Asymmetric Total Syntheses and Structure Elucidations of (+)-Eurotiumide F and (+)-Eurotiumide G

Abstract

Asymmetric total syntheses of dihydropyran containing natural products, (+)-eurotiumide F and (+)-eurotiumide G have been described. These total syntheses revealed the absolute configuration of eurotiumide F and G, and confirmed the reported structure of eurotiumide F and revised the reported structure of eurotiumide G. Highlight of these syntheses is thermal rearrangement with 4-methoxyisochroman-1-one derivative having propargyl ether on phenolic ether under thermal condition to construct dihydropyran ring. X-Ray crystallographic analysis of (+)-eurotiumide G clarified the stereochemistry at the C1-position.

Introduction

Marine-derived fungi have been recognized as excellent sources of a wide variety classes of secondary metabolites, and a number of useful natural products are isolated and identified by the efforts of a lot of researchers in the field of natural products chemistry.^1–3) It is easy to understand that these natural products have a huge potential to become drugs from the fact that some of marine-derived natural products, such as eribulin^4–6) and trabectedin^7,8) have been used as in clinical sites today. In 2014, Wang and colleagues isolated and identified some dihydroisocoumarin derivatives and related compounds as racemate, named eurotiumides, from gorgonian-derived fungus, Eurotium sp. XS-200900E6⁹⁾ (Fig. 1). Among them, they reported that eurotiumide A (1) displayed potent antimicrobial activities against several bacteria, and eurotiumide B (2) exhibited the useful antifouling activity against barnacle Balanus amphitrite. Wang and colleagues separated the racemic eurotiumides by chiral HPLC and obtained both enantiomers of eurotiumide A (1), B (2), C, D, and E, respectively. The absolute configurations of their C3-position were proposed by the comparison of the related known compounds.^10–12) Recently, we reported the first asymmetric total syntheses of eurotiumide A (1) and B (2), and determined the absolute configuration of both two compounds.¹³⁾ Moreover, our total syntheses also revised the relative stereochemistry between C3 and C4 of eurotiumide A (1) and eurotiumide B (2), respectively. On the other hand, eurotiumide F (3) and eurotiumide G (4), which have dihydropyran ring and methoxy lactol moiety, were also isolated as racemate from the same fungus. However, chiral HPLC separation of these compounds could not be succeeded and the absolute configurations and optical properties of them are still unknown. Natural products having dihydropyran ring moiety are reported that they displayed unique biological activities.^14–20) Eurotiumide F (3) and G (4) were evaluated the antimicrobial activities and they displayed medium antimicrobial activities against several strains. As our continuous research, we conducted the asymmetric total syntheses of eurotiumide F (3) and eurotiumide G (4) for elucidating the asymmetric centers of C3- and C4-positions of them, and evaluate the further investigation of biological activities.

Fig. 1. Reported Structures of Eurotiumide A, B, F, and G

Results

Our synthetic plan is shown in Chart 1. Methoxy lactol moiety, which is the common characteristic structure of eurotiumide F (3) and G (4), could be constructed by reduction of the corresponding lactones 5 and 6, and subsequent replacement of the hydroxy group of lactol to methoxy group in acidic MeOH. 5 and 6 are diastereomers at the C3-position, and 5 would be easily obtained from 6 by the sequential hydrolysis and intramolecular Mitsunobu reaction. Dihydropyran moiety of 6 could be constructed by thermal rearrangement with dimethyl propargyl ether compound 7, which would be obtained from cis 4-methoxyisochroman-1-one compound 8 having (3S,4S) stereochemistry.

Chart 1. Retrosynthesis of Eurotiumide F and G

In our recent total syntheses of (–)-eurotiumide A (1) and (+)-eurotiumide B (2),¹³⁾ 8 was one of the key compound which could be prepared in decagram scale and good enantiomeric purity in 4 steps from aldehyde 9^21,22) and 1-phenyl-1H-tetrazol-5-yl (PT)-sulfone 10, by Julia–Kocienski olefination, asymmetric Shi epoxidation,²³⁾ BF₃-OEt₂ mediated epoxide opening, and Pd-catalyzed C1 insertion/lactonization cascade reaction^24–27) (Chart 2). The propargyl moiety was introduced by the treatment of 8 with 3-chloro-3-methylbut-1-yne in the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) and catalytic amount of CuCl₂ to afford propargyl derivative 7 in good yield.²⁸⁾

Chart 2. Preparation of Propargyl Derivative 7

With the desired propargyl derivative 7 in hand, we turned our attention to construction of the dihydropyran ring with thermal rearrangement^29–31) (Table 1). First, we heated 7 in benzene under reflux condition, however, the desired dihydropyran derivative 6 was not obtained at all. Next, we changed solvent from benzene to toluene. As a result, 6 was obtained in 65% (entry 2). Moreover, using xylene as a solvent improved the yield (90%) and shorten the reaction time (entry 3). Finally, we found that treatment of 7 in 1,2-dichlorobenzene at 160°C for 3 h gave the desired 6 in quantitative yield (entry 4). We also tried to use microwave condition (200°C, 300W), and observed the dramatically acceleration effect of this thermal rearrangement, and the reaction completed within 10 min in quantitative yield (entry 5).

Table 1. Examination of Thermal Rearrangement

Entry	Solvent	Temp. (°C)	Time (h)	Yield (%)
1	Benzene	Reflux (80)	48	N.R.
2	Toluene	Reflux (110)	72	65
3	Xylene	Reflux (140)	6	90
4	o-Dichlorobenzene	Reflux (160)	3	Quant
5	o-Dichlorobenzene	200 (MW condition)	0.2	Quant

Next, we constructed the lactol moiety by the reduction of lactone 6 (Chart 3). Treatment of 6 with 1 equiv of diisobutylaluminium hydride (DIBAL) afforded the desired lactol 11 as a sole isomer in good yield. The stereochemistry at the C1-position was assigned as R configuration by the careful nuclear Overhauser effect spectroscopy (NOESY) experiment between 1-OH and H-3, and this C1-stereochemistry is considered to be the result of the convergence to the more thermodynamically stable configuration during the treatment of the saturated aqueous Rochelle salt solution (see details in Experimental).³²⁾ Finally, removal of methoxymethyl (MOM) group and displacement of the hydroxy group at the C1-position to a methoxy group were accomplished by 6 M HCl aq in MeOH to afford (+)-eurotiumide G (4) in good yield. All spectral data of our synthetic eurotiumide G are identified with those of the reported data, however, the X-ray crystallographic analysis³³⁾ of the synthetic (+)-eurotiumide G revealed the stereochemistry of the C1-position as R configuration, not as S configuration reported by Wang et al. and we could determine the all stereochemistries of (+)-eurotiumide G (4) as (1R,3S,4S).

Chart 3. Completion of Total Syntheses of (+)-Eurotiumide G (4) and (–)-Eurotiumide G (13)

(Color figure can be accessed in the online version.)

Toward synthesis of eurotiumide F (3), first, we transformed cis configuration of the C3/C4-positions to trans configuration of them. Hydrolysis of 6 and subsequent intramolecular Mitsunobu reaction gave the corresponding trans 4-methoxyisochroman-1-one compound 5. Reduction of lactone moiety with DIBAL afforded lactol 12 in 79% yield. The stereochemistry among C1-, C3-, and C4-positions were determined as shown in Chart 3 by the NOESY experiment and this C1-stereoselectivity would be attributed that DIBAL reduced the lactone moiety of 5 from the opposite side of the 4-OMe group. However, treatment of 12 with 6 M HCl aq in MeOH for displacement of hydroxyl group at the C1-position to a methoxy group and the subsequent deprotection of MOM group did not give eurotiumide F (3), but (–)-eurotiumide G (13) was obtained.

The plausible mechanism of generation of 13 is shown in Chart 4. The C1-lactol moiety of 12 is easily transformed to oxonium cation intermediate 14. Then MeOH attacks at the C1-position and the more stable methoxy lactol 15 having all equatrial substituents was formed as the result of equilibrium with 15′.³⁴⁾ The Density Functional Theory (DFT) calculation (Method/Basis set b3lyp/6-31 + g(d,p)) displayed that 15 is more stable than 15′ for 3.79 kcal/mol. Next, an ortho-quinone methide 17 is generated by elimination of the C4-methoxy group via the protonated compound 16. In this time, the ortho-quinone methide 17 is considered to take the most stable conformation when pentyl group occupies the equatorial position. Then, MeOH attacks at the C4-position by avoiding axial hydrogen at the C3-position to afford C4-epimerized lactol 18. The DFT calculation (Method/Basis set b3lyp/6-31 + g(d,p)) displayed that 18 is a little more stable than 15 for 0.87 kcal/mol. Finally, the cleavage of MOM group affords (–)-eurotiumide G (13). In other words, 13 was obtained as the result of thermodynamically equilibrium at the C1-position and kinetically nucleophilic attack at the C4-position, respectively.

Chart 4. Plausible Mechanism of Generation of (–)-Eurotiumide G (13)

This isomerization at the C4-position seemed to be completed during the deprotection of MOM group. Based on our previous research of total syntheses of 1 and 2, we found that the isomerization at the C4-position is quite slow under acidic condition with 4-methoxyisochroman-1-one derivative. To avoid this isomerization, we decided to change the order of reduction of lactone moiety and deprotection of MOM group (Chart 5). Treatment of 5 with 6 M HCl aq in MeOH removed MOM group and DIBAL reduction occurred from the opposite side of the 4-OMe group to afford lactol 20. Finally, displacement of a hydroxy group at the C1-position to a methoxy group was conducted under mild condition, and the treatment of 20 with 0.4 M HCl aq in MeOH at 0°C afforded desired (+)-eurotiumide F (3) in moderate yield without isomerization at the C4-position. All spectral data of synthetic eurotiumide F are matched with those of the reported data of eurotiumide F. The absolute configurations of the C1-, C3-, C4-positions of (+)-eurotiumide F were assigned as (1S,3R,4S) by the NOESY experiment.

Chart 5. Completion of Total Synthesis of (+)-Eurotiumide F (3)

In our previous total syntheses of eurotiumide A (1) and B (2), we found that 1 and 2 emitted quite strong fluorescence and the possibility for using natural fluorescent probe. We also evaluated the fluorescent properties of 3 and 4, respectively. Although they emitted weak violet fluorescence, they were not enough to use as natural fluorescent probes.

In conclusion, we achieved the asymmetric total syntheses of (+)-eurotiumide F and (+)-eurotiumide G involving thermal rearrangement for constructing dihydropyran ring in 7 and 4 steps from the known chiral cis 4-methoxyisochroman-1-one compound 8, respectively. Their absolute stereochemistries were determined by using optically active intermediate 6 and relative stereochemistry of the C1-, C3-, and C4-positions of 3 could be assigned by NOESY experiment. Moreover, the X-ray crystallographic analysis of (+)-4 revealed the correct stereochemistry at the C1-position and revised the reported structure of 4. We also found that 3 and 4 emit weak violet fluorescence under UV irradiation. Further investigation of antimicrobial activities is underway in our laboratory.

Experimental

Preparation of Dimethyl Propargyl Ether Compound 7

To a solution of cis 4-methoxyisochroman-1-one compound 8 (150.0 mg, 0.462 mmol) in CH₃CN (12.0 mL) were added CuCl₂ solution (0.85 mL, 0.00231 mmol, 0.0027 M in CH₃CN), DBU (0.242 mL, 1.62 mmol), and 3-chloro-3-methyl-1-butyne (0.18 mL, 1.62 mmol) at 0°C, successively. After stirring for 12 h at room temperature, the reaction was quenched by adding saturated aqueous NH₄Cl. The mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (EtOAc–hexane = 5 : 95 to 10 : 90) to give propargyl derivative 7 (166.1 mg, 92%) as a yellow oil. ¹H-NMR (CDCl₃, 400 MHz) δ: 7.54 (1H, d, J = 9.4 Hz), 7.32 (1H, d, J = 9.4 Hz), 5.22 (2H, s), 4.61 (1H, d, J = 1.6 Hz), 4.22 (1H, ddd, J = 8.5, 5.8, 1.6 Hz), 3.51 (3H, s), 3.31 (3H, s), 2.53 (1H, s), 2.05 (1H, m), 1.80 (1H, m), 1.70 (3H, s), 1.67 (3H, s), 1.46–1.33 (5H, m), 0.90 (3H, t, J = 6.8 Hz). ¹³C-NMR (CDCl₃, 125 Hz) δ: 161.7, 151.2, 149.5, 128.4, 124.7, 120.6, 118.8, 95.2, 86.5, 80.5, 74.7, 74.1, 68.6, 56.7, 56.4, 31.6, 30.6, 29.6, 28.7, 24.9, 22.5, 14.0. IR (KBr) cm⁻¹: 3256, 2373, 1734, 1480, 1258, 1221, 1125. High resolution (HR)-MS (electrospray ionization (ESI)) m/z [M + Na]: 413.1942 (Calcd for C₂₂H₃₀O₆Na: 413.1940). [α]²³_D +115.0 (c 0.95, CH₂Cl₂, 97% enantiomeric excess (ee)).

Preparation of Dihydropyran Derivative 6 with Thermal Condition

A solution of alkyne propargyl derivative 7 (60.0 mg, 0.155 mmol) in o-dichlorobenzene (15 mL) was stirring at 160°C for 3 h. After the mixture was cooled to room temperature, the reaction mixture was directly purified by silica gel column chromatography (EtOAc–hexane = 10 : 90) to give dihydropyran derivative 6 (60.0 mg, quant) as a yellow oil. ¹H-NMR (CDCl₃, 400 MHz) δ: 7.03 (1H, s), 6.29 (1H, d, J = 9.8 Hz), 5.76 (1H, d, J = 9.8 Hz), 5.17 (1H, d, J = 7.2 Hz), 5.16 (1H, d, J = 7.2 Hz), 4.57 (1H, d, J = 1.2 Hz), 4.23 (1H, ddd, J = 8.3, 5.8, 1.6 Hz), 3.50 (3H, s), 3.29 (3H, s), 2.03 (1H, m), 1.79 (1H, m), 1.55 (3H, s), 1.45–1.31 (6H, m), 1.42 (3H, s), 0.90 (3H, t, J = 6.8 Hz). ¹³C-NMR (CDCl₃, 100 Hz) δ: 161.4, 149.8, 147.1, 133.2, 127.5, 124.0, 121.6, 117.1, 114.0, 95.5, 80.5, 77.3, 68.5, 56.5, 56.3, 31.6, 30.7, 28.0, 27.8, 25.0, 22.6, 14.0. IR (KBr) cm⁻¹: 1732, 1600, 1460, 1434, 1293, 1153. HR-MS (ESI) m/z [M + H]: 391.2110 (Calcd for C₂₂H₃₁O₆: 391.2121). [α]²³_D +189.9 (c 0.99, CH₂Cl₂, 97% ee).

Preparation of Dihydropyran Derivative 6 with Microwave Condition

A solution of alkyne propargyl derivative 7 (4.8 mg, 0.0123 mmol) in o-dichlorobenzene (1.2 mL) was stirring at 200°C with microwave (300 W) for 10 min. After the mixture was cooled to room temperature, the reaction mixture was directly purified by silica gel column chromatography (EtOAc–hexane = 10 : 90) to give dihydropyran derivative 6 (4.8 mg, quant) as a yellow oil. These microwave-irradiation reaction was carried out with CEM Discover™SP. All reactions were carried out in sealed heavy-walled Pyrex tubes with constant temperature, pressure, and irradiation power (300 W).

Preparation of Lactol 11

To a solution of dihydropyran derivative 6 (15.4 mg, 0.0394 mmol) in CH₂Cl₂ (0.31 mL) was added DIBAL solution (77 µL, 0.0789 mmol, 1.02 M in Hexane) at −78°C. After stirring for 15 min, saturated aqueous Rochelle salt (0.60 mL) was added to the reaction mixture. After the mixture was stirring for additional 20 min at 0°C, the mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by preparative thin layer chromatography (EtOAc–hexane = 30 : 70) to give lactol 11 (10.9 mg, 70%) as a yellow oil. ¹H-NMR (CDCl₃, 500 MHz) δ: 6.81 (1H, s), 6.28 (1H, d, J = 9.8 Hz), 6.13 (1H, s), 5.65 (1H, d, J = 9.8 Hz), 5.17 (1H, d, J = 6.5 Hz), 5.15 (1H, d, J = 6.5 Hz), 4.29 (1H, d, J = 2.0 Hz), 4.15 (1H, ddd, J = 7.8, 6.3, 1.5 Hz), 3.49 (3H, s), 3.45 (3H, s), 2.83 (1H, s), 1.85 (1H, m), 1.77 (1H, m), 1.47 (1H, m), 1.45 (3H, s), 1.42–1.35 (5H, m), 1.38 (3H, s), 0.91 (3H, t, J = 7.0 Hz). ¹³C-NMR (CDCl₃, 125 Hz) δ: 149.2, 144.2, 131.8, 124.5, 123.5, 122.1, 121.7, 112.1, 95.2, 88.3, 76.5, 71.3, 68.8, 58.1, 56.0, 31.9, 30.7, 28.0, 27.6, 25.5, 22.6, 14.1. IR (KBr) cm⁻¹: 3412, 1731, 1474, 1436, 1272, 1110. HR-MS (ESI) m/z [M + H]: 393.2271 (Calcd for C₂₂H₃₃O₆: 393.2277). [α]²³_D +15.0 (c 0.34, CH₂Cl₂, 97% ee).

Preparation of (+)-Eurotiumide G (4)

To a solution of lactol 11 (6.7 mg, 0.0171 mmol) in MeOH (1.3 mL) was added 6 M aqueous HCl (0.43 mL) at 0°C. After stirring for 1.5 h at room temperature, the reaction was quenched by adding saturated aqueous NaHCO₃. The mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by preparative thin layer chromatography (EtOAc–hexane = 30 : 70) to give (+)-eurotiumide G (4) (3.4 mg, 55%) as a white solid. mp. 111°C. ¹H-NMR (CDCl₃, 500 MHz) δ: 6.59 (1H, s), 6.28 (1H, d, J = 9.6 Hz), 6.02 (1H, s), 5.67 (1H, d, J = 9.6 Hz), 5.56 (1H, s), 4.44 (1H, d, J = 3.6 Hz), 4.21 (1H, td, J = 7.0, 3.6 Hz), 3.54 (3H, s), 3.18 (3H, s), 1.80 (2H, q, J = 7.2 Hz), 1.61 (1H, m), 1.51 (1H, m), 1.44–1.34 (10H, m), 0.93 (3H, t, J = 7.0 Hz). ¹³C-NMR (CDCl₃, 125 Hz) δ: 149.0, 143.3, 132.2, 124.0, 122.4, 122.1, 117.7, 113.1, 95.2, 75.9, 70.1, 68.7, 55.7, 53.7, 31.8, 30.2, 27.8, 27.3, 25.8, 22.6, 14.1. IR (KBr) cm⁻¹: 3339, 1639, 1441, 1261, 1110, 1055. HR-MS (ESI) m/z [M + H]: 363.2174 (Calcd for C₂₁H₃₁O₅: 363.2171). [α]²³_D +20.3 (c 0.81, CH₂Cl₂, 97% ee).

Single crystals of 4, obtained by slow evaporation of a pentane solution of 4, were selected and fitted onto a glass fiber and measured at −173°C with a Bruker Apex II ultra diffractometer using MoKα radiation. Data correction and reduction were performed with the crystallographic package Apex3. The structure was solved and refined with the Bruker SHELXTL software package, using the space group P2₁2₁2₁ with Z = 4 for the formula unit C₂₁H₃₀O₅. The final anisotropic full-matrix least-squares refinement on F² with 241 variables converged at R1 = 3.49%, for the observed data and wR2 = 7.82% for all data. The goodness-of-fit was 0.992. The ORTEP plot was obtained by the program PLATON (A. L. Spek, 2009).

Preparation of trans 4-Methoxyisochroman-1-one Compound 5

To a solution of dihydropyran 6 (138.6 mg, 0.355 mmol) in EtOH (12 mL) was added aqueous KOH (4.5 mL, 2.25 mmol, 0.5 M in H₂O) at room temperature. After stirring for 20 min at 90°C, the reaction mixture was cooled to 0°C and neutralized to pH 5.0 with 1 M aqueous HCl. EtOH was removed under reduced pressure and the aqueous layer was extracted with EtOAc (x3). The combined organic layers were dried over Na₂SO₄, filtered and concentrated under reduced pressure to afford carboxylic acid as a crude product. To a solution of the crude residue in THF (3.6 mL) was added PPh₃ (279.4 mg, 1.07 mmol) and diethylazodicarboxylate (0.45 mL, 0.994 mmol, 2.2 M in toluene) at 0°C. After stirring for 20 min at room temperature, the reaction mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (EtOAc–hexane = 10 : 90 to 20 : 80) to give trans 4-methoxyisochroman-1-one compound 5 (111.5 mg, 80%) as a yellow oil. ¹H-NMR (CDCl₃, 400 MHz) δ: 7.03 (1H, s), 6.29 (1H, d, J = 9.8 Hz), 5.77 (1H, 9.8 Hz), 5.18 (1H, d, J = 6.8 Hz), 5.15 (1H, d, J = 6.8 Hz), 4.71 (1H, ddd, J = 7.8, 6.0, 0.8 Hz), 4.60 (1H, dd, J = 1.2, 0.8 Hz), 3.49 (3H, s), 3.34 (3H, s), 1.50 (1H, m), 1.52 (3H, s), 1.47 (3H, s), 1.39–1.14 (7H, m), 0.85 (3H, t, J = 6.8 Hz). ¹³C-NMR (CDCl₃, 100 Hz) δ: 159.8, 149.7, 148.0, 133.3, 125.5, 124.2, 121.7, 117.7, 113.9, 95.5, 80.2, 70.1, 56.2, 56.0, 32.2, 31.4, 27.85, 27.77, 25.3, 22.4, 13.9. IR (KBr) cm⁻¹: 1730, 1638, 1459, 1434, 1299, 1139. HR-MS (ESI) m/z [M + H]: 391.2116 (Calcd for C₂₂H₃₁O₆: 391.2121). [α]²³_D +175.9 (c 1.03, CH₂Cl₂, 97% ee).

Preparation of Lactol 12

To a solution of trans 4-methoxyisochroman-1-one compound 5 (13.6 mg, 0.0348 mmol) in CH₂Cl₂ (0.27 mL) was added DIBAL solution (68 µL, 0.0697 mmol, 1.02 M in Hexane) at −78°C. After the mixture was stirring for 15 min, saturated aqueous Rochelle salt (0.6 mL) was added to the reaction mixture. After stirring for 20 min at 0°C, the mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by preparative thin layer chromatography (EtOAc–hexane = 30 : 70) to give lactol 12 (10.8 mg, 79%) as a yellow oil. ¹H-NMR (CDCl₃, 400 MHz) δ: 6.78 (1H, s), 6.28 (1H, d, J = 9.8 Hz), 6.12 (1H, d, J = 11.2 Hz), 5.68 (1H, d, J = 9.8 Hz), 5.16 (1H, d, J = 6.6 Hz), 5.13 (1H, d, J = 6.6 Hz), 4.72 (1H, d, J = 11.2 Hz), 4.61 (1H, d, J = 1.6 Hz), 4.44 (1H, td, J = 6.0, 1.6 Hz), 3.48 (3H, s), 3.32 (3H, s), 1.45 (3H, s), 1.40 (3H, s), 1.34–1.24 (8H, m), 0.85 (3H, t, J = 6.8 Hz). ¹³C-NMR (CDCl₃, 100 Hz) δ: 149.1, 143.2, 132.5, 127.3, 122.4, 122.1, 120.6, 111.9, 95.2, 86.9, 76.4, 75.0, 72.7, 56.0, 55.9, 34.2, 31.7, 27.9, 27.7, 25.0, 22.5, 14.0. IR (KBr) cm⁻¹: 3431, 1638, 1479, 1436, 1265, 1105. HR-MS (ESI) m/z [M + H]: 393.2280 (Calcd for C₂₂H₃₃O₆: 393.2277). [α]²³_D +73.2 (c 0.55, CH₂Cl₂, 97% ee).

Preparation of (–)-Eurotiumide G (13)

To a solution of lactol 12 (5.5 mg, 0.0139 mmol) in MeOH (1.0 mL) was added 6 M aqueous HCl (0.35 mL) at 0°C. After stirring for 4 h at room temperature, the reaction was quenched by adding saturated aqueous NaHCO₃. The mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by preparative thin layer chromatography (EtOAc–hexane = 30 : 70) to give (–)-eurotiumide G (13) (2.0 mg, 40%) as a yellow oil. All spectral data except for optical rotation were identified with those of the data of (+)-eurotiumide G (4) prepared by us. [α]²³_D –22.8 (c 0.20, CH₂Cl₂).

Preparation of Free Phenol 19

To a solution of trans 4-methoxyisochroman-1-one compound 5 (204.5 mg, 0.524 mmol) in MeOH (39 mL) was added 6 M aqueous HCl (13.1 mL) at 0°C. After stirring for 4 h at room temperature, the reaction was quenched by adding saturated aqueous NaHCO₃. The mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (EtOAc–hexane = 15 : 85 to 25 : 75) to give free phenol 19 (111.8 mg, 62%) as a yellow amorphous. ¹H-NMR (CDCl₃, 400 MHz) δ: 6.84 (1H, br-s), 6.81 (1H, d, J = 1.6 Hz), 6.25 (1H, d, J = 10.0 Hz), 5.75 (1H, d, J = 10.0 Hz), 4.66 (1H, d, J = 4.0 Hz), 4.63 (1H, m), 3.43 (3H, s), 1.58–1.20 (8H, m), 1.48 (3H, s), 1.46 (3H, s), 0.86 (3H, t, J = 7.0 Hz). ¹³C-NMR (CDCl₃, 100 Hz) δ: 160.7, 149.1, 147.1, 133.5, 124.3, 122.1, 121.4, 119.2, 112.8, 79.8, 77.2, 73.1, 56.3, 32.0, 31.5, 27.7 (overlapped), 25.0, 22.5, 13.9. IR (KBr) cm⁻¹: 3168, 1684, 1598, 1455, 1441, 1305, 1155. HR-MS (ESI) m/z [M + H]: 347.1853 (Calcd for C₂₀H₂₇O₅: 347.1858). [α]²³_D +107.8 (c 1.01, CH₂Cl₂, 97% ee).

Preparation of Lactol 20

To a solution of free phenolic alcohol 19 (46.0 mg, 0.133 mmol) in CH₂Cl₂ (2.7 mL) was added DIBAL solution (2.0 mL, 1.99 mmol, 1.02 M in Hexane) at −78°C over 2 h with a syringe pump. After stirring for 30 min at the same temperature, saturated aqueous Rochelle salt (6.0 mL) was added to the reaction mixture. After stirring for 20 min at 0°C, the mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure to give crude product. Because lactol 20 was unstable on silica gel, the crude product of 20 was used for the next reaction immediately without further purification. ¹H-NMR (CDCl₃, 400 MHz) δ: 7.38 (1H, s), 6.52 (1H, s), 6.26 (1H, d, J = 9.8 Hz), 6.01 (1H, d, J = 3.8 Hz), 5.65 (1H, d, J = 9.8 Hz), 4.71 (1H, d, J = 9.0 Hz), 4.46 (1H, td, J = 9.0, 2.8 Hz), 3.28 (3H, s), 2.94 (1H, d, J = 3.8 Hz), 1.87 (1H, m), 1.65–1.33 (7H, m), 1.46 (3H, s), 1.36 (3H, s), 0.92 (3H, t, J = 7.2 Hz). ¹³C-NMR (CDCl₃, 100 Hz) δ: 149.4, 142.5, 132.1, 125.1, 122.3, 121.9, 117.7, 113.3, 88.1, 76.3, 75.7, 65.5, 51.7, 31.8 (overlapped), 28.0, 27.4, 24.7, 22.6, 14.1. IR (KBr) cm⁻¹: 3431, 1638, 1459, 1361, 1168, 1130. HR-MS (ESI) m/z [M + H]: 349.2018 (Calcd for C₂₀H₂₉O₅: 349.2015). [α]²³_D +83.6 (c 0.64, CH₂Cl₂, 97% ee).

Preparation of (+)-Eurotiumide F (3)

To a solution of crude product of 20 in MeOH (7.5 mL) was added 0.4 M aqueous HCl (2.5 mL) at 0°C. After stirring for 15 min at the same temperature, the reaction was quenched by adding saturated aqueous NaHCO₃. The mixture was extracted with EtOAc (x3) and the combined organic layers were washed with brine, dried over Na₂SO₄, filtered and concentrated under reduced pressure. The residue was purified by preparative thin layer chromatography (EtOAc–hexane = 30 : 70) to give (+)-eurotiumide F (3) (22.2 mg, 46% in 2 steps) as a yellow oil. ¹H-NMR (CDCl₃, 400 MHz) δ: 7.42 (1H, s), 6.51 (1H, s), 6.25 (1H, d, J = 10.0 Hz), 5.64 (1H, d, J = 10.0 Hz), 5.44 (1H, s), 4.69 (1H, d, J = 9.8 Hz), 4.31 (1H, td, J = 9.8, 2.4 Hz), 3.56 (3H, s), 3.24 (3H, s), 1.89 (1H, m), 1.65 (1H, m), 1.55 (1H, m), 1.44 (3H, s), 1.41–1.31 (5H, m), 1.34 (3H, s), 0.93 (3H, t, J = 7.2 Hz). ¹³C-NMR (CDCl₃, 125 Hz) δ: 149.3, 142.9, 132.0, 124.1, 122.4, 122.0, 117.7, 113.2, 95.1, 75.91, 75.89, 64.7, 55.9, 51.2, 31.9, 31.8, 27.8, 27.4, 25.1, 22.7, 14.1. IR (KBr) cm⁻¹: 3395, 1727, 1560, 1460, 1359, 1282, 1132. HR-MS (ESI) m/z [M + H]: 363.2179 (Calcd for C₂₁H₃₁O₅: 363.2171). [α]²³_D +95.3 (c = 0.96, CH₂Cl₂, 97% ee).

Acknowledgments

This work was supported by JSPS KAKENHI Grant Nos. 17K08365 (AN), 16K01927 (KN), and 16H01156 (KN), and the Kurita Water and Environment Foundation and Japan Ecology Foundation. And we acknowledge Tokushima University for their financial support of the Research Clusters program of Tokushima University (No. 1802001) and Research program of development of intelligent Tokushima artificial exosome (iTEX) from Tokushima University.

Conflict of Interest

The authors declare no conflict of interest.

Supplementary Materials

The online version of this article contains supplementary materials.

References and Notes

• 1) Schueffler A., Anke T., Nat. Prod. Rep., 31, 1425–1448 (2014).
• 2) Jin L., Quan C., Hou X., Fan S., Mar. Drugs, 14, 76–100 (2016).
• 3) Blunt J. W., Copp B. R., Keyzers R. A., Munro M. H. G., Prinsep M. R., Nat. Prod. Rep., 34, 235–294 (2017).
• 4) Hirata Y., Uemura D., Pure Appl. Chem., 58, 701–710 (1986).
• 5) Towle M. J., Salvato K. A., Budrow J., et al., Cancer Res., 61, 1013–1021 (2001).
• 6) Wright A. E., Forleo D. A., Gunawardana G. P., Gunasekera S. P., Koehn F. E., McConnell O. J., J. Org. Chem., 55, 4508–4512 (1990).
• 7) Rinehart K. L., Holt T. G., Fregeau N. L., Stroh J. G., Keifer P. A., Sun F., Li H. Li., Martin D. G., J. Org. Chem., 55, 4512–4515 (1990).
• 8) Cuevas C., Francesch A., Nat. Prod. Rep., 26, 322–337 (2009).
• 9) Chen M., Shao C.-L., Wang K.-L., Xu Y., She Z. G., Wang C.-Y., Tetrahedron, 70, 9132–9138 (2014).
• 10) Guimarães K. G., de Souza Filho J. D., dos Marae-Guia T. R., Braga F. C., Phytochemistry, 69, 439–444 (2008).
• 11) Paraschos S., Magiatis P., Kalpoutzakis E., Harvala C., Skaltsounis A. L., J. Nat. Prod., 64, 1585–1587 (2001).
• 12) Kongsaeree P., Prabpai S., Sriubolmas N., Vongvein C., Wiyakrutta S., J. Nat. Prod., 66, 709–711 (2003).
• 13) Nakayama A., Sato H., Karanjit S., Hayashi N., Oda M., Namba K., Eur. J. Org. Chem., 2018, 4013–4017 (2018).
• 14) Yun B.-S., Cho Y., Lee I.-K., Cho S.-M., Lee T. H., Yoo I.-D., J. Antibiot., 55, 208–210 (2002).
• 15) Mokrini R., Mesaoud M. B., Daoudi M., Hellio C., Maréchal J. P., Hattab M. E., O.-Magné A., Piovetti L., Culioli G., J. Nat. Prod., 71, 1806–1811 (2008).
• 16) Castanheiro R. A. P., Silva A. M. S., Campos N. A. N., Nascimento M. S. J., Pinto M. M. M., Pharmaceuticals, 2, 33–43 (2009).
• 17) Sasaki H., Kashiwada Y., Shibata H., Takaishi Y., Phytochemistry, 82, 136–142 (2012).
• 18) Sasaki H., Shibata H., Imabayashi K., Takaishi Y., Kashiwada Y., J. Agric. Food Chem., 62, 6345–6353 (2014).
• 19) Zhou M., Zhou K., Zhao Y.-L., Xiang N.-J., Zhang T.-D., Wang Y.-D., Dong W., Ji B.-K., Li L.-M., Lou J., Li G.-P., Hu Q.-F., Phytochem. Lett., 11, 245–248 (2015).
• 20) Raksat A., Maneerat W., Andersen R. J., Pyne S. G., Laphookhieo S., J. Nat. Prod., 81, 1835–1840 (2018).
• 21) Goddard M.-L., Tabacchi R., Tetrahedron Lett., 47, 909–911 (2006).
• 22) Little A., Porco J. A. Jr., Org. Lett., 14, 2862–2865 (2012).
• 23) Tu Y., Wang Z.-X., Shi Y., J. Am. Chem. Soc., 118, 9806–9807 (1996).
• 24) Ueda T., Konishi H., Manabe K., Angew. Chem. Int. Ed., 52, 8611–8615 (2013).
• 25) Ueda T., Konishi H., Manabe K., Org. Lett., 15, 5370–5373 (2013).
• 26) Konishi H., Manabe K., Synlett, 25, 1971–1986 (2014).
• 27) Haraldsson G. G., Baldwin J. E., Tetrahedron, 53, 215–224 (1997).
• 28) Godfrey J. D. Jr., Mueller R. H., Sedergran T. C., Soundararajan N., Colandrea V. J., Tetrahedron Lett., 35, 6405–6408 (1994).
• 29) Iwai I., Ide J., Chem. Pharm. Bull., 10, 926–933 (1962).
• 30) Iwai I., Ide J., Chem. Pharm. Bull., 11, 1042–1049 (1963).
• 31) Zsindely J., Schmid H., Helv. Chim. Acta, 51, 1510–1514 (1968).
• 32) The DFT calculation (b3lyp/6-31 + G(d,p)) supports that 11 would be more stable than that of the C1-enantiomer of 11 for 4.12 kcal/mol.
• 33) Crystallographic data of 4 have been deposited at the Cambridge Crystallographic Data Centre as supplementary publication numbers CCDC 1918279. Copies of the data can be obtained, free of charge, on application to CCDC, 12 Union Road, Cambridge CB2 1EZ U.K. (Fax: [phone] or e-mail: [e-mail]).
• 34) During this isomerization, only the methoxy lactol 15 could be detected and determined the stereochemistry at the C1-position by observing the NOESY correlation between 1-OMe and H-3.