Concise synthesis of stagonolide-F by ring closing metathesis approach and its biological evaluation

Article abstract of DOI:10.1016/j.bioorg.2008.12.002

The first total synthesis of 9-membered macrolide, stagonolide-F (3), starting from commercially available 1,5-pentane diol is reported. A combination of Jacobsen's hydrolytic kinetic resolution (HKR) and Sharpless epoxidation is used for the creation of two stereogenic centers, while ring-closing metathesis (RCM) strategy was used for the construction of the lactone ring. The molecule synthesized exhibited potent antifungal, antibacterial and cytotoxic activities against all the tested strains.

Full text of DOI:10.1016/j.bioorg.2008.12.002

Bioorganic Chemistry 37 (2009) 46–51
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Concise synthesis of stagonolide-F by ring closing metathesis approach
and its biological evaluation
a,b,
Arun Kumar Perepogu a, D. Raman c, U.S.N. Murty c, , Vaidya Jayathirtha Rao
*
*
a Organic Chemistry Division-II, Indian Institute of Chemical Technology (IICT), Uppal Road, Tarnaka, Hyderabad 500 607, Andhra Pradesh, India
b National Institute of Pharmaceutical Education & Research, Balanagar, Hyderabad 500 037, Andhra Pradesh, India
c Biology Division, Indian Institute of Chemical Technology, Hyderabad 500 607, Andhra Pradesh, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 26 October 2008
Available online 21 January 2009
The first total synthesis of 9-membered macrolide, stagonolide-F (3), starting from commercially avail-
able 1,5-pentane diol is reported. A combination of Jacobsen’s hydrolytic kinetic resolution (HKR) and
Sharpless epoxidation is used for the creation of two stereogenic centers, while ring-closing metathesis
(RCM) strategy was used for the construction of the lactone ring. The molecule synthesized exhibited
potent antifungal, antibacterial and cytotoxic activities against all the tested strains.
Ó 2008 Elsevier Inc. All rights reserved.
Keywords:
Jacobsen’s kinetic resolution
Chemo-selective reduction
Sharpless asymmetric epoxidation
Steglich esterification
Ring closing metathesis
Macrolide
Antimicrobial activity
1. Introduction
ring-closing metathesis as key steps. The retrosynthetic analysis
revealed that 3 could be prepared efficiently by RCM protocol from
Macrolides, particularly lactones with medium-sized rings (8–
10 membered), have continued to attract the attention of both
biologists and chemists during recent years, due to the interesting
biological properties and scarce availability of macrolides. A few
examples, in particular of 10-membered-ring containing macro-
lides that display potent biological activity are putaminoxin [1]
(1), pinolidoxin [2] (2) (Fig. 1). The nonenolide (5S,9R)-5-hydro-
xy-9-methyl-6-nonen-9-olide (3), a diastereomer of aspinolide
[3], is one such example, and has been isolated from stagonospora
circii, a fungal pathogen isolated from cirsium arvense [4].
bis-olefin 21 which in turn could be prepared by Steglich esterifi-
cation of acid 15 and homo allylic alcohol 19. Intermediate 19
could be envisaged from racemic propylene oxide 16, while chiral
TBDPS protected allyl alcohol 15 could be produced from 2,3-epoxy
alcohol 11. Thus in the present strategy (5S)-hydroxy group is in-
stalled through Sharpless epoxidation, while the (9R)-hydroxy
group is introduced by Jacobsen’s hydrolytic kinetic resolution
(Scheme 1).
2. Experimental section
Intrigued by the biological properties and also structural simil-
ilarity with highly potent putominoxin 1, pinolidoxin 2, and other
phytotoxic nonenolides [5] and in continuation of our program to-
wards synthesis of biological active compounds [6], we became
interested in developing a simple and flexible route to the total
synthesis of stagonolide-F (3). However, a few related nonenolides
[7] have been reported starting from chiral pool, we are reporting
here the synthesis and biological screening of stagonolide-F, start-
ing from commercially available 1,5-pentane diol employing Jac-
obsen’s hydrolytic kinetic resolution, Sharpless epoxidation and
2.1. General
NMR spectra were measured on a Gemini 200 MHz Varian
instrument and Avance 300 MHz Bruker UX-NMR instrument in
CDCl3 as reference solvent and chemical shifts were expressed as
d. Coupling constants J are given in Hz. Tetramethyl silane was
used as an internal standard for 1H NMR. Enantiomeric excess is
determined by normal-phase HPLC using Chiralpak AD-H [amylose
tris-(3,5-dimethylphenylcarbamate),
coated on a 5- m silica particle] column from Diacel chemical
250 mm ꢀ 4.6 mm
i.d.,
l
industries Ltd. Mass spectra were recorded on VG Micromass
7070 H (EI and ESI) and Finnigan Mat 1020 Mass (GC–MS) instru-
ments. High-resolution (HR) mass spectra were recorded using a
VG Autospec magnetic sector mass spectrometer (Waters, Man-
* Corresponding author. Address: Organic Chemistry Division-II, Indian Institute
of Chemical Technology (IICT), Uppal Road, Tarnaka, Hyderabad 500 607, Andhra
Pradesh, India. Fax: +91 40 27160757.
E-mail addresses: aruniict@gmail.com (A.K. Perepogu), jrao@iict.res.in (V.J. Rao).
0045-2068/$ - see front matter Ó 2008 Elsevier Inc. All rights reserved.
doi:10.1016/j.bioorg.2008.12.002

A.K. Perepogu et al. / Bioorganic Chemistry 37 (2009) 46–51
47
4.46 (s, 2H, OCH2Ph), 7.25–7.31 (m, 5H, Ar-H) ppm; 13C NMR
(75 MHz, CDCl3) d: 22.7, 29.5, 31.4, 55.9, 58.5, 61.8, 70.0, 72.9,
127.5–128.2, 138.3 ppm; ESI-MS [M+23]+: 259; ESI-HRMS Calcd.
for C14H20O3 Na [M+23]+ 259.1317, found: 259.1310. The enantio-
meric purity was determined by HPLC (Daicel Chiralcel AD column,
15% i-PrOH/hexane, flow rate 1.0 mL/min, Detection wavelength:
OH
H OR
O
HO
HO
H
O
O
O
Me
O
O
O
O
3 Stagonolide F
1 a R = H, putaminoxin
1 b R = Ac, 5-O-Acrtylputaminoxin
270 nm): smajor = 9.9 min; sminor = 11.0 min.
2
Pinolidoxin
2.3. (3S)-7-(Benzyloxy)-1-hepten-3-ol (12)
OH
O
OH
OH
OH
H
O
H
OH
OH
OH
To a stirred solution of epoxy alcohol 11 (2.25 g, 9.5 mmol) in
dry Et2O:CH3CN (5:3), 15 mL were added sequentially Ph3P
(7.46 g, 28.6 mmol), pyridine (3.1 mL, 38.1 mmol) and I2 (4.82 g,
19.06 mmol) at 0 °C. After being stirred for 2 h at 0 °C, H2O
(0.35 mL, 19.06 mmol) was added into the system. The reaction
mixture was refluxed for 6 h at 40 °C, then 20% Na2S2O3 (aq.)
(15 mL) and saturated NaHCO3 (aq.) (15 mL) were added to quench
the reaction and the organic layer was extracted with ether
(3 ꢀ 50 mL). The combined ether extracts were washed with 5%
HCl (4 ꢀ 10 mL), H2O and brine, then dried. Evaporation of the sol-
vent gave the residue, which was flash chromatographed eluting
with hexane and ethylacetate (9:1) gave 12 (2.0 g, 95%) as colorless
oil.
O
O
O
CH3
O
Me
O
CH3
O
6 Stagonolide D
Fig. 1. Phytotoxic nonenolides.
7 Stagonolide E
4 Stagonolide B
5 Stagonolide C
chester, UK). IR Spectra were recorded on Perkin Elmer Model 283B
and Nicolet-740 FT-IR instruments, and band positions were re-
ported in wave numbers (cmꢁ1). Column chromatography was per-
formed using silica gel
procedures and spectral data for compounds 7, 8, 9, 10, 18, 19
and isomer of stagonolide-F (3) are given in Supporting
H (60–120 lm). The experimental
½
a 2D7
ꢂ
¼ þ3:8 ðc ¼ 1; CHCl3Þ; IR (neat)
m
cmꢁ1: 3414, 3066, 3029,
2933, 2859, 1718, 1642, 1453, 1275, 1102, 993; 1H NMR
(300 MHz, CDCl3) d: 1.53–1.68 (m, 6H, 3 ꢀ CH2), 3.47 (t,
J = 6.4 Hz, 2H, CH2CH2O), 4.06–4.12 (m, J = 6.0 Hz, 1H,
CH2CHOHCH), 4.49 (s, 2H, OCH2Ph), 5.07–5.24 (dd, J = 10.3,
17.1 Hz, 2H, CHCH2), 5.80–5.91 (dq, J = 6.2, 10.3, 16.6 Hz, 1H,
CHCHCH2), 7.28–7.33 (m, 5H, Ar-H) ppm; 13C NMR (75 MHz,
CDCl3) d: 22.0, 29.6, 36.7, 70.1, 72.8, 96.1, 114.3, 127.5, 128.2,
138.5, 141.3 ppm; ESI-MS: 243 [M+23]+; ESI-HRMS Calcd. for
C14H20O2Na [M+Na]+: 243.1360, found: 243.1357.
Z
information.
2.2. (2R,3S)-3-[4-(Benzyloxy)butyl]oxiran-2-ylmethanol (11)
To a cooled (ꢁ30 °C) suspension of activated, powdered 4 Å MS
(1 g) in CH2Cl2 (25 mL) were added (+)-DET (0.37 mL, 2.18 mmol),
Ti(OPri)4 (0.53 mL, 1.8 mmol), and cumene hydroperoxide (1.6 mL,
10.8 mmol). After 20 min, a solution of allylic alcohol 10 (2.0 g,
9.0 mmol) in CH2Cl2 (10 mL) was added at ꢁ30 °C over 15 min.
The resulting mixture was stirred at that temperature for 3 h,
quenched with a cold solution of ferrous sulfate and tartaric acid
(stoichiometric amount) in de-ionized water, stirred vigorously
for 30 min, and extracted with ether (50 mL ꢀ 3). The combined or-
ganic layers were treated with a pre-cooled (0 °C) solution of 5 mL
of 30% NaOH (w/v) in brine and stirred for 1 h at rt. The two layers
were separated and the aqueous layer was extracted with ether
(20 mL ꢀ 3). The combined ether layers were washed with brine,
dried over Na2SO4, filtered, and concentrated. The residue was
chromatographed (SiO2, ethyl acetate/hexane = 1.5:8.5) to give 11
(1.7 g, 80%, 80% ee) as colorless syrup.
2.4. (1S)-1-[4-(Benzyloxy)butyl]-2-propenyloxy)(tert-
butyl)diphenylsilane (13)
To a stirred solution of 12 (100 mg, 0.45 mmol) in CH2Cl2
(5 mL), imidazole (50 mg, 0.67 mmol), and TBDPSCl (140 lL,
0.54 mmol) were added at 0 °C and stirred at rt for 3 h. The reac-
tion mixture was treated with satd NH4Cl (5 mL) and extracted
with CH2Cl2 (2 ꢀ 10 mL). The organic layer was washed with water
(2 ꢀ 10 mL), brine (2 ꢀ 10 mL), dried over Na2SO4, evaporated, and
the residue obtained was purified by column chromatography (60–
120 silica gel, 0.3:9.7 ethyl acetate–hexane) to furnish 13 (160 mg,
78%) as colorless syrup.
½
a 2D7
ꢂ
¼ þ17:3 c ¼ 1; CHCl3Þ; IR (neat)
½
a 2D7
ꢂ
¼ ꢁ20:6 ðc ¼ 1; CHCl3Þ; IR (neat)
m
cmꢁ1: 3414, 3063, 2936,
cmꢁ1: 3069, 2933, 2857, 1724, 1463, 1427, 1262, 1108, 999,
2862, 1717, 1454, 1099, 878, 746; 1H NMR (300 MHz, CDCl3) d:
1.56–1.68 (m, 6H, 3 ꢀ CH2), 2.81–2.89 (m, 2H, CH2CHCHCH2),
3.45 (t, J = 5.87 Hz, 2H, CH2CH2O), 3.53–3.87 (m, 2H, CHCH2OH),
m
701; 1H NMR (300 MHz, CDCl3) d: 1.04 (s, 9H, 3 ꢀ CH3), 1.25–
1.46 (m, 6H, 3 ꢀ CH2), 3.30 (t, J = 6.8 Hz, 2H, CH2CH2O), 4.08–4.14
OH
7
6
8
4
3
5
1
9
O
10
2
O
3
O
OH
3
O
O
OTBDPS
OH
O
O
BnO
OH
3
HO
3
16
19
11
15
21
Scheme 1. Retrosynthesis of stagonolide-F (3).

48
A.K. Perepogu et al. / Bioorganic Chemistry 37 (2009) 46–51
(m, 1H, HCOTBDPS), 4.40 (s, 2H, OCH2Ph), 4.91–4.98 (m, 2H,
CHCH2), 5.69–5.80 (dq, J = 6.8, 10.5, 16.6 Hz, 1H, CHCHCH2),
7.21–7.37 (m, 10H, Ar-H), 7.59–7.65 (m, 5H, Ar-H) ppm; 13C
NMR (75 MHz, CDCl3) d: 19.3, 21.0, 27.0, 29.5, 37.3, 70.2, 72.7,
74.5, 114.3, 127.3, 128.2, 129.4, 135.8, 140.7 ppm; ESI-MS: 481
[M+23]+; ESI-HRMS Calcd. for C30H38O2 SiNa [M+Na]+: 481.2518,
found: 481.2583.
CDCl3) d: 1.06 (s, 9H, 3 ꢀ CH3), 1.16 (d, J = 5.85 Hz, 3H, CH3), 2.01
(t, J = 6.5 Hz, 2H, CH2COO), 2.23–2.33 (m, 6H, 3 ꢀ CH2), 4.04–4.13
(m, 2H, 2 ꢀ CH), 4.85–5.09 (m, 4H, olefin), 5.63–5.84 (m, 2H, ole-
fin), 7.29–7.39 (m, 6H, Ar-H), 7.58–7.67 (m, 4H, Ar-H) ppm; 13C
NMR (75 MHz, CDCl3) d: 20.0, 27.0, 29.7, 36.8, 40.3, 74.2, 96.1,
114.6, 117.6, 127.3, 129.4, 135.8, 140.4 ppm; ESI-MS: 473
[M+Na]+; ESI-HRMS Calcd. for C28H38O3SiNa [M+Na]+: 473.2492,
found: 473.2487.
2.5. (5S)-5-[1-(Tert-butyl)-1,1-diphenylsilyl]oxy-6-hepten-1-ol (14)
2.8. (1R)-1-Methyl-3-butenyl-(5S)-5-hydroxy-6-heptenoate (21)
To a stirred solution of 13 (100 mg, 0.22 mmol) in dichloro-
methane–water (19:1, 5 mL), DDQ (0.25 g, 1.11 mmol) was added
and stirred at reflux for 4 h. saturated aq. NaHCO3 solution
(5 mL) was added to the reaction mixture and extracted with
CH2Cl2 (3 ꢀ 10 mL). The combined organic layers were washed
with water (5 mL), brine (5 mL), dried over Na2SO4, and concen-
trated. The crude residue was purified by column chromatography
(silica gel, 60–120 mesh, ethylacetate–hexane, 0.8:9.2) to afford 14
(69 mg, 85%) as colorless syrup.
A solution of 20 (0.10 g, 1.66 mmol) in THF (1 mL) was taken in
a plastic bottle, and HF-pyridine (2–3 drops) was added at 0 °C and
stirred at room temperature for 12 h. The reaction mixture was
quenched with saturated NaHCO3 solution (5 mL) at 0 °C and ex-
tracted with AcOEt (2 ꢀ 50 mL). The organic layer was washed
with saturated CuSO4 solution (5 mL), dried over Na2SO4, and the
residue obtained was purified by column chromatography (silica
gel, 60–120 mesh, ethylacetate–hexane, 2:8) to afford 21
(0.035 g, 75%) as colorless syrup.
½
a 2D7
ꢂ
¼ þ20:9 ðc ¼ 1; CHCl3Þ; IR (neat)
m
cmꢁ1: 3398, 3070, 2931,
2857, 1710, 1642, 1466, 1425, 1216, 1108, 922,701; 1H NMR
(300 MHz, CDCl3) d: 1.06 (s, 9H, 3 ꢀ CH3), 1.25–1.44 (m, 6H,
3 ꢀ CH2), 3.47 (t, J = 6.6 Hz, 2H, CH2CH2OH), 4.08–4.16 (m, 1H,
HCOTBDPS), 4.93–5.02 (m, 2H, CHCH2), 5.68–5.85 (dq, J = 6.6,
10.2, 16.8 Hz, 1H, CHCHCH2), 7.25–7.42 (m, 5H, Ar-H), 7.58–7.67
(m, 5H, Ar-H) ppm; 13C NMR (75 MHz, CDCl3) d: 20.5, 27.0, 29.7,
32.5, 37.1, 62.8, 74.4, 114.4, 127.3, 129.5, 135.9, 140.6 ppm; ESI-
MS: 367 [Mꢁ1]+; ESI-HRMS Calcd. for C23H32O2SiNa [M+Na]+:
391.2058, found: 391.2069.
½
a 2D7
ꢂ
¼ þ6:5 ðc ¼ 0:5; CHCl3Þ; IR (neat)
m : 2923, 2853,
cmꢁ1
2360, 1711, 1459, 1375, 1216; 1H NMR (300 MHz, CDCl3) d: 1.09
(d, J = 6.1 Hz, 3H, CH3), 1.56–1.72 (m, 2H), 1.9 (m, 2H), 2.39 (t,
J = 7.5 Hz, 2H), 2.45–2.60 (m, 2H), 4.0–4.12 (m, 1H), 4.77–4.83
(m, 1H), 5.07, 5.17 (dd, J = 10.5, 16.2 Hz, 2H, olefin), 5.19–5.37
(dd, J = 10.5, 16.6 Hz, 2H, olefin), 5.79–5.91 (dq, J = 5.2, 10.5,
15.8 Hz, 2H, olefin); 13C NMR (75 MHz, CDCl3) d: 18.0, 20.4, 27.8,
29.5, 36.0, 72.6, 80.2, 114.9, 116.8, 135.9, 140.6 ppm; ESI-MS:
235 [M+Na]+; ESI-HRMS Calcd. for C12H20O3Na [M+Na]+:
235.1315, found: 235.5312.
2.6. (5S)-5-[1-(Tert-butyl)-1,1-diphenylsilyl]oxy-6-heptenoic acid
(15)
2.9. (5S,9R)-5-Hydroxy-9-methyl-6-nonen-9-olide (3)
To a stirred solution of 14 (100 mg, 0.27 mmol) in DMF (5 mL)
was added PDC (0.5 g, 1.35 mmol) at room temperature. After
10 h, the mixture was quenched with cold water (5 mL), and ex-
tracted with AcOEt (3 ꢀ 10 mL). The combined organic layer was
washed with KHSO4 (15 mL, 1 mol/L), water (10 mL), and brine
(10 mL), respectively, dried over Na2SO4, filtered, and concentrated
in vacuo. Flash chromatography of the residue over silica gel
(CH2Cl2/MeOH, 40:1) afforded 15 as a colorless oil (77 mg, 75%
yield).
Ester 21 (50 mg, 0.23 mmol) is dissolved in freshly distilled de-
gassed anhydrous CH2Cl2 (100 mL) was treated with Grubb’s cata-
lyst I (22 mg, 0.027 mmol) and heated at reflux for 2 days under
inert atmosphere. Most of the solvent was then distilled off and
the concentrated solution is left to stir at room temperature for
2 h under air bubbling in order to decompose the catalyst. The
reaction mixture was evaporated to dryness to give a brown resi-
due, which was purified by column chromatography (silica gel,
60–120 mesh, ethylacetate–hexane, 3:97) to afford 3 (22 mg,
55%) as colorless syrup.
½
a 2D5
ꢂ
¼ þ20:7 ðc ¼ 1:0; CHCl3Þ; IR (neat)
m
cmꢁ1: 3444, 3070,
2931, 2858, 1707, 1462, 1425, 1257, 1109; 1H NMR (300 MHz,
CDCl3) d: 1.06 (s, 9H, 3 ꢀ CH3), 1.38–1.64 (m, 4H, 2 ꢀ CH2, 2.18 (t,
J = 7.34 Hz, 2H, CH2COOH), 4.11–4.19 (m, 1H, HCOTBDPS), 4.96–
5.06 (m, 2H, CHCH2), 5.69–5.86 (dq, J = 6.6, 11.0, 16.8 Hz, 1H,
CHCHCH2), 7.32–7.43 (m, 5H, Ar-H), 7.60–7.68 (m, 5H, Ar-H); 13C
NMR (75 MHz, CDCl3) d: 19.5, 27.0, 33.8, 36.6, 74.0, 114.7, 127.3,
129.4, 135.8, 140.2, 179.7 ppm; ESI-MS: 405 [M+1]+; ESI-HRMS
Calcd. for C23H30O3SiNa [M+Na]+: 405.1868, found: 405.1861.
½
a 2D5
ꢂ
¼ ꢁ26:3 ðc ¼ 0:5; CHCl3Þ, (lit.[3] ½a D25
ꢁ 27 ðc ¼ 0:1;CHCl3Þ;
ꢂ
IR (neat)
m
cmꢁ1: 3449, 2924, 2853, 1738, 1644, 1461, 1235, 1099;
1H NMR (300 MHz, CDCl3) d: 1.1 (d, J = 6.3 Hz, 3H, CH3), 1.6–1.72
(m, 2H, CH2CHOH), 1.85–2.28 (m, 2H, CH2CH2CH2), 2.31 (t, J = 6.5
Hz, 2H, CH2CO), 2.45–2.65 (m, 2H, CH2CHCH), 3.98–4.0 (m, 1H,
CH2CHOH), 4.8–5.0 (m, 1H, CH3CHOCO), 5.35–5.40 (dd, J = 15.2,
9.2 Hz, 1H, CHOHCHCH), 5.52–5.62 (ddd, J = 15.2, 10.4, 4.2 Hz,
1H, CH2CHCH); 13C NMR (75 MHz, CDCl3) d: 21.3, 30.0, 31.5,
34.3, 35.0, 71.6, 75.4, 131.2, 134.2, 174.8 ppm. ESI-MS: 185
[M+H]+; ESI-HRMS Calcd. for C10H16O3H [M+H]+: 185.1153, found:
185.5150.
2.7. (1R)-1-Methyl-3-butenyl-(5S)-5-[1-(tert-butyl)-1,1-
diphenylsilyl]oxy-6-heptenoate(20)
3. Results and discussion
To a stirred solution of acid 15 (1 g, 2.6 mmol) and DMAP
(64 mg, 0.52 mmol in anhydrous DCM (25 mL) was added alcohol
19 (0.9, 10.4 mmol) taken in DCM at rt. The reaction mixture is
cooled to 0 °C and added DCC (1.0 g, 5.2 mmol) in DCM and stirred
for 10 min and brought to room temperature and stirred overnight.
The white precipitate formed was filtered off and washed with 2N
HCl, 5% NaHCO3 and finally with water. The esterification product
20 is purified by distillation at atmospheric pressure at 125 °C.
(0.8 g, 65%).
In designing a route to 21, we chose racemic propylene oxide 16
as one of the appropriate starting materials (Scheme 2). Thus, com-
mercially available propylene oxide 16 was subjected to Jacobsen’s
hydrolytic kinetic resolution [8] by using (R,R)-Salen-Co-OAc cata-
lyst 4 (Fig. 2) to give (R)-propylene oxide 17 as a single isomer, and
it was isolated from the more polar diol 18 by distillation and the
optical purity was proven by comparison with reported literature
[8]. Our next task was to construct the homoallylic alcohol 19 after
keeping enantiomerically pure epoxide 17 in hand. Thus (R)-pro-
½
a 2D7
ꢂ
¼ þ4:5 ðc ¼ 0:5; CHCl3Þ; IR (neat)
m : 2926, 2855,
cmꢁ1
1735, 1642, 1462, 1425, 1216, 1110, 761; 1H NMR (300 MHz,

A.K. Perepogu et al. / Bioorganic Chemistry 37 (2009) 46–51
49
The reaction was carried out in refluxing benzene for 2 h, which
gave a mixture of trans and cis conjugated ester in 88:12 ratio. The
trans compound 9 was purified and then subjected to chemo-selec-
tive reduction [12] using LAH/AlCl3 in anhydrous ether to afford al-
lyl alcohol 10 in 86% yield. Sharpless asymmetric epoxidation [13]
OH
O
O
i
OH
+
16
18
17
OH
O
ii
of 10 with L-(+)-DET produced 2,3-epoxy alcohol 11 in 85% yield
19
17
with 80% ee. Accordingly, one pot transformation [14] of 2,3-epoxy
alcohol 11 into an allylic alcohol 12 was achieved by the insitu for-
mation of the epoxy iodide and its subsequent reduction with
phosphine hydroxyiodide in 95% yield. Allylic alcohol 12 was trea-
ted with TBDPSCl to afford 13 (78%), which was subjected to deb-
enzylation with DDQ in CH2Cl2–H2O to give 14 (88%). Finally, the
hydroxyl group of 14 was oxidized with pyridinium dichromate
[15] (PDC) to give acid 15 in 75% yield. After successfully obtaining
the alcohol 19 and acid 15 fragments, the coupling reaction was
achieved by employing Steglich esterification [16] (Scheme 4).
Desilylation of 20 was achieved under neutral conditions using
HF-Pyridine to give bis-olefin 21. Finally diene 21 was treated with
Grubb’s first generation catalyst 5 (Fig. 2) under high dilution con-
dition [17] furnished a 10:1 E:Z mixture which on chromatographic
purification gave the target molecule in 55% yield. The physical and
spectral data [20] of 3 are identical to those reported in the litera-
ture [4]. Application of this strategy in the total synthesis of other
analogues is currently in progress.
Scheme 2. Reagents and conditions: (i) (R,R)-Jacobsen catalyst, H2O, rt, 40% and (ii)
Vinylmagnesium bromide, CuI, THF, –78 °C, 85%.
H
H
PCy3
N
O
N
O
Ph
Cl
Cl
M
Ru
PCy3
t
-Bu
t-Bu
t-Bu
t-Bu
M = Co(OAc); (R,R)-1-OAc
Grubs first generation (l) catalyst
5
4
Fig. 2. Catalysts used in synthetic strategy.
The synthesized stagonolide-F (3) was evaluated invitro for
antibacterial and antifungal activity using ‘agar well diffusion anti
microbial assay’. The antibacterial activity was evaluated against
gram-positive bacterial strains Staphylococcus aureus (MTCC 737),
Staphylococcus epidermidis (MTCC 435), and gram-negative strains
Escherichia coli (MTCC 1687), and Pseudomonas aeruginosa (MTCC
1688). The antifungal activity was evaluated against pathogenic
strains Sacharomyces cereviseae (MTCC 36), Candida albicans (MTCC
227), Aspergillus niger (MTCC 1344) and Rhizopus oryzae (MTCC
262). The MIC (minimum inhibitory concentration) values for anti-
bacterial activity were determined using standard broth microdilu-
pylene oxide 17 was treated with vinylmagnesium bromide [9] in
the presence of CuI to give the homoallylic alcohol 19 in excellent
yield (85%).
The synthesis of fragment 15 was initiated from commercially
available 1,5-pentanediol 6 as illustrated in Scheme 3. Thus selective
monoprotection of 6 with benzyl bromide in DMF gave benzyl-ether
7 in 70% yield, which on oxidation under Swern conditions [10] gave
the corresponding aldehyde 8 in 90% yield. Compound 8 was sub-
jected to a two-carbon homologation by means of Wittig reaction
[11] using ethoxycarbonylmethylenetr-iphenylphosphorane.
O
ii
iii
i
HO
HO
OH
3
BnO
OH
BnO
BnO
3
O
OEt
OH
3
3
6
7
8
9
iv
OR
OTBDPS
O
v
vi, vii
viii
BnO
3
3
BnO
OH
BnO
3
3
12 R = H
13 R = TBDPSCl
14
10
11
ix
OTBDPS
O
HO
3
15
Scheme 3. Reagents and conditions: (i) NaH, BnBr, 0 °C – rt 4 h, 70%; (ii) (COCl)2, DMSO, DCM, ꢁ78 °C, 90%; (iii) Ph3P = CHCO2Et, anhyd benzene, reflux 2 h, 88%; (iv) LAH/
AlCl3, THF, 0 °C 1/2 h, 86%; (v) Ti(OPri)4, (+)DET, PhC(CH3)2O2H, dry DCM, ꢁ20 °C, 5 h, 85%, 80% ee; (vi) Ph3P, pyridine, I2, Et2O:CH3CN (5:3), 0 °C, H2O, reflux, 6 h, 95%; (vii)
TBDPSCl, Imidazole, DMF, rt, 78%; (viii) DDQ, CH2Cl2/H2O (19:1), reflux, 3 h, 88% and (ix) PDC, DMF, rt, 75%.
O
OH
OTBDPS
O
OH
i
ii
iii
15
19
+
3
3
O
O
O
O
20
21
3
Scheme 4. Reagents and conditions: (i) DCC, DMAP, CH2Cl2, 0 °C – rt, 65%; (ii) HF-pyridine, CH2Cl2, rt, 12 h, 75% and (iii) (Pcy3)2Cl2Ru = CHPh (12 mole%), CH2Cl2, 40 °C, 48 h,
55%.

50
A.K. Perepogu et al. / Bioorganic Chemistry 37 (2009) 46–51
Table 1
Invitro antimicrobial activity of synthesized stagonolide-F (3).
Microorganism
Antimicrobial activity
Zone of Inhibitiona
MICb
(lg/mL)
Compd. 3
Control
Compd. 3
Control
Bacterial strains
Streptomycin
Nitrofurantoin
Staphylococcus aureus
Staphylococcus epidermis
Escherichia coli
16
13
14
17
25
26
32
28
100
100
100
200
50
50
25
Psudomonas aeruginosa
100
Fungal strains
Clotrimazole
Saccharomyces serviseae
Candida albicans
Aspergillus niger
16
14
16
14
23
22
18
21
Rhizopus orizae
c
Cell line
IC50
(lg/mL)
Compd. 3
Control (Etoposide)
Cytotoxic activity
THP-1d
U-937e
32.67 4.88
34.72 3.45
1.4
1.2
a
ZI: zone of inhibition (diameter in mm).
MIC (minimum inhibitory concentration in
b
lg/mL) was determined as 90% inhibition of growth with respect to positive growth control. Negative control: DMSO, no
inhibition.
c
IC50: inhibitory concentration.
THP-1: human acute monocytic leukemia cell line.
U-937: human leukemic monocyte lymphoma cell line.
d
e
tion technique described by NCCLS [18]. Nitrofurantoin was used
as reference drug. In comparison with the antimicrobial activity,
Clotrimazole was used as reference antifungal drug, while Strepto-
mycin was used as reference antibacterial drug. All the biological
data is depicted in Table 1 as zone of inhibition of growth (ZI)
and minimum inhibitory concentration (MIC) values. From the
activity results, it is observed that stagonolide-F (3) has showed
potent antifungal and antibacterial activity against all the tested
fungal and bacterial strains. The analysis of ZI and MIC values for
antibacterial activity revealed stagonolide-F (3) has more than
60% antibacterial activity against Staphylococcus aureus and moder-
ate activity against other tested bacterial strains in comparison
with the tested reference drug’s antibacterial activity. Zone of inhi-
bition results for antifungal activity revealed compound 3 has
more than 80% activity against Aspergillus niger and more than
60% activity against other fungal strains in comparison with tested
reference drug’s antifungal activity.
Cytotoxic activity was evaluated against THP-1 and U-937 hu-
man cancer cell lines (Human acute monocytic leukemia cell line,
Human leukemic monocyte lymphoma cell line). Cytotoxicty was
measured using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphe-
nyl tetrasolium bromide] assay, according to the method of Mos-
mann [19]. Etoposide was used as reference cytotoxic drug. IC50
values (Inhibotory Concentration) of the test compound 3 is calcu-
lated and presented in Table 1. It is evident from the results that
the test compound has shown, significant decrease in cell viability
in the test cell line in concentration dependant manner.
3 exhibited potent antifungal, significant antibacterial and cyto-
toxic activities against all the tested strains.
Acknowledgments
We thank Director IICT, Project Director NIPER and Head of the
Division Org ll for their encouraging support. PAK thanks CSIR for
fellowship.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bioorg.2008.12.002.
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