A divergent synthesis of minor groove binders with tail group variation

Article abstract of DOI:10.1039/b814452d

A new synthesis of polyamide minor groove binders in which diversity is introduced by the nucleophilic substitution of a 2-sulfido-1,3,2- diazaphospholidinyloxy substituent by volatile secondary amine nucleophiles is described. Such a method has potential

Full text of DOI:10.1039/b814452d

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
A divergent synthesis of minor groove binders with tail group variation†
David Breen, Alan R. Kennedy and Colin J. Suckling*
Received 19th August 2008, Accepted 2nd October 2008
First published as an Advance Article on the web 7th November 2008
DOI: 10.1039/b814452d
A new synthesis of polyamide minor groove binders in which diversity is introduced by the nucleophilic
substitution of a 2-sulfido-1,3,2-diazaphospholidinyloxy substituent by volatile secondary amine
nucleophiles is described. Such a method has potential value for economically investigating
structure–activity relationships in this important class of compounds through library synthesis. As an
example using this method are prepared two new minor groove binders with pyrrolidinyl or piperidinyl
tail groups that are close relatives of highly active antibacterial minor groove binders with morpholinyl
tail groups. The antibacterial activity found against Staphylococcus aureus and Mycobacterium spp.
indicates that the pKa of this set of compounds is not the dominant factor in determining the
antibacterial activity.
Introduction
Polyamide minor groove binders (MGBs) have been found to
have significant anti-infective1,2 and anticancer activity.3,4 These
Scheme 1 Concept for divergent amino tail synthesis.
compounds are composed of aromatic and heteroaromatic amino
acid amides usually with an aromatic ring at the N-terminus
and a tertiary alkylamino, amidino, or guanidine group at the
C-terminus, the so-called tail group. The importance of the pKa of
the tail group has been suggested to be of major importance in the
preparation and substitution of the diazaphosphocyclopentanes
was developed first on monomeric precursors of MGBs and simple
benzenoid analogues.
selectivity and potency of the biological activity of such MGBs.5
In most cases, the synthesis of such compounds has required the
preparation of many intermediates with the specific tail group
of interest.6 In order to investigate more rapidly the variation
of tail groups, it would be advantageous to have a method of
synthesis that inserted the amino tail group at the end of the
synthesis of the MGB. Such a method must be compatible with
critical functional groups in the MGB such as the alkene, which
has a major role in promoting antibacterial activity.2 We describe
here a method suitable for multiple parallel synthesis based upon
phosphorus chemistry that meets these requirements, together
with the biological activity of some of the products.
Chloro-1,3-dimethyl-1,3,2-diazaphosphacyclopentane, 1, was
prepared as described by Anson and McGuigan7 in 62% yield free
from P(V) impurities as shown by the single 31P NMR resonance
at 171.76 ppm. The intermediate phosphite esters, for example 2,
were obtained in 92–95% yield from the corresponding alcohol in
the presence of triethylamine; again, significant oxidation did not
occur as shown by the single 31P resonance at close to 128 ppm
(Scheme 2). The choice of substituted ethanols as substrates
is not essential for the chemistry of the reactions but derives
from the fact that aminoethyl substituted tail groups afford the
strongest antibacterially active compounds;2 this was regarded as
an essential feature for the MGB synthesis.
In accordance with the original concept (Scheme 1), the two-
step cleavage of the phosphite was attempted. Alkylation with
Synthesis
◦
methyl triflate at -75 C afforded the phosphonium salt 3; clear
The starting point was the possibility that a precursor with a
P(III) tail group might be carried through to an MGB oligomer
and the various amino tail groups introduced by a variation of
the Micahelis–Arbuzov reaction (Scheme 1). Many variations of
this strategy were tried including alkyl and aryl phosphites and
phosphoramidates. None was successful either because of poor
reactivity or more commonly because of ready oxidation from
P(III) to P(V) during several of the synthetic steps. Eventually,
the solution to this problem was found through the use of diaza-
phosphacyclopentanes, which were found to have high stability to
oxidation and to provide derivatives capable of substitution. The
evidence for alkylation at phosphorus and not on the ring nitrogen
atoms came from the new methyl resonance which appears as
a doublet due to 31P–1H coupling (dH = 1.34, J = 22.5 Hz).
Subsequent nucleophilic displacement of the phosphonium salt
unfortunately was not successful with morpholine at -78 ◦C,
-40 ◦C, 0 ◦C, and room temperature nor with pyrrolidine, in which
case the 2-phenyltetrahydro-oxazole 4 was obtained at -78 ◦C
and at room temperature together with the phosphonamidate
5 (dP = 40.93); intramolecular nucleophilic displacement by
the amide carbonyl group accounts well for this observation.
This reaction clearly demonstrates the thermodynamic driving
=
force for the C–O cleavage and P O formation planned but the
WestCHEM, Department of Pure & Applied Chemistry, University of
Strathclyde, 296 Cathedral Street, Glasgow, G1 1XL, Scotland. E-mail:
c.j.suckling@strath.ac.uk; Fax: +44 141 548 5743; Tel: +44 141 548 2271
† CCDC reference number 701644. For crystallographic data in CIF or
other electronic format see DOI: 10.1039/b814452d
substrates require modification in order to develop the MGB
synthesis. The phenylethanol derivative 6 (Scheme 3) was used
as a test substrate to avoid the intramolecular cleavage but
unfortunately, clean products were not isolable under a wide
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Scheme 2
Scheme 3
range of reaction conditions using methyl triflate followed by
pyrrolidine. The required transformation was, however, effected
with the phenylethanol derivative 6 using methyl iodide as an
alkylating agent followed by pyrrolidine. Substitution to afford the
intermediate phosphonium salt occurred in close to quantitative
yield and substitution with an excess of pyrrolidine afforded N-(2-
phenyl)ethylpyrrolidine in 28% isolated yield, over the two steps
of P–O cleavage and substitution of the intermediate alkyl iodide
by pyrrolidine. The alkyl iodide is a potential intermediate for the
synthesis of MGBs; however the risk of oxidation to iodine and
subsequent addition to alkene links in the MGBs argued for the
investigation of further possibilities.
The alternative strategy was the activation of the phosphorus
leaving group by oxidation with sulphur.8 In order to replicate
the requirements for MGB synthesis as closely as possible in
the development, the hydroxyethylamidopyrrole 8 was used as
a substrate (Scheme 4). This compound was converted into the
diazaphosphocyclopentyl ester 9 (98%) which was oxidised by
sulfur flowers to afford the P(V) intermediate 10, the structure
of which was confirmed by X-ray crystallography (Fig. 1). In the
crystal, the unit cell contains three molecules, one of which is
shown in Fig. 1.
Scheme 4
This method potentially offers several advantages over the
previous reactions described. Firstly there is no need for a
strong methylating reagent which could be problematic with
certain MGBs such as those containing thiazoles or imidazoles.
Secondly oxidation to P(V) with sulfur leads to a much more
stable intermediate, a white crystalline solid as opposed to an oil
for the previous method. A balancing disadvantage is that the
cleavage step now requires forcing conditions, the phosphorus
already being oxidised to P(V). The conversion of 10 into 12,
for example, required reaction in a sealed tube under reflux for
24–48 h with pyrrolidine as solvent (43%). This may limit the
number of nucleophiles that can be used.
Having demonstrated that suitable cleavage chemistry for
introducing the tail group was available, it was now necessary
to incorporate it into a full MGB synthesis sequence (Scheme 5).
Using the hydroxyethyl nitropyrrole derivative 9 the nitro group
was reduced in high yield to afford the corresponding aminopy-
rrole 13, but it proved impossible to acylate this amine with
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delaying the phosphorus chemistry until later in the sequence with
a protected hydroxyl group (acetate, 17) as a precursor. The target
molecule thus became the MGB acetate 17. This compound was
selected because it embodies features of some of our most active
antibacterial MGBs.2
The alkenyl head group 19 was prepared from 4-
methoxyacetophenone and methyl 6-methylnicotinate using acetic
anhydride as the solvent and a catalytic amount of zinc chlo-
ride (Scheme 6);2,9 the methyl ester was then hydrolysed under
standard basic conditions to give 19 and the required acid
chloride, 20, prepared using thionyl chloride. The pyrrolyl acetate
dimer 21 was prepared from the nitrotrichloroacetylpyrrole 22
and ethanolamine in good yield. The alcohol formed was then
protected with acetyl chloride to give 23. The nitropyrrole dimer
21 was reduced under standard hydrogenation conditions and
coupled with the acid chloride 20 to give the MGB acetate 17,
again using a standard procedure. The acetate protecting group
was hydrolysed in base to give the alcohol 24, the phosphorus tail
group was then attached followed by oxidation with sulphur to give
the key intermediate 18. Attempts to isolate and fully characterise
18 were carried out on HPLC using our standard conditions; this
however led to the hydrolysis of the phosphorus containing ring to
give25. However isolation of 18 was not necessary and substitution
with pyrrolidine and piperidine gave two target MGBs, 26 and 27,
respectively in sufficient yield for initial investigation of biological
activity in comparison with compounds previously prepared by
non-diversifying routes.2 21 forms a key intermediate in this new
procedure, as it is a readily prepared stable crystalline solid which
Fig. 1 View of one of the three crystallographically independent
molecules present in the asymmetric unit of 10. Ellipsoids are drawn at
the 50% probability level.
pyrrole acyl chlorides without complete oxidation of phosphorus
to P(V) (14) as shown by NMR. Therefore reduction and oxidation
were attempted with the P(V) thiophosphonamidate 10 previously
prepared. Reduction with palladium on charcoal and hydrogen
gas was slow and incomplete, no doubt due to the presence of
sulfur, but reduction with sodium borohydride as the hydrogen
source in the presence of palladium on carbon was successful,
with reduction complete in 25 min. A demonstration coupling
reaction with 3-nitropyrrole-2-carbonyl chloride to give 16 was
achieved, albeit in low yield (11%). With the limitations of these
reactions in mind, it was decided to prepare a complete MGB
Scheme 5
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Scheme 6
is stable on the bench for months rather than weeks, and provides
a rapid route for the preparation of a range of final products as
demonstrated.
antibacterial effects.1,2 The acetate and alcohol intermediates 17
and 24 and the by-product 25 importantly showed no antibacterial
activity, emphasising the importance of a cationic group in the pre-
sumed interaction of MGBs with the biological target DNA. On
the other hand, the pyrrolidinoethyl tail group in 26 was associated
with a minimum inhibitory concentration (MIC) of 15.6 mM
against Staphylococcus aureus, 7.8 mM against Staphylococcus
epidermis, and 15.6 mM against Mycobacterium aureum. The
corresponding values for 27 were 3.9, 1.9, and 15.6 mM. The
Biological evaluation
The value of this approach is evidenced from the biological activity
of the two test compounds 26 and 27 against Gram positive
organisms, the class of bacteria for which we observe the highest
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activity of the more potent compound 27 is comparable with
that of some of our similar antibacterial MGBs such as 28, 29,
and 30. Comparable data for 28 are 6.3 mM for S. aureus but
no activity against Mycobacterium fortuitum; similarly for 29,
activity against S. aureus was also high (3 mM) and activity against
M. fortuitum was poorer (50 mM). 30 had a similar profile to 28
and 29 (6.3 mM against S. aureus and 50 mM against M. fortuitum).
In the context of the development of antibacterial MGBs, both of
the new compounds 26 and 27 are significantly active especially
with respect to Mycobacterium aurum, which is of interest in view
of the recurrence of tuberculosis. In making these assessments,
differences in MIC greater than a factor of two can be taken
as significant and differences of a factor of ten or more highly
significant.
depends upon the structure of the rest of the MGB. For example
we have found that the dimethylaminopropyl analogue (31) of 29 is
approximately ten times less active than 29 against Staphylococcus
aureus and inactive against Mycobacterium fortuitum. For the new
compounds 26 and 27, it turns out that tail group variations within
this group of monocyclic alkyl tertiary amines do not have a major
influence on antibacterial activity; in this case, the hydrophobic
character of the N-terminal alkenyl region of the MGB plays the
dominant role in determining antibacterial activity.
Experimental
General
1H, 13C, and 31P-NMR were carried out on a Bruker DPX-
400 MHz spectrometer with chemical shifts given in ppm
(d values), relative to proton and carbon traces in solvent or in
the case of 31P, phosphoric acid was used as an external standard
(d = 0). Coupling constants are reported in Hz. IR spectra were
recorded on a Perkin Elmer, 1 FT-IR spectrometer. Elemental
analysis was carried out on a Perkin Elmer 2400, analyser series
2. Mass spectra were obtained on a Jeol JMS AX505 mass
spectrometer. Anhydrous solvents were obtained from a Puresolv
purification system, from Innovative Technologies, or purchased
as such from Aldrich. Melting points were recorded on a Reichert
hot stage microscope, and are uncorrected. Chromatography was
carried out using 200–400 mesh silica gels, or using reverse phase
HPLC chromatography on a Waters system using a C18 Luna
column with the gradient given in Table 1.
Conclusions
Although developed for specific application in minor groove
binder chemistry, the new route to aminoalkyl groups may have
applications in other fields. In relation to the biological activity,
we have discussed elsewhere the importance of the balance
of physicochemical properties in terms of hydrophobicity and
pKa that promotes antibacterial activity2,4 and others have been
concerned with the influence of tail group structure on DNA
binding.6 Lee and colleagues have found that tighter binding is
found with diaminoalkyl tail groups on the polyamide MGB.6 It
is also notable that many anticancer MGBs of this class bear
amidine or guanidine tail groups.3,4 From the point of view
of selective and strong biological activity, the basicity of the
tail group is one factor that plays an important role. Several
studies suggest that optimum antibacterial activity is found in
compounds with low pKa tail groups such as morpholinoalkyl.1,2
The most active compounds not only possess such a tail group
but also a number of hydrophobic components, in contrast to
the compounds studied by Lee and colleagues.6 The inference can
be drawn that not only is strong binding to DNA important but
also that the MGB must have suitable properties to penetrate
the bacterial cell. The influence of a specific tail group, however,
2-Chloro-1,3-dimethyl-1,3,2-diazaphospholidine 17
A
solution of N,N ¢-dimethylethylenediamine (12.10 ml,
113.4 mmol) and triethylamine (11.14 ml, 78 mmol) in dichlorome-
thane (20 ml), and a solution of phosphorus trichloride (11.88 ml,
136 mmol) in dichloromethane (20 ml) were added simultaneously
to dichloromethane (60 ml) at -40 ◦C. After addition, the solution
was allowed to warm to -30 ◦C, and a further solution of triethy-
lamine (11.14 ml, 78 mmol) in dichloromethane (20 ml) added.
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Table 1 HPLC elution gradient program for MGB purification
1,2,3-Trimethyl-1,3,2-diazaphospholidine 2-oxide 5
Obtained as a white, low-melting solid, m.p. 36–37 ◦C in 83% yield.
dH (CDCl3), 1.34 (3H, d, CH3, J = 22.5), 2.52 (3H, s, CH3), 2.56
(3H, s, CH3), 2.91 (2H, m, CH2), 3.09 (2H, m, CH2). dP (CDCl3),
40.36. LRMS: Found 149.1 (M + H) calculated for C5H13N2OP
148.0.
Time/min
A
B
Flow rate (ml/min)
0
28
33
38
40
90
30
10
90
90
10
70
90
10
10
4
4
4
4
0
2-(Iodoethyl)benzene
A = Water + 0.1% TFA; B = Acetonitrile + 0.1% TFA.
Obtained in 74% yield as an oil. nmax (NaCl): 3061, 2835, 1601,
1584, 1495, 1453, 1236, 618 cm-1. dH (CDCl3), 3.19 (2H, t, CH2,
J = 11.1), 3.38 (2H, t, CH2, J = 10.5), 7.41–7.15 (5H, m, Ar–H).
dC (CDCl3), 5.80 (CH2), 40.52 (CH2), 127.02 (2C), 128.48 (2C),
128.80, 140.77, (Ar). LRMS: Found 233.0 (M + H), 105.1 (M - I)
The solution was then allowed to return to room temperature
over 2 h. The solvent was removed, and the residue extracted with
diethyl ether (3 ¥ 100 ml). The ether fractions were combined and
the solvent removed under reduced pressure to yield the crude
product, from which the desired product was distilled, 62%, b.p.
+
calculated for C13H19N6 231.9, 105.0.
◦
60–62 C @ 1–2 mmHg. nmax (NaCl): 2933, 2451, 1030 cm-1. dH
Preparation of 1-(2-phenylethyl)pyrrolidine 7
(CDCl3), 2.70 (6H, d, 2 ¥ CH3, J = 15.9), 3.29 (4H, d, CH2, J =
7.3). dP (CDCl3), 171.76.
The above cleavage step with methyl iodide was carried out with
the diazaphospholidine 6 (1.00 g, 4.42 mmol) and iodomethane
(275 ml, 4.42 mmol), then before workup a 10 fold excess of
pyrrolidine (3.67 ml, 44.54 mmol) was added and the solution
allowed to stir overnight at room temperature. The solvent was
then removed under reduced pressure, and the residue dissolved
in dichloromethane (20 ml). The solution was then acidified with
dilute hydrochloric acid, and extracted with distilled water (2 ¥
20 ml), the dichloromethane layer was dried and the solvent
removed under reduced pressure to yield 5. The aqueous solution
was then made alkaline with saturated sodium bicarbonate
solution, and extracted with dichloromethane (2 ¥ 20 ml). The
dichloromethane fraction was then dried (Mg2SO4), and the
solvent removed under reduced pressure to yield the desired
product 7 (0.211 g, 27%). nmax (NaCl): 3063, 2788, 1603, 1584,
1496, 1454, 1235 cm-1. dH (CDCl3), 1.82 (4H, m,CH2), 2.61 (4H,
m, CH2), 3.75 (2H, m, CH2), 3.89 (2H, m, CH2), 7.40–7.12 (5H,
m, Ar-H). LRMS: Found 176.1 (M + H) calculated for C12H17N
175.1.
General coupling procedure of 2-chloro-1,3-dimethyl-1,3,2-
diazaphospholidine with an alcohol7
A solution containing the alcohol (6.5 mmol) and triethylamine
(6.5 mmol) in dichloromethane (10 ml) was added dropwise to a
solution of 1 (65 mmol) in dichloromethane (10 ml) at -60 ◦C
under nitrogen. After addition the solution was allowed to return
to room temperature over 40 min. The solution was then extracted
with saturated aqueous sodium hydrogen carbonate (40 ml), the
dichloromethane fraction was dried (Mg2SO4), the residue was
then dissolved in anhydrous diethyl ether (40 ml) filtered and
the solvent removed under reduced pressure to yield the desired
product.
N-{2-[(1,3-Dimethyl-1,3,2-diazaphospholidin-2-yl)oxy]ethyl}-
benzamide 2
Obtained in 93% yield. nmax (NaCl): 3311, 3062, 2873, 1643, 1531,
1487, 1222, 1023 cm-1. dH (CDCl3), 2.65 (3H, s, CH3), 2.69 (3H, s,
CH3), 3.03 (2H, m, CH2), 3.24 (2H, m, CH2), 3.57 (2H, q, CH2,
J = 5.4), 3.72 (2H, q, CH2, J = 5.2), 6.74 (1H, s, NH), 7.36 (2H, t,
Ar–H, J = 6.9), 7.44 (1H, t, Ar–H, J = 7.0), 7.75 (2H, m, Ar–H).
dP (CDCl3), 128.44.
N-(2-Hydroxyethyl)-1-methyl-4-nitro-1H-pyrrole-2-
carboxamide 8
2,2,2-Trichloro-1-(1-methyl-4-nitro-1H-pyrrol-2-yl)ethanone
(1.00 g, 3.70 mmol) was dissolved in dichloromethane (15 ml),
and a solution of ethanolamine (225 ml, 3.70 mmol) in
dichloromethane (5 ml) added. The solution was allowed to stir
for 1 h after which the solvent was removed to yield the p◦ roduct
8 as a pale yellow solid (0.772 g, 98%) m.p. = 158–160 C. nmax
(KBr): 3500–2500, 3358, 3268, 3141, 2947, 1644, 1566, 1313 cm-1.
dH (DMSO): 3.25 (2H, q, CH2, J = 6.0), 3.2 (2H, m, CH2), 3.90
(3H, s, NCH3), 4.69 (1H, s, O–H), 7.44 (1H, d, Ar–H, J = 1.9),
8.10 (1H, d, Ar–H, J = 1.90), 8.36 (1H, s, NH,). LRMS: Found
1,3-Dimethyl-2-(2-phenylethoxy)-1,3,2-diazaphospholidine 6
Obtained in 88% yield as an oil. nmax (NaCl): 3062, 1603, 1583,
1496, 1453, 1231, 1015 cm-1. dH (CDCl3), 2.68 (3H, s, CH3), 2.70
(3H, s, CH3), 2.89 (2H, t, CH2, J = 7.2), 3.01 (2H, m, CH2), 3.29
(2H, m, CH2), 3.88 (2H, q, CH2, J = 7.2), 7.42–7.17 (5H, m,
Ar–H). dP (CDCl3), 129.20.
+
Procedure for the cleavage step with methyl iodide
214.1 calculated for C8H12N3O4 , 214.1.
The above phenylethanol derivative 6 (1.05 g, 4.42 mmol), and
methyl iodide (277 ml, 4.42 mmol) were placed in toluene (15 ml).
The solution was heated to reflux for 4 h under nitrogen, the
solution was then allowed to cool and the solvent removed under
reduced pressure to yield the crude product. This was purified on
silica gel using 1:1 ethyl acetate:hexane, to yield the products 5 and
(2-iodoethyl)benzene.
N-{2-[(1,3-Dimethyl-1,3,2-diazaphospholidin-2-yl)oxy]ethyl}-1-
methyl-4-nitro-1H-pyrrole-2-carboxamide 9
The diazaphospholidine 1 (0.152 g, 1.00 mmol) was dissolved in
dichloromethane (15 ml) and cooled to -60 ◦C under nitrogen.
The pyrrole 8 (0.230 g, 1.00 mmol) and triethylamine (154 ml,
1.20 mmol) in dichloromethane (15 ml) were then added together
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dropwise and the solution allowed to return to room temperature
over 40 min. Sat. sodium bicarbonate solution (20 ml) and brine
(30 ml) were then added and the dichloromethane layer separated
and dried (Mg2SO4). The solvent was removed under reduced
pressure to yield the product 9 as a pale yellow oil (0.322 g, 98%).
pressure. The residue was purified by chromatography on neutral
alumina to yield the product 16 as a bright yellow solid (0.03 g,
11%) m.p. = sublimes 90 ◦C, m.p. 149–151 ◦C. nmax (KBr): 3421,
2928, 1666, 1305, 1028 cm-1. dH (DMSO): 2.50 (3H, s, N–Me),
2.53 (3H, s, NCH3), 3.06 (2H, m, CH2), 3.23 (2H, m, CH2), 3.33
(2H, m, CH2), 3.81 (3H, s, N–Me), 3.87 (2H, m, CH2), 3.95
(3H, s, N–Me), 6.87 (1H, d, Ar–H, J = 1.8 Hz), 7.24 (1H, d,
Ar–H, J = 1.8 Hz), 7.58 (1H, d, Ar–H, J = 1.8 Hz), 8.15 (1H, t,
NH, J = 5.8 Hz), 8.18 (1H, d, Ar–H, J = 1.8 Hz), 10.26 (1H, s,
NH), dP (CDCl3): 81.51, HRFABMS: Found 484.1533 calculated
for C18H27N7O5PS+, 484.1532. Found: C, 44.4; H, 5.9; N, 20.1.
C18H26N7O5PS requires C, 44.72; H, 5.4; N, 20.3%.
n
max (NaCl): 3311, 2873, 1643, 1531, 1222, 1023 cm-1. dH (CDCl3):
2.69 (3H, s, N–Me), 2.73 (3H, s, NCH3), 3.10 (2H, m, CH2), 3.31
(2H, m, CH2), 3.52 (2H, m, CH2), 3.78 (2H, m, CH2), 3.99 (3H, s,
N–Me), 7.07 (1H, d, Ar–H, J = 1.8), 7.55 (2H, m, Ar–H and NH).
dP (CDCl3): 131.26. HRFABMS: Found 330.1283; C12H21N5O4P+
requires 330.1286.
Preparation of N-{2-[(1,3-dimethyl-2-sulfido-1,3,2-
diazaphospholidin-2-yl)oxy]ethyl}-1-methyl-4-nitro-1H-
pyrrole-2-carboxamide 10
Methyl 6-[(E)-2-(4-methoxyphenyl)ethenyl]nicotinate10
4-Methoxybenzaldehyde (2.120 g, 30.40 mmol) and methyl 6-
methylnicotinate (2.305 g, 30.40 mmol) were dissolved in acetic
anhydride (10 ml); zinc chloride (0.050 g, 0.38 mmol) was then
added and the solution refluxed for 72 h. Water (10 ml) was
then added and the solution extracted with ethyl acetate (2 ¥
15 ml); the aqueous layer was then made alkaline with sat. sodium
carbonate solution and extracted with ethyl acetate (2 ¥ 15 ml).
The organic layers were combined, dried, and the solvent removed
under reduced pressure. The residue was then purified by flash
chromatography to yield the product as a brown solid (1.78 g,
23%) m.p. = 173–175 ◦C. nmax (KBr): 2950, 1717, 1595, 1290,
1254 cm-1. dH (DMSO): 3.80 (3H, s, OCH3), 3.88 (3H, s, OCH3),
The pyrrolyl diazaphospholidine 9 (0.417 g, 1.30 mmol), was
dissolved in toluene (10 ml). Sulfur (flowers 0.043 g, 1.30 mmol)
was then added and the solution allowed to stir for 15 min.
The solvent was then removed under reduced pressure and
the residue purified by flash chromatography eluting with ethyl
acetate/hexane (1:1 v/v) to yield the product 10 as a white solid
(0.460 g, 98%) m.p. = 118–120 ◦C. nmax (KBr): 3321, 2870, 1661,
1522, 1309, 1030 cm-1. dH (DMSO): 2.70 (3H, s, N–Me), 2.73
(3H, s, NCH3), 3.21 (2H, m, CH2), 3.33 (2H, m, CH2), 3.67 (2H,
m, CH2), 4.05 (3H, s, N–Me), 4.21 (2H, m, CH2), 6,89 (1H, broad s,
NH), 7.27 (1H, d, Ar–H, J = 1.8), 7.65 (1H, d, Ar–H, J = 1.8).
dP (CDCl3): 84.07. HRFABMS: Found 362.1050; C12H20N5O4PS+
requires 362.1052.
=
7.01 (1H, d, Ar–H, J = 8.8), 7.27 (1H, d, (C C–H), J = 16.0), 7.64
=
(3H, m, Ar–H), 7.80 (1H, d, (C C–H), J = 16.0), 7.01 (1H, d of
d, Ar–H, J = 2.2 and 8.2) 9.04 (1H, s, Ar–H), HRFABMS: Found
+
270.1129 calculated for C16H16NO3 , 270.1130.
1-Methyl-4-nitro-N-[2-(1-pyrrolidinyl)ethyl]-1H-pyrrole-2-
carboxamide 128
6-[(E)-2-(4-methoxyphenyl)ethenyl]nicotinic acid 1910
The pyrrolyl sulfidodiazaphospholidine 10 (0.200 g, 0.51 mmol)
was placed in a sealed thick walled Pyrex tube, pyrrolidine (5 ml)
was then added and the solution heated to 100 ◦C and allowed
to stir for 48 h, after which time the solvent was removed under
reduced pressure, and the residue purified by flash chromatography
to yield the product 12 as an off-white solid (0.133 g, 43%) m.p. =
133–135 ◦C. nmax (KBr): 3321, 2955, 1651, 1306 cm-1. dH (DMSO):
2.78 (4H, m, 2(CH2)), 3.10 (4H, m, 2(N–CH2)), 3.30 (2H, m, CH2),
3.38 (2H, m, CH2), 3.91 (3H, s, N–Me), 6.89 (1H, broad s, NH),
7.44 (1H, d, Ar–H, J = 2.0), 8.13 (1H, d, Ar–H, J = 2.0). LRMS:
Methyl 6-[(E)-2-(4-methoxyphenyl)ethenyl]nicotinate (1.058 g,
4.15 mmol) was dissolved in ethanol (3 ml). A solution of sodium
hydroxide (0.332 g, 8.30 mmol) in water (10 ml) was then added
and the solution was heated under reflux for 2 h. Water (20 ml)
was then added and the solution cooled to 0 ◦C, followed by
acidification using dilute hydrochloric acid, during which the
product precipitated as a yellow solid. (0.77 g, 73%), sublimes
at 200–202 ◦C, m.p. >230◦C. nmax (KBr): 3600–3000, 3107, 2934,
1719, 1595, 1276 cm-1. dH (DMSO): 3.80 (3H, s, OCH3), 7.03 (2H,
+
=
Found 267.2 calculated for C12H19N4O3 , 267.1. Found: C, 53.7;
d, Ar–H, J = 8.7), 7.36 (1H, d, (C C–H), J = 16.2), 7.67 (2H, d,
=
H, 6.5; N, 21.5; C12H18N4O3 requires C, 54.1; H, 6.8; N, 21.0%.
Ar–H, J = 8.7), 7.93 (1H, d, Ar–H, J = 8.4) 7.98 (1H, d, (C C–
H), J = 16.2), 8.44 (1H, d, Ar–H, J = 8.4), 9.01 (1H, d, Ar–H,
+
J = 1.7), HRFABMS: Found 256.0972 calculated for C15H14NO3 ,
N-{2-[(1,3-Dimethyl-2-sulfido-1,3,2-diazaphospholidin-2-
yl)oxy]ethyl}-1-methyl-4-{[(1-methyl-4-nitro-1H-pyrrol-2-
yl)carbonyl]amino}-1H-pyrrole-2-carboxamide 16
256.0974; Found: C, 71.0; H, 5.5; N, 5.4; C15H13NO3 requires C,
70.6; H, 5.1; N, 5.5%.
The pyrrolyl sulfidodiazaphospholidine 10 (0.200 g, 0.56 mmol)
was dissolved in dioxane (10 ml), Pd/C (10%, 0.100 g) was then
added, followed by a solution of sodium borohydride (0.040 g,
0.70 mmol) in water (5 ml). The solution was then allowed
to stir for 1 h. The solution was then filtered directly into a
flask containing 1-methyl-4-nitro-1H-pyrrole-2-carbonyl chloride
(0.104 g, 0.56 mmol). Sat. sodium carbonate solution (2 ml) was
then added and the solution allowed to stir overnight. Water (5 ml)
was added and the solution extracted with ethyl acetate (2 ¥ 20 ml);
the organic layer was dried and the solvent removed under reduced
2-{[(1-Methyl-4-nitro-1H-pyrrol-2-yl)carbonyl]amino}ethyl
acetate 23
The hydroxyethylpyrrole 8 (0.826 g, 5.42 mmol) was dissolved
◦
in dichloromethane (10 ml) and cooled to 0 C. Acetyl chloride
(386 ml, 5.42 mmol) in dichloromethane (2 ml) was then added
dropwise followed by triethylamine (550 ml, 6.50 mmol) and the
solution allowed to return to room temperature over 2 h. The
solvent was then removed under reduced pressure and the residue
purified by flash chromatography to give the product 23 as an
184 | Org. Biomol. Chem., 2009, 7, 178–186
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off-white solid (1.34 g, 97%), m.p. = 127–129 ◦C. nmax (KBr):
3410, 3110, 2952, 1723, 1661, 1308 cm-1. dH (DMSO): 2.01 (3H, s,
CH3), 3.44 (2H, q, CH2, J = 5.7), 3.91 (3H, s, NCH3), 4.10 (2H,
t, CH2, J = 5.7), 7.43 (1H, d, Ar–H, J = 2.0), 8.14 (1H, d, Ar–H,
J = 1.7), 8.54 (1H, t, NH, J = 6.5), HRFABMS: Found 256.0887
N-(5-{[(5-{[(2-Hydroxyethyl)amino]carbonyl}-1-methyl-1H-
pyrrol-3-yl)amino]carbonyl}-1-methyl-1H-pyrrol-3-yl)-6-[(E)-
2-(4-methoxyphenyl)ethenyl]nicotinamide 24
The tetramer 17 (0.100 g, 0.17 mmol) was dissolved in ethanol
(1 ml); a solution of sodium hydroxide (0.020 g, 0.5 mmol) in
water (5 ml) was then added and the solution allowed to reflux
for 2 h. The whole solution was then freeze dried and the residue
purified by HPLC to give the product as an orange solid (0.02 g,
+
calculated for C10H14N3O5 , 256.0889. Found: C, 47.1; H, 5.3; N,
16.2; C10H13N3O5 requires C, 47.1; H, 5.1; N, 16.5%.
◦
2-{[(1-Methyl-4-{[(1-methyl-4-nitro-1H-pyrrol-2-yl)carbonyl]-
amino}-1H-pyrrol-2-yl)carbonyl]amino}ethyl acetate 21
23%), m.p. >230 C, purity by HPLC = 98%. nmax (KBr): 1342,
2926, 1632, 1593, 1261 cm-1. dH (DMSO): 3.24 (2H, q, CH2, J =
5.5), 3.81 (6H, m, NCH3 and OCH3), 3.87 (3H, s, NCH3), 3.46
(2H, t, CH2, J = 5.5), 6.86 (1H, d, Ar–H, J = 1.9), 7.00 (2H, d,
Ar–H, J = 8.8), 7.07 (1H, d, Ar–H, J = 1.9), 7.18 (1H, d, Ar–H,
The acetoxyethyl pyrrole 23 (0.900 g, 3.56 mmol) was dissolved
in methanol (10 ml); Pd/C (10%, 0.450 g) was then added and
the solution was then allowed to stir under hydrogen for 3 h.
The solution was then filtered and the solvent removed under
reduced pressure. The residue was dissolved in DMF (1.5 ml) and
added to a solution of 1-methyl-4-nitro-1H-pyrrole-2-carbonyl
chloride (0.600 g, 3.56 mmol) and N-methylmorpholine (431 ml,
4.27 mmol). The solution was allowed to stir overnight, during
which time the product 21 precipitated as a bright yellow solid
which was collected by filtration (0.86 g, 63%), m.p. = 155–160 ◦C.
=
J = 1.8), 7.24 (1H, d, (C C–H), J = 16.1), 7.35 (1H, d, Ar–H,
=
J = 1.8), 7.66 (3H, m, Ar–H), 7.77 (1H, d, (C C–H), J = 16.0),
7.91 (1H, t, NH, J = 5.6), 8.25 (1H, d of d, Ar–H, J = 2.3 and
8.2), 9.06 (1H, d, Ar–H, J = 2.2), 9.93 (1H, s, NH), 10.46 (1H, s,
+
NH), HRFABMS: Found 543.2314 calculated for C31H33N6O6 ,
543.2311.
nmax (KBr): 3399, 3296, 3129, 2957, 1731, 1657, 1303 cm.-1 dH
.
1-{2-[({4-[({4-[({6-[(E)-2-(4-Methoxyphenyl)ethenyl]-3-
pyridinyl}carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)-
amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)amino]ethyl}pyrrolidi-
nium trifluoroacetate 26 and 1-{2-[({4-[({4-[({6-[(E)-2-(4-
methoxyphenyl)ethenyl]-3-pyridinyl}carbonyl)amino]-1-methyl-
1H-pyrrol-2-yl}carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}-
carbonyl)amino]ethyl}piperidinium trifluoroacetate 27
(DMSO): 2.01 (3H, s, CH3), 3.42 (2H, q, CH2, J = 5.8), 3.81
(3H, s, NCH3), 3.96 (3H, s, NCH3), 4.09 (2H, t, CH2, J = 5.8),
6.87 (1H, d, Ar–H, J = 1.9), 7.23 (1H, d, Ar–H, J = 1.8), 7.58 (1H,
d, Ar–H, J = 2.0), 8.14 (2H, m, Ar–H and NH), 10.25 (1H, s, NH),
+
HREIMS: Found 378.1365 calculated for C16H20N5O6 , 378.1369.
Found: C, 50.5; H, 5.7; N, 18.2; C16H19N5O6 requires C, 50.9; H,
5.1; N, 18.6%.
The tetramer 24 (0.100 g, 0.18 mmol) and triethylamine (40 ml,
0.36 mmol) in dichloromethane (5 ml) was added dropwise to a
solution of 1 (0.040 g, 0.22 mmol) in dichloromethane (10 ml) at
-60 ◦C under nitrogen. After addition the solution was allowed to
return to room temperature over 40 min. Sulfur (flowers 0.028 g,
0.22 mmol) was then added and the solution allowed to stir for
a further 30 min. The dichloromethane was then removed under
reduced pressure to yield an orange residue. This material was
then divided into two portions. The fractions were then refluxed in
either pyrrolidine (2 ml) or piperidine (2 ml) for 24 h, after which
the solvent was removed under reduced pressure, and the residues
purified by HPLC before being freeze dried to yield 26 and 27 as
orange solids.
2-[({4-[({4-[({6-[(E)-2-(4-Methoxyphenyl)ethenyl]-3-pyridinyl}-
carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)amino]-1-
methyl-1H-pyrrol-2-yl}carbonyl)amino]ethyl acetate 17
The alkene dimer 19 (0.067 g, 0.27 mmol) was dissolved in
dichloromethane (2 ml) and thionyl chloride (5 ml) added and the
solution refluxed for 1 h, after which the solvent was removed un-
der reduced pressure to give 6-[(E)-2-(4-methoxyphenyl)ethenyl]
nicotinoyl chloride 20 as a dark orange solid. The pyrrolyl
acetate dimer acetate 21 (0.100 g, 0.27 mmol) was dissolved in
methanol (5 ml), Pd/C, (10%, 0.050 g) was then added, the
solution was then allowed to stir under hydrogen for 3 h, the
solution was then filtered and the solvent removed under vacuum,
the residue was dissolved in DMF (1.5 ml) and added to the
previously prepared 6-[(E)-2-(4-methoxyphenyl)ethenyl]nicotinic
acid chloride; N-methylmorpholine (36 ml, 0.32 mmol) was added
and the solution allowed to stir overnight. The product was then
purified by HPLC and freeze drying the eluate afforded the pure
product as an orange solid (0.04 g, 27%) m.p. >230 ◦C, purity by
HPLC = 97%. nmax (KBr): 3446, 2925, 1637, 1594, 1260 cm.-1 dH
(DMSO): 2.02 (3H, s, CH3), 3.40 (2H, q, CH2, J = 5.7), 3.81 (6H,
m, NCH3 and OCH3), 3.88 (3H, s, NCH3), 4.09 (2H, t, CH2, J =
5.7), 6.89 (1H, d, Ar–H, J = 1.8), 7.00 (2H, d, Ar–H, J = 8.8),
7.10 (1H, d, Ar–H, J = 1.8), 7.23 (1H, d, Ar–H, J = 1.8), 7.26
1-{2-[({4-[({4-[({6-[(E)-2-(4-methoxyphenyl)ethenyl]-3-pyridinyl}-
carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)amino]-1-
methyl-1H-pyrrol-2-yl}carbonyl)amino]ethyl}pyrrolidinium
trifluoroacetate 26
◦
Obtained in 26% yield, m.p. >230 C, purity by HPLC = 98%.
nmax (KBr): 3315, 3126, 2984, 1679, 1530, 1310 cm-1. dH (DMSO):
1.88 (4H, m, 2(CH2)), 2.06 (4H, m, 2(CH2)), 3.22 (2H, q, CH2, J =
5.4), 3.38 (2H, t, CH2, J = 5.4), 3.80 (6H, m, NCH3 and OCH3),
3.89 (3H, s, NCH3), 7.02 (3H, m, Ar–H), 7.09 (1H, d, Ar–H, J =
=
1.9), 7.18 (1H, d, Ar–H, J = 1.8), 7.26 (1H, d, (C C–H), J =
=
(1H, d, (C C–H), J = 16.1), 7.37 (1H, d, Ar–H, J = 1.8), 7.66
16.0), 7.35 (1H, d, Ar–H, J = 1.8), 7.66 (3H, m, Ar–H), 7.77 (1H,
=
=
(3H, m, Ar–H), 7.77 (1H, d, (C C–H), J = 16.1), 8.14 (1H, t, NH,
d, (C C–H), J = 16.0), 7.91 (1H, t, NH, J = 5.6), 8.25 (1H, d of d,
J = 5.6), 8.29 (1H, d of d, Ar–H, J = 2.3 and 8.2 Hz), 9.07 (1H, d,
Ar–H, J = 2.3 and 8.2), 9.06 (1H, d, Ar–H, J = 2.2), 9.93 (1H, s,
NH), 10.46 (1H, s, NH). HRFABMS: Found 596.2941 calculated
for C33 H38 N7 O4, 596.2941.
Ar–H, J = 2.2), 9.98 (1H, s, NH), 10.51 (1H, s, NH). HRFABMS:
+
Found 585.2419 calculated for C31H33N6O6 , 585.2417.
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Biological data
1-{2-[({4-[({4-[({6-[(E)-2-(4-methoxyphenyl)ethenyl]-3-pyridinyl}-
carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)amino]-1-
methyl-1H-pyrrol-2-yl}carbonyl)amino]ethyl}piperidinium
trifluoroacetate 27
MIC data was obtained as described previously.2 In summary,
sample dilutions were typically prepared by dissolving the test
sample (2 mg) in sterile water (10 mL) to provide a working
concentration of 200 mg mL-1. The test wells on each 96 well
microtiter plate were initially inoculated with culture medium
(100 mL) using Mueller–Hinton broth. A solution of each test
sample (100 mL) was added to one row of each plate, and a series
of doubling dilutions was made for successive rows. Incubation
was at 37 ◦C for antibacterial assays and 25 ◦C for antifungal
assays. Plates were inspected visually for growth, and Resazurin
was added to each well; a distinct color change from blue to red
indicated that growth had occurred in an individual well. From
the observed pattern of color, the MIC was determined. All tests
included sterility and growth controls.
Obtained in 22% yield, m.p. >230 ◦C, purity by HPLC = 96%. nmax
(KBr): 3296, 3136, 2875, 1701, 1542, 1298 cm-1. dH (DMSO): 1.67
(6H, m, 2(CH2)), 1.81 (4H, m, 2(CH2)), 2.93 (2H, q, CH2, J = 5.4),
3.19 (2H, t, CH2, J = 5.4), 3.79 (6H, m, NCH3 and OCH3), 3.82
(3H, s, NCH3), 6.98 (3H, m, Ar–H), 7.10 (1H, d, Ar–H, J = 1.9),
=
7.21 (1H, d, Ar–H, J = 1.8), 7.25 (1H, d, (C C–H), J = 16.0),
7.34 (1H, d, Ar–H, J = 1.8), 7.65 (3H, m, Ar–H), 7.75 (1H, d,
=
(C C–H), J = 16.0), 8.26 (2H, m, Ar–H, NH), 9.05 (1H, d, Ar–H,
J = 2.2 Hz), 10.00 (1H, s, NH), 10.49 (1H, s, NH). HRFABMS:
+
Found 610.3093 calculated for C34H40N7O4 , 610.3097.
N-[{2-[({4-[({4-[({6-[(E)-2-(4-Methoxyphenyl)ethenyl]-3-
pyridinyl}carbonyl)amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)-
amino]-1-methyl-1H-pyrrol-2-yl}carbonyl)amino]ethoxy}-
(sulfanyl)phosphoryl]-N,N¢-dimethylmethanediaminium
bis(trifluoroacetate) 25
Acknowledgements
Crystallographic data for 10 was kindly collected by the EPSRC
National Crystallography Service, University of Southampton.
The tetramer 24 (0.100 g, 0.18 mmol) and triethylamine (40 ml,
0.36 mmol) in dichloromethane (5 ml) was added dropwise to a
solution of 1 (0.040 g, 0.22 mmol) in dichloromethane (10 ml) at
-60 ◦C under nitrogen. After addition the solution was allowed to
return to room temperature over 40 min. Sulfur (flowers 0.028 g,
0.22 mmol) was then added and the solution allowed to stir for
a further 30 min. The dichloromethane was then removed under
reduced pressure to yield an orange residue which was purified by
HPLC to yi◦eld the product as a yellow/orange solid (0.02 g, 14%),
m.p. >230 C, purity by HPLC = 94%. nmax (KBr): 3421, 3118,
2487, 1666, 1593, 1261, 1175 cm.-1 dH (DMSO): 2.53 (4H, m, CH2),
2.63 (3H, s, N+CH3), 3.17 (3H, s, N+CH3), 3.46 (4H, m, CH2), 3.79
(6H, m, NCH3 and OCH3), 3.87 (3H, s, NCH3), 6.65 (1H, m, Ar–
H), 6.88 (2H, d, Ar–H, J = 8.7), 7.09 (1H, d, Ar–H, J = 1.6), 7.18
Notes and references
1 R. W. Bu¨rli, Y. Ge, S. White, S. M. Touami, M. Taylor, J. A. Kaizerman
and H. E. Moser, Bioorg. Med. Chem. Lett., 2002, 12, 2591; R. W.
Bu¨rli, D. McMinn, J. A. Kaizerman, W. Hu, Y. Ge, Q. Pack, V. Jiang,
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Lett., 2004, 14, 1253; R. W. Bu¨rli, P. Jones, D. McMinn, Q. Le, J.-X.
Duan, J. A. Kaizerman, S. Difuntorum and H. E. Moser, Bioorg. Med.
Chem. Lett., 2004, 14, 1259; R. W. Bu¨rli, J. A. Kaizerman, J.-X. Duan,
P. Jones, K. W. Johnson, M. Iwamoto, K. Truong, W. Hu, T. Stanton,
A. Chen, S. Touami, M. Gross, V. Jiang, Y. Ge and H. E. Moser, Bioorg.
Med. Chem. Lett., 2004, 14, 2067.
2 C. J. Suckling, D. Breen, A. I. Khalaf, E. Ellis, I. S. Hunter, G. Ford,
C. G. Gemmell, N. Anthony, J.-J. Helsebeux, S. P. Mackay and R. D.
Waigh, J. Med. Chem., 2007, 50, 6116–6125.
3 A. J. Ten Tije, J. Verweij, A. Sparreboom, A. von der Gaast, C. Fowst,
F. Fiorentini, J. Tursi, A. Antonellini, M. Mantel, C. M. Hartman, G.
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Z. Mishal, L. Dassoneville, A. Kumar, C. E. Stephens, Q. Hu, W. D.
Wilson, D. W. Boykin and C. Bailly, Cancer Research, 2002, 62, 7219.
4 C. J. Suckling, Expert Opinion on Therapeutic Patents, 2004, 14, 1693.
5 C. J. Suckling, J. Phys. Org. Chem., 2008, 21, 575.
6 T. Brown, Z. Taherbhai, J. Sexton, A. Sutterfield, M. Turlington, J.
Jones, L. Stallings, M. Stewart, K. Buchmueller, H. Mackay, C. O’Hare,
J. Kluza, B. Nguyen, D. Wilson, M. Lee and J. A. Hartley, Bioorg. Med.
Chem., 2007, 15, 474.
7 C. McGuigan and M. S. Anson, J. Chem. Soc., Perkin Trans., 1989,
715.
8 R. J. M. Hermans and H. M. Buck, J. Org. Chem., 1987, 23, 5150.
9 C. A. Lipinski, J. L. LaMattina and P. J. Oates, J. Med. Chem., 1986,
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10 R. Cluzan and L. Katz, The Boots Company Ltd., 1977, US patent
4,009,174.
=
(2H, m, Ar–H and (C C–H), 7.26 (1H, d, Ar–H, J = 1.5), 7.63
=
(3H, m, Ar–H), 7.75 (1H, d, (C C–H), J = 16.0), 7.91 (1H, m,
NH), 8.25 (1H, d, Ar–H, J = 8.1), 8.58 (2H, broad s, 2(N+H)), 9.06
(1H, s, Ar–H), 9.97 (1H, s, NH), 10.40 (1H, s, NH). HRFABMS:
found 709.2688 calculated for C33H43N8O6PS+, 709.2686
Crystal data for 10: C12H20N5O4PS, Mr = 361.36, monoclinic,
space group P21/n, a = 18.0006(6), b = 10.8282(3), c =
◦
3
˚
˚
25.5950(9) A, b = 98.184(2) , V = 4938.0(3) A , Z = 12, l =
-1
˚
0.71073 A, m = 0.321 mm , T = 120 K; data quality was impaired
by generic twinning. This model was treated as twinned by a 180◦
rotation about 1 0 0. Using the SHELXL-97 HKLF 5 formalism11
gave a twin ratio of 0.852:0.148. In addition two of the three
crystallographically independent molecules show some disorder.
Final refinement to convergence on F2 gave R = 0.0904 (F, 19205
obs. data only) and Rw = 0.2004 (F2, all 28548 data), GOF =
1.085. CCDC reference numbers 701664.†
11 G. M. Sheldrick, SHELXL-97 a program for crystal structure refine-
ment, University of Go¨ttingen, Go¨ttingen, Germany, 2007.
186 | Org. Biomol. Chem., 2009, 7, 178–186
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The Royal Society of Chemistry 2009
©