Syntheses of dinor-cis/iso-12-oxo-phytodienoic acid (dn-cis/iso-OPDAs), ancestral jasmonate phytohormones of the bryophyte Marchantia polymorpha L., and their catabolites

Abstract

In recent years, the biology of the evolutionary origin of phytohormone signaling has made significant progress. Among them, the ligand-receptor co-evolution found in jasmonate signaling has attracted the attention of plant scientists. Dinor-cis-12-oxo-phytodienoic acid (dn-cis-OPDA, 4) and dn-iso-OPDA (5) are ancestral plant hormones of the bryophyte Marchantia polymorpha L. We succeeded in the first practical synthetic supply of these hormones as well as their possible catabolites. These compounds are expected to be useful in the study of ancestral jasmonate signaling in bryophytes.

Introduction

(+)-Jasmonoyl-l-isoleucine (JA-Ile, 1) is a lipid-derived plant phytohormone, implicated in the regulation of plant growth, fertility, and defense against pathogens and insects1,2,3. JA-Ile-mediated signal transduction depends on the COI1-JAZ co-receptor system, composed of an F-box protein CORONATINE INSENSITIVE 1 (COI1) and JASMONATE ZIM-DOMAIN (JAZ) repressor protein4,5,6. After signal transduction, 1 is catabolized into 12-hydroxy-JA-Ile (12-OH-JA-Ile, 2) by CYP94B1/B3, then 12-carboxy-JA-Ile (12-COOH-JA-Ile, 3) by CYP94C1, and then deactivated within a few hours (Fig. 1)7,8,9. Biological studies of the evolution of phytohormone signaling is an important topic in plant science10,11,12,13, and recent achievements in the genomic sequencing of a myriad of plant species have enabled research into the evolutionary origins of phytohormones. Marchantia polymorpha L., a type of bryophyte, has attracted a great deal of attention due to its ancestral signaling module14, and its jasmonate signaling constitutes an intriguing example of ligand-receptor co-evolution, depending on the MpCOI1-MpJAZ co-receptor system (Fig. 1)15. However, JA-Ile, the usual ligand for the COI1-JAZ co-receptor system of vascular plants, cannot be perceived by MpCOI1-MpJAZ–dinor-cis-12-oxo-phytodienoic acid (dn-cis-OPDA, 4) and dn-iso-OPDA (5) are the ligands of MpCOI1-MpJAZ co-receptor instead (Fig. 1). Genetic studies have revealed that the ligand-receptor pair of dn-cis/iso-OPDA and MpCOI1-MpJAZ participates in all the jasmonate responses of M. polymorpha, including defense responses16,17,18. However, biological studies require samples of 4 and 5—in the past, 4 has been synthesized enzymatically19 but is now out of stock; and the only known synthesis of 5 entails an electroorganic reaction20 that cannot be accomplished using normal laboratory equipment (Scheme S1). Accordingly, we developed and report herein the first chemical synthesis of 4 and the first non-electroorganic synthesis of 5. In addition, we synthesized their potent catabolites 16-hydroxy-dinor-cis-OPDA (16-OH-dn-cis-OPDA, 6), 16-carboxy-dinor-cis-OPDA (16-COOH-dn-cis-OPDA, 7), 16-hydroxy-dinor-iso-OPDA (16-OH-dn-iso-OPDA, 8), 16-carboxy-dinor-iso-OPDA (16-COOH-dn-iso-OPDA, 9) (Fig. 1). These potent catabolites will enable further studies on the catabolism of ancestral plant hormones.

Figure 1
figure1

Ligand-receptor co-evolution of jasmonates, JA-Ile (1) in Arabidopsis thaliana, dn-cis-OPDA (4) and dn-iso-OPDA (5) in Marchantia polymorpha L., and their potent catabolites.

Results and discussion

Synthesis of dn-cis-OPDA (4) and its potent catabolites (6 and 7)

Our plan for the synthesis of dn-cis-OPDA (4) and its catabolites (6 and 7) is outlined in Scheme 1. A major initial concern in the syntheses of 4, 6 and 7 was the avoidance of epimerization at C11 which was anticipated to be facile in the presence of either acid or base based on the ready epimerization of the structurally similar 1 to give the more thermodynamically stable trans-1 in a ratio of trans:cis = 95:521. We therefore planned to synthesize 4 according to a procedure similar to that used to synthesize OPDA (10), a congener of 4, developed by Kobayashi et al.22. They avoided epimerization by introducing the ketone at the late stage of synthesis23. Compounds 4 and 6 would be obtained from a common intermediate 11 by Wittig reaction using a different phosphonium salt (Scheme 1), and 7 would be obtained by oxidation of 6.

Scheme 1
scheme1

Synthetic plan for dn-cis-OPDA (4) and its potent catabolites (6 and 7).

Our synthesis of dn-cis-OPDA (4) is summarized in Scheme 2. Allylic substitution of monoacetate 15, prepared by enzymatic reaction22, with TBDPS(CH2)6MgCl, was performed in the presence of CuCN to afford 16. The Mitsunobu reaction of 16, hydrolysis of resulting acetate and Eschenmoser–Claisen rearrangement gave dimethylamide 17. Iodolactonization, elimination with DBU and subsequent reduction with LiAlH4 afforded diol 18. In the iodolactonization step, use of water in place of buffer resulted in the removal of TBDPS group. TES protection of diol 18 and regioselective Swern oxidation of the primary TESOCH2 group24 afforded the common intermediate 11. In pursuit of 4, 11 was treated with [Ph3PPr]+Br and NaHMDS to give diene 20. Deprotection of the silyl groups of 20 with TBAF and subsequent Jones oxidation at − 20 °C afforded dn-cis-OPDA (4, 15 mg in 12 steps and 22% overall yield from 15). In the work up of final Jones oxidation, removal of chromium compounds and sulfuric acid using silica gel caused epimerization of C11 (probably due to exothermic adsorption of sulfuric acid on silica gel)22, but this could be easily avoided by removal of the inorganic substances with water instead. The obtained 4 was quite stable at room temperature under neutral condition and no epimerization at C11 was observed even after 2 weeks25,26,27.

Scheme 2
scheme2

Synthesis of dn-cis-OPDA (4). Reagents and conditions: (a) TBDPSO(CH2)6MgCl,CuCN, THF, − 20 °C; (b) PPh3, AcOH, DIAD, toluene, − 20 °C; (c) LiOH, THF, MeOH, H2O; (d) MeC(OMe)2NMe2, xylene, reflux, 50% (4 steps); (e) I2, buffer (pH 5.0), THF; (f) DBU, THF, reflux; LiAlH4, 78% (2 steps); (g) TESCl, imadizole, DMF, 92%; (h) (COCl)2, DMSO; NEt3, CH2Cl2; (i) [Ph3PPr]+Br, NaHMDS, THF, 71% (2 steps); (j) TBAF, THF, reflux, 91%; (k) Jones reagent, acetone, − 20 °C, 97%.

For the syntheses of 16-OH-dn-cis-OPDA (6) and 16-COOH-dn-cis-OPDA (7), [THPO(CH2)3PPh3]+Br was used in the Wittig reaction with common intermediate 11, leading to diene 21. Deprotection of the silyl groups of 21 with TBAF afforded diol 22, from which the target products 6 and 7 could be obtained by oxidation and deprotection, or vice versa (Scheme 3). Jones oxidation of 22 and subsequent THP deprotection using MgBr2 afforded 16-OH-dn-cis-OPDA (6, 41 mg in 13 steps and 14% overall yield from 15). In the THP deprotection step, some epimerization of C11 took place (cis:trans = ca. 94:6 by NMR). This epimerization and observed diastereomeric ratio were in good accordance with previous literature (cis:trans = ca. 92:8 in NMR)22. cis- and trans-6 were easily separated by chiral HPLC using a CHIRALPAK IA column to obtain pure cis-6 (22.3 mg). Conversely, THP deprotection of 22 using PPTS afforded triol 23, Jones oxidation of which afforded 16-COOH-dn-cis-OPDA (7, 8.3 mg in 13 steps and 7.5% overall yield from 15).

Scheme 3
scheme3

Synthesis of 16-OH-dn-cis-OPDA (6) and 16-COOH-dn-cis-OPDA (7). Reagents and conditions: (a) (COCl)2, DMSO, NEt3, CH2Cl2; (b) [THPO(CH2)3PPh3]+Br-, NaHMDS, THF, 76% (2 steps); (c) TBAF, THF 72%; (d) Jones reagent, acetone, − 20 °C, (e) MgBr2, Et2O, 72% (2 steps); (f) PPTS, MeOH, 35 °C, 54%; (g) Jones reagent, acetone, − 20 °C, 73%.

Synthesis of dn-iso-OPDA (5) and its potent catabolites (8 and 9)

Our plan for the synthesis of dn-iso-OPDA (5) and its potent catabolites (8 and 9) is shown in Scheme 4. In the synthesis of [2H2]-tetrahydrodicanenone (iso-OPDA), Lauchli and Boland Introduced the C1–8 side chain by the 1,4-addition using an organozinc reagent and CuCN28. However, organozinc reagent are difficult to prepare and CuCN is highly toxic. In contrast, Grignard reagents used for the 1,2-addition are easy to prepare and less toxic than CuCN. And potent catabolites 16-OH-dn-iso-OPDA (8) and 16-COOH-dn-iso-OPDA (9) could be obtained from the same starting material 24 by using a different allyl bromide.

Scheme 4
scheme4

Synthetic plan for dn-iso-OPDA (5) and its potent catabolites (8 and 9).

Our synthesis of dn-iso-OPDA (5) is summarized in Scheme 5. Allylation of 1,3-cyclopentanedione 24 followed by methylation of the resulting 27 gave cyclopentenone 28. After the Grignard reaction of 28, dilution of the reaction mixture with hydrochloric acid promoted hydrolysis of the enol ether and the deprotection of the THP group, to give alcohol 30. Finally, Jones oxidation of 30 gave 61 mg of dn-iso-OPDA (5) in only 4 steps from 24.

Scheme 5
scheme5

Synthesis of dn-cis-OPDA (5). Reagents and conditions: (a) cis-1-bromopent-2-ene, K2CO3, H2O, 60 °C, 25%; (b) Me2SO4, K2CO3, acetone, reflux, quant.; (c) THPO(CH2)6MgBr, THF, reflux (d) 2 M HCl aq., 24%; (e) Jones reagent, acetone, 0 °C, 89%.

Next, we synthesized 16-OH-dn-iso-OPDA (8) and 16-COOH-dn-iso-OPDA (9) (Schemes 6, 7). Introduction of the C12–C16 side chain of 24 was first attempted by allylation, but the side chain could not be directly introduced using allylation in order to O-alkylation. Accordingly, we abandoned this approach and sought to construct the C12–C16 side chain by Z-selective cross metathesis29,30,31. Pd-mediated allylation32 of 24 and subsequent methylation gave allylcyclopentenone 32. Introduction of the C1–C6 side chain using THPO(CH2)6MgBr followed by hydrolysis and deprotection of THP group gave alcohol 33. Finally, Jones oxidation of 33 followed by cross metathesis with CH2=CHCH2CH2OAc and deprotection of the acetyl group gave 16-OH-dn-iso-OPDA (8, 15.6 mg in 7 steps and 11% overall yield from 24) as a Z/E mixture (10:1 Z/E). Z/E isomers were easily separated by chiral HPLC using a CHIRALPAK IA column to obtain pure Z-8 (2.4 mg). 16-COOH-dn-iso-OPDA (9, 12:1 Z/E, 6.3 mg) was also obtained from allylcyclopentenone 33 by cross metathesis with CH2=CHCH2CH2OAc, hydrolysis of acetyl group and Jones oxidation (total 7 steps and 3.4% overall yield from 24). Z/E isomers were easily separated by HPLC to obtain pure Z-9 (2.6 mg).

Scheme 6
scheme6

Synthesis of 16-OH dn-iso-OPDA (8). Reagents and conditions: (a) allyl acetate, [Pd(C3H5)Cl]2, dppe, BSA, NaOAc, THF, reflux, 81%; (b) Me2SO4, K2CO3, acetone, reflux, 98%; (c) THPO(CH2)6MgBr, THF, reflux; (d) 1 M HCl aq., reflux, 36% (2 steps); (e) Hoveyda–Grubbs catalyst M2001 Umicore, CH2 = CHCH2CH2OAc; (f) Jones reagent, acetone, − 20 °C; (g) LiOH, H2O, THF, 40% (3 steps).

Scheme 7
scheme7

Synthesis of 16-COOH-dn-iso-OPDA (9). Reagents and conditions: (a) Hoveyda-Grubbs Catalyst M2001 Umicore, CH2=CHCH2CH2OAc. (b) NaOH, H2O, MeOH, 50 °C 32% (2 steps). (c) Jones reagent, acetone, − 20 °C, 36%.

Conclusions

A synthetic supply of jasmonates is indispensable if their study is to be advanced20,30,33,34, and this work finally enables dn-cis/iso-OPDAs (4 and 5) and their potent catabolites (69) to be readily obtained. Our work is expected to accelerate biological studies of the signaling mechanisms of bryophyte hormones, which should lead to a better understanding of the evolutional origins of phytohormone signaling. In particular, study of the catabolism of 4 and 5 should provide insights into the deactivation mechanism of ancestral plant hormones. Biological studies using synthetic 49 are now in progress.

Method

Synthesis of dn-cis-OPDA (4)

To a solution of 20 (57.2 mg, 94.5 µmol) in THF (20 mL) was added 1 M TBAF in THF (1.6 mL, 1.60 mmol). After being stirred at reflux temperature for 1.5 h, the solvent was removed under reduced pressure. The residue was purified by medium-pressure chromatography (Isolera, eluent: 93:7 n-hexane/EtOAc to 40:60 n-hexane/EtOAc) to give a diol intermediate (22.3 mg, 94%) as a colorless oil. [α]D22 + 63.6 (c 1.09, CHCl3). 1H NMR (400 MHz, CDCl3) δH: 6.22 (dd, J = 5.8, 2.6 Hz, 1H), 5.96 (brd, J = 5.8 Hz, 1H), 5.49–5.38 (m, 2H), 4.51 (dd, J = 5.8, 2.6 Hz, 1H), 3.63 (t, J = 6.6 Hz, 2H), 2.52–2.43 (m, 1H), 2.37–2.27 (m, 1H), 2.24–2.04 (m, 4H), 1.66–1.52 (m, 3H), 1.47–1.20 (m, 7H), 0.99 (t, J = 7.5 Hz, 3H); 13C NMR (100 MHz, CDCl3) δC: 141.64, 132.46, 132.01, 127.88, 76.55, 62.99, 46.13, 46.00, 33.52, 32.73, 29.70, 28.05, 25.69, 23.08, 20.78, 14.26; IR (neat) cm–1: 3356, 2931, 2861, 1458, 1053; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H28NaO2: 275.1987, Found: 275.1976.

To a solution of diol intermediate (15.3 mg, 60.6 µmol) in acetone (5.2 mL) was added Jones reagent (4.0 M solution) at − 20 °C until the orange color of the reagent persisted (30 drops). After 10 min of stirring at − 20 °C, i-PrOH was added to quench the remaining reagent. Then, EtOAc/n-hexane (1/1, 20 mL) and H2O (20 mL) were added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by medium-pressure chromatography (Isolera, eluent: 0.1:88:12 AcOH/n-hexane/EtOAc to 0.1:99.9 AcOH/EtOAc) to give 4 (116 mg, 97%) as a colorless oil. Diastereomeric purity of 4 was > 99% by 1H NMR spectroscopy (δH = 7.72 (dd, J = 5.8, 2.7 Hz, 1H) for 4; 7.59 (dd, J = 5.8, 2.6 Hz, 1 H) for the trans isomer). [α]D22 + 135.5 (c 0.75, CHCl3). 1H NMR (400 MHz, CDCl3) δH: 7.72 (dd, J = 5.8, 2.7 Hz, 1H), 6.18 (dd, J = 5.8, 1.6 Hz, 1H), 5.48–5.31 (m, 2H), 3.04–2.94 (m, 1H), 2.56–2.41 (m, 2H), 2.36 (brt, J = 7.1 Hz, 2H), 2.13 (dt, J = 15.8, 7.8 Hz, 1H), 2.05 (quintet, J = 7.5 Hz, 2H), 1.80–1.69 (m, 1H), 1.63 (brt, J = 7.1 Hz, 2H), 1.52–1.29 (m, 4H), 1.23–1.11 (m, 1H), 0.97 (t, J = 7.5 Hz, 3H); 13C NMR (100 MHz, CDCl3) δC: 210.86, 179.32, 166.96, 133.02, 132.50, 126.82, 49.79, 44.20, 33.84, 30.58, 29.18, 27.32, 24.49, 23.76, 20.78, 13.99; IR (neat) cm–1: 3487, 3178, 2935, 1709; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H24NaO3: 287.1623, Found: 287.1618.

Synthesis of 16-OH-dn-cis-OPDA (6)

To a solution of 22 (71.0 mg, 201 µmol) in acetone (20 mL) was added Jones reagent (4.0 M solution, 400 µL, 1.60 mmol) at − 20 °C. After 20 min of stirring at − 20 °C, i-PrOH was added to quench the remaining reagent. Then, EtOAc/n-hexane (1/1, 10 mL) and H2O (20 mL) were added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and filtered. The reaction mixture was concentrated under reduced pressure to give a carboxylic acid intermediate (86.3 mg, mixture) as a colorless oil. The crude product was used for the next reaction without further purification. To a solution of the carboxylic acid intermediate (86.3 mg, mixture) in Et2O (10 mL) was added MgBr2 (90.5 mg, 510 µmol). The solution was stirred at room temperature for 20 min and diluted with Et2O and MeOH. Then, H2O was added and the water layer was extracted with AcOH/EtOAc (1/999). The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and filtered. The residue was purified by medium-pressure chromatography (Isolera, eluent: 0.1:84:16 AcOH/n-hexane/EtOAc to 0.1:99.9 AcOH/EtOAc) to give 6 (40.8 mg, 72% cis/trans = 92/8 mixture) as a colorless oil. cis- and trans-6 were separated by chiral HPLC using a CHIRALPAK IA column (Daicel Co., Ltd., Japan, Φ20 × 250 mm, eluent: 0.1:20:80 AcOH/EtOH/n-hexane, flow rate: 8.0 mL/min, cis-6: Rt = 33.3 min, trans-6: Rt = 23.3 min) to afford pure cis-6 (22.3 mg) as a colorless oil. [α]D20 + 34.8 (c 1.95, CHCl3). 1H NMR (400 MHz, CDCl3) δH: 7.72 (dd, J = 5.8, 2.7 Hz, 1H), 6.18 (dd, J = 5.8, 1.6 Hz, 1H), 5.48–5.31 (m, 2H), 3.04–2.94 (m, 1H), 2.56–2.41 (m, 2H), 2.36 (brt, J = 7.1 Hz, 2H), 2.13 (dt, J = 15.8, 7.8 Hz, 1H), 2.05 (quintet, J = 7.5 Hz, 2H), 1.80–1.69 (m, 1H), 1.63 (brt, J = 7.1 Hz, 2H), 1.52–1.29 (m, 4H), 1.23–1.11 (m, 1H), 0.97 (t, J = 7.5 Hz, 3H); 13C NMR (100 MHz, CDCl3) δC: 210.86, 179.32, 166.96, 133.02, 132.50, 126.82, 49.79, 44.20, 33.84, 30.58, 29.18, 27.32, 24.49, 23.76, 20.78, 13.99; IR (neat) cm–1: 2935, 2873, 1107; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H24NaO4: 303.1572, Found: 303.1565.

Synthesis of 16-COOH-dn-cis-OPDA (7)

To a solution of 23 (10.3 mg, 38.4 µmol) in acetone (6.0 mL) was added Jones reagent (4.0 M solution) at − 20 °C until the orange color of the reagent persisted (37 drops). After 20 min of stirring at − 20 °C, i-PrOH was added to quench the remaining reagent. Then, EtOAc (10 mL) and H2O (60 mL) were added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by medium-pressure chromatography (Isolera, eluent: 0.1:98:2 AcOH/CHCl3/MeOH to 0.1:80:20 AcOH/CHCl3/MeOH) to give 7 (8.3 mg, 73%) as a colorless oil. Diastereomeric purity of 7 was > 98% by 1H NMR spectroscopy (δH = 7.74 (dd, J = 5.9, 2.7 Hz, 1H) for 7; 7.62 (dd, J = 5.9, 2.4 Hz, 1 H) for the trans isomer). [α]D21 + 117.6 (c 0.24, CHCl3). 1H NMR (400 MHz, CDCl3) δH: 7.74 (dd, J = 5.9, 2.7 Hz, 1H), 6.18 (dd, J = 5.9, 1.6 Hz, 1H), 5.68–5.56 (m, 2H), 3.21 (dd, J = 17.2, 5.9 Hz, 1H), 3.11 (dd, J = 17.2, 3.9 Hz, 1H), 3.04–2.93 (m, 1H), 2.56–2.46 (m, 2H), 2.36 (brt, J = 7.1 Hz, 2H), 2.20 (ddd J = 16.7, 10.7, 6.1 Hz, 1H), 1.78–1.54 (m, 3H), 1.52–1.22 (m, 4H), 1.22–1.07 (m, 1H); 13C NMR (100 MHz, CDCl3) δC: 210.45, 179.82, 177.53, 167.19, 132.47, 131.97, 121.59, 49.21, 44.26, 33.82, 32.94, 30.48, 29.03, 27.30, 24.24, 24.15; IR (neat) cm–1: 3448, 3205, 3031, 2935, 1708 (br); HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H22NaO5: 317.1365, Found: 317.1355.

Synthesis of dn-iso-OPDA (5)

To a solution of THPO(CH2)6MgBr (0.85 M in THF, 3.3 mL, 2.80 mmol) was added a solution of 28 (198 mg, 1.10 mmol) in THF (6.5 mL) at reflux temperature under argon atmosphere. After being stirred at 60 °C for 3 h, the reaction mixture was allowed to cool to rt and 2 M HCl aq. (7 mL) was added. After 1.5 h of stirring, H2O was added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. After evaporation, the residue was purified by medium-pressure chromatography (Isolera, eluent: 40:60 n-hexane/EtOAc to EtOAc) to give the oxidized compound. The compound was carried on to the next step.

To a solution of the mixture (79.6 mg) in acetone (6 mL) was added Jones reagent (4.0 M solution, 200 µL, 800 µmol) at 0 °C. After 3.5 h of stirring at 0 °C, i-PrOH was added to quench the remaining reagent. Then, H2O (20 mL) was added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by medium-pressure chromatography (Isolera, eluent: 0.5:50:50 AcOH/n-hexane/EtOAc to 0.5:99.5 AcOH/EtOAc) to give 5 (61.4 mg, 89%) as a colorless oil. 1H NMR (400 MHz, CDCl3); δH 5.37 (ddt, J = 18.0, 7.2, 1.2 Hz, 1H), 5.21 (ddt, J = 17.6 7.2, 1.2 Hz, 1H), 2.93 (d, J = 7.2 Hz, 2H), 2.50 (t, J = 4.4 Hz, 2H), 2.44 (t, J = 7.6 Hz, 2H), 2.39–2.35 (m, 4H), 2.15 (quin, J = 7.6 Hz, 2H), 1.68 (quin, J = 6.4 Hz, 2H), 1.58 (quin, J = 8.0 Hz, 2H), 1.44–1.36 (m. 2H), 0.99 (t, J = 7.6 Hz, 3H); 13C NMR (100 MHz, CDCl3); δC 209.56, 178.73, 174.01, 139.39, 132.34, 125.32, 34.19, 33.71, 31.10, 29.15, 29.12, 27.08, 24.44, 21.24, 20.61, 14.14; IR (neat) cm–1: 3502, 2935, 2866, 1697, 1631; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H24O3: 287.1623, found: 287.1601.

Synthesis of 16-OH-dn-iso-OPDA (8)

A 5 mL pear-shaped flask was charged with 33 (31.2 mg, 140 µmol), CH2 = CHCH2CH2OAc (165 mg, 144 mmol) and Hoveyda–Grubbs Catalyst M2001 Umicore (4.6 mg, 7.27 µmol). After 24 h of stirring, the residue was roughly purified by medium-pressure chromatography (Isolera, eluent: 0.1:99:1 AcOH/CHCl3/MeOH to 0.1:90:10 AcOH/CHCl3/MeOH) to give a mixture (58.2 mg). The crude mixture was used for the next reaction without further purification. To a solution of the mixture (58.2 mg) in acetone (10 mL) was added Jones reagent (4.0 M solution, 95 µL, 380 µmol) at − 20 °C. After 30 min of stirring at − 20 °C, i-PrOH was added to quench the remaining reagent. Then, EtOAc and H2O were added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. The crude mixture (33.0 mg) was used for the next reaction without further purification. To a solution of the mixture (33.0 mg) in THF (4.5 mL) was added 1 M-LiOH solution (510 µL, 510 µmol) and the mixture was stirred for 1.5 h. The reaction mixture was quenched with 1 M HCl aq. and the aqueous layer was extracted with EtOAc. The organic layer was washed with saturated aqueous NaCl, dried over Na2SO4, and filtered. After evaporation, the residue was purified medium-pressure chromatography (Isolera, eluent: 0.1:85:15 AcOH/n-hexane/EtOAc to 0.1:99.9 AcOH/ EtOAc) to give 16-OH-dn-iso-OPDA (8) (15.6 mg, 40% in 3 steps) as a Z/E mixture (Z/E = 10/1). Z/E isomers were separated by chiral HPLC using a CHIRALPAK IA column (Daicel Co., Ltd., Japan, Φ20 × 250 mm, eluent: 0.1:20:80 AcOH/EtOH/n-hexane, flow rate: 8.0 mL/min, E-8: Rt = 18.8 min, Z-8: Rt = 25.5 min) to afford pure Z-8 (2.4 mg) as a colorless oil. 1H NMR (400 MHz, CDCl3) δH: 5.43 (dt, J = 11.0, 7.2 Hz, 1H), 5.37 (dt, J = 11.0, 6.6 Hz, 1H), 3.71 (t, J = 6.2 Hz, 2H), 2.98 (d, J = 6.6 Hz, 2H), 2.52–2.50 (m, 2H), 2.49–2.44 (m, 4H) 2.38–2.36 (m, 4H), 1.69 (quintet, J = 7.6 Hz, 2H), 1.57 (quintet, J = 7.6 Hz, 2H), 1.40 (quintet, J = 7.6 Hz, 2H); 13C NMR (100 MHz, CDCl3) δC:210.23, 178.34, 174.92, 138.75, 128.63, 126.82, 61.97, 34.18, 33.69, 31.03, 30.71, 29.27, 28.94, 27.07, 24.35, 21.71; IR (neat) cm-1: 3433, 2931, 2866, 1701, 1631; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H24NaO4: 303.1572, found: 303.1565.

Synthesis of 16-COOH-dn-iso-OPDA (9)

To a solution of 35 (16.1 mg, 60.4 µmol) in acetone (1.5 mL) was added Jones reagent (4.0 M solution, 150 µL, 600 µmol) at − 20 °C. After 1.5 h of stirring at − 20 °C, i-PrOH was added to quench the remaining reagent. Then, EtOAc and H2O were added and the water layer was extracted with EtOAc. The combined organic layers were washed with saturated aqueous NaCl, dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by medium-pressure chromatography (Isolera, eluent: 0.1:98:2 AcOH/CHCl3/MeOH to 0.1:80:20 AcOH/CHCl3/MeOH) to give 16-COOH-dn-iso-OPDA (9) (6.3 mg, 36%) as a Z/E mixture (Z/E = 12/1). Z/E isomers were separated by HPLC using a Cholester column (Nacalai Tesque, Inc., Japan, Φ20 × 250 mm, eluent: 0.1:30:70 AcOH/MeCN/H2O, flow rate: 8.0 mL/min, E-9: Rt = 25.0 min, Z-9: Rt = 25.9 min) to afford pure Z-9 (2.6 mg) as a colorless oil. 1H NMR (400 MHz, CDCl3) δH: 5.57 (dtt, J = 10.6, 7.3, 1.5 Hz, 1H), 5.48 (dtt, J = 10.6, 7.4, 1.3 Hz, 1H), 3.27 (d, J = 7.3 Hz, 2H), 2.96 (d, J = 7.4 Hz, 2H), 2.53–2.51 (m, 2H), 2.44 (t, J = 7.7 Hz, 2H), 2.40–2.35 (m, 4H), 1.66 (quintet, J = 7.7 Hz, 2H), 1.55 (quintet, J = 7.7 Hz, 2H), 1.38 (quintet, J = 7.7 Hz, 2H); 13C NMR (100 MHz, CDCl3) δC: 209.72, 179.46, 177.13, 175.07, 138.16, 129.97, 121.01, 34.13, 33.71, 32.95, 31.08, 29.27, 28.94, 27.05, 24.23, 21.58; IR (neat) cm−1: 3518, 3190, 2935, 1705, 1628; HRMS (ESI, positive) m/z [M + Na]+ Calcd. for C16H22NaO5: 317.1365, found: 317.1340.

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Acknowledgements

We thank to Prof. Roberto Solano (CNB, Spain) to let us know the significance of these important ancestral hormones. This work was financially supported by a Grant-in-Aid for Scientific Research for MU from JSPS, Japan (Nos. 17H06407, 18KK0162, and 20H00402), for NK (no. 19K06968), Grant-in-Aid for Young Scientists (No. 20K15404) for TK, SUNBOR GRANT (NK), JSPS A3 Foresight Program (MU), and JSPS Core-to-Core Program Asian Chemical Biology Initiative (MU).

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M.U. conceived, designed, and coordinated the research project. N.K. designed the synthetic route of all compounds. M.U., N.K. and T.K. wrote the main manuscript text and all figures. J.W. T.K. and N.K. synthesized dn-cis-12-OPDA and the derivatives, and H.S. and N.K. synthesized dn-iso-12-OPDA and the derivatives. All authors reviewed the manuscript.

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Correspondence to Nobuki Kato or Minoru Ueda.

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Wang, J., Sakurai, H., Kato, N. et al. Syntheses of dinor-cis/iso-12-oxo-phytodienoic acid (dn-cis/iso-OPDAs), ancestral jasmonate phytohormones of the bryophyte Marchantia polymorpha L., and their catabolites. Sci Rep 11, 2033 (2021). https://doi.org/10.1038/s41598-021-81575-z

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