Structure-activity studies on the anti-proliferation activity of ajoene analogues in WHCO1 oesophageal cancer cells

Article abstract of DOI:10.1016/j.ejmech.2012.01.058

The organosulfur compound ajoene derived from the rearrangement of allicin found in crushed garlic can inhibit the proliferation of tumour cells by inducing G₂/M cell cycle arrest and apoptosis. We report on the application of a concise four-step synthesis (Hunter et al., 2008 [1]) that allows access to ajoene analogues with the end allyl groups substituted. A library of twelve such derivatives tested for their anti-proliferation activity against WHCO1 oesophageal cancer cells has identified a derivative containing p-methoxybenzyl (PMB)-substituted end groups that is twelve times more active than Z-ajoene, with an IC₅₀ of 2.1 μM (Kaschula et al., 2011 [2]). Structure-activity studies involving modification of the sulfoxide and vinyl disulfide groups of this lead have revealed that the disulfide is the ajoene pharmacophore responsible for inhibiting WHCO1 cell growth, inducing G ₂/M cell cycle arrest and apoptosis by caspase-3 activation, and that the vinyl group serves to enhance the anti-proliferation activity a further eightfold. Reaction of the lead with cysteine in refluxing THF as a model reaction for ajoene's mechanism of action based on a thiol/disulfide exchange reveals that the allylic sulfur of the vinyl disulfide is the site of thiol attack in the exchange.

Full text of DOI:10.1016/j.ejmech.2012.01.058

European Journal of Medicinal Chemistry 50 (2012) 236e254
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Structureeactivity studies on the anti-proliferation activity of ajoene analogues in
WHCO1 oesophageal cancer cells
,
,
a
a
a
Catherine H. Kaschula b c, Roger Huntera , Nashia Stellenboom , Mino R. Caira , Susan Winks ,
*
Thozama Ogunleye a, Philip Richards a, Jonathan Cotton a, Kani Zilbeyaz a d, Yabing Wang b
,
,
,c
Vuyolwethu Siyo b c, Ellen Ngarande b c, M. Iqbal Parker b
,
,
,c
a Department of Chemistry, University of Cape Town, PD Hahn Building, Rondebosch, 7701 Cape Town, South Africa
b Department of Medical Biochemistry, University of Cape Town, Anzio Rd, Observatory, 7925 Cape Town, South Africa
c International Centre for Genetic Engineering and Biotechnology (ICGEB), Wernher and Beit South, Anzio Rd, Observatory, 7925 Cape Town, South Africa
d Department of Chemistry, Agri Ibrahim Cecen University, 4700 Agri, Turkey
a r t i c l e i n f o
a b s t r a c t
Article history:
The organosulfur compound ajoene derived from the rearrangement of allicin found in crushed garlic can
inhibit the proliferation of tumour cells by inducing G2/M cell cycle arrest and apoptosis. We report on
the application of a concise four-step synthesis (Hunter et al., 2008 [1]) that allows access to ajoene
analogues with the end allyl groups substituted. A library of twelve such derivatives tested for their anti-
proliferation activity against WHCO1 oesophageal cancer cells has identified a derivative containing
p-methoxybenzyl (PMB)-substituted end groups that is twelve times more active than Z-ajoene, with an
Received 28 July 2011
Received in revised form
20 January 2012
Accepted 30 January 2012
Available online 11 February 2012
IC50 of 2.1 mM (Kaschula et al., 2011 [2]). Structureeactivity studies involving modification of the sulf-
Keywords:
oxide and vinyl disulfide groups of this lead have revealed that the disulfide is the ajoene pharmacophore
responsible for inhibiting WHCO1 cell growth, inducing G2/M cell cycle arrest and apoptosis by caspase-3
activation, and that the vinyl group serves to enhance the anti-proliferation activity a further eightfold.
Reaction of the lead with cysteine in refluxing THF as a model reaction for ajoene’s mechanism of action
based on a thiol/disulfide exchange reveals that the allylic sulfur of the vinyl disulfide is the site of thiol
attack in the exchange.
Garlic
Ajoene
End-substituted ajoene derivatives
Structureeactivity
Vinyl disulfide
Anti-cancer
WHCO1 oesophageal cancer cell-line
Ó 2012 Elsevier Masson SAS. All rights reserved.
1. Introduction
(i.e. when crushed garlic is heated in oil) [4,5]. The E-isomer is more
predominant in aged preparations due to its increased chemical
Ajoene was first isolated as an E/Z-mixture by Block and Apitz-
Castro in 1984 [3] as a rearrangement product of the thiosulfinate
compound allicin, which is an organosulfur compound produced
from freshly-crushed garlic. Its structure was established to be an
allyl sulfoxide containing an unusual vinyl disulfide group, the
latter rarely seen in natural product structures. The authors [3,4]
suggested that the key step in ajoene’s biosynthetic pathway
involves the addition of allylsulfenic acid to an S-thioallylated
vinylthionium ion formed from the self-condensation of allicin
(Scheme 1).
stability over its Z-isomer. In Block and Apitz-Castro’s work [4], the
focus of ajoene’s biological activity centred on its anti-thrombotic
activity [6,7], but in subsequent years ajoene has demonstrated
a range of other biological activities including anti-microbial [8],
anti-obesity [9,10], anti-fungal [11e14], and anti-cancer [15e17]
activities, with the latter of particular interest in the current
work. Ajoene has been shown to inhibit tumour cell proliferation
by inducing apoptosis in a number of cell lines, with the IC50
ranging from 5 to 40 mM [1,18e20]. In vivo, ajoene has been shown
to inhibit the growth of skin carcinoma in both rodents and humans
[21,22], and to inhibit tumour cell growth in mouse xenografts by
2e3-fold [18,19].
Several groups have uncovered aspects of mode of action and
molecular targets of ajoene, but a comprehensive picture has yet to
emerge. Ajoene and the related garlic organosulfur compounds
diallyl disulfide and diallyl trisulfide have been shown to induce
cell-cycle arrest in the G2/M phase of the cell cycle [23,24].
Apoptosis has also been demonstrated to proceed via the
Ajoene is not found in freshly crushed garlic but is found in
treated or aged preparations, with increased concentrations in
heated preparations [3,5]. The E-isomer is more abundant in fresh
extracts [5] with the Z-isomer predominating in heated extracts
* Corresponding author. Tel.: þ27 21 650 2544; fax: þ27 21 650 5195.
E-mail address: roger.hunter@uct.ac.za (R. Hunter).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.01.058

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
237
O
S
MeO
-H+
O
S
S
S
HOS
S
S
S
S
(E/Z)-Ajoene
Scheme 1. The key step in ajoene biosynthesis.
OMe
Fig. 1. The chemical structure of the ajoene analogue E/Z-4l.
mitochondrial-dependent caspase cascade with involvement of
mitochondrial membrane permeabilization (MMP), release of
cytochrome c, caspase 3 activation and cleavage of the anti-
apoptotic protein Bcl-2 [19,24e30]. It has been proposed [2,31,32]
that the anti-proliferation activity of garlic organosulfur
compounds may be tied to the chemical reactivity of the disulfide
bond, which may thiolate cysteine residues in proteins. In support
of this hypothesis, the related garlic compound diallyl trisulfide has
obligatory allyl group at the disulfide end of the molecule. Thus, it
was clear that a new synthesis needed to be developed for
accessing derivatives of type 4 shown in Scheme 2 with variable
end-groups. Consideration of the development of such a synthesis
suggested the possibility of using cross-coupling methodology [36]
for accessing a stereo-defined protected substituted vinyl sulfide;
this had the attraction of accessing individual E/Z-stereoisomers of
the targets. However, ultimately it was decided to base the key step
of our synthesis [1] on a reaction studied in the 1960s by Kamp-
meier [37] involving regioselective radical addition of thiolacetic
acid to 1-hexyne at its terminus to form a vinyl thioacetate.
Kampmeier showed that the Z-product was kinetically favoured,
and application of this finding to our system resulted in the
development [1,38] of a moderately Z-stereoselective four-step
synthesis of the ajoene core structure in which the end groups
could be varied, and one that was amenable to production of
derivatives for biological evaluation. A general overview of it is
depicted in Scheme 1.
been found to oxidatively modify
ment to form mixed disulfides at cysteine residues Cys-12
Cys-354 [33]. In another cell-free model, ajoene was found to act
b-tubulin in a cell-free environ-
b
and
b
as a covalent inhibitor and substrate of glutathione reductase
through mixed disulfide formation with Cys58 in the active site of
the enzyme [34].
In 2008 we published a letter [1] on the first synthetic route to
access end-substituted ajoenes (see Scheme 2 for a general
formula), which demonstrated that terminal end substitution
(sulfoxide end, R1) of ajoene results in retention, and in some cases
a modest improvement, in the anti-proliferation activity against
transformed CT-1 fibroblast cells compared to the parent natural
product. We now provide a full account of activity against the
WHCO1 cell-line involving seven of the previously published
analogues together with five new analogues in which both R1 and
R2 are substituted. This study has resulted in identification of our
most active analogue E/Z-4l to date substituted with a p-methox-
ybenzyl (PMB)-group at both terminal positions (Fig. 1). This
analogue is more active than Z-ajoene at inhibiting cancer-cell
proliferation of cultured prostrate, breast, cervical and oesopha-
geal cancer cells, and more specifically displays a twelve-fold
improvement against the WHCO1 oesophageal cancer cell-line
[2]. We also present data on five new analogues of E/Z-4l in
which the central vinyl disulfide/sulfoxide core is varied. The
findings in this paper have enabled the generation of a structur-
eeactivity model for the anti-proliferation activity of ajoene in
WHCO1 cancer cells.
Thus, a thiol R1SH obtained commercially or via reaction of R1Br
with thiourea was propargylated under standard conditions to
form propargylic thioether 1. This underwent regioselective radical
addition with thiolacetic acid in refluxing toluene using ACCN as
radical initiator to give an E/Z-mixture of stereoisomeric vinyl thi-
oacetates 2 in around 60% after chromatography (see Table 1). Low
temperature hydroxide-promoted cleavage of the thioester fol-
lowed by sulfenylation with an appropriate S-alkylated thio-p-tol-
uenesulfonate afforded vinyl disulfide
3 in high yield after
chromatography. Finally, chemoselective sulfide oxidation with m-
CPBA (1.1 equiv) in DCM at low temperature gave substituted
ajoene 4 in yields of around 60e70% after chromatography. In some
cases the polarities of the isomeric E- and Z-sulfoxide products 4
was sufficiently different such that they could be separated chro-
matographically. The overall yield for the sequence was generally
not lower than about 30%, allowing production of sufficient mate-
rial of a range of derivatives for biological evaluation purposes.
Unfortunately, the ajoene parent (R1, R2 ¼ allyl) couldn’t be
2. Results and discussion
accessed by this route, possibly as a result of cyclization of the b-
vinyl radical onto the S-allyl group (R1 ¼ allyl in 2 of Scheme 2) via
2.1. Synthesis aspects
a 5-exo- or 6-endo-trig process [1].
We have previously synthesized seven analogues of 4 in which
only R2 was varied (Table 1, compounds 4aeg) [1]. We now report
the synthesis of a further 5 analogues which probe both the R1 and
R2 terminal positions (Table 1, compounds 4hel). All compounds
have been fully characterised by 1H and 13C NMR spectroscopy (see
Experimental section), and the E/Z-isomer ratios in compounds 3
and 4 were established by virtue of Jcis/Jtrans vinyl couplings in the
2.1.1. Part 1: synthesis of terminally-substituted ajoene analogues
The only known synthesis of ajoene to date is the biomimetic
rearrangement of allicin according to Block [3,4,35], by heating it in
aqueous acetone. Apart from our experience of obtaining low yields
of the natural product using this route, the biomimetic synthesis
demands that an S-allyl group (or substituted allyl) is present in the
substituted thiosulfinate starting material, which results in an
Propargyl Br
CH3COSH, Tol, Δ
R1S
R1S
R1S
R1
SAc
S
R1SH
KOH, MeOH
ACCN (5 mol%)
1
2
O
S
m-CPBA,
R2
DCM, -78°C
(i) KOH, MeOH, -40°C
S
R2
S
S
(ii) p-TolSO2SR2
3
4
Scheme 2. Synthesis of doubly-end substituted ajoenes of type 4.

Table 1
Yields and E/Z ratios for the synthesis of compounds 1aeg, 2aeg, 3ael and 4ael.
O
S
R1S
SAc
R1S
S
R2
R1
R2
R2
S
S
R1
S
3
2
4
Cmpd no
Yield % (E/Z)
Cmpd no
Yield % (E/Z)
Cmpd no
Yield % (E/Z)
Cmpd no
Yield % (E/Z)
3a
97 (4:5)
4a
82 (2:3)
1a
80
2a
2b
73 (1:2)
73 (1:2)
3h
3i
76 (1:1)
76 (2:3)
4h
4i
68 (3:2)
68 (2:3)
CH3
1b
57
3b
89 (1:2)
4b
78 (1:2)
1c
70
57
2c
31 (1:1)
64 (2:3)
3c
89 (1:2)
89 (2:3)
4ca
4d
86b (2:3)
92 (1:2)
1d
2d
3d
1e
1f
69
98
2e
2f
66 (2:3)
53 (1:2)
3e
67 (1:1)
4e
64 (1:2)
3f
86 (3:4)
84 (2:3)
4f
65 (5:6)
85 (1:2)
3g
4g
3j
88 (2:3)
70 (3:4)
4j
71 (2:3)
74 (3:4)
1g
66
2g
55 (2:3)
3k
4k
3l
83 (1:1)
4l
78 (3:2)
a
Isolated as the free alcohol via silyl deprotection with HF in acetonitrile.
Yield refers to the m-CPBA oxidation step on the silyl ether.
b

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
239
1H NMR spectra. Similarly, IR spectroscopy revealed a thioacetate
carbonyl stretching absorption for the vinyl thioesters 2 at around
1700 cmꢀ1. All vinyl thioacetates 2aeg were isolated by column
chromatography as a mixture of stereoisomers and as oils that
couldn’t be separated, but which all gave correct high resolution
mass spectrometry data. E/Z-Ratios revealed a preference on
average of about 2:1 in favour of the Z-isomer, which was pleasing
in view of the report that Z-ajoene is more active than its E-isomer
as an anti-thrombotic agent [4], and that some studies in cancer
have also focused primarily on the Z-isomer [19,30]. Such stereo-
selectivity indicates that the rate of reduction of the intermediate Z-
vinylthioacetate radical by thiolacetic acid is higher than that of the
E-radical in the hydrogen-quenching propagation step, presumably
owing to less steric hindrance between the attached thioacetate
group and the incoming thiolacetic acid (Table 1 summarises the
results).
2g (Table 1) in 80 % yield and as a 3:2 Z:E mixture of stereoisomers,
using a similar sequence to the one used to obtain E/Z-4l except
with alkylation of the enethiolate with PMB chloride to generate 5.
Subsequent selective oxidation of the more nucleophilic sulfide
sulfur of 5 furnished derivative 6 in 82 % yield (Z:E ¼ 3:2) containing
a vinyl sulfide/sulfoxide combination rather than the vinyl disul-
fide/sulfoxide found in E/Z-4l (Scheme 3).
Both compounds 5 and 6 are new compounds that were
comprehensively characterised as an E/Z-isomeric mixture in the
case of 5 and as crystalline, individual stereoisomers for 6 that gave
correct microanalyses. The site-selectivity of oxidation was estab-
lished as before by observation of downfield shifts in the NMR
spectra for the sulfoxide a-hydrogens and carbons originating from
the sulfide sulfur in 5. For compounds 7 and 8 (Scheme 4) con-
taining a saturated disulfide instead of a vinyl disulfide grouping,
hydrogenation of E/Z-3l or E/Z-4l failed to afford the desired
product. Thus, a novel route for synthesis of 7 was developed,
which involved two successive alkylations of 1,3-propane-dithiol in
a one-pot sequence using: i) PMBCl; ii) S-PMB p-toluenesulfo-
nylthioate, to afford 7 in high overall yield (85%). Subsequent
oxidation of the more nucleophilic sulfur of 7 gave sulfoxide/
disulfide 8 after column chromatography lacking the double bond
of the ajoene derivative E/Z-4l. As before, NMR provided compre-
hensive support for the site of oxidation as stated (see
Experimental section), Scheme 4.
With a range of vinyl thioacetates in hand, we set about incor-
porating the vinyl disulfide moiety. Such a functionality was first
synthesized by Brandsma [39] via sulfenylation of
a 1-
alkenethiolate prepared in situ by cleavage of a 1-alkenyl alkyl
sulfide with Li/NH3, and this methodology was subsequently
exploited by Block [40] in his synthesis of cepaene natural products
from onion. In our case, it was found that low temperature thio-
acetate cleavage with KOH to the enethiolate was successful at
around ꢀ40 ꢁC. Reaction with the appropriately substituted S-allyl
p-toluenesulfonylthioate was carried out in each case (2aeg)
resulting in a high-yielding conversion to the vinyl disulfide
products 3ael. Interestingly, and importantly from the bioactivity
discussion that follows, in most cases the starting E/Z-ratio in 2aeg
was retained or only marginally increased in favour of the E-isomer
in 3aeg indicating that enethiolateethioaldehyde tautomerism
with stereoinversion is slow at ꢀ40 ꢁC. Other sulfenylating agents
could be easily accessed, so derivatives 3hel were also prepared in
which R2 was varied, (see Table 1). Once again cis/trans-isomeri-
zation was minimal. All vinyl disulfides 3ael were characterised as
their E/Z-mixtures, and with the aid of HSQC and COSY NMR
spectroscopy, isomers could be fully assigned (see Experimental
section). Completion of the synthesis was achieved by chemo-
selective oxidation of the sulfide sulfur of each derivative 3ael
using m-CPBA at low temperature in dichloromethane. Following
chromatography, target ajoenes 4ael were obtained in an accept-
able yield of around the 70% mark, and in some cases the E/Z-
isomers could be separated by column chromatography (4a, 4c, 4d,
4e, 4h, 4i) or crystallized (4jel) as a mixture of isomers (by 1H
NMR). The E- and Z-isomers of 4c containing a terminal primary
hydroxyl group could be obtained via desilylation (HF/CH3CN) of
the TBS derivative. All compounds were fully characterised by
a range of spectroscopic and analytical methods. The oxidation
resulted in slight fluctuations either way of the E/Z-stereochemistry
ratios going from 3 to 4, which was ascribed to the experimental
conditions and the isolation and purification steps. Evidence for the
2.2. Biological activity
2.2.1. Effect of R1 and R2 substitution
The ajoene analogues 4ael were all tested for their ability to
inhibit the proliferation of WHCO1 oesophageal cancer cells by the
MTT cell proliferation assay. Strength of inhibition is reported as an
IC50, defined as the concentration found to inhibit 50 % of cell
proliferation after 48 h. Some of the isomeric mixtures of 4 were
separable chromatographically and were hence tested as pure
geometric isomers (4a, 4c, 4d, 4e, 4h, 4i), whereas the others (4b,
4f, 4g, 4j, 4k, 4l) were inseparable and were tested as E/Z mixtures
(see Table 2 for anti-proliferation activities reported as an average
and standard deviation of three independent determinations).
For ajoene analogues in which at least one of the allyl groups of
ajoene was varied as propyl and/or methyl 4a, 4h, 4i, and for which
E- and Z-isomers could be separated, retention of activity was
observed against WHCO1 cell proliferation regardless of whether
one or both allyl groups was substituted, complementing our
previous findings on transformed CT-1 cells [1], and implying that
the ajoene pharmacophore does not reside within the terminal end
allyl groups. This finding contrasts with that of Sundaram et al.
[41,42] in which the related garlic compound diallyl disulfide
(DADS) was found to be ineffective at inhibiting growth of human
colon, skin and lung cancer cells when the terminal allyl groups
were substituted for propyl. It is unlikely that this discrepancy is
cell-line specific as 4a was also previously found to be fully active
against CT-1 cells [1]. It would therefore appear that different
mechanisms govern the anti-proliferation activity of ajoene and
DADS in cancer cells. Furthermore, and in agreement with previous
findings on different cancer cell lines [1,2], Z-ajoene and the Z-
analogues were found to be marginally more active at inhibiting
site of oxidation came from the downfield shifts in the sulfoxide
methylene hydrogens and
-carbons in the 1H NMR and 13C NMR
spectra respectively. Furthermore, the -sulfoxide hydrogens of 4
a-
a
a
allylic to the vinyl disulfide double bond always resonated as dia-
stereotopic ABXX0 double double doublets, and these could be
identified for each isomer.
2.1.2. Part 2: synthesis of E/Z-4l analogues
Four structural analogues of E/Z-4l were synthesized (see
Schemes 3 and 4 for structures) including a vinyl sulfide (removal of
one S of the SeS functionality) as well as a saturated disulfide
(removal of the double bond), while keeping everything else
constant. To this end, derivative 6, with the vinyl disulfide of E/Z-4l
replaced by a vinyl sulfide, could be prepared from vinyl thioacetate
i) KOH / MeOH, -40°C
MeO
OMe
O
S
5
ii) PMBCl
2g
S
iii) m-CPBA, DCM, -78°C
6
Scheme 3. Synthesis of disulfides 5 and 6.

240
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
MeO
i) PMBCl, KOH, EtOH
ii) p-TolSO2SPMB,
O
S
S
7
HS
SH
S
8
iii) m-CPBA, DCM, -78°C
OMe
Scheme 4. Synthesis of disulfides 7 and 8.
cell proliferation than E-ajoene and the corresponding E-analogues.
This data supports the hypothesis that stereospecific proteinedrug
In our previous study [1], we identified that p-methoxybenzyl
substitution at R1 (R2 ¼ allyl) (4g) produced the most active
analogue at inhibiting the proliferation of transformed CT1 cells
and with increased lipophilicity appearing to play a role. In the
interactions appear to play
a minor role in ajoene’s anti-
proliferation activity in cancer cells.
Table 2
Effect of R1 and R2 substitution on WHCO1 cell proliferation.
O
S
R2
Cmpd no.
Ajoene
Isomer
IC50 ꢂ SD (
mM)
S
R1
S
4
R1
R2
Z
E
25.0 ꢂ 2.8 [2]
39.0 ꢂ 7.8 [2]
Z
E
23.0 ꢂ 4.2
37.0 ꢂ 4.5
4a
4b
EZ
23.0 ꢂ 6.7
Z
E
38.0 ꢂ 5.9
27.0 ꢂ 5.6
4c
4d
4e
Z
E
21.0 ꢂ 0.6
18.0 ꢂ 5.7
Z
E
33.0 ꢂ 1.4
68.0 ꢂ 15
4f
EZ
EZ
8.9 ꢂ 1.2
7.4 ꢂ 0.7
4g
4h
Z
E
18.0 ꢂ 4.1
24.0 ꢂ 2.8
Z
E
26.0 ꢂ 7.5
28.0 ꢂ 7.2
4i
4j
CH3
EZ
EZ
3.1 ꢂ 1.1
16 ꢂ 1.1
4k
4l
EZ
2.1 ꢂ 0.4 [2]

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
241
Table 3
2.3. Structureeactivity analysis of the pharmacophore
Anti-proliferation activity of E/Z-4l, cisplatin and 5-fluorouracil
on WHCO1 cells.
2.3.1. Importance of the disulfide in E/Z-4l anti-proliferation
activity
Compound name
IC50 ꢂ SD (
mM)
Having established that the R1 and R2 end groups are not critical
for anti-proliferation activity in WHCO1 cells but appear to play
a modulation role, the focus of the structureeactivity analysis was
next directed at the vinyl disulfide and sulfoxide functionalities of
the most active analogue E/Z-4l. Analogues 5e8 were then tested
for WHCO1 anti-proliferation activity by (i) the MTT assay (Table 4),
and (ii) xCelligence (Fig. 3).
E/Z-4l
Cisplatin
5-Fluorouracil
2.1 ꢂ 0.4 [2]
9.2 ꢂ 0.1
7.9 ꢂ 0.6
current study, a similar trend was also observed in WHCO1 cells
(see Table 2 for the IC50s). We therefore selected p-methoxybenzyl
as our active lead for the R1 position, and turned our attention
towards optimizing R2. To this end, a small library of analogues
(4je4l) was prepared in which R2 was varied as benzyl (4j), p-flu-
orobenzyl (4k) and p-methoxybenzyl (E/Z-4l). The geometrical
isomers of these analogues were all inseparable and were hence
tested as E/Z mixtures, which revealed a trend of OMe > H [ F
(Table 2) suggesting that electron-rich end groups may favour
increased activity. In such a way, we identified E/Z-4l as our most
active analogue, which as an E/Z-mixture returned a twelvefold
Analogues lacking the disulfide bond (E/Z-5, Z-6 and E-6) were
found to be inactive both by the MTT assay and xCelligence (Fig. 3),
strongly implying that the pharmacophore resides within the
disulfide. This agrees with reports on the two garlic sulfides diallyl
sulfide (DAS) and S-allyl cysteine, which are also both inactive at
inhibiting the growth of colon, lung and skin cancer cells [42e44],
although DAS is reported to be active at inducing apoptosis in
human colon 320 DM cancer cells [45]. Removal of only the vinyl
group (analogue 8) still rendered the analogue active but caused an
eightfold reduction in activity as measured by the MTT assay, which
was confirmed by xCelligence, supporting the hypothesis that the
vinyl group serves to activate the disulfide pharmacophore.
Analogue E/Z-3l containing an intact vinyl disulfide pharmaco-
phore but lacking the sulfoxide was in fact found to be threefold
more active than E/Z-4l, returning a nanomolar IC50 (700 nM) by
the MTT assay but with a reduced activity when measured by
xCelligence. Interestingly, analogue 7, which contains the disulfide
pharmacophore but lacks both the vinyl and sulfoxide groups, was
found to be inactive by both methods implying some synergy
between these two groups.
increase in activity (2.1
isomer, Z-ajoene (25 M) [2]. This IC50 falls within the in vitro
clinical range where two other oesophageal cancer chemothera-
peutics cisplatin and 5-fluorouracil were found to have an IC50 of
mM) relative to the more active parent
m
9.2
m
M and 7.9
mM respectively on the same cell line (see Table 3).
To further confirm the MTT data, WHCO1 cells were treated
with E/Z-4l, Z-ajoene and E-ajoene, and cell growth, adhesion and
spreading were monitored as a function of time by Roche xCel-
ligence (Fig. 2). When compared to the control cells (pink), 10
of Z-ajoene (green) or 10 M of E-ajoene (blue) was found to
partially inhibit WHCO1 cell-growth, followed by recovery within
two days. Conversely, treatment with 10 M E/Z-4l (red) caused
immediate inhibition of cell growth with very little recovery up to
5 days. (For interpretation of the references to colour in this
paragraph, the reader is referred to the web version of this
article.)
mM
m
2.3.2. The importance of the disulfide in E/Z-4l cell-cycle arrest and
apoptosis
Ajoene and the garlic related organosulfides are reported to
inhibit cancer cell growth by inducing G2/M cell cycle arrest and
apoptosis through caspase-3 activation [19,24,46]. We were
therefore interested in assessing whether E/Z-4l and analogues of
m
Fig. 2. WHCO1 cell growth monitored by Roche xCelligence as a function of time. 24 h after plating, WHCO1 cells (2500 cells per well) were treated with Z-ajoene (10
ajoene (10 M, blue) or E/Z-4l (10 M, red) in 0.1% DMSO and control cells received 0.1% DMSO alone (pink). Cell growth was then monitored continuously for a further 5 days. (For
interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
mM, green), E-
m
m

242
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
Table 4
Structureeactivity pharmacophore analysis of E/Z-4l regarding inhibition of WHCO1 cell proliferation.
Cmpd no.
Chemical structure
WHCO1
IC50 ꢂ SD (
mM)
MeO
S
S
E/Z-3l
0.7 ꢂ 0.3
S
OMe
OMe
MeO
O
S
S
E/Z-4l
2.1 ꢂ 0.4 [2]
S
MeO
MeO
OMe
E/Z-5
Z-6
E-6
7
>200.0
>200.0
>200.0
>200.0
S
S
O
S
S
OMe
OMe
MeO
MeO
O
S
S
S
S
S
OMe
MeO
O
S
S
8
16 ꢂ 3.7
S
OMe
E/Z-4l that lack the disulfide are also active at inducing cell-cycle
arrest or apoptosis. Thus WHCO1 cells were treated either with
10 mM of E/Z-4l, E/Z-3l or the sulfides E-6 and Z-6 for 16 h and
suggesting, similar to ajoene, that E/Z-4l inhibits WHCO1 cell
proliferation by inducing apoptosis through the activation of
caspase-3 (Fig. 6). Both sulfide isomers (E-6 and Z-6) lacking the
disulfide functionality were each found to be inactive at inducing
cell-cycle arrest and apoptosis in WHCO1 cells at the same molar
concentration (Figs. 4 and 5 respectively), although significantly
more cells were found in the S-phase relative tothe untreated DMSO
control. Taken together, these findings support the hypothesis that
the ajoene pharmacophore responsible for anti-proliferation
activity, G2/M cell cycle arrest and induction of apoptosis in
WHCO1 oesophageal cancer cells is the disulfide functional group.
analysed by flow cytometry (Fig. 4). The compounds were also
assayed for their ability to induce apoptosis in WHCO1 cells, which
was quantified using the Roche Cell Death Elisa Assay Kit (Fig. 5).
This assay is a photometric enzyme-immunoassay which quanti-
tates histone-associated DNA fragments (mono- and oligonucleo-
somes) in the cytoplasm after treatment.
In agreement with the MTT and xCelligence cell proliferation
data, the two active compounds that both contain a disulfide (E/Z-4l
and E/Z-3l)were found toinduce apoptosis and G2/M cell cyclearrest
in WHCO1 cells after 16 h treatment. This data supports the
hypothesis that the disulfide but not the sulfoxide is critical for
activity. As a representative disulfide for further analysis, active
analogue E/Z-4l was assayed for its ability to activate caspase 3
enzyme activity in treated WHCO1 cells using the Ac-DEVD-AFC
2.3.3. Reaction of E/Z-4l with N-Boc-L-cysteine ethyl ester
The vinyl disulfide’s enhanced activity compared to that of the
saturated disulfide suggests some type of chemical activation of this
grouping, in which reaction with a thiol group of a host protein
residue seems to be a likely possibility. Indeed, the study by Krauth-
Siegel [34] and co-workers dating back just over ten years revealed
via an X-ray structure that ajoene reacts with glutathione reductase
(GR) to afford allylSOCH2CH]CHSS(Cys58)-GR formed between the
fluorogenic substrate for caspase 3. Treatment with E/Z-4l (25 mM)
for 24 h was found to induce a threefold increase in caspase 3
enzyme activity relative to untreated or DMSO treated cells

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
243
Fig. 3. WHCO1 cell growth following treatment with analogues of E/Z-4l, monitored by xCelligence. A: 24 h after plating, WHCO1 cells (5000 cells per well) were treated with 0.1%
DMSO (control, dark-blue), Z-6 (34 M, light-green), E-6 (34 M, pink), 8 (34 M, light-blue), E/Z-3l (34 M, dark-green), and E/Z-4l (34 M, red). Cell growth was then measured
continuously for a further six days; B: WHCO1 cells (5000 cells per well) treated with 0.1% DMSO (control, dark-blue), 7 (113 M, dark-green), 5 (113 M light-blue), and E/Z-4l
(100 M, red). Each curve represents an average of three replicates. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of
m
m
m
m
m
m
m
m
this article.)
Cys58 of GR and the vinyl sulfurofajoenewith the expulsion of the S-
allyl group. No rationale of this unexpected regioselectivity was
presented in the paper. We thus decided to investigate this reaction
in solution, as it was considered more likely that the allyl sulfur of
ajoene would be the more electrophilic sulfur of the two in view of
the greater leaving ability of the vinylthio moiety in the disulfide
the allyl sulfur, and that the better leaving group in the thiol/disul-
fide exchange is the vinylthio moiety. This would imply that the
product observed by Krauth-Siegel in vitro with the enzyme GR [34]
with reverse regioselectivity was not the primary product of a direct
thiolysis. Krauth-Siegel also demonstrated that GR reduces ajoene
with the formation of single-electron reduced products and super-
oxide anion radical. Together with the thiolysis pathway, such
single-electron processes of the disulfide grouping with production
of ROS leading to apoptosis via the mitochondrial signalling cascade
provide the likelymode of action of ajoene invitroinwhich reactivity
is enhanced by the presence of the vinyl disulfide double bond.
exchange. To this end, N-Boc-L-cysteine ethyl ester as a limiting
reagent was refluxed with the ajoene derivative E/Z-4l (2 equiv) for
24 h inTHF. Onlya single product spot was observed onTLC, and this
was taken to be the primary product of the reaction. Column chro-
matographic purification gave a 52% yield (based on the cysteine
derivative as limiting reagent and at 78% conversion) of para-
methoxybenzyl N-Boc cysteine ethyl ester disulfide (9), indicating
that the preferred site of attack in solution is indeed at the non-vinyl
sulfur (Scheme 5). This finding is in agreement with those of Krauth-
Siegel’s laboratory results [34] on the same process, although Law-
son [47] has reported that in addition to forming S-thioallylcysteine,
reaction of excess cysteine (2 equiv) with ajoene also forms ajocys-
teine containing a disulfide bond with the vinyl sulfur of ajoene,
possibly as a secondary product. Overall, such results lead to the
conclusion that the more electrophilic sulfur of the vinyl disulfide is
2.3.4. Structureeactivity hypothesis for ajoene
Based on the findings in this paper, a structureeactivity hypoth-
esis for ajoene is proposed (see Fig. 7) in which the disulfide is the
pharmacophore responsible for anti-proliferation activity, G2/M cell-
cycle arrest and induction of apoptosis, with the vinyl group playing
a further activation role. The allyl sulfur is hypothesized to be the
more electrophilic sulfur capable of reacting with a free cysteine
residue in a protein to form a mixed disulfide. A Z-configuration
about the vinyl disulfide, electron rich side groups and a lipophilic

244
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
Fig. 4. Flow cytometry of WHCO1 cells treated with analogues of E/Z-4l. WHCO1 cells were treated either with 10
mM E/Z-4l, E/Z-3l, E-6 or Z-6 for 16 h. Cell cycle analysis was then
performed using propidium iodide staining as described in the Experimental Section. Each sample was done in triplicate.

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
245
MeO
O
4l, THF,
Δ
2.5
2.0
1.5
1.0
0.5
0.0
S
N-Boc Cys Et ester
***
S
9
OEt
NHBoc
**
Scheme 5. Reaction of N-Boc cysteine Et ester with E/Z-4l.
cysteine in solution reveals a regioselectivity in the thiol/disulfide
exchange favouring nucleophilic (thiol) attack on the allylic rather
than the vinylic sulfur. However, the extent to which this might be
relevant to events in vitro still remains to be elucidated as does the
reason for the in vitro bioactivity dependency on the ajoene E/Z-
geometry. Future studies will attempt to separate the E/Z-isomeric
mixture of 4l and evaluate the activity of individual isomers. The
use of fluorescently labelled ajoene derivatives to identify molec-
ular targets within the cancer cell will also be explored, as well as
the activity of E/Z-4l in a mouse model.
Fig. 5. Quantification of apoptosis in WHCO1 cells treated with analogues of E/Z-4l.
WHCO1 cells were treated either with 10 M E/Z-4l, E/Z-3l, E-6 or Z-6 for 16 h. Cell
lysate was then collected and apoptosis was quantified by assaying for cytoplasmic
histone-associated DNA fragments using the Roche Cell Death Elisa assay as described
in Materials and Methods. Bars, SD for each sample analyzed in quadruplicate;
**P < 0.01, ***P < 0.005 relative to DMSO control.
m
4. Experimental section
4.1. Cell lines and cell proliferation assays
The oesophageal cancer cell-line WHCO1 was derived from
a biopsy of primary oesophageal squamous cell carcinoma of South
African origin [48]. Cells were all incubated at 37 ꢁC under 5% CO2
and cultured without antibiotics in DMEM (Dulbecco’s Modified
Eagle Medium, Sigma) containing 10% FBS (Fetal Bovine Serum,
Gibco). IC50 determinations were carried out using the Roche MTT
[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]
assay. Briefly, WHCO1 cells were seeded (2 ꢃ103 cells/well) in 96-
surface is generally favourable for increased activity implying a level
of stereospecificity between the target protein and ajoene.
3. Conclusions
A concise, non-stereoselective four-step synthesis has been
developed for accessing derivatives of the natural product ajoene in
which the terminal end-groups (R1 and R2) can be varied whilst
retaining the vinyl disulfide and sulfoxide functional groups. A
structureeactivity study involving a small library of such modified
derivatives against WHCO1 cancer-cell proliferation has revealed
that end-group substitution retains and may enhance bioactivity. In
general, Z-ajoene and the Z-analogues were found to be slightly
more active than E-ajoene and the corresponding E-analogues.
Benzylic end-groups increased activity in which an electron-rich
PMB group at each end (derivative E/Z-4l) returned the highest
activity of twelve-fold compared to the parent ajoene. Varying the
vinyl disulfide and sulfoxide groupings indicates the disulfide to be
the dominant pharmacophore at inhibiting WHCO1 cell prolifera-
tion, inducing G2/M cell cycle arrest and apoptosis by caspase 3
activation, with the vinyl group appearing to play an activation
role; by comparison, the sulfoxide group appears to play a minor
role in the bioactivity in the ajoene series. Reaction of E/Z-4l with
well plates in a total volume of 90
solutions of each compound in media were added to the cells to
give a final concentration of 0.5 200 M compound and 0.1% DMSO.
Control cells received 0.1% DMSO alone. After 48 h incubation,10
of 5 mg/mL MTT was added to each well. After a further 4 h incu-
bation, 100 L of 10% SLS (sodium lauryl sulfate) in 0.01 M HCl was
mL. The following day, DMSO
m
mL
m
added to each well, and plates were incubated overnight. Plates
were read at 595 nm on a Thermo Scientific Multiscan FC plater-
eader and data was analysed by Graphpad prism 4 software using
sigmoidal doseeresponse (variable slope) curve fitting. For the
Roche xCelligence system, WHCO1 cells (2- or 5 ꢃ103 cells per
well) were plated in 96 well E-plates and growth was monitored as
an impedance value continuously for seven days on the RTCA SP
xCelligence system. 24 h after seeding, the compounds were added
according to the method described for the MTT assay.
4.2. Flow cytometry
3.5
WHCO1 cells were seeded for triplicate experiments
(1 ꢃ106 cells, 8 mL) in 100 mm cell culture dishes. 6 h after plating,
DMSO solutions of each compound in media were added to the cells
3.0
**
2.5
2.0
1.5
1.0
0.5
0.0
to give a final concentration of 10 mM compound and 0.1% DMSO.
Control cells received 0.1% DMSO alone. After 16 h treatment, the
cells were lifted by trypsinisation and resuspended in 2 mL cold
PBS, and fixed by adding 8 mL cold 70% ethanol. Cells were pelleted
at 4000 rpm for 5 min, washed twice with 1 mL cold PBS, resus-
pended in 100
bated for 15 min at 37 ꢁC. 20 min prior to analysis, propidium
iodide (10 g/mL) was added and incubated with the sample at
mL PBS containing 50 mg/mL RNase A (Roche), incu-
m
4 ꢁC. Individual samples were run on a Becton Dickinson FACSCa-
libur flowcytometer using a 488 nm coherent laser and the acqui-
sition software Cellquest Pro version 5.2.1. The cell population was
identified and gated (R1) on a forward scatter (FSC) vs. side scatter
(SSC) dot plot in acquisition mode. Fluorescent Channel 2 (FL2) at
575 nm was used for Propidium Iodide detection. A dot plot of FL2A
Media
0.1% DMSO
E/Z-4l
Fig. 6. Quantification of caspase 3 enzyme activity in WHCO1 cells treated with E/Z-4l.
WHCO1 cells were treated with 25 M E/Z-4l for 24 h. Cell lysate was then collected
m
and assayed for caspase 3 enzyme activity using the Roche caspase 3 enzyme assay kit
as described in Materials and Methods. Bars, SD for each sample analyzed in triplicate;
**P < 0.01, relative to DMSO control.

246
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
Non-essential for bioactivity
Activates the disulfide
O
End-group modulates
bioactivity
S
S
S
Z-isomer is more
potent than E-
End-group modulates
bioactivity
The more electrophilic sulfur
of the disulfide pharmacophore
Fig. 7. Proposed structureeactivity hypothesis for Z-ajoene in its inhibition of WHCO1 cancer-cell proliferation.
(area) vs FL2W (width) was used to identify single cells (R2) and to
thus eliminate doublets. A histogram plot of FL2A was used to
enumerate G1/G0, S-phase and G2 populations and a threshold of 52
on the FSC channel was set to remove sample debris. Nile Red
Fluorescent particles were used for instrument standardization,
stability and reproducibility. Data was analysed using MODFIT LT
version 2.0, software.
for 2 h after which the fluorescence was measured again
(t1 ¼120 min). The caspase 3 enzyme activity is calculated as the
rate of change in fluorescence (DFU) between time intervals t0 and t1.
4.5. Statistical analysis
Student’s t test was used for statistical significance of differences
in treated vs untreated samples. P < 0.05 was considered to be
significant.
4.3. Quantification of apoptosis
WHCO1 cells were seeded for quadruplicate experiments
4.6. General synthesis methods
(5 ꢃ103 cells, 90
m
L) in a 96-well cell culture plate. The following
L) were
M compound
day, DMSO solutions of each compound in media (10
m
Thin layer chromatography (TLC) was used to monitor reactions
using aluminium-backed plates coated with silica-gel F254.
added to the cells to give a final concentration of 10
m
and 0.1% DMSO. Control cells received 0.1% DMSO alone. After 16 h
treatment, the cell lysate was collected and pelleted. The cyto-
plasmic fraction (supernatant, 20 mL) was transferred to a strepa-
vadin coated plate for analysis by the Roche Cell Death Elisa Kit
which was performed according to the manufacturer’s instruction.
Compounds on TLC plates were observed by a combination of ultra-
violet light, iodine vapour, or by spraying with a 2.5% solution of
anisaldehyde in a mixture of sulfuric acid and ethanol (1:10 v/v)
and then heating at 150 ꢁC. Column chromatography was per-
formed using silica-gel 60 mesh. Infrared spectra were recorded in
chloroform, dichloromethane or neat on caesium chloride or
sodium chloride plates on an FT-IR spectrophotometer. Elemental
analyses were performed using a CHN elemental analyser. High
resolution mass spectrometry data was obtained using a micromass
spectrometer. 1H and 13C NMR spectra were recorded at 300 MHz
for 1H and 75.5 MHz for 13C or at 400 MHz for 1H and 100.6 MHz for
13C in deuteriochloroform (CDCl3) unless otherwise stated. Chem-
Briefly, the freshly prepared immunoreagent (80 mL) containing
anti-histone biotin and anti-DNA-POD was added to each well and
the plate was incubated on a shaker at room temperature for 2 h.
The solution was then removed by gentle tapping, washed (three
times) and then incubated with ABTS (100
mL) for 15 min. The
reaction was quenched by adding the ABTS stop solution (100
m
L)
after which the absorbance at 405 nm and 495 nm was read using
a Thermoscientific multiskan FC plate reader.
ical shifts are quoted using residual chloroform (
d
7.26 in 1H NMR
and
d
77.00 in 13C NMR) as an internal standard. All chemical shifts
are reported in ppm and all coupling constants are quoted in Hz. All
chemicals and reagents were commercially available. Dichloro-
methane (DCM) and THF were purified using standard methods
and freshly distilled before use. Degassed methanol or toluene as
stated in the text refers to purging the solvent of oxygen with
nitrogen gas.
4.4. Quantification of caspase-3 enzyme activity
WHCO1 cells were seeded (1.8 ꢃ 106 cells) for triplicate experi-
ments in 145 mm cell culture dishes. The following day E/Z-4l in
DMSO was added to give a final concentration of 25 mM and 0.1%
DMSO. Control cells received media alone or 0.1% DMSO in media
alone. Cells were incubated for 24 h and then lifted with trypsin,
pelleted and washed with cold PBS. The cells were then resuspended
4.7. Experimental procedures for synthesis of the ajoene
derivatives(i) Addition of the propargyl group to (1)
in lysis buffer (300
Caspase-3 Activity Assay which was followed according to the
m
L) and stored at ꢀ20 ꢁC ready for the Roche
manufacturer’s instruction. Briefly the provided 96 well plates were
Thiol R1SH or its corresponding isothiouronium salt
(R1SC(¼NH2)NHþ2 Brꢀ) was dissolved in degassed methanol (0.5 M)
at 0 ꢁC and solid KOH added (1.2 equiv for R1SH or 2.5 equiv for the
salt). After 5 min, propargyl bromide (1.5 equiv, 80% in toluene) was
added and the mixture left to warm to room temperature. After
a few hours, TLC had indicated complete propargylation to a less
polar product, whereupon the methanol was removed under
vacuum and the residue extracted from water with ethyl acetate or
dichloromethane (3 times). Following drying and removal of
solvent, the residue was chromatographed with toluene/hexane
mixtures on silica-gel to afford the propargylic sulfide 1 product
discernible by NMR and IR spectroscopy. In some cases the sulfide
could be readily distilled in vacuo.
coated with the anti-caspase-3 coating solution (100 mL per well) at
4 ꢁC overnight. The coating solution was removed by tapping and
non-specific binding was blocked using the provided blocking buffer
(200
tapping followed by washing (three times ꢃ 200
was then added to the wells (200
L) and incubated for 1 h at 37 ꢁC.
The lysate was then removed by tapping followed by washing (three
times, 200 L). The substrate solution was then added (freshly
prepared, 100 L) and the fluorescence reading for time zero
mL, 30 min). The blocking buffer was removed by gentle
mL). The cell lysate
m
m
m
immediately determined (t0 ¼ 0) on a Varian Cary Eclipse Fluores-
cence Spectrophotometer [lex ¼ 390 nm, lem ¼ 505 nm]. The plate
was then covered with an adhesive cover foil and incubated at 37 ꢁC

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
247
4.7.1. 3-(Propylthio)-1-propyne (1a) [49]
dC (100 MHz, CDCl3): 18.3 (C-3), 34.7 (Bn), 55.3 (OCH3), 71.1 (C-1),
79.9 (C-2), 114.0 (Cm), 129.3 (Ci), 130.1 (Co), 158.8 (Cp); HRMS (ES):
m/z 193.0701 [M þ H]þ, C11H13OS requires 193.0687.
Obtained as an orange oil (80%): IR nmax (neat)/cmꢀ1 3304
(C^CH), 2967 (C^C), 746 (CeS); dH (300 MHz, CDCl3): 0.97 (3H, t,
J ¼ 7.3 Hz, H-30), 1.61 (2H, sext, J ¼ 7.3 Hz, H-20), 2.20 (1H, t,
J ¼ 2.7 Hz, H-1), 2.63 (2H, t, J ¼ 7.3 Hz, H-10), 3.20 (2H, d, J ¼ 2.7 Hz,
H-3); dC (75 MHz, CDCl3): 13.3 (C-30), 19.0 (C-3), 22.2 (C-20), 33.6 (C-
10), 70.7 (C-1), 80.1 (C-2).
4.8. (ii) Radical addition step to (2)
The propargylic sulfide was dissolved in degassed toluene
(0.5 M) and AIBN or an equivalent radical initiator (5 mol %) added.
The mixture was heated to 85 ꢁC whereupon thiolacetic acid
(1.1 equiv) in toluene (1 M) was added slowly dropwise over 1 h.
Thereafter, the mixture was left stirring until TLC indicated
maximum conversion of starting material. In some cases, extra
thiolacetic acid was added in order to maximize conversion but
care had to be taken not to significantly promote secondary addi-
tion to afford the bis-addition product. On completion of reaction,
the solvent was evaporated and the residue chromatographed
directly on silica-gel using toluene or ethyl acetate/petroleum ether
mixtures to afford vinyl thioacetate 2 as a w2:1 mixture of Z:E
stereoisomers by 1H NMR spectroscopy.
4.7.2. 3-[(10,10-Dimethylethyl)thio]-1-propyne (1b) [50]
Obtained as a pale-yellow oil (57%): dH (400 MHz, CDCl3): 1.36
(9H, s, H-20), 2.16 (1H, t, J ¼ 3.0 Hz, H-1), 3.24 (2H, d, J ¼ 3.0 Hz, H-3);
dC (100 MHz, CDCl3): 16.4 (C-3), 30.7 (C-20), 43.2 (C-10), 70.4 (C-1),
81.5 (C-2).
4.7.3. 3-[30-((tert-Butyl)dimethylsilyloxy)-prop-1-ylthio]-1-
propyne (1c)
Obtained as a colourless oil (70%): IR nmax (neat)/cmꢀ1 3314
(C^CH), 2931 (C^C), 1256 (Si(CH3)2), 776 (CeS); dH (400 MHz,
CDCl3): 0.06 (6H, s, CH3), 0.90 (9H, s, t-butyl), 1.82 (2H, m, H-20),
2.20 (1H, t, J ¼ 2.6 Hz, H-1), 2.76 (2H, t, J ¼ 7.3 Hz, H-10), 3.24 (2H, d,
J ¼ 2.6 Hz, H-3), 3.70 (2H, t, J ¼ 6.1 Hz, H-30); dC (100 MHz,
CDCl3): ꢀ5.3 (Si-(CH3)2), 18.3 (C-(CH3)3), 19.2 (C-3), 25.9 (C-(CH3)3),
28.2 (C-20), 32.1 (C-10), 61.5 (C-30), 70.8 (C-1), 80.1 (C-2).
4.8.1. (E/Z)-S-[3-(Prop-1-ylthio)propen-1-yl] ethanethioate (2a)
Obtained as a 2:1 Z:E mixture (73%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ1 1705 (C]O); (Z)-2a dH (400 MHz, CDCl3): 0.97 (3H, t,
J ¼ 7.3 Hz, H-60), 1.60 (2H, sext, J ¼ 7.3 Hz, H-50), 2.38 (3H, s, H-1),
2.43 (2H, t, J ¼ 7.3 Hz, H-40), 3.17 (2H, dd, J ¼ 1.1, 7.7 Hz, H-30), 5.86
(1H, dt, J ¼ 7.7, 9.6 Hz, H-20), 6.65 (1H, dt, J ¼ 1.1, 9.6 Hz, H-10); dC
(100 MHz, CDCl3): 13.4 (C-60), 22.8 (C-50), 30.8 (C-1), 30.9 (C-30),
33.2 (C-40),119.1 (C-10),128.9 (C-20),191.3 (C-2); (E)-2a dH (400 MHz,
CDCl3): 0.97 (3H, t, J ¼ 7.3 Hz, H-60), 1.58 (2H, sext, J ¼ 7.3 Hz, H-50),
2.34 (3H, s, H-1), 2.43 (2H, t, J ¼ 7.3 Hz, H-40), 3.20 (2H, dd, J ¼ 1.2,
7.4 Hz, H-30), 5.82 (1H, dt, J ¼ 7.4, 15.6 Hz, H-20), 6.53 (1H, dt, J ¼ 1.2,
15.6 Hz, H-10); dC (100 MHz, CDCl3): 13.4 (C-60), 22.6 (C-50), 30.3 (C-
1), 33.0 (C-40), 34.0 (C-30), 118.8 (C-10), 130.5 (C-20), 192.8 (C-2);
HRMS (EI): m/z 190.0485 [M]þ, C8H14OS2 requires 190.0486.
4.7.4. 3-((30-p-Methoxybenzyloxy)prop-1-ylthio)-1-propyne (1d)
Obtained as a colourless oil (57%): IR nmax (neat)/cmꢀ1 3290
(C^CH), 2858 (C^C), 1098 (CeO), 710 (CeS); dH (400 MHz, CDCl3):
1.92 (2H, quint, J ¼ 6.6 Hz, H-20), 2.22 (1H, d, J ¼ 2.4 Hz, H-1), 2.79
(2H, t, J ¼ 6.6 Hz, H-10), 3.24 (2H, d, J ¼ 2.4 Hz, H-3), 3.55 (2H, t,
J ¼ 6.6 Hz, H-30), 3.80 (3H, s, OCH3), 4.44 (2H, s, Bn), 6.88 (2H, d,
J ¼ 8.6 Hz, Hm), 7.26 (2H, d, J ¼ 8.6 Hz, Ho); dC (75 MHz, CDCl3): 19.2
(C-3), 28.5 (C-20), 29.2 (C-10), 55.2 (OCH3), 68.3 (C-30), 70.9 (C-1),
72.6 (Bn), 80.0 (C-2), 113.8 (Cm), 129.2 (Co), 130.5 (Ci), 159.2 (Cp);
HRMS (EI): m/z 250.1010 [M]þ, C14H18O2S requires 250.1028.
4.7.5. N-(3-Bromopropyl)-phthalimide [51]
4.8.2. (E/Z)-S-[3-[(1,1-Dimethylethyl)thio)propen-1-yl]
Obtained as a flakey-white solid (71%): dH (400 MHz, CDCl3):
2.25 (2H, quint, J ¼ 6.8 Hz, H-2), 3.41 (2H, t, J ¼ 6.8 Hz, H-3), 3.83
(2H, t, J ¼ 6.8 Hz, H-1), 7.71 (2H, m, Ar-H), 7.83 (2H, m, Ar-H).
ethanethioate (2b)
Obtained as a 2:1 Z:E mixture (73%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ1 1705 (C]O); (Z)-2b dH (400 MHz, CDCl3): 1.33 (9H, s, t-
butyl), 2.39 (3H, s, H-1), 3.22 (2H, dd, J ¼ 1.2, 7.5 Hz, H-30), 5.90 (1H,
dt, J ¼ 7.5, 9.6 Hz, H-20), 6.61 (1H, dt, J ¼ 1.2, 9.6 Hz, H-10); dC
(100 MHz, CDCl3): 28.5 (C-30), 30.8 (C-1), 30.9 (t-butyl), 42.8
(C(CH3)3), 118.8 (C-10), 129.3 (C-20), 191.2 (C-2); (E)-2b dH (400 MHz,
CDCl3): 1.32 (9H, s, t-butyl), 2.33 (3H, s, H-1), 3.30 (2H, dd, J ¼ 1.2,
7.5 Hz, H-30), 5.88 (1H, dt, J ¼ 7.5, 15.6 Hz, H-20), 6.62 (1H, dt, J ¼ 1.2,
15.6 Hz, H-10); dC (100 MHz, CDCl3): 30.3 (C-1), 31.0 (C(CH3)3), 31.4
(C-30), 42.9 (C(CH3)3), 118.8 (C-10), 131.2 (C-20), 192.8 (C-2); HRMS
(ES): m/z 221.0696 [M þ O þ H]þ, C9H17O2S2 requires 221.0670.
4.7.6. 3-(30-Phthalimidoprop-1-ylthio)-1-propyne (1e)
Obtained as a white solid (69%) which was recrystallized from
petroleum ether/toluene, mp 61e62 ꢁC; IR nmax (neat)/cmꢀ1 3308
(C^CH), 2944 (C^C), 1709 (C]O), 665 (CeS); dH (400 MHz, CDCl3):
2.01 (2H, quint, J ¼ 7.3 Hz, H-20), 2.17 (1H, t, J ¼ 2.6 Hz, H-1), 2.73
(2H, t, J ¼ 7.4 Hz, H-10), 3.25 (2H, d, J ¼ 2.6 Hz, H-3), 3.81 (2H, t,
J ¼ 7.1 Hz, H-30), 7.69 (2H, m, Ar-H), 7.80 (2H, m, Ar-H); dC (100 MHz,
CDCl3): 19.0 (C-3), 27.7 (C-20), 28.7 (C-10), 37.0 (C-30), 71.1 (C-1), 79.6
(C-2), 123.2 (Ar-C), 132.1 (Cqu), 133.9 (Ar-C), 168.3 (C]O); HRMS (EI):
m/z 260.0744 [M þ H]þ, C14H14NO2S requires 260.0745; found: C,
64.57%; H, 4.99%; N, 5.17%; S, 12.12%; C14H13NO2S requires C,
64.84%; H, 5.05%; N, 5.40%; S, 12.37%.
4.8.3. (E/Z)-S-[3-(3-tert-Butyldimethylsilyloxy-prop-1-ylthio)
propen-1-yl] ethanethioate (2c)
Obtained as a 1:1 Z:E mixture (31%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1704 (C]O); (Z)-2c dH (400 MHz, CDCl3): 0.05 (6H, s,
CH3), 0.89 (9H, s, t-butyl), 1.77 (2H, m, H-50), 2.39 (3H, s, H-1), 2.54
(2H, t, J ¼ 7.3 Hz, H-40), 3.18 (2H, dd, J ¼ 1.0, 7.7 Hz, H-30), 3.67 (2H, t,
J ¼ 6.1 Hz, H-60), 5.87 (1H, dt, J ¼ 7.7, 9.6 Hz, H-20), 6.66 (1H, dt,
J ¼ 1.0, 9.6 Hz, H-10); dC (100 MHz, CDCl3): ꢀ5.3 (Si(CH3)2), 18.3
(C(CH3)3), 25.9 (C(CH3)3), 27.8 (C-50), 30.8 (C-1), 31.1 (C-30), 32.5 (C-
40), 61.6 (C-60), 119.3 (C-10), 128.7 (C-20), 191.2 (C-2); (E)-2c dH
(400 MHz, CDCl3): 0.05 (6H, s, CH3), 0.89 (9H, s, t-butyl), 1.77 (2H,
m, H-50), 2.34 (3H, s, H-1), 2.54 (2H, t, J ¼ 7.3 Hz, H-40), 3.22 (2H, dd,
J ¼ 1.2, 7.4 Hz, H-30), 3.68 (2H, t, J ¼ 6.1 Hz, H-60), 5.83 (1H, dt, J ¼ 7.4,
15.6 Hz, H-20), 6.54 (1H, dt, J ¼ 1.2, 15.6 Hz, H-10); dC (100 MHz,
CDCl3): ꢀ5.3 (Si(CH3)2), 18.3 (C(CH3)3), 25.9 (C(CH3)3), 27.5 (C-50),
30.4 (C-1), 32.4 (C-40), 34.2 (C-30), 61.5 (C-60), 118.9 (C-10), 130.4 (C-
4.7.7. 3-Benzylthio-1-propyne (1f) [52]
Obtained as a yellow solid (98%): dH (400 MHz, CDCl3): 2.29 (1H,
t, J ¼ 2.6 Hz, H-1), 3.09 (2H, d, J ¼ 2.6 Hz, H-3), 3.88 (2H, s, Bn), 7.25
(5H, m, Ar-H); dC (100 MHz, CDCl3): 18.4 (C-3), 35.3 (Bn), 71.2 (C-1),
79.8 (C-2), 127.2 (Ar-C), 128.5 (Ar-C), 129.0 (Ar-C),137.4 (Ar-C);
HRMS (EI): m/z 163.0611 (M þ H)þ, C10H11S requires 163.0581.
4.7.8. 1-Methoxy-4-[(2-propyn-1-ylthio)methyl]-benzene (1g) [53]
Obtained as a light-yellow oil (66%): IR nmax (neat)/cmꢀ1 3282
(C^CH), 2860 (C^C), 663 (CeS); dH (400 MHz, CDCl3): 2.25 (1H, t,
J ¼ 2.6 Hz, H-1), 3.04 (2H, d, J ¼ 2.6 Hz, H-3), 3.77 (2H, s, Bn), 3.80
(3H, s, OCH3), 6.84 (2H, d, J ¼ 8.8 Hz, Hm), 7.24 (2H, d, J ¼ 8.8 Hz, Ho);

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20), 193.9 (C-2); HRMS (EI): m/z 320.1297 [M]þ, C14H28O2S2Si
requires 320.1300.
CDCl3): 30.7 (C-1), 30.8 (C-30), 35.2 (Bn), 55.2 (OCH3),113.9 (Cm),119.5
(C-10),128.6 (C-20),129.8 (Ci),130.0 (Co),158.7 (Cp),191.2 (C-2); (E)-2g
dH (400 MHz, CDCl3): 2.35 (3H, s, H-1), 3.08 (2H, dd, J ¼ 1.1, 7.4 Hz, H-
30), 3.61 (2H, s, Bn), 3.78 (3H, s, OCH3), 5.80 (1H, td, J ¼ 7.4,15.2 Hz, H-
20), 6.49 (1H, dt, J ¼ 1.1,15.2 Hz, H-10), 6.83 (2H, d, J ¼ 8.7 Hz, Hm), 7.22
(2H, d, J ¼ 8.7 Hz, Ho); dC (100 MHz, CDCl3): 30.4 (C-1), 33.1 (C-30),
34.4 (Bn), 55.2 (OCH3), 114.0 (Cm), 119.3 (C-10), 129.8 (Ci), 130.1 (Co),
130.3 (C-20), 158.7 (Cp), 192.9 (C-2); HRMS (ES): m/z 307.0404
[M þ Na þ O]þ, C13H16NaO3S2 requires 307.0439.
4.8.4. (E/Z)-S-[3-(3-p-Methoxybenzyl prop-1-ylthio)-propen-1-yl]
ethanethioate (2d)
Compound 2e was obtained as a 3:2 Z:E mixture (64%) as
a colourless oil: IR nmax (neat)/cmꢀ11702 (C]O); (Z)-2d dH (300 MHz,
CDCl3): 1.86 (2H, m, H-50), 2.34 (3H, s, H-1), 2.56 (2H, t, J ¼ 7.2 Hz, H-
40), 3.18 (2H, dd, J ¼ 1.1, 7.7 Hz, H-30), 3.52 (2H, t, J ¼ 6.2 Hz, H-60),
3.80 (3H, s, OCH3), 4.43 (2H, s, Bn), 5.86 (1H, dt, J ¼ 7.7, 9.6 Hz, H-20),
6.66 (1H, dt, J ¼ 1.1, 9.6 Hz, H-10), 6.87 (2H, d, J ¼ 8.6 Hz, Hm), 7.25
(2H, d, J ¼ 8.6 Hz, Ho); dC (75 MHz, CDCl3): 28.0 (C-50), 29.7 (C-40),
30.8 (C-1), 31.0 (C-30), 55.2 (OCH3), 68.5 (C-60), 72.6 (Bn), 113.8 (Cm),
119.3 (C-10), 128.7 (C-20), 129.2 (Co), 130.3 (Ci), 159.2 (Cp), 192.8 (C-
2); (E)-2d dH (300 MHz, CDCl3): 1.86 (2H, m, H-50), 2.37 (3H, s, H-1),
2.56 (2H, t, J ¼ 7.2 Hz, H-40), 3.21 (2H, dd, J ¼ 1.2, 7.4 Hz, H-30), 3.52
(2H, t, J ¼ 6.2 Hz, H-60), 3.80 (3H, s, OCH3), 4.43 (2H, s, Bn), 5.82 (1H,
dt, J ¼ 7.4, 15.6 Hz, H-20), 6.54 (1H, dt, J ¼ 1.2, 15.6 Hz, H-10), 6.87
(2H, d, J ¼ 8.6 Hz, Hm), 7.25 (2H, d, J ¼ 8.6 Hz, Ho); dC (75 MHz,
CDCl3): 27.7 (C-50), 29.5 (C-40), 30.3 (C-1), 34.1 (C-30), 55.2 (OCH3),
68.4 (C-60), 72.6 (Bn), 113.8 (Cm), 119.0 (C-10), 129.2 (Co), 130.3 (Ci),
130.5 (C-20), 159.2 (Cp), 191.2 (C-2); HRMS (EI): m/z 326.1035 [M]þ,
C16H22O3S2 requires 326.1010.
4.9. (iii) Sulfenylation step to vinyl disulfide (3)
Vinyl thioacetate 2 dissolved in methanol (1 M) was cooled
to ꢀ40 ꢁC using a cooling bath with acetonitrile/liquid nitrogen. KOH
in methanol (1.05 equiv, 1 M) was added slowly via syringe and the
mixture stirred for 20 min before cooling the mixture to ꢀ78 ꢁC
(acetone/liquid nitrogen bath), whereupon S-allyl p-toluenesulfo-
nylthioate in methanol (1.1 equiv, 1 M) was syringed into the solu-
tion. The solution was allowed to warm to 0 ꢁC and stirred at this
temperature for 2 h before adding aqueous NH4Cl after which the
organic product was extracted with ethyl acetate or dichloro-
methane (3 times). Following drying and evaporation of solvent, the
residue was subjected to column chromatography to afford vinyl
disulfide 3.
4.8.5. (E/Z)-S-[3-(3-N-Phthalimidoprop-1-ylthio]propen-1-yl]
ethanethioate (2e)
4.9.1. (E,Z)-4,5,9-Trithiadodeca-1,6-diene (3a)
Obtained as a 3:2 Z:E mixture (66%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1713 (C]O); (Z)-2e dH (300 MHz, CDCl3): 1.93 (2H, q,
J ¼ 7.3 Hz, H-50), 2.36 (3H, s, H-1), 2.49 (2H, t, J ¼ 7.3 Hz, H-40), 3.16
(2H, dd, J ¼ 0.8, 7.6 Hz, H-30), 3.74 (2H, t, J ¼ 7.3 Hz, H-60), 5.82 (1H,
dt, J ¼ 7.6, 9.6 Hz, H-20), 6.62 (1H, dt, J ¼ 0.8, 9.6 Hz, H-10), 7.68 (2H,
m, Ar-H), 7.81 (2H, m, Ar-H); dC (100 MHz, CDCl3): 28.2 (C-50), 28.5
(C-40), 29.7 (C-1), 30.9 (C-30), 37.1 (C-60), 119.6 (C-10), 123.2 (Ar-C),
128.4 (C-20), 132.1 (Cqu), 133.9 (Ar-C), 168.3 (C]O), 191.2 (C]O); (E)-2e
dH (300 MHz, CDCl3): 1.93 (2H, q, J ¼ 7.3 Hz, H-50), 2.29 (3H, s, H-1),
2.47 (2H, t, J ¼ 7.3 Hz, H-40), 3.21 (2H, dd, J ¼ 1.0, 7.6 Hz, H-30), 3.76
(2H, t, J ¼ 7.3 Hz, H-60), 5.79 (1H, dt, J ¼ 7.6, 15.6 Hz, H-20), 6.48 (1H,
dt, J ¼ 1.0, 15.6 Hz, H-10), 7.68 (2H, m, Ar-H), 7.81 (2H, m, Ar-H); dC
(100 MHz, CDCl3): 28.1 (C-50), 28.3 (C-40), 30.3 (C-1), 34.0 (C-30),
37.0 (C-60), 119.2 (C-10), 123.2 (Ar-C), 130.3 (C-20), 132.1 (Cqu), 133.9
(Ar-C), 168.3 (C]O), 192.7 (C]O); HRMS (EI): m/z 335.0659 [M]þ,
C16H17NO3S2 requires 335.0650.
Obtained as a 5:4 Z:E mixture (97%) as a colourless oil: IR nmax
(neat)/cmꢀ1 498 (SeS); (Z)-3a dH (400 MHz, CDCl3): 0.99 (3H, t,
J ¼ 7.3 Hz, H-12), 1.61 (2H, sext, J ¼ 7.3 Hz, H-11), 2.47 (2H, t,
J ¼ 7.3 Hz, H-10), 3.27 (2H, dd, J ¼ 1.1, 7.6 Hz, H-8), 3.36 (2H, dt,
J ¼ 0.9, 7.0 Hz, H-3), 5.18 (2H, m, H-1), 5.68 (1H, dt, J ¼ 7.6, 9.2 Hz,
H-7), 5.85 (1H, m, H-2), 6.22 (1H, dt, J ¼ 1.1, 9.2 Hz, H-6); dC
(100 MHz, CDCl3): 13.5 (C-12), 22.9 (C-11), 29.4 (C-8), 33.4 (C-10),
42.0 (C-3), 118.8 (C-1), 128.7 (C-7), 131.7 (C-6), 132.8 (C-2); (E)-3a
dH (400 MHz, CDCl3): 0.99 (3H, t, J ¼ 7.3 Hz, H-12), 1.61 (2H, sext,
J ¼ 7.3 Hz, H-11), 2.44 (2H, t, J ¼ 7.3 Hz, H-10), 3.18 (2H, dd, J ¼ 1.1,
7.6 Hz, H-8), 3.34 (2H, dt, J ¼ 0.9, 7.0 Hz, H-3), 5.18 (2H, m, H-1),
5.85 (1H, m, H-2), 5.88 (1H, dt, J ¼ 7.6, 14.6 Hz, H-7), 6.11 (1H, dt,
J ¼ 1.1, 14.6 Hz, H-6); dC (100 MHz, CDCl3): 13.5 (C-12), 22.7 (C-11),
33.1 (C-10), 33.5 (C-8), 41.3 (C-3), 118.9 (C-1), 127.5 (C-6), 128.6 (C-
7), 132.8 (C-2); HRMS (ES): m/z 237.0460 [M þ O þ H]þ, C9H17OS3
requires 237.0442.
4.8.6. (E/Z)-S-[3-(Benzylthio)propen-1-yl] ethanethioate (2f)
Obtained as a 2:1 Z:E mixture (53%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1702 (C]O); (Z)-2f dH (300 MHz, CDCl3): 2.38 (3H, s,
CH3), 3.11 (2H, dt, J ¼ 1.3, 7.5, H-30), 3.69 (2H, s, Bn), 5.87 (1H, dt,
J ¼ 7.5, 9.6 Hz, H-20), 6.68 (1H, dt, J ¼ 1.3, 9.6 Hz, H-10), 7.32 (5H, m,
Ar-H); dC (100 MHz, CDCl3): 30.7 (C-30), 30.8 (C-1), 35.8 (C-Bn),
119.7 (C-10), 128.5 (Ar-C), 128.6 (Ar-C) 129.0 (Ar-C), 129.1 (C-20),
137.9 (Ar-C), 191.2 (C-2); (E)-2f dH (300 MHz, CDCl3): 2.37 (3H, s,
CH3), 3.11 (2H, dt, J ¼ 1.3, 7.5 Hz, H-30), 3.68 (2H, s, Bn), 5.83 (1 H, dt,
J ¼ 7.5,15.6 Hz, H-20), 6.52 (1H, dt, J ¼ 1.3,15.6 Hz, H-10), 7.26 (5H, m,
Ar-H); dC (100 MHz, CDCl3): 30.4 (C-1), 33.3 (C-30), 35.0 (Bn), 119.5
(C-10), 127.0 (Ar-C), 128.5 (Ar-C), 128.6 (Ar-C) 130.3 (C-20), 137.9 (Ar-
C), 191.2 (C-2); HRMS (EI): m/z 255.0545 [M þ O þ H]þ, C12H15O2S2
requires 255.0513.
4.9.2. (E,Z)-10,10-Dimethyl-4,5,9-trithiaundeca-1,6-diene (3b)
Obtained as a 3:1 Z:E mixture (89%) as a colourless oil: IR nmax
(neat)/cmꢀ1 500 (SeS); (Z)-3b dH (300 MHz, CDCl3): 1.34 (9H, s, t-
butyl), 3.32 (2H, dd, J ¼ 1.2, 7.4 Hz, H-8), 3.35 (2H, dt, J ¼ 1.2, 7.4 Hz,
H-3), 5.15 (2H, m, H-1), 5.72 (1H, dt, J ¼ 7.4, 9.5 Hz, H-7), 5.83 (1H,
m, H-2), 6.15 (1H, dt, J ¼ 1.2, 9.5 Hz, H-6); dC (75 MHz, CDCl3): 26.9
(C-8), 31.0 (C(CH3)3), 42.0 (C-3), 42.8 (C(CH3)3),118.9 (C-1),129.2 (C-
7), 131.1 (C-6), 132.8 (C-2); (E)-3b dH (300 MHz, CDCl3): 1.33 (9H, s,
t-butyl), 3.26 (2H, dd, J ¼ 1.2, 7.4 Hz, H-8), 3.33 (2H, dt, J ¼ 1.2,
7.4 Hz, H-3), 5.21 (2H, m, H-1), 5.82 (1H, m, H-2), 5.93 (1H, dt,
J ¼ 7.4, 14.7 Hz, H-7), 6.17 (1H, dt, J ¼ 1.2, 14.7 Hz, H-6); dC (75 MHz,
CDCl3): 30.8 (C-8), 31.0 (C(CH3)3), 41.2 (C-3), 42.8 (C(CH3)3), 118.9
(C-1), 127.3 (C-6), 129.2 (C-7), 132.8 (C-2); HRMS (ES): m/z 251.0615
[M þ O þ H]þ, C10H19OS3 requires 251.0598.
4.8.7. (E/Z)-S-[3-(p-Methoxybenzylthio)propen-1-yl] ethanethioate
(2g)
4.9.3. (E,Z)-12-(t-Butyldimethylsilyloxy)-4,5,9-trithiadodeca-1, 6-
diene (3c)
Obtained as a 3:2 Z:E mixture (55%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ11700 (C]O);(Z)-2g dH (400 MHz, CDCl3):2.36 (3H, s, H-1),
3.09 (2H, dd, J ¼ 1.0, 7.4 Hz, H-30), 3.63 (2H, s, Bn), 3.78 (3H, s, OCH3),
5.84 (1H, dt, J ¼ 7.4, 9.5 Hz, H-20), 6.66 (1H, dt, J ¼ 1.0, 9.5 Hz, H-10),
6.83 (2H, d, J ¼ 8.3 Hz, Hm), 7.20 (2H, d, J ¼ 8.3 Hz, Ho); dC (100 MHz,
Compound 3c was obtained as a 2:1 Z:E mixture (89%) as a col-
ourless oil: IR nmax (neat)/cmꢀ1 498 (SeS); (Z)-3c dH (400 MHz,
CDCl3): 0.05 (6H, s, CH3), 0.89 (9H, s, t-butyl), 1.80 (2H, m, H-11),
2.57 (2H, t, J ¼ 7.0 Hz, H-10), 3.27 (2H, dd, J ¼ 1.1, 7.7 Hz, H-8), 3.36
(2H, dt, J ¼ 1.0, 7.4 Hz, H-3), 3.69 (2H, t, J ¼ 6.2 Hz, H-12), 5.18 (2H,

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
249
m, H-1), 5.69 (1H, dt, J ¼ 7.7, 9.3 Hz, H-7), 5.83 (1H, m, H-2), 6.21
(1H, dt, J ¼ 1.1, 9.3 Hz, H-6); dC (100 MHz, CDCl3): ꢀ5.3 (Si(CH3)2),
18.3 (C(CH3)3), 26.0 (C(CH3)3), 27.9 (C-11), 29.5 (C-8), 32.6 (C-10),
42.1 (C-3), 61.6 (C-12), 118.9 (C-1), 128.5 (C-7), 131.9 (C-6), 132.8 (C-
2); (E)-3c dH (400 MHz, CDCl3): 0.06 (6H, s, CH3), 0.90 (9H, s, t-
butyl), 1.80 (2H, m, H-11), 2.54 (2H, t, J ¼ 7.4 Hz, H-10), 3.18 (2H, dd,
J ¼ 1.1, 7.4 Hz, H-8), 3.34 (2H, dt, J ¼ 1.0, 7.4 Hz, H-3), 3.69 (2H, t,
J ¼ 6.2 Hz, H-12), 5.18 (2H, m, H-1), 5.83 (1H, m, H-2), 5.88 (1H, dt,
J ¼ 7.4, 14.7 Hz, H-7), 6.11 (1H, dt, J ¼ 1.1, 14.7 Hz, H-6); dC (100 MHz,
CDCl3): ꢀ5.3 (Si(CH3)2), 18.3 (C(CH3)3), 26.0 (C(CH3)3), 27.5 (C-11),
32.5 (C-10), 33.6 (C-8), 41.3 (C-3), 61.6 (C-12), 118.9 (C-1), 127.6 (C-
6), 128.4 (C-7), 132.8 (C-2); HRMS (ES): m/z 277.1131 [M ꢀ SC3H5]þ,
C12H25OS2Si requires 277.1116.
3.33 (2H, dt, J ¼ 1.0, 7.2 Hz,, H-3), 3.65 (2H, s, Bn), 5.15 (2H, m, H-1),
5.80 (1H, m, H-2), 5.85 (1H, dt, J ¼ 7.6, 14.8 Hz, H-7), 6.05 (1H, dt,
J ¼ 1.0, 14.8, Hz, H-6), 7.27 (5H, m, Ar-H); dC (100 MHz, CDCl3): 32.8
(C-8), 35.2 (Bn), 41.3 (C-3),118.9 (C-1),127.0 (Ar-C),127.9 (C-6),128.1
(C-7),128.5 (Ar-C),128.9 (Ar-C),132.8 (C-2),138.0 (Ar-C); HRMS (EI):
m/z 285.0439 [M þ O þ H]þ, C13H17OS3 requires 285.0442.
4.9.7. (E,Z)-10-(p-Methoxyphenyl)-4,5,9-trithiadeca-1,6-diene (3g)
Obtained as a 3:2 Z:E mixture (84%) as a colourless oil: IR nmax
(neat)/cmꢀ1 495 (SeS); (Z)-3g dH (300 MHz, CDCl3): 3.20 (2H, dd,
J ¼ 1.1, 7.7 Hz, H-8), 3.35 (2H, dd, J ¼ 0.9, 7.5 Hz, H-3), 3.67 (2H, s,
Bn), 3.80 (3H, s, OCH3), 5.15 (2H, m, H-1), 5.68 (1H, dt, J ¼ 7.7, 9.5 Hz,
H-7), 5.81 (1H, m, H-2), 6.21 (1H, dt, J ¼ 1.1, 9.5 Hz, H-6), 6.84 (2H, d,
J ¼ 9.0 Hz, Hm), 7.20 (2H, d, J ¼ 9.0 Hz, Ho); dC (100 MHz, CDCl3): 29.4
(C-8), 35.4 (C-Bn), 42.1 (C-3), 55.3 (OCH3), 114.0 (Cm), 118.9 (C-1),
128.2 (C-7), 128.7 (Ci), 130.0 (Co), 132.1 (C-6), 132.8 (C-2), 158.7 (Cp);
(E)-3g dH (300 MHz, CDCl3): 3.07 (2H, dd, J ¼ 1.1, 7.4 Hz, H-8), 3.35
(2H, dd, J ¼ 0.9, 7.5 Hz,, H-3), 3.63 (2H, s, Bn), 3.80 (3H, s, OCH3), 5.15
(2H, m, H-1), 5.81 (2H, m, H-2, H-7), 6.06 (1H, dt, J ¼ 1.1, 14.7, Hz, H-
6), 6.84 (2H, d, J ¼ 9.0 Hz, Hm), 7.20 (2H, d, J ¼ 9.0 Hz, Ho); dC
(100 MHz, CDCl3): 32.7 (C-8), 34.6 (C-Bn), 41.3 (C-3), 55.3 (OCH3),
114.0 (Cm), 118.9 (C-1), 128.0 (C-6), 128.2 (C-7), 128.7 (Ci), 130.0 (Co),
132.8 (C-2), 158.7 (Cp); HRMS (ES): m/z 225.0436 [M ꢀ C3H5S]þ,
C11H13OS2 requires 225.0408.
4.9.4. (E,Z)-12-(p-Methoxybenzyloxy)-4,5,9-trithiadodeca-1,6-
diene (3d)
Obtained as a 3:2 Z:E mixture (89%) as a colourless oil: IR nmax
(neat)/cmꢀ1 498 (SeS); (Z)-3d dH (400 MHz, CDCl3): 1.87 (2H, m, H-
11), 2.60 (2H, t, J ¼ 7.2 Hz, H-10), 3.27 (2H, dd, J ¼ 1.1, 7.8 Hz, H-8),
3.35 (2H, m, H-3), 3.53 (2H, t, J ¼ 6.2 Hz, H-12), 3.80 (3H, s, OCH3),
4.43 (2H, s, Bn), 5.17 (2H, m, H-1), 5.67 (1H, dt, J ¼ 7.8, 9.2 Hz, H-7),
5.83 (1H, m, H-2), 6.22 (1H, dt, J ¼ 1.1, 9.2 Hz, H-6), 6.88 (2H, d,
J ¼ 8.8 Hz, Hm), 7.25 (2H, d, J ¼ 8.8 Hz, Ho); dC (100 MHz, CDCl3): 28.2
(C-11), 29.5 (C-8), 29.8 (C-10), 42.0 (C-3), 55.3 (OCH3), 68.5 (C-12),
72.6 (Bn), 113.8 (Cm), 118.9 (C-1), 128.4 (C-7), 129.2 (Co), 130.4 (Ci),
131.9 (C-6), 132.8 (C-2), 159.2 (Cp); (E)-3d dH (400 MHz, CDCl3): 1.87
(2H, m, H-11), 2.56 (2H, t, J ¼ 7.4 Hz, H-10), 3.18 (2H, dd, J ¼ 1.1,
7.2 Hz, H-8), 3.33 (2H, m, H-3), 3.53 (2H, t, J ¼ 6.2 Hz, H-12), 3.80
(3H, s, OCH3), 4.43 (2H, s, Bn), 5.17 (2H, m, H-1), 5.83 (1H, m, H-2),
5.87 (1H, dt, J ¼ 7.4, 14.4 Hz, H-7), 6.10 (1H, dt, J ¼ 1.1, 14.4 Hz, H-6),
6.88 (2H, d, J ¼ 8.8 Hz, Hm), 7.25 (2H, d, J ¼ 8.8 Hz, Ho); dC (100 MHz,
CDCl3): 27.8 (C-11), 29.6 (C-10), 33.5 (C-8), 41.2 (C-3), 55.3 (OCH3),
68.4 (C-12), 72.6 (Bn), 113.8 (Cm), 118.9 (C-1), 127.7 (C-6), 128.3 (C-
7), 129.2 (Co), 130.4 (Ci), 132.8 (C-2), 159.2 (Cp); HRMS (ES): m/z
356.0949 [M]þ, C17H24O2S3 requires 356.0938.
4.9.8. (E,Z)-4,5,9-Trithiadodeca-6-ene (3h)
Obtained as a 1:1 Z:E mixture (76%) as a colourless oil: IR nmax
(neat)/cmꢀ1 499 (SeS); (Z)-3h dH (300 MHz, CDCl3): 0.97 (3H, t,
J ¼ 7.3 Hz, H-1/ H-12), 0.99 (3H, t, J ¼ 7.3 Hz, H-1/H-12), 1.61 (2H, m,
H-2/H-11), 1.69 (2H, m, H-2/H-11), 2.47 (2H, t, J ¼ 7.3 Hz, H-10), 2.70
(2H, t, J ¼ 7.3 Hz, H-3), 3.26 (2H, dd, J ¼ 1.1, 7.4 Hz, H-8), 5.66 (1H, dt,
J ¼ 7.4, 9.3 Hz, H-7), 6.23 (1H, dt, J ¼ 1.1, 9.3 Hz, H-6); dC (75 MHz,
CDCl3): 12.9 (C-1/C-12), 13.4 (C-1/C-12), 22.2 (C-2/C-11), 22.9 (C-2/
C-11), 29.4 (C-8), 33.2 (C-10), 41.1 (C-3), 128.2 (C-7), 132.2 (C-6); (E)-
3h dH (300 MHz, CDCl3): 0.97 (3H, t, J ¼ 7.3 Hz, H-1/H-12), 0.99 (3H,
t, J ¼ 7.3 Hz, H-1/H-12), 1.61 (2H, m, H-2/H-11), 1.69 (2H, m, H-2/H-
11), 2.43 (2H, t, J ¼ 7.3 Hz, H-10), 2.68 (2H, t, J ¼ 7.3 Hz, H-3), 3.17
(2H, dd, J ¼ 1.1, 7.8 Hz, H-8), 5.87 (1H, dt, J ¼ 7.8, 14.4 Hz, H-7), 6.11
(1H, dt, J ¼ 1.1, 14.4 Hz, H-6); dC (75 MHz, CDCl3): 13.0 (C-1/C-12),
13.4 (C-1/C-12), 22.3 (C-2/C-11), 22.6 (C-2/C-11), 33.0 (C-10), 33.5
(C-8), 40.4 (C-3), 127.8 (C-6), 127.9 (C-7); HRMS (EI): m/z 222.0566
[M]þ, C9H18S3 requires 222.0571.
4.9.5. (E,Z)-12-Phthalimido-4,5,9-trithiadodeca-1,6-diene (3e)
Obtained as a 1:1 Z:E mixture (67%) as a colourless oil: IR nmax
(neat)/cmꢀ11713 (C]O), 499 (SeS); (Z)-3e dH (400 MHz, CDCl3): 1.85
(2H, q, J ¼ 7.1 Hz, H-11), 2.42 (2H, t, J ¼ 7.1 Hz, H-10), 3.15 (2H, dd,
J ¼ 1.0, 7.7 Hz, H-8), 3.22 (2H, dt, J ¼ 1.0, 7.2 Hz, H-3), 3.67 (2H, t,
J ¼ 7.1 Hz, H-12), 5.05 (2H, m, H-1), 5.55 (1H, dt, J ¼ 7.7, 9.4 Hz, H-7),
5.70 (1H, m, H-2), 6.09 (1H, dt, J ¼ 1.0, 9.4 Hz, H-6), 7.60 (2H, m, Ar-
H), 7.73 (2H, m, Ar-H); dC (100 MHz, CDCl3): 28.0 (C-11), 28.5 (C-10),
29.3 (C-8), 37.0 (C-12), 42.0 (C-3), 118.9 (C-1), 123.1 (Ar-C), 128.1 (C-
7), 132.0 (C-2), 132.1 (C-6), 132.6 (Cqu), 133.8 (Ar-C), 168.1 (C]O); (E)-
3e dH (400 MHz, CDCl3): 1.85 (2H, t, J ¼ 7.1 Hz, H-11), 2.38 (2H, q,
J ¼ 7.1 Hz, H-10), 3.07 (2H, dd, J ¼ 1.0, 7.7 Hz, H-8), 3.19 (2H, dt,
J ¼ 1.0, 7.2 Hz, H-3), 3.67 (2H, t, J ¼ 7.1 Hz, H-12), 5.05 (2H, m, H-1),
5.70 (2H, m, H-2, H-7), 5.97 (1H, dt, J ¼ 1.0, 14.8 Hz, H-6), 7.60 (2H,
m, Ar-H), 7.72 (2H, m, Ar-H); dC (100 MHz, CDCl3): 28.0 (C-11), 28.4
(C-10), 33.2 (C-8), 37.0 (C-12), 41.2 (C-3), 118.8 (C-1), 123.1 (Ar-C),
127.7 (C-6), 127.8 (C-7), 132.1 (C-2), 132.7 (Cqu), 133.9 (Ar-C), 168.1
(C]O); HRMS (EI): m/z 221.0511 [M ꢀ C4H7S2 þ H]þ, C11H11NO2S
requires 221.0510.
4.9.9. (E,Z)-2,3,7-Trithiadeca-4-ene (3i)
Obtained as a 3:2 Z:E mixture (76%) as a colourless oil: IR nmax
(neat)/cmꢀ1 498 (SeS); (Z)-3i dH (400 MHz, CDCl3): 0.98 (3H, t,
J ¼ 7.2 Hz, H-10), 1.60 (2H, m, H-9), 2.44 (3H, s, H-1), 2.47 (2H, t,
J ¼ 7.4 Hz, H-8), 3.26 (2H, dd, J ¼ 1.0, 7.4 Hz, H-6), 5.74 (1H, dt,
J ¼ 7.4, 9.3 Hz, H-5), 6.24 (1H, dt, J ¼ 1.0, 9.3 Hz, H-4); dC (100 MHz,
CDCl3): 13.4 (C-10), 22.9 (C-1/C-9), 22.9 (C-1/ C-9), 29.3 (C-6), 33.2
(C-8), 129.2 (C-5), 130.9 (C-4); (E)-3i dH (400 MHz, CDCl3): 0.98 (3H,
t, J ¼ 7.2 Hz, H-10), 1.60 (2H, m, H-9), 2.40 (3H, s, H-1), 2.46 (2H, t,
J ¼ 7.4 Hz, H-8), 3.20 (2H, dd, J ¼ 1.0, 7.4 Hz, H-6), 5.90 (1H, dt,
J ¼ 7.4, 14.6 Hz, H-5), 6.12 (1H, dt, J ¼ 1.0, 14.6, Hz, H-4); dC
(100 MHz, CDCl3): 13.4 (C-10), 22.1 (C-1), 22.6 (C-9), 33.0 (C-8), 33.4
(C-6), 126.4 (C-4), 128.5 (C-5); HRMS (EI): m/z 194.0248 [M]þ,
C7H14S3 requires 194.0258.
4.9.6. (E,Z)-10-Phenyl-4,5,9-trithiadeca-1,6-diene (3f)
Obtained as a 4:3 Z:E mixture (86%) as a colourless oil: IR nmax
(neat)/cmꢀ1 490 (SeS); (Z)-3f dH (400 MHz, CDCl3): 3.19 (2H, dd,
J ¼ 1.0, 7.6 Hz, H-8), 3.33 (2H, dt, J ¼ 1.0, 7.2 Hz, H-3), 3.69 (2H, s, Bn),
5.15 (2H, m, H-1), 5.66 (1H, dt, J ¼ 7.6, 9.8 Hz, H-7), 5.80 (1H, m, H-
2), 6.20 (1H, dt, J ¼ 1.0, 9.8 Hz, H-6), 7.27 (5H, m, Ar-H); dC (100 MHz,
CDCl3): 29.4 (C-8), 36.0 (Bn), 42.1 (C-3), 118.9 (C-1), 127.0 (Ar-C),
128.2 (C-7), 128.5 (Ar-C), 128.9 (Ar-C), 132.3 (C-6), 132.8 (C-2), 138.0
(Ar-C); (E)-3f dH (400 MHz, CDCl3): 3.06 (2H, dd, J ¼ 1.0, 7.2 Hz, H-8),
4.9.10. (E,Z)-8-(p-Methoxyphenyl)-1-phenyl-2,3,7-trithiaocta-4-
ene (3j)
Obtained as a 3:2 Z:E mixture (88%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ1 499 (SeS); (Z)-3j dH (300 MHz, CDCl3): 3.18 (2H, d,
J ¼ 7.6 Hz, H-6), 3.67 (2H, s, H-8), 3.81 (3H, s, OCH3), 3.94 (2H, s, H-
1), 5.61 (1H, dt, J ¼ 7.6, 9.3 Hz, H-5), 5.99 (1H, d, J ¼ 9.3 Hz, H-4),
6.87 (2H, m, ArOMe-Hm), 7.25 (2H, m, ArOMe-Ho), 7.32 (5H, m, Ar-H);

250
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
dC (75 MHz, CDCl3): 29.2 (C-6), 35.3 (C-8), 43.5 (C-1), 55.2 (OCH3),
113.9 (ArOMe-Cm),127.5 (Ar-C),128.1 (C-5),128.5 (Ar-C),129.3 (Ar-C),
129.8 (ArOMe-Ci), 129.9 (ArOMe-Co), 131.6 (C-4), 136.8 (Ar-C), 158.6
(ArOMe-Ci); (E)-3j dH (300 MHz, CDCl3): 3.02 (2H, d, J ¼ 7.0 Hz, H-6),
3.61 (2H, s, H-8), 3.82 (3H, s, OCH3), 3.94 (2H, s, H-1), 5.82 (1H, dt,
J ¼ 7.0, 14.6 Hz, H-5), 5.92 (1H, d, J ¼ 14.6 Hz, H-4), 6.87 (2H, m,
ArOMe-Hm), 7.25 (2H, m, ArOMe-Ho), 7.32 (5H, m, Ar-H); dC (75 MHz,
cases (see Tables 3 and 4), isomers could be separated by gravity
chromatography using a low flow rate. Yields varied in the 60e90%
range and reaction temperatures for optimal conversion were
substrate-specific.
4.10.1. (E,Z)-4,5,9-Trithiadodeca-1,6-diene 9-oxide (4a) [54]
Obtained as a 3:2 Z:E mixture (82% and separable) as colourless
oils: IR nmax (neat)/cmꢀ1 1021 (S]O), 498 (SeS); (Z)-4a dH (400 MHz,
CDCl3): 1.07 (3H, t, J ¼ 7.4 Hz, H-12), 1.79 (2H, m, H-11), 2.60 (2H, dt,
J ¼ 8.2, 13.7 Hz, H-10a), 2.70 (2H, dt, J ¼ 8.2, 13.7 Hz, H-10b), 3.36
(2H, d, J ¼ 7.3 Hz, H-3), 3.55 (1H, ddd, J ¼ 0.8, 8.0, 13.2 Hz, H-8a),
3.62 (1H, ddd, J ¼ 0.8, 8.0, 13.2 Hz, H-8b), 5.16 (2H, m, H-1), 5.75
(1H, dt, J ¼ 8.0, 9.2 Hz, H-7), 5.82 (1H, m, H-2), 6.53 (1H, d,
J ¼ 9.2 Hz, H-6); dC (100 MHz, CDCl3): 13.3 (C-12), 16.2 (C-11), 42.0
(C-3), 50.9 (C-8), 53.4 (C-10), 118.4 (C-7), 119.2 (C-1), 132.5 (C-2),
138.2 (C-6); HRMS (ES): m/z 259.0263 [M þ Na]þ, C9H16ONaS3
requires 259.0261; (E)-4a dH (400 MHz, CDCl3): 1.06 (3H, t,
J ¼ 7.3 Hz, H-12), 1.80 (2H, m, H-11), 2.58 (1H, m, H-10-a), 2.67 (1H,
m, H-10-b), 3.33 (2H, d, J ¼ 7.4 Hz, H-3), 3.42 (1H, ddd, J ¼ 0.9, 7.9,
13.2 Hz, H-8a), 3.51 (1H, ddd, J ¼ 0.9, 7.9, 13.2 Hz, H-8b), 5.15 (2H, m,
H-1), 5.80 (1H, m, H-2), 5.90 (1H, dt, J ¼ 7.9, 14.6 Hz, H-7), 6.34 (1H,
d, J ¼ 14.6 Hz, H-4); dC (100 MHz, CDCl3): 13.3 (C-12), 16.1 (C-11),
41.2 (C-3), 53.0 (C-10), 54.5 (C-8),117.1 (C-7),119.1 (C-1),132.4 (C-2),
134.2 (C-6); HRMS (ES): m/z 259.0263 [M þ Na]þ, C9H16ONaS3
requires 259.0261.
CDCl3): 32.6 (C-6), 34.5 (C-8), 42.7 (C-1), 55.2 (OCH3), 113.9 (ArOMe
-
Cm), 127.5 (C-4), 127.5 (Ar-C), 128.1 (C-5), 128.5 (Ar-C), 129.3 (Ar-C),
129.8 (ArOMe-Ci), 130.0 (ArOMe-Co), 136.8 (Ar-C), 158.6 (ArOMe-Cp);
HRMS (ES): m/z 365.0695 [M þ O þ H]þ, C18H21O2S3 requires
365.0704.
4.9.11. (E,Z)-1-(p-Fluorophenyl)-8-(p-methoxyphenyl)-2,3,7-
trithiaocta-4-ene (3k)
Obtained as a 4:3 Z:E mixture (70%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ1 498 (SeS); (Z)-3k dH (300 MHz, CDCl3): 3.17 (2H, d,
J ¼ 7.7 Hz, H-6), 3.66 (2H, s, H-8), 3.80 (3H, s, OCH3), 3.89 (2H, s, H-
1), 5.61 (1H, dt, J ¼ 7.7, 9.2 Hz, H-5), 5.96 (1H, dt, J ¼ 1.1, 9.2 Hz, H-4),
6.86 (2H, m, ArOMe-Hm), 7.00 (2H, m, ArF-Hm), 7.25 (4H, m, ArOMe-Ho
and ArF-Ho); dC (75 MHz, CDCl3): 29.2 (C-6), 35.3 (C-8), 42.5 (C-1),
55.2 (OCH3), 113.9 (ArOMe-Cm), 115.4 (d, JCF ¼ 21.1 Hz, ArF-Cm), 128.2
(C-5), 129.8 (ArOMe-Ci), 129.9 (ArOMe-Co), 130.9 (d, JCF ¼ 8.0 Hz, ArF-
Co), 131.4 (C-4), 132.6 (d, JCF ¼ 3.0 Hz, ArF-Ci), 158.6 (ArOMe-Cp), 162.2
(d, JCF ¼ 245.3 Hz, ArF-Cp); (E)-3k dH (300 MHz, CDCl3): 3.02 (2H, dd,
J ¼ 1.1, 7.1 Hz, H-6), 3.60 (2H, s, H-8), 3.81 (3H, s, OCH3), 3.89 (2H, s,
H-1), 5.80 (1H, dt, J ¼ 7.1,14.7 Hz, H-5), 5.90 (1H, d, J ¼ 14.7 Hz, H-4),
6.86 (2H, m, ArOMe-Hm), 7.00 (2H, m, ArF-Hm), 7.25 (4H, m, ArOMe-Ho
and ArF-Ho); dC (75 MHz, CDCl3): 32.6 (C-6), 34.6 (C-8), 41.8 (C-1),
55.2 (OCH3), 113.9 (ArOMe-Cm), 115.4 (d, JCF ¼ 21.1 Hz, ArF-Cm), 127.3
(C-4), 128.3 (C-5), 129.8 (ArOMe-Ci), 129.9 (ArOMe-Co), 130.9 (d,
JCF ¼ 9.5 Hz, ArF-Co), 132.6 (d, JCF ¼ 3.0 Hz, ArF-Ci), 158.6 (ArOMe-Cp),
162.2 (d, JCF ¼ 245.3 Hz, ArF-Cp); HRMS (ES): m/z 405.0450
[M þ O þ Na]þ, C18H19FNaO2S3 requires 405.0429.
4.10.2. (E,Z)-10,10-Dimethyl-4,5,9-trithiaundeca-1,6-diene 9-oxide
(4b)
Obtained as a 2:1 Z:E mixture (78%) and as a colourless oil: IR
nmax (neat)/cmꢀ1 1043 (S]O), 498 (SeS); (Z)-4b dH (400 MHz,
CDCl3): 1.30 (9H, s, CH3), 3.31 (1H, ddd, J ¼ 1.0, 8.1, 13.2 Hz, H-8a),
3.37 (2H, m, H-3), 3.51 (1H, ddd, J ¼ 1.0, 8.1, 13.2 Hz, H-8b), 5.19 (2H,
m, H-1), 5.85 (2H, m, H-2 and H-7), 6.50 (1H, d, J ¼ 9.2 Hz, H-6); dC
(100 MHz, CDCl3): 22.9 (CH3), 42.0 (C-3), 45.8 (C-8), 54.0 (C(CH3)3),
119.1 (C-1), 121.1 (C-7), 132.7 (C-2), 136.7 (C-6); HRMS (ES): m/z
273.0417 [M þ Na]þ, C10H18ONaS3 requires 273.0418; (E)-4b dH
(400 MHz, CDCl3): 1.28 (9H, s, CH3), 3.24 (1H, ddd, J ¼ 1.0, 7.5,
13.0 Hz, H-8a), 3.37 (2H, m, H-3), 3.37 (1H, ddd, J ¼ 1.0, 7.5, 13.0 Hz,
H-8b), 5.19 (2H, m, H-1), 5.85 (1H, m, H-2), 6.02 (1H, dt, J ¼ 7.5,
14.8 Hz, H-7), 6.37 (1H, d, J ¼ 14.8 Hz, H-6); dC (100 MHz, CDCl3):
23.0 (CH3), 41.1 (C-3), 49.5 (C-8), 53.7 (C(CH3)3), 119.2 (C-1), 119.8
(C-7), 132.5 (C-2), 133.0 (C-6); HRMS (ES): m/z 273.0417 [M þ Na]þ,
C10H18ONaS3 requires 273.0418.
4.9.12. (E,Z)-1,8-(Bis-p-methoxyphenyl)-2,3,7-trithia-octa-4-ene
(E/Z-3l)
Obtained as a 1:1 Z:E mixture (83%) as a pale-yellow oil: IR nmax
(neat)/cmꢀ1 500 (SeS); (Z)-E/Z-3l dH (400 MHz, CDCl3): 3.20 (2H, d,
J ¼ 0.8, 7.6 Hz, H-6), 3.68 (2H, s, H-8), 3.80 (3H, s, OCH3), 3.81 (3H, s,
OCH3), 3.91 (2H, s, H-1), 5.63 (1H, dt, J ¼ 7.6, 9.4 Hz, H-5), 6.03 (1H,
d, J ¼ 9.4 Hz, H-4), 6.87 (4H, m, Hm), 7.23 (4H, m, Ho); dC (100 MHz,
CDCl3): 29.4 (C-6), 35.4 (C-8), 43.1 (C-1), 55.3 (OCH3), 114.0 (Cm),
114.1 (Cm), 128.0 (C-5), 128.8 (Ci), 130.0 (Ci), 130.1 (Co), 130.6 (Co),
131.9 (C-4), 158.7 (Cp), 159.2 (Cp); (E)-E/Z-3l dH (400 MHz, CDCl3):
3.03 (2H, d, J ¼ 7.3 Hz, H-6), 3.61 (2H, s, H-8), 3.78 (3H, s, OCH3),
3.80 (3H, s, OCH3), 3.91 (2H, s, H-1), 5.83 (1H, dt, J ¼ 7.3, 14.6 Hz, H-
5), 5.94 (1H, d, J ¼ 14.6 Hz, H-4), 6.87 (4H, m, Hm), 7.23 (4H, m, Ho);
dC (100 MHz, CDCl3): 32.7 (C-6), 34.7 (C-8), 42.3 (C-1), 55.3
(OCH3 ꢃ 2), 114.0 (Cm), 114.1 (Cm), 127.8 (C-4), 128.1 (C-5), 128.8 (Ci),
130.0 (Ci), 130.1 (Co),130.6 (Co), 158.7 (Cp), 159.2 (Cp); HRMS (ES): m/
z 395.0807 [M þ O þ H]þ, C19H23O3S3 requires 395.0809.
4.10.3. (E,Z)-4,8,9-Trithiadodeca-6,11-dien-1-ol 4-oxide (4c)
(E,Z)-12-(t-Butyldimethylsilyloxy)-4,5,9-trithiadodeca-1,6-diene
4-oxide was obtained from the oxidation as a 3:2 Z:E mixture (86%
and separable) as colourless oils: IR nmax (neat)/cmꢀ11027 (S]O), 497
(SeS); (Z)-isomer dH (400 MHz, CDCl3): 0.05 (6H, s, Si(CH3)2), 0.89
(9H, s, CH3),1.97 (2H, m, H-11), 2.79 (2H, m, H-10), 3.37 (2H, dt, J ¼ 1.0,
7.2 Hz, H-3), 3.56 (1H, ddd, J ¼ 1.0, 8.0, 13.3 Hz, H-8a), 3.64 (1H, ddd,
J ¼ 1.0, 8.0, 13.3 Hz, H-8b), 3.75 (2H, m, H-12), 5.18 (2H, m, H-1), 5.79
(2H, m, H-2 and H-7), 6.55 (1H, d, J ¼ 9.6 Hz, H-6); dC (100 MHz,
CDCl3): ꢀ5.4 (Si(CH3)2), 18.2 (C(CH3)3), 25.7 (C-11), 25.9 (C(CH3)3),
42.1 (C-3), 48.4 (C-10), 51.0 (C-8), 61.4 (C-12),118.4 (C-7), 119.2 (C-1),
132.3 (C-2),138.3 (C-6); HRMS (ES): m/z 366.1169 [M]þ, C15H30O2S3Si
requires 366.1177; (E)-isomer dH (300 MHz, CDCl3): 0.05 (6H, s,
Si(CH3)2), 0.88(9H, s, CH3),1.96(2H, m, H-11), 2.76 (2H, m, H-10), 3.34
(2H, dt, J ¼ 1.0, 7.5 Hz, H-3), 3.46 (1H, ddd, J ¼ 1.1, 8.0, 12.7 Hz, H-8a),
3.54 (1H, ddd, J ¼ 1.1, 8.0,12.7 Hz, H-8b), 3.74 (2H, m, H-12), 5.17 (2H,
m, H-1), 5.82 (1H, m, H-2), 5.93 (1H, dt, J ¼ 7.5,15.0 Hz, H-7), 6.36 (1H,
dt, J ¼ 1.1, 14.7 Hz, H-6); dC (75 MHz, CDCl3): ꢀ5.4 (Si(CH3)2), 18.2
(C(CH3)3), 25.7 (C-11), 25.9 (C(CH3)3), 41.3 (C-3), 48.1 (C-10), 54.6 (C-
8), 61.4 (C-12), 117.1 (C-7),119.2 (C-1),132.5 (C-2),134.3 (C-6); HRMS
(ES): m/z 366.1171 [M]þ, C15H30O2S3Si requires 366.1177.
4.10. (iv) Oxidation step to (4)
The vinyl disulfide 3 dissolved in dichloromethane (0.2 M) was
cooled to ꢀ78 ꢁC under nitrogen, and m-CPBA (1.1 equiv) added in
one portion. The reaction was allowed to warm to room tempera-
ture over a few hours until TLC (40% ethyl acetate/petroleum ether)
indicated the consumption of starting material. Saturated aqueous
NaHCO3 was then added and the product extracted into ethyl
acetate or dichloromethane (3 times). The combined organic layers
were dried and concentrated in vacuo to afford a residue, which was
purified on a silica-gel column using petroleum ether/ethyl acetate
mixtures to afford the ajoene derivative as an E/Z mixture. In some

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
251
The TBS ethers were individually deprotected with HF in CH3CN
to afford individual isomers of 4c:
m, Ar-H), 7.85 (2H, m, Ar-H), dC (100 MHz, CDCl3): 22.2 (C-11), 36.9
(C-12), 41.3 (C-3), 48.5 (C-10), 54.7 (C-8), 116.7 (C-7), 119.2 (C-1),
123.4 (Ar-C), 132.0 (C-2), 132.5 (Arqu), 134.2 (Ar-C), 134.7 (C-6),
168.3 (C]O); HRMS (ES): m/z 382.0583 [M þ H]þ, C17H20NO3S3
requires 382.0605.
(Z)-4c was obtained as a colourless oil (84%): IR nmax (neat)/
cmꢀ1 3378 (OH), 1615 (C]C), 1015 (S]O), 497 (SeS); dH (400 MHz,
CDCl3): 2.07 (2H, m, H-11), 2.85 (2H, m, H-10), 2.87 (1H, br s, OH),
3.37 (2H, dt, J ¼ 1.0, 7.3 Hz, H-3), 3.61 (1H, ddd, J ¼ 1.0, 8.0, 13.3 Hz,
H-8a), 3.67 (1H, ddd, J ¼ 1.0, 8.0, 13.3 Hz, H-8b), 3.75 (2H, m, H-
12), 5.18 (2H, m, H-1), 5.73 (1H, dt, J ¼ 8.0, 9.6 Hz, H-7), 5.81 (1H,
ddt, J ¼ 7.3, 10.0, 16.5 Hz, H-2), 6.56 (1H, dt, J ¼ 1.0, 9.6 Hz, H-6); dC
(100 MHz, CDCl3): 26.5 (C-11), 42.1 (C-3), 48.7 (C-10), 50.8 (C-8),
61.0 (C-12), 118.0 (C-7), 119.3 (C-1), 132.6 (C-2), 138.7 (C-6); HRMS
(ES): m/z 275.0229 [M þ Na]þ, C9H16NaO2S3 requires 275.0210;
(E)-4c was obtained as a colourless oil (89%): IR nmax (neat)/cmꢀ1
3481 (OH), 1630 (C]C), 1010 (S]O), 495 (SeS); dH (400 MHz,
CDCl3): 2.03 (2H, m, H-11), 2.82 (2H, m, H-10), 3.25 (1H, br s, OH),
3.33 (2H, d, J ¼ 7.2 Hz, H-3), 3.53 (2H, m, H-8), 3.72 (2H, t,
J ¼ 5.6 Hz, H-12), 5.15 (2H, m, H-1), 5.81 (1H, ddt, J ¼ 7.2, 9.8,
16.9 Hz, H-2), 5.89 (1H, dt, J ¼ 7.6, 14.8 Hz, H-7), 6.37 (1H, d,
J ¼ 14.8 Hz, H-6); dC (100 MHz, CDCl3): 26.2 (C-11), 41.2 (C-3), 48.2
(C-10), 54.3 (C-8), 60.8 (C-12), 116.6 (C-7), 119.2 (C-1), 132.4 (C-2),
134.7 (C-6); HRMS (ES): m/z 275.0201 [M þ Na]þ, C9H16NaO2S3
requires 275.0210.
4.10.6. (E,Z)-10-Phenyl-4,5,9-trithiadeca-1,6-diene 9-oxide (4f)
Obtained as a 6:5 Z:E mixture (65%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1047 (S]O), 493 (SeS); (Z)-4f dH (300 MHz, CDCl3): 3.36
(2H, m, H-3), 3.44 (1H, m, H-8a), 3.56 (1H, m, H-8b), 3.98 (2H, s, Bn),
5.18 (2H, m, H-1), 5.76 (1H, m, H-7), 5.84 (1H, m, H-2), 6.57 (1H, d,
J ¼ 9.6 Hz, H-6), 7.34 (5H, m, Ar-H); dC (75 MHz, CDCl3): 42.1 (C-3),
49.8 (C-8), 57.6 (Bn), 118.3 (C-7), 119.2 (C-1), 128.4 (Ar-C), 129.0 (Ar-
C), 129.9 (Ar-C), 130.1 (Ar-C), 132.6 (C-2), 138.5 (C-6); HRMS (ES): m/
z 285.0452 [M þ H]þ, C13H17OS3 requires 285.0442; (E)-4f IR nmax
(neat)/cmꢀ11634 (C]C),1047 (S]O), 493 (SeS); dH (300 MHz, CDCl3):
3.30 (1H, m, H-8a), 3.36 (2H, m, H-3), 3.49 (1H, m, H-8b), 3.98 (2H, s,
Bn), 5.18 (2H, m, H-1), 5.84 (1H, m, H-2), 5.96 (1H, dt, J ¼ 7.5,
14.7 Hz, H-7), 6.36 (1H, d, J ¼ 14.7 Hz, H-6), 7.34 (5H, m, Ar-H); dC
(75 MHz, CDCl3): 41.3 (C-3), 52.9 (C-8), 57.0 (Bn), 117.0 (C-7), 119.2
(C-1), 128.5 (Ar-C), 129.1 (Ar-C), 129.7 (Ar-C), 130.0 (Ar-C), 132.5 (C-
2), 134.7 (C-6); HRMS (ES): m/z 285.0452 [M þ H]þ, C13H17OS3
requires 285.0442.
4.10.4. (E,Z)-12-(p-Methoxybenxyloxy)-4,5,9-trithiadodeca-1,6-
diene 9-oxide (4d)
4.10.7. (E,Z)-10-(p-Methoxyphenyl)-4,5,9-trithiadeca-1,6-diene 9-
oxide (4g)
Obtained as a 2:1 Z:E mixture (92% and separable) as colourless
oils: IR nmax (neat)/cmꢀ1 1030 (S]O), 498 (SeS); (Z)-4d dH (300 MHz,
CDCl3): 2.06 (2H, m, H-11), 2.79 (2H, m, H-10), 3.36 (2H, dt, J ¼ 1.1,
7.2 Hz, H-3), 3.58 (4H, m, H-8, H-12), 3.80 (3H, s, OCH3), 4.44 (2H, s,
Bn), 5.17 (2H, m, H-1), 5.74 (1H, dt, J ¼ 8.0, 9.3 Hz, H-7), 5.82 (1H, m,
H-2), 6.54 (1H, dt, J ¼ 1.1, 9.3 Hz, H-6), 6.87 (2H, d, J ¼ 8.7 Hz, Hm),
7.24 (2H, d, J ¼ 8.7 Hz, Ho); dC (75 MHz, CDCl3): 23.2 (C-11), 42.1 (C-
3), 48.7 (C-10), 51.0 (C-8), 55.3 (OCH3), 68.1 (C-12), 72.7 (Bn), 113.8
(Cm), 118.4 (C-7), 119.2 (C-1), 129.3 (Co), 130.2 (Ci), 132.6 (C-2), 138.3
(C-6), 159.3 (Cp); HRMS (ES): m/z 395.0791 [M þ Na]þ,
C17H24NaO3S3 requires 395.0785; (E)-4d dH (300 MHz, CDCl3): 2.05
(2H, m, H-11), 2.77 (2H, m, H-10), 3.33 (2H, dt, J ¼ 1.0, 7.5 Hz, H-3),
3.44 (1H, ddd, J ¼ 1.0, 7.8, 13.1 Hz, H-8a), 3.53 (1H, ddd, J ¼ 1.0, 7.8,
13.1 Hz, H-8b), 3.59 (2H, m, H-12), 3.80 (3H, s, OCH3), 4.44 (2H, s,
Bn), 5.17 (2H, m, H-1), 5.82 (1H, m, H-2), 5.92 (1H, dt, J ¼ 7.8,
14.7 Hz, H-7), 6.35 (1H, dt, J ¼ 1.0, 14.7 Hz, H-6), 6.88 (2H, d,
J ¼ 8.7 Hz, Hm), 7.24 (2H, d, J ¼ 8.7 Hz, Ho); dC (75 MHz, CDCl3): 23.1
(C-11), 41.2 (C-3), 48.3 (C-10), 54.6 (C-8), 55.3 (OCH3), 68.0 (C-12),
72.7 (Bn), 113.9 (Cm), 117.1 (C-7), 119.2 (C-1), 129.3 (Co), 130.1 (Ci),
132.5 (C-2), 134.3 (C-6), 159.3 (Cp); HRMS (ES): m/z 395.0790
[M þ Na]þ, C17H24NaO3S3 requires 395.0785.
Obtained as a 2:1 Z:E mixture (85%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1015 (S]O), 495 (SeS); (Z)-4g dH (300 MHz, CDCl3):
3.37 (2H, m, H-3), 3.43 (1H, ddd, J ¼ 1.1, 7.6, 13.5 Hz, H-8a), 3.54
(1H, ddd, J ¼ 1.1, 7.6, 13.5 Hz, H-8b), 3.80 (3H, s, OCH3), 3.92 (2H,
s, Bn), 5.17 (2H, m, H-1), 5.77 (1H, dt, J ¼ 7.6, 9.5 Hz, H-7), 5.83
(1H, m, H-2), 6.56 (1H, dt, J ¼ 1.1, 9.5 Hz, H-6), 6.90 (2H, m, Hm);
7.22 (2H, m, Ho); dC (100 MHz, CDCl3): 42.1 (C-3), 49.5 (C-8), 55.3
(OCH3), 56.9 (Bn), 114.5 (Cm), 118.4 (C-7), 119.2 (C-1), 121.6 (Ci),
131.2 (Co), 132.6 (C-2), 138.2 (C-6), 159.8 (Cp). HRMS (ES): m/z
315.0551 [M þ H]þ, C14H19O2S3 requires 315.0547; (E)-4g dH
(300 MHz, CDCl3): 3.30 (1H, ddd, J 1.1, 8.0, 13.5 Hz, H-8a), 3.36
(2H, m, H-3), 3.45 (1H, ddd, J ¼ 1.1, 8.0, 13.5 Hz, H-8b), 3.80 (3H, s,
OCH3), 3.92 (2H, s, Bn), 5.17 (2H, m, H-1), 5.83 (1H, m, H-2), 5.92
(1H, dt, J ¼ 8.0, 14.7 Hz, H-7), 6.35 (1H, dt, J ¼ 1.1, 14.7 Hz, H-6),
6.90 (2H, m, Hm), 7.22 (2H, m, Ho); dC (100 MHz, CDCl3): 41.3 (C-
3), 52.7 (C-8), 55.3 (OCH3), 56.3 (Bn), 114.5 (Cm), 117.2 (C-7), 119.2
(C-1), 121.5 (Ci), 131.2 (Co), 132.5 (C-2), 134.4 (C-6), 159.8 (Cp);
HRMS (ES): m/z 315.0551 [M þ H]þ, C14H19O2S3 requires
315.0547.
4.10.8. (E,Z)-4,5,9-Trithiadodeca-6-ene 9-oxide (4h)
4.10.5. (E,Z)-12-Phthalimido-4,5,9-trithiadodeca-1,6-diene 9-oxide
(4e)
Obtained as a 2:3 Z:E mixture (68% and separable) as colourless
oils: IR nmax (neat)/cmꢀ11045 (S]O), 498 (SeS); (Z)-4h dH (300 MHz,
CDCl3): 0.98 (3H, t, J ¼ 7.2 Hz, H-1), 1.07 (3H, t, J ¼ 7.5 Hz, H-12), 1.69
(2H, sext, J ¼ 7.2 Hz, H-2), 1.80 (2H, m, H-11), 2.66 (2H, m, H-10),
2.70 (2H, t, J ¼ 7.2 Hz, H-3), 3.54 (1H, ddd, J ¼ 0.8, 8.0, 13.2 Hz, H-8a),
3.64 (1H, ddd, J ¼ 0.8, 8.0,13.2 Hz, H-8b), 5.74 (1H, dt, J ¼ 8.0, 9.5 Hz,
H-7), 6.56 (1H, d, J ¼ 9.5 Hz, H-6); dC (75 MHz, CDCl3): 12.9 (C-1),
13.3 (C-12),16.2 (C-11), 22.2 (C-2), 41.2 (C-3), 50.8 (C-8), 53.3 (C-10),
117.9 (C-7), 138.7 (C-6); HRMS (ES): m/z 146.0211 [M ꢀ PrSOH]þ,
C6H10S2 requires 146.0224; (E)-4h dH (400 MHz, CDCl3): 0.98 (3H, t,
J ¼ 7.3 Hz, H-1), 1.08 (3H, t, J ¼ 7.4 Hz, H-12), 1.69 (2H, sext,
J ¼ 7.3 Hz,, H-2), 1.81 (2H, m, H-11), 2.62 (4H, m, H-10), 2.69 (2H, t,
J ¼ 7.3 Hz, H-3), 3.47 (1H, ddd, J ¼ 1.0, 7.8, 13.0 Hz, H-8a), 3.53 (1H,
ddd, J ¼ 1.0, 7.8, 13.0 Hz, H-8b), 5.92 (1H, dt, J ¼ 7.8, 14.8 Hz, H-7),
6.38 (1H, d, J ¼ 14.8 Hz, H-6); dC (100 MHz, CDCl3): 13.0 (C-1), 13.4
(C-12), 16.2 (C-11), 22.4 (C-2), 40.4 (C-3), 53.0 (C-10), 54.6 (C-8),
116.6 (C-7), 134.7 (C-6); HRMS (ES): m/z 146.0211 [M ꢀ PrSOH]þ,
C6H10S2 requires 146.0224.
Obtained as a 2:1 Z:E mixture (64% and separable) as colourless
oils: IR nmax (CH2Cl2)/cmꢀ1 1047 (S]O), 499 (SeS); (Z)-4e dH
(300 MHz, CDCl3): 2.19 (2H, m, H-11), 2.75 (2H, m, H-10), 3.36 (2H,
dt, J ¼ 1.0, 7.5 Hz, H-3), 3.55 (1H, ddd, J ¼ 0.9, 8.0, 13.2 Hz, H-8a),
3.64 (1H, ddd, J ¼ 0.9, 8.0, 13.2 Hz, H-8b), 3.85 (2H, m, H-12), 5.17
(2H, m, H-1), 5.73 (1H, dt, J ¼ 8.0, 9.5 Hz, H-7), 5.82 (1H, m, H-2),
6.54 (1H, dt, J ¼ 0.9, 9.5 Hz, H-6), 7.73 (2H, m, Ar-H), 7.86 (2H, m,
Ar-H); dC (100 MHz, CDCl3): 22.3 (C-11), 36.9 (C-12), 42.1 (C-3),
48.9 (C-10), 51.1 (C-8), 118.0 (C-7), 119.2 (C-1), 123.4 (Ar-C), 132.0
(C-2), 132.6 (Arqu), 134.1 (Ar-C), 138.7 (C-6), 168.2 (C]O); HRMS
(ES): m/z 382.0583 [M þ H]þ, C17H20NO3S3 requires 382.0605; (E)-
4e dH (300 MHz, CDCl3): 2.18 (2H, m, H-11), 2.71 (2H, m, H-10),
3.33 (2H, d, J ¼ 7.3 Hz, H-3), 3.46 (1H, ddd, J ¼ 1.1, 7.9, 13.1 Hz, H-
8a), 3.54 (1H, ddd, J ¼ 1.1, 7.9, 13.1 Hz, H-8b), 3.86 (2H, m, H-12),
5.17 (2H, m, H-1), 5.78 (1H, dt, J ¼ 7.3, 9.9 Hz,, H-2), 5.90 (1H, dt,
J ¼ 7.9, 14.9 Hz, H-7), 6.35 (1H, dt, J ¼ 0.9, 14.9 Hz, H-6), 7.73 (2H,

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4.10.9. (E,Z)-2,3,7-Trithiadeca-4-ene 7-oxide (4i) [55]
JCF ¼ 9.5 Hz, Ar-CF), 131.1 (Ar-COMe), 132.4 (d, JCF ¼ 3.3 Hz, Ar-CF),
133.7 (C-4), 159.7 (Ar-COMe), 162.3 (d, JCF ¼ 245.1 Hz, Ar-CF); HRMS
(ES): m/z 383.0610 [M þ H]þ, C18H20FO2S3 requires 383.0609;
found: C, 56.40%; H, 5.11%; S, 25.25%. C18H19FO2S3 requires C,
56.68%; H, 5.01%; S, 25.14%.
Obtained as a 3:2 Z:E mixture (68%) as a colourless oil: IR nmax
(neat)/cmꢀ1 1035 (S]O), 499 (SeS); (Z)-4j dH (400 MHz, CDCl3): 1.03
(3H, t, J ¼ 7.4 Hz, H-10), 1.76 (2H, m, H-9), 2.41 (3H, s, H-1), 2.58 (2H,
m, H-8), 3.50 (1H, dd, J ¼ 7.9, 13.2 Hz, H-6a), 3.59 (1H, dd, J ¼ 7.9,
13.2 Hz, H-6b), 5.77 (1H, dt, J ¼ 7.9, 9.2 Hz, H-5), 6.55 (1H, d,
J ¼ 9.2 Hz, H-4); dC (100 MHz, CDCl3): 13.2 (C-10), 16.1 (C-9), 23.0
(C-1), 50.6 (C-6), 53.2 (C-8), 118.9 (C-5), 137.3 (C-4); HRMS (ES): m/z
117.9892 [M ꢀ PrSOH]þ, C4H6S2 requires 117.9911; (E)-4j dH
(400 MHz, CDCl3): 1.04 (3H, t, J ¼ 7.4 Hz, H-10), 1.76 (2H, m, H-9),
2.37 (3H, s, H-1), 2.65 (2H, m, H-8), 3.43 (1H, dd, J ¼ 7.7, 13.1 Hz, H-
6a), 3.52 (1H, dd, J ¼ 7.7, 13.1 Hz, H-6b), 5.91 (1H, dt, J ¼ 7.7, 15.0 Hz,
H-5), 6.34 (1H, d, J ¼ 15.0 Hz, H-4); dC (100 MHz, CDCl3): 13.2 (C-10),
16.0 (C-9), 21.9 (C-1), 53.0 (C-8), 54.4 (C-6), 116.9 (C-5), 132.9 (C-4);
HRMS (ES): m/z 117.9892 [M ꢀ PrSOH]þ, C4H6S2 requires 117.9911.
4.10.12. (E,Z)-1,8-(Bis-p-methoxyphenyl)-2,3,7-trithiaocta-4-ene 7-
oxide (E/Z-4l)
Obtained as a 2:3 Z:E mixture (78%) as a white solid which was
recrystallized from ethanol, mp 111e113 ꢁC: IR nmax (neat)/cmꢀ1
1035 (S]O), 499 (SeS); (Z)-E/Z-4l dH (400 MHz, CDCl3): 3.41 (1H,
ddd, J ¼ 1.0, 7.8, 13.4 Hz, H-6a), 3.51 (1H, ddd, J ¼ 1.0, 7.8, 13.4 Hz, H-
6b), 3.82 (3H, s, OCH3), 3.83 (3H, s, OCH3), 3.89 (1H, s, H-8a), 3.90
(1H, s, H-8b), 3.93 (2H, s, H-1), 5.69 (1H, dt, J ¼ 7.8, 9.6 Hz, H-5), 6.30
(1H, dt, J ¼ 1.0, 9.6 Hz, H-4), 6.86 (2H, d, J ¼ 8.8 Hz, Hm), 6.92 (2H, d,
J ¼ 8.8 Hz, Hm), 7.23 (4H, m, Ho); dC (100 MHz, CDCl3): 43.1 (C-1),
49.5 (C-6), 55.2 (OCH3), 55.3 (OCH3), 56.8 (C-8), 114.0 (Cm), 114.5
(Cm), 118.1 (C-5), 121.7 (Ci), 128.6 (Ci), 130.6 (Co), 131.3 (Co), 138.0 (C-
4), 159.2 (Cp), 159.8 (Cp); HRMS (ES): m/z 395.0808 [M þ H]þ,
C19H23O3S3 requires 395.0809; found: C, 57.76%; H, 5.60%; S,
24.21%; C19H22O3S3 requires C, 57.84%; H, 5.62%; S, 24.38%; (E)-E/Z-
4l IR nmax (neat)/cmꢀ1 1611 (C]C), 1035 (S]O), 499 (SeS); dH
(400 MHz, CDCl3): 3.24 (1H, ddd, J ¼ 1.0, 7.8, 13.2 Hz, H-6a), 3.38
(1H, ddd, J ¼ 1.0, 7.8, 13.2 Hz, H-6b), 3.79 (3H, s, OCH3), 3.84 (3H, s,
OCH3), 3.91 (1H, s, H-8a), 3.92 (1H, s, H-8b), 3.92 (2H, s, H-1), 5.86
(1H, dt, J ¼ 7.8, 14.8 Hz, H-5), 6.19 (1H, dt, J ¼ 1.0, 14.8 Hz, H-4), 6.86
(2H, d, J ¼ 8.8 Hz, Hm), 6.93 (2H, d, J ¼ 8.8 Hz, Hm), 7.23 (4H, m, Ho);
dC (100 MHz, CDCl3): 42.3 (C-1), 52.7 (C-6), 55.2 (OCH3), 55.3
(OCH3), 56.1 (C-8), 114.0 (Cm), 114.5 (Cm), 117.0 (C-5), 121.5 (Ci), 128.6
(Ci), 130.6 (Co), 131.2 (Co), 134.2 (C-4), 159.2 (Cp), 159.8 (Cp); HRMS
(ES): m/z 395.0808 [M þ H]þ, C19H23O3S3 requires 395.0809; found:
C, 57.76%; H, 5.60%; S, 24.21%. C19H22O3S3 requires C, 57.84%; H,
5.62%; S, 24.38%.
4.10.10. (E,Z)-8-(p-Methoxyphenyl)-1-phenyl-2,3,7-trithiaocta-4-
ene 7-oxide (4j)
Obtained as a 3:2 Z:E mixture (71%) as a white solid which was
recrystallized from ethanol, mp 76e78 ꢁC: IR nmax (neat)/cmꢀ1 1034
(S]O), 499 (SeS); (Z)-4j dH (400 MHz, CDCl3): 3.37 (1H, ddd, J ¼ 1.1,
7.8, 13.6 Hz, H-6a;), 3.47 (1H, ddd, J ¼ 1.1, 7.8, 13.6 Hz, H-6b), 3.80
(3H, s, OCH3), 3.88 (1H, s, H-8a), 3.88 (1H, s, H-8b), 3.94 (2H, s, H-1),
5.64 (1H, dt, J ¼ 7.8, 9.6 Hz, H-5), 6.25 (1H, dt, J ¼ 1.1, 9.6 Hz, H-4),
6.90 (2H, m, Ar-H), 7.20 (2H, m, Ar-H), 7.29 (4H, m, Ar-H); dC
(100 MHz, CDCl3): 43.5 (C-1), 49.5 (C-6), 55.3 (OCH3), 56.8 (C-8),
114.4 (Ar-COMe),118.2 (C-5),121.6 (Ar-COMe),127.6 (Ar-CH),128.6 (Ar-
CH), 129.4 (Ar-CH), 131.2 (Ar-COMe), 136.6 (Ar-CH), 137.8 (C-4), 159.8
(Ar-COMe); HRMS (ES): m/z 365.0714 [M þ H]þ, C18H21O2S3 requires
365.0704; found: C, 59.01%; H, 5.48%; S, 26.49%. C18H20O2S3
requires C, 59.31%; H, 5.53%; S, 26.38%; (E)-4j dH (400 MHz, CDCl3):
3.20 (1H, ddd, J ¼ 1.1, 7.7, 13.4 Hz, H-6a), 3.34 (1H, ddd, J ¼ 1.1, 7.7,
13.4 Hz, H-6b), 3.81 (3H, s, OCH3), 3.86 (1H, s, H-8a), 3.87 (1H, s, H-
8b), 3.93 (2H, s, H-1), 5.82 (1H, dt, J ¼ 7.7, 14.8 Hz, H-5), 6.14 (1H, dt,
J ¼ 1.1, 14.8 Hz, H-4), 6.90 (2H, m, Ar-H), 7.20 (2H, m, Ar-H), 7.29 (4H,
m, Ar-H); dC (100 MHz, CDCl3): 42.8 (C-1), 52.7 (C-6), 55.3 (OCH3),
56.2 (C-8), 114.4 (Ar-COMe), 117.2 (C-5), 121.5 (Ar-COMe), 127.6 (Ar-
CH), 128.6 (Ar-CH), 129.4 (Ar-CH), 131.2 (Ar-COMe), 134.1 (C-4), 136.6
(Ar-CH), 159.8 (Ar-COMe); HRMS (ES): m/z 365.0714 [M þ H]þ,
C18H21O2S3 requires 365.0704; found: C, 59.01%; H, 5.48%; S,
26.49%; C18H20O2S3 requires C, 59.31%; H, 5.53%; S, 26.38%.
4.11. Synthesis of sulfenylating agents p-TsSR2
To a solution of potassium p-toluenethiosulfonate (1.3 equiv)
dissolved in DMF (1 M) was added R2X (X ¼ Cl or Br; 1.0 equiv) neat
or in solution (DMF) slowly via syringe. The reaction was stirred at
room temperature for 3 h or heated according to conversion of R2X
by TLC before being quenched with saturated aqueous NaHCO3. The
resulting mixture was extracted with dichloromethane (3 times)
and the combined organic extracts were dried over MgSO4. The
solvent was removed under reduced pressure and the residue
purified on a silica-gel column using hexane/ethyl acetate mixtures
to afford the desired compound.
4.10.11. (E,Z)-1-(p-Fluorophenyl)-8-(p-methoxyphenyl)-2,3,7-
trithiaocta-4-ene 7-oxide (4k)
Obtained as a 4:3 Z:E mixture (74%) as a white solid which was
recrystallized from ethanol, mp 89e91 ꢁC: IR nmax (neat)/cmꢀ1 1035
(S]O), 497 (SeS); (Z)-4k dH (300 MHz, CDCl3): 3.37 (1H, ddd, J ¼ 1.0,
7.8, 13.4 Hz, H-6a), 3.46 (1H, ddd, J ¼ 1.0, 7.8, 13.4 Hz, H-6b), 3.79
(3H, s, OCH3), 3.88 (2H, s, H-8), 3.88 (2H, s, H-1), 5.65 (1H, dt, J ¼ 7.8,
9.3 Hz, H-5), 6.25 (1H, dt, J ¼ 1.0, 9.3 Hz, H-4), 6.89 (2H, m, Ar-H),
6.98 (2H, m, Ar-H), 7.20 (2H, m, Ar-H), 7.26 (2H, m, Ar-H); dC
(75 MHz, CDCl3): 42.5 (C-1), 49.5 (C-6), 55.2 (OCH3), 56.8 (C-8),
114.4 (Ar-COMe), 115.4 (d, JCF ¼ 22.6 Hz, Ar-CF), 118.5 (C-5), 121.5 (Ar-
COMe), 131.0 (d, JCF ¼ 9.5 Hz, Ar-CF), 131.2 (Ar-COMe), 132.4 (d,
JCF ¼ 3.3 Hz, Ar-CF), 137.6 (C-4), 159.7 (Ar-COMe), 162.3 (d,
JCF ¼ 245.1 Hz, Ar-CF); HRMS (ES): m/z 383.0610 [M þ H]þ,
C18H20FO2S3 requires 383.0609; found: C, 56.40%; H, 5.11%; S,
25.25%. C18H19FO2S3 requires C, 56.68%; H, 5.01%; S, 25.14%; (E)-4k
dH (300 MHz, CDCl3): 3.19 (1H, ddd, J ¼ 1.1, 7.7, 13.2 Hz, H-6a), 3.34
(1H, ddd, J ¼ 1.1, 7.7, 13.2 Hz, H-6b), 3.80 (3H, s, OCH3), 3.88 (2H, s,
H-1), 3.89 (2H, s, H-8), 5.83 (1H, dt, J ¼ 7.7, 14.9 Hz, H-5), 6.14 (1H,
dt, J ¼ 1.1, 14.9 Hz, H-4), 6.89 (2H, m, Ar-H), 6.98 (2H, m, Ar-H), 7.20
(2H, m, Ar-H), 7.26 (2H, m, Ar-H); dC (75 MHz, CDCl3): 41.7 (C-1),
52.6 (C-6), 55.2 (OCH3), 56.3 (C-8), 114.4 (Ar-COMe), 115.4 (d,
JCF ¼ 22.6 Hz, Ar-CF), 117.5 (C-5), 121.4 (Ar-COMe), 131.0 (d,
4.11.1. Benzenesulfonothioic acid, S-2-propen-1-yl ester [56]
Obtained as a clear oil (96%): dH (300 MHz, CDCl3): 2.44 (3H, s,
CH3), 3.66 (2H, dd, J ¼ 1.0, 7.2 Hz, H-10), 5.08 (1H, d, J ¼ 10.5 Hz, H-
30), 5.19 (1H, d, J ¼ 17.1 Hz, H-30) 5.70 (1H, m, H-20), 7.33 (2H, d,
J ¼ 7.8 Hz, Hm), 7.79 (2H, d, J ¼ 7.8 Hz, Ho).
4.11.2. Benzenesulfonothioic acid, S-propyl ester [56]
Obtained as a pale-yellow oil (96%): dH (400 MHz, CDCl3): 0.93
(3H, t, J ¼ 7.3 Hz, H-30), 1.64 (2H, s, J ¼ 7.3 Hz, H-20), 2.45 (3H, s, CH3),
2.97 (2H, t, J ¼ 7.3 Hz, H-10), 7.34 (2H, d, J ¼ 8.0 Hz, Hm), 7.81 (2H, d,
J ¼ 8.0 Hz, Ho); dC (100 MHz, CDCl3): 13.1 (C-30), 21.5 (C-20), 22.1
(CH3), 37.9 (C-10), 126.9 (Co), 129.7 (Cm), 142.1 (Ci), 144.6 (Cp).
4.11.3. Benzenesulfonothioic acid, S-methyl ester [57]
Obtained as a colourless oil (96%): dH (300 MHz, CDCl3): 2.43
(3H, s, CH3), 2.48 (3H, s, CH3), 7.34 (2H, d, J ¼ 8.1 Hz, Hm), 7.78 (2H, d,
J ¼ 8.1 Hz, Ho); dC (100 MHz, CDCl3): 17.9 (CH3), 21.5 (CH3), 127.0
(Co), 129.8 (Cm), 140.8 (Ci), 144.8 (Cp).

C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
253
4.11.4. Benzenesulfonothioic acid, S-[(4-fluorophenyl)methyl] ester
[58]
Obtained as a white solid (78%): dH (400 MHz, CDCl3): 2.43 (3H,
s, CH3), 4.23 (2H, s, CH2), 6.91 (2H, m, H-30), 7.15 (2H, m, H-20), 7.27
(2H, d, J ¼ 8.6 Hz, Hm), 7.71 (2H, d, J ¼ 8.6 Hz, Ho).
(2H, s, H-1), 5.70 (1H, dt, J ¼ 7.8, 9.6 Hz, H-4), 6.42 (1H, d,
J ¼ 9.6 Hz, H-3), 6.83 (2H, d, J ¼ 8.7 Hz, Hm), 6.87 (2H, d, J ¼ 8.7 Hz,
Hm), 7.18 (2H, d, J ¼ 8.7 Hz, Ho), 7.21 (2H, d, J ¼ 8.7 Hz, Ho); dC
(75 MHz, CDCl3): 37.6 (C-1), 50.4 (C-5), 55.2 (OCH3), 56.6 (C-7),
114.1 (Cm), 114.3 (Cm), 116.2 (C-4), 122.0 (C-3), 129.3 (Ci), 129.9 (Co),
131.3 (Co), 132.9 (Ci), 158.9 (Cp), 159.6 (Cp); HRMS (ES): m/z
363.1082 (M þ H)þ, C19H23O3S2 requires 363.1089; found: C,
62.79%; H, 6.09%; S, 17.50%. C19H22O3S2 requires C, 62.95%; H,
6.12%, S; 17.69%; (E)-6 (0.035 g, 32%) was obtained as a white solid
which was recrystallized from ethanol, mp 90e92 ꢁC; IR nmax
(CH2Cl2)/cmꢀ1 1033 (S]O), 690 (CeS); dH (300 MHz, CDCl3): 3.24
(1H, dd, J ¼ 7.6, 13.2 Hz, H-5a), 3.38 (1H, dd, J ¼ 7.6, 13.2 Hz, H-5b),
3.76 (3H, s, OCH3), 3.80 (5H, m, OCH3 and H-7), 3.90 (2H, s, H-1),
5.56 (1H, dt, J ¼ 7.6, 15.3 Hz, H-4), 6.29 (1H, d, J ¼ 15.3 Hz, H-3),
6.85 (2H, d, J ¼ 8.6 Hz, Hm), 6.88 (2H, d, J ¼ 8.6 Hz, Hm), 7.13 (2H, t,
J ¼ 8.6 Hz, Ho), 7.26 (2H, t, J ¼ 8.6 Hz, Ho); dC (75 MHz, CDCl3): 36.2
(C-1), 53.6 (C-5), 55.2 (OCH3), 55.7 (C-7), 114.1 (Cm), 114.2 (C-4),
114.4 (Cm), 121.7 (C-3), 128.6 (Ci), 129.9 (Co), 131.2 (Co), 132.9 (Ci),
158.9 (Cp), 159.7 (Cp); HRMS (ES): m/z 363.1082 (M þ H)þ,
C19H23O3S2 requires 363.1089; found: C, 62.61%; H, 5.97%; S,
17.32%. C19H22O3S2 requires C, 62.95%; H, 6.12%; S, 17.69%.
4.11.5. Benzenesulfonothioic acid, S-[(4-methoxyphenyl)methyl]
ester [59]
Obtained as a white solid (74%): dH (400 MHz, CDCl3): 2.44 (3H,
s, CH3), 3.77 (3H, s, OCH3), 4.21 (2H, s, CH2), 6.76 (2H, d, J ¼ 8.6 Hz,
H-30), 7.10 (2H, d, J ¼ 8.6 Hz, H-20), 7.28 (2H, d, J ¼ 8.6 Hz, Hm), 7.74
(2H, d, J ¼ 8.6 Hz, Ho).
4.11.6. Benzenesulfonothioic acid, S-benzyl ester [60]
Obtained as a white solid (95%): dH (400 MHz, CDCl3): 2.43 (3H,
s, CH3), 4.25 (2H, s, CH2), 7.20 (5H, m, Ph), 7.27 (2H, d, J ¼ 8.6 Hz,
Hm), 7.73 (2H, d, J ¼ 8.6 Hz, Ho).
4.12. Synthesis of derivatives of E/Z-4l
4.12.1. (E,Z)-1,8-(Bis-p-methoxyphenyl)-2,6-dithiahepta-3-ene (5) [61]
A solution of S-3-(p-methoxyphenylthio)-prop-1enyl ethane-
thioate 2g (295 mg,1.1 mmol) in methanol (2 mL) was slowly added
at 0 ꢁC to a degassed solution of KOH (123 mg, 2.2 mmol) in
methanol (3 mL). The resulting mixture was stirred at 0 ꢁC under
a nitrogen atmosphere for 10 min. p-Methoxybenzyl chloride
(157 mg, 1.0 mmol) in methanol (1 mL) was then added and the
mixture was stirred for 2 h at room temperature. The solvent was
removed under reduced pressure and the resultant residue was
added to water (20 mL) and the solution acidified with hydrochloric
acid. The compound was extracted with dichloromethane
(3 ꢃ 20 mL), dried with MgSO4 and the solvent removed by rotary
evaporation. The crude product was purified by silica-gel column
chromatography using petroleum ether/ethyl acetate mixtures to
afford a 3:2 Z:E mixture of compound 5 (0.277 g, 80%) as a clear oil:
IR nmax (CH2Cl2)/cmꢀ1 690 (CeS); (Z)-5 dH (400 MHz, CDCl3): 3.08
(2H, d, J ¼ 7.2 Hz, H-5), 3.48 (2H, s, H-7), 3.66 (3H, s, OCH3), 3.69 (3H,
s, OCH3), 3.73 (2H, s, H-1), 5.50 (1H, m, H-4), 5.99 (1H, d, J ¼ 9.2 Hz,
H-3), 6.74 (4H, m, Hm), 7.11 (4H, m, Ho); dC (100 MHz, CDCl3): 30.0
(C-5), 35.1 (C-7), 37.6 (C-1), 55.2 (OCH3 ꢃ 2),114.0 (Cm ꢃ 2),126.1 (C-
4), 127.0 (C-3), 129.2 (Ci),130.0 (Co ꢃ 2), 130.4 (Ci), 158.6 (Cp), 158.8
(Cp); (E)-5 dH (400 MHz, CDCl3): 2.92 (2H, d, J ¼ 7.2 Hz, H-5), 3.43
(2H, s, H-7), 3.67 (3H, s, OCH3), 3.69 (3H, s, OCH3),3.75 (2H, s, H-1),
5.50 (1H, m, H-4), 5.89 (1H, d, J ¼ 15.6 Hz, H-3), 6.74 (4H, m, Hm),
7.11 (4H, m, Ho); dC (100 MHz, CDCl3): 33.5 (C-5), 34.3 (C-7), 36.5 (C-
1), 55.2 (OCH3 ꢃ 2), 113.8 (Cm ꢃ 2), 125.2 (C-4), 126.1 (C-3), 129.2
(Ci), 129.9 (Co ꢃ 2), 130.4 (Ci), 158.6 (Cp), 158.8 (Cp); HRMS (ES): m/z
347.1174 (M þ H)þ, C19H23O2S2 requires 347.1139.
4.12.3. 1,8-(Bis-p-methoxyphenyl)-2,3,7-trithiaoctane (7) [62]
A solution of p-methoxybenzyl chloride (157 mg, 1.00 mmol) in
ethanol (2 mL) was added dropwise at room temperature to a solu-
tion of 1,3-propanedithiol (119 mg, 1.10 mmol) and KOH (112 mg,
2.00 mmol) in ethanol (3 mL) under a N2 atmosphere. The mixture
was allowed to stir for 3 h after which a solution of S-p-methox-
ybenzyl p-toluenesulfonylthioate (339 mg, 1.1 mmol) in ethanol
(2 mL) was added. The resulting mixture was stirred at room
temperature for 3 h. The reaction mixture was added to water
(20 mL) and extracted with dichloromethane (3 ꢃ 20 mL). The
combined organic fractions were then washed with water (30 mL),
brine (20 mL) and aqueous KOH (20 mL) and dried over MgSO4. The
solvent was removed under reduced pressure and the crude residue
was purified by silica-gel column chromatography using petroleum
ether/ethyl acetate mixtures to afford compound 7 (0.323 g, 85%) as
a white solid which was recrystallized from ethanol, mp 41e43 ꢁC; IR
nmax (CH2Cl2)/cmꢀ1 516 (SeS); dH (400 MHz, CDCl3): 1.86 (2H, quint,
J ¼ 7.1 Hz, H-5), 2.46 (2H, t, J ¼ 7.1 Hz, H-6), 2.51 (2H, t, J ¼ 7.1 Hz, H-
4), 3.68 (2H, s, H-8), 3.83 (6H, s, OCH3), 3.88 (2H, s, H-1), 6.88 (4H, d,
J ¼ 8.4 Hz, Hm), 7.26 (4H, m, Ho); dC (100 MHz, CDCl3): 28.2 (C-5), 29.6
(C-6), 35.5 (C-8), 37.0 (C-4), 43.0 (C-1), 55.2 (OCH3 ꢃ 2), 113.8 (Cm),
113.9 (Cm), 129.3 (Ci), 129.8 (Co), 130.2 (Ci), 130.3 (Co), 158.5 (Cp), 159.0
(Cp); HRMS (ES): m/z 419.0794 (M þ Na þ O)þ, C19H24NaO3S3
requires 419.0785; found: C, 59.95%; H, 6.33%; S, 25.30%. C19H24O2S3
requires C, 59.96%; H, 6.36%; S, 25.27%.
4.12.4. 1,8-(Bis-p-methoxyphenyl)-2,3,7-trithiaoctane 7-oxide (8)
4.12.2. (E,Z)-1,7-(Bis-p-methoxyphenyl)-2,6-dithiahepta-3-ene 6-
To a
0 ꢁC solution of 1,8-(bis-p-methoxyphenyl)-2,3,7-tri-
oxide (6)
thiaoctane (75 mg, 0.20 mmol) in dichloromethane (2 mL) was
added solid m-chloroperoxybenzoic acid (32 mg, 0.14 mmol). The
solution was stirred for 1 h, after which aqueous NaHCO3 (8 mL)
was added and the compound extracted with dichloromethane
(3 ꢃ15 mL). The combined organic extracts were dried with MgSO4
and the solvent removed under reduced pressure. The crude
residue was purified by silica-gel chromatography using petroleum
ether/ethyl acetate mixtures to afford compound 8 (0.050 g, 89%) as
a white solid: recrystallized from ethanol/CH2Cl2, mp 108e110 ꢁC;
IR nmax (CH2Cl2)/cmꢀ1 1033 (S]O), 517 (SeS); dH (400 MHz, CDCl3):
2.06 (2H, m, H-5), 2.50 (2H, t, J ¼ 6.8 Hz, H-4), 2.57 (1H, m, H-6a),
2.65 (1H, m, H-6b), 3.81 (3H, s, OCH3), 3.82 (3H, s, OCH3), 3.86 (2H,
s, H-1), 3.92 (1H, d, J ¼ 13.2 Hz, H-8a), 3.96 (1H, d, J ¼ 13.2 Hz, H-8b),
6.87 (2H, d, J ¼ 8.0 Hz, Hm), 6.92 (2H, d, J ¼ 8.0 Hz, Hm), 7.22 (2H, d,
J ¼ 8.0 Hz, Ho), 7.23 (2H, d, J ¼ 8.0 Hz, Ho); dC (100 MHz, CDCl3): 21.8
To
a
0 ꢁC solution of (E,Z)-1,8-(bis-p-methoxyphenyl)-2,6-
dithiahepta-3-ene (115 mg, 0.33 mmol) in dichloromethane
(3 mL) was added solid m-chloroperoxybenzoic acid (67 mg,
0.30 mmol). The solution was stirred for 1 h, after which aqueous
NaHCO3 (8 mL) was added and the compound extracted with
dichloromethane (3 ꢃ 15 mL). The combined organic extracts were
dried with MgSO4 and the solvent removed under reduced pres-
sure. The crude residue was purified by silica-gel chromatography
using petroleum ether/ethyl acetate mixtures to afford compound
(Z/E)-6 as separate isomers. (Z)-6 (0.054 g, 50%) was obtained as
a white solid which was recrystallized from ethanol, mp 78e80 ꢁC;
IR nmax (CH2Cl2)/cmꢀ1 1033 (S]O), 690 (CeS); dH (300 MHz, CDCl3):
3.41 (1H, dd, J ¼ 7.8, 13.2 Hz, H-5a), 3.52 (1H, dd, J ¼ 7.8, 13.2 Hz, H-
5b), 3.75 (3H, s, OCH3), 3.79 (3H, s, OCH3), 3.81 (2H, m, H-7), 3.87

254
C.H. Kaschula et al. / European Journal of Medicinal Chemistry 50 (2012) 236e254
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(C-5), 36.9 (C-4), 42.9 (C-1), 48.9 (C-6), 55.2 (OCH3 ꢃ 2), 57.6 (C-8),
114.0 (Cm), 114.4 (Cm), 121.5 (Ci), 129.2 (Ci), 130.4 (Co), 131.1 (Co),
159.1 (Cp), 159.7 (Cp); HRMS (ES): m/z 419.0737 (M þ Na)þ,
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24.30%. C19H24O3S3 requires C, 57.54%; H, 6.10%; S, 24.25%.
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added N-Boc- -cysteine (55 mg, 0.22 mmol) dissolved in THF
(0.5 mL) at room temperature under a nitrogen atmosphere. The
solution was heated to reflux (70 ꢁC) for 25 h. The mixture was
evaporated under reduced pressure and the residue purified by
silica-gel column chromatography to afford recovered starting
L
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L
(96 mg, 55% recovered) as well as product 9 (36 mg, 52% based on
a 78% conversion of the cysteine) as a clear oil: IR nmax (CHCl3)/cmꢀ1
3364 (NH), 1716 (C]O), 1634 (C]O), 497 (SeS); dH (400 MHz, CDCl3):
1.25 (3H, t, J ¼ 7.1 Hz, H-3b), 1.42 (9H, s, -C(CH3)3), 2.85 (2H, m, H-1),
3.77 (3H, s, OCH3), 3.84 (2H, s, Bn), 4.16 (2H, qu, J ¼ 7.1 Hz, H-3a),
4.46 (1H, m, H-2), 5.24 (1H, br. s., NH), 6.82 (2H, d, J ¼ 8.8 Hz, Hm),
7.21 (2H, d, J ¼ 8.8 Hz, Ho); dC (100 MHz, CDCl3): 14.1 (C-3b), 28.3
(eC(CH3)3), 40.9 (C-1), 43.0 (Bn), 53.0 (C-2), 55.2 (OCH3), 61.7 (C-
3a), 80.1 (eC(CH3)3), 114.0 (Cm), 128.9 (Ci), 130.5 (Co), 155.0 (C]O),
159.2 (Cp), 170.7 (C-3); HRMS (ES): m/z 424.1244 (M þ Na)þ,
C18H27NNaO5S2 requires 424.1228.
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Acknowledgements
This work was supported by the South African Research Chairs
Initiative (SARCHI) of the Department of Science and Technology,
the National Research Foundation (NRF), and research grants from
the Medical Research Council (MRC) of South Africa and the
University of Cape Town. We thank Roche for the loan of the RTCA
SP xCelligence system. We thank Mr Ronnie Dreyer for his assis-
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