Development of a novel hapten for radioimmunoassay of the lignan, enterolactone in plasma (Serum). Total synthesis of (±)-trans-5- carboxymethoxyenterolactone and several analogues

Article abstract of DOI:10.1016/S0040-4020(00)00078-8

A recently developed method for the analysis of the mammalian lignan, enterolactone 1, is based on time-resolved fluoroimmunoassay (TR-FIA) using an europium chelate as a label. This RIA utilizes enterolactone derivatives carrying a carboxylic acid appendage for the production of antiserum and tracer. The synthesis of 5-carboxymethoxyenterolactone 6 and analogues 5, 7 and 8 is described, and their suitability for the method are discussed. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4020(00)00078-8

TETRAHEDRON
Pergamon
Tetrahedron 56 (2000) 1873–1882
Development of a Novel Hapten for Radioimmunoassay of the
Lignan, Enterolactone in Plasma (Serum). Total Synthesis of
(^)-trans-5-Carboxymethoxyenterolactone
and Several Analogues
*
¨
¨
¨ ¨ ¨
Taru Makela, Jorma Matikainen, Kristiina Wahala and Tapio Hase
Organic Chemistry Laboratory, Department of Chemistry, University of Helsinki,
P.O. Box 55 (A.I. Virtasen aukio 1), 00014 Helsinki, Finland
Received 23 June 1999; revised 4 January 2000; accepted 20 January 2000
Abstract—A recently developed method for the analysis of the mammalian lignan, enterolactone 1, is based on time-resolved fluoro-
immunoassay (TR-FIA) using an europium chelate as a label. This RIA utilizes enterolactone derivatives carrying a carboxylic acid
appendage for the production of antiserum and tracer. The synthesis of 5-carboxymethoxyenterolactone 6 and analogues 5, 7 and 8 is
described, and their suitability for the method are discussed. ᭧ 2000 Elsevier Science Ltd. All rights reserved.
Introduction
enterolignans are produced by the action of intestinal micro-
flora on the precursors (i.e. matairesinol 2 and secoiso-
lariciresinol 4) in dietary fiber. The precursor lignans have
also been detected in human urine and plasma (Scheme 1).13,14
Lignans have attracted much interest over the years on
account of their widespread occurrence in various plant
species,1 and their broad range of biological activity.2
Since the discovery of the mammalian lignans enterolactone
1 and enterodiol 3 from human urine,3–6 there has been
much discussion about their biological function. Especially
interesting is their suggested role as antiestrogens and anti-
carcinogens among other possible biological activities.7
Enterolactone 1, alone or together with enterodiol 3, has
more recently been detected in human plasma and other
biological fluids as well.8–10
Available analytical methods for the detection and quantifi-
cation of phytoestrogens in human biological fluids and in
food samples are based on gas–liquid chromatography or
high-performance liquid chromatography alone or in combi-
nation with mass spectrometry.14–19 These expensive and
time-consuming methods are not suitable for screening
purposes in large populations. In addition, these procedures
are not sensitive enough for the assay of conjugated
phytoestrogens in plasma. These disadvantages led to the
application of a new analytical method based on radio-
immunoassay (RIA).20
Human diet has been shown to contain plant lignans, which
act as precursors for the mammalian lignans.8,11,12 The
Scheme 1. 1; 3 : R1ˆOH, R2ˆH; 2; 4 : R1ˆOMe, R2ˆOH; 5 : R1ˆOCH2CO2H; R2ˆR3ˆH; 6 : R1ˆOH; R2ˆH; R3ˆOCH2CO2H; 7 : R1ˆOH;
R2ˆO(CH2)3CO2H; R3ˆH; 8 : R1ˆOH; R2ˆ(CH2)2CO2H; R3ˆH.
Keywords: lignans; lactones; carboxylic acids and derivatives; Michael reactions.
* Corresponding author. E-mail: taru.makela@helsinki.fi
0040–4020/00/$ - see front matter ᭧ 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(00)00078-8

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T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1874
Scheme 2.
The first direct radioimmunoassay for unconjugated and
total daidzein and genistein in human biological fluids
was developed by Adlercreutz et al.21,22 and as the next
step, the use of time-resolved fluoroimmunoassay (TR-
FIA) of plasma enterolactone was recently introduced.23
This rapid technique has the advantages of other nonradio-
isotopic assays, such as stability of the reagent and lack of
radiation. The increase in sensitivity and assay range
compares well with the conventional enzyme immunoassay
(EIA) and fluoroimmunoassay (FIA) methods. The method
has very recently been applied to the quantitative determi-
nation of phytoestrogens in human urine.24
Two approaches for attaching the carboxylate side chain
were studied, either adding it to the finished molecular
framework or incorporating it in one of the starting
materials. The former approach was more successful, par-
ticularly for the attachment of side chain to the aromatic ring
by a carbon–oxygen bond. As a matter of fact, the method
was chosen for general use due to the flexibility in regard to
the aromatic ring substituents. To allow a selective and
controlled introduction of the side chain late in the
synthesis, suitable protection of the phenolic hydroxy
groups was required. For this purpose, benzyl and silyl
ethers were chosen.
An essential part of the work was to find a properly substi-
tuted enterolactone derivative to be coupled to bovine serum
albumin and then used in the immunization of rabbits in
order to create an antiserum.20 The same derivative would
be used in preparing the tracer with europium label. A
number of synthetic enterolactone derivatives carrying
various carboxylic acid side chains were developed for
this purpose. We present here the synthesis of 5-carboxy-
methoxyenterolactone 6 and analogous derivatives 5, 7 and
8, and discuss their suitability for radioimmunoassay.
The latter approach was applied only in one case, where the
side chain was attached directly to the aromatic ring by a
carbon–carbon bond.
Since the carboxylic acid side chain was intended to be
incorporated in the benzyl bromide moiety of the three-
component synthesis (Scheme 2), the synthetic procedure
of properly substituted bromides varied considerably with
the availability of starting materials.
The benzyl bromide 11 for 3-carboxymethoxyenterolactone
5 was readily prepared via silyl ether27 formation and the
reduction of aldehyde 9 to alcohol 10 with NaBH4 (Scheme
3). For the preparation of benzyl bromide 16 for 5-carboxy-
methoxyenterolactone 6, methyl 3,5-dihydroxybenzoate
Results and Discussion
The trans-a,b-dibenzyl-g-butyrolactone framework is
generally obtained by the Michael addition of an anion
derived from dithioacetal to butenolide followed by benzy-
lation in situ (Scheme 2).25,26
Scheme 5. Reagents: (a) benzyl chloride, KOH, DMF, 80–90ЊC (56%); (b)
NaOH, ethanol, H2O, CHCl3, reflux (17%); (c) t-BuPh2SiCl, imidazole,
DMF (71%); (d) NaBH4, ethanol (61%); (e) PBr3, ether, ϩ4ЊC (87%).
Scheme 3. Reagents: (a) t-BuPh2SiCl, imidazole, DMF (99%); (b) NaBH4,
ethanol (99%); (c) PBr3, ether, ϩ4ЊC (86%).
Scheme 4. Reagents: (a) MeOH, conc. H2SO4, reflux (89%); (b) benzyl bromide, K2CO3, acetone, reflux (33%); (c) t-BuPh2SiCl, imidazole, DMF (99%);
(d) LiAlH4, ether, reflux (27%); (e) PBr3, ether, ϩ4ЊC (89%).

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T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1875
Scheme 6. Reagents: (a) n-butyllithium, THF, Ϫ78ЊC, then 2-butenolide, THF, Ϫ78ЊC, then benzyl bromide, 11, HMPA, THF, Ϫ78ЊC (10%); (b) n-Bu4NϩFϪ,
THF (99%); (c) BrCH2CO2Et, K2CO3, KI, acetone, reflux (96%); (d) Raney nickel, ethanol, reflux (43%); (e) KOH (10%), MeOH, H2O (71%).
1228 was monobenzylated29 (Scheme 4). After subsequent
silylation, methyl ester 14 was reduced to alcohol 15 with
LiAlH4. For the preparation of the benzyl bromide 21 for
4-carboxypropoxyenterolactone 7, O-benzylcatechol 1730
was Reimer–Tiemann formylated with CHCl3 in aqueous
NaOH to afford the aldehyde 1831 (Scheme 5). Silyl ether
formation and aldehyde reduction to alcohol 20 was accom-
plished in the usual manner.
Treatment of the corresponding benzyl alcohols 10, 15
and 20 with phosphorous tribromide in ether afforded
the desired benzyl bromides 11, 16 and 21 (Schemes
Scheme 7. Reagents: (a) n-butyllithium, THF, Ϫ78ЊC, then 2-butenolide,
THF, Ϫ78ЊC, then benzyl bromide, 16, HMPA, THF, Ϫ78ЊC (18%); (b) n-
Bu4NϩFϪ, THF (99%); (c) BrCH2CO2Et, K2CO3, KI, acetone, reflux
(99%); (d) Raney nickel, ethanol, reflux (38%); (e) KOH (10%), MeOH,
H2O (50%).
3–5).32
Thioacetal derivatives of dibenzylbutyrolactone 23, 27 and
31 were prepared from the dithioacetal 2233 by reaction with
n-butyllithium and 2-butenolide in THF at Ϫ78ЊC followed
by in situ alkylation with the corresponding benzyl
bromides 11, 16 or 21 in the presence of HMPA (Schemes
6–8).26
Cleavage of the silyl ether protection with tetrabutylammo-
nium fluoride,27 and subsequent addition of ethyl bromo-
acetate or ethyl 4-bromobutyrate afforded the carboxylate
side chain as ethyl ester in the lignan framework. Simul-
taneous desulfurization and debenzylation were achieved by
treatment with Raney nickel in refluxing ethanol.26 Ester
hydrolysis under basic conditions completed the reaction
sequence to afford the free acids, trans-2,3-dibenzylbutyro-
lactones 5, 6 and 7 with the desired side chains.
Scheme 8. Reagents: (a) n-butyllithium, THF, Ϫ78ЊC, then 2-butenolide,
THF, Ϫ78ЊC, then benzyl bromide, 21, HMPA, THF, Ϫ78ЊC (27%); (b)
n-Bu4NϩFϪ, THF (98%); (c) Br(CH2)3CO2Et, K2CO3, KI, acetone, reflux
(98%); (d) Raney nickel, ethanol, reflux (97%); (e) KOH (10%), MeOH,
H2O (54%).
As already mentioned, for the synthesis of a carbon–carbon
Scheme 9. (a) allyl bromide, K2CO3, ethanol, reflux, (75%); (b) NaBH4, ethanol (79%); (c) N,N-dimethyl aniline, reflux; (d) benzyl chloride, t-BuOK, DMF,
ϩ70ЊC (74%); (e) PBr3, ether, 0ЊC (86%).

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T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1876
Scheme 10. Reagents: (a) n-butyllithium, THF, Ϫ78ЊC, then 2-butenolide, THF, Ϫ78ЊC, then benzyl bromide, 40, HMPA, THF, Ϫ78ЊC (41%);
(b) n-Bu4NϩFϪ, AIBN, toluene, 90ЊC (69%); (c) NaBH4, THF, DMSO, 35ЊC, then H2O, 5ЊC, and then H2O2 (30%), NaOH (51%); (d) PDC, DMF (77%);
(e) Raney nickel, ethanol, reflux (26%); (f) CH2N2, ether.
linked carboxylate side chain analogue a completely
different approach was chosen. A functionalized side
chain was generated by the preparation of benzyl bromide
40, available by the Claisen rearrangement of allyl
m-formylphenyl ether as a key step (Scheme 9).34
carboxyl group to the molecular framework, that could be
coupled to amino residues on protein and used after euro-
pium label as a tracer.20
The enterolactone derivative with carboxylic acid side chain
was synthesized by attachment to the benzyl bromide
moiety of the molecular framework either through a
carbon–carbon or carbon–oxygen bond. The most favor-
able positions in the aromatic ring were assumed to be in
ortho- and meta-position with respect to the phenolic
hydroxyl group in order to avoid any steric hindrance and
allow easy access to protein during the coupling procedure.
The length of the chain was not restricted.
3-Allyloxybenzaldehyde 35 was reduced to the alcohol 36,
which in refluxing N,N-dimethylaniline underwent
rearrangement to afford the two isomers of o-allyl phenol
37 and 38 in yields of 44% and 39%, respectively. The
O-benzylated isomer 39 underwent bromination in the
usual manner to afford the benzyl bromide 40.
The tandem conjugate addition reaction with benzyl
bromide 40 afforded the allylic dibenzylbutyrolactone 41
(Scheme 10).26
When the derivatives were used in analytical procedures,
some problems concerning structural features appeared.39
In compounds 7 and 8, where the side chain is located
next to the phenolic hydroxyl group, interaction between
hydroxy and carboxy groups was observed, causing lacto-
nization competing with the protein coupling. This problem
is due to the favorable position of these groups, and the
suitable length of the chain, which allows the intermo-
lecular esterification reaction. The reason for difficulties
concerning compound 5 is probably due to substitution
of one of the phenolic hydroxyl groups, which either
causes recognition problems or problems during europium
labeling.
The phenylthio groups were eliminated with tributyltin
hydride35 in the presence of AIBN initiator affording the
reduced product 42 without affecting the allyl group.36
The desired carboxy group was generated by a reaction
sequence involving the hydroboration–oxidation of alkene
42 to the primary alcohol 43,37 and subsequent oxidation
with pyridinium dichromate in DMF,38 yielding the
carboxylic acid 44. Benzyl ethers were cleaved with
Raney nickel in refluxing ethanol to afford the desired
4-carboxyethylenterolactone 8. As a confirmation of the
successful synthesis of 8, the product was converted
to the dimethyl ether methyl ester 45 by prolonged
exposure to ethereal diazomethane. The mass spectrum
of the resulting product (412, Mϩ) confirmed the methy-
lation of both the carboxyl group and the free phenolic
hydroxy groups.
Taking into consideration the disadvantages discussed
above, compound 6 should have most suitable structure
for the experiments. Hydroxy and carboxy groups are far
enough, and also the chain is short enough to avoid cycli-
zation problems. As a matter of fact, the reported experi-
mental results have demonstrated that the new TR-FIA
method for detection and quantification of plasma entero-
lactone utilized successfully the synthesized derivative 6 in
preparation of antiserum and tracer.23
The developed analytical method utilized an enterolactone
derivative for the production of antiserum and tracer.23 The
synthesis of a carboxyalkyl ether derivative introduces a

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T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1877
Experimental
(m, 6 ArH), 7.68–7.78 (m, 4 ArH); 13C NMR (CDCl3) d
19.38, 26.44, 33.27, 119.82, 120.53, 121.73, 127.85, 129.51,
130.01, 132.70, 135.56, 138.95, 155.78; HRMS m/z calcd
for C23H25OSiBr (Mϩ) 424.0858, found 424.0870.
All experiments were monitored by thin layer chroma-
tography using aluminum based, precoated silica gel sheets
(Merck 60 F254, layer thickness 0.2 mm). Silica gel 60 (230–
400 mesh, Merck) was used for flash column chroma-
tography. Melting points were determined on an Electro-
thermal melting point apparatus in an open capillary tube
and are uncorrected. IR spectra were obtained with a
Methyl 3-benzyloxy-5-hydroxybenzoate (13). To a solu-
tion of 12 (20.0 g, 0.12 mol) in acetone (300 mL) was added
benzyl bromide (20.3 g, 0.12 mol) and powdered K2CO3
(16.4 g, 0.12 mol). The reaction was refluxed for 5 h. The
solid material was filtered off and washed with acetone. The
filtrate, which consisted of starting material and both diben-
zyl and monobenzyl ether, was evaporated. Most of the
dibenzyl ether was removed by crystallization from ethanol,
and the resulting residue was purified by flash column
chromatography (EtOAc–hexane 1:1) to give 13 as a
white solid (10.1 g, 33%): mp 98.5ЊC (lit.40 mp 98–99ЊC);
1H NMR (acetone-d6) d 3.85 (3 H, s, CH3), 5.14 (2 H, s,
CH2), 6.74 (t, Jˆ2.3 Hz, ArH), 7.11 (dd, Jˆ1.1, 2.4 Hz,
ArH), 7.15 (dd, Jˆ1.4, 2.5 Hz, ArH), 7.35–7.51 (m, 5
ArH), 8.78 (1 H, s, OH).
Perkin–Elmer 125 spectrophotometer. H and 13C NMR
1
spectra were recorded on 60 MHz JEOL JNM-PMX 60 or
200 MHz Varian GEMINI spectrometer in CDCl3 or
acetone-d6 and chemical shifts are relative to tetramethyl-
silane (TMS) as an internal standard. EIMS and HRMS
were obtained using JEOL JMS-SX102 spectrometer. THF
was freshly distilled from sodium benzophenone ketyl,
diethyl ether from sodium, and DMF and DMSO from
CaH. Other solvents were of analytical grade. All commer-
cially available chemicals were used as supplied by the
manufacturers.
Methyl 3-benzyloxy-5-tert-butyldiphenylsilyloxybenzo-
ate (14). Following the same procedure as for 9, 13
(10.0 g, 0.039 mol) was converted to 14 as a gum
(19.0 g, 99%), which was used directly in the next step:
1H NMR (CDCl3) d 1.10 (9 H, s, 3 CH3), 3.82 (3 H, s,
CH3), 4.79 (2 H, s, CH2), 6.48 (t, Jˆ2.4 Hz, ArH), 7.14
(dd, Jˆ1.4, 2.2 Hz, ArH), 7.19 (dd, Jˆ1.4, 2.4 Hz,
ArH), 7.24–7.44 (m, 11 ArH), 7.71 (m, 4 ArH); 13C NMR
3-tert-Butyldiphenylsilyloxybenzaldehyde (9). 3-Hydroxy-
benzaldehyde (3.0 g, 0.025 mol) in dry DMF (7 mL) was
stirred at room temperature and tert-butyldiphenylsilyl-
chloride (10.1 g, 0.037 mol) and imidazole (3.3 g,
0.048 mol) was added to the solution. The reaction mixture
was stirred for 5 h, poured into water and extracted with
ether. The organic phase was washed with water and dried
over Na2SO4. Evaporation of the solvent gave 9 as a gum
1
(CDCl3)
d
19.41, 26.46, 52.07, 70.03, 108.89,
(8.8 g, 99%), which was used directly in the next step: H
111.47, 114.06, 127.47, 127.91, 128.01, 128.56, 130.08,
131.83, 132.55, 135.58, 136.49, 156.63, 159.38, 166.81;
HRMS m/z calcd for C31H32O4Si (Mϩ) 496.2070, found
496.2083.
NMR (CDCl3) d 1.12 (9 H, s, 3 CH3), 6.95 (m, ArH), 7.15–
7.48 (m, 10 ArH), 7.68–7.77 (m, 3 ArH), 9.80 (1 H, s,
CHO); 13C NMR (CDCl3) d 19.34, 26.36, 120.35, 122.79,
125.97, 127.94, 129.57, 130.17, 132.25, 135.49, 137.73,
156.29, 192.19; HRMS m/z calcd for C23H24O2Si (Mϩ)
360.1546, found 360.1537.
3-Benzyloxy-5-tert-butyldiphenylsilyloxybenzyl alcohol
(15). A solution of 14 (19.0 g, 0.038 mol) in dry ether
(330 mL) was carefully added to LiAlH4 (3.7 g,
0.097 mol) in dry ether (70 mL), and the reaction mixture
was refluxed for 1 h. The mixture was treated carefully with
ice-water and 1N NaOH (20 mL). The solids were filtered
off and washed with ether, which was dried over MgSO4 and
evaporated. The residue was purified by flash column chro-
matography (pentane–ether 1:1) to give 15 as a white solid
(4.9 g, 27%, mp 57ЊC): 1H NMR (CDCl3) d 1.09 (9 H, s, 3
CH3), 4.46 (1 H, d, Jˆ6.2 Hz, CH2), 4.77 (1 H, s, CH2), 6.28
(t, Jˆ2.2 Hz, ArH), 6.39 (m, ArH), 6.54 (m, ArH), 7.25–
7.43 (m, 11 ArH), 7.68 (d, Jˆ2.0 Hz, 2 ArH), 7.72 (d,
Jˆ1.6 Hz, 2 ArH); 13C NMR (CDCl3) d 19.39, 26.46,
65.10, 69.81, 105.72, 106.49, 110.98, 127.44, 127.82,
127.89, 128.52, 129.97, 132.88, 135.59, 136.86, 143.05,
156.86, 159.78; HRMS m/z calcd for C30H32O3Si (Mϩ)
468.2121, found 468.2128.
3-tert-Butyldiphenylsilyloxybenzyl alcohol (10). To a
stirred solution of 9 (8.8 g, 0.024 mol) in 94% ethanol
(120 mL) was added in small portions NaBH4 (1.85 g,
0.049 mol). The mixture was stirred at room temperature
for 4 h, ethanol evaporated, and the residue was acidified
with 2N H2SO4 and extracted with ether. The organic phase
was washed with brine and dried over MgSO4. Evaporation
of the solvent gave 10 as a gum (8.7 g, 99%), which was
used directly in the next step: 1H NMR (CDCl3) d 1.10 (9 H,
s, 3 CH3), 2.31 (1 H, br s, OH), 4.50 (2 H, s, CH2), 6.62 (m,
ArH), 6.83 (m, 2 ArH), 7.03 (m, ArH), 7.28–7.45 (m, 6
ArH), 7.68–7.78 (m, 4 ArH); 13C NMR (CDCl3) d
19.36, 26.43, 65.03, 118.33, 118.89, 119.55, 127.79,
129.32, 129.93, 132.88, 135.56, 142.34, 155.84;
HRMS m/z calcd for C23H26O2Si (Mϩ) 362.1702, found
362.1699.
3-Benzyloxy-5-tert-butyldiphenylsilyloxybenzyl bromide
(16). Following the same procedure as for 11, 15 (4.89 g,
0.010 mol) was converted to 16 as an oil (4.96 g, 89%),
which was used directly in the next step: 1H NMR
(CDCl3) d 1.09 (9 H, s, 3 CH3), 4.25 (2 H, s, CH2), 4.75
(2 H, s, CH2), 6.28 (m, ArH), 6.44 (m, ArH), 6.55 (m,
ArH), 7.25–7.44 (m, 11 ArH), 7.67 (m, 2 ArH), 7.71
(m, 2 ArH); 13C NMR (CDCl3) d 19.39, 26.46, 33.34,
69.88, 106.57, 108.91, 113.37, 127.49, 127.88, 127.97,
3-tert-Butyldiphenylsilyloxybenzyl bromide (11). To a
stirred solution of 10 (8.8 g, 0.024 mol) in dry ether
(85 mL) in an ice bath was added dropwise PBr3 (4.0 g,
0.015 mol). The solution was kept at ϩ4ЊC for two days.
The reaction mixture was washed with water (5×) and dried
over Na2SO4. Evaporation of the solvent gave 11 as a gum
(8.9 g, 86%), which was used directly in the next step: H
NMR (CDCl3) d 1.10 (9 H, s, 3 CH3), 4.30 (2 H, s, CH2),
6.64 (m, ArH), 6.87 (m, 2 ArH), 7.02 (m, ArH), 7.29–7.48
1

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T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1878
128.54, 130.03, 132.70, 135.58, 136.65, 139.39, 156.77,
159.64.
(^)-2-(3-Hydroxybenzyl)-3-[30-benzyloxy-a,a-bis(phenyl-
thio)benzyl]butyrolactone (24). To a stirred solution of 23
(1.0 g, 1.2 mmol) in dry THF (5 mL) maintained under
argon in an ice bath was added dropwise a 1 M solution
of tetrabutylammoniumfluoride in THF (5.9 mL, 5.9
mmol). The reaction mixture was stirred in an ice bath for
10 min, the bath was removed and stirring continued at
room temperature for 3 h. The reaction was quenched
with water and the mixture was extracted with ether. The
organic phase was washed with water and dried over
Na2SO4. Evaporation of the solvent gave 24 as an amor-
phous solid (0.71 g, 99%), which was used directly in the
next step: 1H NMR (CDCl3) d 2.77 (1 H, dd, Jˆ6.8,
13.2 Hz, H-7), 2.97 (1 H, m, H-80), 3.09 (1 H, dd, Jˆ4.6,
14.1 Hz, H-7), 3.31 (1 H, m, H-8), 3.55 (1 H, dd, Jˆ8.5,
10.2 Hz, H-90), 4.35 (1 H, dd, Jˆ3.4, 10.2 Hz, H-90),
4.98 (2 H, s, CH2), 6.46 (m, ArH), 6.70 (m, ArH), 6.90
(m, 2 ArH), 7.06 (m, 2 ArH), 7.22–7.44 (m, 15 ArH), 7.72
(m, 2 ArH).
3-Benzyloxy-4-tert-butyldiphenylsilyloxybenzaldehyde
(19). Following the same procedure as for 9, 18 (11.7 g,
0.051 mol) was converted to 19 as an oil (16.9 g, 71%),
after purification by flash column chromatography
1
(CH2Cl2): H NMR (CDCl3, 60 MHz) d 1.12 (9 H, s, 3
CH3), 5.16 (2 H, s, CH2), 6.80–8.10 (m, 18 ArH), 10.00
(1 H, s, CHO).
3-Benzyloxy-4-tert-butyldiphenylsilyloxybenzyl alcohol
(20). Following the same procedure as for 10, 19 (8.7 g,
0.019 mol) was converted to 20 as a gum (5.3 g, 61%)
after purification by flash column chromatography (ace-
1
tone–cyclohexane 2:5): H NMR (CDCl3, 60 MHz) d 1.20
(9 H, s, 3 CH3), 2.52 (1 H, br s, OH), 4.45 (2 H, s, CH2), 5.06
(2 H, s, CH2), 6.79 (m, 2 ArH), 7.08 (m, ArH), 7.32–7.63
(m, 11 ArH), 7.84–8.10 (m, 4 ArH); 13C NMR (CDCl3) d
19.60, 26.52, 65.21, 70.55, 113.07, 119.68, 120.09, 127.53,
127.61, 127.69, 128.33, 129.69, 133.41, 134.11, 135.44,
137.12, 144.94, 149.80.
(^)-2-(3-Ethoxycarbonylmethoxybenzyl)-3-[30-benzyl-
oxy-a,a-bis(phenylthio)benzyl]butyrolactone (25). To a
solution of 24 (0.68 g, 1.12 mmol) in acetone (20 mL) was
added ethyl bromoacetate (0.21 g, 1.24 mmol), K2CO3
(0.31 g, 2.25 mmol), and KI (0.09 g, 0.56 mmol). The reac-
tion was refluxed for 4 h. The mixture was cooled, the solids
were filtered off and rinsed with acetone. The solvent was
evaporated and the residue extracted with ether. The organic
phase was washed with 2 N NaOH solution and water, and
dried over Na2SO4. Evaporation of the solvent afforded 25
as a gum (0.75 g, 96%), which was used directly in the next
step: 1H NMR (CDCl3) d 1.29 (3 H, t, Jˆ7.1 Hz, CH3), 2.78
(1 H, dd, Jˆ5.7, 13.5 Hz, H-7), 2.92 (1 H, m, H-80), 3.09 (1
H, dd, Jˆ5.1, 13.6 Hz, H-7), 3.29 (1 H, m, H-8), 3.51 (1 H,
dd, Jˆ8.6, 10.2 Hz, H-90), 4.20–4.35 (3 H, q and dd, over-
lapping, CH2, H-90), 4.51 (2 H, s, CH2), 4.96 (2 H, s, CH2),
6.54 (m, 2 ArH), 6.75–6.91 (m, 2 ArH), 7.06–7.42 (m, 15
ArH), 7.70 (m, 4 ArH); 13C NMR (CDCl3) d 14.08, 36.90,
44.39, 47.43, 61.35, 65.18, 68.02, 70.03, 72.96, 113.77,
115.07, 115.28, 116.32, 121.19, 122.92, 127.53, 128.02,
128.10, 128.67, 128.70, 128.80, 129.47, 129.59,
129.78, 133.01, 136.29, 138.50, 157.99, 158.79, 169.68,
178.50.
3-Benzyloxy-4-tert-butyldiphenylsilyloxybenzyl bromide
(21). Following the same procedure as for 11, 20 (5.3 g,
0.011 mol) was converted to 21 as a white solid (5.1 g,
87%, mp 71ЊC) after crystallization from light petroleum
1
(bp 40–60ЊC): H NMR (CDCl3, 60 MHz) d 1.10 (9 H, s,
3 CH3), 4.61 (2 H, s, CH2), 5.18 (2 H, s, CH2), 6.91 (m, 2
ArH), 7.20 (m, ArH), 7.40–7.80 (m, 11 ArH), 7.83–8.18 (m,
4 ArH); 13C NMR (CDCl3) d 19.61, 26.48, 34.38, 70.61,
114.79, 120.13, 121.83, 127.57, 127.66, 127.78, 128.37,
129.78, 130.74, 133.22, 135.43, 136.90, 145.71, 149.81.
(^)-2-(3-tert-Butyldiphenylsilyloxybenzyl)-3-[30-benzyl-
oxy-a,a-bis(phenylthio)benzyl]butyrolactone (23). To a
stirred solution of 22 (5.0 g, 12 mmol) in dry THF
(30 mL) maintained under argon at Ϫ78ЊC was added a
solution of n-butyllithium (8.5 mL, 12 mmol) in n-hexane.
The resulting solution was stirred for 2.5 h, and a solution of
2-butenolide (1.0 g, 12 mmol) dissolved in dry THF (4 mL)
was added. The reaction mixture was stirred for a further
2.5 h at Ϫ78ЊC and then treated dropwise with a solution of
11 (5.1 g, 12 mmol) and HMPA (2.1 mL, 12 mmol) in dry
THF (10 mL). The reaction mixture was allowed to reach
room temperature overnight, and then the reaction was
quenched with water. The mixture was extracted with
EtOAc, washed with water and dried over Na2SO4.
Evaporation of the solvent left a gum, which was purified
by flash column chromatography (CH2Cl2–pentane 2:1) to
give 23 as an amorphous solid (1.0 g, 10%): IR (film)
(^)-2-(3-Ethoxycarbonylmethoxybenzyl)-3-(30-hydroxy-
benzyl)butyrolactone (26). A suspension of 25 (0.75 g,
1.1 mmol) in ethanol (75 mL) and W-2 Raney nickel
(12 g) was refluxed for 3 h. The catalyst was removed by
filtration and rinsed with acetone. Evaporation of the
solvents gave a gum, which was purified by flash column
chromatography (acetone–cyclohexane 1:2) to give 26 as a
1
1770 cmϪ1 (CvO); H NMR (CDCl3) d 1.09 (9 H, s, 3
gum (0.18 g, 43%): IR (film) 1750, 1735 cmϪ1; H NMR
1
CH3), 2.68 (1 H, dd, Jˆ5.8, 13.8 Hz, H-7), 2.90 (2 H, m
and dd, overlapping, H-80, H-7), 3.25 (1 H, m, H-8), 3.43 (1
H, dd, Jˆ8.5, 10.4 Hz, H-90), 4.32 (1 H, dd, Jˆ3.0, 10.0 Hz,
H-90), 4.96 (2 H, s, CH2), 6.40 (d, Jˆ7.5 Hz, ArH), 6.51 (dd,
Jˆ2.6, 8.1 Hz, ArH), 6.61 (m, 2 ArH), 6.83 (m, 2 ArH),
7.16–7.42 (m, 23 ArH), 7.65 (m, 4 ArH); 13C NMR
(CDCl3) d 19.39, 26.47, 36.74, 44.56, 47.64, 67.97, 70.01,
72.90, 114.97, 116.33, 118.43, 120.90, 121.07, 122.36,
127.54, 127.80, 128.07, 128.64, 128.72, 129.26, 129.51,
129.95, 130.83, 132.28, 132.86, 133.34, 135.55, 135.95,
136.74, 138.25, 139.35, 155.82, 158.76, 178.43.
(acetone-d6) d 1.24 (3 H, t, Jˆ7.0 Hz, CH3), 2.51–3.02 (6
H, m, H-70, H-80, H-8, H-7), 3.89 (1 H, m, H-90), 4.07 (1 H,
m, H-90), 4.20 (2 H, q, Jˆ7.1 Hz, CH2), 4.72 (2 H, s, CH2),
6.59–6.72 (m, 3 ArH), 6.79–6.90 (m, 3 ArH), 7.10 (t,
Jˆ8.1 Hz, ArH), 7.24 (t, Jˆ8.0 Hz, ArH), 8.23 (1 H, s,
OH); HRMS m/z calcd for C22H24O6 (Mϩ) 384.1573,
found 384.1582; EIMS m/z 384 (Mϩ, 92), 277 (31), 193
(37), 191 (20), 108 (100), 107 (62); 13C NMR (acetone-d6)
d 14.43, 35.40, 38.72, 42.19, 46.83, 61.52, 65.73, 71.56,
113.72, 114.38, 116.60, 120.73, 123.44, 130.43, 130.51,
141.19, 141.41, 158.63, 159.40, 169.60, 178.90.

¨
¨
T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1879
(^)-trans-2-(3-Carboxymethoxybenzyl)-3-(30-hydroxy-
benzyl)butyrolactone (5). To a stirred solution of 26
(0.18 g, 0.47 mmol) in aqueous methanol (50%, 28 mL) at
room temperature was added aqueous KOH (10%, 5.5 mL).
The reaction was stirred for 3 h, methanol was evaporated,
and water was added to the residue. The inorganic phase
was washed with ether (5×), acidified with 2N H2SO4 and
extracted with ether. The organic phase was washed with
water and dried over Na2SO4. Evaporation of the solvent
gave 5 as an amorphous solid (0.12 g, 71%): IR (film)
6.15 (m, ArH), 6.28 (m, ArH), 6.45 (m, ArH), 6.90 (m,
ArH), 7.13–7.45 (m, 20 ArH), 7.71 (m, 3 ArH); 13C NMR
(CDCl3) d 14.07, 37.12, 44.40, 47.59, 61.37, 65.17, 68.16,
69.97, 70.01, 72.90, 101.09, 107.78, 109.34, 115.06, 116.32,
121.20, 127.53, 127.56, 128.08, 128.63, 128.66, 128.70,
128.76, 128.87, 129.42, 129.56, 133.13, 135.22, 136.09,
139.29, 158.76, 159.02, 160.06, 168.69, 178.65.
(^)-2-(3-Hydroxy-5-ethoxycarbonylmethoxybenzyl)-3-
(30-hydroxybenzyl)butyrolactone (30). Following the
same procedure as for 26, 29 (1.37 g, 1.7 mmol) was
converted to 30 as an amorphous solid (0.26 g, 38%) after
purification by flash column chromatography (EtOAc–hex-
ane 4:1): IR (film) 1750, 1735 cmϪ1; 1H NMR (acetone-d6)
d 1.24 (3 H, t, Jˆ7.1 Hz, CH3), 2.56 and 2.68 (4 H, m and m,
overlapping, H-70, H-80, H-8), 2.87 (2 H, m, H-7), 3.88 (1 H,
m, H-90), 4.05 (1 H, m, H-90), 4.20 (2 H, q, Jˆ7.1 Hz, CH2),
4.65 (2 H, s, CH2), 6.30 (t, Jˆ2.3 Hz, ArH), 6.36 (dd, Jˆ1.4,
2.2 Hz, ArH), 6.42 (dd, Jˆ1.6, 1.8 Hz, ArH), 6.65 (m, 3
ArH), 7.10 (t, Jˆ8.0 Hz, ArH), 8.22 (1 H, br s, OH) 8.41
(1 H, br s, OH); HRMS m/z calcd for C22H24O7 (Mϩ)
400.1522, found 400.1516; EIMS m/z 400 (Mϩ, 55), 293
(11), 210 (100), 209 (6), 191 (3), 107 (15); 13C NMR
(acetone-d6) d 14.20, 35.27, 38.49, 41.91, 46.47, 61.26,
65.53, 71.30, 101.13, 107.55, 110.39, 114.14, 116.36,
120.54, 130.28, 141.23, 141.63, 158.40, 159.43, 160.29,
169.38, 178.68.
2990 (br), 1760, 1710 cmϪ1 1H NMR (acetone-d6) d
;
2.48–2.79 (4 H, m, H-70, H-80, H-8), 2.85–3.08 (2 H, m,
H-7), 3.89 (1 H, m, H-90), 4.07 (1 H, m, H-90), 4.72 (2 H, s,
CH2), 6.59–6.71 (m, 3 ArH), 6.78–6.92 (m, 3 ArH), 7.09 (t,
Jˆ8.0 Hz, ArH), 7.24 (t, Jˆ8.0 Hz, ArH), 8.03 (1 H, s, OH);
13C NMR (acetone-d6) d 35.42, 38.70, 42.20, 46.75, 65.26,
71.49, 113.50, 114.21, 116.43, 116.56, 120.64, 123.20,
130.30, 130.39, 141.02, 141.28, 158.35, 159.21, 178.68,
180.34; HRMS m/z calcd for C20H20O6 (Mϩ) 356.1260,
found 356.1261; EIMS m/z 356 (Mϩ, 63), 338 (MϪH2O,
3), 249 (27), 191 (18), 165 (28), 108 (100), 107 (42).
(^)-2-(3-Benzyloxy-5-tert-butyldiphenylsilyloxybenzyl)-
3-[30-benzyloxy-a,a-bis(phenylthio)benzyl]butyrolac-
tone (27). In a similar manner as for 23 using benzyl bro-
mide 16 (4.96 g, 9.3 mmol) in place of benzyl bromide 11 to
give 27 as an amorphous solid (1.63 g, 18%): IR (film)
1
1770 cmϪ1 (CvO); H NMR (CDCl3) d 1.08 (9 H, s, 3
CH3), 2.62 (1 H, dd, Jˆ5.5, 13.6 Hz, H-7), 2.90 (2 H, m,
H-80, H-7), 3.24 (1 H, m, H-8), 3.49 (1 H, dd, Jˆ8.4,
10.0 Hz, H-90), 4.32 (1 H, dd, Jˆ2.9, 10.2 Hz, H-90), 4.64
(2 H, s, CH2), 4.95 (2 H, s, CH2), 6.18 (m, 2 ArH), 6.88 (m,
ArH), 7.14–7.39 (m, 30 ArH), 7.69 (m, 4 ArH); 13C NMR
(CDCl3) d 19.42, 26.47, 36.94, 44.56, 47.86, 68.10, 69.63,
70.01, 72.86, 105.55, 109.28, 113.80, 114.98, 116.35,
121.09, 127.39, 127.55, 127.85, 128.06, 128.19, 128.50,
128.65, 128.70, 129.20, 129.48, 130.00, 130.91, 132.21,
132.88, 133.54, 135.22, 135.59, 135.68, 136.76, 136.82,
138.87, 139.47, 156.80, 158.72, 159.52, 178.50.
(^)-trans-2-(3-Hydroxy-5-carboxymethoxybenzyl)-3-(30-
hydroxybenzyl)butyrolactone (6). Following the same
procedure as for 5, 30 (0.22 g, 0.55 mmol) was converted
to 6 as a white solid (0.10 g, 50%, mp 130–133ЊC) after
crystallization from chloroform: IR (film) 2990 (br), 1750,
1710 cmϪ1; 1H NMR (acetone-d6) d 2.56 and 2.68 (4 H, m
and m, overlapping, H-70, H-80, H-8), 2.89 (2 H, m, H-7),
3.89 (1 H, m, H-90), 4.06 (1 H, dd, Jˆ7.0, 8.6 Hz, H-90),
4.67 (2 H, s, CH2), 6.32 (t, Jˆ2.3 Hz, ArH), 6.38 (dd, Jˆ1.4,
2.2 Hz, ArH), 6.43 (dd, Jˆ1.4, 2.0 Hz, ArH), 6.64 (m, 3
ArH), 7.09 (t, Jˆ8.1 Hz, ArH), 8.43 (2 H, s, OH); 13C
NMR (acetone-d6) d 35.50, 38.71, 42.16, 46.65, 65.31,
71.48, 101.16, 107.82, 110.37, 114.23, 116.46, 120.68,
130.40, 141.33, 141.72, 158.37, 159.48, 160.35,
178.72, 180.37; HRMS m/z calcd for C20H20O7 (Mϩ)
372.1210, found 372.1212; EIMS m/z 372 (Mϩ, 43),
354 (MϪH2O, 2), 265 (11), 191 (7), 182 (100), 181 (12),
107 (22).
(^)-2-(3-Benzyloxy-5-hydroxybenzyl)-3-[30-benzyloxy-
a,a-bis(phenylthio)benzyl]butyrolactone (28). Following
the same procedure as for 24, 27 (1.63 g, 1.7 mmol) was
converted to 28 as a gum (1.21 g, 99%), which was used
directly in the next step: 1H NMR (CDCl3) d 2.72 (1 H, dd,
Jˆ6.0, 13.8 Hz, H-7), 2.96 (1 H, m, overlapping, H-80), 3.05
(1 H, dd, Jˆ5.0, 13.0 Hz, H-7), 3.28 (1 H, m, H-8), 3.62 (1
H, dd, Jˆ8.4, 10.2 Hz, H-90), 4.37 (1 H, dd, Jˆ3.5, 10.1 Hz,
H-90), 4.93 (2 H, s, CH2), 4.97 (2 H, s, CH2), 6.02 (m, ArH),
6.20 (m, ArH), 6.36 (m, ArH), 6.90 (m, ArH), 7.16–7.45 (m,
20 ArH), 7.71 (m, 3 ArH).
(^)-2-(3-Benzyloxy-4-tert-butyldiphenylsilyloxybenzyl)-
3-[30-benzyloxy-a,a-bis(phenylthio)benzyl]butyrolac-
tone (31). In a similar manner as for 23 using benzyl bro-
mide 21 (6.7 g, 0.013 mol) in place of benzyl bromide 11 to
give 31 as an amorphous solid (3.26 g, 27%): IR (film)
1
(^)-2-(3-Benzyloxy-5-ethoxycarbonylmethoxybenzyl)-
3-[30-benzyloxy-a,a-bis(phenylthio)benzyl]butyrolac-
tone (29). Following the same procedure as for 25, 28
(1.2 g, 1.7 mmol) was converted to 29 as a gum (1.34 g,
1770 cmϪ1 (CvO); H NMR (CDCl3) d 1.07 (9 H, s, 3
CH3), 2.62 (1 H, dd, Jˆ5.6, 13.6 Hz, H-7), 2.78 (1 H, m,
H-80), 3.02 (1 H, dd, Jˆ4.4, 13.6 Hz, H-7), 3.10–3.28 (2 H,
m, H-8, H-90), 4.28 (1 H, dd, Jˆ2.9, 9.9 Hz, H-90), 4.82 (2
H, s, CH2), 4.97 (2 H, s, CH2), 6.03 (1 H, dd, Jˆ1.8, 8.0 Hz,
ArH), 6.48 (1 H, d, Jˆ8.2 Hz, ArH), 6.57 (1 H, d, Jˆ2.0 Hz,
ArH), 6.90 (m, ArH), 7.05–7.43 (m, 30 ArH), 7.71 (t,
Jˆ7.5 Hz, 3 ArH); 13C NMR (CDCl3) d 19.57, 26.51,
36.65, 44.57, 47.12, 68.09, 70.05, 70.46, 73.02, 114.59,
114.93, 116.35, 120.25, 121.10, 121.80, 127.56, 127.62,
127.66, 127.71, 127.93, 128.09, 128.33, 128.66, 128.72,
1
99%), which was used directly in the next step: H NMR
(CDCl3) d 1.28 (3 H, t, Jˆ7.1 Hz, CH3), 2.75 (1 H, dd,
Jˆ5.7, 13.4 Hz, H-7), 2.99 (1 H, m, overlapping, H-80),
3.08 (1 H, dd, Jˆ5.1, 13.5 Hz, H-7), 3.29 (1 H, m, H-8),
3.60 (1 H, dd, Jˆ8.4, 10.1 Hz, H-90), 4.26 (2 H, q,
Jˆ7.2 Hz, CH2), 4.36 (1 H, dd, Jˆ3.3, 10.2 Hz, H-90),
4.49 (2 H, s, CH2), 4.93 (2 H, s, CH2), 4.96 (2 H, s, CH2),

¨ ¨
T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1880
129.42, 129.54, 129.82, 130.55, 132.38, 132.98, 133.29,
133.44, 135.55, 136.41, 136.73, 136.93, 139.23, 144.43,
149.90, 158.79, 178.87.
(MϪH2O, 5), 314 (35), 191 (7), 180 (28), 123 (100), 107
(20).
3-(2-Propenyloxy)benzaldehyde (35). To a solution of
3-hydroxybenzaldehyde (61.1 g, 0.50 mol) in ethanol
(500 mL) was added allyl bromide (60.5 g, 0.50 mol) and
powdered anhydrous K2CO3 (138.2 g, 1.0 mol). The reac-
tion mixture was refluxed overnight, poured into water and
extracted with ether. The organic phase was washed with
2% KOH and water, and dried over MgSO4. Evaporation
left a crude product, which was distilled at reduced pressure
to afford 35 as a colorless liquid (61.1 g, 75%): bp 97ЊC
(0.85 mm) [lit.41 bp 71–74ЊC (0.3–0.4 mm)]: 1H NMR
(CDCl3) d 4.60 (2 H, d, Jˆ7.0 Hz, CH2), 5.39 (2 H, m,
CH2), 6.02 (1 H, m, CH), 7.18 (m, ArH), 7.45 (m, 3 ArH),
9.97 (1 H, s, CHO); HRMS m/z calcd for C10H10O2 (Mϩ)
162.0681, found 162.0666.
(^)-2-(3-Benzyloxy-4-hydroxybenzyl)-3-[30-benzyloxy-
a,a-bis(phenylthio)benzyl]butyrolactone (32). Following
the same procedure as for 24, 31 (0.8 g, 0.84 mmol) was
converted to 32 as a gum (0.59 g, 98%), which was used
directly in the next step: 1H NMR (CDCl3) d 2.76 (1 H, dd,
Jˆ5.4, 13.6 Hz, H-7), 2.94 (1 H, m, H-80), 3.07 (1 H, dd,
Jˆ4.7, 13.6 Hz, H-7), 3.28 (1 H, m, H-8), 3.52 (1 H, dd,
Jˆ8.4, 10.1 Hz, H-90), 4.33 (1 H, dd, Jˆ3.7, 10.1 Hz, H-90),
4.96 (2 H, s, CH2), 4.97 (2 H, s, CH2), 6.36 (1 H, dd, Jˆ1.8,
8.0 Hz, ArH), 6.66 (1 H, d, Jˆ1.8 Hz, ArH), 6.74 (1 H, d,
Jˆ8.0 Hz, ArH), 6.90 (m, ArH), 7.17–7.41 (m, 21 ArH),
7.72 (m, 2 ArH).
(^)-2-(3-Benzyloxy-4-ethoxycarbonylpropyloxybenzyl)-
3-[30-benzyloxy-a,a-bis(phenylthio)benzyl]butyrolac-
tone (33). In a similar manner as for 25 using ethyl
4-bromobutyrate (0.18 g, 0.91 mmol) in place of ethyl
bromoacetate, 32 (0.59 g, 0.83 mol) was converted to 33
as a gum (0.67 g, 98%) after purification by flash column
chromatography (acetone–cyclohexane 1:2): 1H NMR
(CDCl3) d 1.26 (3 H, t, Jˆ7.0 Hz, CH3), 2.14 (2 H, m,
CH2), 2.55 (2 H, t, Jˆ7.0 Hz, CH2), 2.76 (1 H, m, H-7),
2.93 (1 H, m, H-80), 3.07 (1 H, dd, Jˆ4.6, 12.8 Hz, H-7),
3.27 (1 H, m, H-8), 3.50 (1 H, dd, Jˆ8.4, 10.1 Hz, H-90),
4.04 (2 H, t, Jˆ6.3 Hz, CH2), 4.14 (2 H, q, Jˆ7.1 Hz, CH2),
4.32 (1 H, m, 90-H), 4.97 (2 H, s, CH2), 4.98 (2 H, s, CH2),
6.40 (m, ArH), 6.68 (m, 2 ArH), 6.90 (m, ArH), 7.22–7.44
(m, 23 ArH).
3-(2-Propenyloxy)benzyl alcohol (36). To a stirred solu-
tion of 35 (61.0 g, 0.38 mol) in 94% ethanol (150 mL) at
room temperature was added in small portions NaBH4
(6.5 g, 0.17 mol). The reaction mixture was stirred for 5 h,
glacial acetic acid was added, and the mixture was neutra-
lized with 10% NaHCO3 and extracted with toluene. The
organic phase was dried over MgSO4. Evaporation left a
crude product, which was distilled under reduced pressure
to give 36 as a colorless liquid (48.7 g, 79%): bp 120ЊC
1
(1.3 mm): H NMR (CDCl3) d 2.21 (1 H, br s, OH), 4.55
(2 H, d, Jˆ7.0 Hz, CH2), 4.64 (2 H, s, CH2), 5.38 (2 H, m,
CH2), 6.08 (1 H, m, CH), 6.86 (dd, Jˆ2.1, 8.0 Hz, ArH),
6.94 (m, 2 ArH), 7.28 (t, Jˆ8.0 Hz, ArH); 13C NMR
(CDCl3) d 64.66, 68.55, 112.96, 113.76, 117.52, 119.18,
129.40, 133.16, 142.53, 158.67; HRMS m/z calcd for
C10H12O2 (Mϩ) 164.0837, found 164.0840.
(^)-2-(3-Hydroxy-4-ethoxycarbonylpropyloxybenzyl)-3-
(30-hydroxybenzyl)butyrolactone (34). Following the
same procedure as for 26, 33 (0.67 g, 0.81 mmol) was
converted to 34 as a gum (0.34 g, 97%), which was used
2-(2-Propenyl)-3-hydroxybenzyl alcohol and 3-hydroxy-
4-(2-propenyl)benzyl alcohol (37 and 38). A mixture of
compound 36 (48.3 g, 0.29 mol) and N,N-dimethylaniline
(95.6 g, 0.79 mol) was refluxed overnight. The reaction
mixture was acidified with 1N H2SO4 and extracted with
ether. The organic phase was dried over MgSO4. Evapo-
ration left a crude product, which was purified by flash
column chromatography (CH2Cl2–EtOAc 4:1) to give
compound 37 as an oil (21.3 g, 44%) and compound 38 as
glassy crystals (18.7 g, 39%, mp 64.5–65.5ЊC).
1
directly in the next step: IR (film) 1750, 1735 cmϪ1; H
NMR (CDCl3) d 1.27 (3 H, t, Jˆ7.1 Hz, CH3), 2.16 (2 H,
m, CH2), 2.26–2.61 (4 H, m, H-70, H-8, H-80), 2.52 (2 H, t,
Jˆ6.8 Hz, CH2), 2.79 (1 H, dd, Jˆ7.2, 14.0 Hz, H-7), 3.02
(1 H, dd, Jˆ4.4, 14.6 Hz, H-7), 3.89 (1 H, m, H-90), 4.05–
4.22 (5 H, m, t, q, overlapping, H-90, 2 CH2), 6.40 (m, ArH),
6.56–6.77 (m, 5 ArH), 7.12 (t, Jˆ7.7 Hz, ArH); HRMS m/z
calcd for C24H28O7 (Mϩ) 428.1835, found 428.1823; EIMS
m/z 428 (Mϩ, 15), 191 (3), 115 (100), 107 (6).
Compound 37. 1H NMR (acetone-d6) d 3.46 (2 H, d,
Jˆ6.0 Hz, CH2), 4.03 (1 H, t, Jˆ5.9 Hz, OHalcoholic), 4.60
(2 H, d, Jˆ6.0 Hz, CH2), 4.82–5.02 (2 H, m, CH2), 5.82–
6.08 (1 H, m, CH), 6.78 (d, Jˆ7.7 Hz, ArH), 6.99 (s, ArH),
7.34 (d, Jˆ7.7 Hz, ArH), 8.16 (1 H, s, OHphenolic); 13C NMR
(acetone-d6) d 34.72, 64.60, 114.30, 115.36, 118.77, 125.84,
130.62, 138.42, 142.84, 155.82; HRMS m/z calcd for
C10H12O2 (Mϩ) 164.0837, found 164.0846.
(^)-trans-2-(3-Hydroxy-4-carboxypropoxybenzyl)-3-(30-
hydroxybenzyl)butyrolactone (7). Following the same
procedure as for 5, 34 (0.34 g, 0.79 mmol) was converted
to 7 as greyish solid (0.17 g, 54%, mp 135ЊC) after crystal-
lization from chloroform: IR (film) 2990 (br), 1750,
1
1710 cmϪ1; H NMR (acetone-d6) d 2.07 (2 H, m, over-
lapping, CH2), 2.50–2.70 (6 H, m, overlapping, H-70,
H-80, H-8), 2.53 (2 H, t, Jˆ7.1 Hz, CH2), 2.77–2.99 (2 H,
m, H-7), 3.87 (1 H, m, H-90), 4.04 (2 H, m, overlapping,
H-90), 4.07 (2 H, t, Jˆ6.2 Hz, CH2), 6.59–6.89 (m, 6 ArH),
7.09 (t, Jˆ7.9 Hz, ArH), 8.30 (2 H, br s, OH); 13C NMR
(acetone-d6) d 25.44, 30.77, 34.72, 38.72, 41.99, 46.96,
68.71, 71.45, 113.34, 114.18, 116.41, 117.05, 120.65,
121.31, 130.40, 132.19, 141.40, 146.33, 147.52, 158.38,
174.78, 178.80; HRMS m/z calcd for C22H24O7 (Mϩ)
400.1522, found 400.1526; EIMS m/z 400 (Mϩ, 27), 382
Compound 38. 1H NMR (acetone-d6) d 3.37 (2 H, d,
Jˆ6.2 Hz, CH2), 4.09 (1 H, t, Jˆ6.0 Hz, OHalcoholic), 4.54
(2 H, d, Jˆ6.0 Hz, CH2), 5.10 (2 H, m, CH2), 5.89–6.09 (1
H, m, CH), 6.78 (d, Jˆ8.0 Hz, ArH), 6.80 (m, ArH), 7.06
(dd, Jˆ2.0, 7.7 Hz, ArH), 8.19 (1 H, s, OHphenolic).
3-Benzyloxy-4-(2-propenyl)benzyl alcohol (39). To a
stirred solution of 37 (9.4 g, 0.057 mol) in DMF (95 mL)

¨
¨
T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1881
maintained under argon was added KOBu-t (8.5 g,
0.076 mol) and benzyl chloride (7.2 g, 0.057 mol). The
reaction mixture was stirred at ϩ70ЊC for 2 h, quenched
with water, and extracted with ether. The organic phase
was dried over MgSO4. Evaporation of the solvent left a
crude product, which was purified by flash column chroma-
tography (CH2Cl2–EtOAc 9:1) to afford 39 as a solid
hexane 1:4) to give 42 as an oil (1.8 g, 69%): IR (film)
1
1770, 1610, 1580, 1257, 1021 cmϪ1; H NMR (CDCl3) d
2.41–2.62 (4 H, m, H-70, H-80, H-8), 2.95 (2 H, m, H-7),
3.40 (2 H, d, Jˆ6.8 Hz, CH2), 3.80 (1 H, dd, Jˆ7.2, 9.0 Hz,
H-90), 4.05 (1 H, dd, Jˆ6.4, 9.2 Hz, H-90), 5.00 (2 H, s,
CH2), 5.03 (2 H, s, CH2), 5.07 (2 H, m, overlapping,
CH2), 5.88–6.09 (1 H, m, CH), 6.57 (d, Jˆ7.4 Hz, 2
ArH), 6.70 (d, Jˆ9.2 Hz, 2 ArH), 6.82 (dd, Jˆ2.1, 7.9 Hz,
ArH), 7.06–7.51 (m, 12 ArH); 13C NMR (CDCl3) d 34.07,
34.80, 38.40, 40.90, 46.34, 69.79, 69.90, 71.11, 112.62,
112.85, 115.57, 121.25, 121.71, 127.19, 127.49, 127.70,
127.78, 128.09, 128.54, 128.66, 129.80, 129.94, 136.83,
136.91, 137.25, 139.61, 156.53, 159.04, 178.67; HRMS
m/z calcd for C35H34O4 (Mϩ) 518.2457, found 518.2470;
EIMS m/z 518 (Mϩ, 22), 427 (23), 106 (78), 91 (100).
1
(10.6 g, 74%, mp 42–48ЊC): H NMR (CDCl3) d 3.43 (2
H, d, Jˆ6.7 Hz, CH2), 4.61 (2 H, s, CH2), 5.00–5.11 (2 H,
m, overlapping, CH2), 5.07 (2 H, s, CH2), 5.90–6.10 (1 H,
m, CH), 6.88 (d, Jˆ7.4 Hz, ArH), 6.95 (s, ArH), 7.14 (d,
Jˆ7.6 Hz, ArH), 7.30–7.46 (m, 5 ArH); 13C NMR (CDCl3)
d 34.18, 65.31, 69.87, 110.38, 115.55, 119.26, 127.17,
127.82, 128.53, 129.98, 136.90, 137.27, 140.34, 156.62;
HRMS m/z calcd for C17H18O2 (Mϩ) 254.1307, found
254.1316.
(^)-2-[3-Benzyloxy-4-(3-hydroxypropyl)benzyl]-3-(30-
benzyloxybenzyl)butyrolactone (43). A solution of 42
(1.8 g, 3.5 mmol) in dry THF (40 mL) was added to
NaBH4 (0.09 g, 2.4 mmol). The reaction mixture was heated
to 35ЊC, and dimethyl sulfate (0.26 g, 2.1 mmol) was added
dropwise with stirring during 5 min. The reaction was
stirred for 4 h, cooled to ϩ5ЊC, and quenched carefully
with water. The mixture was allowed to reach room
temperature, and 2N NaOH (1.7 mL) was added in one
portion. The mixture was cooled again, 30% H2O2
(0.6 mL) was added dropwise, and stirring was continued
for 20 min at room temperature. The mixture was extracted
with ether, washed with ice water and saturated NaCl
solution, and dried over MgSO4. Evaporation of the solvent
left a crude product, which was purified by flash column
chromatography (EtOAc–cyclohexane 1:1) to afford 43 as
a gum (0.94 g, 51%): IR (film) 3430 (br), 1760, 1250,
3-Benzyloxy-4-(2-propenyl)benzyl bromide (40). To a
stirred solution of 39 (8.7 g, 0.034 mol) in dry ether
(60 mL) in an ice bath was added dropwise PBr3 (5.6 g,
0.021 mol). The reaction mixture was stirred for 30 min
and quenched with water. The organic phase was washed
with water (5×) and dried over MgSO4. Evaporation of the
solvent left a crude product, which was crystallized from
light petroleum (bp 40–60ЊC) to give 40 as a white powder
(9.3 g, 86%, mp 61–62ЊC): 1H NMR (CDCl3) d 3.41 (2 H,
d, Jˆ6.0 Hz, CH2), 4.48 (2 H, s, CH2), 4.98–5.15 (2 H, m,
overlapping, CH2), 5.09 (2 H, s, CH2), 5.88–6.07 (1 H, m,
CH), 6.90–6.98 (m, 2 ArH), 7.12 (d, Jˆ7.8 Hz, ArH), 7.32–
7.49 (m, 5 ArH); 13C NMR (CDCl3) d 33.87, 34.18, 69.91,
112.23, 115.83, 121.38, 127.21, 127.88, 128.54, 129.63,
130.08, 136.52, 136.93, 137.00, 156.53; HRMS m/z calcd
for C17H17OBr (Mϩ) 316.0463, found 316.0472.
1
1015 cmϪ1; H NMR (CDCl3) d 1.83 (2 H, t, Jˆ7.0 Hz,
(^)-2-[3-Benzyloxy-4-(2-propenyl)benzyl]-3-[30-benzyl-
oxy-a,a-bis(phenylthio)benzyl]butyrolactone (41). In a
similar manner as for 23 using benzyl bromide 40 (3.92 g,
0.012 mol) in place of benzyl bromide 11 to give 41 as a
white amorphous solid (3.7 g, 41%): IR (film) 1770, 1610,
1579, 1253, 1021, 911 cmϪ1; 1H NMR (CDCl3) d 2.82 (1 H,
dd, Jˆ5.8, 13.8 Hz, H-7), 2.93 (1 H, m, H-80), 3.10 (1 H, dd,
Jˆ4.7, 13.6 Hz, H-7), 3.32 (1 H, m, H-8), 3.41 (2 H, d,
Jˆ6.3 Hz, CH2), 3.54 (1 H, m, H-90), 4.35 (1 H, dd,
Jˆ3.1, 10.1 Hz, H-90), 4.93 (2 H, s, CH2), 4.97 (2 H, s,
CH2), 5.02 (2 H, dd, overlapping, CH2), 5.90–6.07 (1 H,
m, CH), 6.45 (d, Jˆ7.7 Hz, ArH), 6.62 (s, ArH), 6.86–6.98
(m, 2 ArH), 7.14–7.40 (m, 23 ArH); 13C NMR (CDCl3) d
33.98, 36.77, 44.47, 47.30, 67.98, 69.70, 69.95, 72.86,
112.58, 114.90, 115.45, 116.29, 121.17, 121.82, 127.09,
127.44, 127.63, 127.72, 127.91, 128.01, 128.44, 128.58,
128.60, 128.66, 129.32, 129.51, 129.91, 130.61, 132.34,
132.94, 135.98, 136.17, 136.63, 136.88, 137.13, 139.17,
156.42, 158.71, 178.66.
CH2), 2.42–2.63 (4 H, m, H-70, H-80, H-8), 2.72 (2 H, t,
Jˆ7.0 Hz, CH2), 2.95 (2 H, m, H-7), 3.56 (2 H, t, Jˆ6.0 Hz,
CH2), 3.81 (1 H, dd, Jˆ7.4, 9.5 Hz, H-90), 4.04 (1 H, dd,
Jˆ6.2, 9.2 Hz, H-90), 5.01 (2 H, s, CH2), 5.03 (2 H, s, CH2),
6.58 (d, Jˆ7.0 Hz, 2 ArH), 6.71 (d, Jˆ7.1 Hz, 2 ArH), 6.83
(m, ArH), 7.06–7.43 (m, 12 ArH); 13C NMR (CDCl3) d
25.73, 32.86, 34.76, 38.38, 40.93, 46.33, 61.90, 69.92,
70.03, 71.12, 112.67, 112.80, 115.66, 121.25, 121.91,
127.36, 127.50, 128.02, 128.12, 128.69, 129.07, 129.82,
130.32, 136.82, 137.01, 139.63, 156.79, 159.07, 178.58;
HRMS m/z calcd for C35H36O5 (Mϩ) 536.2563, found
536.2569; EIMS m/z 536 (Mϩ, 14), 445 (4), 106 (80), 91
(100).
(^)-2-[3-Benzyloxy-4-(2-carboxyethyl)benzyl]-3-(30-benzyl-
oxybenzyl)butyrolactone (44). A stirred mixture of 43
(0.94 g, 1.8 mmol) and pyridinium dichromate (2.3 g,
6.1 mmol) in dry DMF (20 mL) was maintained under
argon at room temperature for two days. The reaction
mixture was poured into water (215 mL), extracted with
ether and dried over MgSO4. Evaporation of the solvent
gave 44 as an oil (0.74 g, 77%), which was used directly
(^)-2-[3-Benzyloxy-4-(2-propenyl)benzyl]-3-(30-benzyl-
oxybenzyl)butyrolactone (42). To a stirred solution of 41
(3.69 g, 5.0 mmol) in dry toluene (60 mL) maintained under
argon at 90ЊC was added in small portions during 30 min, a
mixture of tributyltin hydride35 (6.0 g, 20.1 mmol) and
AIBN (0.22 g, 1.3 mmol). The reaction mixture was stirred
for 2 h, cooled and concentrated under reduced pressure.
The crude product was purified by flash column chroma-
tography (cyclohexane–toluene 1:1) and (EtOAc–cyclo-
in the next step: IR (film) 2950 (br), 1770, 1715, 1270 cmϪ1
;
1H NMR (CDCl3) d 2.32–2.72 (4 H, m, overlapping, H-70,
H-80, H-8), 2.66 (2 H, t, Jˆ7.7 Hz, CH2), 2.91–3.06 (4 H, m
and t, overlapping, H-7, CH2), 3.82 (1 H, dd, Jˆ7.1, 9.1 Hz,
H-90), 4.05 (1 H, dd, Jˆ6.6, 9.2 Hz, H-90), 5.00 (2 H, s,
CH2), 5.07 (2 H, s, CH2), 6.60–6.84 (m, 5 ArH), 7.06–
7.50 (m, 12 ArH); 13C NMR (CDCl3) d 25.60, 33.38,

¨ ¨
T. Makela et al. / Tetrahedron 56 (2000) 1873–1882
1882
¨
11. Axelson, M.; Sjovall, J.; Gustafsson, B.; Setchell, K. D. R.
34.75, 38.36, 40.90, 46.29, 69.70, 69.98, 71.13, 112.53,
112.74, 115.74, 121.29, 121.69, 127.17, 127.53, 127.88,
128.14, 128.63, 128.69, 129.82, 130.20, 136.80, 137.39,
139.63, 156.40, 159.01, 178.60; HRMS m/z calcd for
C35H34O6 (Mϩ) 550.2355, found 550.2349; EIMS m/z 550
(Mϩ, 6), 459 (8), 106 (75), 91 (100).
Nature 1982, 298, 659–660.
12. Borriello, S. P.; Setchell, K. D. R.; Axelson, M.; Lawson, A. M.
J. Appl. Bacteriol. 1985, 58, 37–43.
13. Bannwart, C.; Adlercreutz, H.; Fotsis, T.; Wahala, K.;
Hase, T.; Brunow, G. Finn. Chem. Lett. 1984, 120–125.
14. Adlercreutz, H.; Fotsis, T.; Lampe, J.; Wahala, K.; Makela, T.;
Brunow, G.; Hase, T. Scand. J. Clin. Lab. Invest. 1993, 53 (suppl.
215), 5–18.
15. Adlercreutz, H.; Fotsis, T.; Kurzer, M. S.; Wahala, K.; Makela,
T.; Hase, T. Anal. Biochem. 1995, 225, 101–108.
¨ ¨ ¨
¨ ¨ ¨
¨
¨
(^)-trans-2-[3-Hydroxy-4-(2-carboxyethyl)benzyl]-3-(30-
hydroxybenzyl)butyrolactone (8). A suspension of 44
(0.74 g, 1.3 mmol) in ethanol (60 mL) and W-2 Raney
nickel (7.5 g, 0.13 mol) was refluxed for 3 h. The catalyst
was filtered by rinsing with acetone, and the solvents were
evaporated. The residue was purified by extraction of the
impurities from basic aqueous solution into ether. Acidifi-
cation with 2N H2SO4, and extraction with ether afforded 8
as an amorphous solid (0.13 g, 26%): IR (film) 2990 (br),
1750, 1710 cmϪ1; 1H NMR (acetone-d6) d 2.50–2.72 (6 H,
m, H-70, H-8, H-80, CH2), 2.80–3.12 (4 H, m, H-7, CH2),
3.89 (1 H, dd, Jˆ7.1, 9.1 Hz, H-90), 4.04 (1 H, dd, Jˆ6.6,
9.2 Hz, H-90), 6.58–6.82 (m, 4 ArH), 7.00–7.20 (m, 2 ArH),
7.4 (m, ArH); 13C NMR (acetone-d6) d 26.33, 34.63, 34.98,
38.77, 42.11, 46.90, 71.61, 114.29, 116.50, 117.07, 120.66,
121.72, 130.20, 130.76, 130.83, 138.60, 141.39, 145.58,
158.49, 175.50, 178.28; HRMS m/z calcd for C21H20O5
(MϪH2O) 352.1311, found 352.1299; EIMS m/z 352
(MϪH2O, 57), 324 (4), 245 (23), 191 (11), 161 (32), 107
(26), 91 (100). Compound 8 was methylated for mass spec-
trum by diazomethane in ether solution at room temperature
to give the trimethylated product 45: EIMS m/z 412 (Mϩ).
¨ ¨ ¨
¨
¨
¨ ¨ ¨
16. Adlercreutz, H.; Fotsis, T.; Bannwart, C.; Wahala, K.;
Brunow, G.; Hase, T. Clin. Chim. Acta 1991, 199, 263–278.
17. Murphy, P. A. J. Chromatogr. 1981, 211, 166–169.
18. Barnes, S.; Kirk, M.; Coward, L. J. Agric. Food Chem. 1994,
42, 2466–2474.
19. Franke, A. A.; Custer, L. J.; Cerna, C. M.; Narala, K. Proc.
Soc. Exp. Biol. Med. 1995, 208, 18–26.
20. Wang, W. Q.; Tanaka, Y.; Han, Z. K.; Cheng, J. J. Agric. Food
Chem. 1994, 42, 1584–1587.
21. Lapcık, O.; Hampl, R.; Al-Maharik, N.; Salakka, A.;
Wahala, K.; Adlercreutz, H. Steroids 1997, 62, 315–320.
22. Lapcık, O.; Hampl, R.; Hill, M.; Wahala, K.; Al-Maharik, N.;
Adlercreutz, H. J. Steroid Biochem. Mol. Biol. 1998, 64, 261–268.
23. Adlercreutz, H.; Wang, G. J.; Lapcık, O.; Hampl, R.;
´
¨ ¨ ¨
´
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´
¨ ¨ ¨
¨
¨
Wahala, K.; Makela, T.; Lusa, K.; Talme, M.; Mikola, H. Anal.
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24. Uehara, M.; Lapcık, O.; Hampl, R.; Al-Maharik, N.; Makela,
T.; Wahala, K.; Mikola, H.; Adlercreutz, H. To be published.
´
¨
¨
¨ ¨ ¨
25. Pelter, A.; Ward, R. S.; Satyanarayana, P.; Collins, P. J. Chem.
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26. Adlercreutz, H.; Musey, P. I.; Fotsis, T.; Bannwart, C.;
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¨
¨
Wahala, K.; Makela, T.; Brunow, G.; Hase, T. Clin. Chim. Acta
1986, 158, 147–154.
27. Hanessian, S.; Lavallee, P. Can J. Chem. 1975, 53, 2975–
2977.
We thank Tarja Patama, MSc, for preliminary work in
synthesizing the compound 8.
28. Birkinshaw, J. H.; Bracken J. Chem. Soc. 1942, 368–370.
29. Meltzer, P. C.; Dalzell, H. C.; Razdan, R. K. J. Chem. Soc.,
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