presence of D-Phe seems to
promote an intramolecular 10-membered H-bond. Uncon-
strained mimics 3a,b were shown to be less effective in
stabilizing reverse turn conformations, and the minimum
energy conformers for both compounds were not β-turns.
The stability of the H-bond in 2a was also checked by
means of molecular dynamics11 simulations at 300 K (100 ps
with 1.5 fs time step and without any imposed constraint) in
Having prearranged the S configuration on the (i þ 2) Ala
component, we considered the two possible configurations at
the spiro CR2 atom (Figure 2) in model compounds 2a,b
(Figure 2). To assess the presence of a β-turn-like conforma-
tion, the pseudotorsion angle of the amide backbone β and
(6) Raghavan, B.; Skoblenick, K. J.; Bhagwanth, S.; Argintaru, N.;
Mishra, R. K.; Johnson, R. L. J. Med. Chem. 2009, 52, 2043–2051.
€
€
€ ꢁ
(7) Tomboly, C.; Ballet, S.; Feytens, D.; Kover, K. E.; Bories, A.; Lovas,
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S.; Al-Khrasani, M.; Furst, Z.; Toth, G.; Benyhe, S.; R.; Tourwe, D. J. Med.
Chem. 2008, 51, 173–177.
(8) (a) Lesma, G.; Landoni, N.; Sacchetti, A.; Silvani, A. Tetrahedron
2010, 66, 4474–4478. (b) Lesma, G.; Landoni, N.; Pilati, T.; Sacchetti, A.;
Silvani, A. J. Org. Chem. 2009, 74, 8098–8105. (c) Lesma, G.; Landoni, N.;
Sacchetti, A.; Silvani, A. J. Org. Chem. 2007, 72, 9765–9768. (d) Lesma, G.;
Meschini, E.; Recca, T.; Sacchetti, A.; Silvani, A. Tetrahedron 2007, 63,
5567–5578.
(9) (a) Kotha, S.; Khedkar, P. Synthesis 2008, 18, 2925–2928. (b) Chen,
K. X.; Njoroge, F. G.; Pichardo, J.; Prongay, A.; Butkiewicz, N.; Yao, N.;
Madison, V.; Girijavallabhan, V. J. Med. Chem. 2006, 49, 567–574. (c) Wang,
W.; Cai, M.; Xiong, C.; Zhang, J.; Trivedi, D.; Hruby, V. J. Tetrahedron
(10) Hydrogen bonds are defined as nonbonded contacts between a
nitrogen or oxygen and an hydrogen attached to nitrogen or oxygen,
separated by a distance ranging from 1.6 to 2.1 A and making an X-H--Y
(X, Y = N, O) angle >120°.
(11) Conformational analysis and DFT geometry optimization were
performed with the suite Spartan’08 (Wavefunction, Inc.: Irvine, CA).
Molecular dynamics were performed with MacroModel, version 9.7
ꢁ
2002, 58, 7365–7374. (d) Gibson (nee Thomas), S. E.; Guillo, N.; Jones, J. O.;
€
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2002, 37, 379–389.
(Schrodinger, LLC, New York, 2009).
(12) Halgren, T. A. J. Comput. Chem. 1996, 17, 490–519.
834 J. Org. Chem. Vol. 76, No. 3, 2011

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FIGURE 3. Plot of the structure of 2a obtained by MC/MM analysis and then optimized at the B3LYP-6-31G* level and of its superimposition
with a type II0 β-turn model (in green).
SCHEME 1. Synthetic Sequence
vacuo, in chloroform (noncoordinating solvent), and in
water (coordinating solvent). The results confirmed the
presence of this H-bond in 53%, 74%, and 21% of samples
in vacuo, chloroform, and water, respectively. The minimum
energy conformer was finally optimized at the B3LYP-6-
31G* level, and the dihedral angles of the backbone amide
formylation of 4 with different reagents (HCO2Et, DMF, or
formylpiperidine) and conditions were unsuccessful. So we
planned to get 6 through alcohol 7 and subsequent Swern
oxidation. The required hydroxymethylene moiety was in-
serted by alkylation of 4 with SEM-Cl to give 8, and subse-
quent removal of the trimehylsilylethyl group with BF3.
Alcohol 7 was obtained in 70% overall yield as the single
expected diastereoisomer. Swern oxidation of the primary
alcohol to aldehyde 6, followed by Wittig olefination with
Ph3PCH3Br and KHMDS at low temperature smoothly af-
fordedolefin5. Treatment of 5with NaOH in MeOH gave acid
9, which was esterified with diazomethane to N-Cbz,R-vinyl-
phenylalanine methylester 10 in quantitative overall yield. To
achieve the tetrahydroisoquinoline ring, the Cbz protecting
group was first removed in acid conditions, and the obtained
amino ester 11 was submitted to Pictet-Spengler cyclization
with p-formaldehyde (30% TFA, CH2Cl2). The reaction
proceeded smoothly to give the Tic derivative 12, from which
the R-vinyl acid 13 was derived in high overall yield by N-
acetylation to 14 and subsequent methylester hydrolysis.
were measured. The found values (Φ(iþ1) = 54.9°; ψ(iþ1)
=
-141.3°; Φ(iþ2) = -96.9°; ψ(iþ2) = 43.9°) pointed out a type
II0 β-turn (Figure 3).
The computational evidence prompted us to undertake the
synthesis of 2a.
Following a reported procedure,13,14 D-Phe was first con-
verted into oxazolidinone 4 (Scheme 1). From this intermedi-
ate, we first meant to achieve vinyl derivative 5 through
aldehyde 6. Actually, all attempts to obtain 6 by direct
(13) Seebach, D.; Fadel, A. Helv. Chim. Acta 1985, 68, 1243.
(14) (a) Scheidt, K. A.; Roush, W. R.; McKerrow, J. H.; Selzer, P. M.;
Hansell, E.; Rosenthal, P. J. Bioorg. Med. Chem. 1998, 6, 2477–2494.
(b) Zydowsky, T. M.; Dellaria, J. F., Jr.; Nellans, H. N. J. Org. Chem.
1988, 53, 5607–5616.
J. Org. Chem. Vol. 76, No. 3, 2011 835

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SCHEME 2. Synthetic Sequence
FIGURE 4. Relevant NOE contacts from NOESY (400 MHz, 301
K, 2.0 mM solution CDCl3) of 2a.
To achieve the RCM precursor 15, the carboxylic acid 13
was reacted with N-allyl-alanine methylester 16, readily
prepared as described in the literature15 (Scheme 2). Due to
the steric demand of this condensation, the use of classical
coupling agents (BOP-Cl, HATU, PyBrOP, BOP/HOBT)
did not work, and the desired amide 15 could only be
obtained in 45% yield by using PCl5 as the condensing agent.
With the aim of making the condensation step more efficient,
we also tried to employ the primary amine alanine methyl-
ester, in place of the secondary amine N-allyl-alanine
methylester. Condensation proceeded better (74% yield),
but subsequent allylation of the amide nitrogen of the
obtained dipeptide to give 15 could not be achieved, prob-
ably due again to steric hindrance.
Dipeptide 15 was then easily cyclized by a RCM reaction
by using Grubbs’ second generation catalyst, affording the
desired diazaspiro ring system of compound 17. By conver-
sion of the methyl ester into methylamide 18 upon treatment
with methylamine, and double bond reduction under hydro-
gen atmosphere in the presence of palladium, the target
compound 2a could be achieved.
With tetrapeptide mimic 2a in hand, we performed a
thorough conformational study to confirm the presence of
the intramolecular hydrogen bond attesting a β-turn con-
formation. After full characterization by one- and two-
dimensional NMR analyses, variable temperature (VT) 1H
NMR studies were performed on a 2.0 mM CDCl3 solution
of 2a. The chemical shift of the NH amide proton ranged
from 7.79 (T = 218 K) to 7.44 ppm (T = 328 K). These
values, together with the low VT coefficient (-3.2 ppb/K),
support the involvement of this proton in a hydrogen
bond. Further, titration of the CDCl3 2.0 mM solution
with up to 30% of DMSO-d6, a strongly coordinating and
hydrogen bond-acceptor solvent, produced a low varia-
tion of the NH chemical shift (from 7.53 to 7.72 ppm),16
thus highlighting the stability of the intramolecular hydro-
gen bond.
FIGURE 5. Plot of the molecular structure of compound 2a, as
determined by X-ray crystallography. Atomic displacement para-
meters at the 50% probability level (hydrogen atoms omitted: see
the Supporting Information for details). β-Turned conformation
and intramolecular 10-membered H bond are shown.
detected between both hydrogens H1 and the methyl of the
acetamide group, while two distinct NOE interactions were
observed for H11β with the hydrogen bonded NHMe proton,
and for H11R with the methyl of the Ala residue (Figure 4).
All these observations are in agreement with the presence of a
stable hydrogen bonded conformation.
Also the FT-IR spectrum (2.0 mM solution CHCl3),
showing a unique extensive absorption band at 3345 cm-1
ascribable to an hydrogen-bonded NH stretching, further
attests the presence of a stable secondary structure for 2a.
We could also isolate a single crystal of 2a for crystal-
lographic analysis from a 2-propanol solution (Figure 5). We
were pleased to see that even in a potentially H-bond
disrupting solvent such as 2-propanol, compound 2a crystal-
lizes in a β-turn conformation, characterized by a 2.084 A
intramolecular hydrogen bond between the COi oxygen
atom acceptor and the NHiþ3 hydrogen amide donor. The
hydrogen bond directionality defined by the (i)
O
H-N(iþ3)
3 3 3
3 3 3
angle of 160.65° is almost ideal. Also the CR(i) CR(iþ3)
distanceof 5.584A nicelycomplieswithaβ-turnarrangement.
The dihedral angles (Φ(iþ1) = 49.42°; ψ(iþ1) = -135.48°;
Further evidence for the β-turn-like conformation came
from the NOESY spectrum. Selective NOE contacts were
Φ
(iþ2) = -110.07°; ψ(iþ2) = 40.74°), measured in the solid
state, are very similar to the computed values, thus confirming
a type II0 β-turn-like17 structure for 2a.
(15) (a) Hoffmann, T.; Waibel, R.; Gmeiner, P. J. Org. Chem. 2003, 68,
62–69. (b) Reichwein, J. F.; Liskamp, R. M. J. Eur. J. Org. Chem. 2000, 12,
2335–2344.
(16) For DMSO-d6/CDCl3 solutions, chemical shifts were standardized
by reference to residual proton resonance for CHD2CD3SO (2.49 ppm).
(17) Ideal values for type II0 β-turn: Φ(iþ1) = 60°; ψ(iþ1) = -120°;
Φ
(iþ2) = -80°; ψ(iþ2) = 0°.
836 J. Org. Chem. Vol. 76, No. 3, 2011

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Conclusions
washed with 5% aqueous H3PO4 (20 mL), followed by extrac-
tion with CH2Cl2 (2 ꢀ 20 mL). The combined organic extracts
were dried over Na2SO4 and concentrated in vacuo, to afford 6
In this work we described the design and synthesis of the
new Phe-Ala based diazaspirocyclic lactam 2a. Computa-
tional studies and spectroscopic conformational analyses
clearly indicated that this scaffold adopts a type II0 β-turn
conformation. This peptidomimetic, having a Tic core in the
i þ 2 position, represents a good candidate to replace the β-
turn motif in various Phe-containing G-protein-coupled
receptors ligands. Work to illustrate the application of this
novel diazaspirocyclic lactam to the synthesis of analgesic
tetrapeptide mimics is in progress in our laboratory.
(1.99 g, quantitative yield) as a colorless oil. Rf (hexane/EtOAc
1
4:1) 0.36. [R]25 -38.2 (c 1.0, CHCl3). H NMR (400 MHz,
D
CDCl3) δ 9.40 (s, 1 H), 7.50-7.32 (m, 5H), 7.31-7.13 (m, 5H),
5.66 (s, 1H), 5.20 (d, J = 11.7 Hz, 1H), 5.08 (d, J = 11.7 Hz, 1H),
3.72 (d, J = 13.6 Hz, 1H), 3.56 (br s, 1H), 0.56 (s, 9H); 13C NMR
(100 MHz, CDCl3) δ 192.3, 166.4, 155.5, 134.7-127.8 (12C),
96.1, 76.5, 68.6, 37.4, 37.0, 24.5 (3C). HRMS (ESI) calcd for
C23H25NO5 395.1733, found 395.1713.
Compound 5. The phosphonium salt Ph3PCH3Br (2 mmol)
was quickly transferred to a flame-dried flask and purged with
nitrogen; anhydrous distilled toluene (4 mL) was added. The
resulting suspension was magnetically stirred and treated with a
0.5 M KHMDS solution in toluene (3.8 mL, 1.9 mmol), afford-
ing a brilliant yellow suspension. After 90 min, the reaction
mixture was cooled to -78 °C and a solution of the crude
aldehyde 6 (395 mg, 1.0 mmol, 1 equiv) in anhydrous toluene (3
mL) was added dropwise. After 50 min the temperature was
slowly warmed to -40 °C and, after 30 min, to rt. After 1 h at rt,
the reaction mixture was cooled to -20 °C and quenched with 3
N HCl (6 mL) and saturated NH4Cl (6 mL) aqueous solutions.
The reaction mixture was diluted with EtOAc, the organic layer
separated, and the aqueous one extracted twice with the same
solvent. The combined organic solvents were dried over Na2SO4
and evaporated to dryness under reduced pressure The crude
product was purified by flash chromatography (hexane/EtOAc
9:1), affording the desired vinyl derivative 5 (267 mg, 68% yield)
Experimental Section
Compound 8. A solution of 418 (1.69 g, 4.6 mmol) in anhy-
drous THF (30 mL) was treated dropwise with a 0.5 M KHMDS
solution in toluene (12 mL, 6.0 mmol, 1.3 equiv) at -78 °C and
under N2 atmosphere. After 30 min, SEM-Cl (1.63 mL, 9.22
mmol, d 0.94 g mL-1, 2 equiv) was added dropwise to the
reaction mixture and stirring was continued at -78 °C for an
additional 45 min and then for 2 h at room temperature. The
reaction was quenched with 10% aqueous NaHSO3 (30 mL),
and extracted with EtOAc (3 ꢀ 20 mL). The combined organic
extracts were washed with 10% aqueous NaHSO3, saturated
aqueous NaHCO3, and brine, dried over Na2SO4, and concen-
trated in vacuo to afford compound 8 (2.30 g, quantitative yield)
as a colorless oil, which was used without further purification. Rf
(hexane/EtOAc 9:1) 0.41. [R]25D -23.6 (c 1.1, CHCl3). 1H NMR
(400 MHz, CDCl3) δ 7.40 (br s, 5H), 7.24 (br s, 5H), 5.50 (s, 1H),
5.25 (br s, 2H), 3.80-3.72 (m, 1H), 3.49 (d, J=9.0 Hz, 1H),
3.41-3.15 (m, 4H), 0.90-0.46 (m, 2H), 0.53 (s, 9H), -0.02 (s,
9H). 13C NMR (100 MHz, CDCl3) δ 173.2, 155.4, 135.2, 134.6,
130.7 (2C), 128.4-126.9 (8C), 95.6, 71.9, 68.7, 68.3, 67.3, 38.5,
37.0, 24.7 (3C), 17.7, -1.8 (3C). HRMS (ESI) calcd for
C28H39NO5Si 497.2597, found 497.2581.
as an oil. Rf (hexane/EtOAc 9:1) 0.25. [R]25 -80.2 (c 1.1,
D
CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.47-7.35 (4H),
7.30-7.16 (6H), 5.96 (dd, J = 17.3, 10.7 Hz, 1H), 5.54 (s, 1H),
5.31 (d, J = 11.8 Hz, 1H), 5.27 (d, J = 17.3 Hz, 1H), 5.25 (d, J =
10.7 Hz, 1H), 5.16 (d, J = 11.8 Hz, 1H), 3.50 (d, J = 13.7 Hz,
1H), 3.44 (d, J = 13.7 Hz, 1H), 0.50 (s, 9H). 13C NMR (100
MHz, CDCl3) δ 170.9, 155.7, 137.7, 134.9, 130.9, 128.6-127.0
(10C), 116.7, 94.64, 69.0, 67.8, 40.6, 37.3, 24.4 (3C). HRMS (EI)
calcd for C24H27NO4 393.1940, found 393.1948.
Compound 7. To a solution of 8 (2.30 g, 4.6 mmol) in toluene
(35 mL), at 0 °C and under N2 atmosphere, was added
BF3 Et2O (645 μL, 5.1 mmol, d 1.1 g mL-1, 1.1 equiv). The
Compound 9. A solution of 5 (400 mg, 1.0 mmol) in NaOH 1
M (6.0 mL, 6.0 mmol, 6 equiv) and MeOH (6.0 mL) was stirred
at reflux for 8 h, under N2 atmosphere. After cooling to rt, the
mixture was concentrated in vacuo and 5% aqueous H3PO4 (15
mL) was added until pH 1. The mixture was extracted three
times with CH2Cl2. The combined organic extracts were dried
over Na2SO4 and concentrated in vacuo to afford 9 (331 mg,
3
reaction mixture was stirred for 20 h, and then it was quenched
by the addition of a saturated solution of NaHCO3 (30 mL).
The mixture was extracted with Et2O (3ꢀ15 mL), dried over
Na2SO4, andconcentrated. Purification by Biotage FlashCroma-
tography (hexane/EtOAc 9:1) furnished compound 7 (1.28 g,
70% yield) as a colorless oil. Rf (hexane/EtOAc 4:1) 0.12. [R]25
D
quantitative yield) as an oil. Rf (EtOAc/MeOH 9:1) 0.17. [R]25
-43.4 (c 1.0, CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.42 (br s,
5H), 7.23 (br s, 5H), 5.59 (s, 1H), 5.25 (d, J = 12.2 Hz, 1H), 5.22
(d, J = 12.2 Hz, 1H), 4.18 (br d, J = 10.2 Hz, 1H), 3.80 (d, J =
10.7Hz, 1H), 3.20 (d, J = 14.0Hz, 1H), 3.16 (d, J = 13.8Hz, 1H),
1.81 (br s, 1H), 0.64 (s, 9H). 13C NMR (100 MHz, CDCl3) δ
173.7, 155.6, 135.3, 134.7, 130.9-127.3 (10C), 96.1, 70.1, 67.9,
65.6, 38.3, 37.4, 24.9 (3C). HRMS (ESI) calcd for C23H27NO5Na
(M þ Na) 420.1781, found 420.1787.
D
-38.5 (c 1.5, CHCl3). 1H NMR (400 MHz, CDCl3) δ 9.04 (br s,
1H), 7.48-7.27 (m, 5H), 7.26-7.16 (m, 3H), 7.13-7.02 (m, 2H),
6.12 (dd, J = 17.2, 10.6 Hz, 1H), 5.64 (br s, 1H), 5.38 (d, J = 10.0
Hz, 1H), 5.36 (d, J = 17.5 Hz, 1H), 5.23 (d, J = 12.2 Hz, 1H),
5.11 (d, J = 12.2 Hz, 1H), 3.63 (d, J = 13.5 Hz, 1H), 3.40 (d, J =
13.6 Hz, 1H). 13C NMR (100 MHz, CDCl3) δ 175.6, 154.44
136.1, 134.8, 129.8-126.8 (10C), 116.3, 66.6, 64.6, 40.3, 27.6.
HRMS (ESI) calcd for C19H19NO4 325.1314, found 325.1323.
Compound 10. At 0 °C, under N2 atmosphere, a 2.0 M
TMSCHN2 solution in Et2O (4.2 mL, 8.31 mmol, 4 equiv) was
added dropwise to a stirred solution of 9 (675 mg, 2.08 mmol, 1
equiv) in MeOH (8 mL). After 1 h the reaction mixture was
concentrated in vacuo, affording 10 (706 mg, quantitative yield)
Compound 6. A solution of DMSO (1.0 mL, 14.1 mmol, d 1.1 g
mL-1, 3 equiv) in 2 mL of anhydrous CH2Cl2 was added
dropwise, at -78 °C and under N2 atmosphere, to a solution
of (COCl)2 (820 μL, 9.7 mmol, d 1.5 g mL-1, 2 equiv) in 8 mL of
anhydrous CH2Cl2. The mixture was stirred at -78 °C for 10
min. A solution of 7 (1.91 g, 4.8 mmol, 1 equiv) in 18 mL of
anhydrous CH2Cl2 was added, and the reaction mixture was
stirred at -78 °C and under N2 atmosphere for 1.5 h. Et3N (2.7
mL, 19.3 mmol, d 0.7 g mL-1, 4 equiv) was added and the
temperature was slowly warmed to rt. After 2 h of stirring at rt,
the reaction mixture was diluted with 15 mL of CH2Cl2 and
as an oil. Rf (EtOAc/MeOH 9:1) 0.69. [R]25 -35.6 (c 1.1,
D
1
CHCl3). H NMR (400 MHz, CDCl3) δ 7.47-7.32 (m, 5H),
7.27-7.15 (m, 3H), 7.06-6.97 (m, 2H), 6.10 (dd, J = 17.3, 10.6
Hz, 1H), 5.70 (br s, 1H), 5.33 (d, J = 10.4 Hz, 1H), 5.31 (d, J =
17.4 Hz, 1H), 5.23 (d, J = 12.3 Hz, 1H), 5.11 (d, J = 12.3 Hz,
1H), 3.79 (s, 3H), 3.66 (d, J = 13.5 Hz, 1H), 3.34 (d, J = 13.5 Hz,
1H). 13C NMR (100 MHz, CDCl3) δ 171.7, 154.3, 136.6, 135.4,
129.9-126.9 (11C), 116.1, 66.5, 65.2, 52.8, 40.4. HRMS (ESI)
calcd for C20H21NO4 339.1471, found 339.1466.
(18) Compound 4 was prepared according to ref 14. 1H and 13C NMR
spectra are in accordance with literature. [R]25D þ 10.4 (c 1.0, CHCl3).
J. Org. Chem. Vol. 76, No. 3, 2011 837

Sacchetti et al.
(d, J = 14.8 Hz, 1H), 3.05 (d, J = 14.9 Hz, 1H), 2.32 (s, 3H). 13
NMR (100 MHz, CDCl3) δ 174.9, 170.5, 135.6, 133.3, 133.2,
127.8-125.1 (4C), 114.1, 65.3, 48.0, 38.2, 22.3. HRMS (ESI)
calcd for C14H15NO3 245.1052, found 245.1056.
JOCArticle
Compound 11. Under N2 atmosphere, a 33% HBr solution in
AcOH (3 mL) was added dropwise to a stirred solution of 10
(705 mg, 2.08 mmol) in AcOH (4.2 mL). After 36 h, the reaction
mixture was diluted with H2O (15 mL), aqueous NaHCO3
solution was added until pH 9, and the mixture was extracted
three times with CH2Cl2. The combined organic extracts were
dried over Na2SO4 and concentrated in vacuo to afford 11 (406
C
Compound 15. At 0 °C, under N2 atmosphere, a solution of 13
(308 mg, 1.26 mmol) in anhydrous CH2Cl2 (7 mL) was added to
a solution of PCl5 (262 mg, 1.26 mmol, 1 equiv) in anhydrous
CH2Cl2 (6 mL). After stirring at 0 °C for 10 min and at rt for 3 h,
the reaction mixture was cooled to -10 °C and added to a
suspension of 16 (232 mg, 1.29 mmol, 1 equiv) in dry pyridine
(1.65 mL, 20.43 mmol, d 1.0 g mL-1, 16 equiv) and anhydrous
CH2Cl2 (12 mL). After stirring at -10 °C for 10 min, and at rt for
20 h, water (25 mL) was added, the organic layer was washed
with saturated aqueous NaHCO3 solution, water, and 2.5%
aqueous H3PO4 (15 mL), dried over Na2SO4, and concentrated
in vacuo. The crude was purified by flash chromatography
(EtOAc/hexane 9:1), affording 15 (209 mg, 45% yield) as a
foam. Rf (AcOEt/MeOH 9:1) 0.48. [R]25D -14.0 (c 0.4, CHCl3).
1H NMR (400 MHz, CD3CN, 77 °C) δ 7.36-7.13 (m, 4H), 6.37
(dd, J = 17.3, 10.8 Hz, 1H), 5.91 (m, 1H), 5.35 (d, J = 17.7 Hz,
1H), 5.21 (d, J = 10.5 Hz, 1H), 5.96 (d, J = 10.8 Hz, 1H), 4.81
(br d, J = 17.3 Hz, 2H), 4.61 (br d, J = 15.2 Hz, 2H), 4.36 (br d,
J = 14.7 Hz, 1H), 3.98 (br d, J = 15.9 Hz, 1H), 3.66 (s, 3H), 3.41
(d, J = 15.9 Hz, 1H), 3.12 (d, J = 15.9 Hz, 1H), 2.13 (s, 3H), 1.44
(d, J=6.9 Hz, 3H). 13C NMR (100 MHz, CD3CN, 77 °C) δ
177.5, 173.0, 171.4, 137.7, 135.6, 133.4, 132.24, 130.2-126.5
(4C), 117.4, 115.5, 60.8, 54.8, 51.9, 50.8, 48.4, 37.2, 23.3, 14.2.
HRMS (ESI) calcd for C21H26N2O4 370.1893, found 370.1853.
Compound 17. Under N2 atmosphere, 1,3-bis(2,4,6-trime-
thylphenyl)-2-(imidazolidinylidene)(dichlorophenylmethylene)-
(tricyclohexylphosphine)ruthenium (Grubbs second generation
catalyst) (32 mg, 0.04 mmol, 0.2 equiv) was added to a solution of
15 (69 mg, 0.19 mmol) in anhydrous toluene (6 mL). The reaction
mixture was stirred at reflux for 4 h, then concentrated in vacuo.
The crude was purified by flash chromatography (EtOAc/MeOH
mg, 95% yield) as an oil. Rf (EtOAc/hexane 1:1) 0.23. [R]25
D
-21.5 (c 1.5, CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.34-7.22
(m, 3H), 7.21-7.15 (m, 2H), 6.20 (dd, J = 17.3, 10.6 Hz, 1H),
5.37 (d, J = 17.2 Hz, 1H), 5.22 (d, J = 10.6 Hz, 1H), 3.75 (s, 3H),
3.30 (d, J = 13.2 Hz, 1H), 2.88 (d, J = 13.2 Hz, 1H), 1.87 (br s,
2H). 13C NMR (100 MHz, CDCl3) δ 174.8, 139.9, 135.4,
129.7-126.7 (5C), 114.1, 63.3, 51.9, 45.5. HRMS (ESI) calcd
for C12H15NO2 205.1103, found 205.1102.
Compound 12. Under N2 atmosphere, (CH2O)n (80 mg, 2.53
mmol, 1.8 equiv) was added to a solution of 11 (283 mg, 1.38
mmol) in anhydrous CH2Cl2 (8 mL). The resulting suspension
was treated with TFA (3.5 mL) and magnetically stirred for 24 h.
A saturated aqueous solution of NaHCO3 was added until pH 8.
The reaction mixture was extracted three times with CH2Cl2.
The combined organic extracts were dried over Na2SO4 and
concentrated in vacuo. The crude was purified by flash chro-
matography (EtOAc/hexane 3:7), affording 12 (290 mg, 97%
yield) as a foam. Rf (EtOAc/hexane 1:1) 0.23. [R]25D -23.9 (c 1.3,
1
CHCl3). H NMR (400 MHz, CDCl3) δ 7.22-7.09 (m, 3H),
7.07-6.99 (m, 1H), 5.98 (dd, J = 17.4, 10.7 Hz, 1H), 5.35 (d, J =
17.4 Hz, 1H), 5.28 (d, J = 10.7 Hz, 1H), 4.18 (d, J = 16.3 Hz,
1H), 4.10 (d, J = 16.0 Hz, 1H), 3.72 (s, 3H), 3.37 (d, J = 16.1 Hz,
1H), 3.03 (d, J = 16.1 Hz, 1H), 2.75 (br s, 1H). 13C NMR (100
MHz, CDCl3) δ 174.2, 139.0, 133.9, 132.7, 129.1-126.3 (4C),
116.7, 62.9, 52.8, 44.8, 36.3. HRMS (ESI) calcd for C13H15NO2
217.1103, found 217.1114.
Compound 14. Under N2 atmosphere, DIPEA (2.2 mL, 12.63
mmol, d 0.7 g mL-1, 4 equiv) was added dropwise to a solution
of 12 (686 mg, 3.15 mmol) in anhydrous CH2Cl2 (25 mL). The
reaction mixture was cooled to 0 °C and Ac2O (1.2 mL, 12.63
mmol, d 1.1 g mL-1, 4 equiv) was added dropwise. The
temperature was slowly warmed to rt and the mixture was
magnetically stirred for 20 h, then at reflux for 10 h. The reaction
mixture was diluted with H2O (20 mL), 5% aqueous H3PO4 (20
mL) was added, and the mixture was extracted with CH2Cl2 (3 ꢀ
30 mL). The combined organic extracts were washed with sat.
aqueous NaHCO3, dried over Na2SO4, and concentrated in
vacuo. The crude was purified by flash chromatography
(EtOAc/hexane 2:3), affording 14 (747 mg, 91% yield) as a foam.
95:5), affording 17 (58 mg, 91% yield) as an oil. Rf (EtOAc/
1
MeOH 9:1) 0.26. [R]25 -24.3 (c 0.1, CHCl3). H NMR (400
D
MHz, CDCl3) δ 7.38-7.13 (4H), 5.70 (dt, J = 10.1, 3.2 Hz, 1H),
5.13 (dt, J = 10.1, 2.0 Hz, 1H), 4.68 (d, J = 14.2 Hz, 1H), 4.59 (d,
J = 14.2 Hz, 1H), 4.26 (q, J = 7.1 Hz, 1H), 4.16 (br d, J = 15.4
Hz, 1H), 3.97 (br d, J = 16.1 Hz, 1H), 3.77 (s, 3H), 3.55 (d, J =
14.5 Hz, 1H), 2.88 (d, J = 14.5 Hz, 1H), 2.21 (s, 3H), 1.56 (d, J =
7.1 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ 170.9, 169.1, 168.8,
134.0, 133.9, 127.9-126.6 (4C), 125.2, 120.4, 59.0, 55.5, 51.8,
48.0, 47.51, 40.5, 22.6, 12.9. HRMS (ESI) calcd for C19H22N2O4
342.1580, found 342.1572.
Rf (EtOAc/hexane 8:2) 0.25. [R]25 þ17.5 (c 1.0, CHCl3). H
Compound 18. At 0 °C, under N2 atmosphere, 17 (57 mg, 0.17
mmol) was dissolved in a 8.0 M MeNH2 solution in EtOH (5
mL). The reaction mixture was stirred for 4 h at rt, then
evaporated in vacuo. The crude was purified by flash chroma-
tography (EtOAc/MeOH 9:1), affording 18 (51 mg, 90% yield)
as a foam. Rf (EtOAc/MeOH 85:15) 0.38. [R]25D -146.5 (c 1.5,
CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.60 (br s, 1H),
7.37-7.13 (m, 4H), 5.73 (dt, J = 10.2, 3.2 Hz, 1H), 5.49 (q,
J = 7.4 Hz, 1H), 5.13 (dt, J = 10.2, 2.0 Hz, 1H), 4.72 (d, J =
14.2 Hz, 1H), 4.63 (d, J = 14.2 Hz, 1H), 3.97 (dt, J = 18.0, 2.6,
1H), 3.75 (dt, J = 18.0, 2.6 Hz, 1H), 3.60 (d, J = 14.5, 1H), 2.90
(d, J = 14.5 Hz, 1H), 2.80 (d, J = 4.6 Hz, 3H), 2.24 (s, 3H), 1.47
(d, J = 7.4 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ 170.4,
169.8, 168.5, 133.5, 133.5, 127.9-126.9 (4C), 125.4, 120.3, 59.2,
51.4, 48.3, 43.3, 40.5, 26.1, 22.9, 13.3. HRMS (EI) calcd for
C19H23N3O3 341.1739, found 341.1726.
1
NMR (400 MHz, CDCl3) δ 7.32-D7.18 (m, 3H), 7.18-7.06 (m,
1H), 6.27 (dd, J = 17.3, 10.6 Hz, 1H), 4.90 (d, J = 10.6 Hz, 1H),
4.69 (d, J = 14.6 Hz, 1H), 4.64 (d, J = 14.3 Hz, 1H), 4.60 (d, J =
17.2 Hz, 1H), 3.77 (s, 3H), 3.28 (d, J = 14.8 Hz, 1H), 3.01 (d, J =
14.8 Hz, 1H), 2.27 (br s, 1H). 13C NMR (100 MHz, CDCl3) δ
172.5, 169.1, 135.6, 133.5, 133.1, 127.6-125.2 (4C), 113.8, 64.7,
52.1, 47.8, 38.0, 22.2. HRMS (EI) calcd for C15H17NO3 259.1208,
found 259.1197.
Compound 13. At 0 °C, under N2 atmosphere, a 2.0 M LiOH
solution in H2O (7.44 mL, 14.88 mmol, 6 equiv) was added to a
solution of 14 (644 mg, 2.48 mmol) in THF (30 mL) and MeOH
(4 mL). The reaction mixture was stirred at reflux for 8 h, then
concentrated in vacuo, diluted with H2O (20 mL), and extracted
with EtOAc (2 ꢀ 20 mL). A 5% aqueous H3PO4 solution (20
mL) was added to the aqueous layer and the mixture was
extracted with EtOAc (2 ꢀ 20 mL), washed with brine, dried
over Na2SO4, and concentrated in vacuo to afford 13 (554 mg,
91% yield) as a foam. Rf (EtOAc/MeOH 9:1) 0.01. [R]25D þ3.2
(c 1.3, CH3OH). 1H NMR (400 MHz, CDCl3) δ 9.77 (br s, 1H),
7.41-6.98 (m, 4H), 6.26 (dd, J = 17.3, 10.7 Hz, 1H), 4.93 (d,
J = 10.7 Hz, 1H), 4.66 (br s, 2H), 4.63 (d, J = 17.5 Hz, 1H), 3.36
Compound 2a. To a solution of 18 (30 mg, 0.09 mmol) in
MeOH (2 mL) was added 10% Pd/C (6 mg) and the reaction was
stirred at rt under H2 atmosphere for 24 h. The suspension was
then filtered through a pad of Celite and evaporated under
reduced pressure to afford 2a (30 mg, quantitative yield) as a
white solid. Rf (AcOEt/MeOH 85:15) 0.38. [R]25D -42.4 (c 1.3,
838 J. Org. Chem. Vol. 76, No. 3, 2011

Sacchetti et al.
JOCArticle
CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.54 (br s, 1H),
7.39-7.19 (m, 4H), 5.48 (q, J = 7.0 Hz, 1H), 4.69 (d, J =
14.2 Hz, 1H), 4.59 (d, J = 14.2 Hz, 1H), 3.38 (d, J = 14.8 Hz,
1H), 3.33-3.20 (m, 2H), 3.02 (d, J = 14.8 Hz, 1H), 2.83 (d, J =
4.3 Hz, 3H), 2.25 (s, 3H), 2.10-1.97 (m, 1H), 1.90-1.74 (m,
2H), 1.51-1.44 (m, 1H), 1.41 (d, J=7.0 Hz, 3H). 13C NMR
(100 MHz, CDCl3) δ 172.4, 171.1, 170.2, 134.7, 134.0,
128.5-126.2 (4C), 62.2, 51.7, 49.2, 41.5, 37.9, 31.3, 26.7,
23.7, 20.8, 13.8. HRMS (EI) calcd for C19H25N3O3 343.1896,
found 343.1859.
Supporting Information Available: General procedures and
copy of NMR spectra for all compounds; VT NMR studies,
DMSO-d6 titrations, IR and crystallographic data for com-
pound 2a; computational details. This material is available free
of charge via the Internet at http://pubs.acs.org.
J. Org. Chem. Vol. 76, No. 3, 2011 839