Efficient synthesis of phosphonodepsipeptides derived from norleucine

Article abstract of DOI:10.1016/j.tet.2009.05.051

In the present work, we describe in detail an efficient solution synthesis of norleucine-derived phosphonopeptides mimicking the peptide sequences Nle-Gly(Ala) and Nle-Gly(Ala)-Val. The most efficient strategy involved use of the benzyl group. The synthesis was achieved through BOP-catalysed coupling of the monobenzyl ester of the N-Cbz-protected phosphonate derivative of norleucine with the hydroxyl moieties of derivatised l-lactic or glycolic acid. Subsequently, complete deprotection of the products was achieved in good yields by one-step Pd-catalysed hydrogenolysis. We also prepared the Fmoc-Nle-Ψ[PO(OH)O]-CH₂-COOH synthon and demonstrated that this precursor is a suitable building block for the solid-phase synthesis of cysteine-containing phosphonopeptides.

Full text of DOI:10.1016/j.tet.2009.05.051

Tetrahedron 65 (2009) 6090–6103
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Efficient synthesis of phosphonodepsipeptides derived from norleucine
ˇ
*
´
ˇ
ˇˇ´
´
ˇ
´
´
ˇ
ˇ´ ´ ˇ
Jan Pıcha, Milos Budesınsky, Ivona Hanclova, Miloslav Sanda, Pavel Fiedler, Vaclav Vanek, Jirı Jiracek
´
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, v.v.i., Flemingovo nam. 2, 166 10 Prague 6, Czech Republic
a r t i c l e i n f o
a b s t r a c t
Article history:
In the present work, we describe in detail an efficient solution synthesis of norleucine-derived phos-
phonopeptides mimicking the peptide sequences Nle-Gly(Ala) and Nle-Gly(Ala)-Val. The most efficient
strategy involved use of the benzyl group. The synthesis was achieved through BOP-catalysed coupling of
the monobenzyl ester of the N-Cbz-protected phosphonate derivative of norleucine with the hydroxyl
Received 30 January 2009
Received in revised form 6 May 2009
Accepted 21 May 2009
Available online 27 May 2009
moieties of derivatised
achieved in good yields by one-step Pd-catalysed hydrogenolysis. We also prepared the Fmoc-Nle-
[PO(OH)O]-CH2-COOH synthon and demonstrated that this precursor is a suitable building block for
the solid-phase synthesis of cysteine-containing phosphonopeptides.
Ó 2009 Elsevier Ltd. All rights reserved.
L-lactic or glycolic acid. Subsequently, complete deprotection of the products was
J
1. Introduction
synthesis of new phosphonate dipeptides for use as inhibitors of
VanX D,D-dipeptidase.6,23–26 The preparations of some phosphonate
compounds designed as potential antihypertensive agents are pro-
tected by patents.27–31
The most common strategy to prepare depsiphosphonopeptides
of Structure I (see below) consists of two crucial steps: (i) elonga-
tion of the C- and/or N-terminus of mixed esters, followed by (ii)
selective cleavage of the protecting group R3.11,12
Phosphinopeptides1,2 and phosphonamidopeptides3 belong to
the family of pseudopeptidic compounds, inwhich one peptide bond
is substituted bya non-hydrolysable phosphinate4 –PO(OH)–CH2– or
phosphonamidate3 –PO(OH)–NH– group. Phosphonopeptides,3,5
also called depsiphosphonopeptides,6 bearing the motif –PO(OH)–
O–, are another type of mimic in which a phosphonate ester replaces
the peptide bond. The resemblance of these phosphorus-containing
moieties to the structure of the peptide bond during enzyme-cata-
lysed hydrolysis enables effective mimicry of the structure of the
peptidic substrate in the transition state7–9 (TS). Thus, these com-
pounds may be potent inhibitors of peptidases.4 Phosphonopeptides
were found to be efficacious inhibitors of leucine aminopeptidase,10
pepsin,11,12 carboxypeptidase A9,13–15 and B,16 chymotrypsin,17 serine
proteases,18 thermolysin19 and penicillopepsin.20–22 Recently, it was
R2
O
H
N
Y
R1, R2 = alkyl, aryl
R3 = methyl, benzyl
X, Y = amino acids
P
O
X
OR3
O
R1
The synthesis of the above-mentioned mixed diesters can be ac-
complished by the condensation of an N-protected phosphonate
monoester with the hydroxyl moiety of a derivative of a glycolic,
lactic or mandelic acid using coupling agents such as DIAD/PPh3,31–34
BOP or PyBOP.35,36 The same products have also been prepared from
phosphonochloridates by treatment of monoesters with thionyl
chloride10–12,14,23,24 or by oxidative chlorination with carbon tetra-
chloride.11,18 Next, the demethylation of unsymmetrical diesters us-
ing TMSBr,12,20,22,31,32,34,35 LiSPr,10,23,24 LiOH14,19 or by hydrogenolysis
of the protecting benzyl group6,29 led to the desired phosphonic acid
monoesters. Alternatively, direct monoesterification starting from
phosphonic acid using thionyl chloride,16,25,37 BroP, TPyClU38 or
DCC27,30 has also been reported. Similar results were obtained by the
esterification of the corresponding phosphorous acid with DCC fol-
lowed by oxidationwith sodium periodate.28,39,40 A new approach to
the one-pot preparation of depsiphosphonopeptides involves
a multicomponent condensation of an aldehyde, benzylcarbamate and
carboethoxyalkyl phosphorodichloridate.41 However, this method
shown that vancomycin-resistant bacteria are able to produce
D-Ala-
D-lactate dipeptide instead of -Ala- -Ala dipeptide. -Ala- -lactate
D
D
D
D
dipeptide binds much more weakly to vancomycin, and can be in-
corporated into the peptidoglycan layer. This resulted in the
Abbreviations: BOP, (benzotriazol-1-yloxy)tris(dimethylamino)phosphonium
hexafluorophosphate; BroP, bromotris(dimethylamino)phosphonium hexa-
fluorophosphate; Cbz, benzyloxycarbonyl; DABCO, 1,4-diazabicyclo[2.2.0]octane;
´
DBU, 1,8-diazabicyclo[5.4.0]undec-7-ene; DCC, N,N-dicyclohexylcarbodiimide;
DIAD, diisopropyl azodicarboxylate; DIC, N,N0-diisopropylcarbodiimide; DIPEA, N,N-
diisopropylethylamine; Fmoc, 9-fluorenylmethyloxycarbonyl; HBTA, 1-hydroxy-
benzotriazole; PyBOP, (benzotriazol-1-yloxy)tripyrrolidinophosphonium hexa-
fluorophosphate; TEA, triethylamine; TFA, trifluoroacetic acid; TMSBr, trimethylsilyl
bromide; TPyClU, 1,1,3,3-bis(tetramethylene)chlorouronium tetrafluoroborate;
TrisCl, 2,4,6-triisopropylbenzenesulfonyl chloride.
* Corresponding author. Tel.: þ420 220183441; fax: þ420 220183571.
´ ˇ
E-mail address: jiracek@uochb.cas.cz (J. Jiracek).
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.05.051

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6091
is suitable only for aromatic aldehydes. Campagne et al.42 described
a methodology for the preparation of phosphonopeptides from N-
Fmoc-protected monoesters of phosphonic acids by solid-phase
synthesis using BOP as a condensation agent.
is a suitable building block for the solid-phase synthesis of cysteine-
containing phosphonopeptides.
2. Results and discussion
Herein, we present a detailed description of the synthesis of new
types of phosphonodepsipeptide, Nle
J[PO(OH)O]-Gly(Ala) and
First, we decided to synthesise target compounds 6, 10 and 20a
using the methodology outlined in Scheme 1. The first step of our
effort consisted of the preparation of phosphonate monomethyl
ester 3, which was further transformed in three synthetic steps: (i)
condensation with benzyl glycolate followed by (ii) selective
demethylation or (iii) use in solid-phase synthesis.
Thus, diphenyl ester 1 was prepared through the one-pot con-
densation of valeraldehyde, triphenyl phosphite and benzyl carba-
mate according to the procedure described by Oleksyszyn et al.,45
followed by a slightly modified procedure of transesterification46
with sodium methanolate to yield dimethyl ester 2. Alternatively,
we also prepared compound 2 using a one-pot reaction of valer-
aldehyde, dimethyl phosphite and benzyl carbamate in acetyl
chloride,47 which gave a moderate yield of 36%. The procedure of
Oleksyszyn et al.45 is somewhat longer, but more convenient with
Nle [PO(OH)O]-Gly(Ala)-Val, mimicking the peptide sequences
J
Nle-Gly(Ala) and Nle-Gly(Ala)-Val and, in all cases, having a phos-
phonate analogue of norleucine at the N-terminus. Recently, we
published43 a preliminary report of this synthesis. The presented
compounds may represent potential inhibitors of methionine or
leucine aminopeptidases. The proposed pseudopeptide sequences
should fulfil the structural requirements of methionine aminopep-
tidases; Nle or Met at P1 and a small aliphatic residue,0such as Ala or
0
Gly, in the P1 position. The C-terminal Val at the P2 position was
chosen based on our recent results with statine pseudopeptides as
inhibitors of methionine aminopeptidases.44 We also prepared an
Fmoc-Nle-
block for the efficient solid-phase synthesis of Tyr-Nle-
Gly-Cys-NH2 pseudopeptide, and demonstrated that this precursor
J[PO(OH)O]-CH2-COOH synthon. We used this building
J[PO(OH)O]-
O
O
P
OPh
a
O
P(OPh)3
+
OPh
+
BnO
NH2
CbzHN
1
b
O
P
OCH3
O
c
OCH3
O
HPO(OCH3)2
+
+
BnO
NH2
CbzHN
2
d
O
P
OCH3
O
P
OCH3
O
P
OH
f
e
OH
OH
OCH3
CbzHN
3
FmocHN
NH2
4
5
j
g
O
P
OCH3
8
O
P
OBn
NH2
O
h
O
O
O
OCH3
O
CbzHN
NH2
6
k,l
O
i
OH
O
P
P
O
NH2
O
OH
NH2
OCH3
O
CbzHN
20a
7
O
O
H
N
O
P
H
N
O
P
OCH3
n
m
O
OH
O
OBn
3
OCH3
O
O
NH2
CbzHN
10
9
Scheme 1. Reagents, conditions, yields: (a) AcOH, 80 ꢀC, 2 h (75%); (b) NaOCH3, methanol and dioxane, rt, overnight (77%); (c) AcCl, ꢁ5 to 0 ꢀC, 1 h then rt overnight (36%);
(d) NaOH, water and methanol, 80 ꢀC, 4 h (84%); (e) 10% Pd–C, H2, methanol, rt, 15 psi, overnight (87%); (f) Fmoc-OSu, NaHCO3, water and dioxane, rt, overnight; (g) TrisCl,
1-methylimidazole, glycolyl-resin (Sieber Amide); (h) glycolamide, BOP, TEA, DMF, rt, overnight (81%); (i) 10% Pd–C, H2, methanol, rt, 15 psi, overnight (59%); (j) benzyl
glycolate, BOP, TEA, DMF, rt, overnight (89%); (k) TMSBr, dichloromethane, rt, 3 h; (l) 10% Pd–C, H2, methanol, rt, overnight (3% over two steps); (m) HOCH2CONHCH(S-iPr)COOBn
17a, BOP, TEA, DMF, rt, overnight (67%); (n) 10% Pd–C, H2, methanol, rt, 15 psi, overnight (50%).

6092
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
regard to the yield (58% of 2 over two steps). The hydrolysis of 2 with
NaOH afforded the important monomethyl ester intermediate 3.
Coupling of 3 with benzyl glycolate afforded mixed diester 8,
which was treated with TMSBr. Subsequent hydrogenolysis gave
target compound 20a, but in an unsatisfactory yield (3% after RP-
HPLC purification). The main problem with this method was the
low selectivity of the demethylation agent (TMSBr) and difficulty in
purifying crude compound 20a by flash chromatography (due to
the presence of the polar P–OH group).
Glycolic and L-lactic acid-derived dipeptides, necessary for
coupling with phosphonic acid 16, were synthesised as published
earlier56 (Scheme 3). Due to great differences in the reactivity
of amino and alcohol groups, the hydroxyl functions of glycolic and
L
-lactic acids did not need to be protected. Glycolic or
L-lactic acid
reacted with -valine benzyl ester p-toluenesulfonate using DIC as
L
a coupling reagent to afford 17a or 17b, respectively. Compounds
18a and 18b were prepared in the same manner starting from
-valine methyl ester hydrochloride, but DBU was used instead of
L
Concerning the solid-phase synthesis (SPS) pathway, removal of
the Cbz group through catalytic hydrogenation48 afforded zwit-
terion 4, which was converted into protected phosphonate 5 by
treatment with Fmoc-OSu.49 We tried to use phosphonate synthon
5 in a condensation reaction with the hydroxyl group of glycolic
acid attached via its carboxylic group to Sieber Amide resin using
a mixture of TrisCl and 1-methylimidazole,50 but all attempts failed.
Later, we found that compound 6 was easily available by coupling 3
with glycolamide followed by removal of the Cbz group. Phos-
phonate 10, having the Gly-Val dipeptide sequence at the C-ter-
minus, was obtained in a similar manner.
Taking into account the synthetic problems we encountered
with methyl ester protection of the phosphoryl moiety, we decided
to focus our next effort on the development of a new method, one
that would overcome the disadvantages of the synthetic pro-
cedures described above.
TEA for liberating the product from the salt. Aminolysis of 18a and
18b with ammonia in methanol afforded amides 19a and 19b,
respectively.
R
O
R
O
a
b
H
N
.
OH
H2N
TsOH
HO
OBn
HO
OBn
+
O
O
R = H
R = CH3
17a, R = H
17b, R = CH3
R
O
R
O
H
N
.
OH
H2N
HCl
HO
O
OCH3
HO
OCH3
+
O
18a, R = H
18b, R = CH3
R = H
R = CH3
In this work, we modified previously published methodologies
for the preparation of phosphonodepsipetides reported by Cam-
pagne et al.42 and Isomura et al.6 Instead of a phosphonic methyl
ester moiety, which we used for compound 3, we introduced benzyl
ester protection, which is easily removable through one-step cat-
alytic hydrogenolysis together with the Cbz protecting group.
Scheme 2 describes the synthesis of key intermediate 16, which we
proposed as a precursor for the synthesis of the target compounds.
Diphenylester 11 was prepared as described by Kudzin and
Stec51 using a three-component condensation of valeraldehyde,
triphenyl phosphite and N-phenyl thiourea, which after heating in
concentrated hydrochloric acid afforded free 1-aminophosphonic
acid 12. Reaction of 12 with benzyloxycarbonyl chloride furnished
Cbz-protected acid 13, and after esterification52,53 proceeding un-
der mild conditions with 2 equiv of N,N-diisopropyl-O-benzyli-
sourea54 14, we isolated the corresponding dibenzyl ester 15. Key
intermediate 16 was obtained through the base-catalysed hydro-
lysis of 15 using DABCO.55 Surprisingly, the condensation of valer-
aldehyde, dibenzyl phosphite and benzyl carbamate performed
analogously to the standard method47 (Scheme 2, reaction f) fur-
nished dibenzyl ester 15 in only 5%.
c
R
O
H
N
HO
O
NH2
19a, R = H
19b, R = CH3
Scheme 3. Reagents, conditions, yields: (a) TEA, HBTA, DIC, dichloromethane, 0 ꢀC, 1 h
then rt overnight R¼H (80%), R¼CH3 (88%); (b) DBU, HBTA, DIC, dichloromethane, 0 ꢀC,
1 h then rt overnight R¼H (61%), R¼CH3 (69%); (c) NH3, CH3OH, rt, 6 days, R¼H (60%),
R¼CH3 (79%).
The key intermediate 16 and commercially available benzyl
glycolate, benzyl (S)-lactate, methyl glycolate, methyl (S)-lactate,
glycolamide or (S)-lactamide were coupled using BOP35 to afford
the corresponding mixed phosphonic acid diesters 20a, 20b, 22a,
22b, 24a and 24b, respectively (Scheme 4). Phosphonic acid di-
esters 26a, 26b, 28a, 28b, 30a and 30b were obtained by reaction of
O
P
S
a
O
P(OPh)3
OPh
H2N
NHPh
OPh
PhHN
NH
S
11
O
P
OH
O
P
O
P
OBn
c
b
OH
OH
d
OBn
OH
CbzHN
13
NH2
12
CbzHN
15
f
e
O
P
OBn
O
OH
O
HPO(OBn)2
BnO
NH2
CbzHN
16
Scheme 2. Reagents, conditions, yields: (a) AcOH, 80 ꢀC, 3 h (81%); (b) 35% HCl and AcOH, reflux for 20 h, then 1,2-epoxypropane, ethanol (58%); (c) benzyl chloroformate, Na2CO3,
water and dioxane, 0 ꢀC, 2 h then at rt overnight (88%); (d) N,N0-diisopropyl-O-benzylisourea 14, benzene and DMF, 80 ꢀC, 8 h (75%); (e) DABCO, toluene, 80 ꢀC, 8 h (92%); (f) AcCl,
ꢁ5 to 0 ꢀC, 1 h then rt overnight (5%).

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6093
R
O
R
O
P
OBn
b
OH
a
P
O
O
OBn
O
OH
O
CbzHN
NH2
20a, R = H
20b, R = CH3
21a, R = H
21b, R = CH3
R
O
R
O
OCH3
OCH3
P
O
c
P
O
d
OBn
O
OH
O
CbzHN
NH2
22a, R = H
22b, R = CH3
23a, R = H
23b, R = CH3
R
O
R
O
NH2
NH2
P
O
e
P
O
f
OBn
O
OH
O
CbzHN
NH2
24a, R = H
24b, R = CH3
25a, R = H
25b, R = CH3
16
R
H
O
R
O
O
H
N
O
P
N
O
OBn
P
O
OH
h
g
OBn
O
OH
O
CbzHN
NH2
27a, R = H
27b, R = CH3
26a, R = H
26b, R = CH3
R
H
O
R
O
O
H
N
O
P
N
j
i
O
OCH3
P
O
OCH3
OBn
O
OH
O
CbzHN
CbzHN
NH2
29a, R = H
29b, R = CH3
28a, R = H
28b, R = CH3
R
O
R
O
H
N
O
P
H
O
P
N
O
NH2
l
O
NH2
k
OBn
O
OH
O
NH2
31a, R = H
31b, R = CH3
30a, R = H
30b, R = CH3
Scheme 4. Reagents, conditions, yields: (a) benzyl glycolate or benzyl (S)-lactate, BOP, TEA, DMF, rt, overnight, R¼H (88%), R¼CH3 (86%); (b) 10% Pd–C, H2, methanol, rt, overnight,
R¼H (70%), R¼CH3 (69%); (c) methyl glycolate or methyl (S)-lactate, BOP, TEA, DMF, rt, overnight, R¼H (85%), R¼CH3 (82%); (d) 10% Pd–C, H2, methanol, rt, overnight, R¼H (78%),
R¼CH3 (73%); (e) glycolamide or (S)-lactamide, BOP, TEA, DMF, rt, overnight, R¼H (74%), R¼CH3 (70%); (f) 10% Pd–C, H2, methanol, rt, overnight, R¼H (77%), R¼CH3 (64%); (g)
HOCH2CONHCH(S-i-Pr)COOBn 17a or HOCH(S-CH3)CONHCH(S-i-Pr)COOBn 17b, BOP, TEA, DMF, rt, overnight, R¼H (70%), R¼CH3 (72%); (h) 10% Pd–C, H2, methanol, rt, overnight,
R¼H (65%), R¼CH3 (56%); (i) HOCH2CONHCH(S-i-Pr)COOCH3 18a or HOCH(S-CH3)CONHCH(S-i-Pr)COOCH3 18b, BOP, TEA, DMF, rt, overnight, R¼H (83%), R¼CH3 (78%); (j) 10% Pd–C,
H2, methanol, rt, overnight, R¼H (66%), R¼CH3 (77%); (k) HOCH2CONHCH(S-i-Pr)CONH2 19a or HOCH(S-CH3)CONHCH(S-i-Pr)CONH2 19b, BOP, TEA, DMF, rt, overnight, R¼H (76%),
R¼CH3 (82%); (l) 10% Pd–C, H2, methanol, rt, overnight, R¼H (62%), R¼CH3 (71%).
O
H
N
O
P
O
P
O
P
OBn
OH
c
OBn
a,b
O
O
NH2
SH
O
NH2
O
OH
O
OH
O
O
FmocHN
32
NH
CbzNH
33
20a
OH
Scheme 5. Reagents, conditions, yields: (a) 10% Pd–C, H2, methanol, rt, overnight; (b) Fmoc-OSu, NaHCO3, water–dioxane, rt, overnight (22% through two steps); (c) (i) Cys(Trt)-
resin Sieber Amide (0.9 equiv), BOP (2 equiv)–DIPEA (4 equiv) in DMF, rt overnight, 8 h; (ii) piperidine–DMF; (iii) Fmoc-Tyr(Ot-Bu) (3 equiv), BOP (3 equiv)–DIPEA (6 equiv), rt, 2 and
1 h; (iv) piperidine–DMF; (v) TFA (2%), ethanedithiol (2%), water (1%), triisopropylsilane (2%) in dichloromethane, rt for 20 min, evaporation of the filtrate to dryness, (vi) TFA (95%),
ethanedithiol (2%), water (1%) and triisopropylsilane (2%), evaporation of the filtrate to dryness (45%, yield is given for compound 33 purified by RP-HPLC as a mixture of
diastereoisomers).

6094
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
16 under the same conditions with dipeptides 17a, 17b, 18a, 18b,
19a and 19b, respectively. In all cases, the reactions proceeded
smoothly and in high preparative yields (72–86%). The final step of
the synthesis involved the removal of the benzyl and Cbz groups by
classical catalytic hydrogenation. The final target compounds 21a,
21b, 23a, 23b, 25a, 25b, 27a, 27b, 29a, 29b, 31a and 31b were pu-
rified using RP-HPLC in good yields.
To extend the versatility of our approach, we prepared N-pro-
tected phosphonate synthon 32 by hydrogenolysis of 20a followed
by reaction with Fmoc-OSu, as illustrated in Scheme 5. The pres-
ence of the lypophilic Fmoc group and free phosphonic acid in the
molecule resulted in a difficult purification and rather low pre-
parative yield. On the other hand, synthon 32 appeared to be a very
useful building block for the solid-phase synthesis of pseudo-
peptidic phosphonate inhibitors of various metallopeptidases. As
an example, using a protocol described by Raguin et al.,57 we suc-
cessfully prepared phosphonodepsipeptide 33, which mimics the
peptide sequence of Tyr-Nle-Gly-Cys-amide. Synthon 32 may be
useful in the synthesis of cysteine-containing pseudopeptides,
which are hardly accessible (due to the poisoning effect of sulfur)
min on a C18 Nucleosil column (250ꢂ4 mm, 5
mm) from Watrex
(Praha) using the same gradients and solvents. Eluted compounds
were detected at 218 nm. Melting points were determined on
13
a Boetius block and are uncorrected. 1H and NMR spectra were
measured on
a
Bruker AVANCE-600 spectrometer (1H at
600.13 MHz, 13C at 150.9 MHz) in CDCl3, DMSO-d6, CD3OD or D2O
solution at 300 K. The 2D-H,H-COSY, 2D-H,C-HSQC and 2D-H,C-
HMBC spectra were recorded and used for the structural assign-
ment of proton and carbon signals. IR spectra were recorded on
Bruker IFS 55 Equinox apparatus. HRMS spectra were obtained on
an FTMS mass spectrometer LTQ-orbitrap XL (Thermo Fisher, Bre-
men, Germany) in electrospray ionisation mode. Optical rotation
values were measured on polarimeter AUTOPOL IV (Rudolph Re-
search Analytical, USA) for the sodium D line at 20 ꢀC.
4.2. Synthesis of compounds
4.2.1. Diphenyl [(R,S)-1-(benzyloxycarbonylamino)-
pentyl]phosphonate (1)
Benzyl carbamate (6.04 g, 0.04 mol), triphenyl phosphite
(12.41 g, 0.04 mol) and valeraldehyde (5.17 g, 0.06 mol) were stirred
for 2 h at 80 ꢀC in glacial acetic acid (20 mL). The acetic acid
was evaporated in vacuo and the residue was dissolved in
15 mL of methanol. The solution was left to stand overnight at
5 ꢀC to afford colourless crystals, which were filtered off and
washed with petroleum ether. The pure product was obtained
by recrystallisation from a mixture chloroform–methanol. Yield
13.5 g (75%). Colourless solid, mp 97–98 ꢀC (lit.58 95–97 ꢀC).
Rf¼0.62 (S1). 1H NMR (600 MHz, DMSO): 0.86 (3H, t, J¼7.3,
CH3), 1.26 and 1.34 (2H, m, CH2), 1.34 and 1.45 (2H, m, CH2),
1.80 and 1.90 (2H, m, CH2), 4.30 (1H, m, N–CH–P), 5.09 and 5.13
(2H, d, Jgem¼12.6, –CH2–O), 7.15–7.39 (15H, m, 3ꢂC6H5), 8.08
(1H, d, J¼9.5, NH). 13C NMR (150.9 MHz, DMSO): 13.99 (CH3),
21.67 (CH2), 27.74 (d, J(C,P)¼14.4, CH2), 28.23 (d, J(C,P)¼3.3,
CH2), 48.48 (d, J(C,P)¼158.3, N–CH–P), 66.05 (O–CH2), 120.62 (d,
J(C,P)¼4.0, 2ꢂAr–CH), 120.83 (d, J(C,P)¼3.8, 2ꢂAr–CH), 125.38
(Ar–CH), 125.53 (Ar–CH), 127.93 (2ꢂAr–CH), 128.10 (Ar–CH),
128.57 (2ꢂAr–CH), 130.09 (2ꢂAr–CH), 130.13 (2ꢂAr–CH), 137.17
(Ar–C), 150.12 (d, J(C,P)¼9.6, Ar–C), 150.40 (d, J(C,P)¼9.7, Ar–C),
156.54 (d, J(C,P)¼5.0, O–CO–N). IR (KBr, nmax cmꢁ1) 3270, 1711,
1591, 1544, 1491, 1455, 1253, 1218, 1201, 1163, 1071, 1028, 950,
772, 758, 701, 692, 688. HRMS (ESI) calcd for C25H28NNa O5P
[MþNa]þ 476.1603; found: 476.1597.
by solution-synthesis approaches requiring
hydrogenolysis.
a final catalytic
3. Conclusion
In conclusion, we present herein an efficient method for the
solution synthesis of phosphonodepsipeptides. In comparison to
previously published methods, our approach offers several im-
provements: (i) masking of the amino and hydroxyl moieties by
non-polar protecting groups permits convenient and simple work-
up of intermediates, (ii) the condensation of protected phosphonic
acid 16 with the hydroxyl moiety of glycolic or L-lactic acid de-
rivatives proceeds easily and without sensitivity to steric hindrance
and (iii) the final step of the synthesis, catalytic hydrogenation,
enables the complete removal of the protecting groups under mild
conditions. In addition, N-Fmoc-protected precursor 32 provides
a convenient building block for the solid-phase synthesis of pseudo-
peptide phosphonates without any restriction of sulfur-containing
amino acids.
4. Experimental
4.1. General
Reagents and solvents (Sigma–Aldrich–Fluka) used in this study
were of analytical grade. TLC on silica gel coated aluminium plates
(Fluka) was performed in the following systems (v/v): chloroform–
ethanol 98:2 (S1), chloroform–ethanol 95:5 (S2), isopropyl alcohol–
acetic acid–water 12:3:5 (S3), chloroform–ethanol 90:10 (S4),
isopropyl alcohol–concentrated aqueous ammonia–water 7:1:2
(S5). The compounds were visualised by exposure to UV light at
254 nm, by ninhydrin spraying (dark blue colour of amines) and by
4.2.2. Dimethyl [(R,S)-1-(benzyloxycarbonylamino)-
pentyl]phosphonate (2)
Sodium methoxide (5.6 g, 0.011 mol) was added in one portion
to a stirred solution of compound 1 (9.5 g, 0.021 mol) dissolved in
100 mL of anhydrous methanol and 100 mL of dioxane. The flask
was equipped with a calcium dichloride tube and the reaction was
allowed to proceed at rt overnight. The solvents were evaporated in
vacuo, the residue was taken up in 100 mL of ethyl acetate and
washed with 100 mL of water and twice with 100 mL of a 1% so-
lution of NaOH. The organic layer was separated, dried over Na2SO4,
filtered and concentrated in vacuo. The crude product was purified
by flash chromatography on silica gel using elution with a linear
gradient of ethyl acetate in toluene. Yield 5.3 g (77%).
Alternatively, we also prepared compound 2 as follows. A mix-
ture of benzyl carbamate (6.05 g; 0.04 mol) and dimethyl phosphite
(4.4 g; 0.04 mol) in 50 mL of acetyl chloride was cooled to ꢁ5 ꢀC,
and valeraldehyde (4.31 g, 0.05 mol) was added dropwise over
10 min. The reaction mixture was stirred for 1 h for at 0 ꢀC and then
left to react at rt overnight. The acetyl chloride was evaporated in
vacuo and the crude product was purified by flash chromatography
on silica gel using elution with a linear gradient of ethyl acetate in
toluene. Yield 4.8 g (36%). Colourless oil. Rf¼0.67 (S2). 1H NMR
spraying with
a 1% (v/v) ethanolic solution of 4-(4-nitro-
benzyl)pyridine followed by heating and treating with gaseous
ammonia (blue colour of diesters of phosphonic acids or esters of
carboxylic acids). Dipeptides 19a and 19b were visualised by
spraying with a solution of 10% CuSO4 5H2O in 10% phosphoric acid
followed by heating at 200 ꢀC. Flash chromatography purifications
were carried out on silica gel (40–63
HPLC chromatography was carried out on a C18 Luna column
m) at a flow rate 9 mL/min. Sol-
mm, Fluka). Preparative RP-
(Phenomenex, 250ꢂ21.2 mm, 10
m
vent A: 0.1% TFA. Solvent B: 80% CH3CN, 0.1% TFA. The following
gradients were used; G1: t¼0–15 min (0% B), t¼30 min (20% B),
t¼40 min (40% B), t¼45 min (100% B); G2: t¼0–15 min (0% B),
t¼30 min (20% B), t¼40 min (80% B), t¼45 min (100% B). Analytical
RP-HPLC chromatography was carried out at a flow rate of 1 mL/

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6095
(600 MHz, CDCl3): 0.89 (3H, t, J¼7.2, CH3), 1.32 (2H, m, CH2), 1.34
and 1.45 (2H, m, CH2), 1.57 and 1.84 (2H, m, CH2), 3.71 (3H, d,
J(H,P)¼10.6, OCH3), 3.75 (3H, d, J(H,P)¼10.6, OCH3), 4.10 (1H, m, N–
CH–P), 5.11 and 5.15 (2H, d, Jgem¼12.3, –CH2–O), 5.15 (1H, d, J¼9.5,
NH), 7.31–7.37 (5H, m, C6H5). 13C NMR (150.9 MHz, CDCl3): 13.79
(CH3), 22.12 (CH2), 27.78 (d, J(C,P)¼12.5, CH2), 29.34 (d, J(C,P)¼2.7,
CH2), 47.08 (d, J(C,P)¼156.0, N–CH–P), 52.93 (d, J(C,P)¼6.6, OCH3),
53.10 (d, J(C,P)¼7.2, OCH3), 67.07 (O–CH2), 127.94 (2ꢂAr–CH),
128.12 (Ar–CH), 128.44 (2ꢂAr–CH), 136.20 (Ar–C), 156.03 (d,
J(C,P)¼5.6, O–CO–N). IR (CHCl3, nmax cmꢁ1): 3433, 3020, 2958, 1721,
1511, 1246, 1059, 1042, 836, 698. HRMS (ESI) calcd for C15H25NO5P
[MþH]þ 330.1470; found: 330.1467.
Colourless solid, mp 176–178 ꢀC. Rf¼0.59 (S5). 1H NMR (600 MHz,
DMSO): 0.84 (3H, t, J¼7.0, CH3), 1.22 and 1.29 (2H, m, CH2), 1.22 and
1.34 (2H, m, CH2), 1.56 and 1.68 (2H, m, CH2), 3.54 (3H, d,
J(H,P)¼10.5, OCH3), 3.73 (1H, m, N–CH–P), 4.26 (1H, dd, J¼10.4 and
6.8, –CHaHb-O), 4.28 (1H, dd, J¼10.4 and 7.8, –CHaHb–O), 4.31 (1H,
m, CH (Fmoc)), 7.31 (2H, m, Ar–H), 7.41 (2H, m, Ar–H), 7.56 (1H, d,
J¼9.6, NH), 7.74 (2H, m, Ar–H), 7.88 (2H, m, Ar–H). 13C NMR
(150.9 MHz, DMSO): 14.09 (CH3), 21.84 (CH2), 27.99 (d, J(C,P)¼13.1,
CH2), 28.57 (d, J(C,P)¼3.0, CH2), 46.91 (pCH– (Fmoc)), 47.64 (d,
J(C,P)¼154.7, N–CH–P), 52.20 (d, J(C,P)¼6.2, OCH3), 65.88 (O–CH2),
120.34 (2ꢂAr–CH),125.61 (Ar–CH),125.63 (Ar–CH),127.25 (Ar–CH),
127.28 (Ar–CH), 127.88 (2ꢂAr–CH), 140.94 (2ꢂAr–C), 143.95 (Ar–C),
144.13 (Ar–C), 156.37 (d, J(C,P)¼5.1, O–CO–N). IR (KBr, nmax cmꢁ1):
3285, 2957,1688,1543,1451,1294,1258,1216,1044, 756, 739. HRMS
(ESI) calcd for C21H27NO5P [MþH]þ 404.1627; found: 404.1612.
4.2.3. Methyl hydrogen [(R,S)-1-(benzyloxycarbonylamino)-
pentyl]phosphonate (3)
Phosphonate 2 (3.9 g; 0.012 mol) was dissolved in 25 mL of
methanol and 25 mL of 1 M NaOH. The reaction mixture was heated
for 4 h at 80 ꢀC. The methanol was evaporated in vacuo and the
aqueous layer was extracted with 25 mL of ethyl acetate. The sep-
arated aqueous layer was acidified (pH¼1) using 1 M HCl. The
resulting clear oil was extracted twice with 25 mL of ethyl acetate
and the organic phase was dried over Na2SO4 prior to filtering and
evaporating under reduced pressure. The pure product was
obtained by trituration of the residue at ꢁ20 ꢀC from a mixture of
ethyl acetate and petroleum ether. Yield 3.14 g (84%). Colourless
solid, mp 131–133 ꢀC. Rf¼0.71 (S5). 1H NMR (600 MHz, CDCl3): 0.87
(3H, t, J¼7.2, CH3), 1.27 and 1.34 (2H, m, CH2), 1.32 and 1.42 (2H, m,
CH2), 1.52 and 1.87 (2H, m, CH2), 3.70 (3H, d, J(H,P)¼10.8, OCH3),
4.09 (1H, m, N–CH–P), 5.11 and 5.13 (2H, d, Jgem¼12.3, –CH2–O), 5.22
(1H, d, J¼10.2, NH), 7.29–7.35 (5H, m, C6H5). 13C NMR (150.9 MHz,
CDCl3): 13.84 (CH3), 22.17 (CH2), 27.87 (d, J(C,P)¼12.8, CH2), 29.23
(d, J(C,P)¼2.7, CH2), 47.32 (d, J(C,P)¼158.0, N–CH–P), 52.59 (d,
J(C,P)¼6.9, OCH3), 67.14 (O–CH2), 127.95 (2ꢂAr–CH), 128.13 (Ar–
CH), 128.48 (2ꢂAr–CH), 136.18 (Ar–C), 156.19 (d, J(C,P)¼5.7, O–CO–
N). IR (KBr, nmax cmꢁ1): 3280, 2952, 1692, 1547, 1286, 1257, 1221,
1047, 1034, 978, 835, 754, 695. HRMS (ESI) calcd for C14H23NO5P
[MþH]þ 316.1314; found: 316.1323.
4.2.6. General procedure for the preparation of compounds
7, 8 and 9
Monoester 3 (0.5 g, 1.6 mmol), BOP (1.04 g, 2.4 mmol) and the
hydroxy component (2.4 mmol) were dissolved in 10 mL of dried
DMF and TEA (0.64 g, 6.3 mmol) was added in one portion. The
reaction mixture was allowed to react at rt overnight, and the
solvent was evaporated to provide a black oil that was purified by
column chromatography.
4.2.6.1. {[(R,S)-1-(Benzyloxycarbonylamino)-pentyl(methoxy)phosphoryl]-
oxy}acetamide (7). Compound 7 was prepared by the reaction of 3 with
glycolamide. Flash chromatography: linear gradient of ethanol in chlo-
roform. Yield 0.49 g (81%). Colourless oil, Rf¼0.47 (S4). 1H NMR
(600 MHz, CDCl3): mixture of diastereoisomers (w3:2), only signals of
the major isomer are given: 0.90 (3H, t, J¼7.2, CH3), 1.30 and 1.36 (2H, m,
CH2), 1.34 and 1.47 (2H, m, CH2), 1.58 and 1.92 (2H, m, CH2), 3.75 (3H, d,
J(H,P)¼10.6, OCH3), 4.11 (1H, m, N–CH–P), 4.36 (1H, dd, J¼15.3 and 3.8,
O–CHaHb–CO), 4.65 (1H, dd, J¼15.3 and 8.0, O–CHaHb–CO), 5.10 and
5.13 (2H, d, Jgem¼12.2, –CH2–O), 5.94 (1H, d, J¼9.8, NH), 6.07 and 6.98
(2H, br, CONH2), 7.31–7.37 (5H, m, C6H5). 13C NMR (150.9 MHz, CDCl3):
13.76 (CH3), 22.02 (CH2), 27.86 (d, J(C,P)¼13.3, CH2), 28.70 (d, J(C,P)¼2.9,
CH2), 48.22 (d, J(C,P)¼156.6, N–CH–P), 53.33 (d, J(C,P)¼7.4, OCH3), 63.78
(d, J(C,P)¼7.4, O–CH2–CO), 67.37 (O–CH2–C), 128.02 (2ꢂAr–CH), 128.33
(Ar–CH), 128.53 (2ꢂAr–CH), 136.09 (Ar–C), 156.56 (d, J(C,P)¼4.1, O–CO–
N), 170.14 (d, J(C,P)¼7.5, N–CO–C). IR (KBr, nmax cmꢁ1) 3470, 3410, 3279,
3220, 1716, 1696, 1685, 1662, 1545, 1498, 1456, 1377, 1246, 1071, 1033,
982, 753, 701. HRMS (ESI) calcd for C16H25N2NaO6P [MþNa]þ 395.1348;
found: 395.1343.
4.2.4. Methyl hydrogen [(R,S)-1-aminopentyl] phosphonate (4)
Monoester 3 (2 g, 0.006 mol) was dissolved in 80 mL of meth-
anol, and then 10% Pd–C (0.15 g) was added. The reaction mixture
was vigorously stirred and left to react under an atmosphere of
hydrogen (15 psi) at rt overnight. The catalyst was filtered off
through Celite and the filter was washed with 100 mL of methanol.
The solvent was evaporated in vacuo, and pure product was
obtained by crystallisation from a mixture of methanol and petro-
leum ether. Yield 1 g (87%). Colourless solid, mp 240–242 ꢀC.
Rf¼0.75 (S3). 1H NMR (600 MHz, CD3OD): 0.96 (3H, t, J¼7.3, CH3),
1.40 (2H, m, CH2), 1.47 and 1.52 (2H, m, CH2), 1.71 and 1.94 (2H, m,
CH2), 3.09 (1H, m, N–CH–P), 3.64 (3H, d, J(H,P)¼10.3, OCH3). 13C
NMR (150.9 MHz, CD3OD): 14.15 (CH3), 23.55 (CH2), 29.50 (d,
J(C,P)¼8.5, CH2), 29.91 (d, J(C,P)¼2.0, CH2), 49.41 (d, J(C,P)¼145.6,
N–CH–P), 52.08 (d, J(C,P)¼6.2, OCH3). IR (KBr, nmax cmꢁ1): 3008 br,
2959, 1645, 1539, 1469, 1205, 1087, 1039, 782. HRMS (ESI) calcd for
C6H17NO3P [MþH]þ 182.0946; found: 182.0941.
4.2.6.2. Benzyl {[(R,S)-1-(benzyloxycarbonylamino)-pentyl(methoxy)-
phosphoryl]oxy}acetate (8). Compound 8 was prepared by the re-
action of 3 with benzyl glycolate. Flash chromatography: linear
gradient of ethyl acetate in toluene. Yield 0.64 g (89%). Colourless
oil, Rf¼0.67 (S2). 1H NMR (600 MHz, CDCl3): mixture of di-
astereoisomers (59:41), only signals of the major isomer are given:
0.88 (3H, t, J¼7.2, CH3),1.28 and 1.34 (2H, m, CH2),1.34 and 1.43 (2H,
m, CH2), 1.57 and 1.87 (2H, m, CH2), 3.76 (3H, d, J(H,P)¼10.9, OCH3),
4.18 (1H, m, N–CH–P), 4.63 (1H, dd, J¼16.2 and 11.0, O–CHaHb–CO),
4.67 (1H, dd, J¼16.2 and 11.5, O–CHaHb–CO), 5.09 and 5.13 (2H, d,
Jgem¼12.2, –CH2–O), 5.19 and 5.21 (2H, d, Jgem¼12.0, –CH2–O), 5.25
(1H, d, J¼9.5, NH), 7.17–7.37 (10H, m, 2ꢂC6H5). 13C NMR
(150.9 MHz, CDCl3): 13.83 (CH3), 22.16 (CH2), 27.89 (d, J(C,P)¼12.5,
CH2), 29.53 (d, J(C,P)¼2.3, CH2), 47.90 (d, J(C,P)¼156.8, N–CH–P),
52.78 (d, J(C,P)¼7.5, OCH3), 62.37 (d, J(C,P)¼6.0, O–CH2–CO), 67.09
(O–CH2–C), 67.36 (O–CH2–C), 127.97 (Ar–CH), 128.10 (Ar–CH),
128.45 (2ꢂAr–CH), 128.46 (2ꢂAr–CH), 128.51 (2ꢂAr–CH), 128.64
(2ꢂAr–CH), 134.75 (Ar–C), 136.27 (Ar–C), 156.14 (d, J(C,P)¼5.7, O–
CO–N), 168.55 (d, J(C,P)¼4.2, O–CO–C). IR (CHCl3, nmax cmꢁ1) 3431,
3068, 3031,1758,1721,1511,1456,1240,1099,1046, 698. HRMS (ESI)
calcd for C23H30NNaO7P [MþNa]þ 486.1658; found: 486.1652.
4.2.5. Methyl hydrogen [(R,S)-1-(9H-fluoren-9-
ylmethoxycarbonylamino)-pentyl] phosphonate (5)
Fmoc-OSu (1.63 g, 0.0048 mol) in 10 mL of dioxane was added
dropwise to a solution of monoester 4 (0.8 g, 4.4 mmol) in 15 mL of
saturated NaHCO3. The reaction mixture was left to react at rt
overnight, and then the dioxane was evaporated in vacuo and the
solution was acidified (pH¼1) using 1 M HCl. The resulting pre-
cipitate was decanted and stirred in 20 mL of ethyl acetate. The
crystals were filtered off and the pure product was recrystallised
from a mixture of methanol and ethyl acetate. Yield 0.95 g (51%).

6096
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
4.2.6.3. Benzyl (2S)-2-[({[((R,S)-1-benzyloxycarbonylamino)-pentyl-
(methoxy)phosphoryl]oxy}acetyl)amino]-3-methylbutanoate (9).
Compound 9 was prepared by the reaction of 3 with dipeptide 16a.
Flash chromatography: linear gradient of ethyl acetate in toluene.
Yield 0.70 g (79%). Pale yellow oil, Rf¼0.33 (S2). Mixture of four
diastereomers; assignment of individual isomers was not possible;
only groups and/or regions of signals are given: 1H NMR (600 MHz,
CDCl3): 0.88–0.90 (3H, t, J¼7.3, CH3), 0.91–0.94 (6H, d, J¼7.0, 2ꢂCH3
(Val)), w1.29 and w1.36 (2H, m, CH2), w1.36 and w1.46 (2H, m,
CH2), w1.60 and w1.87 (2H, m, CH2), 2.20–2.25 (1H, m, CH (Val)),
3.75, 3.755, 3.79 and 3.80 (3H, d, J(H,P)¼10.7, OCH3), 4.14–4.19 (1H,
m, N–CH–P), 4.47–4.65 (2H, m, O–CH2–CO), 4.59–4.61 (1H, dd, N–
CH–CO), 5.09–5.19 (4H, m, 2ꢂO–CH2), 5.19, 5.23, 5.34 and 5.38 (1H,
d, J¼10.0, O–CO–NH), 7.11, 7.13 and 7.22 (1H, d, J¼8.5, CO–NH), 7.15–
7.36 (10H, m, 2ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.76 (CH3),
17.52–17.88 and 18.83–18.94 (2ꢂCH3 (Val)), 22.03–22.06 (CH2),
27.33–27.85 (CH2), 28.72–29.13 (CH2), 30.96–31.22 (CH (Val)),
47.02–48.46 (N–CH–P), 53.34–53.66 (OCH3), 56.78–56.98 (N–CH–
CO), 63.90–64.19 (O–CH2–CO), 66.95–67.36 (2ꢂO–CH2–C6H5),
127.86–137.79 (2ꢂC6H5), 156.02–156.19 (O–CO–NH), 167.10–167.25
(CO–NH), 171.22–171.34 (CO–O). IR (CHCl3, nmax cmꢁ1) 3431, 1722,
1685, 1511, 1456, 1393, 1382, 1235, 1069, 1043, 978, 698. HRMS (ESI)
calcd for C28H39N2NaO8P [MþNa]þ 585.2342; found: 585.2338.
14.06 (CH3), 22.74 (CH2), 29.52 (d, J(C,P)¼12.5, CH2), 31.89 (CH2),
50.61 (d, J(C,P)¼138.7, N–CH–P). IR (KBr, nmax cmꢁ1)z3000 br, 2295
br, 1650, 1601, 1530, 1469, 1178, 1032, 924, 564, 511, 493. HRMS (ESI)
calcd for C5H15NO3P [MþH]þ 168.0790; found: 168.0782.
4.2.9. [(R,S)-1-(Benzyloxycarbonylamino)-pentyl]phosphonic
acid (13)
Phosphonic acid 12 was dissolved in a 20% aqueous solution of
sodium carbonate (150 mL) and a solution of benzyloxycarbonyl
chloride (11.46 g, 0.067 mol) in 50 mL of dioxane was added
dropwise at 0 ꢀC over 1 h. After stirring for 2 h at 0 ꢀC, the mixture
was allowed to stand overnight at rt. The dioxane was evaporated
under reduced pressure and the acidity of the aqueous solution was
adjusted to pH¼1 while efficiently ice-cooling. The solution was
extracted three times with 200 mL of ethyl acetate, and the com-
bined organic layers were dried over Na2SO4 before filtering and
removing the solvent in vacuo. Trituration of the residue from
a mixture of ethyl acetate and petroleum ether at ꢁ20 ꢀC afforded
the pure product. Yield 14.9 g (88%). Colourless solid, mp 100–
102 ꢀC. Rf¼0.45 (S5). 1H NMR (600 MHz, D2OþNaOD): 0.85 (3H, t,
J¼7.0, CH3), 1.21–1.37 (4H, m, 2ꢂCH2), 1.52 and 1.82 (2H, m, CH2),
3.69 (1H, ddd, J¼15.8, 11.5 and 3.0, N–CH–P), 5.04 and 5.19 (2H, d,
Jgem¼12.6, –CH2–O), 7.37–7.43 (5H, m, C6H5). 13C NMR (150.9 MHz,
D2OþNaOD): 14.03 (CH3), 22.20 (CH2), 28.55 (d, J(C,P)¼12.8, CH2),
29.90 (d, J(C,P)¼2.1, CH2), 50.29 (d, J(C,P)¼149.9, N–CH–P), 67.51
(O–CH2), 128.16 (2ꢂAr–CH), 128.90 (Ar–CH), 129.38 (2ꢂAr–CH),
4.2.7. Diphenyl [(R,S)-1-(3-phenylthioureido)-pentyl]-
phosphonate (11)
The compound was prepared according to Kudzin and Stec.51 A
solution of triphenyl phosphite (37.2 g, 0.12 mol), valeraldehyde
(12.9 g, 0.15 mol) and N-phenyl thiourea (18.2 g, 0.12 mol) was
heated for 4 h at 80 ꢀC in 100 mL of glacial acetic acid. After cooling
to rt, water was added (20 mL) and the solution was allowed to
stand overnight at rt. The precipitate was filtered off, washed with
a mixture of acetic acid–water (40 mL, 1:1) and dried over P2O5.
Recrystallisation from a mixture of chloroform and methanol yiel-
ded pure product. Yield 44 g (81%). Colourless solid, mp 174–175 ꢀC
(lit.51 161–162 ꢀC). Calcd for C24H27N2O3PS (454.52): C 63.24%, H
5.99%, N 6.16%. Found: 63.37%, 6.03%, 6.08%. Rf¼0.48 (S1). 1H NMR
(600 MHz, DMSO): 0.87 (3H, t, J¼7.2, CH3), 1.31 and 1.47 (2H, m,
CH2), 1.40 (2H, m, CH2), 1.84 and 1.98 (2H, m, CH2), 5.63 (1H, m, N–
CH–P), 7.13, 7.33 and 7.49 (1H, 2H and 2H, m, N–C6H5), 7.19, 7.23 and
7.39 (2H, 1H and 2H, m, O–C6H5), 7.24 and 7.41 (3H and 2H, m, O–
C6H5), 8.25 (1H, d, J¼9.8, NH), 9.72 (1H, s, NH). 13C NMR (150.9 MHz,
DMSO): 14.02 (CH3), 22.08 (CH2), 27.48 (d, J(C,P)¼12.8, CH2), 29.31
(d, J(C,P)¼3.2, CH2), 50.58 (d, J(C,P)¼155.1, N–CH–P), 120.70 (d,
J(C,P)¼3.8, 2ꢂAr–CH), 120.87 (d, J(C,P)¼3.9, 2ꢂAr–CH), 123.20
(2ꢂAr–CH), 124.65 (Ar–CH), 125.58 (Ar–CH), 125.68 (Ar–CH),
128.79 (2ꢂAr–CH), 130.11 (2ꢂAr–CH), 130.21 (2ꢂAr–CH), 139.48
(Ar–C), 150.00 (d, J(C,P)¼9.6, Ar–C), 150.26 (d, J(C,P)¼9.7, Ar–C),
181.79 (d, J(C,P)¼8.1, N–CS–N). IR (KBr, nmax cmꢁ1) 3291, 3070, 3057,
3041, 1597, 1589, 1488, 1337, 1235, 1209, 1159, 955, 945, 767, 709,
691, 487. HRMS (ESI) calcd for C24H27N2NaO3PS [MþNa]þ 477.1378;
found: 477.1364.
137.25 (Ar–C), 158.79 (d, J(C,P)¼6.0, O–CO–N). IR (KBr, nmax cmꢁ1
)
3343, 3282, 2900 br, 1694, 1531, 1456, 1378, 1277, 1040, 736, 699.
HRMS (ESI) calcd for C13H19NO5P [MꢁH]þ 300.1001; found:
300.1007.
4.2.10. N,N0-Diisopropyl-O-benzylisourea (14)
The compound was prepared according to Mathias et al.54
Benzyl alcohol (17.95 g, 0.17 mol) was added dropwise to a stirred
mixture of N,N0-diisopropylcarboimide (20.9 g, 0.17 mol) and CuCl
(0.5 g) at 0 ꢀC over a period of 30 min. The cooling was removed and
then stirring was continued overnight at rt. Thereafter, the reaction
mixture was taken up in 100 mL of hexane and the cuprous salts
were filtered off over a pad of neutral alumina. The solvent was
evaporated under reduced pressure and the rest of the hexane was
removed under high vacuum. Yield 37 g (95%). Colourless oil,
Rf¼0.30 (S4). 1H NMR (600 MHz, CDCl3): 1.12 (12H, d, J¼6.5,
4ꢂCH3), 3.20 and 3.47 (2H, m, 2ꢂpCH–N), 5.10 (2H, s, –CH2–O), 7.28
(1H, m, Ar–H), 7.34 (2H, m, Ar–H), 7.38 (2H, m, Ar–H). 13C NMR
(150.9 MHz, CDCl3): 24.00 (2ꢂCH3), 24.25 (2ꢂCH3), 43.42 (pCH–N),
46.24 (pCH–N), 66.66 (O–CH2), 127.32 (Ar–CH), 127.56 (2ꢂAr–CH),
128.19 (2ꢂAr–CH), 137.89 (Ar–C), 151.50 (N–C(O)]N). IR (CCl4, nmax
cmꢁ1) 3446, 3091, 3067, 3034, 2967, 1669, 1467, 1388, 1366, 1320,
1306, 1124, 1029, 732, 696. HRMS (ESI) calcd for C14H23N2 [MþH]þ
235.1810; found: 235.1805.
4.2.11. Dibenzyl [(R,S)-1-(benzyloxycarbonylamino)-
pentyl]phosphonate (15)
4.2.8. [(R,S)-1-Aminopentyl]phosphonic acid (12)
Phosphonic acid 13 (14.9 g, 0.049 mol) and isourea 14 (23.19 g,
0.099 mol) were heated at 80 ꢀC in a mixture of benzene (200 mL)
and DMF (60 mL) over a period of 12 h. The solvents were evapo-
rated in vacuo to afford the crude product, which was purified by
flash chromatography on silica gel using elution with a linear gra-
dient of ethyl acetate in toluene. Trituration at ꢁ20 ꢀC from a mix-
ture of ethyl acetate and petroleum ether furnished pure product.
Yield 17.9 g (75%). Colourless solid, mp 65–67 ꢀC. Rf¼0.64 (S2). 1H
NMR (600 MHz, CDCl3): 0.85 (3H, t, J¼7.2, CH3), 1.24 and 1.30 (2H,
m, CH2), 1.32 and 1.42 (2H, m, CH2), 1.53 and 1.84 (2H, m, CH2), 5.63
(1H, dtd, J¼16.4, 10.6, 10.6 and 3.8, N–CH–P), 4.95 (1H, d, J¼10.6,
NH), 4.97 (2H, m, O–CH2), 4.98 (1H, dd, J¼11.7 and 7.4, O–CHaHb),
5.03 (1H, dd, J¼11.7 and 8.5, O–CHaHb), 5.06 (2H, s, O–CH2), 7.29–
The compound was prepared according to Kudzin and Stec.51
Ester 11 (44 g, 0.10 mol) was heated for 20 h at reflux with 200 mL
of 35% HCl and 30 mL of glacial acetic acid. The solvents were
evaporated under reduced pressure and the oily residue was taken
up in 200 mL of ethanol. The ethanolic solution of the crude
product was treated with methyloxirane until pHz6 was reached.
The precipitate of the product was filtered off, and then washed
with ethanol and diethyl ether. Yield 9.4 g (58%). Colourless solid,
mp 260–264 ꢀC (lit.51 262–264 ꢀC). Rf¼0.68 (S3). 1H NMR (600 MHz,
D2OþNaOD): 0.89 (3H, t, J¼7.0, CH3), 1.28–1.36 (2H, m, CH2), 1.30
and 1.49 (2H, m, CH2), 1.32 and 1.74 (2H, m, CH2), 2.49 (1H, td,
J¼10.4, 10.4 and 3.3, N–CH–P). 13C NMR (150.9 MHz, D2OþNaOD):

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6097
7.33 (15H, m, 3ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.80 (CH3),
22.12 (CH2), 27.80 (d, J(C,P)¼12.5, CH2), 29.45 (d, J(C,P)¼2.9, CH2),
47.78 (d, J(C,P)¼155.4, N–CH–P), 67.05 (O–CH2), 67.75 (d, J(C,P)¼6.5,
O–CH2), 67.86 (d, J(C,P)¼7.1, O–CH2), 127.96 (6ꢂAr–CH), 128.13
(2ꢂAr–CH), 128.40 (Ar–CH), 128.42 (Ar–CH), 128.46 (2ꢂAr–CH),
128.52 (Ar–CH), 128.54 (2ꢂAr–CH), 135.98 (d, J(C,P)¼6.0, Ar–C),
136.04 (d, J(C,P)¼5.7, Ar–C), 136.18 (Ar–C), 155.97 (O–CO–N). IR
(KBr, nmax cmꢁ1): 3208, 3063, 3048, 1722, 1550, 1497, 1455, 1290,
1260, 1222, 1215, 1046, 1018, 748, 732, 701, 696, 548, 536, 461, 456.
HRMS (ESI) calcd for C27H32NNaO5P [MþNa]þ 504.1916; found:
504.1908.
m, J¼6.8(6ꢂ) and 4.9, pCH–), 4.13 (2H, s, O–CH2–CO), 4.62 (1H, dd,
J¼9.2 and 4.9, N–CH–CO), 5.16 and 5.20 (2H, d, Jgem¼12.2, O–CH2),
7.13 (1H, br d, J¼9.2, NH), 7.32–7.38 (5H, m, C6H5). 13C NMR
(150.9 MHz, CDCl3): 17.53 (CH3), 18.93 (CH3), 31.18 (pCH–), 56.60
(N–CHo), 62.06 (CH2OH), 67.11 (O–CH2), 128.32 (2ꢂAr–CH), 128.46
(Ar–CH), 128.57 (2ꢂAr–CH), 135.13 (Ar–C), 171.73 (CO–O), 172.08
(CO–N). IR (CHCl3, nmax cmꢁ1) 3410, 2970, 1736, 1674, 1526, 1500,
1456, 1393, 1384, 1196, 1071, 1003, 698. HRMS (ESI) calcd for
20
C14H19NNaO4 [MþNa]þ 288.1212; found: 288.1207. [
a]
ꢁ3.0 (c
D
0.710, CHCl3).
Alternatively, compound 15 was prepared as follows. A mixture
of benzyl carbamate (3.02 g, 0.02 mol) and dibenzyl phosphite
(5.25 g, 0.02 mol) in 50 mL of acetyl chloride was cooled to ꢁ5 ꢀC
and valeraldehyde (2.15 g, 0.025 mol) was added dropwise over
10 min. The reaction mixture was stirred for 1 h at 0 ꢀC and left to
react at rt overnight. The acetyl chloride was evaporated in vacuo
and the crude product was purified by flash chromatography on
silica gel using elution with a linear gradient of ethyl acetate in
toluene. Yield 0.45 g (5%).
4.2.14. Benzyl (2S)-2-{[(2S)-2-hydroxypropanoyl]amino}-3-
methylbutanoate (17b)
Dipeptide 17b was prepared in the same manner as 17a by the
reaction of -lactic acid (0.28 g, 3.1 mmol), -valine benzyl ester p-
L
L
toluensulfonate (1.17 g, 3.1 mmol), 1-hydroxybenzotriazole (0.42 g,
3.1 mmol), triethylamine (0.31 g, 3.1 mmol) and DIC (0.78 g,
6.2 mmol). Yield 0.76 g (88%). Semisolid, Rf¼0.40 (S2). 1H NMR
(600 MHz, CDCl3): 0.87 (3H, d, J¼6.9, CH3), 0.92 (3H, d, J¼6.9, CH3),
1.43 (3H, d, J¼6.8, CH3), 2.22 (1H, m, J¼6.9(6ꢂ) and 4.9, pCH–), 3.37
(1H, d, J¼4.8, OH), 4.27 (1H, qd, J¼6.8(3ꢂ) and 4.8, pCH–), 4.59 (1H,
dd, J¼9.1 and 4.9, N–CH–CO), 5.14 and 5.20 (2H, d, Jgem¼12.3, O–
CH2), 7.09 (1H, br d, J¼9.1, NH), 7.32–7.38 (5H, m, C6H5). 13C NMR
(150.9 MHz, CDCl3): 17.48 (CH3), 18.95 (CH3), 21.27 (CH3), 31.22
(pCH–), 56.54 (N–CHo), 67.08 (O–CH2), 68.48 (pCH–O), 128.32
(2ꢂAr–CH), 128.45 (Ar–CH), 128.58 (2ꢂAr–CH), 135.18 (Ar–C),
171.87 (CO–O), 174.70 (CO–N). IR (CHCl3, nmax cmꢁ1) 3413, 2970,
1736, 1672, 1522, 1456, 1392, 1382, 1374, 1265, 1195, 1082, 1030,
4.2.12. Benzyl hydrogen [(R,S)-1-(benzyloxycarbonylamino)-
pentyl]phosphonate (16)
Dibenzyl ester 15 (17.9 g, 0.037 mol) and DABCO (6.23 g,
0.055 mol) in toluene (150 mL) were heated at 80 ꢀC over a period
of 12 h. The solvent was removed in vacuo, and the residue was
taken up in 100 mL of ethyl acetate and washed with 30 mL of 1 M
HCl. The organic phase was dried over Na2SO4 prior to filtration
and removal of the solvent in vacuo. Trituration of the crude
compound at ꢁ20 ꢀC from a mixture of ethyl acetate and petro-
leum ether afforded pure product. Yield 13.4 g (92%). Colourless
solid, mp 98–100 ꢀC. (lit.39 154–156 ꢀC). Calcd for C20H26NO5P
(391.39): C 61.37%, H 6.70%, N 3.58%. Found: 61.48%, 6.80%, 3.53%.
Rf¼0.86 (S5). 1H NMR (600 MHz, DMSO): 0.83 (3H, t, J¼7.1, CH3),
1.22 and 1.29 (2H, m, CH2), 1.22 and 1.35 (2H, m, CH2), 1.55 and
1.70 (2H, m, CH2), 3.80 (1H, dddd, J¼15.3, 11.6, 9.7 and 3.4, N–CH–
P), 4.91 (1H, dd, J¼12.1 and 7.1, O–CHaHb), 4.95 (1H, dd, J¼12.1
and 6.9, O–CHaHb), 5.04 (2H, s, O–CH2), 7.30–7.35 (10H, m,
2ꢂC6H5), 7.49 (1H, d, J¼9.7, NH). 13C NMR (150.9 MHz, DMSO):
14.08 (CH3), 21.82 (CH2), 28.03 (d, J(C,P)¼13.3, CH2), 28.55 (d,
J(C,P)¼3.0, CH2), 48.09 (d, J(C,P)¼155.3, N–CH–P), 65.63 (O–CH2),
66.35 (d, J(C,P)¼5.7, O–CH2), 127.57 (2ꢂAr–CH), 127.77 (2ꢂAr–
CH), 127.89 (Ar–CH), 128.04 (Ar–CH), 128.54 (2ꢂAr–CH), 128.56
(2ꢂAr–CH), 137.44 (Ar–C), 137.60 (d, J(C,P)¼6.5, Ar–C), 156.49 (d,
J(C,P)¼5.0, O–CO–N). IR (KBr, nmax cmꢁ1) 3291, 2800–2500 br,
1724, 1711, 1685, 1541, 1497, 1456, 1244, 1218, 1052, 1035, 1025,
737, 697. HRMS (ESI) calcd for C20H25NO5P [MꢁH]þ 390.1470;
found: 390.1478.
1002, 698. HRMS (ESI) calcd for C15H21NNaO4 [MþNa]þ 302.1368;
20
found: 302.1362. [
a]
ꢁ12.5 (c 0.584, CHCl3).
D
4.2.15. Methyl (2S)-2-[(hydroxyacetyl)amino]-3-methyl-
butanoate (18a)
Dipeptide 18a was prepared in the same manner as dipeptide
17a by the reaction of glycolic acid (0.52 g, 6.8 mmol), L-valine
methyl ester hydrochloride (1.14 g, 6.8 mmol), 1-hydroxybenzo-
triazole (0.92 g, 6.8 mmol), DBU (1.03 g, 6.8 mmol) and DIC
(1.03 g, 8.2 mmol). Yield 0.79 g (61%). Semisolid, Rf¼0.40 (S4). 1H
NMR (600 MHz, CDCl3): 0.94 (3H, d, J¼6.8, CH3), 0.96 (3H, d,
J¼6.8, CH3), 2.20 (1H, m, J¼6.8(6ꢂ) and 5.1, pCH–), 3.75 (3H, s,
OCH3), 4.12 (1H, dd, J¼15.0 and 5.5, O–CHaHb), 4.15 (1H, dd,
J¼15.0 and 5.0, O–CHaHb), 4.22 (1H, br t, J¼5.5 and 5.0, OH), 4.56
(1H, dd, J¼9.1 and 5.1, N–CH–CO), 7.21 (1H, br d, J¼9.1, NH). 13C
NMR (150.9 MHz, CDCl3): 17.69 (CH3), 18.90 (CH3), 31.12 (pCH–),
52.19 (OCH3), 56.63 (N–CHo), 62.02 (O–CH2), 172.26 and 172.27
(CO–O and CO–N). IR (CHCl3, nmax cmꢁ1) 3409, 2970, 1739, 1673,
1527, 1439, 1393, 1374, 1272, 1153, 1074. HRMS (ESI) calcd for
20
C8H15NNaO4 [MþNa]þ 212.0899; found: 212.0893. [
a
]
þ13.0 (c
D
0.432, CHCl3).
4.2.13. Benzyl (2S)-2-[(hydroxyacetyl)amino]-3-methyl-
butanoate (17a)
4.2.16. Methyl (2S)-2-{[(2S)-2-hydroxypropanoyl]amino}-3-
A mixture of glycolic acid (0.52 g, 6.8 mmol),
L-valine benzyl
methylbutanoate (18b)
ester p-toluensulfonate (2.58 g, 6.8 mmol), 1-hydroxybenzotriazole
(0.91 g, 6.8 mmol) and triethylamine (0.69 g, 6.8 mmol) in 50 mL of
dichloromethane was cooled to 0 ꢀC and DIC (1.72 g, 13.6 mmol)
was added dropwise. The reaction mixture was stirred at 0 ꢀC for
2 h and then at rt overnight. The solvent was removed in vacuo and
the residue was dissolved in 50 mL of cold ethyl acetate. Crystals of
diisopropylurea were filtered off and the filtrate was washed suc-
cessively with a saturated solution of citric acid and then with
a saturated solution of sodium hydrogencarbonate. The separated
organic layer was dried over Na2SO4 and concentrated to dryness.
The crude product was purified by flash chromatography on silica
gel using elution with a linear gradient of ethyl acetate in toluene.
Yield 1.45 g (80%). Semisolid, Rf¼0.31 (S2). 1H NMR (600 MHz,
CDCl3): 0.88 (3H, d, J¼6.8, CH3), 0.93 (3H, d, J¼6.8, CH3), 2.22 (1H,
Dipeptide 18b was prepared in the same manner as dipeptide
18a by the reaction of -lactic acid (0.61 g, 6.8 mmol), -valine
L
L
methyl ester hydrochloride (1.14 g, 6.8 mmol), 1-hydroxybenzo-
triazole (0.92 g, 6.8 mmol), DBU (1.03 g, 6.8 mmol) and DIC (1.03 g,
8.2 mmol). Yield 0.95 g (69%). Semisolid, Rf¼0.47 (S4). 1H NMR
(600 MHz, CDCl3): 0.92 (3H, d, J¼6.9, CH3), 0.95 (3H, d, J¼6.9, CH3),
1.45 (3H, d, J¼6.9, CH3), 2.20 (1H, m, J¼6.9(6ꢂ) and 5.0, pCH–), 3.75
(3H, s, OCH3), 4.28 (1H, q, J¼6.9(3ꢂ), pCH–O), 4.54 (1H, dd, J¼9.1
and 5.0, N–CH–CO), 7.10 (1H, br d, J¼9.1, NH). 13C NMR (150.9 MHz,
CDCl3): 17.64 (CH3), 18.92 (CH3), 21.25 (CH3), 31.17 (pCH–), 52.21
(OCH3), 56.57 (N–CHo), 68.48 (O–CHo), 172.52 (CO–O), 174.74 (CO–
N). IR (CHCl3, nmax cmꢁ1) 3413, 2970, 1738, 1672, 1523, 1439, 1373,
1270, 1154, 1119. HRMS (ESI) calcd for C9H17NNaO4 [MþNa]þ
20
226.1055; found: 226.1050. [
a
]
þ3.2 (c 0.437, CHCl3).
D

6098
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
4.2.17. (2S)-2-[(Hydroxyacetyl)amino]-3-methylbutanamide (19a)
Compound 18a (1.5 g, 7.9 mmol) was stirred over one week in
50 mL of a saturated solution of ammonia in methanol. The solvent
was evaporated in vacuo and the crude product was purified by flash
chromatography on silica gel using elution with a linear gradient of
ethanol in chloroform. Yield 0.83 g (60%). Colourless solid, mp 148–
151 ꢀC. Rf¼0.22 (S4). 1H NMR (600 MHz, DMSO): 0.80 (3H, d, J¼6.7,
CH3), 0.85 (3H, d, J¼6.7, CH3), 1.96 (1H, m, J¼6.7(6ꢂ) and 6.1, pCH–),
3.81 (1H, dd, J¼16.0 and 5.8, O–CHaHb), 3.86 (1H, dd, J¼16.0 and 5.8,
O–CHaHb), 4.17 (1H, dd, J¼9.2 and 6.1, N–CH–CO), 5.66 (1H, t, J¼5.8,
OH), 7.42 (1H, d, J¼9.2, NH), 7.14 and 7.56 (2H, d, Jgem¼1.8, CONH2).
13C NMR (150.9 MHz, DMSO): 18.01 (CH3),19.56 (CH3), 31.32 (pCH–),
56.73 (N–CHo), 61.55 (CH2–OH), 171.75 (CO–NH), 173.02 (CO–NH2).
IR (KBr, nmax cmꢁ1) 3370, 3206, 2973, 2965, 1680, 1617, 1550, 1426,
in toluene. Yield 0.64 g (86%). Colourless oil, Rf¼0.84 (S2). 1H NMR
(600 MHz, CDCl3): mixture of four diastereomers, assignment of
individual isomers was not possible, only regions of signals are
given: w0.85 (3H, t, J¼7.3, CH3), 1.42–1.44 (3H, d, J¼7.0, CH3), w1.25
(2H, m, CH2),1.30–1.40 (2H, m, CH2), w1.55 and w1.85 (2H, m, CH2),
4.10–4.25 (1H, m, N-CH-P), w4.32 (1H, br q, J¼7.0, O–CH–CO), 4.10–
4.25 and 5.05–5.45 (4H, m, 2ꢂO–CH2–CO), 4.92–5.00 (2H, m, P–O–
CH2), 4.90, 5.21, 5.28 and 5.45 (1H, br dd, NH–CO), 7.29–7.39 (15H,
m, 3ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): w13.8 (CH3), 18.9–20.3
(CH3), w22.1 (CH2), 27.7–28.0 (CH2), 29.3–29.8 (CH2), 47.6–48.9 (N–
CH–P), 66.77 (O–CH–CO), 66.9–68.0 (2ꢂO–CH2), 70.54, 70.71, 70.88
and 71.06 (P–O–CH2), 127.79–128.62 (15ꢂAr–CH), 134.9–136.4
(3ꢂAr–C), 155.9–156.25 (O–CO–NH), 175.5 and 170.75–171.1 (CO–
O). IR (CHCl3, nmax cmꢁ1) 3430, 3033, 1757, 1729, 1510, 1456, 1380,
1250, 1098, 1041, 999, 698. HRMS (ESI) calcd for C30H36NNaO7P
[MþNa]þ 576.2127; found: 576.2121.
1390,1372,1252,1230,1082, 666. HRMS (ESI) calcd for C7H14N2NaO3
20
[MþNa]þ 197.0902; found: 197.0897. [
a]
þ3.2 (c 0.495, CH3OH).
D
4.2.18. (2S)-2-{[(2S)-2-Hydroxypropanoyl]amino}-3-
methylbutanamide (19b)
4.2.19.3. Methyl {[(R,S)-1-(benzyloxycarbonylamino)-pentyl(benzyloxy)-
phosphoryl]oxy}acetate (22a). Compound 22a was prepared by the
reaction of 16 and methyl glycolate (0.18 g, 1.95 mmol). Flash chro-
matography: linear gradient of ethyl acetate in toluene. Yield 0.51 g
(85%). Colourless oil, Rf¼0.54 (S2). 1H NMR (600 MHz, CDCl3): mixture
of diastereoisomers w2:1, only signals of the major isomer are given:
0.87 (3H, t, J¼7.1, CH3),1.27 and 1.34 (2H, m, CH2), 1.34 and 1.44 (2H, m,
CH2),1.57 and 1.88 (2H, m, CH2), 3.75 (3H, s, OCH3), 4.20 (1H, m, N–CH–
P), 4.47 (1H, dd, J¼16.2 and 11.0, O–CHaHb–CO), 4.60 (1H, dd, J¼16.2
and 11.7, O–CHaHb–CO), 5.08 and 5.12 (2H, d, Jgem¼12.2, OCH2), 5.13
(2H, m, OCH2), 5.24 (1H, br dd, J¼10.2 and 1.5, NH), 7.30–7.38 (10H, m,
2ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.81 (CH3), 22.14 (CH2), 27.84
(d, J(C,P)¼12.8, CH2), 29.54 (CH2), 48.17 (d, J(C,P)¼156.2, N-CH-P),
52.39 (OCH3), 62.07 (d, J(C,P)¼6.3, O–CH2), 67.04 (O–CH2), 67.79 (d,
J(C,P)¼7.4, O–CH2), 127.92–128.59 (10ꢂAr–CH), 135.85 (d, J(C,P)¼5.9,
Ar–C), 136.28 (Ar–C), 156.14 (d, J(C,P)¼5.9, O–CO–N), 169.10 (d,
J(C,P)¼4.4, O–CO). IR (CHCl3, nmax cmꢁ1) 3431, 3031, 1762, 1721, 1510,
1456, 1442, 1384, 1234, 1101, 1039, 1010, 697. HRMS (ESI) calcd for
C23H29NO7P [MꢁH]þ 462.1682; found: 462.1674.
Dipeptide 19b was prepared from compound 18b (1.5 g,
7.3 mmol) in the same manner as compound 19a. Yield 1.1 g (79%).
Colourless solid, mp 140–143 ꢀC. Rf¼0.25 (S4). 1H NMR (600 MHz,
DMSO): 0.79 (3H, d, J¼6.7, CH3), 0.84 (3H, d, J¼6.7, CH3), 1.22 (3H, d,
J¼6.8, CH3), 1.94 (1H, m, J¼6.7(6ꢂ) and 6.2, pCH–), 3.97 (1H, qd,
J¼6.8(3ꢂ) and 5.2, O–CH–CO), 4.16 (1H, dd, J¼9.4 and 6.2, N–CH–
CO), 5.72 (1H, d, J¼5.2, OH), 7.40 (1H, d, J¼9.4, NH), 7.14 and 7.55
(2H, d, Jgem¼1.8, CONH2). 13C NMR (150.9 MHz, DMSO): 17.89 (CH3),
19.53 (CH3), 21.56 (CH3), 31.48 (pCH–), 56.54 (N–CHo), 67.67 (O–
CHo), 173.03 (CO–NH2), 174.41 (CO–NH). IR (KBr, nmax cmꢁ1) 3389,
3361, 3300, 3201, 2976, 2961, 1667, 1648, 1625, 1550, 1432, 1236,
1142, 1125, 661. HRMS (ESI) calcd for C8H16N2NaO3 [MþNa]þ
20
211.1059; found: 211.1053. [
a
]
ꢁ13.2 (c 0.608, CH3OH).
D
4.2.19. General procedure for the preparation of compounds 20a,
20b, 22a, 22b, 24a, 24b, 26a, 26b, 28a, 28b, 30a and 30b
Monoester 16 (0.5 g, 1.3 mmol), the hydroxy component
(1.95 mmol) and BOP (0.86 g, 1.95 mmol) were dissolved in 10 mL
of dry DMF, and TEA (0.53 g, 5.2 mmol) was added in one portion.
The reaction mixture was stirred at rt overnight, the solvent was
evaporated and the resulting black oil was purified by flash chro-
matography on silica gel.
4.2.19.4. Methyl (2S)-2-{[(R,S)-1-(benzyloxycarbonylamino)-pentyl-
(benzyloxy)phosphoryl]oxy} propanoate (22b). Compound 22b was
prepared by the reaction of 16 and methyl (S)-lactate (0.20 g,
1.95 mmol). Flash chromatography: linear gradient of ethyl acetate
in toluene. Yield 0.50 g (82%). Colourless oil, Rf¼0.61 (S2). 1H NMR
(600 MHz, CDCl3): mixture of four diastereomers, assignment of
individual isomers was not possible, only regions of signals are
given: 0.86–0.87 (3H, t, J¼7.0, CH3), 1.41–1.49 (3H, d, J¼7.0, CH3),
1.25–1.33 (2H, m, CH2), 1.33–1.43 (2H, m, CH2), w1.55 and w1.85
(2H, m, CH2), 3.67–3.74 (3H, s, OCH3), 4.12–4.25 (1H, m, N–CH–P),
4.88–4.98 (1H, m, O–CH–CO), 5.05–5.21 (4H, m, 2ꢂO–CH2), 4.88,
5.16, 5.29 and 5.44 (1H, dd, NH–CO), 7.30–7.37 (10H, m, 2ꢂC6H5).
13C NMR (150.9 MHz, CDCl3): 13.81–13.84 (CH3), 18.96, 19.02, 19.08
and 19.37 (CH3), 22.0–22.2 (CH2), 27.7–28.0 (CH2), 29.3–29.9 (CH2),
47.6–48.8 (N-CH-P), 52.6–52.4 (OCH3), 66.9–68.1 (2ꢂO–CH2),
70.49, 70.67, 70.81 and 70.93 (d, Jw6.5, O–CH–CO), 127.8–128.6
(10ꢂAr–CH), 135.8–136.4 (2ꢂAr–C), 155.9–156.2 (O–CO–NH),
171.4–171.7 (CO–O). IR (CHCl3, nmax cmꢁ1) 3430, 3031, 1751, 1721,
1511, 1456, 1439, 1380, 1247, 1055, 1041, 997, 697. HRMS (ESI) calcd
for C24H32NNaO7P [MþNa]þ 500.1814; found: 500.1807.
4.2.19.1. Benzyl {[(R,S)-1-(benzyloxycarbonylamino)-pentyl(benzyloxy)-
phosphoryl]oxy}acetate (20a). Compound 20a was prepared by the
reaction of 16 and benzyl glycolate (0.32 g, 1.95 mmol). Flash
chromatography: linear gradient of ethyl acetate in toluene. Yield
0.60 g (88%). Colourless oil, Rf¼0.76 (S2). 1H NMR (600 MHz, CDCl3):
mixture of diastereoisomers (w2:1), only signals of the major iso-
mer are given: 0.86 (3H, t, J¼7.1, CH3), 1.25 and 1.32 (2H, m, CH2),
1.32 and 1.42 (2H, m, CH2), 1.56 and 1.87 (2H, m, CH2), 4.18 (1H, m,
N–CH–P), 4.50 (1H, dd, J¼16.3 and 10.8, O–CHaHb–CO), 4.62 (1H,
dd, J¼16.3 and 11.5, O–CHaHb–CO), 5.06–5.19 (6H, m, 3ꢂ–CH2–O),
5.18 (1H, d, J¼9.5, NH), 7.29–7.36 (15H, m, 3ꢂC6H5). 13C NMR
(150.9 MHz, CDCl3): 13.84 (CH3), 22.15 (CH2), 27.85 (d, J(C,P)¼12.8,
CH2), 29.56 (d, J(C,P)¼2.1, CH2), 48.21 (d, J(C,P)¼156.3, N–CH–P),
62.21 (O–CH2–CO), 67.08 (O–CH2), 67.36 (O–CH2), 67.80 (d,
J(C,P)¼7.4, O–CH2), 127.99–128.67 (15ꢂArCH), 125.85 (d, J(C,P)¼5.9,
Ar–C), 136.30 (2ꢂAr–C), 156.14 (d, J(C,P)¼5.7, O–CO–N), 168.57
(–CO–O). IR (CHCl3, nmax cmꢁ1) 3431, 3034, 1759, 1721, 1510, 1456,
1381, 1252, 1098, 1039, 1010, 1002, 698. HRMS (ESI) calcd for
C29H34NNaO7P [MþNa]þ 562.1971; found: 562.1964.
4.2.19.5. {[((R,S)-1-Benzyloxycarbonylamino)-pentyl(benzyloxy)-
phosphoryl]oxy}acetamide (24a). Compound 24a was prepared by
the reaction of 16 and glycolamide (0.15 g, 1.95 mmol). Flash
chromatography: linear gradient of ethanol in chloroform. Yield
0.42 g (74%). Pale yellow oil, Rf¼0.23 (S2). 1H NMR (600 MHz,
CDCl3): mixture of diastereoisomers w2:1, only signals of the major
isomer are given: 0.87 (3H, t, J¼7.3, CH3), 1.24–1.32 (2H, m, CH2),
1.31–1.44 (2H, m, CH2), 1.49 and 1.89 (2H, m, CH2), 4.11 (1H, m, N–
4.2.19.2. Benzyl (2S)-2-{[(R,S)-1-(benzyloxycarbonylamino)-pentyl-
(benzyloxy)phosphoryl]oxy} propanoate (20b). Compound 20b was
prepared by the reaction of 16 and benzyl (S)-lactate (0.39 g,
1.95 mmol). Flash chromatography: linear gradient of ethyl acetate

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6099
CH–P), 4.14 (1H, dd, J¼15.2 and 5.6, O–CHaHb–CO), 4.53 (1H, dd,
J¼15.2 and 7.8, O–CHaHb–CO), 5.08 (4H, m, 2ꢂO–CH2), 5.93 (1H, d,
J¼9.8, NH), 5.96 and 6.88 (2H, br, CONH2), 7.30–7.36 (10H, m,
2ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.74 (CH3), 21.99 (CH2),
27.82 (d, J(C,P)¼13.4, CH2), 28.55 (d, J(C,P)¼3.0, CH2), 48.41 (d,
J(C,P)¼156.4, N–CH–P), 63.42 (d, J(C,P)¼7.6, O–CH2), 67.33 (O–CH2),
68.51 (d, J(C,P)¼7.2, O–CH2), 127.93–128.92 (10ꢂAr–CH), 135.57 (d,
J(C,P)¼4.6, Ar–C), 136.08 (Ar–C), 156.37 (d, J(C,P)¼3.5, O–CO–N),
169.86 (d, J(C,P)¼8.8, CO–NH2). IR (CHCl3, nmax cmꢁ1) 3484, 3431,
1718, 1694, 1579, 1511, 1456, 1403, 1380, 1241, 1066, 1037, 970, 698.
HRMS (ESI) calcd for C22H29N2NaO6P [MþNa]þ 471.1661; found:
471.1651.
(1H, m, N–CH–CO), 4.86–4.90 (1H, m, O–CH–CO), 5.00–5.17 (6H, m,
3ꢂO–CH2), 7.30–7.39 (15H, m, 3ꢂC6H5), 7.60, 7.70, 7.72 and 7.73
(1H, d, Jw9.5, CO–NH), 8.16, 8.21, 8.23 and 8.28 (1H, d, Jw8.2, CO–
NH). 13C NMR (150.9 MHz, CDCl3): 14.0–14.3 (CH3), 18.1–19.1
(2ꢂCH3), 19.8–20.2 (CH3), w21.7 (CH2), w27.9 (CH2), w28.2 (CH2),
30.0–30.4 (–CHo), 47.2–48.7 (N–CH–P), 56.87, 57.52, 57.60 and
57.68 (N–CH–CO), 65.8–67.2 (3ꢂO–CH2), 71.3–71.7 (O–CH–CO),
127.7–128.7 (15ꢂAr–CH), 136.0–137.2 (3ꢂAr–C), 156.4–156.6 (O–
CO–NH), 170.7–171.5 (CO–NH and CO–O). IR (CHCl3, nmax cmꢁ1
)
3429, 3033, 1736, 1724, 1682, 1514, 1456, 1392, 1376, 1247, 999, 698.
HRMS (ESI) calcd for C35H45N2NaO8P [MþNa]þ 675.2811; found:
675.2808.
4.2.19.6. (2S)-2-{[((R,S)-1-Benzyloxycarbonylamino)-pentyl(benzyloxy)-
phosphoryl]oxy} propionamide (24b). Compound 24b was prepared
by the reaction of 16 and (S)-lactamide (0.17 g, 1.95 mmol). Flash
chromatography: linear gradient of ethanol in chloroform. Yield
0.45 g (70%). Pale yellow oil, Rf¼0.29 (S2). 1H NMR (600 MHz,
DMSO): mixture of four diastereomers, assignment of individual
isomers was not possible, only regions of signals are given: 0.82–
0.83 (3H, t, J¼7.3, CH3), w1.22 (2H, m, CH2), 1.28–1.35 (3H, d, J¼6.8,
CH3), w1.32 (2H, m, CH2), 1.55 and 1.69 (2H, m, CH2), 3.90–4.02 (1H,
m, N–CH–P), 4.57–4.66 (1H, m, O–CH–CO), 5.01–5.09 (4H, m, O–
CH2), 7.30–7.38 (10H, m, 2ꢂC6H5), 7.40–7.46 (2H, CO–NH2), 7.72–
7.78 (1H, d, J¼9.5, NH). 13C NMR (150.9 MHz, DMSO): 14.01–14.03
(CH3), 19.98–20.32 (CH3), w21.70 (CH2), 27.76–28.22 (2ꢂCH2),
47.11–48.39 (N–CH–P), 65.84–66.02 (O–CH2), 67.16–67.36 (O–CH2),
71.70–72.16 (O–CH–CO),127.77–128.68 (10ꢂAr–CH),136.66–137.23
(2ꢂAr–C), 156.46–156.67 (O–CO–NH), 172.43–172.66 (CONH2). IR
(CHCl3, nmax cmꢁ1) 3430, 1712, 1697, 1587, 1511, 1456, 1378, 1088,
1038, 1007, 999, 980, 698. HRMS (ESI) calcd for C23H31N2NaO6P
[MþNa]þ 485.1817; found: 485.1810.
4.2.19.9. Methyl (2S)-2-[({[((R,S)-1-benzyloxycarbonylamino)-pentyl-
(benzyloxy)phosphoryl]oxy} acetyl)amino]-3-methylbutanoate (28a).
Compound 28a was prepared by the reaction of 16 and dipeptide 18a
(0.37 g, 1.95 mmol). Flash chromatography: linear gradient of ethyl
acetate in toluene. Yield 0.60 g (83%). Colourless oil, Rf¼0.36 (S2). 1H
NMR (600 MHz, CDCl3): mixture of four diastereomers, assignment
of individual isomers was not possible, only regions of signals are
given: 0.86–0.89 (3H, t, J¼7.3, CH3), 0.89–0.96 (6H, d, J¼7.0, CH3),
w1.26 and w1.33 (2H, m, CH2), w1.33 and w1.45 (2H, m, CH2), 1.56–
1.60 and 1.82–1.90 (2H, m, CH2), 2.15–2.23 (1H, m, –CHo), 3.70–3.72
(3H, s, OCH3), 4.15–4.20 (1H, m, N–CH–P), 4.25–4.57 (2H, dd, J w 15.0
and 8.0, O–CH2–CO), 4.47–4.52 (1H, N–CH–CO), 5.06–5.17 (4H, m,
2ꢂO–CH2), 4.96, 5.01 and 5.13 (1H, br d, Jw10.0, O–CO–NH), 7.06,
7.07, 7.17 and 7.21 (1H, br d, Jw9.0, CO–NH), 7.31–7.39 (10H, m,
2ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.75–13.83 (CH3), 17.79–
18.07 and 18.84–19.01 (2ꢂCH3), 22.06–22.13 (CH2), 27.75–27.87
(CH2), 28.74–29.15 (CH2), 30.87–31.11 (–CHo), 47.44–48.75 (N–CH–
P), 52.08–52.18 (OCH3), 56.89–57.14 (N–CH–CO), 63.57–64.00 (O–
CH2–CO), 67.25–67.45 and 68.61–68.87 (2ꢂO–CH2), 127.94–128.92
(10ꢂAr–CH), 135.46–136.05 (2ꢂAr–C), 156.00–156.20 (O–CO–NH),
4.2.19.7. Benzyl (2S)-2-[({[((R,S)-1-benzyloxycarbonylamino)-pentyl-
(benzyloxy)phosphoryl]oxy} acetyl)amino]-3-methylbutanoate (26a).
Compound 26a was prepared by the reaction of 16 and dipeptide
17a (0.51 g, 1.95 mmol). Flash chromatography: linear gradient of
ethyl acetate in toluene. Yield 0.58 g (70%). Colourless oil, Rf¼0.43
(S2). 1H NMR (600 MHz, CDCl3): mixture of four diastereomers, as-
signment of individual isomers was not possible; only regions of
signals are given: w0.87 (3H, t, J¼7.0, CH3), 0.84–0.93 (6H, d, J¼7.0,
CH3), w1.26 and w1.32 (2H, m, CH2), w1.33 and w1.44 (2H, m, CH2),
w1.82 and w1.89 (2H, m, CH2), 2.17–2.24 (1H, m, –CHo), 4.15–4.20
(1H, m, N–CH–P), 4.25–4.43 (2H, O–CH2–CO), w4.57 (1H, m, N–CH–
CO), 4.93–5.12 (1H, br d, CO–NH), 5.06–5.18 (6H, m, O–CH2–CO),
7.07–7.23 (1H, br d, CO–NH), 7.30–7.37 (15H, m, 3ꢂC6H5). 13C NMR
(150.9 MHz, CDCl3): w13.8 (CH3), 17.6–19.0 (2ꢂCH3), 22.0–22.1
(CH2), 27.7–27.9 (CH2), 28.7–29.2 (CH2), 30.9–31.2 (–CHo), 47.4–48.0
(N–CH–P), 56.9–57.1 (N–CH–CO), 63.6–64.0 (O–CH2–CO), 66.9–68.9
(3ꢂO–CH2), 127.9–129.0 (15ꢂAr–CH), 135.2–136.1 (3ꢂAr–C), 156.0–
156.2 (O–CO–NH), 167.1–167.2 (CO–NH), 171.2–171.4 (CO–O). IR
(CHCl3, nmax cmꢁ1) 3431, 1735, 1723, 1685, 1510, 1456, 1393, 1376,
1247, 1198, 1148, 1067, 1040, 1009, 1000, 970, 698. HRMS (ESI) calcd
for C34H43N2NaO8P [MþNa]þ 661.2655; found: 661.2650.
167.03–167.20 (CO–NH),171.80–171.99 (CO–O). IR (CHCl3, nmax cmꢁ1
)
3431, 1740, 1722, 1684, 1510, 1456, 1439, 1375, 1067, 1039, 1009, 1000,
698. HRMS (ESI) calcd for C28H38N2O8P [MꢁH]þ 561.2366; found:
561.2355.
4.2.19.10. Methyl (2S)-2-{[(2S)-2-{[((R,S)-1-benzyloxycarbonylamino)-
pentyl(benzyloxy) phosphoryl]oxy}propanoyl]amino}-3-methylbutanoate
(28b). Compound 28b was prepared by the reaction of 16 and di-
peptide 18b (0.39 g, 1.95 mmol). Flash chromatography: linear
gradient of ethyl acetate in toluene. Yield 0.58 g (78%). Colourless
oil, Rf¼0.44 (S2). 1H NMR (600 MHz, CDCl3): mixture of four di-
astereomers, only regions of signals are given: w0.86 (3H, t, J¼7.3,
CH3), 0.88–0.94 (6H, d, J¼7.0, CH3), 1.54, 1.48, 1.45 and 1.40 (3H, d,
J¼7.0, CH3), w1.25 and w1.33 (2H, m, CH2), w1.32 and w1.46 (2H,
m, CH2), w1.50 and w1.83 (2H, m, CH2), 2.15–2.22 (1H, m, –CHo),
3.72, 3.70, 3.68 and 3.66 (3H, s, OCH3), 4.15–4.22 (1H, m, N–CH–P),
4.46–4.50 (1H, m, N–CH–CO), 4.81–4.90 (1H, br d, O–CH–CO), 5.05–
5.15 (4H, m, O–CH2), 5.05, 5.10, 5.25 and 5.45 (1H, br d, J w 10.0,
O–CO–NH), 6.99, 7.06, 7.09 and 7.39 (1H, br d, Jw9.0, CO–NH), 7.30–
7.38 (10H, m, 2ꢂC6H5). 13C NMR (150.9 MHz, CDCl3): 13.74–13.78
(CH3), 17.67–17.89 and 18.86–18.95 (2ꢂCH3), 19.86–20.17 (CH3),
22.02–22.08 (CH2), 27.73–27.93 (CH2), 28.84–29.44 (CH2), 30.90–
31.19 (–CHo), 47.61–49.14 (N–CH–P), 52.09–52.25 (OCH3), 56.74–
57.03 (N–CH–CO), 67.12–67.25 and 68.25–68.64 (2ꢂO–CH2),
72.95–73.35 (O–CH–CO), 127.91–128.72 (10ꢂAr–CH), 135.61–
136.22 (2ꢂAr–C), 155.94–156.22 (O–CO–NH), 170.20–170.47 (CO–
NH), 171.93–172.39 (CO–O). IR (CHCl3, nmax cmꢁ1) 3430, 1741, 1722,
1684, 1512, 1456, 1439, 1375, 1088, 1026, 998, 697. HRMS (ESI) calcd
for C29H41N2NaO8P [MþNa]þ 599.2498; found: 599.2492.
4.2.19.8. Benzyl (2S)-2-{[(2S)-2-{[((R,S)-1-benzyloxycarbonylamino)-
pentyl(benzyloxy) phosphoryl]oxy}propanoyl]amino}-3-methylbutanoate
(26b). Compound 26b was prepared by the reaction of 16 and di-
peptide 17b (0.55 g, 1.95 mmol). Flash chromatography: linear
gradient of ethyl acetate in toluene. Yield 0.60 g (72%). Colourless
oil, Rf¼0.33 (S2). 1H NMR (600 MHz, CDCl3): mixture of four di-
astereomers, assignment of individual isomers was not possible;
only regions of signals are given: 0.80–0.83 (3H, t, J¼7.0, CH3), 0.83–
0.85 (6H, d, J¼7.0, CH3), 1.27–1.31 (3H, d, J¼6.8, CH3), w1.22 (2H, m,
CH2), w1.19 and w1.33 (2H, m, CH2), w1.57 and w1.68 (2H, m, CH2),
2.05–2.12 (1H, m, –CHo), 3.88–3.99 (1H, m, N–CH–P), 4.20–4.26
4.2.19.11. (2S)-2-[({[((R,S)-1-Benzyloxycarbonylamino)-pentyl(ben-
zyloxy)phosphoryl]oxy}acetyl) amino]-3-methylbutanamide (30a).
Compound 30a was prepared by the reaction of 16 and dipeptide

6100
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
19a (0.34 g, 1.95 mmol). Flash chromatography: linear gradient of
ethanol in chloroform. Yield 0.53 g (76%). Semisolid, Rf¼0.40 (S4).
1H NMR (600 MHz, DMSO): mixture of four diastereomers, as-
signment of individual isomers was not possible, only regions of
signals are given: 0.80–0.86 (9H, m, 3ꢂCH3), w1.22 (2H, m, CH2),
w1.22 and w1.34 (2H, m, CH2), w1.58 and w1.71 (2H, m, CH2),
1.97–2.02 (1H, m, –CHo), 3.95–4.02 (1H, N–CH–CO), 4.16–4.20 (1H,
m, N–CH–P), 4.40–4.50 (2H, dd, O–CH2–CO), 5.03–5.11 (4H, m,
2ꢂO–CH2), w7.14 and w7.52 (2H, d, J¼1.8, CONH2), 7.30–7.38 (10H,
m, 2ꢂC6H5), 7.73, 7.74, 7.75, 7.76, 7.81, 7.83, 7.84 and 7.88 (2H, d,
2ꢂCO–NH). 13C NMR (150.9 MHz, DMSO): 14.01–14.02 (CH3),
17.95–18.03 and 19.49 (2ꢂCH3), 21.74 (CH2), 27.89–28.39 (2ꢂCH2),
30.62–30.68 (–CHo), 47.84, 47.88, 47.90 and 47.96 (d, J(C,P)w156,
N–CH–P), 57.36 and 57.39 (N–CH–CO), 63.80–63.97 (O–CH2–CO),
65.89–65.96 and 67.11–67.28 (2ꢂO–CH2), 127.83–128.67 (10ꢂAr–
CH), 136.68–137.23 (2ꢂAr–C), 156.44–156.50 (O–CO–NH), 167.25–
J(C,P)¼8.6, CH2), 27.76 (d, J(C,P)¼2.2, CH2), 46.34 (d, J(C,P)¼154.0,
N–CH–P), 53.50 (d, J(C,P)¼7.0, OCH3), 64.26 (d, J(C,P)¼6.5, O–CH2–
CO), 169.80 (d, J(C,P)¼4.4, N–CO–C). IR (KBr, nmax cmꢁ1) 3190, 1685,
1627, 1205, 1178, 1044, 801. HRMS (ESI) calcd for C8H19N2NaO4P
[MþNa]þ 261.0980; found: 261.0974.
4.2.20.2. (2S)-2-[({[(R,S)-1-Aminopentyl(methoxy)phosphoryl]oxy}
acetyl)amino]-3-methylbutanoic acid (10). Phosphonate 10 was
prepared by hydrogenolysis of 9 (0.6 g, 1.1 mmol). Yield 179.7 mg
(50%). Semisolid, tR¼43.4 min, G1. 1H NMR (600 MHz, CDCl3):
Mixture of four diastereomers, assignment of individual isomers
was not possible, only regions of signals are given: 0.90–0.91 (3H,
t, J¼7.3, CH3), 0.94–0.99 (6H, d, J¼7.0, 2ꢂCH3), w1.35 (2H, m, CH2),
1.43–1.58 (2H, m, CH2), 1.87–2.00 (2H, m, CH2), 3.60–3.70 (1H, m,
N–CH–P), 3.82–3.89 (3H, d, J(H,P)w11.0, OCH3), 4.28–4.43 (1H, m,
N–CH–CO), 4.63–5.03 (2H, m, O–CH2–CO), 7.51, 7.61, 7.93 and 8.04
(1H, br d, Jw7.5, CO–NH). 13C NMR (150.9 MHz, CDCl3): w13.3
(CH3), 17.5–18.8 (2ꢂCH3), w21.8 (CH2), w27.3 (CH2), w27.6 (CH2),
47.2–48.2 (N–CH–P), 53.0–54.1 (OCH3), 57.7–58.8 (N–CH–CO),
64.0–66.0 (O–CH2), 168.5–170.0 (CO–NH), w175.0 (COOH). IR
(KBr, nmax cmꢁ1) 3300, 3185, 1730, 1670, 1554, 1203, 1047, 799.
HRMS (ESI) calcd for C13H28N2O6P [MþH]þ 339.1685; found:
339.1679.
167.43 (CO–NH), 172.74 and 172.76 (CO–NH2). IR (CHCl3, nmax cmꢁ1
)
3428, 3410, 3323, 1717, 1695, 1676, 1593, 1512, 1456, 1391, 1375,
1066, 1041, 1009, 1000, 698. HRMS (ESI) calcd for C27H37N3O7P [M-
H]þ 546.2369; found: 546.2375.
4.2.19.12. (2S)-2-{[(2S)-2-{[((R,S)-1-Benzyloxycarbonylamino)-pentyl-
(benzyloxy)phosphoryl]oxy} propanoyl]amino}-3-methylbutanamide
(30b). Compound 30b was prepared by the reaction of 16 and di-
peptide 19b (0.37 g, 1.95 mmol). Flash chromatography: linear
gradient of ethanol in chloroform. Yield 0.59 g (82%). Semisolid,
Rf¼0.21 (S2). 1H NMR (600 MHz, DMSO): mixture of four di-
astereomers; assignment of individual isomers was not possible,
only regions of signals are given: w0.82 (3H, t, J¼7.3, CH3), 0.81–
0.86 (6H, d, J¼7.0, CH3), w1.22 (2H, m, CH2), w1.21 and w1.33 (2H,
m, CH2), 1.30–1.35 (3H, d, J¼6.8, CH3), 1.50–1.69 (2H, m, CH2), 1.95–
2.01 (1H, m, –CHo), 3.90–4.01 (1H, m, N–CH–P), 4.15 (1H, dd, J¼9.0
and 6.4, N–CH–CO), 4.82 (1H, m, O–CH–CO), 5.01–5.09 (4H, m, O–
CH2), 7.11–7.15 and 7.47–7.51 (2H, d, Jw2.0, CONH2), 7.30–7.38 (10H,
m, 2ꢂC6H5), 7.64, 7.68, 7.70, 7.72, 7.73, 7.74, 7.75 and 7.79 (2H, d,
Jw9–10, O–CO–NHþCO–NH). 13C NMR (150.9 MHz, DMSO): 13.98–
14.05 (CH3), 17.99–18.15 and 19.46–19.51 (2ꢂCH3), 19.92–20.29
(CH3), 21.69–21.74 (CH2), 27.72–28.40 (2ꢂCH2), 30.72–30.97
(–CHo), 47.28–48.57 (N–CH–P), 57.24–57.45 (N–CH–CO), 65.86–
65.98 and 67.02–67.34 (2ꢂO–CH2), 71.95–72.43 (O–CH–CO),
127.72–129.16 (10ꢂAr–CH), 136.75–137.61 (2ꢂAr–C), 156.43–
156.51 (O–CO–NH), 170.10–170.26 (CO–NH), 172.69–172.75 (CO–
NH2). IR (CHCl3, nmax cmꢁ1) 3430, 3410, 3329, 1717, 1691, 1680, 1512,
1456, 1392, 1375, 1242, 1086, 1038, 1025, 1007, 999, 697. HRMS (ESI)
calcd for C28H40N3NaO7P [MþNa]þ 584.2502; found: 584.2502.
4.2.20.3. {[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}acetic acid
(21a). Phosphonate 21a was prepared by hydrogenolysis of 20a
(0.35 g, 0.81 mmol). Yield 102.3 mg (70%). Colourless powder,
tR¼28.4 min, G1. 1H NMR (600 MHz, DMSO): 0.86 (3H, d, J¼7.3,
CH3), 1.26 (2H, m, CH2), 1.36 and 1.42 (2H, m, CH2), 1.59 and 1.77
(2H, m, CH2), 3.07 (1H, m, N–CH–P), 4.40 (2H, d, J¼10.9, O–CH2–CO),
8.02 (2H, br, NH2). 13C NMR (150.9 MHz, DMSO): 14.01 (CH3), 22.22
(CH2), 27.95 (d, J(C,P)¼8.1, CH2), 28.55 (CH2), 47.79 (d, J(C,P)¼145.2,
N–CH–P), 61.91 (O–CH2), 172.35 (COOH). IR (KBr, nmax cmꢁ1) 3265,
3200–2700 br, 2658, 2568, 1720, 1620, 1535, 1204, 1180, 1058, 1012,
949, 893, 817. HRMS (ESI) calcd for C7H15NO5P [MꢁH]þ 224.0688;
found: 224.0693.
Alternatively, phosphonate 21a was prepared from diester 8.
Compound 8 (0.5 g, 1.2 mmol) was dissolved in 5 mL of dry
dichloromethane and TMSBr (0.59 g, 3.9 mmol) was added in one
portion. The reaction mixture was stirred for 3 h at rt until the
starting material disappeared (TLC monitoring). The reaction was
quenched by adding 5 mL of 1 M HCl. The organic phase was sep-
arated, washed with brine and dried over Na2SO4. The solvent was
removed in vacuo to give 0.5 g of an oil, which was hydrogenated
according to the method described above. The crude product was
purified by RP-HPLC. Yield 2.8 mg (3% over two steps).
4.2.20. General procedure for the preparation of compounds 6, 10,
21a, 21b, 23a, 23b, 25a, 25b, 27a, 27b, 29a, 29b, 31a and 31b
The protected precursor was dissolved in 80 mL of methanol
and then 10% Pd–C (0.15 g) was added. The mixture was vigorously
stirred and allowed to react under an atmosphere of hydrogen
(15 psi) at rt overnight. The catalyst was filtered off through Celite
and the filter was washed with 100 mL of methanol. The filtrate was
concentrated in vacuo and the crude product was purified using
preparative RP-HPLC. Finally, the target compounds were lyophi-
lised from water.
4.2.20.4. (2S)-2-{[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}pro-
panoic acid (21b). Phosphonate 21b was prepared by hydro-
genolysis of 20b (0.25 g, 0.45 mmol). Yield 74.5 mg (69%). Colourless
powder, tR¼31.5 min, G1. 1H NMR (600 MHz, D2O): 0.90 (3H, d,
J¼7.3, CH3), 1.53 (3H, d, J¼7.0, CH3), 1.36 (2H, m, CH2), 1.38 and 1.48
(2H, m, CH2), 1.73 and 1.92 (2H, m, CH2), 3.33 (1H, m, N–CH–P), 4.86
(1H, dq, J¼8.3 and 7.0(3ꢂ), O–CH–CO). 13C NMR (150.9 MHz, D2O):
15.67 (CH3), 22.03 (d, J¼4.4, CH3), 24.32 (CH2), 30.24 (d, J¼9.2, CH2),
30.28 (CH2), 51.32 (d, J¼148.7, N–CH–P), 72.11 (d, J¼6.0, O–CHo),
179.21 (d, J¼4.3, COOH). IR (KBr, nmax cmꢁ1) 3200–2700 br, 2629,
2594, 1719, 1699, 1639, 1536, 1190, 1177, 1049, 989, 806. HRMS (ESI)
calcd for C8H17N2O5P [MꢁH]þ 238.0844; found: 238.0849.
4.2.20.1. {[(R,S)-1-Aminopentyl(methoxy)phosphoryl]oxy}acetamide
(6). Phosphonate 6 was prepared by hydrogenolysis of 7 (0.3 g,
0.81 mmol). Yield 113.5 mg (59%). Semisolid, tR¼34.8 min, G1. 1H
NMR (600 MHz, DMSO): mixture of diastereoisomers (w2:1), only
signals of the major isomer are given: 0.87 (3H, t, J¼7.3, CH3), 1.29
(2H, m, CH2), 1.37 and 1.43 (2H, m, CH2), 1.68 and 1.79 (2H, m, CH2),
3.74 (1H, m, N–CH–P), 3.78 (3H, d, J(H,P)¼10.9, OCH3), 4.50 (2H, m,
4.2.20.5. Methyl {[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}acetate
(23a). Phosphonate 23a was prepared by hydrogenolysis of 22a
(0.45 g, 0.97 mmol). Yield 182 mg (78%). Colourless powder,
tR¼33.2 min, G1. 1H NMR (600 MHz, DMSO): 0.86 (3H, t, J¼7.3, CH3),
1.25 (2H, m, CH2),1.25 and 1.42 (2H, m, CH2),1.55 and 1.74 (2H, m, CH2),
2.91 (1H, m, N–CH–P), 3.65 (3H, s, OCH3), 4.41 (1H, dd, J¼16.4 and 9.8,
CO–CH2–O), 7.58 and 7.72 (2H, br, CONH2), 8.47 (2H, br, NH2). 13
C
NMR (150.9 MHz, DMSO): 13.83 (CH3), 21.92 (CH2), 27.37 (d,

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6101
O–CHaHb–CO), 4.43 (1H, dd, J¼16.4 and 9.8, O–CHaHb–CO), 7.83 (2H,
br, NH). 13C NMR (150.9 MHz, DMSO): 14.04 (CH3), 22.28 (CH2), 28.11
(d, J(C,P)¼8.0, CH2), 28.85 (CH2), 48.25 (d, J(C,P)¼141.3, N–CH–P), 51.95
(OCH3), 61.62 (d, J(C,P)¼5.3, O–CH2), 171.43 (d, J(C,P)¼4.6, O–CO). IR
(KBr, nmax cmꢁ1) 3200–2700 br, 2647,1759,1735,1630,1536,1230,1107,
1073. HRMS (ESI) calcd for C8H19NO5P [MþH]þ 240.1001; found:
240.0996.
4.2.20.11. Methyl (2S)-2-[({[(R,S)-1-aminopentyl(hydroxy)phosphoryl]-
oxy}acetyl)amino]-3-methylbutanoate (29a). Phosphonate 29a was
prepared by hydrogenolysis of 28a (0.4 g, 0.71 mmol). Yield 158.7 mg
(66%). Colourless powder, tR¼43.3 min, G1. 1H NMR (600 MHz,
DMSO): two diasteroisomers ca. 1:1: 0.86 (3H, t, J¼7.3, CH3), 0.88 and
0.89 (6H, d, J¼7.0, CH3), 1.27 (2H, m, CH2), w1.40 (2H, m, CH2), 1.59
and 1.77 (2H, m, CH2), 2.06 (1H, m, –CHo), 3.13 (1H, m, N–CH–P), 3.65
(3H, s, OCH3), 4.21 and 4.22 (1H, dd, Jw8.0 and 6.2, N–CH–P), 4.44
(2H, d, J¼10.8, O–CH2–CO), 8.09 (2H, br, NH2), 8.52 and 8.53 (1H, br d,
Jw8.0, CO–NH). 13C NMR (150.9 MHz, DMSO): 13.96 (CH3), 18.32 and
19.14 (2ꢂCH3), 22.16 (CH2), 27.86 (d, J(C,P)¼8.1, CH2), 28.36 and 28.38
(CH2), 30.22 and 30.23 (–CHo), 47.67 (d, J(C,P)¼145.2, N–CH–P), 52.14
(OCH3), 57.59 and 57.61 (N–CH–CO), 62.95 (d, J(C,P)¼5.6, O–CH2–CO),
w170.48 (CO–NH), 171.85 (CO–O). IR (CHCl3, nmax cmꢁ1) 3420, 3200–
2700 br, 2565, 1736, 1674, 1534, 1440, 1394, 1376, 1171, 1057, 985.
HRMS (ESI) calcd for C13H27N2NaO6P [MþNa]þ 361.1504; found:
361.1498.
4.2.20.6. Methyl (2S)-2-{[(R,S)-1-aminopentyl(hydroxy)phosphoryl]-
oxy}propanoate (23b). Phosphonate 23b was prepared by hydro-
genolysis of 22b (0.4 g, 0.84 mmol). Yield 154.2 mg (73%).
Colourless powder, tR¼36.6 min, G1. 1H NMR (600 MHz, CDCl3):
mixture of diastereoisomers w3:2, only signals of the major isomer
are given: 0.90 (3H, t, J¼7.3, CH3), 1.36 (2H, m, CH2), 1.40–1.48 (2H,
m, CH2),1.51 (3H, d, J¼7.0, CH3),1.74 and 1.91 (2H, m, CH2), 3.29 (1H,
ddd, J¼13.2, 9.0 and 5.2, N–CH–P), 3.79 (3H, s, OCH3), 4.88 (1H, m,
O–CH–CO). 13C NMR (150.9 MHz, CDCl3): 15.69 (CH3), 21.80 (d,
J(C,P)¼4.7, CH3), 24.35 (CH2), 30.30 (d, J(C,P)¼8.9, CH2), 30.44 (d,
J(C,P)¼1.9, CH2), 51.24 (d, J(C,P)¼146.6, N–CH–P), 55.73 (OCH3),
72.69 (d, J(C,P)¼6.0, O–CHo), 177.70 (d, J(C,P)¼3.8, O–CO). IR (KBr,
nmax cmꢁ1) 3200–2700 br, 2608, 1755, 1739, 1638, 1532, 1439, 1380,
1212, 1162, 1072, 1057, 982. HRMS (ESI) calcd for C9H20NNaO5P
[MþNa]þ 276.0977; found: 276.0972.
4.2.20.12. Methyl (2S)-2-{[(2S)-2-{[(R,S)-1-aminopentyl(hydroxy)-
phosphoryl]oxy}propanoyl] amino}-3-methylbutanoate (29b). Phos-
phonate 29b was prepared by hydrogenolysis of 28b (0.4 g,
0.6 mmol). Yield 188.1 mg (77%). Colourless powder, tR¼44.7 min,
G1. 1H NMR (600 MHz, DMSO): two diasteroisomers ca. 1:1: 0.85
(3H, t, J¼7.3, CH3), 0.87–0.89 (6H, d, J¼7.0, CH3), 1.26 (2H, m, CH2),
1.36 and 1.41 (2H, m, CH2), 1.38 (3H, d, J¼6.8), 1.61 and 1.76 (2H, m,
CH2), 2.07 (1H, m, –CHo), 3.10 and 3.21 (1H, m, N–CH–P), 3.64 (3H,
s, OCH3), 4.17 and 4.18 (1H, dd, Jw8.0 and 6.5, N–CH–CO), 4.85 (1H,
m, O–CH–CO), 8.09 and 8.16 (2H, br, NH2), 8.35 and 8.37 (1H, br d,
Jw8.0, CO–NH). 13C NMR (150.9 MHz, DMSO): 13.91 and 13.92
(CH3), 18.45, 18.46 and 19.17 (2ꢂCH3), 20.35 and 20.38 (d,
J(C,P)¼2.0, CH3), 22.08 and 22.10 (CH2), 27.72 and 27.77 (d,
J(C,P)w5.0, CH2), 28.14 and 28.32 (d, J(C,P)¼1.8, CH2), 30.13 and
30.15 (–CHo), 47.62 (d, J(C,P)¼148.3, N–CH–P), 52.10 (OCH3), 57.72
and 57.73 (N–CH–CO), 70.34 and 70.84 (d, J(C,P)w6.0, O–CH–CO),
171.84 and 171.88 (CO–O), 172.33 and 172.43 (d, J(C,P)w3.8, CO–
NH). IR (CHCl3, nmax cmꢁ1) 3420, 3200–2700 br, 2565, 1736, 1674,
1534, 1440, 1394, 1376, 1171, 1057, 985. HRMS (ESI) calcd for
C14H29N2NaO6P [MþNa]þ 375.1661; found: 375.1655.
4.2.20.7. {[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}acetamide
25a. Phosphonate 25a was prepared by hydrogenolysis of 24a
(0.25 g, 0.56 mmol). Yield 95.1 mg (77%). Colourless powder,
tR¼21.5 min, G1. 1H NMR (600 MHz, DMSO): 0.86 (3H, t, J¼7.4, CH3),
1.27 (2H, m, CH2), 1.37 and 1.43 (2H, m, CH2), 1.60 and 1.77 (2H, m,
CH2), 3.14 (1H, m, N–CH–P), 4.29 (2H, d, J¼9.6, O–CH2–CO), 7.52 and
7.66 (2H, br, CONH2), 8.16 (2H, br, NH2). 13C NMR (150.9 MHz,
DMSO): 13.95 (CH3), 22.13 (CH2), 27.82 (d, J(C,P)¼8.4, CH2), 28.30
(CH2), 47.52 (d, J(C,P)¼146.4, N–CH–P), 67.20 (d, J(C,P)¼5.9, O–CH2),
172.11 (d, J(C,P)¼5.3, CO–NH2). IR (KBr, nmax cmꢁ1) 3383, 3238,
3205, 3200–2700 br, 2694, 2621, 2584, 1677, 1639, 1541, 1213, 1166,
1088, 1060. HRMS (ESI) calcd for C7H16N2O4P [MꢁH]þ 223.0848;
found: 223.0852.
4.2.20.8. (2S)-2-{[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}-
propionamide (25b). Phosphonate 25b was prepared by hydro-
genolysis of 24b (0.4 g, 0.87 mmol). Yield 128.2 mg (64%).
Colourless powder, tR¼29.4 min, G1. 1H NMR (600 MHz, DMSO):
0.86 (3H, t, J¼7.3, CH3),1.27 (2H, m, CH2),1.39 (2H, m, CH2),1.39 (3H,
d, J¼6.8, CH3), 1.61 and 1.77 (2H, m, CH2), 3.26 (1H, m, N–CH–P),
4.67 (1H, qd, J¼6.8(3ꢂ) and 6.6, O–CH–CO), 7.47 and 7.58 (2H, br,
CONH2), 8.28 (3H, br, NH3). 13C NMR (150.9 MHz, DMSO): 13.92
(CH3), 20.46 (d, J(C,P)¼4.0, CH3), 22.07 (CH2), 27.67 (d, J(C,P)¼9.2,
CH2), 28.00 (CH2), 47.54 (d, J(C,P)¼150.7, N–CH–P), 70.63 (d,
J(C,P)¼6.3, O–CHo), 174.31 (d, J(C,P)¼4.3, CO–NH2). IR (KBr, nmax
cmꢁ1) 3389, 3242, 3200–2700 br, 2696, 2615, 1679, 1655, 1607,
1530, 1205, 1072, 973. HRMS (ESI) calcd for C8H19N2NaO4P
[MþNa]þ 261.0980; found: 261.0975.
4.2.20.13. (2S)-2-[({[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}-
acetyl)amino]-3-methylbutanamide (31a). Phosphonate 31a was
prepared by hydrogenolysis of 30a (0.25 g, 0.46 mmol). Yield
91.5 mg (62%). Colourless powder, tR¼23.4 min, G2. 1H NMR
(600 MHz, DMSO): mixture of two diastereomers (1:1), some sig-
nals were doubled: 0.835 and 0.865 (6H, d, J¼6.8, 2ꢂCH3), 0.86 (3H,
t, J¼7.3, CH3), 1.27 (2H, m, CH2), 1.38 and 1.42 (2H, m, CH2), 1.60 and
1.77 (2H, m, CH2), 2.01 (1H, m, –CHo), 3.16 (1H, br, N–CH–P),
4.155þ4.165 (1H, dd, J¼9.2 and 6.1, N–CH–CO), 4.41 (1H, dd, J¼15.4
and 10.9, O–CHaHb–CO), 4.46 (1H, dd, J¼15.4 and 10.9, O–CHaHb–
CO), 7.14 and 7.60 (2H, d, J¼1.5, CONH2), 8.11þ8.12 (1H, d, J¼9.0, CO–
NH), 8.17 (3H, br, NH3). 13C NMR (150.9 MHz, DMSO): 13.94 (CH3),
17.90þ17.93 and 19.49þ19.52 (2ꢂCH3), 22.14þ22.15 (CH2), 27.84 (d,
J(C,P)¼8.1, CH2), 28.38 (CH2), 30.56þ30.63 (–CHo), 47.54þ47.58 (d,
J(C,P)¼146.2þ146.4, N–CH–P), 57.54þ57.56 (N–CH–CO), 63.14 (d,
J(C,P)¼5.0, O–CH–CO), 169.89þ169.91 (d, J(C,P)¼7.4þ7.2, CO–NH),
172.70 (CO–NH2). IR (KBr, nmax cmꢁ1) 3333, 3195, 3200–2700 br,
1671, 1540, 1431, 1207, 1172, 1071, 981. HRMS (ESI) calcd for
C12H26N3NaO5P [MþNa]þ 346.1508; found: 346.1502.
4.2.20.9. (2S)-2-[({[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]oxy}ac-
etyl)amino]-3-methylbutanoic acid (27a). Phosphonate 27a was
prepared by hydrogenolysis of 26a (0.3 g, 0.47 mmol). Yield 99 mg
(65%). Colourless powder, tR¼39.7 min, G1. 1H, 13C NMR and HRMS
see lit. 18. IR (KBr, nmax cmꢁ1) 3360, 3200–2700 br, 2659, 1714, 1648,
1542, 1224, 1203, 1072.
4.2.20.14. (2S)-2-{[(2S)-2-{[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]-
oxy}propanoyl]amino}-3-methylbutanamide (31b). Phosphonate 31b
was prepared by hydrogenolysis of 30b (0.25 g, 0.44 mmol). Yield
106.6 mg (71%). Colourless powder, tR¼25.7 min, G2. 1H NMR
(600 MHz, DMSO): mixture of two diastereomers (1:1), most of the
signals were doubled: 0.835þ0.866 (6H, br d, J¼6.8, 2ꢂCH3),
0.854þ0.856 (3H, t, J¼7.3, CH3), 1.26 (2H, m, CH2), 1.36 (3H, d, J¼6.8,
4.2.20.10. (2S)-2-{[(2S)-2-{[(R,S)-1-Aminopentyl(hydroxy)phosphoryl]-
oxy}propanoyl]amino}-3-methylbutanoic acid (27b). Phosphonate
27b was prepared by hydrogenolysis of 26b (0.35 g, 0.54 mmol).
Yield 101.6 mg (56%). Colourless powder, tR¼42.3 min, G1. 1H, 13C
NMR and HRMS see lit. 18. IR (KBr, nmax cmꢁ1) 3419, 3200–2700 br,
2587, 1723, 1665, 1538, 1394, 1376, 1209, 1168, 1055, 1029, 989.

6102
J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
CH3), 1.38 and 1.42 (2H, m, CH2), 1.60 and 1.76 (2H, m, CH2), 2.05
(1H, m, –CHo), 3.05þ3.13 (1H, br, N–CH–P), 4.11þ4.115 (1H, dd,
J¼9.2 and 5.9, N–CH–CO), 4.73þ4.735 (1H, p, J¼6.8 (O–CH–CO)),
7.12þ7.135 and 7.715þ7.73 (2H, d, J¼1.7, CONH2), 7.82þ7.83 (1H, d,
J¼9.2, CO–NH), 8.05þ8.085 (3H, br, NH3). 13C NMR (150.9 MHz,
DMSO): 13.92þ13.94 (CH3), 17.82þ17.83 and 19.52 (2ꢂCH3),
20.46þ20.49 (d, J(C,P)¼5.0, CH3), 22.10þ22.12 (CH2), 27.82þ27.85
(d, J(C,P)¼7.5þ8.4, CH2), 28.48þ28.58 (CH2), 30.48þ30.49 (–CHo),
47.71þ47.74 (d, J(C,P)¼147.3þ145.8, N–CH–P), 57.47þ57.48 (N–CH–
CO), 70.64þ71.07 (O–CH–N), 172.07þ172.09 (d, J(C,P)¼3.8, CO–NH),
172.96þ173.00 (CO–NH2). IR (KBr, nmax cmꢁ1) 3406, 3194, 3200–
2700 br, 1671, 1533, 1206, 1171, 1071, 982. HRMS (ESI) calcd for
C13H28N3NaO5P [MþNa]þ 360.1664; found: 360.1660.
DMF (1 mL for 5 and 20 min). The resin was thoroughly washed
with dichloromethane. Then, a mixture (2 mL) of TFA (2%), etha-
nedithiol (2%), water (1%), and triisopropylsilane (2%) in dichloro-
methane was added for 20 min at rt. The resin was filtered and
washed, and the filtrate was evaporated to dryness. A mixture
(2 mL) of TFA (95%), ethanedithiol (2%), water (1%) and triisopro-
pylsilane (2%) was added to the filtrate for 1 h at rt. The reaction
mixture was evaporated to dryness. The crude compound was
triturated repeatedly in diethyl ether. The diethyl ether was re-
moved and compound 33 purified with RP-HPLC at a flow rate of
3 mL/min on a C18 Nucleosil column (250ꢂ8 mm, 5
mm) from
Watrex (Praha) using the following gradient (solvents are the same
as described above): t¼0 min (0% B), t¼35 min (50% B), t¼40 min
(100% B), t¼41 min (0% B). Yield (20 mg, 45%, for a mixture of two
diastereoisomers). Colourless powder. HRMS (ESI) calcd for
C19H32N4O7PS [MþH]þ 491.1729; found: 419.1727.
4.2.21. {[(R,S)-1-(9H-Fluoren-9-ylmethoxycarbonylamino)-
pentyl(hydroxy)phosphoryl]oxy} acetic acid (32)
Compound 22a (1 g, 1.85 mmol) was dissolved in 80 mL of
methanol and then 10% Pd–C (0.15 g) was added, and the reaction
mixture was vigorously stirred and allowed to react at rt overnight
under an atmosphere of hydrogen (15 psi). The catalyst was filtered
off through Celite and the filter was washed with 100 mL of
methanol. The filtrate was evaporated in vacuo to give 400 mg of
crude compound 21a, which was directly used for the next reaction
without further purification. Compound 21a (0.4 g, 1.8 mmol) was
dissolved in 5 mL of saturated NaHCO3 and Fmoc-OSu (0.61 g,
1.8 mmol) in 10 mL of dioxane was added dropwise. The reaction
mixture was allowed to react at rt overnight, the dioxane was
evaporated in vacuo and the solution was acidified (pH¼1) using
1 M HCl. The separated semisolid was extracted with 2ꢂ20 mL of
ethyl acetate. The combined organic layers were dried over Na2SO4
before filtration and removal of the solvent in vacuo, which gave an
oil that was purified by reverse-phase chromatography. Yield
183.5 mg (22%). Colourless powder, tR¼46.9 min, G2. 1H NMR
(600 MHz, DMSO): 0.85 (3H, t, J¼7.1, CH3), 1.23 and 1.30 (2H, m,
CH2), 1.22 and 1.35 (2H, m, CH2), 1.55 and 1.72 (2H, m, CH2), 3.76
(1H, dtd, J¼16.0, 9.6(2ꢂ) and 3.2, N–CH–P), 4.21 (1H, m, CH (Fmoc)),
4.24 (1H, dd, J¼10.2 and 5.4, –CHaHb–O), 4.27 (1H, dd, J¼10.2 and
8.0, –CHaHb–O), 4.38 (1H, dd, J¼16.2 and 9.4, CO–CHaHb–O), 4.43
(1H, dd, J¼16.2 and 9.4, CO–CHaHb–O), 7.31 (1H, m, Ar–H), 7.33 (1H,
m, Ar–H), 7.41 (2H, m, Ar–H), 7.52 (1H, d, J¼9.6, NH), 7.72 (1H, m,
Ar–H), 7.74 (1H, m, Ar–H), 7.89 (2H, m, Ar–H). 13C NMR (150.9 MHz,
DMSO): 14.08 (CH3), 21.83 (CH2), 27.98 (d, J(C,P)¼13.4, CH2), 28.59
(d, J(C,P)¼2.3, CH2), 46.89 (pCH– (FMOC)), 48.16 (d, J(C,P)¼155.9,
N–CH–P), 61.83 (d, J(C,P)¼5.3, O–CH2), 65.98 (O–CH2), 120.35
(2ꢂAr–CH), 125.59 (Ar–CH), 125.64 (Ar–CH), 127.26 (Ar–CH), 127.31
(Ar–CH), 127.89 (2ꢂAr–CH), 140.93 (Ar–C), 140.94 (Ar–C), 143.92
(Ar–C), 144.19 (Ar–C), 156.39 (d, J(C,P)¼5.3, O–CO–N), 170.28 (d,
J(C,P)¼5.4, COOH). IR (KBr, nmax cmꢁ1) 3394, 3299, 3200–2700 br,
3042, 1725, 1537, 1451, 1295, 1243, 1214, 1098, 1033, 993, 759, 740.
HRMS (ESI) calcd for C22H25NO7P [MþNa]þ 446.1369; found:
446.1365.
Acknowledgements
This project was supported by a grant from the Grant Agency of
the Czech Republic (203/06/1405, to J.J.), by the Chemical Genetics
Consortium of the Ministry of Education, Youth and Sports of the
Czech Republic (LC060777, to J.J.) and by Research Project Z4 055
0506 of the Academy of Sciences of the Czech Republic (to IOCB).
References and notes
1. Dive, V.; Georgiadis, D.; Matziari, M.; Makaritis, A.; Beau, F.; Cuniasse, P.; Yiotakis,
A. Cell. Mol. Life Sci. 2004, 61, 2010–2019.
2. Yiotakis, A.; Georgiadis, D.; Matziari, M.; Makaritis, A.; Dive, V. Curr. Org. Chem.
2004, 8, 1135–1158.
3. Kafarski, P.; Lejczak, B. Aminophosphinic and Aminophosphonic Acids. Chem-
istry and Biological Activity. In Synthesis of Phosphono- and Phosphinopeptides;
Kukhar, V. P., Hudson, H. R., Eds.; John Wiley: Chichester, UK, 2000; pp 173–203.
4. Collinsova, M.; Jiracek, J. Curr. Med. Chem. 2000, 7, 629–647.
5. Kafarski, P.; Lejczak, B. Aminophosphinic and Aminophosphonic acids. Chem-
istry and Biological Activity. In The Biological Activity of Phosphono- and Phos-
phinopeptides; Kukhar, V. P., Hudson, H. R., Eds.; John Wiley: Chichester, UK,
2000; pp 407–442.
6. Isomura, S.; Ashley, J. A.; Wirsching, P.; Janda, K. D. Bioorg. Med. Chem. Lett.
2002, 12, 861–864.
7. Wolfenden, R.; Radzicka, A. Curr. Opin. Struct. Biol. 1991, 1, 780–787.
8. Schramm, V. L.; Horenstein, B. A.; Kline, P. C. J. Biol. Chem.1994, 269,18259–18262.
9. Phillips, M. A.; Kaplan, A. P.; Rutter, W. J.; Bartlett, P. A. Biochemistry 1992, 31,
959–963.
10. Giannousis, P. P.; Bartlett, P. A. J. Med. Chem. 1987, 30, 1603–1609.
11. Bartlett, P. A.; Giangiordano, M. A. J. Org. Chem. 1996, 61, 3433–3438.
12. Bartlett, P. A.; Hanson, J. E.; Giannousis, P. P. J. Org. Chem. 1990, 55, 6268–6274.
13. Hanson, J. E.; Kaplan, A. P.; Bartlett, P. A. Biochemistry 1989, 28, 6294–6305.
14. Song, J. B.; Hah, S. H.; Suh, J. Bull. Korean Chem. Soc. 2004, 25, 1703–1706.
15. Kim, H. D.; Lipscomb, W. N. Biochemistry 1991, 30, 8171–8180.
16. Buckman, B. O.; Emayan, K.; Islam, A.; May, K.; Bryant J.; Mohan, R.; West, C. L.;
Yuan, S. Plasma carboxypeptidase B inhibitors. U.S. and international patent
application PCT/US03/08587 (WO 03/080631 A2), 2003.
17. Bartlett, P. A.; Lamden, L. A. Bioorg. Chem. 1986, 14, 356–377.
18. Sampson, N. S.; Bartlett, P. A. Biochemistry 1991, 30, 2255–2263.
19. Morgan, B. P.; Scholtz, J. M.; Ballinger, M. D.; Zipkin, I. D.; Bartlett, P. A. J. Am.
Chem. Soc. 1991, 113, 297–307.
20. Meyer, J. H.; Bartlett, P. A. J. Am. Chem. Soc. 1998, 120, 4600–4609.
21. Ding, J. H.; Fraser, M. E.; Meyer, J. H.; Bartlett, P. A.; James, M. N. G. J. Am. Chem.
Soc. 1998, 120, 4610–4621.
22. Smith, W. W.; Bartlett, P. A. J. Am. Chem. Soc. 1998, 120, 4622–4628.
23. Yang, K. W.; Brandt, J. J.; Chatwood, L. L.; Crowder, M. W. Bioorg. Med. Chem. Lett.
2000, 10, 1085–1087.
24. Ellsworth, B. A.; Tom, N. J.; Bartlett, P. A. Chem. Biol. 1996, 3, 37–44.
25. Chang, Y. P.; Tseng, M. J.; Chu, Y. H. Anal. Biochem. 2006, 359, 63–71.
26. Bussiere, D. E.; Pratt, S. D.; Katz, L.; Severin, J. M.; Holzman, T.; Park, C. H. Mol.
Cells 1998, 2, 75–84.
27. Karanewsky, D. S.; Petrillo, E. W., Jr. Phosphonyl hydroxyacyl amino acid de-
rivatives as antihypersensives. European patent application 83303592.6 (0 097
534 A2), 1983.
28. Loots, M. J.; Karanewsky, D. S. Substituted aminoalkanoylaminoalkyl phos-
phonate angiotensin converting enzyme inhibitors. European patent applica-
tion 87305099.1 (0 249 445 A2), 1987.
29. Karanewsky, D. S. and Petrillo, E. W., Jr. Orally active phosphonyl hydroxyacyl
prolines and processes for preparing phosphonyl hydroxyacyl prolines. Euro-
pean patent application 86310043.4 (0 229 520 A1), 1986.
4.2.22. Phosphonodepsipeptide 33
Compound 32 (49 mg, 0.11 mmol) was added to a mixture of
Cys(Trt)-resin (Sieber Amide, Merck Novabiochem, 0.1 mmol) and
BOP (89 mg, 0.2 mmol) in DMF (1 mL). The reaction was initiated by
the addition of DIPEA (51 mg, 0.4 mmol). The reaction was allowed
to proceed at rt overnight and for 8 h under the same conditions.
The resin was filtered and washed, and the N-terminal Fmoc group
was cleaved off with 30% (v/v) piperidine in DMF (1 mL for 5 and
20 min). Fmoc-Tyr(OtBu) (138 mg, 0.3 mmol) was added to the
mixture of resin and BOP (0.133 g, 0.3 mmol) in DMF (1 mL). The
reaction was initiated by the addition of DIPEA (76 mg, 0.6 mmol).
The reaction was allowed to proceed at rt for 2 h and for 1 h under
the same conditions. The resin was filtered and washed, and the N-
terminal Fmoc group was cleaved off with 30% (v/v) piperidine in

J. P´ıcha et al. / Tetrahedron 65 (2009) 6090–6103
6103
30. Karanewsky, D. S.; Petrillo, E. W., Jr. Phosphonyl hydroxyacyl amino acid de-
rivatives as antihypersensitive. U.S. patent 4,703,043, 1987.
31. Campbell, D. A. Methods for the synthesis of phosphonate esters; U.S. patent
5,420,328, 2008.
48. King, A. O.; Shinkai, I. Handbook of Reagents for Organic Synthesis. Oxidizing
and Reducing Agents. In Palladium on carbon; Burke, S. D., Danheiser, R. L., Eds.;
John Wiley: Chicester, UK, 2005; pp 280–285.
49. Raillard, S. P. Handbook of Reagents for Organic Synthesis. Reagents for Gly-
coside, Nucleotide, and Peptide Synthesis In N-(9-Fluorenylmethoxy-
carbonyl)oxysuccimide; Crich, D., Ed.; John Wiley: Chichester, UK, 2005;
pp 359–360.
50. Reese, C. B.; Zhang, P. Z. J. Chem. Soc., Perkin Trans. 1 1993, 2291–2301.
51. Kudzin, Z. H.; Stec, W. J. Synthesis 1978, 469–472.
52. Hoffmann, M. J. Prakt. Chem. 1988, 330, 820–824.
53. Mucha, A.; Kafarski, P.; Plenat, F.; Cristau, H. J. Phosphorus Sulfur 1995, 105,
187–193.
32. Campbell, D. A. J. Org. Chem. 1992, 57, 6331–6335.
33. Campbell, D. A.; Bermak, J. C. J. Org. Chem. 1994, 59, 658–660.
34. Wagner, J.; Lerner, R. A.; Barbas, C. F. Bioorg. Med. Chem. 1996, 4, 901–916.
35. Campagne, J. M.; Coste, J.; Jouin, P. J. Org. Chem. 1995, 60, 5214–5223.
36. Campagne, J. M.; Coste, J.; Jouin, P. Tetrahedron Lett. 1993, 34, 6743–6744.
37. Hoffmann, M. Aust. J. Chem. 1988, 41, 605–607.
38. Galeotti, N.; Coste, J.; Bedos, P.; Jouin, P. Tetrahedron Lett. 1996, 37, 3997–3998.
39. Karanewsky, D. S.; Badia, M. C. Tetrahedron Lett. 1986, 27, 1751–1754.
40. Sampson, N. S.; Bartlett, P. A. J. Org. Chem. 1988, 53, 4500–4503.
41. Xu, J. X.; Gao, Y. H. Synthesis 2006, 783–788.
54. Mathias, L. J.; Fuller, W. D.; Nissen, D.; Goodman, M. Macromolecules 1978, 11,
534–539.
42. Campagne, J. M.; Coste, J.; Jouin, P. Tetrahedron Lett. 1995, 36, 2079–2082.
43. Picha, J.; Budesinsky, M.; Sanda, M.; Jiracek, J. Tetrahedron Lett. 2008, 49, 4366–
4368.
55. Hoffmann, M. Phosphorus Sulfur 1998, 134, 109–118.
56. Toth, I.; Anderson, G. J.; Hussain, R.; Wood, I. P.; Fernandez, E. D.; Ward, P.;
Gibbons, W. A. Tetrahedron 1992, 48, 923–930.
44. Mitra, S.; Dygas-Holz, A. M.; Jiracek, J.; Zertova, M.; Zakova, L.; Holz, R. C. Anal.
Biochem. 2006, 357, 43–49.
45. Oleksyszyn, J.; Subotkowska, L.; Mastalerz, P. Synthesis-Stuttgart 1979, 985–986.
46. Sellergren, B.; Karmalkar, R. N.; Shea, K. J. J. Org. Chem. 2000, 65, 4009–4027.
47. Yuan, C. Y.; Wang, G. H.; Chen, S. J. Synthesis 1990, 522–524.
57. Raguin, O.; Fournie-Zaluski, M. C.; Romieu, A.; Pelegrin, A.; Chatelet, F.; Pelaprat,
D.; Barbet, J.; Roques, B. P.; Gruaz-Guyon, A. Angew. Chem., Int. Ed. 2005, 44,
4058–4061.
58. Consiglio, G. A.; Failla, S.; Finocchiaro, P.; Siracusa, V. Phosphorus Sulfur 1998,
143, 159–166.