A three-component Mannich-type condensation leading to phosphinic dipeptides-extended transition state analogue inhibitors of aminopeptidases

Article abstract of DOI:10.1016/j.tetlet.2011.04.037

N-Protected α-aminoalkylphosphinic acids bearing a P-H function were found to be novel practical building blocks in three-component condensations with formaldehyde and secondary amines (amino acids). Such Mannich-type N-phosphonomethylation is a common approach for phosphorus acid derived substrates and leads to multifunctional (phosphonic/amino/carboxylic) compounds of diverse relevance. The utility of this reaction was examined for construction, in a single synthetic step, of advanced phosphinic pseudodipeptides designed to act as extended transition state analogue inhibitors of selected aminopeptidases. Phosphinomethylation of primary amino acids was less efficient and yielded mixtures of products which were separated into individual components, and their structures identified.

Full text of DOI:10.1016/j.tetlet.2011.04.037

Tetrahedron Letters 52 (2011) 3141–3145
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Tetrahedron Letters
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A three-component Mannich-type condensation leading to phosphinic
dipeptides—extended transition state analogue inhibitors of aminopeptidases
Anna Dziełak a, Małgorzata Pawełczak b, Artur Mucha a,
⇑
a
_
´
Department of Bioorganic Chemistry, Faculty of Chemistry, Wrocław University of Technology, Wybrzeze Wyspianskiego 27, 50-370 Wrocław, Poland
b Institute of Chemistry, University of Opole, Oleska 48, 45-052 Opole, Poland
a r t i c l e i n f o
a b s t r a c t
Article history:
N-Protected
a-aminoalkylphosphinic acids bearing a P–H function were found to be novel practical
Received 21 February 2011
Revised 24 March 2011
Accepted 8 April 2011
Available online 15 April 2011
building blocks in three-component condensations with formaldehyde and secondary amines (amino
acids). Such Mannich-type N-phosphonomethylation is a common approach for phosphorus acid derived
substrates and leads to multifunctional (phosphonic/amino/carboxylic) compounds of diverse relevance.
The utility of this reaction was examined for construction, in a single synthetic step, of advanced phos-
phinic pseudodipeptides designed to act as extended transition state analogue inhibitors of selected ami-
nopeptidases. Phosphinomethylation of primary amino acids was less efficient and yielded mixtures of
products which were separated into individual components, and their structures identified.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Mannich condensation
Phosphinomethylation
Aminopeptidases
Transition state inhibitors
Phosphinic dipeptide analogues have been reported to be potent
competitive inhibitors of the M17 cytosolic leucine aminopeptidase
(E.C.3.4.11.1, LAP) and the M1 microsomal alanyl aminopeptidase
(E.C.3.4.11.2, APN/CD13), the most recognized representatives of
metallo-containing exopeptidases of biomedical importance.1
These phosphorus compounds are believed to act as high energy
transition state (TS) analogues in amide hydrolysis.2 Accordingly,
the pseudodipeptides 1 containing hydrophobic P1 and P10 resi-
dues, which were linked via a –P(O)(OH)–CH2– moiety mimicking
the peptide bond, exhibited inhibition constants for LAP and APN
in the nanomolar range (Scheme 1).3 The same ligands 1 were also
found to be very effective towards Plasmodium falciparum M1 and
M17 recombinant enzymes.4 Interestingly, phosphonamidate
dipeptides 2, containing a direct nitrogen-to-phosphorus bond
[–P(O)(OH)–NH–], were predicted to be more favourable TS ana-
logues. As calculated theoretically, they should be an order of mag-
nitude more potent inhibitors of LAP than phosphinates owing to
the energy gain from a very specific hydrogen bond NHꢀ ꢀ ꢀO@C that
followed interaction of the substrate with the enzyme Leu360 upon
cleavage.3 Unfortunately, the P–N moiety adjacent to a free amino
group was discovered to be extremely labile in water (pH <11)
which excluded application of phosphonamidate compounds such
as 2 in kinetic assays.5
3, Scheme 1] as inhibitors of LAP and APN aminopeptidases. The
novel pseudodipeptidic backbone should combine the advantages
of hydrolytically stable phosphinates with the enhanced active site
interactions characteristic for phosphonamidates. Optimal hydro-
phobic residues were selected as side-chain substituents of the
modified compounds: 2-phenylethyl for the P1 position and ben-
zyl, p-hydroxybenzyl or isobutyl for P10. A simple and versatile
synthetic strategy, the three-component Mannich-type condensa-
tion of
a-aminoalkylphosphinate 4, formaldehyde and an amino
acid, was examined as a method to achieve the target compounds
(Scheme 1).
The traditional synthesis of phosphinic dipeptide analogues in-
volves a multistep process. Two building blocks bearing an appro-
priate P1 (frequently an N-protected
a-aminoalkyl-H-phosphinic
acid6) and P10 fragment (e.g., an acrylate7) are synthesized individ-
ually. In the final step, the nucleophilic phosphinate, upon activa-
tion (most commonly via silylation), adds to the double bond in a
Michael reaction.8 As this methodology does not allow simple var-
iation of the substituents, many efforts have been dedicated to the
development of alternative synthetic approaches.9 To achieve the
structures designed here, we intended to investigate a convenient,
Mannich-type, three-component condensation of a P–H compound,
formaldehyde and an amine. Presently, there is only one paper
describing a corresponding structure, which was prepared via
alkylation of an amino ester with a chloromethylphosphinic acid.10
The Mannich reaction is frequently applied for partial and exhaus-
tive phosphonomethylation11 of primary and secondary amines/
amino acids (glyphosate herbicide, antiscale agents, water soften-
ers, etc.), under acidic or basic catalysis.12 Typically, phosphorus
In the present work, we report the synthesis and validation of
extended TS analogues [–P(O)(OH)–CH2–NH–, general structure
⇑
Corresponding author. Tel.: +48 71 320 3446; fax: +48 71 320 2427.
E-mail address: artur.mucha@pwr.wroc.pl (A. Mucha).
0040-4039/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2011.04.037

3142
A. Dziełak et al. / Tetrahedron Letters 52 (2011) 3141–3145
Y
P1'
O OH
P
H
N
H2N
COOH
extended
transition state
O OH
P
X
H2N
COOH
3
P1
1a, X = CH , Y = H
2
K = 66 nM (LAP)
i
3-CC
K = 276 nM (APN)
i
O OH
P
H
H
N
1b, X = CH , Y = OH
2
K = 67 nM (LAP)
i
PG
K = 36 nM (APN)
i
2, X = NH, hydrolytically not stable
4
+ CH2O
amino acid
+
Scheme 1. The structures of the phosphinic (1) and phosphonamidate (2) lead compounds, and the target phosphinic dipeptides with a modified backbone (3). The planned
synthetic strategy to achieve the designed compounds via a three-component condensation starting from phosphinic acid 4.
acid or its esters (dialkyl phosphites), but also hypophosphorus
acid, P–H phosphines and phosphinoxides, are used. Herein, accord-
existed as two isomers (trans and cis forms of the carbamate14) as
was evident in the 31P NMR spectrum (1:0.4 ratio). This is somewhat
surprising since usually the cis isomer content is negligible. It seems
that for this compound, the presence of the N-benzyl group stabi-
lizes the cis carbamate arrangement. Indeed, after removal of the
Cbz group, both forms coalesced into a single 31P NMR signal.
ing to the best of our knowledge, N-protected
a-aminoalkyl-
H-phosphinic acids were subjected to these conditions for the first
time. Unprotected, enantiomerically pure amino acids were se-
lected as the amino components. This makes the P10 building blocks
easily accessible and stereochemically defined. It also increases the
availability and structural diversity of the target compounds, which
in turn allows extensive optimization of the P10–S10 ligand–protein
interactions.
To optimize the conditions for the condensation of 5 with L-leu-
cine (to avoid N-methylation), the reaction time was fixed for 1 h
at reflux. The condensation proceeded slowly at lower tempera-
ture, whereas prolongation of refluxing increased the complexity
of the product mixture. Similar complications were observed when
a high excess of formaldehyde was used, and only two equivalents
of CH2O proved to be the optimum amount. Typically, a water solu-
tion of formaldehyde was added in two portions to a refluxing
water/AcOH/HClconcd mixture (1:1:0.05, v/v) containing both ami-
no acid components.15 The separated product still consisted of
several compounds. After removal of the benzyloxycarbonyl pro-
tection, these components were purified by HPLC, characterized
and identified. Their structures (8–14) are presented in Scheme 3.
The phosphonic analogue of phenylglycine 8 predominated in
the HPLC amino acid fraction. This oxidized form of the phosphorus
substrate represented 25–30% of the total content. As various
N-methylated products (e.g., 10, 12 and 14, Scheme 3) were also
present, the reduction of formimines (or formiminium cations) to
the corresponding N-methyl derivatives must have accompanied
the oxidation reaction (Scheme 4). Such an observation was
reported earlier in the literature.16
The analogue of phenylglycine 5,13 easily obtained on multi-
gram scale, was selected as a model phosphinate for preliminary
optimization of the reaction conditions. The substrate was reacted
in water with excess formaldehyde and an amino acid (L-leucine or
N-benzylglycine) possessing a primary or secondary amino group,
respectively (Scheme 2). It was necessary to use an organic solvent
to achieve satisfactory solubility of the phosphorus component.
Thus, acetic acid was added for this purpose. However, its acidity
was not sufficient to ensure effective catalysis and addition of a
few drops of conc. HCl was also required. Basic catalysis was ex-
cluded to avoid racemization of the amino acid. Additionally, ester-
ification of the phosphorus component would be required to
maintain its reactivity under alkaline conditions.
Initially, the reaction was performed with a 10-fold molar excess
of formaldehyde and was heated under reflux for 5–6 h. For the ami-
no acid containing a primary amino moiety, unexpected N-methyl-
ated pseudodipeptide 6 (Scheme 2) crystallized preferentially from
the complex mixture of organophosphorus products (see below for
discussion), and was isolated in 30% yield. In contrast, phosphi-
nomethylation of secondary N-benzylglycine was not problematic
and led to the desired product. The above-mentioned conditions
afforded the pure target product 7 that crystallized after cooling,
without any work-up, in a 70% yield. Interestingly, compound 7
The N-methylation involved the amino group of both substrates
and was combined with other side-reactions. Cleavage of the
Cbz protection under acidic conditions was then evident as the
first reaction. Condensation of only two components, namely
(deprotected) phosphinic phenylglycine with formaldehyde
was also apparent. These reactions led to P-hydroxymethylated
O OH
P
O OH
O OH
P
H
N
H
N
H
N
N
COOH
P
N
COOH
Cbz
Cbz
H
Gly(N-Bzl), CH O
2
Cbz
L-Leu, CH O
2
a (30%)
a (70%)
6
5
7
Scheme 2. The products of condensation of 1-N-benzyloxycarbonylamino(phenyl)methane-H-phosphinic acid (5) with
L-leucine or N-benzylglycine and formaldehyde.
Reagents and conditions: (a) amino acid (2 equiv), CH2O (36–38% aqueous solution, 10 equiv), H2O/AcOH/HClconcd (1:1:0.05, v/v), reflux, 5 h.

A. Dziełak et al. / Tetrahedron Letters 52 (2011) 3141–3145
3143
O OH
P
O OH
P
H
N
H2N
OH
OH
R
8
9, 10
R = H or Me
L-Leu, CH O
2
5
a, b
R
N
O OH
P
HO
NH
H2N
COOH
P
O
O
P
N
R
13, 14
OH
11, 12
Scheme 3. Identification of the products of condensation of 1-N-benzyloxycarbonylamino(phenyl)methane-H-phosphinic acid (5) with a primary amino acid. Reagents and
conditions: (a) -leucine (2 equiv), CH2O (36–38% aqueous solution, 2 equiv), H2O/AcOH/HClconcd (1:1:0.05, v/v), reflux, 1 h; (b) HBr (33% solution in AcOH), room
L
temperature, 2 h, then HPLC separation.
Such a modus operandi was followed to isolate the target prod-
ucts in order to verify their inhibitory activity towards LAP and
O OH
P
H
O OH
P
OH
APN aminopeptidases.
L-Leucine, L-phenylalanine, D-phenylalanine,
L-tyrosine and N-benzylglycine were selected as amino acid sub-
redox
strates bearing a hydrophobic residue potentially interacting with
the S10 subsite. The reactions were carried out starting from the
phosphinic analogue of homophenylalanine 4 (Scheme 5) using
the optimized conditions described above. To obtain the N-methyl
dipeptides (17 and 19), phenylalanine enantiomers were reacted
with 4 and a higher excess of CH2O under harsher conditions, like
those used for N-benzylglycine. After Cbz cleavage the final prod-
ucts 15–21 were purified (e.g., full data for compound 20 are
supplied15).
The results of the inhibition of cytosolic leucine and alanyl ami-
nopeptidase are presented in Table 1. The tested pseudodipeptides
appeared to be moderate competitive inhibitors of LAP and APN.
Two compounds, 20 and 21, exhibited slow-binding kinetics. This
observation is not unexpected since for tighter binding organo-
phosphorus inhibitors a change in enzyme conformation or slow
displacement of a water molecule from the active site are usually
suggested as responsible factors.3,19 The affinity of the inhibitors
was not improved when compared to the data obtained for the
lead compounds. Apparently, the backbone elongation disturbed
the optimal binding mode by distortion of the complexation of
the phosphorus fragment by the zinc ion/ions and/or interactions
in the P10–S10 region. This might be due to the fact that binding
of phenylalanine analogues to APN was not stereospecific within
H
N
OH
H
N
- H O
2
N
Scheme 4. A redox process leading to oxidation of the H-phosphinic acid to its
phosphonic analogue, accompanied with formimine reduction to N-methyl
derivatives.
a
-aminophosphinates 9 and 10, traces of which were isolated. It
was also clearly demonstrated by separation and identification of
significant quantities (10–15% of the total yield) of the [2+2] cyclo-
addition products 13 and 14. Related eight-membered (and great-
er) rings were reported in the literature as the result of [2+2+2]
condensation of hypophosphorus acid, formaldehyde and gly-
cine,17 or ethylenediamine-N,N-diacetic acid18 (as an amino group
donor). Here, the presence of both reactive NH2 and PH functions in
the same starting molecule was the only difference. The product 14
was present as a diastereoisomeric mixture, as was evident by the
1:1 peak ratio in the 31P NMR spectrum. Attempts to synthesize the
eight-membered ring compound 13 from the unprotected
aminophosphinate and formaldehyde were not successful. The
complexity of the obtained mixture provided evidence that this
process probably occurs via a polycondensation pathway.
The required aminomethylphosphinic backbone product 11 was
also obtained and was accompanied by its N-methylated analogue
12. The tertiary amine 12 predominated when a 10-fold molar ex-
cess of formaldehyde was added (compare Scheme 2). The amount
of unwanted product was reduced significantly when the opti-
mized two equivalents of CH2O were used. Disappointingly, the
target compound constituted only roughly 10–20% of the total
mixture. Each of the pseudodipeptides was composed of two dia-
stereoisomers in 1 to 1 ratio. For 11, it was possible to separate
the individual isomers by careful collection of the HPLC fractions.
However, it became obvious that this procedure could not be rec-
ommended as a general method to synthesize pseudodipeptides
containing aminomethylphosphinic fragments. Although clean,
and giving easy to separate, structurally complex products in one
step starting from secondary amines or amino acids, the designed
three-component reaction could not be applied simply for amino
acids bearing a primary amino group. The overall yields of the de-
sired products were poor and the mixtures obtained needed to be
separated cautiously into individual compounds by HPLC.
the S10 pocket. The Ki value for the
identical to that of -Phe, as were their N-methylated forms (com-
pare 16 vs 18 and 17 vs 19). Quite surprisingly, the non-natural P10
configuration of 18 (Ki = 7.66 M) was visibly more preferred by
LAP than 16 (the form, Ki = 91.1 M).
L-Phe pseudopeptide was almost
D
l
L
l
Interestingly, the studied compounds appeared to be more po-
tent towards APN than LAP; this is typically difficult to achieve
(complexation of two zinc ions present in the active site of LAP is
much more effective than complexation of only the one of APN).
The affinity measured for both enzymes differed by 1–2 orders of
magnitude in the cases of the best ligands 15, 20 and 21 (Ki = 1–
3 lM for APN). Such activity and selectivity factors seem to be a
good starting point to improve the potency of the compounds to-
wards this individual molecular target.
In summary, a three-component condensation of N-protected
a-aminophosphinic acid, formaldehyde and a secondary amine/
amino acid was found to be an extremely convenient one-step
method for the preparation of multifunctional compounds that
contain an N–C–P–C–N system. Here, this has been applied to
synthesize phosphinic dipeptide analogues with a modified back-
bone. They have been examined as extended transition state ana-
logue inhibitors of cytosolic leucine and microsomal alanyl

3144
A. Dziełak et al. / Tetrahedron Letters 52 (2011) 3141–3145
R1
O OH
H2N
P
N
COOH
O OH
P
H
H
N
H
N
COOH
a, b
R2
+
CH2O
+
R1
Cbz
P1'
R2
1
2
15, R = H,
R
R
= i-Bu
= Bzl
(L-Leu)
1
2
2
L-Leu
L-Phe
D-Phe
L-Tyr
Gly(N-Bzl)
16, R = H,
(L-Phe)
1
17, R = Me, R = Bzl
[L-Phe(N-Me)]
(D-Phe)
1
2
2
18, R = H,
R
= Bzl
1
19, R = Me, R = Bzl
[D-Phe(N-Me)]
1
2
2
20, R = H,
R
= p-HO-Bzl (L-Tyr)
[Gly(N-Bzl)]
1
21, R = Bzl, R = H
Scheme 5. The synthesis of the designed compounds.15 Reagents and conditions: (a) amino acid (2 equiv), CH2O (36–38% aqueous solution, 2 equiv for 15, 16, 18 and 20 or
10 equiv for 17, 19 and 21), H2O/AcOH/HClconcd (1:1:0.05, v/v), reflux, (1 h for 15, 16, 18 and 20 or 5 h for 17, 19 and 21); (b) HBr (33% solution in AcOH), room temperature,
2 h, then preparative HPLC.
R.; Cuniasse, P.; Yiotakis, A.; Dive, V.; Rio, M.-C.; Basset, P.; Moras, D. J. Mol. Biol.
2001, 307, 577–686; Selkti, M.; Tomas, A.; Gaucher, J.-F.; Prangé, T.; Fournié-
Zaluski, M.-C.; Chen, H.; Roques, B.-P. Acta Crystallogr., Sect. D 2003, 59, 1200–
1205.
Table 1
Inhibition of the mammalian (porcine kidney) leucine (LAP) and alanyl aminopep-
tidases (APN) by extended TS phosphinic pseudodipeptides (for experimental details
of the assays and LAP preparation see Refs. 3 and 20).
3. Grembecka, J.; Mucha, A.; Cierpicki, T.; Kafarski, P. J. Med. Chem. 2003, 46,
2641–2655.
31P NMR
(D2O, ppm, 121.5 MHz)
Ki (lM)
4. Skinner-Adams, T. S.; Lowther, J.; Teuscher, F.; Stack, C. M.; Grembecka, J.;
Compound
Mucha, A.; Kafarski, P.; Trenholme, K. R.; Dalton, J. P.; Gardiner, D. L. J. Med.
Chem. 2007, 50, 6024–6031.
LAP
APN
5. Mucha, A.; Grembecka, J.; Cierpicki, T.; Kafarski, P. Eur. J. Org. Chem. 2003, 24,
4797–4803.
6. Baylis, E. K.; Campbell, C. D.; Dingwall, J. G. J. Chem. Soc., Perkin Trans. 1 1984,
2845–2853.
15
16
17
18
19
20
40.46 and 39.94 (1:1)a
21.54 and 21.45 (1:0.9)
21.06 and 20.93 (1:1.1)
40.38 and 39.96 (1:1)a
40.39 and 40.10 (1:0.9)a
21.05 and 20.95 (1:1)
226
2.89
9.33
19.2
9.39
17.9
91.1
41.8
7.66
17.5
33.2
7. Stetter, H.; Kuhlmann, H. Synthesis 1979, 29–30.
ˇ
8. Yiotakis, A.; Vassiliou, S.; Jirácek, J.; Dive, V. J. Org. Chem. 1996, 61, 6601–6605;
4.59 (initial)b
Yiotakis, A.; Georgiadis, D.; Matziari, M.; Makaritis, A.; Dive, V. Curr. Org. Chem.
2004, 8, 1135–1158. and references cited therein.
1.06 (steady state)
21
21.00
52.4
3.11 (initial)b
9. For selected examples, see: Matziari, M.; Georgiadis, D.; Dive, V.; Yiotakis, A.
Org. Lett. 2001, 3, 659–662; Makaritis, A.; Georgiadis, D.; Dive, V.; Yiotakis, A.
Chem. Eur. J. 2003, 9, 2079–2094; Matziari, M.; Beau, F.; Cuniasse, P.; Dive, V.;
Yiotakis, A. J. Med. Chem. 2004, 47, 325–336; Matziari, M.; Yiotakis, A. Org. Lett.
2005, 7, 4049–4052; Vassiliou, S.; Xeilari, M.; Yiotakis, A.; Grembecka, J.;
Pawełczak, M.; Kafarski, P.; Mucha, A. Bioorg. Med. Chem. 2007, 15, 3187–3200.
2.90 (steady state)
a
NaOD was added to achieve solubility.
Slow binding mechanism of type A.
b
_
10. Kiss, T.; Farkas, E.; Jezowska-Bojczuk, M.; Kozłowski, H.; Kowalik, E. J. Chem.
Soc., Dalton Trans. 1990, 377–379.
aminopeptidases. The most interesting aspects of the structure–
activity relationship concerned their low micromolar affinity to-
wards monozinc APN with significantly poorer affinity to LAP.
Increasing the scope of this synthetic method to a wider variety
of amine substrates could be profitable in other fields of study, for
example in the construction of novel metal complexing agents or
multifunctional molecular receptors. Disappointingly, the applica-
tion of primary amino acids led to complicated mixtures of
products under standard conditions. Their separation and identifi-
cation provided confirmation of the occurrence of side-reactions.
These involve random combination of phosphine/imine redox pro-
cesses, leading to phosphinate P–H oxidation and N-methylation,
with N-deprotection of the phosphorus substrate and different
variants of polycondensation. Introduction of easily accessible
N-benzyl amino acids might be suggested as a suitable protecting
group strategy (since it may be removed simultaneously with
Cbz) in an alternative synthetic pathway leading to the target
compounds.
11. Moedritzer, K.; Irani, R. R. J. Org. Chem. 1966, 31, 1603–1607.
12. Aminophosphonic and Aminophosphinic Acids. Chemistry and Biological Activity;
Kukhar, V. P., Hudson, H. R., Eds.; John Wiley & Sons: Chichester, 2000.
13. Mucha, A.; Kafarski, P.; Plenat, F.; Cristau, H.-J. Phosphorus, Sulfur, Silicon 1995,
105, 187–193.
14. Hirschmann, R.; Yager, K. M.; Taylor, C. M.; Witherington, J.; Sprengeler, P. A.;
Phillips, B. W.; Moore, W.; Smith, A. B., III J. Am. Chem. Soc. 1997, 119, 8177–
8190.
15. A representative phosphinomethylation procedure of a primary amino acid with
1-(N-benzyloxycarbonylamino)alkane-H-phosphinic acid and formaldehyde,
and the separation of products.
A phosphinic acid (3.0 mmol) and an amino acid (9.0 mmol) were dissolved
in a hot H2O/AcOH/HClconcd mixture (10:10:0.5 mL). Formaldehyde (36–38%
aqueous solution, 6.0 mmol, 0.5 mL) was added in two portions to the stirred
solution. Following addition the mixture was refluxed for 1 h and then cooled
to room temperature. The solvents were evaporated to ca. 20–25% of the
starting volume. The solid was precipitated by addition of H2O (50 mL),
filtered and dried in the air. The Cbz protection was removed by the action of
HBr (33% solution in AcOH, 10 mL per 1 g) for 2 h at room temperature. The
acids were removed under reduced pressure and the residue was triturated
with Et2O. The resulting solid was filtered, dissolved in a small quantity of
H2O/MeCN and subjected to preparative HPLC (Varian ProStar apparatus with
ProStar 325 UV/vis detector using a Dynamax 250 ꢁ 21.4 Microsorb 300-10
C18 column).
2-(S)-N-{[(1’-(R,S)-amino-3’-phenylpropyl)(hydroxy)phosphinyl]methyl}amino-3-
(p-hydroxyphenyl)-propionic acid (20). 1H NMR (ppm, D2O, 300.1 MHz): 7.11
(m, 5H, C6H5), 6.91 and 6.64 (2 ꢁ d, J = 8.0 Hz, 2H and 2H, C6H4), 4.09 (m, 1H,
NCHCO), 3.14 (m, 1H, CHP), 3.02 (m, 4H, PCH2 and p-HO–C6H4CH2), 2.63 and
2.50 (2 ꢁ m, 1H and 1H, CH2C6H5), 1.95 and 1.80 (2 ꢁ m, 1H and 1H,
CHCH2CH2). 31P NMR (ppm, D2O, 121.5 MHz): 21.05 and 20.95 (1:1). 13C NMR
(ppm, D2O + NaOD, 151.0 MHz): 181.82 and 181.91 (C@O), 164.52 (C4, C6H4),
142.48 (C1, C6H5), 130.41 and 130.39 (C2, C6H4), 128.71 (C2, C6H5), 128.66
(C3, C6H5), 126.08 (C4, C6H5), 124.01 (C1, C6H4), 118.61 (C3, C6H4), 67.91 and
67.82 (2 ꢁ d, JP = 9.1 Hz, NCHCO), 49.12 and 49.02 (2 ꢁ d, JP = 99.7 Hz and
98.2 Hz, CHP), 45.17 and 45.07 (2 ꢁ d, JP = 95.1 Hz and 99.7 Hz, PCH2), 38.22
and 38.14 (p-HO–C6H4CH2), 32.21 (d, JP = 12.1 Hz, CH2C6H5), 31.98 (d,
JP = 6.0 Hz, CHCH2CH2). HRMS (ESI) m/z: [M+H]+ calcd for C19H26N2O5P:
393.1579, observed: 393.1539. HPLC purification (v/v, H2O/MeCN, both
containing 0.1% of trifluoroacetic acid): gradient 85:15 (0 min)?75:25
(35 min), tr = 27.2 min.
Acknowledgement
This work was supported by a grant from the Polish Ministry of
Science and Higher Education (N N302 159937).
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3145
Phosphinomethylation of
a
A
(3.0 mmol) and a secondary amino acid (6.0 mmol) were dissolved in a hot
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solution, 30.0 mmol, 3.0 mL) was added dropwise to the stirred solution.
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temperature. The precipitated solid was filtered and dried in the air. The
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