Mendeleev
Communications
Mendeleev Commun., 2008, 18, 295–296
Synthesis of alafosfalin and its phosphinic
analogue and their fungicidal activity
Radii M. Khomutov,*a Tatyana I. Osipova,a Elena N. Khursa and Vitalii G. Dzhavakhiyab
a V. A. Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991 Moscow, Russian
Federation. Fax: +7 499 135 1405; e-mail: khomutov@genome.eimb.relarn.ru
b All-Russian Research Institute of Phytopathology, 143050 Golitsyno, Moscow Region, Russian Federation.
Fax: +7 496 233 409 02; e-mail: vitaly@vniif.rosmail.com
DOI: 10.1016/j.mencom.2008.11.001
A new convenient method for the synthesis of the phosphonic and phosphinic analogues of L-Ala-L-Ala has been developed and
their fungicidal activity has been demonstrated.
Alafosfalin, i.e., (R)-[1-N-(S-alanyl)aminoethylphosphonic acid]
1, has high antibacterial activity (ID50 = 0.1–4 μg cm–3 for gram-
negative bacteria†) and provides a good example of a successful
solution to the problem of transportation of compounds that
are active in vitro into a bacterial cell.1 Its phosphine analogue,
(R)-[1-N-(S-alanyl)aminoethylphosphinic acid] 2, is also efficient
against pathogenic bacteria.2 In the known synthesis of com-
pound 1, original racemic 1-aminoethylphosphonic acid 3 was
obtained from an ester of α-bromopropionic acid (four stages).3
After that, the diastereomeric salts of N-benzyloxycarbonyl
derivative 3 with (S)-methylbenzylamine were separated by
crystallisation, protection was removed, the (R)-isomer of acid
3 was acylated with an activated ester of N-benzyloxycarbonyl-
L-alanine, and compound 1 was obtained after protection removal
(four stages).4 Phosphinic analogue 2 was synthesised by a
similar procedure including the stage of isomer separation.2,5
Aiming at practical aspects of the utilisation of phosphorus
analogues of dipeptides, we developed a new synthesis of
analogues 1 and 2 starting from racemic 1-aminoethylphos-
phinic acid 4 obtained in one stage from acetaldoxime and
H3PO2.6 The key stage of the synthesis involves the separation
of diastereomeric N-benzyloxycarbonyl derivatives of dipeptide
5 by usual crystallisation from water.‡ Protection removal results
in target analogue 2,§ which was oxidised to give alafosfalin 1¶
(Scheme 1). The suggested synthetic method is based on acces-
sible phosphinate 4, excludes laborious crystallisation of salts
with chiral amines, considerably reduces the number of stages,
and allows the synthesis of analogues 1 and 2 to be performed
using a common procedure.
biosynthesis of the melanin pigment, which is a component
of the cytoderm in fungi.8 In in vitro experiments with the
Magnaporthe grisea (Hebert) Barr fungi, the originator of
rice blast, we compared the effect of (R)-phosphinate 4 and
(R)-acid 3, as well as analogues 1 and 2, on the pigmentation
and growth of mycelium under various growing conditions on
solid media in Petri dishes.†† On an agarized medium containing
no amino acids, the minimum suppressing concentrations (MSC)
‡
A mixture of compound 4 (818 mg, 7.5 mmol), NaHCO3 (1.26 g,
15 mmol) and 25% aqueous dioxane (20 ml) was stirred at 40 °C until
dissolution and cooled to 20 °C, then a solution of Z-(L)-Ala-OSu (2.4 g,
7.5 mmol) in dioxane (40 ml) was added over 30 min with stirring. The
mixture was stirred for 20 h at 20 °C, concentrated to 25 ml in vacuo,
and extracted with ethyl acetate (2×10 ml); the aqueous portion was
acidified with conc. HCl to pH 1 and kept for 12 h at +4 °C. The residue
was separated, washed with ice water (2×2.5 ml), and dried in vacuo
over KOH to give 1.62–1.75 g of a raw mixture of diastereomers 5,
mp 158–166 °C (decomp.). 155 mg of this mixture was dissolved in water
(6 ml) with stirring at 90 °C and the solution was kept for 12 h at 20 °C;
the precipitate was separated, washed with hot water (90–95 °C, 2×1 ml)
on a warm filter, and dried in vacuo over KOH to give 44 mg of com-
pound 5 [39% with respect to the (R)-isomer 4], mp 183–186 °C (decomp.)
[lit.,2 180–182 °C (decomp.)]. [a]D22 –61 (c 2, AcOH) {lit.,2 [a]D22 –61
(c 2, AcOH)}. Rf 0.66 (A; PriOH–25% NH4OH–H2O, 7:1:2).
§
Compound 5 (220 mg, 0.7 mmol) was added at +4 °C with stirring to
2 ml AcOH saturated with HBr. After keeping for 30 min at 20 °C, the
mixture was evaporated to dryness in vacuo. The residue was triturated
with diethyl ether, dissolved in 5 ml of anhydrous EtOH, and a solution
of propylene oxide in acetone was added until formation of a precipitate
stopped. After keeping for 1 h at +4 °C, the precipitate was separated,
washed with acetone and diethyl ether, and dried in vacuo over KOH to
give 106 mg (84%) of compound 2, mp 273–277 °C (decomp.) [lit.,2
276 °C (decomp.)]. [a]D22 –79 (c 2, H2O) {lit.,2 [a]D22 –80.1 (c 2, H2O)}.
Rf 0.52 (A).
It has been shown previously that the fungicidal activity of
acid 4 is due to the inhibition of the polyketide pathways of
Z-NH
¶
Compound 5 (220 mg, 0.7 mmol) was treated with AcOH saturated
PO2H2
H
Z-L-Ala-OSu
HBr /AcOH
N
with HBr similarly to the procedure described above. The residue was
triturated with diethyl ether and dissolved in water (3 ml), Br2 (0.05 ml)
was added, the mixture was washed for 30 min at 20 °C and evaporated to
dryness in vacuo. The residue was dissolved in water (5 ml) and evaporated
to dryness in vacuo; after repeating this operation, the residue was dissolved
in 5 ml of anhydrous EtOH. Alafosfalin 1 (124 mg, 90%) was isolated
similarly to the procedure described above; mp 293–297 °C (decomp.)
[lit.,4 295–297 °C (decomp.)]. [a]D22 –44.7 (c 2, H2O) {lit.,4 [a]D20 –45.4
(c 1, H2O)}. Rf 0.17 (A).
†† The (R) isomers of acids 3 and 4 were obtained using the procedures
from refs. 5 and 4, respectively. The effect of analogues on the growth
and pigmentation of the mycelium of phytopathogenic fungi Magnaporthe
grisea was studied according to ref. 7.
NH2
O
PO2H2
rac-4
5
NH2
NH2
H
N
H
N
Br2 / H2O
O
PO2H2
O
PO3H2
2
1
Scheme 1
†
ID50 represents the concentration that inhibited growth to 50% with
respect to the reference.
– 295 –
© 2008 Mendeleev Communications. All rights reserved.
Khomutov, Radii M.
