Divergent total synthesis of crassalactones B and C and evaluation of their antiproliferative activity

Article abstract of DOI:10.1016/j.tet.2015.05.040

A divergent total synthesis of cytotoxic natural products (+)-crassalactones B (2) and C (3) has been achieved by utilizing diacetone d-glucose (4) as a chiral precursor. The key steps of the synthesis of both targets 2 and 3 were a stereo-selective addit

Full text of DOI:10.1016/j.tet.2015.05.040

Tetrahedron 71 (2015) 4581e4589
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Tetrahedron
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Divergent total synthesis of crassalactones B and C and evaluation
of their antiproliferative activity
a
a
ꢀ a
ꢁ
ꢀ a
Goran Benedekovic , Ivana Kovacevic , Mirjana Popsavin , Jovana Francuz ,
a,
ꢀ b
ꢀ b
*
Vesna Kojic , Gordana Bogdanovic , Velimir Popsavin
a
ꢀ
Department of Chemistry, Biochemistry and Environmental Protection, Faculty of Sciences, University of Novi Sad, Trg Dositeja Obradovica 3, 21000
Novi Sad, Serbia
b Oncology Institute of Vojvodina, Put doktora Goldmana 4, 21204 Sremska Kamenica, Serbia
a r t i c l e i n f o
a b s t r a c t
Article history:
A divergent total synthesis of cytotoxic natural products (þ)-crassalactones B (2) and C (3) has been
Received 26 March 2015
Received in revised form 26 April 2015
Accepted 11 May 2015
achieved by utilizing diacetone D-glucose (4) as a chiral precursor. The key steps of the synthesis of both
targets 2 and 3 were a stereo-selective addition of phenyl magnesium bromide to a dialdose derivative,
a regioselective introduction of the cinnamic acid residue, and a stereospecific furano-lactone ring for-
mation by cyclocondensation of a suitable hemiacetal derivative with Meldrum’s acid. No protection is
necessary for the synthesis of the (þ)-crassalactone C (3), except the diacetonide function that is already
present in the commercially available starting material 4. Preparation of (þ)-crassalactone B (2) from the
same starting material, requires the use of a single silyl ether protecting group throughout the synthesis.
The synthesized natural products were evaluated for their in vitro antiproliferative activity against PC3,
HT29 and A549 human tumour cell lines.
Available online 15 May 2015
Keywords:
Styryl lactones
(þ)-Crassalactone B
(þ)-Crassalactone C
Polyalthia crassa
Ó 2015 Elsevier Ltd. All rights reserved.
Antitumour agents
Protecting-group-free synthesis
1. Introduction
B (2) starting from D
-glucose.8 Immediately thereafter a new total
synthesis of (þ)-crassalactones B and C starting from diacetone
The Annonaceae family consists of 120 genera and over 2000
species. Polyalthia is one of the Annonaceae genera with over 20
species distributed throughout tropical and subtropical region of
Asia. Species from the Polyalthia genus have been widely used in
traditional medicine in several Asian countries.1 Chemical constit-
uents of the Polyalthia species exhibit a variety of pharmacological
activities, such as cytotoxic,2 antimicrobial,3 antimalarial,4 and anti-
HIV.5 A number of styryl lactones have been recently isolated from
the tropical plant Polyalthia crassa.6 The bioassay-directed separa-
tion of the ethyl acetate extract of the leaves and twigs led to the
isolation of the known antitumour agent (þ)-goniofufurone
(1, Fig.1), along with several newcompounds including the 5-O- and
7-O-cinnamoyl derivatives of 1, as (þ)-crassalactones B (2) and C (3),
respectively. Their structures were elucidated on the basis of spec-
troscopic methods, while their absolute configurations were re-
solved by a (non-selective) chemical transformation of natural
(þ)-goniofufurone. We have recently reported a few approaches to
D
-glucose,9 as well as a novel total synthesis of (ꢀ)-crassalactone C
from tartaric acid10 has been reported by other groups. Finally, an
efficient semi-synthesis of both 2 and 3 starting from (þ)-goniofu-
furone has been very recently completed in our laboratory.11 Herein,
we disclose a newand concise total synthesis of (þ)-crassalactones B
(2) and C (3) starting from D-glucose. A brief study of their anti-
tumour activity against several human cancer cell lines is an addi-
tional objective of this work.
2. Results and discussion
2.1. Chemical synthesis
As both natural products 2 and 3 display a clear structural
similarity we wanted to design a divergent approach to 2 and 3
(þ)-crassalactone C (3) starting from
D
-xylose,7 as well as a pre-
liminaryaccount relatedthe first total synthesisof (þ)-crassalactone
* Corresponding author. Tel.: þ 381 21 485 27 68; fax: þ381 21 454 065; e-mail
Fig. 1. Styryl lactones from Polyalthia crassa: (þ)-goniofufurone (1) and (þ)-crassa-
lactones B (2) and C (3).
address: velimir.popsavin@dh.uns.ac.rs (V. Popsavin).
http://dx.doi.org/10.1016/j.tet.2015.05.040
0040-4020/Ó 2015 Elsevier Ltd. All rights reserved.

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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4582
from a common intermediate. It was assumed that the required
[3.3.0] bicyclic lactone core may be built up through the cyclo-
condensation of Meldrum’s acid with a protected sugar lactol
derivative.12e14 Accordingly, we designed the retrosynthetic con-
cept shown in Scheme 1. For both natural products 2 and 3, lactone
ring-removal would give rise to the lactol derivatives 2a and 3a,
respectively. These can be prepared by a number of selective
chemical transformations of the benzylic alcohol 5. Synthesis of
divergent intermediate 5 is visualized from the commercially
When diacetone-D-glucose (4) was treated with periodic acid in
dry ethyl acetate, and the crude product 4a was allowed to react
with phenyl magnesium bromide in ether, the known12,15 benzylic
alcohols 5 and 6 were obtained (Scheme 2), with -ido-derivative 6
L
as the major product (6/5 w3:1). Since only the minor diastereomer
5 has the correct configuration for a synthesis of both 2 and 3, we
were looking for a way to shift the stereomeric ratio towards 5.
Accordingly we have studied the influence of several solvents16 and
found that the best stereoselectivity in favour of -gluco-isomer 5
D
available diacetone-
D
-glucose (4) through the Grignard addition of
(5/6 w3:1) was obtained when the reaction is performed in
a mixture of anhydrous THF and toluene, in the presence of TMEDA
PhMgBr to protected dialdose 4a (Scheme 2).12,15
Scheme 1. Retrosynthetic analysis of (þ)-crassalactones B and C.
Scheme 2. Reagents and conditions: (a) H5IO6, EtOAc, rt, 24 h; (b) PhMgBr, Et2O, 0 ꢁC, 4 h, then rt 20 h, 60% of 6, 19% of 5 (both from 4); (c) PhMgBr, 1:1 THF/PhMe, TMEDA, 0 ꢁC, 4 h,
rt, 72 h, 49% of 5, 15% of 6 (both from 4); (d) PCC, H5IO6, EtOAc, 0 ꢁC, 2 h, 57%; (e) DDQ, 10:1 CH2Cl2/H2O, rt, 72 h, 79%; (f) NaBH4, L-tartaric acid, THF, ꢀ7 ꢁC/rt, 24 h for 7, 78% of 5, 7%
of 6, 4 h for 8, 92% of 11, 2% of 12; (g) NaBH4, silica gel, hexane, rt, 26 h, 86% of 5, 9% of 6; (h) cinnamoyl chloride, DMAP, CH2Cl2, 0 ꢁC, 45 min, 75%; (i) cinnamic acid, DMAP, DCC,
CH2Cl2, rt, 20 h for 4, 98% of 9, 50 h for 13, 96% of 14; (j) H5IO6, EtOAc, rt, 1.5 h, 93%; (k) (i) PhMgBr, THF, PhMe, 0 ꢁC, 3 h; (ii) PCC, CH2Cl2, reflux, 4 h, 34% from 2 steps; (l) BuPh2SiCl,
imdazole, CH2Cl2, rt, 48 h, 97% from 5, 90% from 11.

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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4583
as a N-donor ligand.17 The combined yield of stereoisomers 5 and 6
was 64%. Somewhat lower selectivity (5/6 w2:1), but a higher
combined yield of 5 and 6 (80%) was achieved when the Grignard
reaction was performed in THF as a solvent. Additional efforts to
improve the epimeric ratio in favour of 5 possessing the correct
stereochemistry for 2 and 3 were unsuccessful. However, a more
efficient access to 5 was possible by using an alternative two-step
sequence that involved selective oxidation of the benzylic hy-
droxyl function in 6 followed by stereoselective reduction of the
resulting prochiral ketone 7. Thus, oxidation of 6 under the Hunsen
conditions18 gave the corresponding ketone 7 that was obtained in
57% yield. However, a DDQ-oxidation of 6 gave ketone 7 in 79%
yield. Stereocontrolled reduction of the ketone functionality in 7
was achieved using the method of Yatagai and Ohnuki for prochiral
in 11 from taking part in ensuing reactions, it was blocked as a silyl
ether by treatment of 11 with tert-butyldiphenylsilyl chloride to
give the key intermediate 14 in 90% yield (31% overall yield from 4).
An alternative and more efficient route to the intermediate 14
started with selective silylation of the benzylic hydroxyl group in 5
to give the corresponding silyl ether 13 in 97% yield. Further es-
terification of 13 with cinnamic acid (DCC, DMAP) gave the key
chiral intermediate 14 in 96% yield. Thus, the six-step sequence
based on 13 as an intermediate represents a more convenient route
to the key intermediate 14, since it provided a considerably higher
overall yield (64% from 4) compared to the six-step sequence car-
ried out via the cinnamoate 9 as an intermediate (31% from 4).
The conversion of the intermediate 14 to (þ)-crassalactones B
and C is outlined in Scheme 3. Hydrolytic removal of the iso-
propylidene protecting group in 14 gave the expected lactol 15
(92%), which upon treatment with Meldrum’s acid in the presence
of triethylamine gave the protected furano-lactone 16 in 65% yield.
We initially carried out the TBDPS cleavage in 16 using TBAF, which
surprisingly gave (þ)-crassalactone C (3) as the major product
(43%), accompanied by a minor amount (14%) of (þ)-crassalactone
B (2). Both the 1H and 13C NMR data of compound 3 were consistent
with naturally occurring (þ)-crassalactone C and its physical
properties were in good agreement with those reported pre-
viously.16 Product 3 was presumably formed by a competitive
intramolecular cinnamate migration process in 2, via the cyclic
orthoester intermediate 16a (Scheme 3). A similar intramolecular
O-acyl rearrangement during the silyl deprotection with TBAF has
already been described in the literature.23 Moreover, treatment of
16 with SOCl2 in MeOH gave a 56% yield of (þ)-crassalactone B (2).
ketones.19 Thus, the reduction of 7 with NaBH4 in the presence of
L-
tartaric acid gave a mixture of the required alcohol 5 and the al-
ternative stereoisomer 6 in 78 and 7% yields, respectively. Slightly
better yield of desired stereoisomer 5 is obtained if the reduction of
ketone 7 was carried out with NaBH4 in dry hexane in the presence
of silica gel.20 This procedure provided the requisite intermediate 5
in 89% yield (61% from 4), accompanied with a minor amount of
epimer 6 (9%). Apart from the better yield, the main advantage of
this procedure in relation to the previous one is the usage of
a simple experimental technique and inexpensive reagents. Treat-
ment of ketone 7 with cinnamoyl chloride, gave the corresponding
3-O-cinnamoyl derivative 8 in 75% yield (36% from 4). An alterna-
tive but slightly less efficient synthesis of 8 is accomplished through
a three-step sequence that commenced with esterification of
diacetonide 4 with trans-cinnamic acid under the Neises-Steglich
conditions,21 to provide an almost quantitative yield of the ex-
pected cinnamic ester 9. The terminal isopropylidene function in 9
was directly converted into an aldehyde group, in a single step, by
treatment with periodic acid in dry ethyl acetate,22 whereupon the
expected aldehyde 10 was obtained in 93% yield. Addition of phe-
nylmagnesium bromide in THF, to a solution of 10 in toluene, led to
a mixture of two epimeric alcohols. The mixture was not separated
but was oxidised with PCC in dichloromethane to give the corre-
sponding ketone 8 in 34% yield (31% from 4). The final reduction of
Our optical rotation value is greater than the reported value for
20
isolated (þ)-crassalactone B {lit.6
[
a
]
D
þ8.0 (c 0.5, EtOH); this
work: [
a
]
20 þ45.7 (c 0.5, EtOH)}, but was in reasonable agreement
D
with the reported data of synthetic 2 {lit.9 [
a
]
D
20 þ31.6 (c 1.0, CHCl3);
20
this work: [
a
]
þ35.5 (c 1.0, CHCl3)}. The melting point and NMR
D
data of synthesized sample 2 were in full agreement with those
reported previously.6,9,16
This synthesis of natural product 2 was successfully accom-
plished in nine steps with an overall yield of 19% starting from
prochiral ketone 7 with NaBH4/
L
-tartaric acid took place stereo-
commercially available diacetone-D-glucose (4). However, the nat-
selectively to afford 11 and 12 in 92 and 2% respective yields. The
intermediate 11 was thus obtained in 28% overall yield with respect
to the starting compound 4. However, when this four-step se-
quence was carried out without purification of the intermediates 8
and 10 the desired product 11 was obtained in a slightly higher
overall yield (34% from 4). To prevent the benzylic hydroxyl group
ural product 2 was also prepared starting from the keto-lactone 17.
Compound 17 is readily available from 7-epi-(þ)-goniofufurone
through a regioselective oxidation of the benzylic hydroxyl group at
the C-7 position.24 Treatment of 17 with cinnamoyl chloride and
DMAP in dichloromethane, gave the corresponding 5-O-cinnamoyl
derivative 18 in a yield of 84%. Stereocontrolled reduction of the
Scheme 3. Reagents and conditions: (a) 90% aq TFA, CH2Cl2, 10 ꢁC, 0.5 h, 92%; (b) Meldrum’s acid, Et3N, DMF, 46e48 ꢁC, 64 h, 65%; (c) TBAF, AcOH, THF, rt, 144 h, 43% of 3, 14% of 2;
(d) SOCl2, MeOH, rt, 6 h, 56%; (e) cinnamoyl chloride, DMAP, MeCN, 0.5 h at 0 ꢁC, then 1 h at rt, 84%; (f) NaBH4, L-tartaric acid, THF, 2 h at ꢀ7 ꢁC, then 1 h at 0 ꢁC, 74%.

ꢀ
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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
Table 1
ketone functionality in 18 under the Yatagai-Ohnuki conditions
gave the natural product 2 as the only stereoisomer in 74% yield.
The new synthesis of crassalactone C (3) proceeded in nine
linear steps, with 15% overall yield from the same starting com-
pound 4. The preceding preparation of 3 was accomplished starting
Antiproliferative activities of natural products 1, 2, and 3
Compounds
IC50
PC3
, m
Ma
HT-29
0.59
>100
2.5
A549
(þ)-Goniofufurone (1)
(þ)-Crassalactone B (2)
(þ)-Crassalactone C (3)
>100
25
31
35
4.4
2.6
from D
-xylose in 8% overall yield over ten linear steps.7
An alternative and more efficient synthesis of 3 is presented in
Scheme 4. Treatment of 5 with cinnamoyl chloride in anhydrous
1,2-dichloroethane gave the corresponding 5-O-cinnamoyl de-
rivative 19 (55%) along with a minor amount of 3,5-di-O-cinnamoyl
derivative 20 (12%). Hydrolytic removal of the isopropylidene
protecting group in 19 gave the known7 lactol 19a (85%), which
upon treatment with Meldrum’s acid in the presence of triethyl-
amine gave the natural product 3 in 61% yield (20% overall yield
form 4). It appears that this seven-step sequence represents the
most efficient synthesis of natural product 3 so far. An important
feature of this new approach is that no protection is necessary for
the synthesis of the (þ)-crassalactone C except the diacetonide
function that is already present in the commercially available
starting material 4.25 Finally, a simple semi-synthesis of (þ)-cras-
salactone C (3) was also accomplished by treatment of (þ)-gonio-
fufurone (1) with cinnamoyl chloride in boiling acetonitrile,
whereby the target 3 was obtained in 79% yield.
a
IC50 is the concentration of compound required to inhibit the cell growth by 50%
compared to an untreated control. Values are means of three independent experi-
ments done in quadruplicates. Coefficients of variation were <10%.
(þ)-Goniofufurone (1) showed a moderate cytotoxicity against this
cell line, while (þ)-crassalactones B (2) and C (3) exhibited a potent
activity with growth inhibitory concentrations in the range of low
micromolar values (IC50 4.4 and 2.6 mM, respectively). From Table 1
can be further seen that both natural products 2 and 3 demon-
strated significantly higher potency with respect to control com-
pound 1 (8- and 14-fold respectively).
3. Conclusion
In conclusion we have demonstrated an efficient four-step
conversion of diacetone- -glucose (4) into the benzylic alcohol 5,
D
2.2. In vitro antitumour activity
the key divergent intermediate for a new total synthesis of
(þ)-crassalactones B (2) and C (3). Further, a series of selective
chemical transformations were carried out to construct the ad-
vanced intermediate 14 containing the tetrahydrofuran core of 2.
Compound 14 was finally converted to the final precursor of
(þ)-crassalactones B and C (compound 16), through a two-step
sequence that involves hydrolytic removal of the isopropylidene
protecting group, followed by a successive cyclocondensation of the
resulting lactol 15 with Meldrum’s acid. The intermediate 16 can be
converted into (þ)-crassalactone B (2) or C (3). Selective access to
either 2 or 3 was accomplished by simply changing the conditions
for TBDPS cleavage in the final intermediate 16. The most efficient
route to target 2 was achieved in 15% overall yield from 4. This new
synthesis of 2 requires the use of a single silyl ether protecting
group throughout the synthesis. Finally, we have achieved a prac-
tical, step-economic, and protecting-group-free synthesis of
We have previously found that both natural products 2 and 3
exhibited a potent cytotoxic activity against several cancer cell
lines.11 Hence, we wanted to explore their cytotoxic effects against
the following additional human tumour cell lines: prostate cancer
cells (PC3), colon adenocarcinoma (HT29) and lung adenocarci-
noma epithelial cell line (A549). Cell growth inhibition was evalu-
ated by using the standard MTTcolorimetric assay after exposure of
cells to the test compounds for 72 h (þ)-Goniofufurone (1) was
used as the positive control in this assay. The results are shown in
Table 1.
As shown in Table 1, crassalactones B (2) and C (3) exhibited
a moderate antiproliferative activity toward PC3 cancer cells with
respective IC50 values 25 and 31 mM. However, the control com-
pound 1 was completely inactive against the same cell line. In
contrast, (þ)-goniofufurone (1) showed submicromolar activity
(þ)-crassalactone C (3) in 20% overall yield from diacetone-
D-glu-
against HT-29 cell line (IC50 0.59
m
M). (þ)-Crassalactone C (3)
cose (4). The synthesis of both 2 and 3 is highly efficient and
competitive with previous reports in literature. In vitro anti-
proliferative activities of 2 and 3 against three human tumour cell
lines were recorded and compared with those observed for
demonstrated a potent cytotoxicity (IC50 2.5
salactone B (2) was completely inactive against this cell line. A549
cancer cells were sensitive to all three tested compounds.
m
M), while (þ)-cras-
Scheme 4. Reagents and conditions: (a) cinnamoyl chloride, 1,2-dichloroethane, 70 ꢁC, 20 h, 55% of 19, 12% of 20; (b) 50% aq TFA, rt, 18 h, 85%; (c) Meldrum’s acid, Et3N, DMF,
44e46 ꢁC, 72 h, 61%; (d) cinnamoyl chloride, MeCN, reflux, 2 h, 79%.

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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4585
(þ)-goniofufurone (1). Against A549 cells natural products 2 and 3
showed 8- and 14 times greater potency compared to the parent
compound 1.
1H, OH), 4.60 (d, 1H, J3,4¼3.6 Hz, H-2), 4.63 (d, 1H, J3,4¼2.6 Hz, H-3),
5.40 (d, 1H, J3,4¼2.6 Hz, H-4), 6.09 (d, 1H, J1,2¼3.6 Hz, H-1),
7.50e8.11 (m, 5H, Ph). 13C NMR (62.9 MHz, CDCl3):
d 26.1 and 26.9
(CMe2), 76.5 (C-3), 82.1 (C-2), 84.4 (C-4), 105.2 (C-1), 112.1 (CMe2),
128.7, 129.0, 134.1, 135.2 (Ph), 196.4 (C-7). HRMS (ESIꢀ) calcd for
4. Experimental section
4.1. General
C
14H15O5 263.0925, found 263.0923 (MꢀH). Eluted second was
unreacted starting material 6 (0.020 g, 9%).
€
Melting points were determined on a Buchi 510 apparatus and
4.3.2. Procedure B. To a solution of 6 (0.08 g, 0.30 mmol) in a 10:1
mixture CH2Cl2/H2O (5 mL), was added DDQ (0.136 g, 0.60 mmol)
and the mixture was stirred at room temperature for 72 h. The
mixture was poured in 1% aqueous NaHCO3 (100 mL) and extracted
with CH2Cl2 (3ꢂ20 mL) and then with EtOAc (20 mL). The com-
bined extracts were dried and evaporated, and the residue purified
by flash column chromatography (2:1 Et2O/light petroleum-
/Et2O). Pure ketone 7 (0.063 g, 79%) was first isolated as a col-
ourless syrup. Eluted second was unreacted starting material 6
(0.014 g, 18%). Rf¼0.27 (3:2 toluene/Et2O).
were not corrected. Optical rotations were measured on P 3002
(Kruss) and Autopol IV (Rudolph Research) polarimeters at 20 ꢁC.
€
1H (250 MHz) and 13C (62.9 MHz) NMR spectra were recorded on
a Bruker AC 250 E instrument and chemical shifts are expressed in
parts per million downfield from TMS. IR spectra were recorded
with an FTIR Nexus 670 spectrophotometer (Thermo-Nicolet). High
resolution mass spectra (ESI) of synthesized compounds were ac-
quired on a Agilent Technologies 1200 series instrument equipped
with Zorbax Eclipse Plus C18 (100 mmꢂ2.1 mm i.d. 1.8
mm) column
and DAD detector (190e450 nm) in combination with a 6210 time-
of-flight LC/MS instrument (ESI) in the positive ion mode. Flash
column chromatography was performed using Kieselgel 60
(0.040e0.063, E. Merck). All organic extracts were dried with an-
hydrous Na2SO4. Organic solutions were concentrated in a rotary
evaporator under reduced pressure at a bath temperature below
35 ꢁC.
4.4. 1,2-O-Isopropylidene-5-C-phenyl-a-D-gluco-pentofur-
anose (5)
4.4.1. Procedure A. To a solution of 4 (1.00 g, 3.84 mmol) in dry
EtOAc (100 mL), was added H5IO6 (0.788 g, 3.46 mmol) and the
mixture was stirred at room temperature for 20 h. The suspension
was filtered, then concentrated to syrup (crude 4a) and dried in
vacuum (10 mm Hg) for 1 h. To the stirred 2 M solution of PhMgBr
in THF (14.4 mL, 28.80 mmol) was added TMEDA (5.18 mL,
34.56 mmol). After being stirred at room temperature for 1 h the
solution was cooled to 0 ꢁC under atmosphere of nitrogen. To the
mixture was added a solution of crude 4a in dry toluene (14 mL)
during 10 min and the resulting solution was stirred at 0 ꢁC for 4 h,
then at room temperature for 72 h. The mixture was poured into
cold (þ4 ꢁC) 10% aqueous HCl (150 mL) and extracted first with
CH2Cl2 (3ꢂ35 mL) and then with EtOAc (1ꢂ30 mL). The combined
extracts were washed with 2% aqueous NaOH (100 mL), dried and
evaporated, and the residue purified by flash column chromatog-
raphy (2:1/4:1 Et2O/light petroleum). The major stereoisomer 5
4.2. 1,2-O-Isopropylidene-5-C-phenyl-a-L-ido-pentofuranose
(6)
To a solution of 4 (0.50 g, 1.92 mmol) in dry EtOAc (50 mL), was
added H5IO6 (0.394 g, 1.73 mmol) and the mixture was stirred at
room temperature for 20 h. The suspension was first filtered, and
then concentrated to syrup. The residue was dried in vacuum
(10 mm Hg) for 1 h. The remaining crude aldehyde 4a (0.42 g) was
dissolved in dry Et2O (50 mL) and the solution cooled to 0 ꢁC under
atmosphere of nitrogen. To the mixture was added a commercial
3 M solution of PhMgBr in Et2O (6.50 mL, 19.5 mmol) previously
cooled to 0 ꢁC. The reaction was stirred at 0 ꢁC for 4 h, then at room
temperature for 20 h. The mixture was poured into cold (þ5 ꢁC)
saturated aqueous NH4Cl (300 mL) and extracted first with Et2O
(2ꢂ50 mL) and then with CH2Cl2 (2ꢂ50 mL). The combined extracts
were dried and evaporated, and the residue purified by flash col-
umn chromatography (2:1 Et2O/light petroleum/Et2O). The minor
stereoisomer 5 (0.097 g, 19%) was first isolated as a colourless solid.
(0.497 g, 49%) was first isolated as a colourless solid, Rf¼0.32 (1:1
25
toluene/EtOAc), mp 106e107 ꢁC (Me2CO/light petroleum), [
a]
D
25
ꢀ12.2 (c 1, MeOH), [
a
]
D
ꢀ22.9 (c 0.5, CHCl3), lit.12 mp 104e106 ꢁ
C
(EtOH), [
a]
25 ꢀ43.4 (c 1.3, MeOH); lit.15 mp 102e103 ꢁC (iPr2O/light
D
25
petroleum), [
a
]
ꢀ26.0 (c 0.8, CHCl3). Eluted second was pure 6
D
(0.155 g, 15%), isolated as a colourless solid. NMR data matched
Eluted second was pure 6 (0.307 g, 60%), isolated as a colourless
what was previously reported for both 5 and 6.16
25
solid, Rf¼0.25 (1:1 toluene/EtOAc), mp 166e168 ꢁC (EtOH), [
a]
D
þ22.9 (c 1, MeOH), [
a
]
þ7.3 (c 1, CHCl3); lit.12 mp 162e165 ꢁC
4.4.2. Procedure B. To a solution of L-tartaric acid (0.199 g,
25
D
25
(EtOH), [
a
]
þ32.1 (c 1.34, MeOH), lit.15 mp 159e160 ꢁC (EtOH),
1.32 mmol) in dry THF (3 mL) was added NaBH4 (0.032 g,
1.32 mmol) portionwise and the suspension was heated at reflux
for 2 h before being cooled to ꢀ7 ꢁC. A solution of 7 (0.07 g,
0.26 mmol) in dry THF (2 mL) was added dropwise and the mixture
was stirred at ꢀ7 ꢁC for 0.5 h, then at 0 ꢁC for 1 h, and finally at room
temperature for 24 h. The mixture was evaporated with silica gel
(1 g) and the residue purified by flash column chromatography (7:3
light petroleum/Et2O). The major stereoisomer 5 (0.055 g, 78%) was
first eluted, followed by a minor amount of 6 (0.005 g, 7%). Com-
pound 5: Rf¼0.32 (1:1 toluene/EtOAc).
D
25
[
a]
þ25.0 (c 1.1, CHCl3). NMR data for 5 and 6 matched those
D
previously reported.16
4.3. 1,2-O-Isopropylidene-5-C-phenyl-a-D-xylo-pentofurano-
5-ulose (7)
4.3.1. Procedure A. To a cooled (0 ꢁC) and stirred solution of 6
(0.210 g, 0.79 mmol) in dry CH3CN (27 mL) was added H5IO6
(0.234 g, 1.02 mmol). A solution of PCC (0.010 g; 0.056 mmol) in dry
CH3CN (5 mL) was added during 1 min and the resulting reaction
mixture was stirred vigorously at 0 ꢁC for 2 h. The mixture was
poured to 10% aq NaCl (150 mL) extracted with EtOAc (3ꢂ50 mL).
The combined organic solutions were dried and evaporated. Flash
column chromatography (1:1 Et2O/PE). of the residue gave pure 7
(0.119 g; 57%) as a colourless syrup Crystallization from CHCl3 gave
4.4.3. Procedure C. To a suspension of ketone 7 (0.174 g, 0.66 mmol)
in dry hexane (15 mL) was added NaBH4 (0.013 g, 0.34 mmol) and
silica gel (0.75 g). After the reaction was stirred at 22ꢁ C for 20 h,
after the next two hours an additional amount of NaBH4 (0.013 g,
0.34 mmol) was added in two equal portions and the stirring at
room temperature was continued for the next 6 h. The mixture was
evaporated and the solid residue was purified by flash column
chromatography (2:1 Et2O/light petroleum/Et2O). The major
colourless needles, mp 97 ꢁC, [
a
]
25 ꢀ61.8 (c 0.5, CHCl3), Rf¼0.27 (3:2
D
toluene/Et2O). IR (CHCl3): 3444 (br), 1695, 1597 cmꢀ1
.
1H NMR
(250 MHz, CDCl3):
d
1.35 and 1.57 (2ꢂs, 3H each, CMe2), 3.23 (br s,

ꢀ
G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4586
stereoisomer 5 (0.155 g, 89%) was first eluted, followed by a minor
amount of 6 (0.015 g, 9%). Compound 5: Rf¼0.32 (1:1 toluene/
EtOAc).
NMR (62.9 MHz, CDCl3): d 26.2 and 26.9 (CMe2), 77.6 (C-3), 81.7
(C-4), 83.2 (C-2), 105.1 (C-1), 112.7 (CMe2), 116.2 (C-20), 128.1, 128.2,
128.7, 128.9, 130.7, 133.7, 135.3 (Ph), 146.4 (C-30), 165.1 (C-10), 192.8
(C-5). HRMS (ESI) calcd for C23H22NaO6 417.1309, found 417.1310
(MþþNa).
4.5. 3-O-Cinnamoyl-1,2:5,6-di-O-isopropylidene-a-D-gluco-
furanose (9)
4.7.2. Procedure B. A solution of 7 (0.225 g, 0.85 mmol), cinnamoyl
chloride (0.181 g, 1.09 mmol) and DMAP (0.160 g, 1.31 mmol) in dry
CH2Cl2 (20 mL) was stirred vigorously at 0 ꢁC for 45 min. The
mixture was partitioned between H2O (150 mL) and CH2Cl2
(60 mL), the organic layer was separated and the aqueous phase
extracted with EtOAc (40 mL). The combined organic solutions
were washed with 10% aq NaCl (100 mL), dried and evaporated.
Flash column chromatography (CH2Cl2) of the residue gave pure 8
(0.253 g, 75%) as a colourless syrup.
A solution of 4 (1.0 g, 3.84 mmol), cinnamic acid (1.253 g,
8.46 mmol), DCC (1.903 g, 9.22 mmol) and DMAP (1.878 g,
15.37 mmol) in dry CH2Cl2 (100 mL) was stirred at room temper-
ature for 20 h. The mixture was poured in water (300 mL) and
extracted with CH2Cl2 (2ꢂ50 mL), the combined organic solutions
were washed with 10% aqueous NaCl (250 mL), dried and evapo-
rated. Flash column chromatography of the residue (3:1 light pe-
25
troleum/Et2O) gave pure 9 (1.476 g, 98%) as a colourless syrup, [a]
D
ꢀ48.7 (c 1, CHCl3), Rf¼0.32 (3:1 light petroleum/Et2O). IR (neat):
1720, 1637, 1578 1162 cmꢀ1. 1H NMR (250 MHz, CDCl3):
1.31, 1.32,
d
4.8. 3-O-Cinnamoyl-1,2-O-isopropylidene-5-C-phenyl-
gluco-pentofuranose (11)
a-D-
1.42, 1.54 (4ꢂs, 3H each, 2ꢂCMe2), 4.03e4.05 (m, 2H, 2ꢂH-6),
4.26e4.37 (m, 2H, H-4 and H-5), 4.58 (d, 1H, J1,2¼3.7 Hz, H-2), 5.39
(d, 1H, J3,4¼2.2 Hz, H-3), 5.93 (d, 1H, J1,2¼3.7 Hz, H-1), 6.44 (d, 1H,
4.8.1. Procedure A. To
a solution of L-tartaric acid (0.19 g,
0
0
0
0
0
J2 ,3 ¼16.0 Hz, H-2 ), 7.35e7.59 (m, 5H, Ph), 7.72 (d, 1H, J2 ,3 ¼16.0 Hz,
1.27 mmol) in dry THF (3 mL) was added NaBH4 (0.048 g,
1.27 mmol) portionwise and the suspension was heated at reflux
for 2 h before being cooled toꢀ7 ꢁC. A solution of 8 (0.12 g,
0.30 mmol) in dry THF (3 mL) was added dropwise and the tem-
perature maintained at ꢀ7 ꢁC for 4 h. The solution was allowed to
warm to room temperature and evaporated with silica gel (1 g). The
residue was purified by flash chromatography (2:1/1:2 light pe-
H-30). 13C NMR (62.9 MHz, CDCl3):
d
25.2, 26.1, 26.6, 26.8 (2ꢂCMe2),
67.0 (C-6), 72.4 (C-5), 76.1 (C-3), 79.7 (C-4), 83.3 (C-2), 105.0 (C-1),
109.2 and 112.2 (2ꢂCMe2), 117.0 (C-20), 128.1, 128.9, 130.6, 133.9
(Ph), 146.0 (C-30), 165.4 (C-10). HRMS (ESI) calcd for C21H26NaO7
413.1571, found 413.1577 (MþþNa).
4.6. 3-O-Cinnamoyl-1,2-O-isopropylidene-
dialdo-1,4-furanose (10)
a
-
D
-xylo-pento-
troleum/Et2O) to give pure 11 (0.111 g, 92%), as a colourless syrup,
25
[
a
]
þ18.0 (c 0.5, CHCl3), Rf¼0.6 (1:1 light petroleum/Et2O). IR
D
(neat): 3483 (br), 1716, 1636, 1579, 1162 cmꢀ1. 1H NMR (250 MHz,
CDCl3):
To a solution of 9 (0.446 g, 1.14 mmol) in dry EtOAc (45 mL) was
added H5IO6 (0.338 g, 1.48 mmol). The mixture was stirred vigor-
ously at room temperature for 1.5 h. The suspension was filtered,
then concentrated under vacuum. The residue was purified by flash
d
1.32 and 1.52 (2ꢂs, 3H each, CMe2), 4.46 (dd, 1H, J3,4¼2.5,
J4,5¼8.6 Hz, H-4), 4.67 (d, 1H, J1,2¼3.7 Hz, H-2), 4.74 (d, 1H,
J4,5¼8.7 Hz, H-5), 5.51 (d, 1H, J3,4¼2.5 Hz, H-3), 5.97 (d, 1H,
0
0
0
J1,2¼3.6 Hz, H-1), 6.53 (d, 1H, J2 ,3 ¼15.9 Hz, H-2 ), 7.29e7.64 (m,
10H, 2ꢂPh), 7.82 (d, 1H, J2 ,3 ¼16.0 Hz, H-3 ). 13C NMR (62.9 MHz,
0
0
0
column chromatography (3:2 light petroleum/Et2O) to give pure 10
25
(0.337 g, 93%) as a colourless syrup, [
a]
ꢀ11.2 (c 0.5, CHCl3),
CDCl3): d 26.2 and 26.6 (CMe2), 70.6 (C-5), 76.7 (C-3), 82.3 (C-4),
D
Rf¼0.26 (3:2 light petroleum/Et2O). IR (neat): 1720 (br s), 1636,
83.2 (C-2), 104.9 (C-1), 112.2 (CMe2), 116.4 (C-20), 126.8, 128.1, 128.3,
128.5, 129.0, 130.9, 133.8, 140.3 (Ph), 147.1 (C-30), 166.8 (C-10). HRMS
(ESI) calcd for C23H24NaO6 419.1465, found 419.1460 (MþþNa).
1578, 1161 cmꢀ1. 1H NMR (250 MHz, CDCl3):
d
1.34 and 1.53 (2ꢂs,
3H each, CMe2), 4.67 (d, 1H, J1,2¼3.6 Hz, H-2), 4.81 (d, 1H,
J3,4¼3.4 Hz, H-3), 5.64 (d, 1H, J3,4¼3.4 Hz, 0 H-4), 6.13 (d, 1H,
Eluted second was pure 12 (0.002 g, 2%), isolated as a colourless oil,
20
0
0
J1,2¼3.6 Hz, H-1), 6.34 (d, 1H, J2 ,3 ¼16.0 Hz, H-2 ), 7.33e7.56 (m, 5H,
[a
]
ꢀ87.4 (c 0.5, CHCl3); Rf¼0.25 (1:1 light petroleum/Et2O). IR
D
Ph), 7.67 (d, 1H, J2 ,3 ¼16.0 Hz, H-3 ), 9.70 (s, 1H, H-5). 13C NMR
(neat): nmax 3474, 1716, 1636, 1578, 1496, 1161. 1H NMR (CDCl3):
0
0
0
(62.9 MHz, CDCl3):
d
26.2 and 26.8 (CMe2), 77.1 (C-4), 83.0 (C-2),
d
1.32 and 1.56 (2ꢂs, H each, CMe2), 4.48 (dd, 1H, J3,4¼2.9,
83.4 (C-3), 105.6 (C-1), 112.9 (CMe2), 116.0 (C-20), 128.2, 128.9, 130.8,
133.7 (Ph), 146.8 (C-30), 165.3 (C-10), 196.8 (C-5). HRMS (ESI) calcd
for C17H19O6 319.1176, found 319.1161 (MþþH).
J4,5¼8.5 Hz, H-4), 4.56 (d, 1H, J1,2¼3.6 Hz, H-2), 4.87 (d, 1H,
J3,4¼2.9 Hz, H-3), 5.07 (d, 1H, J4,5¼8.5 Hz, H-5), 6.04 (d, 1H,
0
0
0
J1,2¼3.6 Hz, H-1), 6.51 (d,1H, J2 ,3 ¼16.0 Hz, H-2 ), 7.29e7.65 (m,10H,
Ph), 7.76 (d, 1H, J2 ,3 ¼16.0 Hz, H-3 ). 13C NMR (CDCl3):
d
26.2 and
0
0
0
4.7. 3-O-Cinnamoyl-1,2-O-isopropylidene-5-C-phenyl-
xylo-pentofuran-5-ulose (8)
a-
D-
26.7 (CMe2), 72.6 (C-5), 75.9 (C-3), 83.6 (C-4), 83.7 (C-2), 104.9
(C-1), 112.4 (CMe2), 116.6 (C-20), 126.7, 128.3, 128.7, 128.8, 129.0,
130.9, 133.9, 138.7 (Ph), 146.6 (C-30), 165.3 (C-10). HRMS (ESI) calcd
for C23H24NaO6: 419.1465, found: 419.1444 (MþþNa).
4.7.1. Procedure A. To a cooled (0 ꢁC) and stirred solution of 10
(0.333 g, 1.05 mmol) in dry toluene (25 mL) was added 1 M solution
of PhMgBr in THF (2.1 mL, 2.1 mmol). The mixture was stirred at
4.8.2. Procedure B. To a solution of 9 (0.72 g, 1.84 mmol) in dry
EtOAc (70 mL), NaIO4 (0.395 g, 1.84 mmol) and H5IO6 (0.378 g,
1.66 mmol) were consecutively added and the mixture was stirred
at room temperature for 3 h. The suspension was filtered, then
concentrated under vacuum. The remaining crude aldehyde 10
(0.63 g) was taken up in dry toluene (60 mL) and the solution
cooled to 0 ꢁC under atmosphere of nitrogen. To the solution was
added 1 M solution of PhMgBr in THF (5 mL, 5 mmol) and the
mixture was stirred at 0 ꢁC for 3 h, then neutralized with glacial
AcOH and poured in 15% aqueous NH4Cl (250 mL). The mixture was
extracted with CH2Cl2 (3ꢂ80 mL), the extracts were combined,
dried and evaporated. The residue (0.85 g) was taken up in dry
DMSO (10 mL) and treated with Ac2O (3.5 mL, 37.03 mmol) while
stirring at room temperature for 20 h. The mixture was poured in
0
ꢁC under atmosphere of nitrogen for 3 h, then neutralized with
glacial AcOH (1 mL) and poured in 15% aqueous NH4Cl (200 mL).
The mixture was extracted with CH2Cl2 (3ꢂ60 mL), the organic
solutions were combined, dried and evaporated. The residue was
taken up in dry CH2Cl2 (30 mL) and to the solution was added PCC
(0.76 g, 3.68 mmol). After the mixture heated to reflux for 4 h, it was
concentrated with silica gel (1 g) and the residue purified by flash
chromatography (CH2Cl2). Pure 8 (0.14 g, 34%) was obtained as
25
a colourless syrup, [
a
]
ꢀ108.1 (c 1, CHCl3), Rf¼0.25 (CH2Cl2). IR
D
(neat): 1718, 1635, 1597, 1156 cmꢀ1 1H NMR (250 MHz, CDCl3):
.
d
1.39 and 1.63 (2ꢂs, 3H each, CMe2), 4.70 (d, 1H, J1,2¼3.6 Hz, H-2),
5.71e5.80 (m, 2H, H-3 and H-4), 6.22 (d, 1H J1,2¼3.6 Hz, H-1)0, 6.23
(d, 1H, J2 ,3 ¼16.0 Hz, H-2 ), 7.33e8.01 (m, 11H, 2ꢂPh and H-3 ). 13C
0
0
0

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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4587
10% aqueous NaCl (200 mL) and extracted with CH2Cl2 (3ꢂ70 mL).
The combined organic solutions were washed with water (200 mL),
dried and evaporated. The remaining crude ketone 8 (0.95 g) was
(0.27 g, 2.22 mmol) in dry CH2Cl2 (30 mL) was stirred at room
temperature for 50 h. The mixture was poured in water (150 mL)
and extracted with CH2Cl2 (3ꢂ30 mL). The combined organic so-
lutions were washed with 10% aqueous NaCl (100 mL), dried and
evaporated. Flash column chromatography of the residue (5:1 light
petroleum/Et2O) gave pure 14 (0.337 g, 96%) as a colourless oil.
Rf¼0.23 (4:1 light petroleum/Et2O).
dissolved in dry THF (6 mL) and treated with NaBH4/L-tartaric acid
reagent system as described above (Procedure A). The reduction
was carried out at ꢀ7 ꢁC for 1 h, then at 0 ꢁC for 1 h and finally at
room temperature for 18 h. The mixture was evaporated with silica
gel (1 g) and the residue was purified by flash chromatography
(19:1 toluene/Et2O) to give slightly contaminated product 11
(0.40 g). Repeated chromatographic purification (7:3 light petro-
leum/Et2O) gave pure 11 (0.255 g, 35% from 9). Rf¼0.6 (1:1 light
petroleum/Et2O).
4.11. 5-O-(tert-Butyldiphenylsilyl)-3-O-cinnamoyl-5-C-phe-
nyl-D-gluco-pentofuranose (15)
To a cooled (0 ꢁC) and stirred solution 14 (0.53 g, 0.84 mmol) in
CH2Cl2 (0.5 mL) was added a cooled (10 ꢁC) solution of 90% aqueous
TFA (14 mL). After stirring for 0.5 h, the mixture was concentrated
by co-distillation with toluene and the residue purified by flash
chromatography (49:1 CH2Cl2/MeOH). Pure 15 (0.459 g, 92%) was
obtained as a colourless solid. The product 15 was isolated as a 5:1
4.9. 5-O-(tert-Butyldiphenylsilyl)-1,2-O-isopropylidene-5-C-
phenyl-a-D-gluco-pentofuranose (13)
To a solution of 5 (0.5 g, 1.88 mmol) and imidazole (0.32 g,
4.69 mmol) in dry CH2Cl2 (30 mL) was slowly added TBDPSCl
(0.98 mL, 3.76 mmol). After being stirred at room temperature for
48 h, the mixture was poured in water (300 mL) and extracted with
CH2Cl2 (3ꢂ50 mL). The organic layers were combined, dried and
evaporated, and the residue purified by flash column chromatog-
raphy (5:1 light petroleum/Et2O). Pure 13 (0.92 g, 97%) was ob-
mixture of
a- and b-anomers. Data for mixture of isomers: mp
25
58e60 ꢁC, Rf¼0.28 (1:1 light petroleum/Et2O), [
a
]
þ57.7 (c 2,
D
H2O). IR (neat): 3438, 1789, 1713, 1638, 1169 cmꢀ1. HRMS (ESI) calcd
for C36H38NaO6Si 617.2330, found 617.2332 (MþþNa). Data for the
major a d 0.95 (s, 9H, CMe3),
-anomer: 1H NMR (250 MHz, CDCl3):
3.89 (br s, 2H, 2ꢂOH), 4.03 (dd, 1H, J1,2¼4.2, J2,3¼1.6 Hz, H-2), 4.77
(dd, 1H J3,4¼3.4, J4,5¼8.1 Hz, H-4), 4.91 (d, 1H, J4,5¼8.2 Hz, H-5), 5.28
(d, 1H, J1,2¼4.4 Hz, H-1), 5.37 (dd, 1H, J2,3¼1.5, J3,4¼3.2 Hz, H-3), 6.15
tained as a colourless oil, [
a
]
25 ꢀ43 (c 2, CHCl3), Rf¼0.38 (CH2Cl2). IR
(neat): 3448 (br), 1590 cmꢀD1. 1H NMR (250 MHz, CDCl3):
d 1.09 (s,
(d, 1H, J2 ,3 ¼16.0 Hz, H-2 ), 7.12e7.70 (m, 21H, Ph and H-3 ). 13C
0
0
0
0
9H, CMe3), 1.31 and 1.41 (2ꢂs, 3H each, CMe2), 4.00 (dd, 1H, J3,4¼1.8,
J4,5¼2.7 Hz, H-4), 4.29 (br d, 1H, J3,4¼1.8 Hz, H-3), 4.57 (d, 1H,
J1,2¼3.6 Hz, H-2), 5.26 (d, 1H, J4,5¼2.7 Hz, H-5), 5.36 (br s, 1H, OH),
6.07 (d, 1H, J1,2¼3.6 Hz, H-1), 7.15e8.75 (m, 15H, 3ꢂPh). 13C NMR
NMR (62.9 MHz, CDCl3): d 19.2 (CMe3), 26.7 (CMe3), 73.0 (C-5), 75.0
(C-2), 79.0 (C-3), 80.9 (C-4), 95.9 (C-1), 117.1 (C-20), 127.1, 128.0,
128.1, 128.2, 128.9, 129.3, 129.4, 129.6, 130.6, 132.8, 133.3, 134.0,
135.7, 140.8 (Ph), 145.5 (C-30), 166.5 (C-10). Data for the minor
(62.9 MHz, CDCl3):
d 19.2 (CMe3), 26.0 and 26.6 (CMe2), 26.8
(CMe3), 75.2 (C-3), 77.2 (C-5), 81.7 (C-4), 85.1 (C-2), 104.7 (C-1),
111.3 (CMe2), 126.8, 127.3, 127.5, 127.9, 128.3, 129.7, 130.0, 131.8,
132.0, 135.9, 136.2, 139.4 (Ph). HRMS (ESI) calcd for C30H36NaO5Si
527.2224, found 527.2222 (MþþNa).
b
-anomer: 13C NMR (62.9 MHz, CDCl3):
d
103.0 (C-1).
4.12. 3,6-Anhydro-7-O-(tert-Butyldiphenylsilyl)-5-O-cinna-
moyl-2-deoxy-7-C-phenyl-D-glycero-D-ido-heptono-1,4-
lactone (16)
4.10. 5-O-(tert-Butyldiphenylsilyl)-3-O-cinnamoyl-1,2-O-iso-
propylidene-5-C-phenyl-
a
-
D
-gluco-pentofuranose (14)
To a solution of 15 (0.375 g, 0.63 mmol) in dry DMF (8 mL), was
added Meldrum’s acid (0.091 g, 0.63 mmol) and dry Et3N (0.1 mL,
0.72 mmol). The mixture was stirred for 64 h at 46e48 ꢁC and then
evaporated. The residue was purified by flash chromatography (1:1
light petroleum/Et2O) to afford pure 16 (0.254 g, 65%) as a colour-
4.10.1. Procedure A. To a solution of 11 (0.25 g, 0.63 mmol) and
imidazole (0.06 g, 0.82 mmol) in dry CH2Cl2 (10 mL) was added
TBDPSCl (0.17 mL, 0.63 mmol) After being stirred for 20 h at room
temperature an additional amount of imidazole (0.116 g, 1.7 mmol)
and TBDPSCl (0.17 mL, 0.63 mmol) was added and the mixture was
stirred at room temperature for 20 h. After 40 h, a new portion of
imidazole (0.044 g, 0.63 mmol) and TBDPSCl (0.085 mL, 0.32 mmol)
was added and the stirring was continued at for the next 7 h. The
mixture was poured in water (250 mL), extracted with CH2Cl2
(2ꢂ40 mL) and EtOAc (40 mL) successively, and dried. After evap-
oration of the solvent, the crude product was subjected to flash
less oil, [
a
]
25 þ41 (c 1, CHCl3), Rf¼0.27 (1:1 light petroleum/Et2O). IR
D
(neat): 1780, 1753, 1637, 1153 cmꢀ1
.
1H NMR (250 MHz, CDCl3):
d
0.91 (s, 9H, CMe3), 2.52 (br d, 1H, J2a,2b¼18.9 Hz, H-2a), 2.65 (dd,
1H, J2a,2b¼18.9, J2b,3¼6.1 Hz, H-2b), 4.51 (dd, 1H, J5,6¼2.6,
J6,7¼8.6 Hz, H-6), 4.80 (br t, 1H, J3,4¼J2b,3¼4.6 Hz, H-3), 4.89 (d, 1H,
J6,7¼8.6 Hz, H-7), 4.97 (d, 1H, J3,4¼4.6 Hz, H-4), 5.66 (d, 1H,
0
0
0
J5,6¼2.6 Hz, H-5), 6.13 (d, 1H, J2 ,3 ¼16.1 Hz, H-2 ), 7.08e8.62 (m,
21H, Ph and H-30). 13C NMR (62.9 MHz, CDCl3):
d 19.2 (CMe3), 26.8
column chromatography (9:1/1:1 light petroleum/Et2O) to give
(CMe3), 35.7 (C-2), 72.6 (C-7), 75.2 (C-5), 76.5 (C-3), 82.7 (C-6), 84.9
(C-4), 116.8 (C-20), 127.1, 127.8, 128.2, 128.23, 128.3, 129.0, 129.4,
129.5, 130.8, 132.8, 133.1, 134.0, 135.8, 136.0, 140.6 (Ph), 145.8 (C-30),
165.4 (C-10), 175.1 (C-1). HRMS (ESI) calcd for C38H38NaO6Si
641.2330, found 641.2338 (MþþNa).
25
pure 14 (0.36 g, 90%) as a colourless oil, [
a
]
þ18.6 (c 2, CHCl3),
D
Rf¼0.23 (4:1 light petroleum/Et2O). IR (neat): 1718,1638,1160 cmꢀ1
.
1H NMR (250 MHz, CDCl3):
d
0.93 (s, 9H, CMe3), 1.30 and 1.58 (2ꢂs,
3H each, CMe2), 4.57 (d, 1H, J1,2¼3.7 Hz, H-2), 4.70 (dd, 1H, J3,4¼2.6,
J4,5¼8.6 Hz, H-4), 4.95 (d, 1H, J4,5¼8.6 Hz, H-5), 5.47 (d, 1H,
J3,4¼2.6 Hz, H-3)0, 5.77 (d, 1H, J1,2¼3.7 Hz, H-1), 60.15 (d, 1H,
4.13. 3,6-Anhydro-5-O-cinnamoyl-2-deoxy-7-C-phenyl-L-ido-
hept-7-ulosono-1,4-lactone (18)
J2 ,3 ¼16.0 Hz, H-2 ), 7.08e8.65 (m, 21H, 4ꢂPh and H-3 ). 13C NMR
0
0
(62.9 MHz, CDCl3):
d 19.2 (CMe3), 26.2 and 26.7 (CMe2), 26.74
(CMe3), 72.6 (C-5), 76.4 (C-3), 82.1 and 82.9 (C-2 and C-4), 104.4
(C-1), 112.0 (CMe2), 117.3 (C-20), 127.0, 127.8, 128.0, 128.1, 128.13,
128.9, 129.2, 129.4, 130.5, 132.9, 133.1, 134.0, 135.8, 136.0, 140.6 (Ph),
145.2 (C-30), 165.7 (C-10). HRMS (ESI) calcd for C39H42NaO6Si
657.2643, found 657.2651 (MþþNa).
A solution of 17 (0.040 g, 0.16 mmol), trans-cinnamoyl chloride
(0.035 g, 0.21 mmol) and DMAP (0.030 g, 0.24 mmol) in dry MeCN
(10 mL) was stirred at 0 ꢁC for 0.5 h and then at room temperature
for 1 h. The mixture was poured to water (150 mL) and extracted
with EtOAc (3ꢂ30 mL). The combined organic phases were washed
with 10% aq NaCl (100 mL), dried and evaporated, and the residue
purified by flash column chromatography (2:1 Et2O/light petro-
leum), to give pure 18 (0.051 g, 84%) as a solid. Recrystallization
4.10.2. Procedure B. A solution of 13 (0.28 g, 0.55 mmol), cinnamic
acid (0.18 g, 1.22 mmol), DCC (0.275 g, 1.33 mmol) and DMAP

ꢀ
4588
G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
20
from Et2O gave colourless needles, mp 108 ꢁC, [
a
]
ꢀ92.2 (c 0.5,
128.1, 128.3, 128.4, 128.5, 128.7, 128.8, 130.3, 130.5, 134.0, 134.1,
137.8 (3ꢂPh), 145.5 and 146.2 (2ꢂC-30), 165.0 and 167.1 (2ꢂC-10).
HRMS (ESI): m/z 549.1886 (MþþNa), calcd for C32H30NaO7:
549.1884. Eluted next was pure 19 (0.103 g, 55%; 63% when cal-
D
CHCl3); Rf¼0.30 (2:1 Et2O/light petroleum). IR (CHCl3): nmax 1791
(C]O, lactone), 1716 (C]O, ester), 1635 (C]C, cinnamate), 1497
(Ph) and 1147 (OC]O, ester). 1H NMR (CDCl3): 2.83 (d, 1H,
J2a,2b¼18.9 Hz, H-2a), 2.92 (dd, 1H, J2a,2b¼18.6, J2b,3¼2.1 Hz, H-2b),
5.04 (d, 1H, J5,6¼4.0 Hz, H-4), 5.19e5.31 (m, 1H, H-3), 5.78 (d, 1H,
J5,6¼4.9 Hz, H-6),0 6.04 (d, 1H, J5,6¼4.9 Hz, H-5), 6.13 (d, 1H,
culated to reacted 5), as a colourless oil, [
a
]
20 þ43.1 (c 1.0, CHCl3);
D
Rf¼0.25 (1:1 hexane/Et2O). IR (film): nmax 3453 (OH), 1712 (C]O,
ester), 1636 (C]C, cinnamate) and 1164 (OC]O, ester). 1H NMR
J2 ,3 ¼16.0 Hz, H-2 ), 7.32e7.95 (m, 6H, Ph and H-3 ). 13C NMR
(CDCl3): 35.9 (C-2), 76.5 (C-5), 78.3 (C-3), 81.8 (C-6), 85.7 (C-4),
115.5 (C-20), 127.9, 128.2, 128.8, 130.8, 133.6, 133.9, 135.0 (2ꢂPh),
146.8 (C-30), 164.6 (C-10), 174.2 (C-1), 193.4 (C-7). HRMS (ESI): m/z
379.1193 (MþþH), calcd for C22H19O6: 379.1176.
(CDCl3):
d
1.32 and 1.52 (2ꢂs, 3H each, CMe2), 4.28 (d, 1H,
0
0
0
J3,4¼2.0 Hz, H-3), 4.48 (dd, 1H, J3,4¼2.1, J4,5¼9.3 Hz, H-4), 4.62
(d, 1H, J1,2¼3.6 Hz, H-2), 5.96 (d, 1H, J1,2¼3.5 Hz, H-1), 6.12 (d, 1H,
0
0
0
J4,5¼9.2 Hz, H-5), 6.49 (d, 1H, J2 ,3 ¼16.0 Hz, H-2 ), 7.29e7.61
(m, 10H, 2ꢂPh), 7.77 (d, 1H, J2 ,3 ¼16.0 Hz, H-3 ). 13C NMR (CDCl3):
0
0
0
d
26.1 and 26.7 (CMe2) 72.8 (C-5), 73.9 (C-3), 81.6 (C-4), 84.5 (C-2),
4.14. (D)-Crassalactone B (2)
104.9 (C-1), 111.6 (CMe2), 116.8 (C-20), 126.7, 127.6, 128.2, 128.5,
128.6, 128.9, 128.9, 133.8, 136.9 (2ꢂPh), 146.8 (C-30), 167.1 (C-10).
HRMS (ESI): m/z 419.1450 (MþþNa), calcd for C23H24NaO6:
419.1465. The unreacted starting material 5 (0.015 g, 12%) was
eluted last.
4.14.1. Procedure A. To a solution of 16 (0.09 g, 0.14 mmol) in dry
MeOH (5 mL) was added SOCl2 (0.1 mL, 1.38 mmol). After being
stirred at room temperature for 6 h, the mixture was poured in
water (50 mL), then neutralized with 10% aqueous NaHCO3
(2.5 mL), and extracted with CH2Cl2 (3ꢂ30 mL). The organic ex-
tracts were combined, dried and evaporated, and the residue pu-
rified by flash column chromatography (97:3 CH2Cl2/Me2CO) to
afford slightly impure (þ)-crassalactone B (2). Repeated chroma-
tography using the same solvent system gave pure 2 (0.031 g, 56%)
4.16. (D)-Crassalactone C (3)
4.16.1. Procedure A. To a stirred solution of 16 (0.079 g, 0.13 mmol)
in a mixture of THF (2 mL) and AcOH (0.15 mL, 2.5 mmol) was
added TBAF (1.53 mL,1.53 mmol) in three equal portions over 48 h.
After stirring at room temperature for additional 48 h, a new
portion of TBAF (0.51 mL, 0.51 mmol) was added and the mixture
stirred at room temperature for the next 24 h. An additional
amount of TBAF (0.46 mL, 0.46 mmol) was added to the solution
and the stirring continued for additional 24 h (total reaction time:
144 h). The mixture was evaporated and the residue purified by
flash column chromatography (97:3/ 19:1 CH2Cl2/Me2CO).
Eluted first was pure 2 (0.007 g, 14%), isolated as a colourless solid.
Data for 2 reported above. Eluted second was impure (þ)-crassa-
lactone C (3). The sample 3 was partitioned between CH2Cl2
(30 mL) and 1% aqueous NaHCO3 (50 mL), the organic layer was
separated, and the aqueous phase extracted with CH2Cl2
(2ꢂ30 mL). The combined organic extracts were dried and evap-
orated, and the residue purified by flash chromatography (Et2O).
Pure 3 (0.021 g, 43%) was obtained as a colourless solid. Re-
as a colourless solid. Recrystallization from Et2O gave colourless
20
needles of pure 2, mp 173ꢀ174 ꢁC, [
a]
þ35.5 (c 1.0, CHCl3), lit.9
D
20
20
168ꢀ171 ꢁC (EtOH), [
a]
þ31.6 (c 1.0, CHCl3), [
a
]
þ45.7 (c 0.5,
D
D
20
EtOH), lit.6 171e173 ꢁC, [
a
]
þ8.0 (c 0.5, EtOH); Rf¼0.38 (97:3
D
CH2Cl2/Me2CO). IR (KBr): 3444 (br), 1789, 1717, 1636 cmꢀ1. 1H and
13C NMR spectral data for 2 were in full agreement with those re-
ported previously.16 HRMS (ESI): m/z 403.1143 (MþþNa), calcd for
C
22H20NaO6: 403.1152.
4.14.2. Procedure B. To a stirred suspension of
L-tartaric acid
(0.065 g, 0.43 mmol) in dry THF (2.5 mL), was added NaBH4
(0.017 g, 0.45 mmol) in two equal portions and the suspension was
heated at reflux for 2 h before being cooled to ꢀ7 ꢁC. A solution of
18 (0.035 g, 0.09 mmol) in dry THF (1.5 mL) was added dropwise
and the temperature maintained at ꢀ7 ꢁC for 2 h and then at 0 ꢁ
C
for the next 1 h. The mixture was evaporated with silica gel (1 g)
and the residue was purified on
a
column of flash silica
crystallization from diethyl ether gave colourless crystals, mp
25
(49:1/97:3 CH2Cl2/Me2CO). Eluted first was unreacted starting
compound 18 (0.008 g, 23%). Eluted second was pure natural
product 2 (0.026 g, 74%), isolated as a colourless syrup. Rf¼0.38
(97:3 CH2Cl2/Me2CO).
153 ꢁC, [
a
]
þ111.6 (c 0.5, EtOH), Rf¼0.46 (1:1 light petroleum/
D
EtOAc); lit.6 mp 147e150 ꢁC (EtOH), [
a
]
30 þ98.4 (c 0.5, EtOH), lit.9
D
mp 145e148 ꢁC; [
spectral data for 3 were in full agreement with those reported
a
]
þ108.0 (c 0.37, CHCl3). 1H and 13C NMR
D
previously.16
4.15. 5-O-Cinnamoyl-1,2-O-isopropylidene-5-C-phenyl-
gluco-pentofuranose (19)
a-D-
4.16.2. Procedure B. A solution of 19 (0.145 g, 0.37 mmol) in 50% aq
TFA (4 mL) was stirred at room temperature for 18 h. The volatiles
were removed by co-distillation with toluene and the residue
purified by flash column chromatography (7:3 EtOAc/hexane). Pure
19a (0.111 g, 85%) was obtained a colourless solid. HRMS (ESI): m/z
379.1150 (MþþNa), calcd for C20H20NaO6: 379.1152. To a solution of
19a (0.034 g, 0.10 mmol) in dry DMF (1 mL) was added dry Et3N
(0.027 mL, 0.19 mmol) and Meldrum’s acid (0.028 g, 0.19 mmol).
The mixture was stirred for 72 h at 44ꢀ46 ꢁC and then evaporated.
The residue was purified by flash column chromatography (17:3
Et2O/hexane) to afford pure 3 (0.022 g, 61%). Rf¼0.46 (1:1 light
petroleum/EtOAc).
A solution of 5 (0.125 g, 0.47 mmol) and trans-cinnamoyl
chloride (0.090 g, 0.54 mmol) in dry 1,2-dichloroethane (12.5 mL)
was stirred at 70 ꢁC for 20 h. The mixture was evaporated and the
residue was purified by flash column chromatography (2:3/1:1
Et2O/light petroleum). Dicinnamoate 20 (0.030 g, 12%) was first
eluted. After crystallization from CH2Cl2/hexane pure 20 was
20
obtained as a white powder, mp 137 ꢁC, [
a
]
ꢀ138.8 (c 0.25,
D
CHCl3), lit.9 125ꢀ126 ꢁC, lit.9
[
a
]
20 ꢀ297.1 (c 0.23, CHCl3); Rf¼0.28
D
(1:5 Et2O/light petroleum). IR (KBr): nmax 1719 (C]O, ester), 1635
(C]C, cinnamate), 1497 (Ph) and 1161 (OC]O, ester). 1H NMR
(CDCl3):
d
1.34 and 1.56 (2ꢂs, 3H each, CMe2), 4.66 (d, 1H,
J1,2¼3.7 Hz, H-2), 4.80 (dd, 1H, J3,4¼3.0, J4,5¼9.3 Hz, H-4), 5.61 (d,
1H, J3,4¼3.0 Hz, H-3), 6.02 (d, 1H, J1,2¼3.7 Hz, H-1), 6.15 (d, 1H,
4.16.3. Procedure C. A solution of 1 (0.025 g, 0.10 mmol) and trans-
cinnamoyl chloride (0.083 g, 0.50 mmol) in dry MeCN (4 mL) was
vigorously stirred under reflux for 2 h. The mixture was poured to
1% aq NaHCO3 (75 mL) and extracted with EtOAc (3ꢂ25 mL). The
combined organic solutions were washed with 10% aq NaCl (50 mL)
dried and evaporated and the residue purified on a column of flash
0
0
0
J4,5¼9.3 Hz, H-5), 6.48 (d, 1H, J2 ,3 ¼16.1 Hz, 0H-2 ), 7.32e7.68 (m,
15H, 3ꢂPh), 7.71 (d, 1H, J2 ,3 ¼16.1 Hz, H-3 ). 13C NMR (CDCl3):
0
0
d
26.2 and 26.7 (CMe2) 72.1 (C-5), 75.8 (C-3), 80.6 (C-4), 83.2 (C-2),
105.1 (C-1), 112.2 (CMe2), 116.7 and 117.3 (2ꢂC-20), 127.3, 127.4,

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G. Benedekovic et al. / Tetrahedron 71 (2015) 4581e4589
4589
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troleum/EtOAc).
4.17. Cell lines
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Supplementary data (General experimental procedures for
preparation of known intermediates and copies of 1H, and 13C NMR
spectra of key compounds.) related to this article can be found at
http://dx.doi.org/10.1016/j.tet.2015.05.040. These data include MOL
files and InChiKeys of the most important compounds described in
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