Stereoconservative synthesis of orthogonally protected γ-functionalized amino acids using N-tritylserine derivatives

Full text of DOI:10.1021/jo960624g

J . Org. Chem. 1996, 61, 6067-6070
6067
Sch em e 1. Nu cleop h ilic Su bstitu tion of th e
Ster eocon ser va tive Syn th esis of
Or th ogon a lly P r otected γ-F u n ction a lized
Am in o Acid s Usin g N-Tr itylser in e
Der iva tives
Lea vin g Gr ou p (LG) by a Selection of
Ma lon a te-Rela ted Nu cleop h iles
Christophe Dugave* and Andre´ Menez
CEA/ Saclay, De´partement d’Inge´nierie et
d’Etude des Prote´ines (DIEP) Baˆtiment 152,
F91191 Gif-sur-Yvette, France
Received April 3, 1996
Use of commercial optically pure R-amino acids as a
“chiral pool” is a conceptually attractive approach that
has known much progress and applications for the
elaboration of unusual and non-natural amino acids1 and
many other interesting compounds.2 Several synthetic
equivalents for â-alanyl cation have been developed for
this goal. However, the ring-opening of cyclic electro-
philes such as serine â-lactone,3 aziridines,4 or serine
sulfamidate5 by soft basic nucleophiles such as malonates
was unfruitful or difficult.6 Despite interesting applica-
tions with organocuprates,7 attempted substitution on to
â-haloalanines by malonates failed and gave the corre-
sponding dehydroalanine.8 One way to overcome these
difficulties may be to use open-chain electrophilic serine
derivatives and to block the amine moiety with an
appropriate bulky protecting group such as N-(triphenyl-
methyl)9 or N-(phenylfluorenyl).10 Very recently (1996),
Cherney and Wang demonstrated that the Mitsunobu
reaction can be performed efficiently with N-trityl and
N-(phenylfluorenyl)serine esters.9b The N-trityl group
prevents R-proton abstraction by non-nucleophilic bases
and protects the R-ester from saponification.11 Moreover,
this group is resistant toward strong bases and most
reductive agents but can be easily removed under mildly
acidic conditions or by hydrogenolysis. Thus, the well-
known functional diversity of malonate compounds could
be readily exploited for generating unusual and non-
natural amino acids especially in the glutamate series.
We report herein on the synthesis of a variety of
orthogonally protected R-amino acids, functionalized at
the γ-position, using N-tritylserine derivatives.
The nucleophilic displacement on mesylate 1a and
iodides 2a and 2b by malonate related anions was
investigated. The starting compounds 1a and 2a /b were
prepared in almost quantitative yields from the easily
available N-tritylserine methyl and benzyl esters (Scheme
1). These reagents can be stored at -18 °C for several
months, without detectable degradation.12 Initially, we
studied the reactivity of these compounds toward sodium
dimethylmalonate under anhydrous conditions (THF-
HMPA). Sterically hindered mesylate 1a did not react
with sodium dimethylmalonate below 50 °C. At higher
temperatures, it cyclized and gave aziridine-2-carboxylate
3a as the only product. On the other hand, all attempts
to use the corresponding triflate (prepared in situ) at low
temperatures failed and led to complex mixtures of
unidentified products. In contrast, compound 2a reacted
with sodium malonates at 30-40 °C and provided an
easily separable mixture of aziridine 3a (30%) and
triester 4a (65%) (Scheme 1, Table 1).
* To whom correspondence should be addressed. Tel.: 33-1-69-08-
52-24. Fax: 33-1-69-08-90-71. E-mail: christophe.dugave@cea.fr.
(1) Williams, R. M. Synthesis of optically active R-amino acids. In
Organic Chemistry Series; Baldwin, J . E., Magnus, P. D., Eds.;
Pergamon Press: New York, 1989; Vol. 7.
(2) Coppola, G. M.; Schuster, H. F. Asymetric Synthesis: Construc-
tion of Chiral Molecules Using Amino Acids; J ohn Wiley & Sons: New
York, 1987.
(3) (a) Arnold, L. D.; May, R. G.; Vederas, J . C. J . Am. Chem. Soc.
1988, 110, 2237-2241 and references cited therein. (b) Ratemi, E. S.;
Vederas, J . C. Tetrahedron Lett. 1994, 35, 7605-7608. (c) Shao, H.;
Wang, S. H. H.; Lee, C.-W.; O¨ sapay, G.; Goodman, M. J . Org. Chem.
1995, 60, 2956-2957.
(4) (a) Nakajima, K.; Okima, K. Bull. Chem. Soc. J pn. 1983, 56,
1565-1566 and references cited therein. (b) Baldwin, J . E.; Adlington,
R. M.; O’Neil, I. A.; Schofield, C.; Spivey, A. C.; Sweeney, J . B. J . Chem.
Soc., Chem. Commun. 1989, 1852-1854. (c) Sato, K.; Kozikowski, A.
P. Tetrahedron Lett. 1989, 4073-4076. (d) Baldwin, J . E.; Spivey, A.
C.; Schofield, C. J . Sweeney, J . B. Tetrahedron 1993, 49, 6309-6330
and references cited therein. (e) Church, N. J .; Young, D. W. Tetra-
hedron Lett. 1995, 36, 151-154. (f) Dure´ault, A.; Tranchepain, I.;
Depezay, J .-C. J . Org. Chem. 1989, 54, 5324-5330. (g) For a recent
review regarding aziridines, see: Tanner, D. Angew. Chem., Int. Ed.
Engl. 1994, 33, 599-619.
To check that basic conditions and heating did not
epimerize the R-carbon, an aliquot of compound 4a was
totally hydrolyzed and decarboxylated (4 h in refluxing
6 N HCl) to glutamic acid. The hydrolysate was quan-
titatively derivatized using (+)-FLEC as previously re-
ported,13 and the enantiomeric excess was determined by
HPLC to be 97% relative to authentic samples of D, L, and
DL-glutamate. Compound 3a was deprotected under mild
(5) Baldwin, J . E.; Spivey, A. C.; Schofield, C. J . Tetrahedron:
Asymmetry 1990, 1, 881-884.
(6) Bouayad, Z.; Chanet-Ray, J .; Ducher, S.; Vessie`re, R. J . Hetero-
cycl. Chem. 1991, 28, 1757-1767.
(7) Bajgrowicz, J . A.; El Hallaoui, A.; J acquier, R.; Pigie`re, C.;
Viallefont, P. Tetrahedron 1985, 41, 1833-1843 and references cited
therein.
(8) (a) Weinstein, B.; Watrin, K. G.; Loie, J .; Martin, J . C. J . Org.
Chem. 1976, 41, 3634-3635. (b) Boggs, N. T., III; Goldsmith, B.;
Gawley, R. E.; Koehler, K. A.; Hiskey, R. G. J . Org. Chem. 1979, 44,
2262-2269.
(11) Dugave, C; Kessler, P.; Colas, C.; Hirth, C. Tetrahedron Lett.
1994, 35, 9199-9202.
(12) Traces of aziridine carboxylate 3a /b were detected after
6
months at -18 °C. 1H and 13C NMR spectra of compounds 2a and 2b
showed considerable doubling of several signals, which suggests the
existence of at least two distinct rotamers. This was not observed for
the more hindered corresponding mesylates 1a /b. See: Son, J .-K.;
Kalvin, D.; Woodard, R. W. Tetrahedron Lett. 1988, 29, 4045-4048.
(13) Einarsson, S.; J osephsson, B.; Mo¨ler, P.; Sanchez, D. Anal.
Chem. 1987, 59, 1191-1195.
(9) (a) Zee-Cheng, R. K-Y.; Olson, R. E. Biochem. Biophys. Res.
Commun. 1980, 94, 1128-1132. (b) Cherney, R. J .; Wang, L. J . Org.
Chem. 1996, 61, 2544-2546.
(10) (a) Koskinen, A. M. P.; Rapoport, H. J . Org. Chem. 1989, 54,
1859-1866. (b) Atfani, M.; Lubell, W. D. J . Org. Chem. 1995, 60, 3184-
3188.
S0022-3263(96)00624-X CCC: $12.00 © 1996 American Chemical Society

6068 J . Org. Chem., Vol. 61, No. 17, 1996
Notes
Ta ble 1. P r od u cts of th e Nu cleop h ilic Atta ck of
Com p ou n d s 2a /b by Ma lon a te-Rela ted An ion sa
Sch em e 2. Syn th esis of Boc- a n d F m oc-P r otected
l-γ-Ca r boxyglu ta m a te a n d L-γ-Cya n oglu ta m a te
fr om N-Tr ityl Com p ou n d s 4a /b, 5b, a n d 6a
3
compd
ee
(a /b)
entry
E1CH2E2
Methodsb (a /b %)
(a /b %)
1
2
3
4
5
6
7
8
9
CH2(CO2Me)2
Ac
A
A
30/22 4a /b (65/74) 97/97
CH2(CO2tBu)2
MeO2CCH2CN
t-BuO2CCH2CN
CH2(CN)2
CH3COCH2CO2tBu
PhO2SCH2CN
Ph2CdNCH2CO2Et
Ph2CdNCH2CN
16
20
12
11
40
50
<5
8
5b (78)
6a (74)
7a (86)
8a (77)
9b (57)
10a (40)
11a (30)
12a (90)
99d
98
98
98
ND
ND
f
A
Ae
A
A
B
C16
f
a
All experiments were carried out on a 1 mmol scale. Yields
are given for silica gel flash chromatography purified products;
ee were calculated from HPLC analysis of FLEC derivatives.
b
Methods: A (NaH/THF-HMPA 80:20, 40 °C, 12 h); B (LDA/
THF-HMPA 80-20, 40 °C, 12 h); C (Bu4NHSO4/10% NaOH-
CH2Cl2, rt 4 h). c Dibenzyl, tert-butyl ethyl, and benzyl methyl
malonates gave similar results. >99% after recrystallization.e At
d
30 °C, 4 h. f Decomposed in refluxing 6 N HCl.
acidic conditions (5% formic acid in dichloroethane)14 and
saponified and was analyzed as described above. For
higher temperatures and prolonged times of reaction, a
serious decrease of yields and enantiomeric excess was
observed, whereas the proportion of optically pure aziri-
dine increased (e.g., 3a , 51% yield, ee ) 100%; 4a , 45%
yield, ee ) 58% after 24 h at 60 °C). This indicates that
substitution and cyclization are competitive reactions and
racemization occurs after the substitution step. More-
over, we confirmed that compound 3a is not an interme-
diate in a cyclization/ring-opening process since it did not
react at 65 °C (2 days) with a 10-fold excess of reagent.
No significant difference in reactivity was observed
between methyl and benzyl esters.
Our study was extended to a series of malonate-related
anions and some other nucleophiles (Table 1). Thus,
malonic-branched amino acids were isolated in moderate
to good yields (30-90%) and typically 97-99% enantio-
meric excess, except for too rigid or too bulky nucleophiles
such as Meldrum’s acid, bis(phenylsulfonyl)methane, and
tetraethylmethylene diphosphonate (data not shown).
With most malonates, the use of a slight excess of sodium
hydride led to traces of the corresponding lactams. As
anticipated, the marked tendency of the malonic center
to epimerize precluded the separation of the diastereo-
mers obtained from unsymmetrical nucleophiles (Table
1, entries 3, 4, 6-9).15 Diaminoglutarate derivative 12a
was obtained in high yield (90%) as a 1:1 mixture of
diastereomers (determined by NMR). Although depro-
tection resulted in a complete degradation of the product,
we assume that the very mild conditions employed (phase
transfer at room temperature for 4 h)16 did not epimerize
the chiral center.17
acidic detritylation14 of triester 4a , protection with di-
tert-butyl dicarbonate and regioselective hydrolysis of the
R-methyl ester by R-Chymotrypsin19 (Scheme 2) afforded
compound 13 (ee > 99%). Although many preparations
of optically pure Gla have been reported in the literature,
most of them require either resolution of racemates20 or
asymetric induction.21 Multistep synthesis of Gla deriva-
tives starting from glutamate,9a pyroglutamate,22 or
proline23 are difficult or relatively more complex than the
route we describe here. In a similar way the 4-cyano-
glutamate 18, which has shown interesting pharmaco-
logical properties,24 was synthesized from compound 6a .
However, in this latter case, diester 16 was isolated in a
disappointing yield (31%) along with the corresponding
lactam 17 (22%) resulting from an intramolecular addi-
tion of the intermediate free amine to the cyano group.
In summary, the nucleophilic substitution of N-trityl-
3-iodoalanine esters 2a /b by malonate, related anions is
an appropriate approach for synthesizing various γ-func-
tionalized amino acids in reasonable yields and almost
without epimerization of the R-center. Moreover, it
As a preliminary application of the method, the natural
but unusual amino acid 4-carboxyglutamate (Gla)18 was
readily prepared. Boc-Gla(OMe)2-OH (13) and Fmoc-Gla-
(OtBu)2-OH (14) were obtained from compounds 4b and
5b in 97% and 90% yields, respectively, and each in >98%
ee as monitored by HPLC analysis of the FLEC-Glu
derivatives (Scheme 2). Following a different approach,
(19) Lalonde, J . J .; Berbreiter, D. E.; Wong, C. H. J . Org. Chem.
1988, 53, 2323-2327.
(20) (a) Ma¨rki, W.; Oppliger, Max; Thanei, P; Schwyzer, R. Helv.
Chim. Acta 1977, 60(3), 798-806. (b) Clape`s, P.; Valverde, I.; J aime,
C.; Torres; J . L. Tetrahedron Lett. 1996, 37, 417-418.
(21) (a) Oppliger, Max; Schwyzer, R. Helv. Chim. Acta 1977, 60(1),
43-47. (b) Schuerman, M. A.; Keverline, K. I.; Hiskey, R. G. Tetrahe-
dron Lett. 1995, 31, 825-828.
(22) (a) Danishefsky, S.; Berman, E.; Clizbe, L. A.; Hirama, M. J .
Am. Chem. Soc. 1979, 101, 4385-4386. (b) Atwood, M. R.; Carr, M.
G.; J ordan, S. Tetrahedron Lett. 1990, 31, 283-284. (c) Efenberger,
F.; Mu¨ller, W.; Keller, R.; Wild, W.; Ziegler, T. J . Org. Chem. 1990,
55, 3064-3067.
(14) Sieber, P.; Riniker, B. Tetrahedron Lett. 1987, 28, 6031-6034.
(15) Dugave, C.; Dubois, J .; Bory, S.; Gaudry, M.; Marquet, A. Bull.
Soc. Chim. Fr. 1991, 128, 381-386.
(16) O’Donnell, M. J .; Eckrih, T. M. Tetrahedron Lett. 1978, 4625-
4628.
(23) Tanaka, K-I.; Yoshifuji, S.; Nitta, Y. Chem. Pharm. Bull. 1986,
34, 3879-3884.
(17) Dugave, C. J . Org. Chem. 1995, 60, 601-607.
(18) Morris, H. R.; Thompson, M. R.; Dell, A. Biochem. Biophys. Res.
Commun. 1975, 62, 856-861.
(24) Rosowsky, A.; Bader, H.; Freisheim, J . H. J . Med. Chem. 1991,
34, 203-208.

Notes
J . Org. Chem., Vol. 61, No. 17, 1996 6069
temperature. A solution of iodide 2a /b (1 mmol) in THF (5 mL)
was added, and the mixture was stirred for 12 h at 40 °C. The
reaction was quenched with saturated aqueous ammonium
chloride, and the product was extracted twice with diisopropyl
ether (2 × 25 mL). The combined organic layers were washed
six times with saturated aqueous ammonium chloride (6 × 10
mL) and were dried over sodium sulfate. After evaporation
under reduced pressure the product was purified by silica gel
flash chromatography.
Meth od B. N-(Diphenylmethylene)glycine ethyl ester (278
mg, 1 mmol) was treated with freshly prepared LDA in a mixture
of dry THF-HMPA 60:40 (5 mL) for 1 h at -78 °C. After
dropwise addition of compound 2a (451 mg, 1 mmol) dissolved
in THF (5 mL), the mixture was stirred overnight at room
temperature. The reaction was quenched with saturated aque-
ous ammonium chloride, and the product was extracted as
described for method A.
Meth od C. A mixture of compound 2a (451 mg, 1 mmol),
N-(diphenylmethylene)acetonitrile (225 mg, 1 mmol), and tetra-
butylammonium hydrogen sulfate (410 mg, 1.2 mmol) dissolved
in dichloromethane (14 mL) was treated with 10% sodium
hydroxide (6 mL) for 4 h at room temperature. The organic layer
was dried over sodium sulfate, and the volume was reduced in
vacuo.
allows an orthogonality of protecting groups and hence
complements previous methods for synthesizing unusual
amino acids.
Exp er im en ta l Section
Gen er a l P r oced u r es. All reagents employed were of ana-
lytical grade and were purchased from Aldrich Chemical Co. and
Lancaster Synthesis Ltd. Serine derivatives were purchased
from Aldrich Chemical Co. and Bachem. THF was distilled
before use from sodium-benzophenone, and DMF was dried over
activated 4 Å molecular sieves. HMPA was distilled from
sodium and kept over 4 Å molecular sieves. All other solvents
were of analytical grade and were used without further purifica-
tion. Flash chromatography was performed on 40-60 µm (230-
400 mesh) Merck silica gel. NMR δ and J values are given in
ppm and Hz, respectively. Melting points were measured on a
hot stage Kofler apparatus and are uncorrected. Mass combus-
tion analyses were carried out by the “Service de Microanalyse”
of the ICSN (Gif-sur-Yvette, France).
(S)-1-Meth yl-N-(tr ip h en ylm eth yl)-3-iod oa la n in a te (2a )/
(S)-1-Ben zyl-N-(tr iph en ylm eth yl)-3-iodoalan in ate (2b). Com-
pound 1a /b (30 mmol) was treated with sodium iodide (49 g,
300 mmol) in dry acetone (300 mL) under argon for 24 h at room
temperature. The volume was concentrated in vacuo at room
temperature. The product, dissolved in diethyl ether, was
washed with 10% sodium thiosulfate until the color faded and
with brine. The organic layer was dried over sodium sulfate,
and the solvent was evaporated under reduced pressure. 2a
(S )-1,5,5′-Tr im e t h yl-N -(t r ip h e n ylm e t h yl)-4-ca r b oxy-
glu ta m a te (4a ) (Meth od A). Purification by silica gel flash
chromatography (solvent: ethyl acetate-cyclohexane 10:90) gave
3a (103 mg, 30%) and 4a (yellowish solid, 311 mg, 65%): mp )
97-99 °C; [R]25 ) +56° (c ) 1.0, chloroform); 1H NMR (CDCl3)
D
δ 7.46 (bd, J ) 6.7, 6H), 7.30-7.14 (m, 9H), 3.77 (s, 3H), 3.70 (s,
3H), 3.65 (d, J ) 7.1, 1H), 3.45 (t, J ) 5.9, 1H), 3.15 (s, 3H),
2.71 (b, 1H), 2.38 (m, 2H); 13C NMR (CDCl3) δ 174.4, 169.5, 169.3,
145.5, 128.7, 127.8, 126.4, 71.2, 54.3, 52.8 + 52.7, 51.7, 48.1,
34.2; MS (DCI, NH3) m/z 476 (MH+, 25), 243 (Tr+, 100), 234
(M2H+ - Tr+, 10). Anal. Calcd for C28H29NO6: C, 70.72; H,
6.15; N, 2.95. Found: C, 70.71; H, 6.18; N, 3.11.
(pale yellow paste, 13.51 g, quantitative): [R]25 ) +21° (c )
D
1.0, chloroform); 1H NMR (CDCl3) (two rotamers 1:1) δ 7.52-
7.43 (m, 6H), 7.32-7.19 (m, 9H), 4.39 (dd, J ) 6.2, J ′ ) 8.3) +
3.48 (dd, J ) 3.5, J ′ ) 7.0) (1H), 3.76 (s) + 3.30 (s) (3H), 3.35
(dd, J ) 3.5, J ′ ) 6.2) + 3.21 (dd, J ) 7.0, J ′ ) 9.8) (1H), 2.70
(dd, J ) 8.3, J ′ ) 13.0) + 2.54 (dd, J ) 6.2, J ′ ) 13.0) (1H), 2.3
(bs, 1H); 13C NMR (CDCl3) δ 172.7 + 171.3, 145.6 + 145.5,
128.6-126.5 (complex), 71.1 + 70.8, 56.3, 52.9, 52.0, 48.4, 20.2,
9.6; MS (DCI, NH3) m/z 472 (MH+, 1.5), 344 (MH+ - HI, 0.75),
243 (Tr+, 100), 230 (M - Tr+, 10). 2b (pale yellow paste, 15.61
(S)-1-Ben zyl-5,5′-d im et h yl-N-(t r ip h en ylm et h yl)-4-ca r -
boxyglu ta m a te (4b) (Meth od A). Purification by silica gel
flash chromatography (solvent: ethyl acetate-cyclohexane 10:
90) gave 3b (82 mg, 22%) and 4b (white gum, 409 mg, 74%):
[R]25D ) +20° (c ) 1.0, chloroform); 1H NMR (CDCl3) δ 7.46 (dd,
J ) 1.4, J ′ ) 8.4, 2H), 7.38-7.34 (m, 8H), 7.27-7.15 (m, 10H),
5.15 (ds, 2H), 4.00 (m, 1H), 3.67 (s, 3H), 3.54 (s, 3H), 3.21 (m,
1H), 2.50-2.31 (m, 2H), 1.83 (b, 1H); 13C NMR (CDCl3) δ 172.4,
170.9, 166.2, 145.6 + 145.3, 135.4 + 135.1, 128.7-126.3 (com-
plex), 71.2, 66.9, 52.7, 51.8, 45.5 + 45.3, 34.1; MS (DCI, NH3)
m/z 552 (MH+, 80), 310 (M2H+ - Tr+), 243 (Tr+, 100). Anal.
Calcd for C34H33NO6: C, 74.03; H, 6.03; N, 2.54. Found: C,
74.28; H, 6.27; N, 2.33.
g, 95%): [R]25 ) +5° (c ) 1.0, chloroform); 1H NMR (CDCl3)
D
(two rotamers 2:1) δ 7.50 (bd, J ) 7.1, 3H), 7.42-7.35 (m, 3H),
7.33-7.14 (m, 14H), 5.22 (d, J ) 13.3) + 4.69 (AB, J AB ) 12.2,
δA ) 4.79, δB ) 4.60) (2H), 4.41 (m) + 3.53 (m) (1H), 3.33-3.15
(m, 1H), 2.95-2.6 (bm, 1H), 1.57 + 2.24 (2bs, 1H); 13C NMR
(CDCl3) δ 172.0 + 170.6, 145.6 + 145.4, 135.2 + 135.1, 129.5-
126.5 (complex), 71.0 + 70.8, 67.4 + 67.1, 56.1, 53.4, 48.5, 20.3,
9.7; MS (DCI, NH3) m/z 548 (MH+, 1), 420 (MH+ - HI, 0.75),
306 (M - Tr+, 15), 243 (Tr+, 100).
En a n tiom er ic Excesses Deter m in a tion .13 Dep r otection
of Azir id in es. Compounds 3a /b (1 mg) were treated with TFA-
CH2Cl2 50:50 (2 mL) for 5 min at room temperature. After
removal of the solvent under reduced pressure, the product was
stirred for 1 h with 1 N lithium hydroxide (2 mL). Hyd r olysis
of Com p ou n d s 4-9. Compounds 4-9 (1 mg) suspended in 6
N hydrogen chloride (500 µL) were heated for 4 h at 100 °C.
Der iva tiza tion . After neutralization with 1 N acetic acid, the
product was freeze-dried. The residue was dissolved in a 1 M
borate buffer pH 6.35 (100 µL), and the solution was treated
with an 18 mM solution of (+)-1-(9-fluorenyl)ethyl chloroformate
in acetone ((+)-FLEC-Cl) (500 µL) for 4 min at room tempera-
ture. The aqueous layer, washed twice with pentane, was
analyzed by HPLC using a spherisorb octyl 3 µm analytical
column (150 × 4.6 mm); linear gradient 3a /b, acetonitrile-THF-
50 mM acetic acid pH 4.35 (0.5 mL/min, 150-200 bars): 0-8
min (8:17:75), 8-22 min (8:17:75 to 0:25:75), 22-45 min (0:25:
75); 4a /b-9a /b, acetonitrile-THF-50 mM acetic acid pH 4.35
(0.5 mL/min, 150-200 bars): 0-8 min (8:17:75), 8-22 min (8:
17:75 to 0:30:70), 22-45 min (0:30:70 to 0:40:60); detection at
265 nm detection at 265 nm. Retention times (rt): 3a /b, 20.24
( 0.18 min ((+)-FLEC-D-Azy), 21.02 ( 0.26 min ((+)-FLEC-L-
Azy); 4a /b-9a /b, 21.67 ( 0.57 min ((+)-FLEC-D-Glu), 23.06 (
0.52 min ((+)-FLEC-L-Glu).
(S)-1-Ben zyl-5,5′-d i-ter t-bu tyl-N-(tr iph en ylm eth yl)-4-ca r -
boxyglu ta m a te (5b) (Meth od A). Purification by silica gel
flash chromatography (solvent: ethyl acetate-cyclohexane 5-95)
gave 3b (67 mg, 16%) and 5b (yellowish crystals, 497 mg, 78%):
mp 121-122 °C; [R]25 ) +7° (c ) 1.0, chloroform); 1H NMR
D
(CDCl3) δ 7.49 (bd, J ) 7.2) + 7.38 (bd, J ) 7.2) + 7.34-7.10
(m) (20H), 5.22 (s, 2H), 3.69 (m, 1H), 3.29 (m, 1H), 2.39 (m) +
2.28 (b) (3H), 1.39 (s, 9H), 1.24 (s, 9H); 13C NMR (CDCl3) δ 172.9,
167.0, 166.8, 146.8, 145.7 + 145.5, 135.6, 129.3, 128.8-126.2
(complex), 81.5 + 81.4, 71.1, 66.7, 54.4, 50.3, 45.4, 27.7 + 27.6;
MS (DCI, NH3) m/z 636 (MH+, 35), 394 (M2H+ - Tr+, 4), 243
(Tr+, 100). Anal. Calcd for C40H45NO6: C, 75.56; H, 7.13; N,
2.20. Found: C, 75.89; H, 7.21; N, 1.93.
(2S ,4R S )-1,5-Dim e t h yl-N -(t r ip h e n ylm e t h yl)-4-cya n o-
glu ta m a te (6a ) (Meth od A). Purification by silica gel flash
chromatography (solvent: ethyl acetate-cyclohexane 20-80)
gave 3a (70 mg, 20%) and 6a (yellow gum, 328 mg, 74%): IR
(CHCl3) 2260; 1H NMR (CDCl3) δ 7.48-7.41 (m, 6H), 7.36-7.16
(m, 9H), 4.22 (dd, J ) 7.2, J ′ ) 8.3, 1H), 3.82 + 3.80 (2s, 3H),
3.69 (s, 3H), 3.21 (m, 1H), 2.73-2.60 (m, 2H), 2.03 (b, 1H); 13C
NMR (CDCl3) δ 171.2, 170.4, 165.7 + 165.6, 145.1, 128.6-126.6
(complex), 115.0 + 114.7, 71.0, 53.8 + 53.7, 46.1 + 45.4, 42.7 +
42.5, 37.4 + 37.1; MS (DCI, NH3) m/z 443 (MH+, 4), 243 (Tr+,
100), 201 (M2H+ - Tr+, 5). Anal. Calcd for C27H26N2O4: C,
73.28; H, 5.92; N, 6.33. Found: C, 73.01; H, 6.06; N, 5.97.
(S)-1,5-Dim et h yl-N-(ter t-b u t yloxyca r b on yl)-4-ca r b oxy-
glu ta m a te 13 fr om 4b. A mixture of compound 4b (954 mg,
1.5 mmol) and diterbutyl dicarbonate (242 mg, 1.65 mmol) was
vigorously stirred with palladium black (90 mg, 10% w/w) in a
Meth od A. All reactions were carried out in dried glassware
under an argon atmosphere. All solvents used were dried by
the usual procedures. The malonate (1 mmol) was added to a
suspension of 80% sodium hydride (1.0 mmol) in THF-HMPA
60-40 (5 mL), and the mixture was stirred 30 min at room

6070 J . Org. Chem., Vol. 61, No. 17, 1996
Notes
mixture of ethyl acetate-methanol 50:50 (75 mL) under hydro-
gen (2.5 bars) for 4 h at room temperature. Filtration of the
catalyst, elimination of the solvent under reduced pressure, and
silica gel flash chromatography afforded 13 (colorless oil, 310
+ 52.5, 52.4, 51.7, 48.3, 31.5, 28.2. MS (DCI, NH3) m/z 351
(MNH4+, 100), 334 (MH+, 95), 295 (MNH4+ - (CH3)2CdCH2, 55),
278 (MH+ - (CH3)2CdCH2, 25), 234 (MH+ - (CH3)2CdCH2
-
CO2, 15), 174 (MH+ - (CH3)2CdCH2 - CO2 - HCO2Me, 10).
Anal. Calcd for C14H23NO8: C, 50.44; H, 6.95; N, 4.20. Found:
C, 50.58; H, 7.27; N, 3.90.
mg, 97%): [R]25 ) -1.9° (c ) 1.0, methanol); 1H NMR (CDCl3)
D
δ 7.98 (b, 1H), 5.26 (bd, 1H), 4.40 (bm, 1H), 3.76 (s, 3H), 3.75 (s,
3H), 3.58 (t, J ) 6.8, 1H), 2.58 (m, 1H), 2.25 (m, 1H), 1.44 (s
(S)-5,5′-Dim eth yl-N-(ter t-bu tyloxyca r bon yl)-4-ca r boxy-
glu ta m a te (13) fr om 15. Compound 32 (333 mg, 1 mmol)
dissolved in DMSO (10 mL) was added to a 0.1M KH2PO4 buffer,
and the pH was adjusted to 7.6. R-Chymotrypsin (100 mg, type
II 54 u/mg) was added, and the mixture was stirred at 37 °C.
The reaction was monitored by TLC. After complete consump-
tion of the starting compound, the reaction was quenched with
10% citric acid and the product was extracted four times with
dichloromethane. The organic layer was dried over sodium
sulfate, and the volume was evaporated under reduced pressure.
Silica gel flash chromatography (solvent: chloroform-methanol-
acetic acid 94:5:1) gave 30 (colorless oil, 318 mg, quantitative):
9H); 13C NMR (CDCl3) δ 175.1, 169.5, 169.1, 155.5, 80.4, 52.8,
+
51.6, 48.3, 39.6, 31.1, 28.2; MS (DCI, NH3) m/z 337 (MNH4
,
40), 320 (MH+, 100), 281 (MNH4 - (CH3)2CdCH2, 60), 264
(MH+ - (CH3)2CdCH2, 55), 220 (MH+ - (CH3)2CdCH2 - CO2,
15). Anal. Calcd for C12H21NO8: C, 46.90; H, 6.89; N, 4.56.
Found: C, 46.74; H, 7.35; N, 4.20.
+
(S)-5,5′-Di-ter t-bu tyl-N-(9-flu or en ylm eth yloxyca r bon yl)-
4-ca r boxyglu ta m a te (14). Compound 5b (954 mg, 1.5 mmol)
was vigorously stirred with palladium black (90 mg, 10% w/w)
in a mixture of ethyl acetate-methanol 50:50 (75 mL) under
hydrogen (2.5 bars) for 8 h at room temperature. The solvent
was evaporated under reduced pressure, and the residue was
treated with Fmoc-N-hydroxysuccinimide (580 mg, 1.65 mmol)
and diisopropylethylamine (295 µL, 1.65 mmol after neutrality)
in dry THF for 2 h at room temperature. The reaction was
quenched by addition of 10% citric acid, the product was
extracted twice with diethyl ether, and the organic layer was
dried over sodium sulfate. Evaporation of the solvent under
reduced pressure and silica gel flash chromatography afforded
14 (colorless paste, 713 mg, 90%): [R]25D ) 0° (c ) 1, CHCl3); 1H
NMR (CDCl3): δ 7.75 (d, J ) 7.4, 2H), 7.59 (d, J ) 5.7, 2H),
7.42-7.16 (m, 4H), 5.56 (bd, 1H), 4.44 (dd, J ) 7.0, J ′ ) 10.2,
2H), 4.33 (t, J ) 10.2, 1H), 4.21 (t, J ) 7.0, 1H), 2.54-2.43 (m,
1H), 2.27-2.15 (m, 1H), 1.46 (s 18H); 13C NMR (CDCl3) δ 175.8,
168.4, 168.2, 156.2, 143.8, 143.6, 141.2, 127.7, 127.1, 125.2, 125.1,
119.0, 82.4, 67.3, 52.4, 50.8, 47.0, 39.6, 30.7, 27.8; MS (DCI, NH3)
[R]25 ) -1.8° (c ) 1.0, methanol); NMR data were identical to
D
those described above.
(2S,4RS)-1,5-Dim et h yl-N-(ter t-b u t yloxyca r b on yl)-4-cy-
a n oglu ta m a te (16). Compound 6a (2.2 g, 5 mmol) was treated
by formic acid as reported above. Silica gel flash chromatogra-
phy (solvent: ethyl-acetate-hexane 25:75) yielded 16 (amor-
phous solid, 465 mg, 31%) and 17 (colorless oil, 332 mg, 27%).
1
16: mp ) 83-84 °C; IR (CHCl3) 2250; H NMR (CDCl3) δ 5.22
(b, 1H), 4.51 (bm, 1H), 3.85 + 384 (2s, 3H), 3.80 + 3.79 (2s, 3H),
3.78-3.67 (m, 1H), 2.68-2.47 (m, 1H), 2.42-2.19 (m, 1H), 1.46
(s, 9H); 13C NMR (CDCl3) δ 173.0, 171.7, 171.1, 152.0, 83.3, 52.3,
50.1, 30.1, 27.9, 27.3, 26.8; MS (DCI, NH3) m/z 301 (MH+, 100),
245 (MH+ - (CH3)2CdCH2, 35), 201 (MH+ - (CH3)2CdCH2
-
CO2, 30). Anal. Calcd for C13H20N2O6: C, 51.99; H, 6.71; N,
9.33. Found: C, 51.56; H, 6.43; N, 9.69.
m/z 543 (MNH4
,
55), 526 (MH+, 45), 487 (MNH4
-
(2S,4RS)-5-Met h yl-N-(ter t-b u t yloxyca r b on yl)-4-cya n o-
glu ta m a te (18). Compound 16 (150 mg, 1 mmol) was treated
as reported for compound 13 from 15. Silica gel flash chroma-
tography (solvent: ethyl acetate-acetic acid 99:1) yielded 18
(colorless oil, 123 mg, 86%): IR (CHCl3) 2250; 1H NMR (CDCl3)
δ 5.25 (b, 1H), 4.46 (bm, 1H), 3.85 + 3.84 (ds, 3H), 3.74 (dd, J )
5.7, 8.5, 1H), 2.62 (m, 1H), 2.39 (m, 1H), 1.47 (s, 9H); 13C NMR
(CDCl3) δ 174.2, 173.5, 155.6, 115.7, 81.1, 53.8, 33.9, 32.0, 28.2,
24.5; MS (DCI, NH3) m/z ) 304 (MNH4+, 55), 287 (MH+, 20),
+
+
(CH3)2CdCH2, 20), 470 (MH+ - (CH3)2CdCH2, 30), 443 (MNH4
- (CH3)2CdCH2 - CO2, 6), 426 (MH+ - (CH3)2CdCH2 - CO2,
+
7), 414 (MH+ - 2 (CH3)2CdCH2, 10), 387 (MNH4 - 2
+
(CH3)2CdCH2 - CO2, 10), 370 (MH+ - 2 (CH3)2CdCH2 - CO2,
55), 179 (Fl+, 100). Anal. Calcd for C29H35NO8: C, 66.27; H,
6.71; N, 2.61. Found: C, 66.71; H, 7.19; N, 2.39.
(S )-1,5,5′-Tr im e t h yl-N -(t er t -b u t yloxyca r b on yl)-4-ca r -
boxyglu ta m a te (15). Compound 4a (142.5 mg, 0.3 mmol) was
stirred for 15 min in a mixture of dichloroethane-formic acid
95:5 (12 mL) at room temperature. After evaporation of the
solvent in vacuo, the residue was dissolved in DMF (10 mL) and
was neutralized with triethylamine. The product was treated
with di-tert-butyl dicarbonate (75 mg, 0.33 mmol) and triethyl-
amine (46 µL, 0.33 mmol) overnight at room temperature. The
solvent was evaported under reduced pressure, and the product
was dissolved in ethyl acetate and washed with 10% citric acid.
Drying of the organic layer over sodium sulfate, evaporation of
the solvent, and purification by silica gel flash chromatography
260 (MNH4 - CO2, 85), 243 (MH+ - CO2, 100). Anal. Calcd
+
for C12H18N2O6: C, 50.35; H, 6.34; N, 9.78. Found: C, 49.87;
H, 6.78; N, 9.36.
Ackn owledgm en t. We gratefully acknowledge Gilles
Meunier for help in optical purity determination.
Su p p or tin g In for m a tion Ava ila ble: Description of 1H
and 13C NMR spectra and characterization of compounds 1,
3, and 7-12 (4 pages). This material is contained in librairies
on microfiche, immediately follows this article in the microfilm
version of the journal, and can be ordered from the ACS; see
any current masthead page for ordering information.
(solvent: ethyl acetate-cyclohexane 25:75) afforded 15 (colorless
1
oil, 96 mg, 96%): [R]25 ) -17° (c ) 1.0, chloroform); H NMR
D
(CDCl3) δ 5.02 (b, 1H), 4.39 (bm, 1H), 3.76 (s, 3H), 3.75 + 3.74
(ds, 3H), 3.53 (t, J ) 7.0, 1H), 2.53 (m, 1H), 2.20 (m, 1H), 1.44
(s 9H); 13C NMR (CDCl3) δ 172.2, 169.4, 169.0, 155.2, 80.2, 52.8
J O960624G