An alkanesulfonamide 'safety-catch' linker for solid-phase synthesis

Article abstract of DOI:10.1021/jo981990y

An alkanesulfonamide 'safety-catch' linker has been developed for tethering carboxylic acids to support. Acylation of the sulfonamide support provides a support-bound N-acylsulfonamide that is stable to both basic and strongly nucleophilic reaction condit

Full text of DOI:10.1021/jo981990y

2322
J . Org. Chem. 1999, 64, 2322-2330
An Alk a n esu lfon a m id e “Sa fety-Ca tch ” Lin k er for
Solid -P h a se Syn th esis
Bradley J . Backes and J onathan A. Ellman*
Department of Chemistry, University of California, Berkeley, California 94720
Received October 1, 1998
An alkanesulfonamide “safety-catch” linker has been developed for tethering carboxylic acids to
support. Acylation of the sulfonamide support provides a support-bound N-acylsulfonamide that is
stable to both basic and strongly nucleophilic reaction conditions. At the end of a solid-phase
synthesis sequence, treatment with iodoacetonitrile provides N-cyanomethyl derivatives that can
be cleaved by nucleophiles under mild reaction conditions to release the target compounds. Coupling
conditions have been developed to load Boc- and Fmoc-amino acids to the alkanesulfonamide resin
with high loading efficiencies and minimal racemization. A number of support-bound amino acids
incorporating diverse side-chain functionality have been subjected to a short synthesis sequence,
followed by iodoacetonitrile activation and nucleophilic displacement to provide dipeptide products.
All 20 of the proteinogenic amino acids, when suitably protected, are compatible with the loading
and activation steps. Finally, displacement with various nucleophiles including amines, R-amino
acid methyl esters, and R-amino acid coumarin derivatives is demonstrated.
Sch em e 1
In tr od u ction
The utility of small molecule libraries in drug research
programs has caused a resurgence in the field of solid-
phase organic synthesis.1,2 The emergence of novel linkers
and linkage strategies has been a central component of
these developments.3 “Safety-catch” linker strategies in
which a stable linkage is activated for cleavage by a
discrete chemical modification have proven to be par-
ticularly useful.4,5 These linkers are often compatible with
a broad range of reactions, yet target compounds can be
released from support under mild conditions after the
activation step. In addition, the activation step can
potentially provide a reactive linkage functionality for
cleavage with diverse nucleophiles. Therefore, an ad-
ditional element of diversity can be added to the structure
of the target compounds in the cleavage step. Activation
can also facilitate the synthesis of cyclic compounds by
intramolecular attack of a nucleophile upon the linkage
functionality liberating the target compounds from sup-
port.
Kenner’s sulfonamide “safety-catch” linker6 1 for teth-
ering carboxylic acids to support is unique in that it
shows stability to both strongly acidic and strongly basic
reaction conditions (Scheme 1). Kenner employed an
arenesulfonamide linker for solid-phase peptide chemis-
try in which activation by diazomethane treatment
provided an N,N-methylacylsulfonamide 3 that can be
cleaved with forcing conditions by aminolysis, hydrazi-
nolysis, and saponification to provide primary amides,
hydrazones, and carboxylic acids, respectively. Limita-
tions encountered in Kenner’s work include poor loading
efficiencies, racemization in the loading step, and poor
reactivity of the N,N-methylacylsulfonamide 3 that is
obtained upon diazomethane activation. We have utilized
a modified version of this linker for small-molecule
organic synthesis efforts to prepare substituted aryl
acetic acid derivatives that represent an important class
of antiinflammatory drugs.7 Basic reaction conditions
including enolate alkylation and Suzuki cross-coupling
reaction conditions were demonstrated. We have also
developed an activation method in which N-alkylation
of the N-acylsulfonamide with a haloacetonitrile provides
an extremely reactive N,N-cyanomethylacylsulfonamide
4 for mild nucleophilic displacement.8,9 The linkage is
reactive enough to allow for the use of limiting quantities
of nucleophilic amines in the cleavage step to provide
pure amide products in solution. Thus, target compounds
can be isolated without purification, greatly simplifying
compound isolation.10
* To whom correspondence should be addressed. Tel: 510-642-4488.
Fax: 510-642-8369. E-mail: jellman@uclink.berkeley.edu.
(1) Thompson, L. A.; Ellman, J . A. Chem. Rev. 1996, 96, 555.
(2) Balkenhohl, F.; von dem Bussche-Hunnefeld, C.; Lansky, A.;
Zechel, C. Angew. Chem., Int. Ed. Engl. 1996, 35, 8.
(3) Backes, B. J .; Ellman, J . A. Curr. Opin. Chem. Biol. 1997, 1, 86.
(4) Gayo, L. M.; Suto, M. J . Tetrahedron Lett. 1997, 38, 211.
(5) Brown, A. R.; Rees, D. C.; Rankovic, Z.; Morphy, J . R. J . Am.
Chem. Soc. 1997, 119, 3288.
We were interested in extending the use of sulfonamide
linkers to include the loading, activation, and cleavage
(7) Backes, B. J .; Ellman, J . A. J . Am. Chem. Soc. 1994, 116, 11171.
(8) Backes, B. J .; Virgilio, A. A.; Ellman, J . A. J . Am. Chem. Soc.
1996, 118, 3055.
(9) For other applications of the N-cyanomethyl activation method,
see: (a) Thompson, L. A.; Moore, F. L.; Moon, Y.-C.; Ellman, J . A. J .
Org. Chem. 1998, 63, 2066. (b) Link, A.; van Calenbergh, S.; Herdewijn,
P. Tetrahedron Lett. 1998, 39, 5175.
(6) Kenner, G. W.; McDermott, J . R.; Sheppard, R. C. J . Chem. Soc.,
Chem. Commun. 1971, 636.
10.1021/jo981990y CCC: $18.00 © 1999 American Chemical Society
Published on Web 03/05/1999

Alkanesulfonamide “Safety-Catch” Linker
J . Org. Chem., Vol. 64, No. 7, 1999 2323
Sch em e 2a
of amino acid derivatives. The use of an amino acid as
the initial, linker-bound building block has been the most
widely employed strategy for the solid-phase synthesis
of small organic molecules.1,2,11 Amino acids are utilized
extensively in organic synthesis,12 and the commercial
availability of numerous, diverse derivatives makes them
ideal for use in combinatorial synthesis efforts.13 Exten-
sion of the safety-catch linker to include amino acids
would also clearly be applicable to solid-phase peptide
synthesis for segment condensation strategies14 and for
the synthesis of cyclic peptides15 and peptide amides.16
In a previous paper,8 we demonstrated that the halo-
acetonitrile activation method can be extended to include
carboxylic acids that possess R-electron-withdrawing
groups, such as N-protected amino acids, by employing
an alkanesulfonamide linker.17 After activation of sup-
port-bound amino acids by iodoacetonitrile treatment,
cleavage was demonstrated with amines and amino acid
methyl esters. Here, a detailed experimental procedure
for the synthesis of an alkanesulfonamide handle and
optimized procedures for the attachment of the initial
amino acid that provide minimal racemization and high
loading efficiencies are reported. As well, the generality
of the iodoacetonitrile activation procedure is demon-
strated by the preparation of a number of dipeptide
amides that incorporate diverse functionality. Finally,
displacement of the support-bound N,N-cyanomethyl-
acylsulfonamide active ester is accomplished with a
variety of nucleophiles including amines, R-amino methyl
esters, and R-amino acid coumarin derivatives.
a
Key: (a) SOCl2, MeOH; (b) Cl2, H2O; (c) NH3, Et2O; (d) KOH,
H2O; (e) aminomethyl polystyrene resin, DICI, HOBt, DMF.
sulfonamide handle: (1) The synthesis sequence must be
short and simple and allow for the preparation of
significant quantities of material without the need for a
column chromatography step. (2) The handle must be
loaded to support in an efficient manner to provide an
alkanesulfonamide resin uncontaminated with byprod-
ucts. To meet these criteria, 3-carboxypropanesulfona-
mide18 was prepared as a handle for loading to amino-
methyl resins (Scheme 2). Starting from commercially
available 4,4′-dithiobutyric acid (6), esterification with
thionyl chloride in MeOH followed by distillation provides
diester 7 in high yield. Oxidation with Cl2 gas in water
followed by extractive isolation then gives 2 equiv of
sulfonyl chloride 8. Aminolysis of 8 provides sulfonamide
9 with product isolation accomplished by extraction.
Saponification and isolation by precipitation of the KCl
salts provides the 3-carboxypropanesulfonamide handle
10 in good yield. The handle can be quantitatively loaded
to aminomethyl resin using standard DICI (N,N′-diiso-
propylcarbodiimide) and HOBt (N-hydroxybenzotriazole)
coupling conditions to provide alkanesulfonamide resin
11 as monitored by a bromophenol blue test.19,20 Oligo-
merization does not occur in the coupling step since the
alkanesulfonamide does not react with HOBt esters (vide
infra).
Op tim iza tion of F ir st Am in o Acid Loa d in g. Cou-
pling conditions were sought to load the first amino acid
to alkanesulfonamide linker 11 with high loading ef-
ficiencies and low levels of racemization. In initial
studies, 11 proved quite unreactive, failing to react using
DICI and HOBt activation. Acylation with HOBt esters
proceeds only with the addition of catalytic DMAP,
indicating that the reactivity of the alkanesulfonamide
is comparable to that of an alcohol. Linker 11 was
subjected to several coupling conditions (Table 1), fol-
lowed by activation with iodoacetonitrile and nucleophilic
displacement with MeO-Leu-NH2 to provide dipeptide 14
(Scheme 3). Reversed-phase HPLC analysis was used to
determine the extent of racemization (% D,L-epimer
observed). Acylation of 11 using symmetrical anhydrides
Resu lts a n d Discu ssion
Syn th esis of a n Alk a n esu lfon a m id e Ha n d le. Two
criteria were considered for the synthesis of an alkane-
(10) Alternative strategies to simplify compound isolation such as
the use of support-bound scavengers have been developed. Kaldor S.
W.; Siegel M. G.; Fritz, J . E.; Dressman, B. A.; Hahn, P. J . Tetrahedron
Lett. 1996, 37, 7193.
(11) Selected examples follow. Benzodiazepines and hydantoins: (a)
DeWitt, S. W.; Kiely, J . S.; Stankovic, C. J .; Schroeder, M. C.; Reynolds-
Cody, D. M.; Pavia, M. R. Proc. Natl. Acad. Sci. U.S.A. 1993, 90, 6909.
Hydantoins: (b) Dressman, B. A.; Spangle, L. A.; Kaldor, S. W.
Tetrahedron Lett. 1996, 37, 937. Diketopiperizines: (c) Gordon D. W.;
Steele, J . Bioorg. Med. Chem. Lett. 1995, 5, 47. Pyrrolidinones: (d)
Murphy, M. M.; Schullek, J . R.; Gordon, E. M.; Gallop, M. A. J . Am.
Chem. Soc. 1995, 117, 7029. Thiazolidinones and 4-metathiazines: (e)
Holmes, C. P.; Chinn, J . P.; Look, G. C.; Gordon, E. M.; Gallop, M. A.
J . Org. Chem. 1995, 60, 7328. â-Lactams: (f) Ruhland, B.; Bhandari,
E. M.; Gordon, E. M.; Gallop, M. A. J . Am. Chem. Soc. 1996, 118, 253.
(12) Sardina, J .; Rapoport, H. Chem. Rev. 1996, 96, 1825.
(13) Gordon, E. M.; Gallop, M. A.; Patel, D. V. Acc. Chem. Res. 1996,
29, 144.
(14) For reviews, see: (a) Lloyd-Williams, P.; Albericio, F.; Giralt,
E. Tetrahedron 1993, 49, 11065. (b) Benz, H. Synthesis 1994, 337. For
the use of Kaiser’s oxime linker, see: Kaiser, E. T.; Mihara, H.; Laforet,
G. A.; Kelly, J . W.; Walters, L.; Findeis, M. A. Sasaki, T. Science 1989,
243, 187.
(15) For the use of Kaiser’s oxime linker see: Osapay, G.; Taylor,
J . W. J . Am. Chem. Soc. 1990, 112, 6046.
(16) For the use of Kaiser’s oxime linker, see: (a) Voyer, N.; Lavoie,
A.; Pinette, M.; Bernier, J . Tetrahedron Lett. 1994, 35, 355. For “safety-
catch” strategies to prepare peptide amides and cyclic peptides, see:
(b) Marshall, D. L.; Liener, I. E. J . Org. Chem. 1970, 35, 867. (c)
Flanigan, E.; Marshall, G. R. Tetrahedron Lett. 1970, 21, 2403. (d)
Lewalter, J .; Birr, C. Liebigs Ann. Chem. 1970, 740, 48. (e) Weieland,
T.; Lewalter, J .; Birr, C. Liebigs Ann. Chem. 1970, 740, 31.
(17) In
a previous report (ref 8), we demonstrated that in the
iodoacetonitrile step an alkanesulfonamide linker, in contrast to
arenesulfonamide linkers, provides for the complete N-alkylation of
N-acylsulfonamides derived from carboxylic acids that possess R-electron-
withdrawing groups. This is due to the greater basicity and, therefore,
greater nucleophilicity of the alkanesulfonamide (methanesulfonamide
DMSO pKa 17.5) vs the arenesulfonamide (benzenesulfonamide DMSO
pKa 16.0). Bordwell, F. G. Acc. Chem. Res. 1988, 21, 456.
(18) The procedure to prepare 3-carboxypropanesulfonamide is
patterned after Beck’s procedure for the synthesis of 2-carboxyethane-
sulfonamide. Beck, M. J . Org. Chem. 1968, 33, 897.
(19) Bunin, B. A. The Combinatorial Index; Academic Press: San
Diego, 1998; pp 213-236.
(20) This resin is now commercially available; Novabiochem catalog
no. 01-64-0152.

2324 J . Org. Chem., Vol. 64, No. 7, 1999
Backes and Ellman
Ta ble 1. Op tim iza tion of F ir st Am in o Acid Loa d in g of Alk a n esu lfon a m id e Resin 11 (Sch em e 3)a
yield
entry
coupling methodb
base
solvent
DMF
CH2Cl2
DMF
DMF
DMF
DMF
DMF
DMF
DMF
14,c %
DL,d
%
1
2
3
4
5
6
7
8
9
10
11
12
13
14
(Boc-Phe-O)2 (5 equiv), DMAP (cat.)
(Boc-Phe-O)2 (5 equiv), DMAP (cat.)
Boc-Phe-OH (5 equiv), HATU (5 equiv)
Boc-Phe-OH (5 equiv), PyBrop (5 equiv)
Boc-Phe-OPfp (5 equiv), DMAP (cat.)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv), HOBt (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv), HOAt (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
2 × Boc-Phe-OH (5 equiv), PyBop (5 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
collidine (10 equiv)
Proton Sponge (10 equiv)
NMM (10 equiv)
90
nd
nd
nd
7-10
5.5
2.5
3.1
<0.5
2.5
2.0
1.4
1.5
<0.5
0.85
5.2
0.7
<0.5
<5
<10
<10
80
<30
<10
<20
nd
<50
90
DMF
DMF
DMF
THF
i-Pr2EtN (10 equiv)
i-Pr2EtN (10 equiv)
CH2Cl2, -20 °C
a
Abbreviations used: nd ) not determined; pfp ) pentafluorophenyl; Proton Sponge ) 1,8-bis(N,N-dimethylamino)naphthalene; NMM
) N-methylmorpholine. Couplings endured 2-4 h. c Yields are based upon the initial aminomethyl resin substitution level and represent
a mass balance of purified material. % D,L-epimer is determined by reversed-phase HPLC analysis.
b
d
Sch em e 3a
a
Key: (a) coupling conditions (Table 1); (b) ICH2CN, i-Pr2EtN, NMP; (c) Leu-OMe, CH2Cl2.
with DMAP in DMF or CH2Cl2 (Table 1, entries 1 and 2)
provides excellent loading efficiency, but significant
epimerization. The use of coupling agents HATU (O-(7-
azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexaflu-
orophosphate) and PyBrop (bromotris(pyrrolidino)phos-
phonium hexafluorophosphate) also results in significant
levels of racemization (Table 1, entries 3 and 4). Acylation
with the pentafluorophenyl active ester of Boc-Phe-OH
and catalytic DMAP leads to significantly less racemiza-
tion but poor loading efficiency (Table 1, entry 5).
Moderate success is realized using PyBOP (benzotriazole-
1-yloxytris(pyrrolidino)phosphonium hexafluorophos-
phate) with i-Pr2EtN in DMF (Table 1, entry 8), in which
a double coupling provides a 80% yield of 14 with only
1.4% of the D,L-epimer being observed. Attempts to lower
the level of racemization obtained with PyBOP using
different bases (Table 1, entries 10-12) or solvents (Table
1, entry 13) did not reduce racemization and resulted in
lower yields. In general, the PyBOP coupling yields fall
with the addition of HOBt and HOAt (7-aza-1-benzo-
triazole) (Table 1, entries 6 and 7), and the coupling yields
are most dependable when PyBOP is added directly to
the reaction mixture without a premix protocol. This
indicated that the O-acyl phosphonium adduct is the
active intermediate. Indeed, Patel and co-workers have
utilized PyBOP and i-Pr2EtN in CH2Cl2 at -20 °C for
the acylation of alcohols in high yields.21 Under these
conditions, the O-acyl phosphonium adduct is proposed
to be the reactive intermediate. When these conditions
are employed (Table 1, entry 14), a good yield of dipeptide
14 is observed and racemization is not detected. Further
optimization of loading efficiencies has been performed
using Fmoc-amino acids since quantitation of the loading
level by spectrophotometric methods19 is both accurate
and straightforward.
Loa d in g Efficien cy for F m oc-Am in o Acid s. When
our most effective loading conditions in CH2Cl2 are
employed (Table 1, entry 14), a double coupling procedure
utilizing 5 equiv of amino acid in each step is required
to obtain high loading efficiencies. As well, many Fmoc-
amino acid derivatives are insoluble in CH2Cl2 at -20
°C. Dramatic improvements in both coupling yields and
solubility properties are realized when CHCl3 rather than
CH2Cl2 is employed as solvent in the coupling step.
Treatment of resin 11 with Fmoc-Phe-OH (3 equiv),
PyBOP (3 equiv), and i-Pr2EtN (5 equiv) with CHCl3 as
solvent at -20 °C for 8 h provides excellent loading
efficiencies (89%).22 This is noteworthy since Fmoc-Phe-
OH is one of the most insoluble Fmoc-amino acids.23
Coupling with Fmoc-Asp(O-t-Bu)-OH employing the iden-
tical coupling procedure results in a loading efficiency of
92%. To determine the extent of racemization for each
of these couplings, dipeptide amides 18a ,b have been
prepared (Scheme 4). The R-amino group of Fmoc-amino
acids 15a ,b are deprotected with 20% piperidine in DMF
for 30 min. After several DMF washes, Boc-L-Phe-OH is
coupled using DICI, HOBt activation in DMF with a
short premixing period of 2 min. Activation with 1 M
iodoacetonitrile and i-Pr2EtN with NMP (1-methyl-2-
pyrrolidinone) as solvent provides the support-bound
N,N-cyanomethylacylsulfonamide active esters 17a ,b.
The support-bound dipeptides are cleaved with 5 equiv
of benzylamine in THF to provide dipeptide amides 18a
in 85% yield (<0.5% D,L-epimer) and 18b in 83% yield
(22) The loading yields are determined by piperidine treatment of
small aliquots of resin followed by spectrophotometric analysis of the
piperidine-bifulvulene adduct in solution (ref 19).
(23) Fields G. B.; Noble R. L. Int. J . Peptide Protein Res. 1990, 35,
161.
(21) Kim, M. H.; Patel, D. V. Tetrahedron Lett. 1994, 35, 5603.

Alkanesulfonamide “Safety-Catch” Linker
J . Org. Chem., Vol. 64, No. 7, 1999 2325
Sch em e 4a
a
Key: (a) Fmoc-Phe-OH (3 equiv) or Fmoc-Asp(O-t-Bu)-OH (3 equiv), PyBOP (3 equiv), i-Pr2EtN (5 equiv), CHCl3 -20 °C, 8 h; (b) 20%
piperidine in DMF; (c) Boc-Phe-OH, DICI, HOBt, DMF (2 min premix); (d) ICH2CN, i-Pr2EtN, NMP; (e) BnNH2 (5 equiv), THF.
Sch em e 5a
Ta ble 2. Cou p lin g of F m oc-Am in o Acid s to
Alk ylsu lfon a m id e Resin 11 (Sch em e 5)a
resin product
Fmoc-amino acid
yield,b,c
%
15c
15d
15e
15b
15f
15g
15h
15i
Fmoc-Ala-OH
93
85
81
92
>95
81
Fmoc-Arg(Pbf)-OH
Fmoc-Asn(Trt)-OH
Fmoc-Asp(O-t-Bu)-OH
Fmoc-Cys(Trt)-OH
Fmoc-Glu(O-t-Bu)-OH
Fmoc-Gln(Trt)-OH
Fmoc-Gly-OH
Fmoc-His(Trt)-OH
Fmoc-His(Boc)-OH
Fmoc-Ile-OH
Fmoc-Leu-OH
Fmoc-Lys(Boc)-OH
Fmoc-Met-OH
Fmoc-Phe-OH
Fmoc-Pro-OH
a
Key: (a) Fmoc-amino acid (3 equiv), PyBOP (3 equiv), i-Pr2EtN
(5 equiv), CHCl3 -20 °C, 8 h.
77
89e
>95
89
(0.8% D,L-epimer). It is important that the coupling
reaction is filtered immediately after the 8 h coupling
period. Couplings allowed to proceed for longer reaction
times, or allowed to come to room temperature, while
providing a modest increase in loading efficiencies, result
in increased levels of racemization (18a , 1.7% D,L-epimer).
To demonstrate the generality of our coupling condi-
tions, a number of Fmoc-protected amino acids have been
coupled to the alkanesulfonamide resin 11 (Scheme 5,
Table 2). In most cases, good loading efficiencies are
achieved in a single coupling step. For sterically hindered
Fmoc-amino acids where couplings are less efficient, such
as Fmoc-Ile-OH (65%), a double coupling provides useful
levels of substitution (80%). For Fmoc-Gly-OH it is
necessary to employ DMF as solvent to increase solubil-
ity.
15j
15k
15l
65 (80)d
80
15m
15n
15o
15a
15p
15q
15r
15s
15t
80
>95
89
44 (65)d
77
Fmoc-Ser(O-t-Bu)-OH
Fmoc-Thr(O-t-Bu)-OH
Fmoc-Trp(Boc)-OH
Fmoc-Tyr(O-t-Bu)-OH
Fmoc-Val-OH
55 (70)d
80
76
15u
69 (84)d
a
Abbreviation used: Pbf ) 2,2,4,6,7-pentamethyldihydroben-
zofuran-5-sulfonyl. Yields determined by Fmoc analysis. c Cou-
b
d
plings were terminated after 8 h. Yield for a double coupling.
e DMF is employed as solvent with Fmoc-Gly-OH (4 equiv) and
i-Pr2EtN (6 equiv).
Syn th esis of Dip ep tid e Am id es. Fmoc-amino acid
loaded resins 15a -u have been subjected to the short
peptide amide synthesis sequence described previously
(Scheme 4) to demonstrate the compatibility of the
iodoacetonitrile activation step with various side chain
functionality. Fmoc removal, Boc-Phe-OH coupling, io-
doacetonitrile activation, and benzylamine cleavage are
performed as described in the previous section with the
exception that the activated support-bound dipeptides
17a -u are cleaved with a limiting amount of benzyl-
amine (ca. 0.5 equiv) in THF to provide dipeptides amides
18a -u in solution (Table 3). The THF solutions are then
filtered through a small plug of silica to remove resin
particulates followed by elution with excess THF. Re-
versed-phase HPLC analysis at 254 nm is utilized to
gauge the purity of the peptide products. The dipeptide
amide THF solutions are concentrated and the yields are
determined on the basis of the limiting reagent, benzyl-
amine, for the mass balance of recovered material.
Concentrated samples were directly submitted for el-
emental analysis without further purification. High levels
of purity are observed for most products. It should be
noted that for cases in which the amino acid-substituted
resin employed may contain free, unreacted alkane-
sulfonamide (Fmoc-Ile 15l), no benzylamide product
derived from Boc-Phe-OH coupling to the free alkylsul-
fonamide is observed. Yields are uniformly high for the
dipeptide products 18a -u , and satisfactory elemental
analyses were obtained for a significant number of
derivatives (18 out of 20). For those dipeptide products
where elemental analyses were not satisfactory, 1H NMR
showed a high level of purity. It is notable that significant
S-alkylation of the thioether-containing side chains of
Met and Cys(Trt) did not occur in the iodoacetonitrile
activation step, and dipeptide products 18o and 18f were
obtained in good yields. While loading proceeds in high
efficiency for Fmoc-His(Trt)-OH (Table 2, >95%), no

2326 J . Org. Chem., Vol. 64, No. 7, 1999
Backes and Ellman
Ta ble 3. Cou p lin g, Activa tion , a n d Nu cleop h ilic
Disp la cem en t w ith Su p p or t-Bou n d F m oc-Am in o Acid s
(Sch em e 4)a
Sch em e 6
resin
starting
material
support-bound
Fmoc-amino acid
dipeptide yield,b
HPLC
product
%
purity,c %
15c
15d
15e
15b
15f
15g
15h
15i
Fmoc-Ala
18c
18d
18e
18b
18f
18g
18h
18i
90
83
69d
78
54d
90
93
89
-
91
89
91
89
78
89
95
96
96
83
89
88
94
93
89
90
87
92
94
96
-
83
90
95
94
75
91
93
95
94
97
94
92
Fmoc-Arg(Pbf)
Fmoc-Asn(Trt)
Fmoc-Asp(O-t-Bu)
Fmoc-Cys(Trt)
Fmoc-Glu(O-t-Bu)
Fmoc-Gln(Trt)
Fmoc-Gly
Fmoc-His(Trt)
Fmoc-His(Boc)
Fmoc-Ile
Fmoc-Leu
Fmoc-Lys(Boc)
Fmoc-Met
Fmoc-Phe
Fmoc-Pro
15j
15k
15l
18j
18k
18l
15m
15n
15o
15a
15p
15q
15r
15s
15t
18m
18n
18o
18a
18p
18q
18r
18s
18t
Fmoc-Ser(t-Bu)
Fmoc-Thr(t-Bu)
Fmoc-Trp(Boc)
Fmoc-Tyr(t-Bu)
Fmoc-Val
15u
18u
a
Abbreviations used: Pbf ) 2,2,4,6,7-pentamethyldihydroben-
b
zofuran-5-sulfonyl. Yields of analytically pure material based
upon BnNH2 as the limiting agent. c Reversed-phase HPLC analy-
d
sis with detection at 254 nm. Unsatisfactory elemental analysis.
dipeptide product was obtained upon activation and
cleavage. It is likely that, in this case, alkylation of the
imidazole functionality occurs in the iodoacetonitrile
activation step and decomposition follows. However, high
loading efficiencies and high yields of dipeptide product
are observed when commercially available Fmoc-His-
(Boc)-OH is employed.
amino acids have been subjected to a short synthesis
sequence to provide support-bound dipeptides. Treatment
with iodoacetonitrile then provides an activated N-
cyanomethyl derivative that can be cleaved with a variety
of nucleophiles to provide dipeptide C-terminal deriva-
tives. Cleavage with amines, amino acid esters, and
aniline proceeds in high yields to provide amide, peptide,
and anilide products, respectively. All 20 of the protei-
nogenic amino acids, when suitably protected, are com-
patible with the loading and activation steps.
These methods should be applicable to the solid-phase
synthesis of several classes of small organic molecules
in which amino acids are employed as the initial linker-
bound building block. As well, these methods are clearly
applicable to solid-phase peptide chemistry for use in
segment condensation strategies and for the synthesis
of cyclic peptides and peptide amides. In future work, we
will apply these general strategies to synthesize fluoro-
genic peptide substrates for the preparation of positional-
scanning combinatorial libraries to determine protease
specificities.24
Disp la cem en t w ith Va r iou s Nu cleop h iles. The
condensation of amino acid methyl esters with activated
support-bound Boc-amino acids to provide dipeptide
esters (Table 1) and the cleavage of support-bound
dipeptides 17a -u with benzylamine (Scheme 4, Table 3)
occur at ambient temperature and have been demon-
strated to be racemization-free and high yielding. Poor
nucleophiles such as aniline also displace 17a to provide
peptide anilide 20 in 84% yield, although forcing condi-
tions are required (Scheme 6). Attempts to displace 17a
with 4-nitroaniline failed to give the desired product.
Fluorogenic peptide substrates derived from 4-nitro-
aniline and 7-amino-4-methylcoumarin have many uses
in enzymology. Because displacement of 17a with 7-amino-
4-methylcoumarin also failed to provide the anilide
product, we sought an alternative route to provide these
fluorogenic peptide substrates. Anilide 22, prepared by
coupling 7-amino-4-methylcoumarin and lysine, can be
condensed with 17a at ambient temperatures to provide
a high yield of tripeptide 23.
Exp er im en ta l Section
Con clu sion
Gen er a l Meth od s. Unless otherwise noted, materials were
obtained from commercial suppliers and used without further
purification. Aminomethyl Merrifield resin was purchased
from Novabiochem, and the aminomethyl substitution level
of the resin was determined (0.84 mequiv/gram) by exhaustive
An alkanesulfonamide linker has been developed for
solid-phase synthesis efforts. Numerous Fmoc-amino acid
derivatives can be loaded efficiently without significant
levels of racemization being observed. To load sterically
hindered amino acids that typically provide less efficient
couplings, a double coupling protocol provides useful
levels of substitution. A number of the support-bound
(24) Rano T. A.; Timkey, T.; Peterson, E. P.; Rotonda, J .; Nicholson,
D. W.; Becker, J . W.; Chapman, K. T.; Thornberry, N. A. Chem. Biol.
1997, 4, 149.

Alkanesulfonamide “Safety-Catch” Linker
J . Org. Chem., Vol. 64, No. 7, 1999 2327
acylation with Fmoc-Ala-OH followed by a spectrophotometric
Fmoc-quantitation assay.19 Fmoc-amino acids were purchased
from Novabiochem. Anhydrous NMP and DMF were pur-
chased from Aldrich. Tetrahydrofuran (THF) was distilled
under N2 from sodium/benzophenone prior to use, and CH2-
Cl2 was distilled under N2 from CaH2. Iodoacetonitrile and
chloroform were filtered through a small plug of basic alumina
prior to use. Chromatography was carried out using Merck
60 230-240 mesh silica gel according to the procedure of
Still.25 Thin-layer chromatography was carried out on Merck
60 F254 250-µm silica gel plates. IR spectra were recorded neat
(for oils) and as films from CH2Cl2 or CHCl3 (for crystalline
compounds), and only partial data are reported. 1H and 13C
NMR spectra were acquired in CDCl3, unless otherwise stated.
NMR chemical shifts are reported in ppm downfield from an
internal solvent peak, or TMS, and J values are in hertz.
Elemental analyses were performed by M-H-W Labs, Phoe-
nix, AZ. All solid-phase yields reported are based upon the
initial aminomethyl substitution level of the resin and con-
stitute a mass balance of analytically pure material from
support, unless otherwise stated. All HPLC assays were
performed utilizing a Rainin Dynamax Microsorb C18 reversed-
phase analytical column.
Gen er a l Meth od s for Solid -P h a se Syn th esis. Overhead
mechanical stirring was utilized for reactions in which >2 g
resin was employed. When employing overhead stirring, it is
important that the stirring paddle does not contact the bottom
of the flask to prevent degradation of the resin. Reactions
employing <2 g of resin were performed in 24/40 50 mL round-
bottom flasks or fritted polypropylene syringe cartridges with
agitation provided by a magnetic stir bar (Fisher Scientific,
13 mm × 3.2 mm octagonal stir bar) or a stream of N2,
respectively. Solvents were not distilled for resin washes and
were removed from the resin by a filtration cannula (Phar-
macia, Uppsala, Sweden) for reactions performed in round-
bottom flasks or by suction provided by an aspirator for
reactions performed in syringe cartridges. In cases where >2
g of resin was employed, washes were applied to the resin in
a fritted glass funnel. Individual resin washes endured 4-5
min, unless otherwise stated.
mL). The flask was cooled with an ice bath, and ammonia gas
was bubbled in slowly until the formation of precipitate ceased.
The solution was concentrated, and H2O (10 mL) was added.
The aqueous layer was extracted with hot EtOAc (3 × 50 mL),
and the organic layers were pooled, dried with Na2SO4, and
concentrated to give 3.4 g (81%) of a light yellow oil that
solidified upon standing. The product was sufficiently pure by
NMR analysis and was taken on to the next step. To provide
a sample of analytical purity, 2 g of material was purified by
column chromatography (5 cm × 20 cm eluted with 70:30
EtOAc/hexanes) to yield 9 as a colorless solid: mp 49-50 °C;
IR 3323, 1734 cm-1 1H NMR (400 MHz, DMSO-d6) δ 1.89
;
(quintet, 2, J ) 7.5), 2.45 (t, 2, J ) 7.5), 2.96 (t, 2, J ) 7.5),
3.56 (s, 3), 6.78 (s, 2); 13C NMR (101 MHz, DMSO-d6) δ 19.7,
31.9, 51.8, 53.9, 173.2. Anal. Calcd for C5H11SO4N: C, 33.14;
H, 6.19; N, 7.73. Found: C, 32.95; H, 6.04; N, 7.50.
3-Ca r boxyp r op a n esu lfon a m id e (10). To a 100 mL round-
bottom flask were added 9 (3.85 g, 21.3 mmol) and an aqueous
KOH solution (1.8 M, 25 mL). The flask was fitted with a
condenser, and the mixture was heated at reflux for 4 h
followed by cooling with an ice bath. An aqueous HCl solution
was added dropwise (1.8 M, 25 mL), and the solution was
concentrated under aspirator pressure to a volume of 2-3 mL.
An equal volume of acetone was then added, and the KCl salts
were filtered. The solution was concentrated and dried in a
desiccator under vacuum with P2O5 to give 3.0 g (85%) of 10
as a colorless solid. A sample was prepared for elemental
analysis by filtration through a plug of silica (20:80 MeOH/
CH2Cl2): mp 93-95 °C; IR 1738 cm-1 1H NMR (400 MHz,
;
DMSO-d6) δ 1.84 (quintet, 2, J ) 7.4), 2.34 (t, 2, J ) 7.4), 2.95
(t, 2, J ) 7.4), 6.75 (s, 2); 13C NMR (101 MHz, DMSO-d6) δ
19.7, 32.7, 54.1, 78.7, 174.3. Anal. Calcd for C4H9SO4N: C,
28.73; H, 5.42; N, 8.38. Found: C, 28.66; H, 5.52; N, 8.16.
Alk a n esu lfon a m id e Resin 11. To a 250 mL three-neck
round-bottom flask fitted with an overhead stirrer were added
aminomethyl polystyrene resin (15 g, 5.9 mmol), 10 (3.1 g, 16
mmol), DICI (2.4 mL, 16 mmol), HOBt (2.1 g, 16 mmol), and
DMF (130 mL). After the slurry was stirred for 4 h, the
reaction was complete as determined by a bromophenol blue
test.19 The resin was washed with DMF (3 × 100 mL), CH2Cl2
(3 × 100 mL), and MeOH (3 × 100 mL) and dried in a
desiccator under vacuum with P2O5.
Iod oa ceton itr ile Activa tion . A general procedure for
iodoacetonitrile activation to prepare a support-bound N,N-
cyanomethylacylsulfonamide is described. The resin-bound
N-acylsulfonamide (500 mg, 0.25 mmol) was washed with
several portions of NMP. To the swollen resin were added NMP
(4 mL) and i-Pr2EtN (240 µL, 1.25 mmol). After filtration
through an alumina basic plug proir to use, iodoacetonitrile
(367 µL, 5 mmol) was added to the reaction mixture, and the
reaction flask was shielded from light. The resin was agitated
for 24 h, filtered, and washed with NMP (5 × 5 mL, >10 min
wash times) and CH2Cl2 (3 × 5 mL) or THF (3 × 5 mL).
Dim eth yl 4,4′-Dith iobu tyr a te (7). To a 500 mL round-
bottom flask were added 4,4′-dithiobutyric acid 6 (21 g, 0.089
mol), SOCl2 (15 mL, 0.20 mol), and MeOH (200 mL). The flask
was fitted with a reflux condenser, and the solution was heated
at reflux for 4 h, after which the flask was cooled with an ice
bath. Water (100 mL) was added very slowly, and the solution
was extracted with CH2Cl2 (2 × 200 mL). The organic layer
was washed with 1 M NaHCO3 (3 × 200 mL), dried with Na2-
SO4, and concentrated to give 23.5 g of unpurified material.
The crude material was distilled (bp 136-138 °C, 0.65 mm)
to afford 21.4 g (90%) of 7 as a colorless oil: IR 1738 cm-1; 1H
NMR (400 MHz) δ 2.00 (quintet, 4, J ) 7.2), 2.42 (t, 4, J )
7.2), 2.69 (t, 4, J ) 7.2), 3.65 (s, 6); 13C NMR (101 MHz) δ
24.2, 32.3, 37.8, 51.6, 173.4. Anal. Calcd for C10H18S2O4: C,
45.09; H, 6.81. Found: C, 44.88; H, 6.64.
Op tim iza tion of Resin 11 Loa d in g P r oced u r e (Ta ble
1). Coupling experiments were performed using a symmetrical
anhydride, a pentafluorophenol ester, or Boc-Phe-OH (0.3-
0.5 M) and the solvent, base, and additives listed in Table 1
with resin 11 (200 mg, 0.14 mmol). Typically, a 2-4 h reaction
time was employed, after which the resin was washed with
NMP (3 × 4 mL) and activated for cleavage with iodoaceto-
nitrile. Leu-OMe (124 mg, 0.85 mmol) in CH2Cl2 (1.5 mL) that
had been free-based by extraction was added to the reaction
flask, and the mixture was stirred for 12 h. The resin was
filtered and washed with CH2Cl2 (3 × 5 mL), and the organic
layers were combined, washed with 0.1 M NaHSO4 (2 × 20
mL), and dried with Na2SO4. The unpurified material was
subjected to HPLC analysis to determine the percent race-
mization in the initial coupling step. Optically pure L,L- and
D,L-epimers of Boc-Phe-Leu-OMe were prepared as standards
in solution using HATU and collidine in DMF26 and subjected
to HPLC analysis (MeOH/H2O-0.1% TFA, 10%-100% for 60
min, 1 mL/min, 260 nm). The L,L-epimer eluted at 48.6 min,
Meth yl 4-(Ch lor osu lfon yl)bu tyr a te (8). To a 300 mL
three-neck round-bottom flask were added 7 (14.6 g, 55.0
mmol) and H2O (100 mL). Cl2 gas was bubbled in slowly with
vigorous stirring for 5 h. It is important that the chlorine gas
is added at a slow rate. After N2 gas was bubbled through the
solution for 1 h to remove excess Cl2, the solution was extracted
with CH2Cl2 (2 × 100 mL), dried with Na2SO4, and concen-
trated to give 14.6 g (67%) of unpurified material. The product
was sufficiently pure by NMR analysis and was taken on to
the next step. A small amount was distilled (bp 98-99 °C, 0.65
mm) to provide a sample for elemental analysis: IR 1733 cm-1
;
1H NMR (400 MHz) δ 2.33 (quintet, 2, J ) 6.9), 2.42 (t, 2, J )
6.9), 3.68 (s, 3), 3.78 (t, 2, J ) 6.9); 13C NMR (101 MHz) δ
19.8, 31.0, 52.0, 64.1, 172.1. Anal. Calcd for C5H9SO4: C, 29.93;
H, 4.52. Found: C, 29.74; H, 4.61.
Meth yl 4-Su lfa m oylbu tyr a te (9). To a 100 mL round-
bottom flask were added 8 (4.65 g, 23.2 mmol) and Et2O (50
(25) Still, W. C.; Kahn, M.; Mitra, A. J . Org. Chem. 1978, 43, 2923.
(26) Carpino, L. A.; Elfaham, A. J . Org. Chem. 1994, 59, 695.

2328 J . Org. Chem., Vol. 64, No. 7, 1999
Backes and Ellman
and the D,L-epimer eluted at 49.3 min. For cases (Table 1) in
which yields are given, purification on silica gel (1.5 × 10 cm
eluted with 20:80 EtOAc/hexanes) provided Boc-L-P h e-L-Leu -
OMe (11): mp 101-104 °C; IR 3294, 2959, 1751, 1684, 1652
156.6, 170.2, 171.1, 172.8. Anal. Calcd for C29H39N3O6: C,
66.20; H, 7.48; N, 7.99. Found: C, 66.08; H, 7.60; N, 7.66.
P r ep a r a tion of Dip ep tid e Am id es 18a -u . The general
procedures for Fmoc deprotection, Boc-Phe-OH coupling, io-
doacetonitrile activation, and BnNH2 displacement were fol-
lowed with the exception that a limiting amount of BnNH2
was employed in the cleavage step. A stock solution of freshly
distilled BnNH2 in THF (100 mg in a 25 mL volumetric flask,
0.037 M) was prepared, and 1.00 mL (0.037 mmol) of this
solution was added to resins 17a -u . The resin-containing flask
was flushed with argon gas, and the reaction mixture was
stirred for 24 h. The solution was then filtered through a small
silica plug, and several volumes of THF (total volume 25 mL)
were applied. The solution was subjected to reversed-phase
HPLC analysis (MeOH/H2O-0.1% TFA, 10-100% for 30 min,
1 mL/min, 254 nm detection for 40 min) to determine the
purity. The samples were then concentrated to obtain a yield
based upon BnNH2. The sample was submitted for 1H NMR
and elemental analysis without further purification.
cm-1 1H NMR (300 MHz) δ 0.90 (d, 6, J ) 6.6), 1.40-1.44
;
(m, 10), 1.50-1.53 (m,1), 1.52-1.55 (m, 1), 3.07 (t, 2, J ) 7.0),
3.69 (s, 3), 4.33-4.35 (m, 1), 4.50-4.52 (m, 1) 4.97 (bs, 1), 6.25
(d, 1, J ) 8.1), 7.18-7.31 (m, 5); 13C NMR (101 MHz) δ 21.9,
22.7, 24.6, 28.2, 38.1, 41.7, 50.7, 55.7, 52.2, 80.2, 126.9, 128.6,
129.4, 136.6, 155.6, 170.9, 172.8.
Op tim ized Resin 11 Loa d in g P r oced u r e for F m oc-
Am in o Acid s. To a 50 mL round-bottom flask were added
resin 11 (200 mg, 0.15 mmol), CHCl3 (1.7 mL) that had been
filtered through an alumina basic plug, i-Pr2EtN (143 µL, 0.750
mmol), and an Fmoc-amino acid (0.45 mmol). The reaction
mixture was stirred for 10 min followed by cooling to -20 °C.
After 20 min, PyBop (234 mg, 0.450 mmol) was added to the
reaction mixture as a solid, and the reaction mixture was
stirred for 8 h, filtered, and washed with CHCl3 (3 × 5 mL). It
is important to terminate the reaction after 8 h in order to
minimize racemization. For coupling with Fmoc-Gly-OH, 4
equiv of amino acid and 6 equiv of i-Pr2EtN were employed
with DMF as solvent, and the reaction mixture was allowed
to come to room temperature overnight after the 8 h coupling
period.
Gen er a l P r oced u r e for th e P r ep a r a tion of Dip ep tid e
P r od u cts 18a a n d 18b. To a 12 mL syringe reactor cartridge
was added a support-bound Fmoc-amino acid resin 15a or 15b
(150 mg, ca. 0.075 mmol), followed by solvation with DMF. A
20% piperidine in DMF solution (2 mL) was added to the
reactor, and the mixture was agitated for 40 min. The resin
was then filtered and washed with DMF (3 × 2 mL). To a
separate 10 mL flask were added Boc-L-Phe-OH (160 mg, 0.60
mmol), HOBt (81 mg, 0.60 mmol), DICI (94 µL, 0.60 mmol),
and DMF (2 mL). After a 2-3 min premix period, the solution
was added to the resin-containing syringe reactor followed by
agitation for 2-3 h. A bromophenol blue test was employed
to determine reaction completion.19 The resin was filtered and
washed with DMF (3 × 3 mL) and NMP (3 × 3 mL). After
activation with iodoacetonitrile, the resin was washed with
NMP (4 × 3 mL, 10 min/wash), CH2Cl2 (3 × 3 mL), and THF
(3 × 3 mL) and transferred to a 50 mL round-bottom flask. To
the resin-containing flask were added THF (2 mL) and
benzylamine (40 µL, 0.38 mmol), and the reaction mixture was
stirred for 4 h. The mixture was taken up in CH2Cl2 (20 mL),
extracted with 1 M NaHSO4 (3 × 20 mL), dried with Na2SO4,
and concentrated. The unpurified material was analyzed for
epimerization by HPLC assay in which authentic samples of
L,L-epimers and D,L-epimers of the dipeptide amides served
as standards. Silica gel column chromatography followed to
provide the respective products in analytically pure form.
Boc-L-P h e-L-Ala -NHBn (18c). To N-cyanomethylated resin
17c was added benzylamine in THF (0.028 M solution, 1.67
mL, 0.047 mmol) to provide 18 mg (90%) of compound 18c as
a colorless solid: reversed-phase HPLC analysis (22.4 min,
1
94%); TLC (40:60 EtOAc/hexanes, Rf ) 0.44); H NMR (300
MHz) δ 1.34 (d, 3, J ) 7.4), 1.35 (s, 9), 3.01-3.04 (m, 2), 4.29-
4.31 (m, 1) 4.38 (d, 2, J ) 5.8), 4.44-4.51 (m, 1), 4.89 (bs, 1),
6.36 (d, 1, J ) 7.2), 6.59 (bs, 1), 7.15 (d, 2, J ) 6.1), 7.22-7.34
(m, 8). Anal. Calcd for C24H31N3O4: C, 67.70; H, 7.34; N, 9.87.
Found: C, 67.90; H, 7.17; N, 9.68.
Boc-L-P h e-L-Ar g(P bf)-NHBn (18d ). To N-cyanomethy-
lated resin 17d was added benzylamine in THF (0.037 M
solution, 1.0 mL, 0.037 mmol) to provide 25 mg (86%) of
compound 18d as a colorless solid: reversed-phase HPLC
analysis (26.1 min, 93%); TLC (70:30 EtOAc/hexanes, Rf )
0.20); 1H NMR (300 MHz) δ 1.30 (s, 9), 146-1.53 (m, 4), 165-
1.69 (m, 1), 1.81 (m, 6), 1.83-1.89 (m, 1), 2.08 (m, 3), 2.47 (s,
3), 2.55 (s, 3), 2.83-2.93 (m,3), 3.07 (dd, 1, J ) 5.3, J ) 13.7),
3.22-3.25 (m, 2), 4.29-4.36 (m, 3), 4.47-4.50 (m, 1), 5.18 (bs,
1), 6.19-6.26 (m, 3), 7.10-7.12 (m, 2), 7.17-7.31 (m, 8), 7.33-
7.42 (m, 2). Anal. Calcd for C41H56N6SO7: C, 63.30; H, 7.26;
N, 10.80. Found: C, 63.53; H, 7.31; N, 10.91.
Boc-L-P h e-L-Asn (Tr t)-NHBn (18e). To N-cyanomethy-
lated resin 17e was added benzylamine in THF (0.037 M
solution, 1.0 mL, 0.037 mmol) to provide 16 mg (69%) of
compound 18e as a colorless solid: reversed-phase HPLC
analysis (31.5 min, 89%); TLC (70:30 EtOAc/hexanes, Rf )
0.50); 1H NMR (300 MHz) δ 1.26 (s, 9), 2.50 (dd, 1, J ) 5.5, J
) 15.3), 2,87 (dd, 1, J ) 8.0, J ) 13.7), 3.04-3.15 (m, 2), 4.22-
4.29 (m, 2), 4.40-4.49 (m, 1), 4.73-4.76 (m, 1), 4.80 (d, 1, J )
4.1), 6.89 (s, 1), 7.05 (bs, 1), 7.11-7.17 (m, 11), 7.22-7.33 (m,
14), 7.58-7.60 (bs, 1).
Boc-L-P h e-L-Asp (O-t-Bu )-NHBn (18b). To N-cyanomethy-
lated resin 17b was added benzylamine in THF (0.037 M
solution, 1.0 mL, 0.037 mmol) to provide 15 mg (79%) of
compound 18b as a colorless solid: reversed-phase HPLC
analysis (26.4 min, 91%); TLC (40:60 EtOAc/hexanes, Rf )
Boc-L-P h e-L-P h e-NHBn (18a ). Cleavage of resin 17a (150
mg, 0.088 mmol) provided 39 mg (85%) of 18a as a colorless
solid after purification on silica gel (1.5 × 10 cm eluted with
30:70 EtOAc/hexanes): mp 129-131 °C; IR 1687, 1645, 1530
1
cm-1; H NMR (300 MHz) δ 1.23 (s, 9), 2.87 (dd, 1, J ) 7.0, J
1
0.36); H NMR data were identical to those described previ-
) 13.5), 2.98-3.01 (m, 2), 3.25 (dd, 1, J ) 4.0, J ) 13.5), 4.22-
4.29 (m, 2), 4.33 (dd, 1, J ) 5.8, J ) 13.8), 4.66-4.4.67 (m, 1),
4.77 (bs, 1), 6.3 (bs, 1), 6.44 (bs, 1), 7.00-7.03 (m, 2), 7.09 (d,
2, J ) 7.8), 7.14 (d, 2, J ) 6.0), 7.16-7.20 (m, 9); 13C NMR
(101 MHz) δ 27.2, 37.2, 37.4, 42.7, 54.6, 57.9, 79.7, 126.4, 126.4,
126.7, 127.0, 127.1, 128.0, 128.1, 128.2, 128.9, 129.0, 136.9,
138.1, 165.1, 171.7, 172.8. Anal. Calcd for C30H35N3O4: C,
71.80; H, 7.03; N, 8.38. Found: C, 71.60; H, 6.83; N, 8.39.
Boc-L-P h e-L-Asp (O-t-Bu )-NHBn (18b). Cleavage of resin
17b (150 mg, 0.085 mmol) provided 37 mg (83%) of 18b as a
colorless solid after purification on silica gel (1.5 × 10 cm
eluted with 30:70 EtOAc/hexanes): mp 153-155 °C; IR 1724,
1691, 1648, 1535 cm-1; 1H NMR (300 MHz) δ 1.32 (s, 9), 1.39
(s, 9), 2.43 (dd, 1, J ) 5.7, 17.0), 2.97-2.99 (m, 2), 3.10 (dd, 1,
J ) 5.4, 13.5), 4.26-4.28 (m, 2), 4.40 (dd, 1, J ) 6.1, 15.0),
4.74-4.76 (m, 1), 4.86 (bs, 1), 6.99 (bs, 1), 7.17-7.30 (m, 11);
13C NMR (101 MHz) δ 26.9, 27.2, 36.5, 37.3, 42.8, 49.8, 56.4,
79.6, 81.2, 126.4, 126.8, 127.0, 128.0, 128.1, 129.0, 137.1, 138.2,
ously. Anal. Calcd for C29H39N3O6: C, 66.20; H, 7.48; N, 7.99.
Found: C, 66.02; H, 7.49; N, 7.76.
Boc-L-P h e-L-Cys(Tr t)-NHBn (18f). To N-cyanomethylated
resin 17f was added benzylamine in THF (0.037 M solution,
1.0 mL, 0.037 mmol) to provide 14 mg (54%) of compound 18f
as a colorless solid: reversed-phase HPLC analysis (22.4 min,
1
87%); TLC (40:60 EtOAc/hexanes, Rf ) 0.42); H NMR (300
MHz) δ 1.23 (s, 9), 2.41 (dd, 1, J ) 5.0, 12.4), 2.84 (dd, 1, J )
7.8, 14.0), 2.90-2.93 (m, 1) 3.03 (dd, 1, J ) 5.9, 14.0), 4.20-
4.33 (m, 3), 4.45 (dd, 1, J ) 5.7, 16.6), 4.76 (m, 1), 6.29 (d, 1,
J ) 7.0), 6.73 (bs, 1), 7.13 (d, 2, J ) 7.6), 7.18-7.33 (m, 15)
7.36-7.39 (m, 8).
Boc-L-P h e-L-Gln (Tr t)-NHBn (18g). To N-cyanomethy-
lated resin 17g was added benzylamine in THF (0.037 M
solution, 1.00 mL, 0.037 mmol) to provide 25 mg (93%) of
compound 18g as a colorless solid: reversed-phase HPLC
analysis (29.1 min, 94%); TLC (70:30 EtOAc/hexanes, Rf )
0.46); 1H NMR (300 MHz) δ 1.30 (s, 9), 1.82-1.99 (m, 2), 2.25-

Alkanesulfonamide “Safety-Catch” Linker
J . Org. Chem., Vol. 64, No. 7, 1999 2329
2.34 (m, 1), 2.45-2.53 (m, 1), 2.86 (dd, 1, J ) 8.0, 13.9), 3.06
(dd, 1, J ) 5.4, 13.9), 4.23-4.29 (m, 2), 4.31-4.35 (m, 2), 4.85
(d, 1 J ) 6.9), 6.92 (bs, 1), 7.04 (s, 1), 7.10-7.13 (m, 2), 7.17-
7.31 (m, 23), 7.37 (bs, 1). Anal. Calcd for C45H48N4O5: C, 74.50;
H, 6.67; N, 7.73. Found: C, 74.48; H, 6.57; N, 7.54.
a colorless solid: reversed-phase HPLC analysis (25.1 min,
75%); TLC (40:60 EtOAc/hexanes, Rf ) 0.24); H NMR (300
MHz) δ 1.34 (s, 9), 2.00-2.08 (m, 5), 2.42-2.48 (m, 2), 3.06 (d,
2, J ) 6.5), 4.26-4.32 (m, 1), 4.38 (d, 2, J ) 5.6), 4.59-4.63
(m, 1), 4.88 (bs, 1), 6.81-6.83 (m, 2), 7.15-7.18 (m, 2), 7.22-
7.33 (m, 8). Anal. Calcd for C26H35N3O4S: C, 64.30; H, 7.26;
N, 8.65. Found: C, 64.15; H, 7.33; N, 8.38.
1
Boc-L-P h e-L-Glu (O-t-Bu )-NHBn (18h ). To N-cyanomethy-
lated resin 17h was added benzylamine in THF (0.037 M
solution, 1.00 mL, 0.037 mmol) to provide 18 mg (90%) of
compound 18h as a colorless solid: reversed-phase HPLC
analysis (24.5 min, 92%); TLC (40:60 EtOAc/hexanes, Rf )
Boc-L-P h e-L-P h e-NHBn (18p). To N-cyanomethylated resin
17p was added benzylamine in THF (0.028 M solution, 1.67
mL, 0.047 mmol) to provide 21 mg (88%) of compound 18p as
a colorless solid: reversed-phase HPLC analysis (24.9 min,
1
0.32); H NMR (300 MHz) δ 1.35 (s, 9), 1.42 (s, 9), 1.90-1.95
1
(m, 1), 2.05-2.09 (m, 1), 2.21-2.23 (m, 1), 2.30-2.33 (m, 1),
3.04 (d, 2, J ) 6.6), 4.30-4.33 (m, 1), 4.37-4.46 (m, 3), 4.93
(d, 1, J ) 4.8), 6.86 (bs, 1), 7.08 (bs, 1), 7.14-7.17 (m, 2), 7.19-
7.27 (m, 8). Anal. Calcd for C30H41N3O6: C, 66.70; H, 7.66; N,
7.79. Found: C, 67.00; H, 7.78; N, 7.86.
91%); TLC (40:60 EtOAc/hexanes, Rf ) 0.32); H NMR data
were identical to those provided previously. Anal. Calcd for
C30H35N3O4: C, 71.80; H, 7.03; N, 8.38. Found: C, 71.69; H,
6.87; N, 8.34.
Boc-L-P h e-L-P r o-NHBn (18p). To N-cyanomethylated resin
17p was added benzylamine in THF (0.028 M solution, 1.67
mL, 0.047 mmol) to provide 20 mg (95%) of compound 18p as
a colorless solid: reversed-phase HPLC analysis (24.3 min,
Boc-L-P h e-L-Gly-NHBn (18i). To N-cyanomethylated resin
17i was added benzylamine in THF (0.028 M solution, 1.67
mL, 0.047 mmol) to provide 17 mg (89%) of compound 18i as
a colorless solid: reversed-phase HPLC analysis (23.4 min,
1
93%); TLC (70:30 EtOAc/hexanes, Rf ) 0.44); H NMR (300
1
96%); TLC (70:30 EtOAc/hexanes, Rf ) 0.32); H NMR (300
MHz) δ 1.40 (s, 9), 1.82-1.86 (m, 2), 2.32-2.35 (m, 1), 2.92 (d,
2, J ) 7.3), 2.66-2.98 (m, 1), 3.35-3.43 (m, 1), 3.50-3.60 (m,
1), 4.3 (dd, 1, J ) 5.5, 14.9), 4.47 (dd, 1, J ) 6.1, 14.9), 4.57-
4.69 (m, 2), 5.26 (d, 1, J ) 10.5), 7.07 (bs, 1), 7.10-7.35 (m,
10). Anal. Calcd for C26H33N3O4: C, 69.10; H, 7.37; N, 9.31.
Found: C, 69.01; H, 7.10; N, 9.33.
MHz) δ 1.33 (s, 9), 2.97 (dd, 1, J ) 5.5, 15.5), 3.09 (dd, 1, J )
6.6, 15.5), 3.82 (dd, 1, J ) 5.7, 16.7), 3.96 (dd, 1, J ) 6.1, 16.7),
4.22-4.29 (m, 1), 4.35 (dd, 1, J ) 5.3, 15.0), 4.44 (dd, 1, J )
6.2, 15.0), 6.52 (bs, 1), 4.97 (d, 1, J ) 6.3), 6.81 (bs, 1), 7.15-
7.18 (m, 2), 7.22-7.33 (m, 8). Anal. Calcd for C23H29N3O4: C,
67.10; H, 7.10; N, 10.20. Found: C, 67.20; H, 6.90; N, 10.02.
Boc-L-P h e-L-His(Boc)-NHBn (18k ). To N-cyanomethy-
lated resin 17k was added benzylamine in THF (0.028 M
solution, 1.67 mL, 0.047 mmol) to provide 20 mg (91%) of
compound 18k as a colorless solid: reversed-phase HPLC
analysis (26.4 min, 83%); TLC (70:30 EtOAc/hexanes, Rf )
0.28); 1H NMR (300 MHz) δ 1.22 (s, 9), 1.66 (s, 9), 2.77 (dd, 1,
J ) 4.9, 14.7), 2.99 (dd, 1, J ) 8.6, 14.1), 3.19 (dd, 1, J ) 4.9,
14.1), 3.29 (dd, 1, J ) 4.9, 14.7), 4.28 (dd, 1, J ) 4.7, 15.3),
4.33-4.36 (m, 1), 4.52 (dd, 1, J ) 7.1, 15.3), 4.70-4.79 (m, 1),
5.01 (d, 1, J ) 4.7), 6.92 (d, 2, J ) 7.5), 7.14-7.36 (m, 10),
7.88 (s, 1), 8.31 (d, 1, J ) 7.2). Anal. Calcd for C32H41N5O6: C,
64.96; H, 6.98; N, 11.84. Found: C, 64.96; H, 7.18; N, 11.84.
Boc-L-P h e-L-Ile-NHBn (18l). To N-cyanomethylated resin
17l was added benzylamine in THF (0.037 M solution, 1.00
mL, 0.037 mmol) to provide 18 mg (89%) of compound 18l as
a colorless solid: reversed-phase HPLC analysis (27.3 min,
Boc-L-P h e-L-Ser (t-Bu )-NHBn (18q). To N-cyanomethy-
lated resin 17q was added benzylamine in THF (0.028 M
solution, 1.67 mL, 0.047 mmol) to provide 22 mg (96%) of
compound 18q as a colorless solid: reversed-phase HPLC
analysis (27.1 min, 95%); TLC (40:60 EtOAc/hexanes, Rf )
0.30); 1H NMR (300 MHz) δ 1.09 (s, 9), 1.28 (s, 9), 3.05 (dd, 1,
J ) 7.6, 14.0), 3.12 (dd, 1, J ) 6.1, 14.0), 3.26-3.30 (m, 1),
3.93-3.95 (m, 1), 4.30-4.34 (m, 2), 4.44-4.46 (m, 1), 4.57 (dd,
1, J ) 6.3, 15.2), 4.92 (bs, 1), 6.75 (d, 1, J ) 7.4), 6.98 (bs, 1),
7.17-7.20 (m, 2), 7.22-7.40 (m, 8). Anal. Calcd for
C
28H39N3O4: C, 67.50; H, 7.90; N, 8.44. Found: C, 67.34; H,
7.75; N, 8.29.
Boc-L-P h e-L-Th r (t-Bu )-NHBn (18r ). To N-cyanomethy-
lated resin 17r was added benzylamine in THF (0.028 M
solution, 1.67 mL, 0.047 mmol) to provide 23 mg (96%) of
compound 18r as a colorless solid: reversed-phase HPLC
analysis (28.9 min, 94%); TLC (40:60 EtOAc/hexanes, Rf )
1
1
90%); TLC (40:60 EtOAc/hexanes, Rf ) 0.38); H NMR (300
0.42); H NMR (300 MHz) δ 0.96 (d, 2, J ) 6.2), 1.16 (s, 9),
MHz) δ 0.83-0.87 (m, 6), 0.92-0.96 (m, 1), 1.35 (s, 9), 1.36-
1.39 (m, 1), 2.02-2.04 (m, 1), 3.03 (dd, 1, J ) 4.5, 17.8), 3.06
(dd, 1, J ) 3.9, 17.8), 4.26-4.28 (m, 2), 4.38 (d, 2, J ) 5.7), 4.9
(bs, 1), 6.43 (d, 1, J ) 8.3), 6.53 (bs, 1), 7.75-7.17 (m, 2), 7.19-
7.33 (m, 8). Anal. Calcd for C27H37N3O4: C, 69.30; H, 7.98; N,
8.94. Found: C, 69.45; H, 8.03; N, 8.87.
Boc-L-P h e-L-Leu -NHBn (18l). To N-cyanomethylated resin
17m was added benzylamine in THF (0.028 M solution, 1.67
mL, 0.047 mmol) to provide 21 mg (91%) of compound 18m as
a colorless solid: reversed-phase HPLC analysis (26.2 min,
91%); TLC (40:60 EtOAc/hexanes, Rf ) 0.40); 1H NMR δ 0.89
(d, 6, J ) 6.4), 1.36 (s, 9), 1.41-1.50 (m, 2), 1.67-1.73 (m, 1),
3.03 (dd, 1, J ) 2.8, 13.0), 3.06 (dd, 1, J ) 2.3, 13.0), 4.28-
4.31 (m, 1), 4.44 (d, 2, J ) 4.7), 4.48-4.50 (m, 1), 4.90 (bs, 1),
6.21 (d, 1, J ) 8.3), 6.55 (bs, 1), 7.17-7.19 (m, 2), 7.21-7.34
(m, 8). Anal. Calcd for C27H37N3O4: C, 69.30; H, 7.98; N, 8.99.
Found: C, 69.32; H, 7.77; N, 8.93.
1.36 (s, 9), 3.05-3.10 (m, 2), 4.25-4.27 (m, 2), 4.35-4.37 (m,
1), 4.41 (d, 2, J ) 5.7), 4.9 (bs, 1), 6.96 (d, 1, J ) 5.7), 7.17-
7.19 (d, 2, J ) 8.0), 7.22-7.34 (m, 9). Anal. Calcd for
C
29H40N3O5: C, 68.20; H, 7.90; N, 8.23. Found: C, 67.96; H,
7.96; N, 8.24.
Boc-L-P h e-L-Tr p (Boc)-NHBn (18s). To N-cyanomethy-
lated resin 17s was added benzylamine in THF (0.037 M
solution, 1.00 mL, 0.037 mmol) to provide 20 mg (83%) of
compound 18s as a colorless solid: reversed-phase HPLC
analysis (29.2 min, 97%); TLC (40:60 EtOAc/hexanes, Rf )
1
0.40); H NMR (300 MHz) δ 1.06 (s, 9), 1.64 (s, 9), 2.96-3.01
(m, 3), 3.36-3.38 (m, 1), 4.19-4.22 (m, 2), 4.33-4.35 (m, 1),
4.68 (d, 1, J ) 5.9), 4.75-4.79 (m, 1), 6.40-6.45 (m, 2), 7.0 (d,
2, J ) 6.1), 7.14-7.37 (m, 12), 8.10 (d, 1, J ) 8.1). Anal. Calcd
for C38H44N4O6: C, 69.90; H, 6.79; N, 8.58. Found: C, 69.56;
H, 6.84; N, 8.81.
Boc-L-P h e-L-Tyr (t-Bu )-NHBn (18t). To N-cyanomethy-
lated resin 17t was added benzylamine in THF (0.028 M
solution, 1.67 mL, 0.047 mmol) to provide 24 mg (89%) of
compound 18t as a colorless solid: reversed-phase HPLC
analysis (28.6 min, 94%); TLC (40:60 EtOAc/hexanes, Rf )
0.38); 1H NMR (300 MHz) δ 1.27 (s, 9), 1.31 (s, 9), 2.86 (dd, 1,
J ) 7.3, J ) 13.6), 2.96-3.05 (m, 2), 3.12 (dd, 1, J ) 8.8, J )
14.0), 4.22-4.38 (m, 3), 4.61-4.63 (m, 1), 4.83 (d, 1, J ) 6.0),
6.34 (bs, 1), 6.82 (d, 2, J ) 8.5), 6.92 (d, 2, J ) 8.3), 7.12-7.20
(m, 4), 7.30-7.35 (m, 7). Anal. Calcd for C34H43N3O5: C, 71.10;
H, 7.55; N, 7.32. Found: C, 71.10; H, 7.55; N, 7.32.
Boc-L-P h e-L-Lys(Boc)-NHBn (18n ). To N-cyanomethy-
lated resin 17n was added benzylamine in THF (0.028 M
solution, 1.67 mL, 0.047 mmol) to provide 24 mg (89%) of
compound 18n as a colorless solid: reversed-phase HPLC
analysis (25.3 min, 94%); TLC (70:30 EtOAc/hexanes, Rf )
1
0.50); H NMR (300 MHz) δ 1.23 (pentet, 2, J ) 7.5), 1.36 (s,
9) 1.40-1.42 (m, 10), 1.53-1.63, (m, 1), 1.86-1.98 (m, 2), 2.95-
3.05 (m, 5), 4.31-4.43 (m, 4), 4.64-4.66 (m, 1), 5.02 (bs, 1),
6.54 (d, 1, J ) 8.0), 6.70 (bs, 1), 7.14 (d, 2, J ) 5.8), 7.21-7.34
(m, 8). Anal. Calcd for C31H46N4O6: C, 65.20; H, 8.12; N, 9.82.
Found: C, 65.38; H, 7.96; N, 9.67.
Boc-L-P h e-L-Val-NHBn (18u ). To N-cyanomethylated resin
17u was added benzylamine in THF (0.037 M solution, 1.00
mL, 0.037 mmol) to provide 15 mg (88%) of compound 18u as
a colorless solid: reversed-phase HPLC analysis (23.7 min,
Boc-L-P h e-L-Met-NHBn (15o). To N-cyanomethylated resin
17o was added benzylamine in THF (0.037 M solution, 1.00
mL, 0.037 mmol) to provide 14 mg (78%) of compound 18o as

2330 J . Org. Chem., Vol. 64, No. 7, 1999
Backes and Ellman
1
92%); TLC (40:60 EtOAc/hexanes, Rf ) 0.36); H NMR (300
MHz) δ 0.82 (d, 3, J ) 6.9), 0.89 (d, 3, J ) 6.9), 1.34 (s, 9),
2.27-2.29 (m, 1), 3.02 (dd, 1, J ) 5.3, 19.4), 3.05 (dd, 1, J )
6.2, 19.4), 4.25-4.40 (m, 4), 4.49, (bs, 1), 6.43 (d, 1, J ) 8.2),
6.53 (bs, 1), 7.17-7.20 (m, 2), 7.22-7.33 (m, 8). Anal. Calcd
for C26H35N3O4: C, 68.80; H, 7.78; N, 9.26. Found: C, 68.65;
H, 7.67; N, 9.10.
mmol), DMF (1.5 mL), and coumarin derivative 22 (128 mg,
0.340 mmol). After being stirred for 15 h, the reaction mixture
was diluted with EtOAc (20 mL), extracted with 1 M NaHSO4
(3 × 20 mL), dried (Na2SO4), and concentrated. Purification
on silica gel (1.5 × 10 cm eluted with 50:50 to 90:10 EtOAc/
hexanes) provided 59 mg (86%) of 23 as a colorless solid: mp
1
143-145 °C; IR 1719, 1707, 1686, 1648, 1581, 1530 cm-1; H
Boc-L-P h e-L-P h e-NHP h (20). To a 50 mL round-bottom
flask were added N-cyanomethylated resin 17a (150 mg, 0.086
mmol), dioxane (1.2 mL), and aniline 20 (128 mg, 1.2 mmol).
The mixture was heated with stirring for 15 h at reflux, diluted
with CH2Cl2 (20 mL), extracted with 1 M NaHSO4 (3 × 20
mL), dried with Na2SO4, and concentrated. Purification on
silica gel (1.5 × 10 cm eluted with 30:70 to 50:50 EtOAc/
hexanes) provided 33 mg (84%) of 23 as a colorless solid: mp
NMR (300 MHz) δ 1.22-1.29 (m, 11), 1.41-1.56 (m, 10), 1.56-
1.62 (m, 1), 2.01-2.05 (m, 2), 2.38 (d, 2, J ) 1.0), 2.93-3.08
(m, 2), 3.10-3.17 (m, 2), 3.24-3.27 (m, 1), 4.16-4.17 (m, 1),
4.50-4.59 (m, 2), 4.67 (bs, 1), 4.98 (bs, 1), 6.16 (d, 2, J ) 1.0),
6.54 (bs, 1), 6.98-7.00 (m, 2), 7.11-7.19 (m, 3), 7.22-7.26 (m,
3), 7.29-7.34 (m, 2), 7.50 (d, 2, J ) 9.1), 7.83 (s, 1), 7.98 (s, 2),
8.99 (s, 1); 13C NMR (101 MHz) δ 18.6, 23.5, 28.1, 28.4, 29.4,
30.9, 36.4, 37.5, 40.2, 54.4, 54.7, 57.0, 79.1, 81.3, 107.6, 113.2,
115.9, 116.2, 124.9, 127.5, 128.7, 129.0, 129.1, 129.2, 129.3,
131.2, 135.4, 141.9, 152.5, 154.1, 156.0, 156.5, 161.4, 170.6,
171.0, 172.8. Anal. Calcd for C44H55N5O9: C, 66.20; H, 6.95;
N, 8.78. Found: C, 66.01; H, 6.59; N, 8.80.
1
183-185 °C; IR 1690, 1648, 1600, 1534 cm-1; H NMR (300
MHz) δ 1.25 (s, 9), 2.96-3.04 (m, 3), 3.33-3.36 (m, 1), 4.28-
4.30 (m, 1), 4.81-4.85 (m, 2), 6.39 (bs, 1), 7.02-7.18 (m, 3),
6.39 (bs, 1), 7.22-7.32 (m, 10), 7.31 (d, 2, J ) 6.9), 8.09 (s, 1);
13C NMR (101 MHz) δ 28.1, 37.5, 37.7, 54.3, 56.2, 80.9, 120.2,
124.4, 127.1, 127.3, 128.7, 128.8, 128.9, 129.3, 129.4, 136.0,
136.1, 137.6, 155.8, 168.8, 171.3. Anal. Calcd for C29H33N3O4:
C, 73.80; H, 7.05; N, 8.91. Found: C, 73.78; H, 7.17; N, 8.81.
Cou m a r in Tr ip ep tid e (23). To a 50 mL round-bottom
flask were added N-cyanomethylated resin 17a (150 mg, 0.086
Ack n ow led gm en t. Support from the NIH is grate-
fully acknowledged. B.J .B. thanks the NSF and Abbott
Laboratories for a graduate fellowship.
J O981990Y