- Email: [email protected]
Synthesis and absolute configuration of syringolide 2, an elicitor from Pseudomonas syringae pv. tomato
Tetrahedron Letters. Vol. 36, No. 18, pp. 3201-3202. 1995
Pergamon
ElsevierScienceLtd Printedin Great Britain 0040-4039/95$9.50+0.00 0040-4039(95)00512-9
Synthesis and Absolute Configuration of S y r i n g o l i d e 2, an Elicitor from Pseudomonas syringae pv. tomato
Shigefumi Kuwahara,* Masahiko Moriguchi, Kazuhiro Miyagawa, Masako Konno, and 0samu Kodama Laboratoryof AgriculturalChemicals, Faculty of Agriculture,IbarakiUniversity,Ami-machi,Inashiki-gun,lbaraki300-03, Japan
Abstract: The enantioselectivesynthesis of syringolide 2, one of the two elicitors produced by Pseudomonas syringae pv. tomato, was accomplishedin 11 steps from dielhyll~-tartrate.
In 1993 Keen et aL isolated two C-glycosides possessing a new ring system as specific elicitors from Pseudomonas syringae pv. tomato and named them syringolide 1 (1) and syringolide 2 (2).1, 2 Their absolute configurations were deduced from the assumption that the incorporated xylulose moiety should be the naturally occurring D-form. These elicitors are produced through the action of avirulence gene (avrD) of P. syringae pv. tomato and function as specific molecular signals which cause hypersensitive responses specifically in soybean plants carrying the Rpg4 disease resistance gene. 2 Much interest is now being attracted in the particular receptor protein which may recognize the syringolides as specific signals, and the synthesis of radiolabelled syfingolides is expected to provide a powerful tool for identifying the receptor. First of all, we carried out the synthesis of the naturally occurring enantiomer of syringolide 2 in order to establish the synthetic route leading to the syringolides.
HO H 0
0
R' " ' ~ ' ~ H "" o HO H
0
n-CTH15"~v ~ 0 o . ~ OH 3,J~ , HO'" 5
1: R=n-CsHn
OH
2:R=n-C7H15
3
According to the biosynthetic pathway proposed by Keen et al., 1,2 we chose 3 as the key intermediate of our synthesis of 2. The intramolecular Knoevenagel condensation of 3 followed by conjugate addition of 5'OH to the resulting ct-acyl-ct,13-unsatumted lactone and hemiacetalization of T-OH to C3-carbonyl function should give rise to 2 judging from the generality that the c/s-fused oxabicyclo[3.3.0]octane system is thermodynamically more favored than the corresponding tram-isomer. Our synthesis began with the selective protection 3 of the known diol 4a 4 obtained almost quantitatively from diethyl D-tam'ate in 2 steps (Scheme 1). The Swern oxidation of the resulting alcohol 4b gave 5, which was then converted into protected D-xylulose 6a by the addition of (1-ethoxyethoxy)methyUithium5 followed by the Swern oxidation. Selective deprotection of 6a gave 6b, 6 which was esterified with 3-oxodecanoic acid 7 to afford 7, a protected form of 3. The Knoevenagel condensation of 7 into 8 was achieved by simply mixing 7 with silica gel in n-hexane-EtOAc.6 Although the direct conversion of 8 into 2 using aqueous acidic conditions
3201
3202
OMOM
RO~o
H
b)
OMOM
~ a) 4b R=TBS 4a R=H
OMOM TBSO,v~CHO OMOM
c), d ) ~
MOMO 0 TBSO 1 ~ , .OR OMOM
5
6aR=EE 6b R=H
O
O
0
0
n.CrHt51Jl"-.~ 0 n-C-,,H.~s~ e) ~ 0..~ OTBS f) ~ MOMO",..I.y MOMO'÷'~ J
7
OMOM
MOMO¢-t-,,,.~OTBS
MeO u
g)
0
n'C7H15' ( ~~" O ~-
H" "(3 HO H
8
9
Scheme 1. a) Nail, TBSCI, 0"C, 30 min, it, 30 rain, 91% ; b) Swern oxidation, -78'C, 2h; c) Bu3SnCH2OEE, n-BuLi, THF, -78"C, lh, then Swern oxidation, 70% from 4b d) PITS, EtOH, rt, 2.5h, 90%; e) 3-oxodecanoic acid, DCC, DMAP, CH2C12, rt, lh; f) SiO2 (Merck Kieselgel 60 Art. 7734, 30 g / g of 7), n-hexane-EtOAc (8:1), rt, 24h, 56% from 6b, g) Dowex 50W-X8 (100 g / g of 8), MeOH, rt, 34h, 36%. was unsuccessful, the methyl ether of 2 (i.e. 9) was obtained in about 35 % yield by treating with Dowex 50X8 or Amberlyst 15E in dry methanol. Finally, 9 was treated with p - T s O H in acetone-water for 16h at room temperature to give 2 in 5 1 % yield as colorless needles (m.p. 118-120.5"C), w h o s e 1H and 13C N M R spectra were identical with those of natural syringolide 2.8 The specific rotation of synthetic syringolide 2 ([ct]o 22 -79 ° (c=0.26, CHCI3)) was in good accord with that of natural syringolide 2 ([ct]o 24 -75.91 ° (c=0.22, CHCI3)) 2 and a preliminary experiment s h o w e d that the synthetic sample had almost the same biological activity as natural syringolide 2. These results confirmed the absolute configuration of syringolide 2 as 2 S , 3 R , 2 ' R , 3 ' S , 4 ' R .
Acknowledgements: We are grateful to Dr. S. Kawasaki, National Institute of Agrobiological Resources, for his helpful advice and bioassay. Our thanks are due to Prof. J. J. Sims, University of California, Riverside, for copies of N M R spectra of natural syringolides. References and Notes 1. Smith, M. J.; Mozzola, E. P.; Sims, J. J.; Midland, S. L.; Keen, N. T.; Burton, V.; Stayton, M. M. TetrahedronLett. 1993, 34, 223-226. 2. Midland, S. L.; Keen, N. T.; Sims, J. J.; Midland, M. M.; Stayton, M. M.; Burton, V.; Smith, M.J.; Mazzola, E. P.; Graham, K. J.; Clardy,J. J. Org. Chem. 1993, 58, 2940-2945. 3. McDougal, P. G.; Rico, 1. G.; Oh, Y.-I.; Condon, B. D. J. Org. Chem. 1986, 51, 3388-3390. 4. lida, H.; Yamazaki, N.; Kibayashi, C. J. Org. Chem. 1986, 51, 1069-1073. 5. Still, W. C. J. Am. Chem. Soc. 1978, 100, 1481-1487. 6. 6b: IR (film) Vmax 3485, 1725 era-l; 1H NMR (CDCI3) b 0.07 (3H, s), 0.08 (3H, s), 0.89 (9H, s), 3.04 (1H, t, J=5.0 Hz, OH), 3.30 (3H, s), 3.43 (3H, s), 3.70 (1H, dd, J=8.0, 10.0 Hz), 3.76 (IH, dd, J=5.5, 10.0 I-Iz), 4.01 (1H, ddd, J=3.0, 5.5, 8.0 Hz), 4.43 (1H, d, J=3.0 Hz), 4.50 (1H, dd, J=5.0, 19.5 Hz), 4.56 (1H, d, J=7.0 Hz), 4.58 (1H, dd, J=5.0, 1%5 Hz), 4.65 (1H, d, J=7.0 Hz), 4.74 (2H, s). 8: IR (film) Vmax 1770, 1690, 1625 era-l; 1H NMR (CDCI3) 6 0.08 (3H, s), 0.12 (3H, s), 0.88 (3H, t, J=7 Hz), 0.92 (9H, s), 1.23-1.36 (8H, m), 1.58-1.64 (2H, m), 2.90-3.02 (2H, m), 3.24 (3H, s), 3.37 (3H, s), 3.78 (IH, rid, J=7.5, 10.0 I--tz), 3.82 (1H, dd, J=6.0, 10.0 Hz), 4.01 (1H, ddd, J=2.5, 6.0, 7.5 Hz), 4.52 (1H, d, J=7.0 Hz), 4.64 (1H, d, J=7.0 Hz), 4.66 (1H, d, J=7.0 Hz), 4.68 (1H, d, J=7.0 I-lz), 4.98 (1H, d, J=19.5 Hz), 5.11 (1H, dd, J=l.5, 19.5 Hz), 5.48 (1H, dd, J=l.5, 2.5 Hz). 7. This acid was prepared by y-alkylation of the dianion of methyl acetoacetate with 1-iodohexane followed by saponification with aq. KOH in EtOH. Epstein, J; Cannon, P., Jr.; Swidler, R.; Baraze, A. J. Org. Chem. 1977, 42, 759762. 8. The melting point of our synthetic syringolide 2 was somewhat lower than the reported value (m.p. 123- 124"C),2 although the synthetic sample seemed to be very pure as judged by its 1H and 13C NMR spectra. We could not purify it further owing to its instability on our recrystallization conditions.
(Received in Japan 6 January 1995; revised 2 March 1995; accepted 9 March 1995)
Pergamon
ElsevierScienceLtd Printedin Great Britain 0040-4039/95$9.50+0.00 0040-4039(95)00512-9
Synthesis and Absolute Configuration of S y r i n g o l i d e 2, an Elicitor from Pseudomonas syringae pv. tomato
Shigefumi Kuwahara,* Masahiko Moriguchi, Kazuhiro Miyagawa, Masako Konno, and 0samu Kodama Laboratoryof AgriculturalChemicals, Faculty of Agriculture,IbarakiUniversity,Ami-machi,Inashiki-gun,lbaraki300-03, Japan
Abstract: The enantioselectivesynthesis of syringolide 2, one of the two elicitors produced by Pseudomonas syringae pv. tomato, was accomplishedin 11 steps from dielhyll~-tartrate.
In 1993 Keen et aL isolated two C-glycosides possessing a new ring system as specific elicitors from Pseudomonas syringae pv. tomato and named them syringolide 1 (1) and syringolide 2 (2).1, 2 Their absolute configurations were deduced from the assumption that the incorporated xylulose moiety should be the naturally occurring D-form. These elicitors are produced through the action of avirulence gene (avrD) of P. syringae pv. tomato and function as specific molecular signals which cause hypersensitive responses specifically in soybean plants carrying the Rpg4 disease resistance gene. 2 Much interest is now being attracted in the particular receptor protein which may recognize the syringolides as specific signals, and the synthesis of radiolabelled syfingolides is expected to provide a powerful tool for identifying the receptor. First of all, we carried out the synthesis of the naturally occurring enantiomer of syringolide 2 in order to establish the synthetic route leading to the syringolides.
HO H 0
0
R' " ' ~ ' ~ H "" o HO H
0
n-CTH15"~v ~ 0 o . ~ OH 3,J~ , HO'" 5
1: R=n-CsHn
OH
2:R=n-C7H15
3
According to the biosynthetic pathway proposed by Keen et al., 1,2 we chose 3 as the key intermediate of our synthesis of 2. The intramolecular Knoevenagel condensation of 3 followed by conjugate addition of 5'OH to the resulting ct-acyl-ct,13-unsatumted lactone and hemiacetalization of T-OH to C3-carbonyl function should give rise to 2 judging from the generality that the c/s-fused oxabicyclo[3.3.0]octane system is thermodynamically more favored than the corresponding tram-isomer. Our synthesis began with the selective protection 3 of the known diol 4a 4 obtained almost quantitatively from diethyl D-tam'ate in 2 steps (Scheme 1). The Swern oxidation of the resulting alcohol 4b gave 5, which was then converted into protected D-xylulose 6a by the addition of (1-ethoxyethoxy)methyUithium5 followed by the Swern oxidation. Selective deprotection of 6a gave 6b, 6 which was esterified with 3-oxodecanoic acid 7 to afford 7, a protected form of 3. The Knoevenagel condensation of 7 into 8 was achieved by simply mixing 7 with silica gel in n-hexane-EtOAc.6 Although the direct conversion of 8 into 2 using aqueous acidic conditions
3201
3202
OMOM
RO~o
H
b)
OMOM
~ a) 4b R=TBS 4a R=H
OMOM TBSO,v~CHO OMOM
c), d ) ~
MOMO 0 TBSO 1 ~ , .OR OMOM
5
6aR=EE 6b R=H
O
O
0
0
n.CrHt51Jl"-.~ 0 n-C-,,H.~s~ e) ~ 0..~ OTBS f) ~ MOMO",..I.y MOMO'÷'~ J
7
OMOM
MOMO¢-t-,,,.~OTBS
MeO u
g)
0
n'C7H15' ( ~~" O ~-
H" "(3 HO H
8
9
Scheme 1. a) Nail, TBSCI, 0"C, 30 min, it, 30 rain, 91% ; b) Swern oxidation, -78'C, 2h; c) Bu3SnCH2OEE, n-BuLi, THF, -78"C, lh, then Swern oxidation, 70% from 4b d) PITS, EtOH, rt, 2.5h, 90%; e) 3-oxodecanoic acid, DCC, DMAP, CH2C12, rt, lh; f) SiO2 (Merck Kieselgel 60 Art. 7734, 30 g / g of 7), n-hexane-EtOAc (8:1), rt, 24h, 56% from 6b, g) Dowex 50W-X8 (100 g / g of 8), MeOH, rt, 34h, 36%. was unsuccessful, the methyl ether of 2 (i.e. 9) was obtained in about 35 % yield by treating with Dowex 50X8 or Amberlyst 15E in dry methanol. Finally, 9 was treated with p - T s O H in acetone-water for 16h at room temperature to give 2 in 5 1 % yield as colorless needles (m.p. 118-120.5"C), w h o s e 1H and 13C N M R spectra were identical with those of natural syringolide 2.8 The specific rotation of synthetic syringolide 2 ([ct]o 22 -79 ° (c=0.26, CHCI3)) was in good accord with that of natural syringolide 2 ([ct]o 24 -75.91 ° (c=0.22, CHCI3)) 2 and a preliminary experiment s h o w e d that the synthetic sample had almost the same biological activity as natural syringolide 2. These results confirmed the absolute configuration of syringolide 2 as 2 S , 3 R , 2 ' R , 3 ' S , 4 ' R .
Acknowledgements: We are grateful to Dr. S. Kawasaki, National Institute of Agrobiological Resources, for his helpful advice and bioassay. Our thanks are due to Prof. J. J. Sims, University of California, Riverside, for copies of N M R spectra of natural syringolides. References and Notes 1. Smith, M. J.; Mozzola, E. P.; Sims, J. J.; Midland, S. L.; Keen, N. T.; Burton, V.; Stayton, M. M. TetrahedronLett. 1993, 34, 223-226. 2. Midland, S. L.; Keen, N. T.; Sims, J. J.; Midland, M. M.; Stayton, M. M.; Burton, V.; Smith, M.J.; Mazzola, E. P.; Graham, K. J.; Clardy,J. J. Org. Chem. 1993, 58, 2940-2945. 3. McDougal, P. G.; Rico, 1. G.; Oh, Y.-I.; Condon, B. D. J. Org. Chem. 1986, 51, 3388-3390. 4. lida, H.; Yamazaki, N.; Kibayashi, C. J. Org. Chem. 1986, 51, 1069-1073. 5. Still, W. C. J. Am. Chem. Soc. 1978, 100, 1481-1487. 6. 6b: IR (film) Vmax 3485, 1725 era-l; 1H NMR (CDCI3) b 0.07 (3H, s), 0.08 (3H, s), 0.89 (9H, s), 3.04 (1H, t, J=5.0 Hz, OH), 3.30 (3H, s), 3.43 (3H, s), 3.70 (1H, dd, J=8.0, 10.0 Hz), 3.76 (IH, dd, J=5.5, 10.0 I-Iz), 4.01 (1H, ddd, J=3.0, 5.5, 8.0 Hz), 4.43 (1H, d, J=3.0 Hz), 4.50 (1H, dd, J=5.0, 19.5 Hz), 4.56 (1H, d, J=7.0 Hz), 4.58 (1H, dd, J=5.0, 1%5 Hz), 4.65 (1H, d, J=7.0 Hz), 4.74 (2H, s). 8: IR (film) Vmax 1770, 1690, 1625 era-l; 1H NMR (CDCI3) 6 0.08 (3H, s), 0.12 (3H, s), 0.88 (3H, t, J=7 Hz), 0.92 (9H, s), 1.23-1.36 (8H, m), 1.58-1.64 (2H, m), 2.90-3.02 (2H, m), 3.24 (3H, s), 3.37 (3H, s), 3.78 (IH, rid, J=7.5, 10.0 I--tz), 3.82 (1H, dd, J=6.0, 10.0 Hz), 4.01 (1H, ddd, J=2.5, 6.0, 7.5 Hz), 4.52 (1H, d, J=7.0 Hz), 4.64 (1H, d, J=7.0 Hz), 4.66 (1H, d, J=7.0 Hz), 4.68 (1H, d, J=7.0 I-lz), 4.98 (1H, d, J=19.5 Hz), 5.11 (1H, dd, J=l.5, 19.5 Hz), 5.48 (1H, dd, J=l.5, 2.5 Hz). 7. This acid was prepared by y-alkylation of the dianion of methyl acetoacetate with 1-iodohexane followed by saponification with aq. KOH in EtOH. Epstein, J; Cannon, P., Jr.; Swidler, R.; Baraze, A. J. Org. Chem. 1977, 42, 759762. 8. The melting point of our synthetic syringolide 2 was somewhat lower than the reported value (m.p. 123- 124"C),2 although the synthetic sample seemed to be very pure as judged by its 1H and 13C NMR spectra. We could not purify it further owing to its instability on our recrystallization conditions.
(Received in Japan 6 January 1995; revised 2 March 1995; accepted 9 March 1995)