A total synthesis of the marine alkaloid ningalin B from (S)-proline

Article abstract of DOI:10.1071/CH09158

The title alkaloid 1 has been synthesized from (S)-proline (2) using a sequence of reactions involving oxidative bromination (of 2), N-alkylation, SuzukiMiyaura cross-coupling, and bromination steps. CSIRO 2009.

Full text of DOI:10.1071/CH09158

RESEARCH FRONT
Full Paper
CSIRO PUBLISHING
www.publish.csiro.au/journals/ajc
Aust. J. Chem. 2009, 62, 683–691
A Total Synthesis of the Marine Alkaloid Ningalin B
from (S)-Proline
Katrin Hasse,A Anthony C. Willis,A and Martin G. BanwellA,B
AResearch School of Chemistry, Institute of Advanced Studies, The Australian National University,
Canberra, ACT 0200, Australia.
BCorresponding author. Email: mgb@rsc.anu.edu.au
The title alkaloid 1 has been synthesized from (S)-proline (2) using a sequence of reactions involving oxidative bromination
(of 2), N-alkylation, Suzuki–Miyaura cross-coupling, and bromination steps.
Manuscript received: 16 March 2009.
Final version: 12 May 2009.
HO
HO
OH
Introduction
D
C
The marine alkaloid ningalin B (1, Fig. 1) was isolated, together
with three related natural products, by Fenical et al. from an
ascidian belonging to the genus Didemnum and collected at the
Ningaloo Reef in Western Australia.[1] Its structure was first
established by detailed NMR analyses and then proved through a
totalsynthesiscarriedoutbyBogerandcoworkers(seebelow).[2]
Biogenetically speaking, the compound is probably derived via a
series of oxidative coupling and condensation reactions involv-
ing the amino acid 3,4-dihydroxyphenylalanine (DOPA). These
same sorts of processes are thought to lead to structurally related
alkaloids such as the lamellarins, the lukianols, the polycitrins,
and the storniamides, all of which have attracted considerable
attention because of their interesting biological properties.[3]
Although ningalin B may be implicated in the pronounced metal-
sesquestering properties of the producing organism,[1] it does
not appear to display many other notable biological effects. It
is certainly not particularly cytotoxic, at least as determined
by its evaluation in a cancer cell-line panel.[2] The permethyl
ether of ningalin B is similarly non-cytotoxic but it and sev-
eral amide derivatives do show a significant capacity to reverse
multidrug resistance by, for example, resensitizing a resistant
human colon cancer cell line HCT116/VM46 to vinblastine and
doxorubicin.[2,4,5] As such, this ningalin B derivative behaves
in a similar fashion to certain of the lamellarins and their
congeners.[3]
HO
B
O
A
N
O
E
HO
OH
1
Fig. 1.
thus requiring that a decarboxylation reaction is carried out
at a relatively late stage in the synthesis. As with all but
one of the other syntheses, Iwao’s reaction sequence provides
ningalin B permethyl ether as the immediate precursor to the
natural product 1.
The Bullington synthesis of ningalin B[7] is quite distinct in
that a symmetrical intermediate is not involved and no decar-
boxylation step is required. It is also particularly concise and
starts with a base-promoted [2 + 3] ‘cycloaddition’ reaction
between the relevant α-cyanostilbene and methyl isocyano-
acetate to give a trisubstituted pyrrole that is readily N-alkylated,
demethylated, and lactonized to give the target compound.
Gupton’s synthesis of ningalin B permethyl ether involves
the application of his versatile vinylogous iminium salt deriva-
tive methodology for forming substituted pyrroles.[8] As with
the Bullington approach, symmetrical intermediates are not
involved and the starting material is the readily prepared 1,2-
diarylethanone derivative desoxyveratroin. The final step in the
reaction sequence is a ‘Steglich-type’lead tetraacetate-mediated
lactonization[9] of a 3-arylpyrrole-2-carboxylic acid derivative.
The Steglich synthesis of ningalin B, which was reported in
2006,[9] is modelled on the proposed biogenesis of the natural
product and also delivers a symmetrical tetrasubstituted pyrrole
Ningalin B has been the subject of several synthetic stud-
ies, the first of which was reported by Boger et al.[2] in
2000. This group employed a heterocyclic azadiene Diels–Alder
cycloaddition/zinc-metal induced ring-contraction protocol to
construct a tetrasubstituted pyrrole from a tolan precursor. Var-
ious manipulations of this pyrrole, including those involving
N-alkylation, lactonization and decarboxylation steps, provided
the permethyl ether of ningalin B. Exhaustive demethylation of
this last compound using boron tribromide then gave, in 98%
yield, ningalin B itself.
Iwao’s synthesis of target 1, reported in 2003,[6] has some
similarities to Boger’s approach[2] in that a symmetrical 2,5-
dicarboalkoxy-substituted pyrrole intermediate is involved and
© CSIRO 2009
10.1071/CH09158
0004-9425/09/070683

RESEARCH FRONT
684
K. Hasse, A. C. Willis, and M. G. Banwell
(i) SOCl2/MeOH
(ii) (Boc)2O
Br
that requires a late-stage decarboxylation. An especially attrac-
tive feature of this work is the use of a lead tetraacetate-mediated
process to establish the lactone moiety within target 1.
Our continuing interest in the chemistry of pyrroles and in the
synthesis of marine alkaloids containing this ring system[10,11]
has prompted us to establish a synthesis of ningalin B which
we now report. Our approach uses (S)-proline as the precursor
to the pyrrole ring of target 1 and avoids the need to employ
a decarboxylation step in the synthetic sequence. Details are
reported in the following sections.
NBS
CO2H
CO2Me
CO2Me
N
H
N
N
Boc
Boc
2
3
4
ZnBr2
Br
Br
CO2Me
N
H
CO2Me
N
5
DIAD,
Ph3P
ϩ
Results and Discussion
OH
MeO
The early stages of our synthesis of ningalin B are shown in
Scheme 1 and involve the conversion of (S)-proline 2 into the 3-
bromo-2-carbomethoxypyrrole 4 using a novel oxidative bromi-
nation protocol first described by Easton et al.[12] and recently
refined by us.[11] The substrate 3 required for this key step
was prepared by converting acid 2 into the corresponding and
well-known[11] methyl ester using thionyl chloride/methanol.
Boc-protection of the secondary amine residue within this last
compound was achieved under standard conditions and the ensu-
ing carbamate 3 treated with N-bromosuccinimide (NBS) in
refluxing carbon tetrachloride. By such means, compound 4 was
obtained in 72% yield[11] and the Boc-group associated with it
was readily cleaved with zinc bromide to give the pivotal pyrrole
5 (89%), the structure of which had been confirmed by a single-
crystal X-ray analysis carried out during our earlier studies.[11]
The N-alkylation of compound 5 with the β-phenethyl alco-
hol 6 was readily achieved under Mitsunobu conditions using
diisopropyl azodicarboxylate (DIAD) and so gave the previ-
ously reported[11] compound 7 in 75% yield. All the spectral
data acquired on pyrrole 7 were in accord with the assigned
structure. In particular, the electrospray ionization (ESI) mass
spectrum of this material displayed molecular associated ions at
m/z 390 and 392 (corresponding to [M + Na]+) while the 13C
NMR spectrum displayed the expected 16 signals, 10 of which
appeared in the aromatic region.
OMe
MeO
7
OMe
6
Scheme 1.
O
B
O
OH
8
X
O
ϩ
Pd[0],
K2CO3
N
Br
O
CO2Me
N
MeO
OMe
MeO
NBS,
DMF
9
X ϭ H
OMe
10 X ϭ Br
7
Scheme 2.
The planned elaboration of pyrrole 7 to ningalin B was
explored through the model study outlined in Scheme 2. Thus,
the former compound was subjected to a Suzuki–Miyaura cross-
coupling, under standard conditions, with the commercially
available aryl boronic acid 8[13] incorporating an o-hydroxy
group that it was expected would participate, after the C–C bond-
forming process, in a spontaneous lactonization reaction with
the ester residue originally present in substrate 7. In accord with
such expectations, the product of the cross-coupling process was
lactone 9 and this was obtained in 79% yield.
C11
C10
C9
C12
C13
Br14
C4
C8
O7
C3
C2
C5
C6
N1
C16
O15
The reaction of compound 9 with ca. two molar equivalents
of NBS in DMF at 0◦C provided a single mono-brominated
derivative in 74% yield and while it was expected, on electronic
grounds, to be the required one, namely compound 10, this could
be readily proved by single-crystal X-ray analysis. The derived
ORTEP is shown in Fig. 2 and this clearly reveals that the desired
bromopyrrole 10 had been obtained. The acquisition of com-
pound 10, incorporating a bromine in the correct position for a
second Suzuki–Miyaura cross-coupling that would allow instal-
lation of the isolated aryl unit of ningalin B, was a pleasing out-
come and the adaptation of this chemistry to the synthesis of the
natural product was thus pursued as described in the following
sections.
C17
C18
C23
C22
C19
C20
C25
C21
O24
O26
C27
Fig. 2. Molecular structure of compound 10 (C21H14BrNO4) with
labelling of selected atoms. Anisotropic displacement ellipsoids show 30%
probability levels. Hydrogen atoms are drawn as circles with small radii.

RESEARCH FRONT
Total Synthesis of Ningalin B
685
The completion of the synthesis of ningalin B from
(DHP) in the presence of an acid catalyst.[14] Bromination of
compound 12 using NBS in DMF proceeded smoothly to give
the required bromide 13[14] in 83% yield over the two steps.
Reaction of a mixture of halide 13 and tri-isopropylborate with
n-BuLifollowedbyacidicworkupthenaffordedHandy’sboronic
acid 14, whichwasimmediatelyesterifiedusingpinacol. Bysuch
means, the ester 15 was obtained as a light-yellow oil and in 69%
overall yield from precursor 13. The electron-impact (EI) mass
spectrum of compound 15 showed a molecular ion at m/z 280
while the 1H NMR spectrum showed two one-proton singlets in
the aromatic region (δ 6.99 and 6.44), two three-proton singlets
at δ 3.86 and 3.84 and a 12-proton singlet at δ 1.21, as would be
expected for the assigned structure.
With compounds 7 and 15 in hand, these were subjected to
a Suzuki–Miyaura cross-coupling process (Scheme 4) and this
provided the anticipated lactone 16, albeit in only 43% yield.The
NMR spectral data obtained on this product were completely
consistent with the assigned structure but final confirmation of
this followed from a single-crystal X-ray analysis. The derived
ORTEP is shown in Fig. 3.
compound
7 required securing quantities of the 4,5-
dimethoxysubstituted derivative of aryl boronate 8 or an equiv-
alent thereof. Handy et al. have described[14] the preparation of
such a species, namely 2-hydroxy-4,5-dimethoxyphenyl boronic
acid, so we sought to obtain this compound by the reported
means, then use it in a Suzuki–Miyaura cross-coupling reac-
tion with compound 7. In our hands, however, this boronic acid
proved unstable and was, therefore, difficult to purify. Accord-
ingly, we pursued the synthesis of the corresponding pinacolato
derivative using a modification of Handy’s protocols.[14] The
reaction sequence used is shown in Scheme 3 and starts with the
conversion, under standard conditions, of commercially avail-
able 3,4-dimethoxyphenol 11 into the corresponding THP ether
12 by reacting the former compound with 3,4-dihydro-2H-pyran
OMe
OMe
OMe
NBS,
DMF
DHP,
Hϩ
OMe
OMe
OMe
Br
Various modifications to the procedure used for the conver-
sion 7 + 15 → 16 failed to provide a better yield of the coupling
OH
OTHP
OTHP
11
12
13
BuLi, B(OiPr)3
then H3Oϩ
Br
OMe
CO2Me
N
OMe
OMe
OMe
Pinacol,
Et2O
OMe
OMe
ϩ
O
B
O
B
O
OH
(HO)2B
O
OH
OH
MeO
OMe
15
14
7
15
Scheme 3.
Pd[0],
Cs2CO3
MeO
OMe
C28
O27
O29
X
C30
MeO
MeO
C10
O
ϩ
C11
C12
N
O
B(OH)2
C9
C8
18
C13
C7
C6
C5
Pd[0],
K2CO3
O1
C2
MeO
C3
OMe
N4
16 X ϭ H
17 X ϭ Br
NBS,
DMF
O14
C15
HO
MeO
HO
MeO
OH
OMe
C16
C17
HO
MeO
C22
C21
C18
C19
O
O
N
O
N
C24
O
BBr3
C20
O23
O25
C26
HO
MeO
OH
OMe
19
Fig. 3. Molecular structure of compound 16 (C23H23NO6) with labelling
of selected atoms.Anisotropic displacement ellipsoids show 30% probability
levels. Hydrogen atoms are drawn as circles with small radii.
1
Scheme 4.

RESEARCH FRONT
686
K. Hasse, A. C. Willis, and M. G. Banwell
OMe
Br
product. Nevertheless, sufficient quantities of compound 16
could be accumulated by this means to allow completion of
the synthesis of ningalin B. Towards such ends, this material
was brominated with NBS in DMF and the ensuing bromide 17
(97%) was then cross-coupled with the commercially available
arylboronic acid 18[13] in the presence of potassium carbonate
and tetra-n-butylammonium bromide (TBAB) to give the previ-
ously reported[2] ningalin B permethyl ether 19 in 69% yield.
Following a protocol established by Boger et al.,[2] treatment of
this last compound with boron tribromide in dichloromethane at
−78◦ to 18◦C gave ningalin B 1 in 96% yield.
OMe
ϩ
CO2Me
O
B
N
Boc
O
OH
4
15
Pd[0],
Cs2CO3
MeO
OMe
OH
The 13C and 1H NMR spectral data derived from compounds
19 and 1 (Tables 1 and 2, respectively) compare very favourably
with those reported previously. The slight variations between the
carbon chemical shifts recorded for naturally derived ningalin B
and those arising from the synthetic material can be attributed
to the differing sample concentrations and solvent systems used
to record the cited data.
ϩ
O
MeO
OMe
N
X
O
20 X ϭ Boc
21 X ϭ H
6
ZnBr2
DIAD,
Ph3P
MeO
In an effort to establish an improved synthesis of ningalin B,
we investigated alternate points in the abovementioned reaction
sequence at which the pivotal Suzuki–Miyaura cross-coupling
reaction involving compound 15 could be carried out. In
particular, the linking of substrates 4 and 15 by such means was
studied (Scheme 5) and this allowed the preparation of lactone
20 in 55% yield. This was accompanied by ∼14% of the free
pyrrole 21. Removal of the Boc group within compound 20 was
readily achieved in 84% yield using zinc bromide and the result-
ing pyrrole 21 was N-alkylated with alcohol 6, under the same
conditions used earlier to produce compound 7, thus affording
lactone 16 in 54% yield. Despite these efforts, the yield of com-
pound 16 obtained using the route shown in Scheme 5 is only
marginally better than that observed earlier (33% versus 27%).
OMe
O
N
O
MeO
OMe
16
Scheme 5.
Experimental
General Procedures
Summary and Conclusions
A comparison of the key features of all the reported synthe-
ses of ningalin B, including the one detailed herein, is provided
in Table 3. The Steglich[9] and Bullington[7] syntheses are the
shortest (each involves just four steps) and particularly notable
for their efficiency. The Steglich synthesis[9] is remarkable for
the rapidity with which the basic framework of the target com-
poundisassembledbuttheBullingtonapproachislikelytobethe
more effective for preparing analogues of ningalin B in which the
substituents on the C and D rings are different. It is also worth
noting that the Bullington,[7] Boger,[2] and Gupton[8] synthe-
ses share a common sequence of ring-forming events (Table 3)
and that this is related to the one used by Steglich and cowork-
ers. These involve a mid-stage pyrrole-forming step while the
construction of the B-ring lactone is left to the end of the syn-
thetic sequence. In contrast, the synthesis of target 1 reported
herein starts with an intact pyrrole and uses this ring system as
a template for introducing the remaining structural components
of ningalin B. Although the present approach is not as efficient
as some of the others described to date, it should be particularly
useful for the preparation of ningalin B analogues in which the
substitution patterns associated with the C, D, and E rings are
quite different from one another.
Unless otherwise specified, 1H and 13C NMR spectra were
recorded at 18◦C in base-filtered CDCl3 on a Varian Mercury
or Inova 300 spectrometer operating at 300 MHz for proton and
75 MHz for carbon nuclei. In certain cases, a Varian Inova 500
spectrometer, operating at 500 MHz for proton and 125 MHz
for carbon nuclei, was used. For 1H NMR spectra, signals aris-
ing from the residual protio-forms of the solvent were used as
1
the internal standards. H NMR data are recorded as follows:
chemical shift (δH) (multiplicity, coupling constant(s) J (Hz),
relative integral) where multiplicity is defined as: s = singlet;
d = doublet; t = triplet; q = quartet; m = multiplet or combina-
tions of the above. The residual CHCl3 peak (δ 7.26), residual
DMSO peak (δ 2.50) and the residual MeOH peak (δ 3.30)
were used as references for 1H NMR spectra. The central peak
(δ 77.0) of the CDCl3 ‘triplet’ and the central peak (δ 39.5)
of the [D6]DMSO ‘heptet’ were used as references for proton-
decoupled 13C NMR spectra. The data for 13C NMR spectra are
given as: chemical shift (δC), (protonicity), where protonicity is
defined as: C = quaternary; CH = methine; CH2 = methylene;
CH3 = methyl.AssignmentsofsignalsobservedinvariousNMR
spectra were often assisted by conducting Attached Proton Test
(APT) experiments. Infrared spectra (νmax) were recorded on a
Perkin–Elmer 1800 Series Fourier-transform (FT)-IR Spectro-
meter. Samples were analysed as thin films on NaCl plates.AVG
FisonsAutoSpec mass spectrometer was used to obtain low- and
high-resolution EI mass spectra. Low- and high-resolution ESI
mass spectra were obtained on aVG Quattro II triple-quadrupole
Work directed towards exploitation of the present protocols
for the preparation of biologically active analogues of ningalin B,
especially permethyl ethers related to compound 19,[2] is now
under way in these laboratories. Results will be reported in due
course.

RESEARCH FRONT
Total Synthesis of Ningalin B
687
Table 1. Comparison of the 13C NMR data recorded for naturally occurring ningalin B (1), synthetically derived 1, and
permethyl ether 19
δC
Natural 1A
Synthetic 1
(present work)B
Synthetic 1
(Boger work)C
Synthetic 19
(present work)D
Synthetic 19
(Boger work)E
Synthetic 19
(Gupton work)F
156.0
146.8
146.0
145.9
145.8
145.5
144.4
142.9
133.5
130.2
127.6
126.4
120.6
120.0
117.7
116.9
116.4
116.1
114.8
112.7
110.6
109.3
104.0
50.9
155.2
146.4
145.6
145.5
145.4
145.1
144.2
142.5
133.1
129.6
126.8
125.8
121.2
120.1
119.6
117.4
116.7
116.2
116.0
114.4
110.1
109.0
103.9
50.4
155.0
146.2
145.4
145.3
145.2
144.9
144.0
142.3
133.0
129.4
126.6
125.6
121.0
119.9
119.4
117.2
116.4
116.0
115.8
114.2
109.9
108.8
103.7
50.2
155.7
149.2
149.1
149.0
148.7
148.0
146.4
145.8
132.2
130.8
127.4
126.9
122.3
121.1
119.3
115.1
113.2
112.2
111.4
111.2
110.7
104.9
100.8
56.4
155.7
149.2
149.1
149.0
148.7
148.0
146.4
145.8
132.1
130.7
127.4
126.9
122.3
121.1
119.3
115.1
113.2
112.3
111.4
111.2
110.6
104.9
100.8
56.3
155.5
149.1
148.9
148.8
148.6
147.8
146.2
145.6
131.9
130.6
127.2
126.7
122.1
120.0
119.1
114.9
113.1
112.1
111.3
111.1
110.4
104.8
100.6
56.1
37.9
37.6
37.4
56.3
56.2
56.0
56.1(1) (1C)
56.1(0) (2C)
56.0
51.3
38.0
56.1 (3C)
55.9
56.0
51.3
38.0
55.8
51.0
37.8
AData from ref. [1] and recorded in 5:1 v/v [D6]DMSO/CD3OD at 125 MHz.
BData arising from work reported in the present paper and recorded in 79% [D6]DMSO/21% CD3OD at 75 MHz.
CData from ref. [2] and recorded in 83% [D6]DMSO/17% CD3OD at 100 MHz.
DData arising from work reported in the present paper and recorded in CDCl3 at 75 MHz.
EData from ref. [2] and recorded in CDCl3 at 100 MHz.
FData from ref. [8] and recorded in CDCl3 at 125 MHz.
MS instrument operating in positive ionization mode. Melting
points were measured on an Optimelt automated melting point
system and are uncorrected. Analytical TLC was performed on
aluminium-backed 0.2-mm thick silica gel 60 F254 plates as sup-
plied by Merck. Eluted plates were visualized using a 254-nm
UV lamp and/or by treatment with a suitable dip followed
by heating. These dips included phosphomolybdic acid/ceric
sulfate/sulfuric acid (conc.)/water (37.5 g:7.5 g:37.5 g:720 mL)
or potassium permanganate/potassium carbonate/5% sodium
hydroxide aqueous solution/water (3 g:20 g:5 mL:300 mL). The
retardation factor (RF) values cited here have been rounded at
the first decimal point. Flash chromatographic separations were
carried out following protocols defined by Still et al.[15] using sil-
ica gel 60 (0.040–0.0063 mm) as the stationary phase and using
the AR- or HPLC-grade solvents indicated. Starting materials
and reagents were generally available from the Sigma–Aldrich,
Merck, TCI, Strem, or Lancaster chemical companies and were
used as supplied. Drying agents and other inorganic salts were
purchased from the AJAX, BDH, or Unilab chemical compa-
nies.Tetrahydrofuran (THF), dichloromethane, and diethyl ether
were dried using a Glass Contour solvent purification system
that is based on a technology originally described by Grubbs
et al.[16] Methanol was distilled from its magnesium alkoxide
salt.Wherenecessary, reactionswereperformedunderanitrogen
atmosphere.
Synthetic Studies
Compound 7
DIAD (428 mg, 2.12 mmol) was added, dropwise, to a mag-
netically stirred solution of compound 5 (215 mg, 1.06 mmol),
triphenylphosphine (556 mg, 2.12 mmol), and alcohol
6
(386 mg, 2.12 mol) in THF (10 mL) maintained at 18◦C. The
resulting solution was left stirring at this temperature for 15 h
then concentrated, under reduced pressure, onto TLC-grade sil-
ica (1.5 g). The solid thus obtained was added to the top of a
flashchromatographycolumnthatwasthensubjectedtogradient
elution (silica, CH2Cl2 → 9:1 v/v CH2Cl2/ethyl acetate gradient
elution). Concentration of the appropriate fractions (RF 0.5) then
afforded carboxylate 7[11] (291 mg, 75%) as a clear, colourless
oil. The spectral data derived from this material were identical,
in all respects, with those reported[11] for an authentic sample.

RESEARCH FRONT
688
K. Hasse, A. C. Willis, and M. G. Banwell

RESEARCH FRONT
Total Synthesis of Ningalin B
689
Compound 9
(2.21 mL, 9.62 mmol) in THF (70 mL) maintained at −78◦C,
and stirring was continued at this temperature for 0.66 h. The
reaction mixture was then allowed to warm to 18◦C over 12 h.
After the addition of HCl (20 mL of a 1 M aqueous solution)
and stirring for another 0.5 h, the phases were separated and the
aqueous one was diluted with brine (20 mL) and extracted with
THF (3 × 50 mL). The combined organic phases were washed
with brine (1 × 50 mL) before being dried (Na2SO4), filtered,
and concentrated under reduced pressure to give a yellow, mud-
like solid (2.12 g). This solid was dissolved in diethyl ether
(80 mL). Then pinacol (1.78 g, 15.1 mmol) was added and the
ensuing slurry was stirred magnetically for 24 h. Concentration
of the resulting mixture under reduced pressure, subjection of
the residue thus obtained to flash chromatography (silica, 1:1 v/v
ethyl acetate/hexane → ethyl acetate gradient elution) and con-
centration of the appropriate fractions (RF 0.6 in ethyl acetate)
afforded compound 15 (1.70 g, 69% over three steps) as a clear,
colourless oil (Found: M+•, 280.1483. C14H21BO5 requires
A mixture of pyrrole 7 (45 mg, 0.12 mmol) in THF/water
(2.0 mL of a 3:1 v/v mixture), boronic ester 8[13] (81 mg,
0.37 mmol), K2CO3 (85 mg, 0.62 mmol), TBAB (8 mg,
20 mol-%), and Pd(PPh3)4 (7 mg, 5 mol-%) was irradiated in
a microwave reactor at 70◦C for 1 h. The cooled reaction mix-
ture was diluted with water (40 mL) and ethyl acetate (50 mL).
The separated aqueous phase was extracted with ethyl acetate
(3 × 40 mL) and the combined organic phases were washed
with brine (1 × 20 mL), then dried (Na2SO4), before being fil-
tered and concentrated under reduced pressure. The ensuing
yellow oil was subjected to flash chromatography (silica, 1:1 v/v
pentane/diethyl ether elution) and concentration of the appropri-
ate fractions (RF 0.2) gave compound 9 (34 mg, 79%) as white
crystalline solid. The spectral data derived from this material
were identical, in all respects, with those reported[11] for an
authentic sample.
M
+•, 280.1482). δH (300 MHz, CDCl3) 7.65 (s, 1H), 6.99 (s,
Compound 10
1H), 6.44 (s, 1H), 3.86 (s, 3H), 3.84 (s, 3H), 1.35 (s, 12H). δC
(75 MHz, CDCl3) 159.8 (C), 154.1 (C), 142.7 (C), 116.4 (CH),
99.7 (CH), 84.5 (C), 56.6 (CH3), 56.0 (CH3), 25.0 (CH3) (one
signal obscured or overlapping). νmax(NaCl)/cm−1 3447, 2978,
2936, 1622, 1511, 1459, 1414, 1387, 1300, 1250, 1237, 1211,
1158, 1140, 992, 859. m/z (EI, 70 eV) 280 (M+•, 80%), 223 (42),
180 (100), 165 (55), 129 (78), 85 (55).
A magnetically stirred solution of pyrrole 9 (68 mg,
0.095 mmol) in DMF (2 mL) was cooled to 0◦C then treated,
in one portion, with NBS (36 mg, 0.2 mmol) and the ensuing
mixture was allowed to warm to 18◦C. After stirring at this
temperature for 18 h, the reaction mixture was diluted with
water (30 mL) and extracted with ethyl acetate (4 × 40 mL). The
combined organic phases were washed with water (1 × 40 mL)
and brine (1 × 20 mL) before being dried (MgSO4), filtered,
and concentrated under reduced pressure to give a light-
yellow oil. Subjection of this material to flash chromatography
(silica, 3:1 → 1:2 v/v pentane/diethyl ether gradient elution) and
concentration of the appropriate fractions (RF 0.3 in 1:2 v/v
pentane/diethyl ether) gave a crystalline solid. Recrystalliza-
tion (pentane/diethyl ether) of this material afforded com-
pound 10 (62 mg, 74%) as a white, crystalline solid, mp 142–
Compound 16
A solution of pyrrole 7 (23 mg, 0.06 mmol), caesium carbon-
ate (102 mg, 0.31 mmol), TBAB (4 mg, 0.01 mmol, 20 mol-%),
and compound 15 (61 mg, 0.22 mmol) in THF/H2O (0.5 mL of
4:1 v/v mixture) was placed in a glass reaction tube that was
flushed with nitrogen. (PPh3)4Pd (7 mg, 0.01 mmol, 10 mol-%)
was then added to the tube which was flushed with nitrogen
once more then sealed and the resulting mixture subjected to
microwave irradiation (150W, 80◦C, 1 min ramp time) for 2 h.
To effect full conversion, the cooled reaction mixture was treated
with further aliquots of boronic ester 15, caesium carbonate,
and (PPh3)4Pd (same amounts as above). The resulting mixture
was irradiated under the abovementioned conditions for another
2 h then cooled, diluted with H2O (30 mL) and extracted with
ethyl acetate (4 × 30 mL). The combined organic phases were
washed with brine (1 × 20 mL) before being dried (Na2SO4),
filtered, and concentrated under reduced pressure. The ensuing
light-yellow oil was subjected to flash chromatography (silica,
3:1 v/v pentane/diethyl ether → diethyl ether gradient elution)
and concentration of the appropriate fractions (RF 0.2 in 1:3 v/v
pentane/diethyl ether) afforded a white solid. Recrystalliza-
tion of this material (pentane/diethyl ether) afforded the title
compound 16 (11 mg, 43%) as white prisms, mp 159–160◦C
(Found: M+•, 409.1527. C23H23NO6 requires M+•, 409.1525).
δH (300 MHz, CDCl3) 7.08 (s, 1H), 6.92 (s, 1H), 6.82 (d, J 2.7,
1H), 6.75 (d, J 8.1, 1H), 6.63 (dd, J 8.1 and 2.1, 1H), 6.56 (d,
J 2.1, 1H), 6.42 (d, J 2.7, 1H), 4.60 (t, J 6.9, 2H), 3.95 (s, 3H),
3.92 (s, 3H), 3.83 (s, 3H), 3.76 (s, 3H), 3.07 (t, J 6.9, 2H). δC
(75 MHz, CDCl3) 155.6 (C), 149.6 (C), 149.1 (C), 147.9 (C),
146.5 (C), 146.4 (C), 133.1 (CH), 131.4 (C), 130.7 (C), 121.1
(CH), 115.1 (C), 112.2 (CH), 111.4 (CH), 110.4 (C), 104.3 (CH),
100.9 (CH), 100.7 (CH), 56.6 (CH3), 56.4 (CH3), 56.1 (CH3),
56.0 (CH3), 51.2 (CH2), 38.0 (CH2). νmax(NaCl)/cm−1 2936,
1705, 1516, 1426, 1261, 1236, 1205, 1192, 1155, 1066, 1029,
153◦C (Found: M+•, 427.0417. C21H1879BrNO4 requires M+•
,
427.0419). δH (300 MHz, CDCl3) 8.55 (d, J 8.3, 1H), 7.40 (m,
2H), 7.30 (m, 1H), 6.89 (s, 1H), 6.77 (d, J 8.1, 1H), 6.66 (dd, J 8.1
and 1.8, 1H), 6.58 (d, J 1.8, 1H), 4.61 (t, J 6.9, 2H), 3.84 (s, 3H),
3.80 (s, 3H), 3.05 (t, J 6.9, 2H). δC (75 MHz, CDCl3) 154.4,
151.4, 149.2, 148.1, 132.9, 130.1, 128.6, 126.7, 124.3, 122.9,
121.1, 117.6, 117.4, 116.2, 112.1, 111.5, 90.5, 56.1, 56.0, 51.5,
37.9. νmax(NaCl)/cm−1 3119, 2999, 2955, 2834, 1717, 1612,
1591, 1516, 1447, 1377, 1262, 1236, 1157, 1050, 1028, 964,
805, 766, 751, 733. m/z (EI, 70 eV) 429 and 427 (M+•, 26%),
164 (100), 151 (48).
Compound 12
Acetal 12 was prepared in near-quantitative yield from phenol
11 according to the method of Handy et al.[14] The spectral data
recorded on product 12 matched those reported previously.[14]
Compound 13
Bromide 13 was prepared in ∼83% yield from acetal 12
according to the method of Handy et al.[14] The spectral data
recorded on product 13 matched those reported previously.[14]
Compound 15
n-Butyllithium(1.6 Msolutioninhexane, 7.1 mL, 11.4 mmol)
was added dropwise over 0.25 h to a magnetically stirred solution
of compound 13 (2.79 g, 8.80 mmol) and tri-isopropyl borate

RESEARCH FRONT
690
K. Hasse, A. C. Willis, and M. G. Banwell
1004. m/z (EI, 70 eV) 409 (M+•, 100%), 258 (28), 245 (76), 164
(40), 151 (53).
18◦C over 16 h. The reaction mixture was then diluted with
methanol (1.5 mL) and concentrated under reduced pressure
to give ningalin B (14 mg, 96%) as an amorphous and dark-
yellow solid (Found: [M + H]+, 462.1182. C25H19NO8 requires
[M + H]+, 462.1189). δH (300 MHz, CDCl3) see Table 2. δC
(75 MHz, CDCl3) seeTable 1. νmax(NaCl)/cm−1 2920, 1601. m/z
(ESI) 484 ([M + Na]+, 16%), 484 ([M + H]+, 85), 279 (100),
149 (20).
Compound 17
N-Bromosuccinimide (12 mg, 0.07 mmol) was added, in one
portion, to a magnetically stirred solution of compound 16
(13 mg, 0.03 mmol) in anhydrous DMF (0.3 mL) maintained
at 0◦C. The ensuing mixture was allowed to warm to 18◦C,
stirred at this temperature for 4 h, then diluted with H2O
(10 mL) and extracted with ethyl acetate (4 × 20 mL). The
combined organic phases were washed with H2O (1 × 30 mL)
and brine (1 × 20 mL) before being dried (Na2SO4), filtered,
and concentrated under reduced pressure. Subjection of the
ensuing residue to flash chromatography (silica, 1:3 → 1:1 v/v
ethyl acetate/hexane gradient elution) and concentration of the
appropriate fractions (RF 0.3 in 1:1 v/v ethyl acetate/hexane)
afforded bromide 17 (15 mg, 97%) as a white solid, mp 142–
Compound 20
A solution of pyrrole 4 (27 mg, 0.09 mmol) in THF/H2O
(0.6 mL of 5:1 v/v mixture) was treated with caesium carbonate
(145 mg, 0.45 mmol), TBAB (6 mg, 0.02 mmol, 20 mol-%),
and boronate ester 15 (94 mg, 0.37 mmol).The resulting mixture
was flushed with nitrogen then (PPh3)4Pd (11 mg, 0.01 mmol,
11 mol-%) was added and the reaction vessel again flushed
with nitrogen then sealed. The reaction mixture was sub-
jected to microwave irradiation (150W, 70◦C, 1 min ramp
time) for 1 h then cooled and further portions of boronate 15
(80 mg, 0.29 mmol), caesium carbonate (128 mg, 0.39 mmol)
and (PPh3)4Pd (10 mg, 0.01 mmol, 10 mol-%) were added.
The resulting mixture was, once again, irradiated under the
abovementioned conditions for 1 h. The cooled reaction mix-
ture was then diluted with H2O (50 mL) and extracted with
ethyl acetate (4 × 30 mL). The combined organic phases were
washed with brine (1 × 30 mL) before being dried (Na2SO4),
filtered and concentrated under reduced pressure. The ensuing
light-yellow oil was subjected to flash chromatography (sil-
ica, CH2Cl2 → 9:1 v/v CH2Cl2/ethyl acetate gradient elution)
to afford two fractions, A and B.
144◦C (Found: M+•, 487.0639. C23H22BrNO6 requires M+•
,
487.0630). δH (300 MHz, CDCl3) 8.02 (s, 1H), 6.93 (s, 1H),
6.89 (s, 1H), 6.78 (d, J 8.1, 1H), 6.65 (dd, J 8.1 and 1.5, 1H),
6.59 (d, J 1.5, 1H), 4.61 (t, J 6.9, 2H), 3.98 (s, 3H), 3.94 (s,
3H), 3.86 (s, 3H), 3.81 (s, 3H), 3.06 (t, J 6.9, 2H). δC (75 MHz,
CDCl3) 154.7 (CO), 149.8 (C), 149.2 (C), 148.1 (C), 146.4 (C),
146.1 (C), 132.8 (CH), 130.2 (CH), 127.3 (C), 121.1 (CH), 115.3
(C), 112.1 (CH), 111.4 (CH), 109.7 (C), 104.0 (C), 100.7 (CH),
89.5 (C), 56.5 (CH3), 56.4 (CH3), 56.1 (CH3), 56.0 (CH3), 51.5
(CH2), 38.0 (CH2). νmax(NaCl)/cm−1 2925, 1711, 1516, 1463,
1414, 1263, 1221, 1157, 1115, 1029, 1012, 966. m/z (EI, 70 eV)
489 and 487 (M+•, 70 and 69%), 323 and 321 (73 and 74), 164
(100), 151 (95).
Concentration of fraction A (RF 0.5 in 9:1 v/v CH2Cl2/ethyl
acetate) afforded carboxylate 20 (17 mg, 55%) as a white solid,
Ningalin B Permethyl Ether 19
no melting point, decomposition above 250◦C (Found: M+•
,
345.1212. C18H19NO6 requires M+•, 345.1212). δH (300 MHz,
CDCl3) 7.75 (d, J 3.3, 1H), 7.09 (s, 1H), 6.90 (s, 1H), 6.66 (d,
J 3.3, 1H), 3.97 (s, 3H), 3.93 (s, 3H), 1.67 (s, 9H). δC (75 MHz,
CDCl3) 152.9 (C), 150.9 (C), 148.0 (C), 147.6 (C), 146.5 (C),
136.5 (C), 132.6 (CH), 115.8 (C), 108.8 (C), 104.5 (CH), 104.3
(CH), 100.5 (CH), 86.1 (C), 56.6 (CH3), 56.5 (CH3), 28.0 (CH3).
νmax(NaCl)/cm−1 1735, 152, 1453, 1406, 1339, 1270, 1224,
1158, 995, 847. m/z (EI, 70 eV) 345 (M+•, 10%), 245 (90), 205
(88), 154 (100), 139 (75), 129 (57), 57 (62).
A mixture of pyrrole 17 (26 mg, 0.05 mmol), boronic
acid 18[13] (29 mg, 0.11 mmol), K2CO3 (37 mg, 0.19 mmol),
TBAB (3 mg, 20 mol-%), and Pd(PPh3)4 (6 mg, 10 mol-%)
in THF/water (0.65 mL of a 4:1 v/v mixture) was irradiated
at 80◦C for 1 h in a microwave reactor. The cooled reac-
tion mixture was diluted with water (20 mL) and ethyl acetate
(20 mL) then the separated aqueous phase was extracted with
ethyl acetate (3 × 30 mL). The combined organic phases were
washed with brine (1 × 20 mL) before being dried (Na2SO4),
filtered then concentrated under reduced pressure. The ensu-
ing yellow oil was subjected to flash chromatography (sil-
ica, dichloromethane → 9:1 v/v dichloromethane/ethyl acetate
gradient elution) and concentration of the appropriate frac-
tions (RF 0.3 in 1:9 v/v dichloromethane/ethyl acetate) gave a
white, crystalline solid. Recrystallization (ethyl acetate/hexane)
of this material afforded compound 19 (20 mg, 69%) as a
white crystalline solid, mp 190–191◦C (lit.[2] mp 186–187◦C)
(Found: M+•, 545.2050. C31H31NO8 requires M+•, 545.2050).
δH (300 MHz, CDCl3) see Table 2. δC (75 MHz, CDCl3) see
Table 1. νmax(NaCl)/cm−1 2927, 1709, 1515, 1464, 1415, 1259,
1244, 1174, 1139, 1027, 731. m/z (EI, 70 eV) 545 (M+•, 100%),
394 (50), 381 (73).
Concentration of fraction B (RF 0.3 in 9:1 v/v CH2Cl2/ethyl
acetate) afforded compound 21 (3 mg, 14%) as a white solid,
no melting point, decomposition above 250◦C (Found: M+•
,
245.0688. C13H11NO4 requires M+•, 245.0688). δH (300 MHz,
CDCl3) 9.98 (br s, NH, 1H), 7.17 (s, 1H), 7.10 (s, 1H), 6.97
(s, 1H), 6.68 (2, 1H), 3.99 (s, 3H), 3.94 (s, 3H). δC (75 MHz,
CDCl3) 156.3 (C), 149.7 (C), 146.7 (C), 146.3 (C), 130.7 (C),
129.0 (CH), 116.5 (C), 110.4 (CH), 104.6 (CH), 103.0 (CH),
101.1 (CH), 56.6 (CH3), 56.4 (CH3). νmax(NaCl)/cm−1 3242,
2919, 1722, 1625, 1535, 1498, 1446, 1421, 1296, 1261, 1222,
1187, 1155, 1109, 1065, 1006, 772. m/z (EI, 70 eV) 245 (M+•
,
100%), 230 (25), 129 (35), 85 (40), 69 (34).
Compound 21
Ningalin B (1)
Zinc bromide (20 mg, 0.087 mmol) was added to a mag-
netically stirred solution of pyrrole 20 (10 mg, 0.029 mmol) in
CH2Cl2 (2 mL) maintained under nitrogen at 18◦C. After 2.5 h
the reaction mixture was treated with H2O (5 mL) and extracted
with CH2Cl2 (3 × 20 mL). The combined organic phases were
washed with brine (1 × 10 mL) before being dried (Na2SO4),
Following the procedure detailed by Boger et al.,[2] a mag-
netically stirred solution of compound 19 (17 mg, 0.03 mmol) in
CH2Cl2 (3 mL) maintained under nitrogen was cooled to −78◦C,
then treated with BBr3 (0.23 mL of a 2 M solution in hexane,
0.47 mmol) and the ensuing mixture then allowed to warm to

RESEARCH FRONT
Total Synthesis of Ningalin B
691
filtered, and concentrated under reduced pressure. Subjection
of the ensuing residue to flash chromatography (silica, 3:1 v/v
pentane/diethyl ether → diethyl ether → 1:1 v/v diethyl ether/
ethyl acetate gradient elution) and concentration of the appro-
priate fractions (RF 0.3 in diethyl ether) afforded compound 21
(6 mg, 84%) as a white, crystalline solid.This material was iden-
tical, in all respects, with an authentic sample of compound 21
obtained via the procedure described immediately above.
Acknowledgements
We thank the Gottlieb Daimler and Karl Benz Foundation for the provision
of a PhD stipend to K.H. and the Australian Research Council for a grant
(DP0771749).
References
[1] H. Kang, W. Fenical, J. Org. Chem. 1997, 62, 3254. doi:10.1021/
JO962132+
[2] D. L. Boger, D. R. Soenen, C. W. Boyce, M. P. Hedrick, Q. Jin, J. Org.
Chem. 2000, 65, 2479. doi:10.1021/JO9916535
Compound 16
DIAD (22 mg, 0.11 mmol) was added dropwise to a mag-
netically stirred solution of compound 21 (18 mg, 0.073 mmol),
triphenylphosphine (29 mg, 0.11 mmol), and alcohol 6 (20 mg,
0.11 mmol) in THF (1 mL) maintained at 18◦C. The resulting
solution was left stirring at this temperature for 15 h then, so
as to ensure complete conversion, the same quantities of DIAD
and triphenylphosphine specified above were again added to the
reaction mixture. This was allowed to stir at 18◦C for a further
15 h then concentrated under reduced pressure onto TLC-grade
silica gel (0.4 g).The free-flowing solid thus obtained was added
to the top of a flash chromatography column. Elution of the col-
umn (with 3:1 v/v pentane/diethyl ether → diethyl ether gradient
elution) and concentration of the appropriate fractions (RF 0.3
in diethyl ether) afforded lactone 16 (16 mg, 54%) as a white,
crystalline solid.This material was identical, in all respects, with
an authentic sample of compound 16 obtained via the procedure
described above.
[3] For a recent and comprehensive review of this topic see: H. Fan,
J. Peng, M. T. Hamann, J.-F. Hu, Chem. Rev. 2008, 108, 264.
doi:10.1021/CR078199M
[4] D. R. Soenen, I. Hwang, M. P. Hedrick, D. L. Boger, Bioorg. Med.
Chem. Lett. 2003, 13, 1777. doi:10.1016/S0960-894X(03)00294-4
[5] H. Tao, I. Hwang, D. L. Boger, Bioorg. Med. Chem. Lett. 2004, 14,
5979. doi:10.1016/J.BMCL.2004.10.002
[6] M. Iwao,T.Takeuchi, N. Fujikawa,T. Fukuda, F. Ishibashi,Tetrahedron
Lett. 2003, 44, 4443. doi:10.1016/S0040-4039(03)01031-1
[7] J. L. Bullington, R. R. Wolff, P. F. Jackson, J. Org. Chem. 2002, 67,
9439. doi:10.1021/JO026445I
[8] J. T. Gupton, S. C. Clough, R. B. Miller, J. R. Lukens, C. A. Henry,
R. P. F. Kanters, J. A. Sikorski, Tetrahedron 2003, 59, 207.
doi:10.1016/S0040-4020(02)01475-8
[9] C. Peschko, C. Winklhofer, A. Terpin, W. Steglich, Synthesis 2006,
3048.
[10] (a) M. G. Banwell, B. L. Flynn, E. Hamel, D. C. R. Hockless, Chem.
Commun. 1997, 207. doi:10.1039/A606793J
(b) M. Banwell, B. Flynn, D. Hockless, Chem. Commun. 1997, 2259.
doi:10.1039/A705874H
(c) M. G. Banwell, A. M. Bray, A. C. Willis, D. J. Wong, New J. Chem.
1999, 23, 687. doi:10.1039/A903330K
(d) M. R. Heinrich, W. Steglich, M. G. Banwell, Y. Kashman, Tetra-
hedron 2003, 59, 9239. doi:10.1016/J.TET.2003.02.005
(e) D. M. Pinkerton, M. G. Banwell, A. C. Willis, Org. Lett. 2007, 9,
5127. doi:10.1021/OL7024313
X-Ray Crystallographic Studies on Compounds 10 and 16
Crystal Data
Compound 10: C21H18BrNO4, M = 428.28, T = 200(1) K,
triclinic, space group P1, Z = 2, a = 7.3505(3), b = 8.9299(4),
c = 14.1446(7) Å, α = 97.197(2)◦, β = 96.320(3)◦, γ =
93.102(3)◦,V = 913.36(7) Å3, Dx = 1.557 g cm−3, 4201 unique
data (2θmax = 55.2◦); R = 0.037 (for 3120 reflections with
I > 2.0σ(I)), Rw = 0.082 (all data), S = 0.93.
[11] L. C. Axford, K. E. Holden, K. Hasse, M. G. Banwell, W. Steglich,
J. Wagler, A. C. Willis, Aust. J. Chem. 2008, 61, 80. doi:10.1071/
CH07402
[12] V. A. Burgess, C. J. Easton, M. P. Hay, J. Am. Chem. Soc. 1989, 111,
1047. doi:10.1021/JA00185A039
[13] This compound was purchased from Boron Molecular Pty
Ltd, Noble Park, VIC 3174, Australia (Available online at:
http://www.boronmolecular.com; accessed 19 May 2009).
[14] S. T. Handy, Y. Zhang, H. Bregman, J. Org. Chem. 2004, 69, 2362.
doi:10.1021/JO0352833
Compound 16: C23H23NO6, M = 409.44, T = 200 K,
monoclinic, space group C2/c, Z = 8, a = 33.6193(7), b =
7.24830(10), c = 16.4673(3) Å, β = 95.6376(12)◦, V = 3993.39
(12) Å3, Dx = 1.362 g cm−3, 4609 unique data (2θmax = 55.2◦);
R = 0.035 (for 2895 reflections with I > 2.0σ(I)), Rw = 0.083
(all data), S = 0.79.
[15] W. C. Still, M. Kahn, A. Mitra, J. Org. Chem. 1978, 43, 2923.
doi:10.1021/JO00408A041
Structure Determination
[16] A. B. Pangborn, M. A. Giardello, R. H. Grubbs, R. K. Rosen,
F. J. Timmers, Organometallics 1996, 15, 1518. doi:10.1021/
OM9503712
[17] DENZO-SMN. Z. Otwinowski W. Minor, in Methods in Enzymol-
ogy, Volume 276: Macromolecular Crystallography, Part A (Eds
C. W. Carter, Jr, R. M. Sweet) 1997, pp. 307–326 (Academic Press:
New York, NY).
[18] A. Altomare, G. Cascarano, C. Giacovazzo, A. Guagliardi,
M. C. Burla, G. Polidori, M. Camalli, J. Appl. Cryst. 1994, 27, 435.
doi:10.1107/S002188989400021X
[19] P. W. Betteridge, J. R. Carruthers, R. I. Cooper, K. Prout, D. J. Watkin,
J. Appl. Cryst. 2003, 36, 1487. doi:10.1107/S0021889803021800
Images were measured on a Nonius Kappa CCD diffracto-
meter (MoKα, graphite monochromator, λ 0.71073 Å) and
the data extracted using the DENZO package.[17] The struc-
ture solutions were by direct methods (SIR92).[18] The struc-
tures of compounds 10 and 16 were refined using the
CRYSTALS program package.[19] Atomic coordinates, bond
lengths and angles, and displacement parameters have been
deposited at the Cambridge Crystallographic Data Cen-
tre (CCDC numbers 723557 and 723558 for compounds
10 and 16, respectively). These data can be obtained
free-of-charge from www.ccdc.cam.ac.uk/data_request/cif, by
emailing data_request@ccdc.cam.ac.uk, or by contacting the
Cambridge Crystallographic Data Centre, 12 Union Road,
Cambridge CB2 1EZ, UK; fax: +44 1223 336033.