A biosynthetically inspired synthesis of (?)-berkelic acid and analogs

Article abstract of DOI:10.1016/j.tet.2018.01.021

We describe a complete account of our total synthesis and biological evaluation of (?)-berkelic acid and analogs. We delineate a synthetic strategy inspired by a potentially biomimetic union between the natural products spicifernin and pulvilloric acid. A

Full text of DOI:10.1016/j.tet.2018.01.021

Accepted Manuscript
A biosynthetically inspired synthesis of (–)-berkelic acid and analogs
Christopher F. Bender, Christopher L. Paradise, Vincent M. Lynch, Francis K.
Yoshimoto, Jef K. De Brabander
PII:
S0040-4020(18)30054-1
10.1016/j.tet.2018.01.021
TET 29236
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 30 August 2017
Revised Date: 5 January 2018
Accepted Date: 10 January 2018
Please cite this article as: Bender CF, Paradise CL, Lynch VM, Yoshimoto FK, De Brabander JK, A
biosynthetically inspired synthesis of (–)-berkelic acid and analogs, Tetrahedron (2018), doi: 10.1016/
j.tet.2018.01.021.
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ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT
A biosynthetically inspired synthesis of (–)-berkelic acid and analogs
Christopher F. Bendera, Christopher L. Paradisea, Vincent M. Lynchb, Francis K. Yoshimotoa, Jef K. De
Brabandera,*
aDepartment of Biochemistry, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-9038, USA
bDepartment of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712, USA
* Corresponding author. E-mail: Jef.DeBrabander@UTSouthwestern.edu
Keywords: Natural products; cycloaddition; cycloisomerization; quinone methide; spiroketals
Abstract
We describe a complete account of our total synthesis and biological evaluation of (–)-berkelic acid and analogs. We delineate a synthetic
strategy inspired by a potentially biomimetic union between the natural products spicifernin and pulvilloric acid. After defining optimal
parameters, we executed a one-pot silver-mediated in situ dehydration of an isochroman lactol to methyl pulvillorate, the
cycloisomerization of a spicifernin-like alkynol to the corresponding exocyclic enol ether, and a subsequent cycloaddition to deliver the
tetracyclic core of berkelic acid. Our studies confirm that the original assigned berkelic acid structure is not stable and equilibrates into a
mixture of 4 diastereomers, fully characterized by X-ray crystallography. In addition to berkelic acid, C22-epi-berkelic acid, and nor-
berkelic acids, we synthesized C26-oxoberkelic acid analogs that were evaluated against human cancer cell lines. In contrast to data
reported for natural berkelic acid, our synthetic material and analogs were found to be devoid of activity.
1. Introduction
In 2006 Stierle et al. disclosed the tetracyclic chroman/isochroman/spiroketal natural product (–)-berkelic acid (Figure 1) isolated from
the fermentation broth of an extremophilic Penicillium fungus encountered in the Berkeley Pit Lake in Butte, Montana.1 The Berkeley Pit
Lake, which formed by groundwater seepage into an abandoned copper pit mine, is currently the United States’ largest superfund cleanup
site containing approximately 30 billion gallons of highly acidic (pH 2.5), heavy metal-contaminated (arsenic, copper, cadmium, cobalt,
iron, manganese, and zinc) water.2 Berkelic acid was found to be a moderate inhibitor of MMP-3 (1.87 µM) and the cysteine protease
caspase-1 (98 µM), and, in testing against the NCI 60 cell line panel, was reported to possess selective activity against the human ovarian
cancer cell line OVCAR-3 (GI50 91 nM). However, a subsequent analysis of fully synthetic (–)-berkelic acid from the Snider group in the
NCI 60 cell line panel indicated no activity against any of the cell lines (up to 10 µM) including OVCAR-3.3
The initially assigned structure of (–)-berkelic acid (1) was determined by NMR experiments, but did not address the configuration of
the C22-quaternary stereocenter or the overall absolute configuration. Subsequent work of the Fürstner group led to a revised structure of
berkelic acid (2) through an elegant synthetic, NMR, and crystallographic study culminating initially in the synthesis of both C22-epimers
of the corresponding methyl esters of ent-2,4 and subsequently (–)-berkelic acid (2).5 These efforts established the absolute configuration of
five stereocenters and a revision of the configuration at C18 and C19 in the original assigned structure. In fact, Fürstner noticed that an
advanced tetracyclic intermediate with configuration reminiscent of the original structure could not be prepared as a single diastereomer.
An unfavorable eclipsing interaction interaction between C16 and C25 instead provided a driving force for equilibration into a nearly
statistical mixture of four diasteromers epimeric at C15, C17, and C18.6,7 Unfortunately, insufficient spectroscopic differences between two
synthetic methyl berkelates epimeric at C22, lack of an authentic sample, and the inability to selectively saponify the C1-methyl benzoate
prevented an unambiguous assignment of the C22 stereocenter. Later, Snider and coworkers reported the first total synthesis of berkelic
1

ACCEPTED MANUSCRIPT
acid, confirming Fürstner’s structural revision and putatively assigning the quaternary stereocenter as C22-S.3,8 As the C22 stereocenter was
introduced via a non-selective Kiyooka aldol reaction,9 the stereochemical assignment relied upon correlation of the resultant diastereomers
to a model compound. Our subsequent synthesis of both C22-epimers of berkelic acid fully corroborated the Snider assignment.10 Finally, a
gram-scale total synthesis was reported in 2012 by Fañanás and Rodríguez that centered on a silver-catalyzed addition/cyclization cascade
of appropriately substituted alkyne and aldehyde precursors to set four new chiral centers in a 2:1 diastereomeric ratio.11,12,13
O
O
MeO2C
Me
MeO2C
Me
1 CO2H
CO2H
22
O
O
H
19
O
OH
O
OH
7
3
24Me
Me
16
15
25
Me
Me
5
H
O
H
O
H
eclips
9
original structure
of berkelic acid (1)
(Stierle)
revised structure
of berkelic acid (2)
(Fürstner, Snider, De Brabander)
14
Me
Me
Figure 1. Original and revised structures of berkelic acid.
Constitutionally, the originally proposed structure of (–)-berkelic acid (1) emerges as an amalgamation of two other natural products,
namely pulvilloric acid (4)14 and spicifernin (3). The co-isolation of berkelic acid with spiciferone A (5),1 which previously had been
isolated alongside spicifernin (3) from a Cochliobolus fungus,15 is indeed suggestive of a proposal that berkelic acid could be the end-
2
product of such a biosynthetic merger (Scheme 1). Biosynthetic studies utilizing 13C- and H-labeling have specified that both spicifernin
(3) and spiciferone A (5) are generated from a common hexaketide precursor.15,16 Thus, the presence of spiciferone A (5) in the berkelic
acid culture medium suggests that the Berkeley Pit Lake Penicillium sp. might also possess the biosynthetic machinery to produce
spicifernin (3). To account for the stereochemical discrepancy between natural spicifernin (3) and (–)-berkelic acid (2), which display an
epimeric relationship at C18 and C19 (berkelic acid numbering), Snider proposed that spicifernin (3) or an immediate biosynthetic
precursor could undergo a double epimerization (C18 and C19) followed by a reduction to provide a correctly configured bis-epi-
deoxyspicifernin (6) for addition to pulvilloric acid (4).3,17 We proposed an alternative hypothesis where spicifernin (3) would be unified
with pulvilloric acid (4) first, perhaps via its dehydrated enollactone 7 in a formal cycloaddition with the ortho-quinone methide tautomer of
pulvilloric acid, followed by a double epimerization (at C18 and C19) and final reduction.18,19
Given berkelic acid’s unique structural features, stereochemical questions, biological activity, and potential biosynthetic origin from
combination of two other natural products, we embarked on a synthetic program that culminated in the total synthesis of (–)-berkelic acid
(2) communicated several years ago.10 Here, we present a full account of our extensive research program including the development of a
novel silver-catalyzed dehydration/cycloisomerization/cycloaddition cascade to forge the tetracyclic chroman/isochroman/spiroketal
structure embedded within berkelic acid, the synthesis and full characterization of diastereomers of the originally proposed structure, C22-
geminal dimethyl substituted analogs, and C26-oxoberkelates to explore the proposed biosynthesis, and biological evaluation of synthetic
berkelic acid and analogs.
2

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Scheme 1. Biosynthetic proposal for the synthesis of (–)-berkelic acid (2) from the natural products spicifernin (3) and pulvilloric acid (4).
2. Results and discussion
Initially, we became interested in a synthetic campaign toward berkelic acid due to a unique constellation of structural features that we
perceived as a fruitful platform to implement methodology developed in our group. In our initial decisively non-biomimetic analysis, we
thus perceived linear polyhydroxy alkyne 9 as a suitable direct precursor to berkelic acid (2) if conditions could be identified to forge a one-
pot cycloisomerization / dehydrative cycloetherification as depicted in Scheme 2. The dividend pay-off would be even higher if thus
identified reaction conditions could operate on a protecting group free substrate with all final functionality at the correct oxidation state (i.e.
substrate 9). Based on studies from our group, we had considerable confidence in executing such a strategy. Indeed, as shown in the box in
Scheme 2, we developed methodology to exploit internal alkynes as a potential nucleus for metal-catalyzed remolding of linear substrates
to cyclic/polycyclic structures. In 2006, we reported on the metal-catalyzed cycloisomerization of dihydroxy-alkynes to form spiroketals
under mild conditions (relevant to path a, 10 → 11)20 followed by propargylic cycloetherification in 2008 (relevant to transformation 10 →
12)21 and a tandem combination that would yield 13 directly from 10 (via 12; propargylic substitution preceding cycloisomerization).22 En
route to berkelic acid, we envisioned polyhydroxy-alkyne precursor 9 to be available from the addition of terminal alkyne 14 to aldehyde
15 (C15-C16 bond formation). However, if we were to implement a protecting group free synthesis, this aldehyde 15 would entirely exist
as the corresponding non-electrophilic lactol 16, leading us to contemplate the natural product pulvilloric acid (4, the dehydration product
of 16) as a viable electrophilic coupling partner instead. It is at this point that our synthetic analysis metamorphosed to one that appeared
more reminiscent of the biosynthetic pathway proposed in Scheme 1 and contemplated that a metal-catalyzed 5-exo-dig hydroalkoxylation
3

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of alkynol 14 should deliver enol 17,20 a material that would combine with the ortho-quinone methide tautomer of pulvilloric acid (4) via a
formal cycloaddition to deliver directly the chroman/isochroman/spiroketal tetracycle of berkelic acid (i.e. bypassing proposed precursor 9).
Ideally, a one-pot operation would be identified that enables (1) the dehydration of lactol 16 to pulvilloric acid 4, (2) the cycloisomerization
of 14 to 17, and (3) the final cycloaddition between 4 and 17 to deliver berkelic acid (2).23
O
O
path a
11
O
OH
O
O
OH
OH
ref. 22
ref. 20
7
26
HO
OH
a
OH
15
HO2C
OH
9
ref. 21
path b
HO
O
OH
26
MeO2C
13
OH
7
b
HO
10
Me
Me
OH
6
15
O
9
12
Me
OH
C5H11
9
CO2H
HO2C
HO
OH
HO
O
OH
O
OH
MeO2C
C-C bond
formation
cycloisomerization
dehydration
Me
Me
HO
+
OH
16
15
Me
O
C5H11
C5H11
14
15
16
O
MeO2C
CO2H
cycloisomeriztion
−H2O
O
H
O
OH
Me
Me
CO2H
CO2H
O
O
OH
HO
O
O
O
Me
MeO2C
Me
O
O
H
+
15
cycloaddition
Me
C5H11
16
4: pulvilloric
acid
Me
2: (−)-berkelic acid
17
C5H11
C5H11
Scheme 2. Retrosynthetic analysis and strategy.
Our synthetic studies began with the development of a concise route to a lactol similar to 16 as a precursor to pulvilloric acid (4). As
shown in Scheme 3, selective triflation of commercially available methyl 2,4,6-trihydroxybenzoate 18 provided triflate 19, which readily
underwent Suzuki-Miyaura cross coupling with known (E)-1-heptenylboronic acid (20)24 to yield styrene derivative 21 in 83% yield from
commercial 18.25 Installation of the homobenzylic alcohol was best accomplished via a three-step sequence from 21 including MOM
protection of the free phenols, styrene epoxidation with m-CPBA, and benzylic hydrogenolysis providing alcohol 22 in 76% overall yield
from 21.26 Methanolysis of the MOM protecting groups and methyl ester delivered alcohol 23 in 97% yield and was followed by a
condensation with triethyl orthoformate via a procedure adapted from the synthesis of pulvilloric acid14c to deliver the ethanol adduct of
methyl pulvillorate 24 in excellent yield.
With a viable racemic route established, we next explored the possibility to intercept racemic homobenzylic alcohol 22 for a late-stage
enzymatic resolution. We subjected (±)-22 to a set of 21 different lipases for acetylation of the homobenzylic alcohol with vinyl acetate and
identified three enzymes that provided the desired acetate 25 at a synthetically practical rate. Of these, a lyophilized formulation of lipase
from Alcaligenes sp. exhibited high selectivity and scalability, allowing for the preparation of (R)-25 and (S)-22 in 95% and 93% ee,
respectively.27 As it was ultimately discovered that berkelic acid was of the C9-(R) configuration,3-5 a Mitsunobu esterification of the
enantioenriched homobenzylic alcohol (S)-22 (93% ee) with acetic acid provided additional (R)-25 in 78% yield, for an overall yield of
85% of (R)-25 from (±)-22. Global deprotection of (R)-25 with acidic MeOH gave (R)-23 in quantitative yield. Treatment with
triethylorthoformate as described above afforded (R)-24. Expecting potential difficulties with a selective late-stage methyl benzoate
4

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saponification based on precedent by Fürstner and Snider,4,8 we briefly explored alternative benzoate esters at this stage. For example,
transesterification of 21 using catalytic Bu2SnO afforded 2-(trimethylsilyl)ethyl (TMSE) benzoate 26 in good yield (Scheme 5).28
Elaboration via the method described for 23 supplied racemic 27 for enzymatic resolution. However, screening of the same set of 21 lipases
as above did not identify a single lipase to affect this transformation. Literature suggests this failure is likely due to the occupation of the
lipase active site by the TMSE ester, preventing transesterification.29 At this point, we decided to move on with methyl benzoate (R)-24 and
force ourselves to find a solution for its selective removal at a later stage.
CO2Me
OH
CO2Me
OH
CO2Me
OMOM
CO2Me
OH
CO2Me
OH
HO
HO
MOMO
HO
HO
HO
HO
EtO
b 91%
c-e
f
g
B(OH)2
76%
97%
99%
OR
O
20
nC5H11
21
nC5H11
22
nC5H11
nC5H11
nC5H11
18: R = H
a
19: R = OTf
23
24
(R)-24
91%
m
86%
h
g
99%
CO2TMSE
OH
CO2TMSE
OH
CO2Me
OMOM
CO2Me
CO2Me
OH
HO
HO
HO
MOMO
HO
MOMO
AcO
OMOM
HO
HO
i-l
o
+
70%
99%
51%
46%
93% ee
95% ee
nC5H11
nC5H11
26
nC5H11
nC5H11
(R)-25
nC5H11
(R)-23
n
78%
27
(S)-22
Scheme 3. Synthesis of lactol 24, a precursor to pulvilloric acid (4). Reagents and conditions: (a) Tf2O, lutidine, CH2Cl2, 0 °C, 16 h (91%); (b) 20, Pd(dppf)Cl2 (5
mol%), K2CO3, THF/H2O (10:1), reflux, 2.5 h (91%); (c) MOMCl, i-Pr2NEt, CH2Cl2, 0 °C → rt, 18 h; (d) m-CPBA, CH2Cl2, rt, 5 h (84%, 2 steps); (e) Pd/CaCO3, H2
(1 atm), MeOH, rt, 20 h (90%); (f) 10-CSA, MeOH, rt, 1 d (97%); (g) (EtO)3CH, TFA, rt, 18 h (99%); (h) Bu2SnO (1 mol%), TMSE-OH, PhMe, 100 °C, 16 h (86%);
(i) MOMCl, i-Pr2NEt, CH2Cl2, 0 °C → rt, 24 h; (j) m-CPBA, CH2Cl2, rt, 4 h; (k) Pd/CaCO3 (cat.), H2 (1 atm), MeOH, rt, 24 h; (l) HCl/MeOH, rt, 16 h (70%, four
steps); (m) Alcaligenes sp. lipases (10 weight%), vinyl acetate, MTBE, 4Å MS, rt, 7 d; (n) DEAD, PPh3, AcOH, PhMe (0.1 M), 0 °C →rt (78%); (o) HCl/MeOH, rt,
16 h (99%).
At the time we started our synthetic studies in early 2007, the stereochemistry at the quaternary center (C22) was not assigned. In light
of that, and our desire to explore a rather bold approach as outlined in Scheme 2, we decided to perform our exploratory chemistry toward a
gem-dimethyl substituted berkelic acid analog (i.e. nor-berkelic acid). The synthesis of the required alkyne coupling partner analogous to
14 (Scheme 2) is depicted in Scheme 4. To this end, formation of the corresponding phosphonium bromide from commercially available α-
bromo ketone 29,30 followed by ylide formation and Wittig olefination with known 2-(4-methoxybenzyloxy)acetaldehyde (30)31 gave α,β-
unsaturated ketone 31 in 58% yield over three steps. Alternatively, a more efficient one-step titanium-mediated dehydrative aldol
condensation between commercially available methyl ketone 28 and aldehyde 30 yielded enone 31 in one step (69% yield). Next, a
conjugate addition of the enamine derived in situ from propanal and diethylamine to enone 31 afforded aldehyde 32 in acceptable yield but
poor diastereoselectivity (75%, dr = 2:1).32,33 The anti/syn ratio further eroded to unity during the ensuing Seyferth-Gilbert homologation of
aldehyde 32 with the Ohira-Bestmann reagent.34 NMR analysis of the chromatographically homogeneous material obtained following
oxidative deprotection with DDQ indicated that the product exists as a mixture of open-chain alkynols syn-33 and anti-33 (1:1) and 33-
lactol isomers (verified by gHMBC).35 Samples of 33 enriched in each diastereomer (anti-33:syn-33 - 14:1; anti-33:syn-33 - 1:3.8) were
obtained by repeated flash chromatography with arbitrary division of material and served to confirm anti-33 as the less polar diastereomer.
5

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OHC
OPMB
CHO
a-c (from 29),
or d (from 28)
O
O
O
e
MeO2C
OPMB
MeO2C
MeO2C
+
58-69%
75%
anti:syn = 2:1
30
Me Me
Me Me
31
Me Me
X
OPMB
Li
Me
O
28: X = H
29: X = Br
HO
MeO2C
32
k
h
Ti
TMS
f, g
Me Me
lactol-33
TMS
H
Me
anti:syn anti:syn
> 20:1 > 8:1
70%
38
34
anti:syn
= 1:1
Me
Me
(S,S)-35
(±)-35
TMS
59% ee
Me
OPMB
OH
OH
Me
O
R
O
O
j
OP(O)(OEt)2
MeO2C
MeO2C
MeO2C
73% (from 31)
anti:syn = 10:1
37
Me Me
Me Me
anti-33
Me Me
syn-33
35: R = TMS
36: R = H
Me
Me
i
Scheme 4. Synthesis of gem-dimethyl substituted alkynol 33. Reagents and conditions: (a) 29, PPh3, THF, reflux, 1 h (71%); (b) NaOH, MeOH, rt, 2 h (c) 30, PhMe,
reflux, 4 h (81%, 2 steps); (d) 28, TiCl4, THF, 0 °C, 30 min; Et3N, –78 °C, 45 min; 30 (1.2 eq), –78 °C → rt, 2.5 h (69%); (e) EtCHO, Et2NH, dioxane, rt, 21 h (75%,
dr = 2:1); (f) Ohira-Bestmann reagent, K2CO3, MeOH, 0 °C, 30 min, rt, 17 h (78%, dr = 1:1); (g) DDQ, CH2Cl2:H2O (10:1), rt, 3.5 h (90%, dr = 1:1); (h) preform
cuprate with 34, CuBr·DMS, THF, –78 °C, 1 h; then add to 31, THF, –78 °C, 2 h; (i) K2CO3, MeOH, rt, 4 h; (j) DDQ, CH2Cl2, H2O, rt, 2.5 h (73%, three steps); (k)
37, Ti(Oi-Pr)4, i-PrMgCl, Et2O, –45 °C, 2 h; then 31, –45 °C, 15 min → –20 °C, 3 h (54%).
The above enamine-based route was designed with the prospect to eventually control diastereo- and facial selectivity using an
enantioselective organocatalytic variant. However, the ease of epimerization of the aldehyde α-stereocenter during the subsequent
alkynylation forced us to consider an alternative approach. Conceptually, 1,4-addition of a metalated propargyl/allenyl species to enone 31
should deliver an α-methyl substituted alkyne straightforwardly. Although initial exploration of reagents more commonly used for the
propargylation of aldehydes (Marshall chemistry,36 allenylstannanes,37 allenylboronic acids,38 allenylzinc species,39 and allenyl/propargyl
lithiates)40 proved frustrating (no Michael addition products 35 or 36), traction was gained when moving to cuprate chemistry. After
substantial experimentation, we found that addition of 4 equivalents of a cuprate generated from lithiate 34 and CuBr·DMS to enone 31 in
THF at –78 °C yielded racemic TMS-alkyne 35 with the desired anti-isomer dominating in a ratio of ~8-10:1.41,42 In contemplating an
enantioselective route, the synthesis of configurationally defined and stable allenyl/propargyl cuprate species from propargyllithium species
34 appeared daunting.43 Ultimately, we encountered a manuscript disclosed by Sato and coworkers describing the conjugate addition of
configurationally stable homochiral allenyltitanium species to alkylidenemalonates.44 Using their conditions, we found that treatment of 31
with allenyltitanium reagent 38, derived from 37 (>98% ee),45 provided (S,S)-35 in moderate yield and enantiomeric excess (54% yield; ee
= 59%)46 but excellent diastereoselectivity (anti:syn > 20:1).47 Although 31 was completely consumed, a mixture of byproducts including
the 1,2-addition product (~20%) was formed, leading to the diminished yield with respect to copper-mediated conjugate propargylation.
Treatment of the crude racemic alkyne (±)-35 with methanolic base to remove the TMS (→ 36) was followed by oxidative PMB-removal to
yield the final alkyne (±)-anti-33 in 73% yield from enone 31 (dr = 10:1). Enantioenriched (S,S)-35 on the other hand provided
enantioenriched (S,S)-anti-33 in 92% yield (2 steps; 59% ee; dr >20:1).
As noted above, our initial approach to berkelic acid entailed the addition of a spicifernin-like alkyne to an electrophilic pulvilloric acid-
like synthon, followed by a final cyclopropargylation/ cycloisomerization of the thus formed linear alkyne coupled intermediate (Scheme 2,
path a). As a prelude to explore this coupling reaction, we attempted to protect the free alcohol of model alkynol 33 (anti:syn = 1:1) as a
methyl ether 39 (or acetal 40) with Ag2O and MeI (Scheme 5A).48 Instead, we obtained a crude mixture containing compounds consistent
with structure 41 (diastereomers) and some endocyclic enol ether 42 (crude NMR, not isolated, assignment tentative). It was this
observation that led us to explore the possibility for a 1-pot metal-mediated cascade entailing: (1) cycloisomerization of an alkynol, (2)
dearomatization of a lactol to a quinone methide, and (3) in situ cycloaddition to deliver the berkelic acid scaffold (Scheme 2, path b). After
6

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a brief survey of transition metal sources,49 we found that AgSbF6 stroke an ideal balance in effecting both the alkynol cycloisomerization
and the quinone methide formation from lactol and acetal precursors, respectively (vide infra). As shown in Scheme 5A, in situ NMR
monitoring of a solution of 33 (anti:syn = 1.1:1) and AgSbF6 (1.4 eq) in d10-Et2O showed 100% conversion to a mixture of compounds
featuring resonances consistent with enol ether 41 within 5 min (no signals indicative of 42). When using sub-stoichiometric AgSbF6 (0.3
eq) a far more complex mixture was observed after 5 min that simplified over time (≤2.5 h) to a mixture similar to that observed with
supra-stoichiometric AgSbF6 but significant accumulation of endocyclic enol ether 42. Although it has been reported that the carboxylic
acid corresponding to 24 will yield the quinone methide pulvilloric acid (4) upon removal of ethanol under ultra high vacuum,14c we opted
to explore the possibility of generating quinone methide 43 in situ via a Lewis acid catalyzed dearomatization.50 Gratifyingly, through
1
titration experiments in CD2Cl2 monitored by H NMR, we found that the alkynophilic AgSbF6, effective for the cycloisomerization of
alkynol 33, enjoyed sufficient hard Lewis acidity to promote a concentration-dependent equilibrium between (±)-24 and quinone methide
43 (Scheme 5B).
Scheme 5. One-pot Ag-catalyzed synthesis of nor-berkelic acid methyl esters (44/45) from alkynol (±)-33 and lactol (±)-24.
With the above defined parameters in hand, we were more than pleased to observe that stirring an ethereal solution of racemic alkynol
33 (anti:syn = 1.85:1) and racemic lactol 24 in the presence of AgSbF6 (2.5 eq.) at room temperature cleanly converted to a mixture of five
methyl berkelates 44 (~60% of total) and 45a-d (~40% of total, 2.5:1.0:1.5:1.2 ratio) comprising >95% of the aromatic containing products
(Scheme 5C, entry 1).51,52 The same results were obtained with an equimolar mixture of diastereomerically enriched (±)-anti-33 (anti:syn =
14:1) and (±)-24 (Scheme 5C, entry 2). Repeated semi-preparative HPLC yielded pure samples of (±)-44 (60% isolated) and the four
individual diastereomers 45a-d, the structures of which were unambiguously assigned by X-ray crystallography (44 and 45a-c) and
multidimensional NMR studies (45d; Figure 2). Diastereomer 44 corresponds to the revised structure of berkelic acid, whereas
7

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diastereomers 45a-d correspond to the equilibrating diastereomers identified by Fürstner when attempting to synthesize the originally
assigned structure of berkelic acid.4,5 In agreement, when individual diastereomer 45a (originally assigned berkelate stereochemistry) was
stirred at room temperature with HCl/MeOH, an equilibrium was established yielding a mixture of diastereomers 45a-d in the same
2.5:1.0:1.5:1.2 ratio as observed by NMR of the crude reaction mixture obtained according to Scheme 5C.53 These and other published
data,3-8 thus establish that the C15, C17 and C18 stereocenters can epimerize under the reaction conditions, and therefore, theoretically
either anti- or syn-33 could provide access to the correctly configured diastereomer 44. To investigate this idea, a mixture of (±)-24 (1.0
eq), AgSbF6 (1.5 eq), and (±)-syn-33 (1.1 eq, syn:anti = 3.8:1) in Et2O was stirred at room temperature, which led to a complex mixture
comprised of ~25% berkelates in addition to previously unobserved, non-berkelate byproducts (crude NMR, Scheme 5C, entry 3).
However, pre-mixing an ethereal solution of syn-33 (dr = 3.8:1, 1.3 eq) and AgSbF6 (2.0 eq) for 30 min prior to the introduction of (±)-24
(1.0 eq) generated a mixture similar (~6:4, ~80% of mixture, crude NMR) to that observed previously with alkynol anti-33 (Scheme 5C,
entry 4). These results are consistent with a 5-exo-dig hydroalkoxylation in which alkynol anti-33 undergoes cyclization at a rate
competitive with berkelate formation, while syn-33 does not. This is likely to the result of a build-up of an unfavorable eclipsing interaction
during the cycloisomerization of syn-33 versus anti-33. Pre-incubating syn-33 with AgSbF6 for 30 min enables a more advanced conversion
to in situ formed cyclic enol ether 41 prior to the addition of acetal 24.
CO2Me
OH
CO2Me
OH
CO2Me
OH
CO2Me
OH
CO2Me
OH
O
O
O
O
O
O
O
O
O
O
O
O
O
O
O
R
R
R
R
R
Me
Me
Me
Me
Me
rac-44
45a
45b
45c
45d
C5H11
C5H11
C5H11
C5H11
C5H11
Figure 2. Representations of X-ray crystal structures of 44 and 45a-c and NMR structure of 45d (diagnostic nOe correlations are indicated with blue arrows).
In an attempt to render the reaction catalytic, we were surprised to find that stirring alkynol (±)-33 (anti:syn = 2.7:1; 1.2 eq.) and acetal
(±)-24 in the presence of a substoichiometric amount of AgSbF6 (0.4 eq.) generated an inseparable mixture of 3 fused tetracyclic ring
products (60% yield)54 from which the major endo-isomer 46 was crystalized by slow evaporation from a EtOAc/hexanes/dichloromethane
solution. X-ray crystallographic analysis unambiguously determined the structure to be the one shown in Scheme 6. Of the two conceivable
pathways, interception of the quinone methide 43 derived from acetal 24 with either a silver enolate 47 derived from β-keto ester 33 or the
endocyclic enol ether 48 derived from a silver-mediated dehydration of lactol-33, we prefer the silver enolate pathway (via 47). Indeed,
under otherwise identical conditions, suprastoichiometric AgSbF6 provided spirocyclic structures (44/45) via 33-derived exocyclic enol
ether 41 (Scheme 5C), but the hydroxymethylene required for this cycloisomerization (33 → 41) would already be sequestered if the
reaction proceeds through endocyclic enol 48.55 Moreover, given the bidentate nature of β-keto esters it is not unreasonable to envision that
33 would sequester the first equivalent of Ag(I) as the silver enolate 47, leading to fused products (e.g. 46) in reaction with quinone
methide 43. In agreement with our in situ NMR-studies (Scheme 5A) further cycloisomerization of the γ-hydroxy alkynol in 47 to an
8

ACCEPTED MANUSCRIPT
exocyclic enol ether is slow with substoichiometric AgSbF6 possible due to a decreased alkynophilicity of bidentate-sequestered Ag (cf.
47). Longer incubation times or suprastoichiometric AgSbF6 (Scheme 5A) are thus required to complete the cycloisomerization to
exocyclic enol ether 41 en route to the spirocyclic berkelate products 44/45 (Scheme 5C).56
Ag
OH
Me
O
O
MeO
Me Me
OH
Me
O
47
MeO2C
Me Me
anti-33
Me
AgSbF6 (0.4 eq.), Et2O, rt, 13 h
Me
O
CO2Me
OH
MeO2C
O
CO2Me
HO
EtO
OH
H
anti:syn =
2.7:1
H
O
Me
C5H11
O
HO
O
46
MeO2C
Me
nC5H11
(±)-24
+ 2 other diastereomers
Me
lactol-33
Me
O
MeO2C
Me
X-ray structure of 46
Me
Me
48
Scheme 6. Reaction of alkynol (±)-33 and lactol (±)-24 with catalytic AgSbF6 produces fused instead of spirocyclic products.
As noted in the introduction, attempts by Fürstner and coworkers to selectively hydrolyze the phenolic methyl ester of berkelic acid
methyl ester were unsuccessful. They, and later Snider and coworkers, solved this issue by resorting to orthogonal ester protecting groups
(phenolic benzyl and allyl ester, respectively).3-8 We decided to reexamine the issue of selective demethylation and after some
experimentation found that heating a solution of nor-berkelic acid methyl ester 44 with excess (Bu3Sn)2O in toluene provided a mixture of
the desired nor-berkelic acid 49 (42% isolated) and a decarboxylated byproduct 50 (34% isolated) in a ratio of ~1.5:1 (based on crude
NMR, Scheme 7).57 Byproduct 50 arises from double methyl ester cleavage and spontaneous decarboxylation at the C28 terminus.
O
O
MeO2C
Me
MeO2C
Me
CO2Me
OH
CO2H
O
H
O
H
O
O
OH
Me
Me
Me
Me
(Bu3Sn)2O, PhMe,
115 C, 8 h
O
H
O
H
49
(±)-44
(42% isolated)
C5H11
C5H11
CO2H
~1.5:1 ratio
(crude NMR)
O
Me
O
H
O
OH
Me
Me
O
H
50
(34% isolated)
X-ray structure of nor-berkelic acid 49
C5H11
Scheme 7. Synthesis of nor-berkelic acid 49 via chemoselective methyl ester cleavage.
With a convergent streamlined synthetic strategy established, we set out to tackle the total synthesis of berkelic acid containing the
quaternary C22-stereocenter. To that end, we decided to exploit the asymmetric enamine alkylation method developed by Koga and
coworkers to install the C22-quaternary stereocenter.58 Accordingly, valine-derived enamine 52 was prepared by Lewis acid-catalyzed
condensation of commercially available 2-ethyl-3-oxobutanoate (51) and (L)-t-butyl valinate in 82% yield (Scheme 8). Sequential lithiation
of a toluene solution of 52 with LDA in the presence of lithium-coordinating THF (2.6 eq), followed by alkylation with methyl iodide
9

ACCEPTED MANUSCRIPT
provided α-quaternary substituted imine derivative 53 (88%, dr > 15:1). Hydrolysis of this material gave crude β-keto ester 54, which was
converted to bromine-containing cyclic hydrazone 55 (40% yield) for single crystal X-ray analysis, thus confirming the (S)-absolute
configuration in agreement with the Koga model for stereoinduction.58 Continuing with the synthesis, titanium-mediated dehydrative aldol
condensation of β-keto ester 54 with aldehyde 30 yielded enone 56 in 42% overall yield from enamine 52 (three steps, no intermediate
purifications).59 Conjugate propargylation of 56 using the homochiral allenyl titanate (R)-38 as described in Scheme 4 afforded an
inseparable mixture of 57a,b in a ratio of 2:1 (anti:syn ˃ 20:1) and 52% yield.60 Sequential TMS-removal and oxidative PMB-deprotection
delivered alkynols 58a,b in 91% yield (2:1 ratio, inseparable, in equilibrium with corresponding hemiketals). Stirring an ethereal mixture of
alkynols 58a,b (2 eq) and enantiopure lactol (R)-24 (1 eq) in the presence of AgSF6 (3 eq) provided a mixture of methyl berkelates 59 (from
58a) and 60a-d (from 58b). Unlike the corresponding nor-berkelates 44 / 45a-d, individual diastereomers could not be obtained from this
inseparable mixture. Crude NMR-analysis indicated a ratio of berkelic acid methyl ester (59) versus four other diastereomers 60a-d in a
ratio of 3:2, similar to that observed for nor-berkelates 44 / 45a-d.61 Treatment of the crude mixture of 59 / 60a-d with (Bu3Sn)2O in
toluene provided pure (–)-berkelic acid (2) in 47% isolated yield over the two steps from lactol (R)-24. The reaction was interrupted at
partial conversion to prevent saponification and subsequent decarboxylation at the C28 terminus as was observed for nor-berkelic acid 49
(Scheme 7). The recycled material was subjected to another round of (Bu3Sn)2O treatment to yield additional (–)-berkelic acid (2) for a total
isolated yield of 59% from lactol (R)-24 based on 1 recycle. Given that at the outset of our program, the absolute configuration at the
quaternary C22-stereocenter remained in doubt, we also prepared the corresponding C22-epi-berkelic acid 61 from the enantiomeric
alkynols ent-58a,b (53%, 70% after 1 recycle from (R)-24).62 Only the C22-S diastereomer 2 displayed spectral data fully congruent with
natural (–)-berkelic acid (2),1 thus unambiguously establishing the complete stereochemical configuration of the natural product. The
spectral data and optical rotation of this synthetic (–)-berkelic acid (2) is in full agreement with the data reported for natural and synthetic (–
)-berkelic acid from the Snider, Fürstner, and Fañanàs groups.1,3,5,11 The spectral data for C22-epi-berkelic acid (61) matched those reported
for the same material synthesized by Snider and• Fürstner.3,5
Br
CO2tBu
iPr
CO2tBu
iPr
O
HN
N
O
MeO2C
MeO2C
MeO2C
Me
MeO2C
a
b
c
e
N
N
Me
Me
Me
Me
Me
Me
crude
O2N
Me
Me
Me
O
82%
dr > 15:1
40%
Me
Me
Me
51
52
53
54
55
88%
Ti
42%
(from 52)
d
H
TMS
OR1
Me
OR1
Me
R2
R2
38
O
O
O
Me
MeO2C
MeO2C
Me
MeO2C
f-h 50%
anti:syn = 20:1
Me
Me
+
Me
Me
Me
OPMB
57a (R1 = PMB, R2 = TMS) 57b
58a (R1 = R2 = H) 58b
56
X-ray structure of 55
inseparable (ratio 2:1)
CO2Me
O
O
O
MeO2C
Me
MeO2C
MeO2C
Me
CO2R
CO2Me
CO2H
O
HO
EtO
OH
O
O
O
i
OH
O
OH
O
OH
Me
Me
Me
Me
Me
Me
Me
ratio 59:60
~ 3:2
O
(R)-24
inseparable
mixture
H
H
O
H
O
H
O
H
nC5H11
C5H11
C5H11
C5H11
60a-d (from 58b)
61: C22-epi-berkelic acid
59: R = Me (from 58a)
2: R = H; berkelic acid
j
47% (from 24; 59% after 1 recycle)
10

ACCEPTED MANUSCRIPT
Scheme 8. Synthesis of (−)-berkelic acid (2). Reagents and conditions: (a)
L-t-Bu-valinate (0.99 eq), BF3·OEt2 (5 mol%), PhH, 80 °C, 14 h; (b) LDA, PhMe, –78 °C,
1 h; then THF (2.6 eq), 1 h; then MeI, 16 h (88%); (c) HCl/H2O, THF, rt, 1 h; (d) TiCl4, Et3N, 4Å MS, THF, –78 °C, 1.5 h; aldehyde 30 (1.2 eq) –78 °C → rt, 3 h
(48% over two steps); (e) (4-bromo-2-nitrophenyl)hydrazine hydrochloride, EtOH, reflux, 2 d (40%); (f) (R)-38 (1.2 eq), THF, –40 °C → –20 °C, 3 h (52%); (g)
K2CO3, MeOH, rt, 4 h; (h) DDQ, CH2Cl2, H2O, rt, 2.5 h (91%, two steps); (i) 58a,b (dr 2:1, 2.0 eq.), (R)-24 (1.0 eq), AgSbF6 (3.0 eq), Et2O, rt, 4 h; (j) (Bu3Sn)2O,
PhMe, 115 °C, 8 h (47% over two steps; 59% after 1 recycle of recovered 59/60).
As noted in the introduction, berkelic acid (2) appears to be comprised of two other natural products, namely spicifernin (3) and the
quinone methide pulvilloric acid (4), with the caveat that the C18 and C19 stereocenters (berkelic acid numbering, see Fig. 1) of spicifernin
(3) are of opposite configuration to those found in berkelic acid (2) and a lower oxidation state at C26. Snider had proposed the possibility
of a biosynthetic transformation of spicifernin (3) to bis-epi-deoxyspicifernin (6, Scheme 1), followed by incorporation into berkelic acid
(2). We offered the possibility of incorporating spicifernin (3) first into bis-epi-oxoberkelic acid (8), followed by epimerization (C18 and
C19) and reduction at C26 (Scheme 1). We, and others had already demonstrated that C15, C17, and C18 are all epimerizable within the
context of the berkelic acid tetracyclic scaffold.5,10 The higher oxidation state at C26 (oxo) is thus expected to enable the required additional
epimerization at C19 to complete the berkelic acid stereochemical constellation. To test this hypothesis, we embarked on the synthesis of
C26-oxoberkelate analogs with C22-gem dimethyl-substituted quaternary carbon (63, Scheme 9) using the silver mediated cycloaddition
strategy employed for berkelic acid synthesis (Schemes 5C and 8).
O
OH
O
O
O
O
O
MeO2C
MeO2C
MeO2C
Me
a-c
d
CO2Me
OH
CO2Me
OH
O
H
HO
EtO
O
Me Me
(±)-33
Me Me
Me
Me
60%
50%
Me
62
Me
anti:syn = 10:1
O
O
H
63a-e
nC5H11
C5H11
(±)-24
O
O
O
O
O
O
O
O
O
O
CO2Me
CO2Me
OH
CO2Me
OH
CO2Me
CO2Me
OH
O
O
H
O
O
O
H
R
R
R
R
R
O
OH
O
O
O
OH
O
Me
Me
Me
Me
Me
H
H
H
O
H
O
H
O
H
O
H
O
H
63a
63b
63c
63d
63e
C5H11
C5H11
C5H11
C5H11
C5H11
O
O
f
O
O
e
CO2Me
OH
CO2Me
OH
~1:1
O
H
O
R
R
40%
67%
O
O
O
MeO2C
Me
R =
Me
Me
Me
O
H
HO
H
63f
64
C5H11
C5H11
Scheme 9. Synthesis of oxoberkelates 63a-f. Reagents and conditions: (a) DMP, CH2Cl2, rt, 1 h; (b) NaH2PO4·H2O, NaHClO2·H2O, 2-methyl-2-butene, t-BuOH,
H2O, 0 °C → rt, 1 h (90%, two steps); (c) NaAuCl4·2H2O (5 mol%), MeCN, H2O, rt, 2.5 h (67%); (d) (±)-24 (1.0 eq), (±)-62 (1.2 eq), AgSbF6 (1.4 eq), Et2O, rt, 3 h
(50% 63a-e); (e) HCl, Et2O, CH2Cl2, rt, 24 h (67%); (f) AgSbF6 (1 eq), CD2Cl2, rt, 3 h (40%, 63f). The structure of 63b could not be determined unambiguously.
The requisite spicifernin-like γ-lactone 62 was prepared from nor-berkelic acid precursor 33 (racemic) in 60% overall yield by double
oxidation (Dess-Martin and Lindgren)63,64 and gold-catalyzed cycloisomerization.65 Treatment of racemic lactone 62 (1.2 eq) with racemic
lactol 24 (1 eq) in the presence of AgSbF6 (1.4 eq) at room temperature in ether for 3 hours produced a mixture containing at least six
oxoberkelate diastereomers 63a-f in 61% yield after flash chromatography. Normal phase semi-preparative HPLC purification with two
different eluents yielded pure diastereomers 63a (4% isolated), 63b (4% isolated), 63c (13% isolated), 63d (9% isolated), and 63e (2%
11

ACCEPTED MANUSCRIPT
isolated).66 Diastereomer 63e has the same relative configuration as berkelic acid.67 Of all the other diastereomers, only 63c was obtained in
quantities sufficient enough for equilibration studies. Unfortunately, attempts to equilibrate this diastereomer to 63e (with berkelic acid
configuration) have thus far been unsuccessful. For example, treatment of 63c with Bronsted acid (HCl, CH2Cl2) – conditions that were
effective for the equilibration of berkelate 45a into the 4 berkelate diastereomers 45a-d (Scheme 4) – resulted in isochroman ring-opening
to yield elimination product 64, whereas treatment with AgSbF6 yielded a 1:1 equilibrium mixture of 63c (configuration similar to 45d) and
63f (configuration similar to 45c) from which 63f could be isolated in 40% yield. No equilibration via epimerization at C17, C18, or C19
could be detected. While these initial results do not disprove our biosynthetic hypothesis involving oxoberkelic acid intermediates en route
to berkelic acid, our inability to identify suitable epimerization conditions appears to bolster the proposal by Snider that epimerization at
C18 and C19 occurs before incorporation of spicifernin into berkelic acid.
During the course of our synthetic campaign toward (–)-berkelic acid (2) described herein, we prepared a number of structural analogs –
notably nor-berkelic acid (49), a decarboxylated nor-berkelic acid analog 50, nor-berkelic acid methyl ester (44) and four diastereomers
45a-d, C22-epi-berkelic acid 61, and six diastereomeric oxoberkelic acid methyl ester analogs 63a-f. In the original isolation paper,
berkelic acid (2) was shown to be a selective inhibitor of the human ovarian cancer cell line OVCAR-3 with a GI50 of 91 nM.1 Later, Snider
reported that synthetic (–)-berkelic acid (2) and C22-epi-berkelic acid (61) failed to register any antiproliferative activity when tested at a10
µM single concentration in the NCI human disease-oriented 60-cell line panel, which includes the OVCAR-3 cell line.3 Recently Rodríguez
et al. reported similarly that their synthetic (–)-berkelic acid (2) and corresponding methyl ester 59 were devoid of activity against the
OVCAR-3 cell line.68 We therefore decided to test our synthetic (–)-berkelic acid (2), C22-epi-berkelic acid (61), nor-berkelic acid (49),
decarboxylated nor-berkelic acid (50), nor-berkelic acid methyl ester (44) and diastereomer 45a, and oxoberkelic acid methyl ester
diastereomer 63c against the HeLa (cervical cancer), A549 (adenocarcinomic human alveolar basal epithelial cells), and OVCAR-3 cancer
cell lines. Unfortunately, none of these compounds demonstrated any activity when measured up to 10µM concentrations. These results in
combination with previous literature observations indicate that the original sample of natural (–)-berkelic acid (2) was either contaminated
with a minor potent metabolite, or that there was a methodological problem with the assay.69
3. Conclusion
In conclusion, we have developed a scalable, highly convergent synthesis of berkelic acid and analogs based on a key silver-mediated
cascade involving the cycloisomerization of an alkynol to an exocyclic enol ether, quinone methide formation from an aromatic isochroman
lactol, and subsequent cycloaddition to provide berkelic acid methyl ester in a one-pot operation. This highly efficient approach, inspired by
the notion that berkelic acid constitutionally appeared as a combination of two other natural products spicifernin and pulvilloric acid,
enabled the total synthesis of (–)-berkelic acid (2) in 10 steps overall longest linear sequence (LLS, 17 total steps) and 10% overall yield.
This compares favorable to the total synthesis described by Fürstner (19 steps LLS, 26 total steps, 5% overall yield),5 Snider (13 steps LLS,
20 total steps, 2% overall yield),3 and Fañanás (10 steps LLS, 18 total steps, 12% overall yield).11 Furthermore, we observed that
substoichiometric amounts of AgSbF6 during the key-step led to fused tetracyclic products instead of the tetracyclic
spiroketal/chroman/isochroman ring system present in berkelic acid. In addition to berkelic acid and its C22-epimer, we also synthesized
several analogs including nor-berkelic acid, a decarboxylated nor-berkelic acid analog, nor-berkelic acid methyl ester and four
diastereomers, and six diastereomeric oxoberkelic acid analogs. The latter were prepared to explore an alternative to Snider’s biosynthetic
proposal to reconcile the opposite C18/C19 configuration of spicifernin to that found in berkelic acid.3,18 Our studies with nor-berkelic acid
analogs (quaternary gem-dimethyl at C22, versus quaternary Me, Et) enabled us to obtain X-ray quality crystal structures of naturally
configured nor-berkelic acid and several diasteromeric methyl esters for the first time. These unambiguously confirm the observations by
Fürstner4 and Snider3 that the originally assigned configuration of berkelic acid1 is not a thermodynamically stable entity and readily
12

ACCEPTED MANUSCRIPT
equilibrates into a more or less statistical mixture of four diastereomers. Finally, we tested berkelic acid, C22-epi-berkelic acid and several
analogs for activity against several human cancer cell lines, and found that in agreement with previous observations, synthetic berkelic acid
and the various analogs were inactive. This indicates that the natural sample of berkelic acid contained a minor potent contaminant, or a
methodological problem with the bioassay.
Conflict of interest
The authors declare no conflict of interest.
Acknowledgements
This work was supported by the Robert A. Welch Foundation (I-1422). C.F.B. thanks the NIH for a postdoctoral fellowship
(T32CA12433401). We thank Dr. Radha Akella for X-ray analysis of 44 and 46. X-ray data for 55, 45a-c, and 49 were collected using an
instrument purchased with funds provided by the NSF (0741973). We thank Clinton Taylor and Prof. Noelle Williams (UTSW) for
biological testing. J. K. De Brabander holds the Julie and Louis Beecherl, Jr., Chair in Medical Science.
Appendix A – Supplementary Data
Supplementary data associated with this article can be found in the online version, at
CCDC 1571417 (44, C29H40O9), CCDC 1571415 (45a, C29H40O9), CCDC 1571419 (45b, C29H40O9), CCDC 1571416 (45c,
C29H40O9), CCDC 1571420 (46, C29H38O8) and CCDC 1571418 (49, C28H38O9) contain the supplementary crystallographic data for
this paper. These data can be obtained free of charge from the Cambridge Crystallographic Data Centre via
www.ccdc.cam.ac.uk/data_request/cif.
References and notes
1
Stierle, A. A.; Stierle, D. B.; Kelley, K. J. Org. Chem. 2006, 71, 5357–5360.
2
Montana Bureau of Mines and Geology, Berkeley Pit and Butte Mine-Flooding Operable Unit.
http://www.mbmg.mtech.edu/env/env-berkeley.asp (accessed December 22, 2009).
3
Wu, X.; Zhou, J.; Snider, B. B. J. Org. Chem. 2009, 74, 6245–6252.
4
Buchgraber, P.; Snaddon, T. N.; Wirtz, C.; Mynott, R.; Goddard, R.; Fürstner, A. Angew. Chem. Int. Ed. 2008, 47,
8450–8454.
5
Snaddon, T. N.; Buchgraber, P.; Schulthoff, S.; Wirtz, C.; Mynott, R.; Fürstner, A. Chem. Eur. J. 2010, 16, 12133–
12140.
6
Snider et al. previously demonstrated that the C15 and C17 stereocenters could be epimerized under acidic conditions
in a model for the chroman/isochroman/spiroketal core lacking both C18 and C19 substituents; see: Zhou, J.; Snider,
B. B. Org. Lett. 2007, 9, 2071–2074.
13

ACCEPTED MANUSCRIPT
7
Huang and Pettus were first to demonstrate a thermodynamically unfavorable eclipsing interaction of a C18-Me
substituent in a tricyclic chroman/spiroketal model system (lacking the isochroman ring) with relative configuration at
C17 and C18 as originally proposed for natural berkelic acid, leading to epimerization at the spiroketal C17-center
under acidic conditions; see: Huang, Y.; Pettus, T. R. R. Synlett 2008, 1353–1356.
8
9
Wu, X.; Zhou, J.; Snider, B. B., Angew. Chem. Int. Ed. 2009, 48, 1283–1286.
Kiyooka, S.-I.; Maeda, H.; Hena, M. A.; Uchida, M.; Kim, C.-S.; Horiike, M. Tetrahedron Lett. 1998, 39, 8287–8290.
10 Bender, C. F.; Yoshimoto, F. K.; Paradise C. L.; De Brabander, J. K. J. Am. Chem. Soc. 2009, 131, 11350–11352.
11 Fañanás, F. J.; Mendoza, A; Arto, T.; Temelli, B.; Rodríguez, F. Angew. Chem. Int. Ed. 2012, 51, 4930–4933.
12 For formal total syntheses, see: (a) Wenderski, T. A.; Marsini, M. A.; Pettus, T. R. R. Org. Lett. 2011, 13, 118–121; (b)
McLeod, M. C.; Wilson, Z. E.; Brimble, M. A. Org. Lett. 2011, 13, 5382–5385; (c) McLeod, M. C.; Wilson, Z. E.;
Brimble, M. A. J. Org. Chem. 2012, 77, 400–416.
13 For model studies and fragment synthesis, see references 6 and 7, and: (a) Marsini, M. A.; Huang, Y.; Lindsey, C. C.;
Wu, K.-L.; Pettus, T. R. R. Org. Lett. 2008, 10, 1477–1480; (b) Stiasni, N.; Hiersemann, M. Synlett 2009, 2132–2136;
(c) Barluenga, J.; Mendoza, A.; Rodríguez, F.; Fañanás, F. J. Angew. Chem. Int. Ed. 2009, 48, 1644–1647; (d)
Wilson, Z. E.; Brimble, M. A. Org. Biomol. Chem. 2010, 8, 1284–1286; (e) Brimble, M. A.; Haym, I.; Sperry, J.; Furkert,
D. P. Org. Lett. 2012, 14, 5820–5823.
14 Isolation: (a) Brian, P. W.; Curtis, P. J.; Hemming, H. G.; Norris, G. L. F. Trans. Br. Mycol. Soc. 1956, 40, 369–374; (b)
Barber, J.; Bradley, S. M.; Pope, S. M. Mycotox. Res. 1986, 2, 25–32; Synthesis and absolute configuration: (c) Rödel,
T.; Gerlach, H. Liebigs Ann./Recueil 1997, 213–216.
15 (a) Nakajima, H.; Fujimoto, H.; Matsumoto, R.; Hamasaki, T. J. Org. Chem. 1993, 58, 4526–4528; (b) Nakajima, H.;
Nakamura, S.; Fujimoto, H.; Baba, T., K.; Hamasaki, T. J. Chem. Soc., Perkin Trans. 1, 1994, 1865–1869.
16 Hamasaki proposes a cyclic common intermediate (ref. 15), whereas Snider has provided a reinterpretation starting
from an acyclic common intermediate (ref. 3).
17 Given that synthetic studies have demonstrated that the C18 stereocenter of berkelic acid is epimerizable under acidic
conditions,4,5,8 a C19-epi-spicifernin would also present a viable precursor towards berkelic acid. One can also not
exclude that spicifernin exists as a pair of anti-diastereomers (C18, C19) and that the C18, C19-bis-epimer of 3
(precursor to 6) has been overlooked during the isolation studies.
18 Razzak, M.; De Brabander, J. K. Nat. Chem. Biol. 2011, 7, 865–875.
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19 A related proposal for the biosynthesis of cytosporolides A-C from a formal cycloaddition between the caryophyllene
derived natural product fuscoatrol and the ortho-quinone methide derived from natural product CJ-12,373 (identical to
pulvilloric acid except for carrying a heptyl instead of pentyl side-chain) has been formulated by: Spence, J. T. J.;
George, J. H. Org. Lett. 2011, 13, 5318–5321.
20 Liu, B.; De Brabander, J. K. Org. Lett. 2006, 8, 4907–4910.
21 (a) De Brabander, J. K.; Liu, B.; Qian, M. Org. Lett. 2008, 10, 2533–2536; For applications to other heterocycles, see:
(b) Liang, Q.; De Brabander, J. K. Tetrahedron 2011, 67, 5046–5053; (c) Paradise, C. L.; Sarkar, P. R.; Razzak, M.;
De Brabander, J. K. Org. Biomol. Chem. 2011, 9, 4017–4020.
22 Liang, Q.; Qian, M.; Razzak, M.; De Brabander, J. K. Chem. Asian J. 2011, 6, 1958–1960.
23 Barluenga et al. (ref. 13c) described a palladium-catalyzed multicomponent reaction between an alkynol, ortho-
hydroxybenzaldehyde, and orthoformate for the synthesis of spirocyclic chromans that was relevant towards the
approach to berkelic acid by Fañanás et al. (ref 11). Although the individual precursors differ, both the Fañanás
synthesis and ours proceed via a similar formal cycloaddition between in situ generated enolether and ortho-quinone
methide partners.
24 Molander, G. A.; Bernadi, C. R. J. Org. Chem. 2002, 67, 8424–8429.
25 We previously reported the use of the corresponding trifluoroborate for cross coupling with an aryl triflate en route to
the synthesis of gustastatin. However, this reagent failed to perform in the cross coupling with triflate 19. See: García-
Fortanet, J; Debergh, J. R.; De Brabander, J. K. Org. Lett. 2005, 7, 685–688.
26 For a similar sequence, see ref. 25.
27 For the use of this enzyme for the resolution of benzylic and homobenzylic alcohols, see: Naemura, K.; Murata, M.;
Tanaka, R.; Yano, M.; Hirose, K.; Tobe, Y. Tetrahedron: Asymmetry 1996, 7, 3285–3295.
28 Transesterification to an allyl benzoate was also successful, but would prove incompatible with later steps in the
sequence.
29 This is likely a result of the stereoelectronic similarities between TMSE esters and methoxyethyl esters, the latter
being employed as a carboxylate protecting group when enzymatic hydrolysis is the desired means of removal. For a
review of the enzymatic deprotection of methoxyethyl ester, see: Gewehr, M.; Kunz, H. Synthesis 1997, 1499–1511.
30 Contour-Galcera, M.-O.; Poitout, L.; Moinet, C.; Morgan, B.; Gordon, T.; Roubert, P.; Thiurieau, C. Bioorg. Med.
Chem. Lett. 2001, 11, 741–745.
31 Chenevert, R.; Dasser, M. J. Org. Chem. 2000, 65, 4529–4531.
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32 (a) Hagiwara, H.; Okabe, T.; Hakoda, K.; Hosji, T.; Ono, H.; Kamat, V. P.; Suzuki, T.; Ando, M. Tetrahedron Lett.
2001, 42, 2705–2707; (b) Hagiwara, H.; Okabe, T.; Ono, H.; Kamat, V. P.; Hosji, T.; Suzuki, T.; Ando, M. J. Chem.
Soc., Perkin Trans. 1 2002, 895–900.
33 Monitoring of the reaction by 1H NMR in d8-dioxane indicated an initial dr of ~3:1, eroding to 2:1 at full conversion.
34 (a) Ohira, S. Synth. Commun. 1989, 19, 561–564; (b) Mueller, S.; Liepold, B.; Roth, G. J.; Bestmann, H. J. Synlett
1996, 521–522.
35 See supporting information.
36 For a review on the propargylation of aldehydes, see: Marshall, J. A. J. Org. Chem. 2007, 72, 8153–8166.
37 (a) Marshall, J. A.; Wang, X. J. Org. Chem. 1992, 57, 1242–1252; (b) Marshall, J. A.; Perkins, J. F.; Wolf, M. A. J. Org.
Chem. 1995, 60, 5556–5559.
38 (a) Ikeda, N.; Arai, I.; Yamamoto, H. J. Am. Chem. Soc. 1986, 108, 483–486; (b) Corey, E. J.; Yo, C.-M.; Lee, D.-H. J.
Am. Chem. Soc. 1990, 112, 878–879.
39 (a) Marshall, J. A.; Adams, N. D. Org. Lett. 2000, 2, 2897–2900; (b) Marino, J. P.; McClure, M. S.; Holub, D. P.;
Comasseto, J. V.; Tucci, F. C. J. Am. Chem. Soc. 2002, 124, 1664–1668.
40 Huynh, C.; Linstrumelle, G. J. Chem. Soc., Chem. Commun. 1983, 1133–1134.
41 Poor regioselectivity (1,4 vs. 1,2) was obtained using CuI (2.5:1), CuBr (2.5:1) and CuCN (5.0:1), while use of
CuBr·DMS afforded a >20:1 ratio of regioisomers as determined by 1H NMR of the crude reaction product.
42
For examples of 1,2-additions to aldehydes with similarly prepared cuprates, see: Alouane, N.; Vrancken, E.;
Mangeney, P. Synthesis 2007, 1261–1264.
43 To the best of our knowledge, there are no reported examples of enantioselective additions with allenyl/propargyl
cuprates (1,2- or 1,4). Our lab is currently exploring the use of chiral ligands for copper.
44 Song, Y.; Okamoto, S.; Sato, F. Org. Lett. 2001, 3, 3543–3545.
45 Phosphonate 37 (>98% ee) was prepared from (S)-4-(trimethylsilyl)-3-butyn-2-ol in one step according to Song et
al.,44 and is known to occur without erosion of enantiopurity. Chirality transfer of phosphonate 37 to allenyltitanium 38
is >97%.44 For the synthesis of (S)- and (R)-4-(trimethylsilyl)-3-butyn-2-ol in multigram quantities via enzymatic
resolution, see: Marshall, J. A.; Chobanian, H. Org. Synth. 2005, 82, 43–54.
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46 Using reagents 37/38 Sato reports yields of 82-96% and ee’s >92% in Michael additions to alkyl- and aryl-substituted
alkylidenemalonates.44
47 As noted in the main text, we have had no success with other allenyl organometallic species (Sn, B, Zn, In) commonly
used for the propargylic addition to aldehydes.36-39
48 Paquette, L. A.; Hu, Y.; Luxenburger, A.; bishop, R. L. J. Org. Chem. 2007, 72, 209–222.
49 Surveyed metals include AuCl3, Ag2CO3, AgSbF6, and AgPF6.
50 For a review of o-quinine methides, see: Van De Water, R. W.; Pettus, T. R. R. Tetrahedron 2002, 58, 5367–5405.
51 Monitoring the reaction by NMR (d10-Et2O) indicated the initial formation (< 5 min) of a major diastereomer different
than 44 and 45a-d, which disappeared at full conversion. This indicates that at least part of the reaction funnels
through a kinetic cycloaddition product that differs from the final observed diastereomers.
52 Lowering the reaction temperature resulted in poorer conversion and increased by-product formation. A brief survey of
solvents indicated that only benzene produced results comparable to Et2O.
53 Fürstner observed a slightly different 1.1: 1.0:1.3:1.6 ratio of diastereomers at equilibrium. His stereochemical
assignments relied on NMR studies.4,5
1
54 The crude H NMR spectra show only minor variations suggesting that the three products are simply stereoisomers.
The ratio of the compounds varies in CDCl3 at room temperature, ultimately reaching a presumed thermodynamic
mixture (ratio 1.4:1.1:1.0).
55 It is hard to envision a pathway from endocyclic enol ether 48 to exocyclic enol ether 41, merely by increasing the
amount of AgSbF6.
56 Increasing amounts of AgSbF6 would also shift the equilibrium from 33-lactol (via open chain isomer 33) to silver
enolate 47, thereby increasing the concentration of primary alcohol required for cycloisomerization to exocyclic enol
ether 41.
57 Mata, E. G.; Mascaretti, O. A. Tetrahedron Lett. 1988, 29, 6893–6896.
58 Ando, K.; Takemasa, Y.; Tomioka, K.; Koga, K. Tetrahedron 1993, 49, 1579–1588.
59 Due to volatility, 54 was not isolated but used as a crude concentrated THF solution.
60 This 2:1 ratio of 57a:57b indicates a mismatch with the chiral quaternary center given that the same reaction with the
achiral gem-dimethyl substituted substrate 31 delivered anti-35 with 59% ee (a 4:1 ratio of enantiomers, Scheme 4).
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Reaction of 56 with the achiral allenyl cuprate (as in Scheme 4) delivered a 1:1 ratio of 57a:57b but with a lower
anti:syn selectivity (5:1).
61 Full structural assignment of diastereomers 60a-d was impossible due to our inability to separate them. However, we
presume that their relative stereochemistry represents that of the four nor-berkelate diastereomers 45a-d by
comparison of the 1H NMR spectrum of the crude mixture of 59 / 60a-d to the 1H NMR spectra of pure 44 and 45a-d.
Specifically, correlation to the diagnostic phenolic and C15 1H resonances was used.
62 Alkynols ent-58a,b were prepared in a manner similar to 58a,b, but starting from D-valine derived enamine ent-52.
Full experimental details can be found in the supporting information to reference 10.
63 Dess, D. B.; Martin, J. C. J. Am. Chem. Soc. 1991, 113, 7277–7287.
64 Kraus, G. A.; Roth, B. J. Org. Chem. 1980, 45, 4825–4830.
65 Genin, E.; Toullec, P. Y.; Antoniotti, S.; Brancour, C.; Genêt, J.-P.; Michelet, V. J. Am. Chem. Soc. 2006, 128, 3112–
3113.
66 A sixth diastereomer (63f) was observable in the 1H NMR spectrum of the crude mixture but could not be isolated in
pure form. Its identity was confirmed by comparison with the NMR spectrum of pure 63f obtained from the
equilibration of diastereomer 63c.
67 The structure of oxoberkelates 63a,c-f was assigned via a combination of 1H, 13C, 1H-1H gCOSY, 1H-13C gHSQC, 1H-
13C gHMBC NMR experiments and correlation to spectra obtained for the nor-berkelic acid methyl ester diastereomers
44 and 45a-d. The structure of oxoberkelate diastereomer 62b could not be assigned unambiguously beyond the
relative C9-C15 cis-stereochemistry of the isochroman ring system.
68 Arto, T.; Sáenz de Santa-María, I.; Chiara, M.-D.; Fañanás, F. J.; Rodríguez, F. Eur. J. Org. Chem. 2016, 5876–5880.
69 The original isolation paper mentions the testing of natural berkelic acid in the NCI 60-cell line panel with selective
activity for the OVCAR-3 cell line. However, no actual data or description of the assay are included in the manuscript
or associated supporting information file.1
18