Reaction of lupane and oleanane triterpenoids with Lawesson's reagent

Article abstract of DOI:10.1016/j.tet.2008.02.023

The reactions of selected triterpenic oxo compounds with Lawesson's reagent were investigated. We examined sulfurization of some oxygen compounds and for these reactions several hindered ketones, one aldehyde, α-hydroxyketones, esters, or anhydrides were

Full text of DOI:10.1016/j.tet.2008.02.023

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Tetrahedron 64 (2008) 3736e3743
www.elsevier.com/locate/tet
Reaction of lupane and oleanane triterpenoids with Lawesson’s reagent
Miroslav Kvasnica a,b, Iva Rudovska b, Ivana Cisarova c, Jan Sarek b,
*
a Institute of Organic Chemistry and Biochemistry, Academy of Sciences, Flemingovo n. 2, 16610 Prague 6, Czech Republic
b Department of Organic and Nuclear Chemistry, Faculty of Science, Charles University in Prague, Hlavova 8, 12843 Prague 2, Czech Republic
c Department of Inorganic Chemistry, Faculty of Science, Charles University in Prague, Hlavova 8, 12843 Prague 2, Czech Republic
Received 20 November 2007; received in revised form 21 January 2008; accepted 7 February 2008
Available online 9 February 2008
Abstract
The reactions of selected triterpenic oxo compounds with Lawesson’s reagent were investigated. We examined sulfurization of some oxygen
compounds and for these reactions several hindered ketones, one aldehyde, a-hydroxyketones, esters, or anhydrides were chosen. We synthe-
sized 15 new sulfur derivatives, including thioketone 16, dimeric sulfides 17e19, and thiaderivatives 20e22. We also observed unusual trans-
formations, which afforded oxathiaphosphinines 23a, 23b, and dithiaphospholanes 24. The prepared compounds failed to demonstrate any
significant cytotoxic activity.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: Terpenoids; Lawesson’s reagent; Ketones; Sulfur
1. Introduction
antitumor activity of several triterpenic derivatives has been
found. This fact has encouraged us to prepare newly modified
skeleton of betulinines. Thionation is one possible modifica-
tion that could have some influence on antitumor activity in
spite of lipophilicity that is an inherent feature of thionation.
Herein we report on the reaction of several lupane and 18a-
oleanane carbonyl derivatives with Lawesson’s reagent to
form new sulfur analogues. In addition, we made a cytotoxic
screening of prepared compounds. Only several lupane and
18a-oleanane triterpenoids with CeS bonds have previously
been published.19
Lawesson’s reagent1 is a widely used thionation reagents.2
It is used especially for thionation of carbonyl compounds (ke-
tones, esters, amides, anhydrides, etc.)3e7 and alcohols.8,9 Un-
like other thionation reagents, e.g., phosphorus pentasulfide,10
reactions with Lawesson’s reagent are useful because of their
high yields, convenient handling, and especially soft thiona-
tion reactions, e.g., in the case of isoprenoids.11,12 In some
cases, the main disadvantage of this reagent is the formation
of by-products or stable heterocyclic intermediates.12
2. Results and discussion
As starting materials for preparation of triterpenoids used
for thionation reactions we used betulin (1) easily accessible
from the birch bark (Betula pendula) by extraction with etha-
nol14 and betulinic acid (2) from the bark of the plane tree
(Platanus hispanica) isolated by MeOH extraction.18 Accord-
ing to known procedures starting with betulin (1), we synthe-
sized unsaturated ketone 3,20 ketone 4,21 heptanorketone 6,14
seco diketones 7 and 8,20 seco diketone 9,22 hydroxyketones
11 and 12,23 ketoacid 13, anhydride 14, and b-ketoester
Triterpenoids are an important group of natural compounds
possessing a variety of biological activities.13 Recently, we
have examined the structureeactivity relationships in lupane
and 18a-oleanane derivatives (betulinines).14e18 Significant
* Corresponding author. Tel./fax: þ420 221 951 332.
E-mail address: jan.sarek@gmail.com (J. Sarek).
0040-4020/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.02.023

M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
3737
15.14 From betulinic acid (2) we prepared benzyl-ester 524 and
unsaturated aldehyde 10.25
yields (approximately 80%). The mechanism of this reaction
is based on dimerization of saturated thioketones. Dimeriza-
tion of these thioketones is enabled by ready formation of their
enol form. This was explained previously.12
Similarly, reactions of seco diketones 7e9 resulted in un-
saturated cyclic sulfides 20e22 (Scheme 3). Although the
mechanism is the same as for saturated ketones 4e6, no dimer
products were observed. Similar reactions are used for the
preparation of substituted thiophenes.26,27 No lactone oxygen
was replaced in the case of diketone 8.
The reaction procedure for thionation was identical for all
derivatives and was carried out in refluxing toluene with equi-
molar amount of Lawesson’s reagent. Only in the case of low
reactivity, toluene was replaced by o-xylene.
Reaction of unsaturated ketone 3 afforded only expected
thioketone 16 (Scheme 1). However, 12 h reflux in o-xylene
led to full conversion and due to the formation of by-products,
the reaction had to be quenched earlier.
Scheme 3. Reagents and conditions: (a) LR, toluene, reflux, 5 h.
Scheme 1. Reagents and conditions: (a) LR, o-xylene, reflux, 12 h.
In previous examples, we only used ketones with no other
functional groups nearby. If there is another functional group
(e.g., double bond, hydroxyl group) we can expect some un-
typical products. This was proved for unsaturated aldehyde
10 and hydroxyketones 11 and 12.
Opposite to ketone 3, reaction of saturated ketones 4e6
gave unexpected dimer sulfides 17e19 (Scheme 2) in good
Reaction of aldehyde 10 with Lawesson’s reagent afforded
only two isomeric oxathiaphosphinines 23a and 23b (Scheme
4). The mechanism of formation of these products is as fol-
lows. Firstly, the phosphorus atom of the ylide attacks the car-
bonyl oxygen to create a PeO bond followed by migration of
double bond. The last step is closing of the cycle by formation
Scheme 4. Reagents and conditions: (a) LR, toluene, reflux, 4 h.
Scheme 2. Reagents and conditions: (a) LR, toluene, reflux, 5 h.

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M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
of a CeS bond (Scheme 5). The absolute configuration on the
phosphorus atom was determined by X-ray analysis of 23b,
which has configuration S (Fig. 1).
On the other hand, the reaction of hydroxyketones 11 and
12 resulted in complicated mixtures of products. Using
HPLC we were able to isolate only one major product from
each reaction. We obtained dithiaphospholane 24 from hy-
droxyketone 11, and dioxaphospholane 25 from hydroxyke-
tone 12 (Scheme 6). The reason why we isolated just these
two compounds could be explained by the different retention
times of these two derivatives in contrast to the other ones
in mixture.
Reaction of ketoacid 13 did not lead to sulfur containing
derivative but only to diene 26 (Scheme 7). It was found
that decarboxylation and deoxygenation with migration of
double bonds occurred during reaction with Lawesson’s re-
agent. The reaction was very fast. It took only about 2 h and
the yield was approximately 60%.
While in previous cases we obtained derivatives with a mod-
ified triterpenic skeleton, the reactions of anhydride 14 and
b-ketoester 15 afforded only products with modified acetate
groups (Scheme 7). The attempt to prepare thioanhydride re-
sulted only in 3b-thioacetate 27 in low yield (about 45%).
Starting anhydride 14 composed the rest of the reaction mix-
ture. Prolongation of reaction time to 50 h does not lead to
Scheme 6. Reagents and conditions: (a) LR, toluene, reflux, 5 h.
higher yield. In the case of b-ketoester 15, we obtained a
mixture of three compounds identified as bis(thioacetate)
28a, 28-thioacetate 28b, and 3b-thioacetate 28c after 15 h.
No substitution in other positions was observed in both cases.
Long reaction times (>10 h) made it possible to modify acetyl
groups whereas the 4 h reaction of aldehyde 10 did not af-
forded any thioacetate. The combination of thioacetates of
b-ketoester 15 is probably caused by the low steric hindrance
of the 28-acetyl group in contrast to anhydride 14.
Scheme 5. Probably mechanism of formation of oxathiaphosphinines 23a and 23b.
Figure 1. ORTEP drawing of oxathiaphosphinine 23b with atom numbering scheme. Thermal ellipsoids are drawn at the 50% probability level.

M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
3739
measured on ZAB-EQ (VG Analytical, Ltd., Manchester).
IR spectra were recorded on Nicolet Avatar 370 FT-IR spec-
trometer, using CHCl3 as a solvent. TLC was performed on
Kieselgel 60 F254 (Merck) sheets; detected by UV fluorescence
and spraying 10% sulfuric acid with heating to 110e200 ꢀC.
Used HPLC system consisted of High Pressure Pump Gilson
(model 361), Inject Valve Rheodyne, Preparative Column
(25ꢁ250 mm) with silica gel filling (Biospher 7 mm; Labio),
´ ˇ´
DifferentialeRefractometrical Detector (Laboratornı prıstroje,
Praha, CR) connected with PC (software Chromulan) and
Automatic Fraction Collector Gilson (model 246). Lawesson’s
reagent was purchased from SigmaeAldrich.
4.2. General procedure for reaction of triterpenic carbonyl
compounds with Lawesson’s reagent
Lawesson’s reagent (1 mmol) was added to a solution of tri-
terpenic ketone (1 mmol) in toluene or o-xylene (8 mL). The
reaction mixture was refluxed for several hours (mentioned
in each experiment). After that, the mixture was filtered
through silica gel in toluene, solvent was evaporated under re-
duced pressure, and crude product was chromatographed using
HPLC (mobile phase hexaneeethyl acetate, ratio mentioned in
each experiment).
Scheme 7. Reagents and conditions: (a) LR, toluene, reflux, 2 h; (b) LR,
o-xylene, reflux, 50 h (for 14) or 15 h (for 15).
3. Conclusion
In this work we showed that Lawesson’s reagent is suitable
for the preparation of different sulfur derivatives of triterpe-
noids. Triterpenic ketones are the best starting derivatives.
The limitations of this thionation are other functional groups
nearby. Nevertheless, it could be a possible way to prepare tri-
terpenic phosphorus heterocycles. Stable carbonyl derivatives
did not undergo transformation with Lawesson’s reaction;
only acetates could be modified under this condition. Synthetic
results of this work could be useful for other chemists working
on the field of isoprenoid chemistry. It could be used not only
for isoprenoid chemistry (reaction with a-hydroxyketones,
a,b-unsaturated aldehydes or ketones, seco diketones, etc.).
The preparation of new sulfur and/or phosphorus containing
heterocycles is interesting for broader organic synthesis.
All prepared sulfur compounds 16e22, 23a, 23b, 24, 25,
27, 28aec, and diene 26 were characterized by spectroscopic
methods and tested for in vitro antitumor activity on CEM leu-
kemia cells. Unfortunately, the prepared triterpenoids failed to
demonstrate any antitumor activity.
4.2.1. Thioketone 16
Starting with ketone 3 (250 mg, 0.57 mmol) in o-xylene
(12 h), chromatography (phase 17:3) and crystallization from
MeOH gave thioketone 16 (155 mg, 60%).
Mp 111e113 ꢀC; [a]D20 ꢂ117 (c 0.36). IR (CHCl3): 1687,
1591, 1453, 1385, 1302, 1253, 1031 cmꢂ1
.
1H NMR
(CDCl3): d¼0.81 (s, 3H), 0.88 (s, 3H), 0.94 (s, 3H), 1.14 (s,
3H), 1.16 (s, 3H), 1.19 (d, J¼7.2 Hz, 3H), 1.20 (d, J¼7.2 Hz,
3H), (7ꢁCH3), 2.41 (dt, J¼13.7, 4.7 Hz, 1H, H-6a), 2.62 (d,
J¼19.7 Hz, 1H, H-1a), 2.885 (d, J¼19.5 Hz, 1H, H-6b),
2.892 (td, J¼14.0, 3.4 Hz, 1H, H-1b), 3.41 (septet,
J¼7.2 Hz, 1H, H-4), 3.47 (d, J¼7.9 Hz, 1H, H-28a), 3.55 (br
s, 1H, H-19a), 3.79 (dd, J¼7.8, 1.7 Hz, 1H, H-28b). MS-EI:
m/z (%)¼454 (14) [Mþ], 439 (100), 396 (2), 383 (1), 367
(2), 257 (11), 245 (48), 215 (9), 205 (10), 192 (19). Anal.
Calcd for C30H46OS: C, 79.23; H, 10.20; S, 7.05. Found: C,
79.15; H, 10.22; S, 6.93.
4. Experimental
4.2.2. Dimer sulfide 17
4.1. General
Starting with ketone 4 (300 mg, 0.68 mmol) in toluene
(5 h), chromatography (phase 24:1) and crystallization from
MeOH gave dimer sulfide 17 (220 mg, 74%).
Mp>300 ꢀC (decomp.); [a]D20 þ52 (c 0.50). IR (CHCl3):
Melting points were determined on Kofler block and are un-
corrected. Optical rotatory power was measured on an Autopol
III (Rudolph Research, Flanders, NJ) polarimeter as CHCl3 so-
lutions. 1H, 13C and 31P NMR spectra were recorded on Varian
UNITY Inova 400 (400 MHz for H), using CDCl3 as a sol-
vent. Chemical shifts are expressed in parts per million with
tetramethylsilane as an internal standard for H spectra. 13C
NMR spectra are referenced to CDCl3 (77.00 ppm) and 31P
NMR spectra are referenced to H3PO4 (0 ppm) as an external
standard. Mass spectra (EI) were measured on INCOS 50 (Fin-
nigan MAT) mass spectrometer. Mass spectra (FAB) were
1
1621, 1450, 1384, 1036, 968, 816, 766. H NMR (CDCl3):
d¼0.81 (s, 6H), 0.85 (s, 6H), 0.92 (s, 6H), 0.93 (s, 6H), 1.00
(s, 6H), 1.09 (s, 6H), 1.16 (s, 6H), (14ꢁCH3), 2.15 (dd,
J¼17.2, 6.7 Hz, 2H, 2ꢁH-1a), 3.46 (d, J¼7.8 Hz, 2H, 2ꢁ
H-28a), 3.56 (br s, 2H, 2ꢁH-19a), 3.79 (d, J¼7.8 Hz, 2H,
2ꢁH-28b), 5.86 (dd, J¼6.4, 1.8 Hz, 2H, 2ꢁH-2). MS-FAB:
m/z (%)¼878 (92) [Mþ], 455 (100), 423 (73). Anal. Calcd
for C60H94O2S: C, 81.94; H, 10.77; S, 3.65. Found: C,
81.71; H, 10.95; S, 3.58.
1
1

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M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
4.2.3. Dimer sulfide 18
1.88 (d, J¼16.8 Hz, 1H, H-1a), 2.15 (d, J¼17.6 Hz, 1H, H-
1b), 2.32 (dd, J¼16.8, 7.0 Hz, 1H, H-7a), 2.34 (septet,
J¼6.8 Hz, 1H, H-4), 3.95 (br s, 1H, H-19a), 5.41 (dd,
J¼6.4, 2.4 Hz, 1H, H-2), 5.79 (dd, J¼6.1, 2.3 Hz, 1H, H-6).
MS-EI: m/z (%)¼468 (100) [Mþ], 453 (2), 425 (3), 399 (1),
267 (3), 233 (14), 219 (16), 207 (13), 189 (19). Anal. Calcd
for C30H44O2S: C, 76.87; H, 9.46; S, 6.84. Found: C, 76.77;
H, 9.48; S, 6.78.
Starting with ketone 5 (300 mg, 0.55 mmol) in toluene
(5 h), chromatography (phase 24:1) and crystallization from
mixture MeOHeCHCl3 gave dimer sulfide 18 (243 mg, 81%).
Mp 170e173 ꢀC; [a]D20 þ49 (c 0.28). IR (CHCl3): 1717,
1641, 1456, 1375, 1153, 1128, 891. 1H NMR (CDCl3): d¼0.78
(s, 6H), 0.82 (s, 6H), 0.94 (s, 6H), 1.01 (s, 6H), 1.09 (s, 6H),
1.68 (s, 6H), (12ꢁCH3), 1.82e1.94 (m, 4H), 2.00 (dd, J¼17.1,
6.6 Hz, 2H, 2ꢁH-1a), 2.21 (dt, J¼12.2, 3.7 Hz, 2H), 2.28 (td,
J¼12.4, 3.3 Hz, 2H), 2.65 (m, 2H), 3.02 (dt, J¼11.1, 4.7 Hz,
2H), 4.59 (m, 2H, H-29a), 4.72 (br d, J¼2.1 Hz, 2H,
H-29b), 5.09 (d, J¼12.4 Hz, 2H, 2ꢁAr-CH), 5.15 (d,
J¼12.4 Hz, 2H, Ar-CH), 5.70 (dd, J¼6.4, 0.9 Hz, 2H, 2ꢁ
H-2), 7.29e7.38 (m, 10H, 2ꢁAr-H). MS-FAB: m/z (%)¼1088
(2) [MþH]þ, 559 (3), 527 (1), 91 (100). Anal. Calcd for
C74H102O4S: C, 81.72; H, 9.45; S, 2.95. Found: C, 81.66; H,
9.56; S, 2.88.
4.2.7. Sulfide 22
Starting with diketone 9 (500 mg, 0.90 mmol) in toluene
(5 h), chromatography (phase 10:1) and crystallization from
MeOH gave cyclic sulfide 22 (408 mg, 82%).
Mp 186e188 ꢀC; [a]D20 ꢂ96 (c 0.34). IR (CHCl3): 1725,
1602, 1453, 1382, 1253, 1031, 975. 1H NMR (CDCl3):
d¼0.84 (s, 3H), 0.86 (s, 3H), 0.90 (s, 3H), 0.92 (s, 3H), 1.13
(d, J¼6.8 Hz, 6H), 1.14 (s, 3H), 2.05 (s, 3H), 2.06 (s, 3H), (9ꢁ
CH3), 2.31 (dd, J¼16.5, 7.5 Hz, 1H, H-22a), 2.39 (septet,
J¼6.9 Hz, 1H, H-20), 2.84 (dd, J¼15.3, 3.2 Hz, 1H, H-22b),
3.96 (d, J¼11.1 Hz, 1H, H-28a), 4.07 (d, J¼11.1 Hz, 1H,
H-28b), 4.47e4.52 (m, 1H, H-3a), 5.49 (dd, J¼7.6, 2.3 Hz,
1H, H-21). MS-EI: m/z (%)¼556 (37) [Mþ], 541 (2), 513
(1), 496 (10), 436 (2), 293 (100), 233 (57), 219 (11), 203
(13), 189 (22). Anal. Calcd for C34H52O4S: C, 73.34; H, 9.41;
S, 5.76. Found: C, 73.21; H, 9.50; S, 5.88.
4.2.4. Dimer sulfide 19
Starting with ketone 6 (300 mg, 0.77 mmol) in toluene
(5 h), chromatography (phase 24:1) and crystallization from
mixture MeOHeCHCl3 gave dimer sulfide 19 (230 mg, 77%).
Mp 237e238 ꢀC; [a]D20 ꢂ8 (c 0.80). IR (CHCl3): 1722,
1
1626, 1372, 1256, 1029. H NMR (CDCl3): d¼0.84 (s, 6H),
0.85 (s, 6H), 0.87 (s, 6H), 0.88 (s, 6H), 1.11 (s, 6H), (10ꢁ
CH3), 1.96 (m, 2H, H-12a), 2.05 (s, 6H, 2ꢁCH3), 2.14 (m,
2H, H-17b), 2.59 (m, 2H, H-17a), 3.29 (ddd, J¼14.4, 4.8,
2.0 Hz, 2H, H-12b), 4.47e4.52 (m, 2H, 2ꢁH-3a). MS-FAB:
m/z (%)¼774 (3) [Mþ], 403 (28), 371 (61). Anal. Calcd for
C50H78O4S: C, 77.47; H, 10.14; S, 4.14. Found: C, 77.59; H,
10.22; S, 4.01.
4.2.8. Reaction of unsaturated aldehyde 10
Starting with aldehyde 10 (300 mg, 0.56 mmol) in toluene
(4 h) and chromatography (phase 6:1) gave two products.
(P-R) Oxathiaphosphinine 23a: (164 mg, 40%), mp 212e
213 ꢀC (MeOHeCHCl3); [a]D20 ꢂ18 (c 0.54). IR (CHCl3):
1724, 1651, 1595, 1463, 1258, 1115, 1030. 1H NMR
(CDCl3): d¼0.84 (s, 3H), 0.85 (s, 6H), 0.97 (s, 3H), 1.03 (s,
3H), 2.04 (s, 3H), 2.08 (s, 3H), (7ꢁCH3), 2.42 (td, J¼11.1,
6.1 Hz, 1H, H-19b), 3.33 (t, J¼16.5 Hz, 1H, H-29a), 3.71
(dd, J¼15.9, 12.7 Hz, 1H, H-29b), 3.85 (d, J¼10.8 Hz, 1H,
H-28b), 3.87 (s, 3H, OCH3), 4.22 (d, J¼10.8 Hz, 1H,
H-28a), 4.49 (dd, J¼11.1, 5.3 Hz, 1H, H-3a), 6.40 (d, J¼
20.3 Hz, 1H, H-30), 6.99 (dd, J¼8.7, 3.2 Hz, 2H, 2ꢁAr-H),
7.92 (dd, J¼14.7, 8.7 Hz, 2H, 2ꢁAr-H). 31P{1H} NMR
(CDCl3): d¼80.53 (s, 1P). MS-EI: m/z (%)¼742 (3) [Mþ],
727 (1), 588 (9), 556 (4), 540 (10), 496 (1), 480 (7), 465
(5), 437 (6), 407 (5), 261 (12), 215 (15), 202 (87), 189
(100). Anal. Calcd for C41H59PO6S2: C, 66.28; H, 8.00; S,
8.63. Found: C, 66.11; H, 8.21; S, 8.69.
(P-S) Oxathiaphosphinine 23b: (170 mg, 41%), mp 229e
231 ꢀC (MeOHeCHCl3); [a]D20 ꢂ32 (c 0.57). IR (CHCl3):
1725, 1650, 1594, 1464, 1254, 1111, 1029. 1H NMR
(CDCl3): d¼0.80 (s, 3H), 0.84 (s, 6H), 0.85 (s, 3H), 1.00 (s,
3H), 2.05 (s, 3H), 2.07 (s, 3H), (7ꢁCH3), 2.40 (td, J¼11.4,
6.0 Hz, 1H, H-19b), 3.10 (ddd, J¼15.9, 9.9, 1.7 Hz, 1H,
H-29a), 3.23 (dd, J¼20.8, 16.2 Hz, 1H, H-29b), 3.78 (d, J¼
11.0 Hz, 1H, H-28a), 3.85 (s, 3H, OCH3), 4.20 (d, J¼11.1 Hz,
1H, H-28b), 4.45e4.50 (m, 1H, H-3a), 6.47 (dd, J¼20.6,
1.7 Hz, 1H, H-30), 6.96 (dd, J¼8.7, 3.4 Hz, 2H, 2ꢁAr-H),
7.77 (dd, J¼14.3, 8.9 Hz, 2H, 2ꢁAr-H). 31P{1H} NMR
(CDCl3): d¼78.70 (s, 1P). MS-EI: m/z (%)¼742 (2) [Mþ],
4.2.5. Sulfide 20
Starting with diketone 7 (250 mg, 0.55 mmol) in toluene
(5 h), chromatography (phase 19:1) and crystallization from
isopropanol gave cyclic sulfide 20 (192 mg, 77%).
Mp 165e167 ꢀC; [a]D20 þ15 (c 0.38). IR (CHCl3): 1602,
1
1453, 1385, 1030. H NMR (CDCl3): d¼0.81 (s, 3H), 0.90
(s, 3H), 0.94 (s, 3H), 1.04 (s, 3H), 1.09 (s, 3H), 1.11 (d,
J¼6.8 Hz, 6H), (7ꢁCH3), 1.88 (d, J¼16.8 Hz, 1H, H-1a),
2.19 (d, J¼17.4 Hz, 1H, H-1b), 2.33 (dd, J¼17.2, 7.0 Hz, 1H,
H-7a), 2.34 (septet, J¼6.8 Hz, 1H, H-4), 3.46 (d, J¼7.8 Hz,
1H, H-28a), 3.54 (br s, 1H, H-19a), 3.80 (d, J¼7.9 Hz, 1H,
H-28b), 5.42 (dd, J¼7.0, 2.0 Hz, 1H, H-2), 5.80 (dd, J¼6.1,
2.3 Hz, 1H, H-6). MS-EI: m/z (%)¼454 (100) [Mþ], 439 (4),
411 (7), 383 (2), 269 (3), 233 (18), 219 (29), 207 (19), 192
(11). Anal. Calcd for C30H46OS: C, 79.23; H, 10.20; S, 7.05.
Found: C, 79.09; H, 10.11; S, 7.19.
4.2.6. Sulfide 21
Starting with diketone 8 (100 mg, 0.21 mmol) in toluene
(5 h), chromatography (phase 19:1) and crystallization from
isopropanol gave cyclic sulfide 21 (75 mg, 75%).
Mp 286e288 ꢀC; [a]D20 þ40 (c 0.50). IR (CHCl3): 1764,
1
1603, 1450, 1386, 1119, 971. H NMR (CDCl3): d¼0.85 (s,
3H), 0.97 (s, 3H), 0.98 (s, 3H), 1.04 (s, 3H), 1.08 (s, 3H),
1.108 (d, J¼6.8 Hz, 3H), 1.110 (d, J¼6.8 Hz, 3H), (7ꢁCH3),

M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
3741
727 (1), 588 (13), 556 (8), 540 (11), 496 (2), 480 (7), 465 (5),
437 (6), 407 (5), 261 (12), 215 (17), 202 (73), 189 (100). Anal.
Calcd for C41H59PO6S2: C, 66.28; H, 8.00; S, 8.63. Found: C,
66.15; H, 8.18; S, 8.60.
from mixture MeOHeCHCl3 gave thioacetate 27 (231 mg,
45%).
Mp 239e241 ꢀC; [a]D20 þ94 (c 0.40). IR (CHCl3): 1788,
1
1742, 1626, 1459, 1365, 1282, 1272, 1057, 909. H NMR
(CDCl3): d¼0.87 (s, 3H), 0.92 (s, 3H), 0.93 (s, 3H), 0.94 (s,
3H), 1.12 (s, 3H), 1.14 (d, J¼6.8 Hz, 3H), 1.31 (d, J¼6.8 Hz,
3H), (7ꢁCH3), 1.77 (td, J¼13.1, 3.4 Hz, 1H), 1.86e1.95
(m, 3H), 2.02 (s, 3H, COCH3), 2.05 (m, 1H), 2.53 (td,
J¼14.5, 3.5 Hz, 1H), 2.57 (s, 3H, COCH3), 2.73 (dd, J¼12.5,
2.3 Hz, 1H, H-13b), 3.25 (septet, J¼6.9 Hz, 1H, H-20), 3.89
(d, J¼11.0 Hz, 1H, H-28a), 4.54 (d, J¼11.0 Hz, 1H, H-28b),
5.15 (dd, J¼11.9, 4.6 Hz, 1H, H-3a). MS-EI: m/z (%)¼586
(not found) [Mþ], 510 (24), 469 (100), 425 (12), 291 (6),
265 (21), 231 (5), 217 (7), 205 (64), 189 (45). Anal. Calcd
for C34H50O6S: C, 69.59; H, 8.59; S, 5.46. Found: C, 69.41;
H, 8.69; S, 5.30.
4.2.9. Reaction of hydroxyketone 11
Starting with ketone 11 (300 mg, 0.66 mmol) in toluene
(5 h), chromatography (phase 4:1) and crystallization from
MeOH gave dithiaphospholane 24 (225 mg, 52%).
Mp 266e268 ꢀC; [a]D20 þ21 (c 0.53). IR (CHCl3): 1593,
1497, 1295, 1256, 1179, 1097, 1030, 830. 1H NMR
(CDCl3): d¼0.76 (s, 3H), 0.90 (s, 3H), 0.91 (s, 3H), 1.01 (s,
3H), 1.02 (s, 3H), 1.14 (s, 3H), 1.19 (s, 3H), (7ꢁCH3), 3.44
(d, J¼7.8 Hz, 1H, H-28a), 3.51 (br s, 1H, H-19a), 3.76 (dd,
J¼7.8, 0.4 Hz, 1H, H-28b), 3.87 (s, 3H, OCH3), 4.28 (s, 1H,
H-3b), 5.86 (d, J¼4.6 Hz, 1H, H-1), 6.99 (dd, J¼8.9,
3.4 Hz, 2H, 2ꢁAr-H), 8.07 (dd, J¼15.0, 8.9 Hz, 2H, 2ꢁ
Ar-H). 31P{1H} NMR (CDCl3): d¼83.55 (s, 1P). MS-EI: m/z
(%)¼656 (100) [Mþ], 623 (12), 454 (23), 421 (9), 235 (7),
215 (5), 201 (12), 191 (5). Anal. Calcd for C37H53PO2S3: C,
67.64; H, 8.13; S, 14.64. Found: C, 67.51; H, 8.22; S, 14.58.
4.2.13. Reaction of b-ketoester 15
Starting with b-ketoester 15 (500 mg, 0.97 mmol) in o-xy-
lene (15 h), chromatography (phase 6:1) gave three products.
Bis(thioacetate) 28a: (143 mg, 27%), mp 200e202 ꢀC
(MeOHeCHCl3); [a]2D0 þ25 (c 1.13). IR (CHCl3): 1736,
1
4.2.10. Reaction of hydroxyketone 12
1715, 1452, 1270, 1255, 1027. H NMR (CDCl3): d¼0.87 (s,
Starting with ketone 12 (300 mg, 0.66 mmol) in toluene
(5 h), chromatography (phase 4:1) and crystallization from
MeOH gave dioxaphospholane 25 (177 mg, 43%).
Mp 109e111 ꢀC; [a]D20 þ46 (c 0.17). IR (CHCl3): 1595,
3H), 0.93 (s, 6H), 0.97 (s, 3H), 1.13 (s, 3H), (5ꢁCH3), 1.76
(td, J¼13.3, 3.4 Hz, 1H), 1.87 (ddd, J¼13.3, 8.1, 4.0 Hz,
1H), 1.97e2.06 (m, 2H), 2.51 (dt, J¼15.1, 4.6 Hz, 1H), 2.56
(s, 6H, 2ꢁCSCH3), 2.60 (dd, J¼11.9, 3.5 Hz, 1H, H-13b),
3.78 (s, 3H, OCH3), 4.78 (d, J¼11.3 Hz, 1H, H-28a), 4.90
(d, J¼11.4 Hz, 1H, H-28b), 5.15 (dd, J¼11.8, 4.6 Hz, 1H,
H-3a). MS-EI: m/z (%)¼550 (not found) [Mþ], 475 (100),
431 (11), 417 (22), 399 (37), 371 (13), 339 (12), 339 (11),
271 (15), 231 (8), 205 (14), 191 (15). Anal. Calcd for
C30H46O5S2: C, 65.42; H, 8.42; S, 11.64. Found: C, 65.29;
H, 8.59; S, 11.58.
1
1457, 1260, 1120, 1029, 864. H NMR (CDCl3): d¼0.80 (s,
3H), 0.93 (s, 3H), 0.94 (s, 3H), 1.02 (s, 3H), 1.05 (s, 3H),
1.10 (s, 3H), 1.15 (s, 3H), (7ꢁCH3), 1.99 (d, J¼15.9 Hz,
1H, H-1a), 2.32 (d, J¼15.9 Hz, 1H, H-1b), 3.46 (d,
J¼7.8 Hz, 1H, H-28a), 3.54 (br s, 1H, H-19a), 3.78 (d,
J¼7.5 Hz, 1H, H-28b), 3.86 (s, 3H, OCH3), 6.96 (dd, J¼8.7,
3.4 Hz, 2H, 2ꢁAr-H), 7.83 (dd, J¼14.7, 8.9 Hz, 2H, 2ꢁ
Ar-H). 31P{1H} NMR (CDCl3): d¼103.63 (s, 1P). MS-EI: m/z
(%)¼624 (100) [Mþ], 609 (32), 593 (4), 438 (1), 282 (3),
205 (3), 189 (2). Anal. Calcd for C37H53O4PS: C, 71.12; H,
8.55; S, 5.13. Found: C, 71.01; H, 8.66; S, 4.98.
Thioacetate 28b: (108 mg, 21%), mp 251e253 ꢀC
(MeOHeCHCl3); [a]2D0 þ38 (c 1.28). IR (CHCl3): 1731 sh,
1455, 1376, 1255, 1031. 1H NMR (CDCl3): d¼0.846 (s,
3H), 0.854 (s, 3H), 0.90 (s, 3H), 0.96 (s, 3H), 1.12 (s, 3H),
2.05 (s, 3H), (6ꢁCH3), 2.50 (dt, J¼15.2, 4.8 Hz, 1H), 2.56
(s, 3H, CSCH3), 2.59 (dd, J¼12.0, 3.6 Hz, 1H, H-13b), 3.78
(s, 3H, OCH3), 4.48 (dd, J¼10.8, 5.2 Hz, 1H, H-3a), 4.78
(d, J¼11.4 Hz, 1H, H-28a), 4.90 (d, J¼11.4 Hz, 1H, H-28b).
MS-EI: m/z (%)¼534 (not found) [Mþ], 474 (8), 459 (13),
431 (12), 415 (5), 398 (9), 355 (7), 271 (100), 213 (13), 204
(14), 190 (63). Anal. Calcd for C30H46O6S: C, 67.38; H,
8.67; S, 6.00. Found: C, 67.21; H, 8.81; S, 5.88.
Thioacetate 28c: (165 mg, 45%), mp 194e195 ꢀC
(MeOHeCHCl3); [a]2D0 þ49 (c 0.26). IR (CHCl3): 1738,
1713, 1453, 1368, 1269, 1252, 1029. 1H NMR (CDCl3):
d¼0.87 (s, 3H), 0.93 (s, 6H), 0.95 (s, 3H), 1.14 (s, 3H),
(5ꢁCH3), 1.76 (td, J¼13.3, 3.4 Hz, 1H), 1.83e1.89 (m, 1H),
2.06 (s, 3H, COCH3), 2.41e2.49 (m, 1H), 2.57 (s, 3H,
CSCH3), 2.61 (dd, J¼11.9, 3.5 Hz, 1H, H-13b), 3.77 (s, 3H,
OCH3), 4.45 (d, J¼11.1 Hz, 1H, H-28a), 4.56 (d,
J¼11.1 Hz, 1H, H-28b), 5.15 (dd, J¼11.8, 4.6 Hz, 1H,
H-3a). MS-EI: m/z (%)¼534 (not found) [Mþ], 459 (100),
417 (12), 399 (4), 255 (11), 231 (5), 205 (7), 191 (6). Anal.
4.2.11. Reaction of ketoacid 13
Starting with acid 13 (500 mg, 0.98 mmol) in toluene (2 h),
chromatography (phase 9:1) and crystallization from isopropa-
nol gave diene 26 (273 mg, 62%).
Mp 228e230 ꢀC; [a]D20 þ28 (c 0.36). IR (CHCl3): 1723,
1
1602, 1465, 1371, 1257, 1028. H NMR (CDCl3): d¼0.62 (d,
J¼6.8 Hz, 3H), 0.87 (s, 3H), 0.88 (s, 3H), 0.89 (s, 3H), 0.91
(d, J¼6.8 Hz, 3H), 0.98 (s, 3H), 1.00 (s, 3H), (7ꢁCH3),
1.95e2.20 (m, 5H), 2.06 (s, 3H, COCH3), 2.34 (m, 1H,
H-22b), 2.90 (m, 1H, H-19b), 4.49e4.54 (m, 1H, H-3a), 5.40
(dd, J¼4.6, 3.1 Hz, 1H, H-12). MS-EI: m/z (%)¼452 (100)
[Mþ], 437 (13), 409 (81), 392 (12), 377 (7), 349 (45), 267
(18), 255 (19), 239 (4), 213 (6), 202 (10), 189 (15). Anal. Calcd
for C31H48O2: C, 82.24; H, 10.69. Found: C, 82.38; H, 10.77.
4.2.12. Reaction of anhydride 14
Starting with anhydride 14 (500 mg, 0.88 mmol) in o-xy-
lene (50 h), chromatography (phase 6:1) and crystallization

3742
M. Kvasnica et al. / Tetrahedron 64 (2008) 3736e3743
Calcd for C30H46O6S: C, 67.38; H, 8.67; S, 6.00. Found: C,
67.28; H, 8.77; S, 5.93.
atmosphere at 100% humidity. At the end of the incubation pe-
riod, the cells were assayed using MTT. Aliquots (10 mL) of
the MTT stock solution were pipetted into each well and incu-
bated for a further 1e4 h. After this incubation period forma-
zan produced was dissolved by the addition of 100 mL/well of
10% aq SDS (pH¼5.5), followed by a further incubation at
37 ꢀC overnight. The optical density (OD) was measured at
540 nm with a Labsystem iEMS Reader MF. Tumor cell sur-
vival (TCS) was calculated using the following equation:
TCS¼(ODdrug-exposed well/mean ODcontrol wells)ꢁ100%. The
TCS50 value, the drug concentration lethal to 50% of the tu-
mor cells, was calculated from appropriate doseeresponse
curves.
4.3. X-ray crystallographic data28
Single crystals of 23b were grown from mixture methanole
chloroform.
Crystal data for 23b: C41H59O6PS2, M¼742.97, Ortho-
˚
˚
rhombic, P212121 (No 19), a¼8.9959(1) A, b¼17.9408(2)ꢂA3,
3
˚
˚
c¼24.2538(3) A, V¼3914.41(8) A , Z¼4, Dx¼1.261 Mg m
.
A colorless crystal of dimensions 0.15ꢁ0.17ꢁ0.60 mm was
mounted on glass capillary with epoxy glue and measured at
Nonius Kappa CCD diffractometer by monochromatized Mo
˚
Ka radiation (l¼0.71073 A) at 150(2) K. An absorption was
neglected (m¼0.222 mmꢂ1); a total of 62,352 measured reflec-
tions in the range h¼ꢂ11 to 11, k¼ꢂ23 to 23, l¼ꢂ31 to 31
(qmax¼27.5ꢀ), from which 8957 were unique (Rint¼0.048),
8159 observed according to the I>2s(I) criterion. Cell param-
eters from 5032 reflections (q¼1e27.5ꢀ). The structure was
solved by direct methods (SIR92)29 and refined by full-matrix
least squares based on F2 (SHELXL97).30 The hydrogen
atoms were fixed into idealized positions (riding model) and
assigned temperature factors either Hiso(H)¼1.2 Ueq(pivot
atom) or Hiso(H)¼1.5 Ueq(pivot atom) for methyl moiety.
The refinement converged (D/smax¼0.000) to R¼0.0382 for
observed reflections and wR¼0.0945, GOF¼1.03 for 459 pa-
rameters and all 8957 reflections. The final difference map dis-
played no peaks of chemical significance (Drmax¼1.02,
Acknowledgements
This study was supported by MPO project (FT-TA/027)
from which chemicals and material were paid, Czech Science
Foundation (301/03/1570) supported the biological testing,
and Czech Science Foundation (203/05/P025) supported the
spectroscopic experiments. Indirect costs were paid from the
Czech Ministry of Education (MSM 6198959216). We are
grateful to Stanislav Hilgard for measurement of IR spectra.
We also thank Bohunka Sperlichova for measurement of opti-
cal rotation.
Supplementary data
Drmin¼ꢂ0.34 e Aꢂ3). The absolute configuration was deter-
˚
13C NMR data of compounds 16e22, 23a, 23b, 24e27, and
28aec are provided. Supplementary data associated with this
article can be found in the online version, at doi:10.1016/
j.tet.2008.02.023.
mined unambiguously from anomalous dispersion in agree-
ment to the known configuration of lupane moiety (chiral
parameter ꢂ0.06(5)).
ꢀ
˚
Selected bond lengths (A) and angles ( ): C(19)eC(22)
1.518(3), C(22)C(23) 1.507(3), S(1)eC(23) 1.841(2), S(1)eP(1)
2.0696(7), P(1)eO(1) 1.6275(16), O(1)eC(24) 1.412 (3),
C(22)eC(24) 1.321(3), S(2)eP(1) 1.9271(8), P(1)eC(35)
1.7944(19). C(19)eC(22)eC(23) 117.29(16), S(1)eC(23)eC(22)
115.31 (13), P(1)eS(1)eC(23) 95.74(7), S(1)eP(1)eO(1)
103.73(6), P(1)eO(1)eC(24) 121.93(13), O(1)eC(24)e(22)
128.37(18), C(23)eC(22)eC(24) 125.02(18), C(19)eC(22)eC(24)
117.67 (17), O(1)eP(1)eC(35) 104.52(9), S(1)eP(1)eS(2)
113.76(3), S(2)eP(1)eO(1) 110.20(6).
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