Taurospongins B and C, new acetylenic fatty acid derivatives possessing a taurine amide residue from a marine sponge of the family Spongiidae

Abstract

Two new acetylenic fatty acid derivatives possessing a taurine amide residue, taurospongins B (1) and C (2), have been isolated from an Okinawan marine sponge of the family Spongiidae. The gross structures of 1 and 2 were elucidated on the basis of their spectral data, especially 2D NMR and FABMS/MS data. The absolute configurations for 1 and 2 were established by chemical means. Taurospongin C (2) showed inhibitory activity against Cryptococcus neoformans.

---

Introduction

Marine sponges of the family Spongiidae have been demonstrated to be a rich source of unique bioactive meroterpenoids^1,2 and acetylenic fatty acid derivatives.^3,4 During our search for bioactive metabolites from marine organisms, we investigated the extract of a sponge family Spongiidae (SS-1202), which resulted in the isolation of two new acetylenic fatty acid derivatives possessing a taurine amide residue, taurospongins B (1) and C (2). Here we describe the isolation and structure elucidation of 1 and 2 (Fig. 1).

Fig. 1 Taurospongins B (1), C (2), and A (3).

Results and discussion

The sponge family Spongiidae collected at Okinawa, was extracted with MeOH. After evaporation, the MeOH extract was partitioned stepwise between organic solvents (EtOAc and n-BuOH) and H₂O. A part of n-BuOH soluble materials was fractionated by gel filtration (Sephadex LH-20, MeOH). A fraction eluted in a relatively early stage was purified by C₁₈ column chromatography (MeOH–H₂O) and SiO₂ column chromatography (CH₃Cl–MeOH) to afford taurospongins B (1, 1.7 mg, 0.00048%, wet weight) and C (2, 4.2 mg, 0.0012%).

Taurospongin B (1) was obtained as an optically active colorless amorphous solid. The molecular formula of 1 was revealed to be C₃₈H₆₉NO₇S by HRESIMS data [m/z 682.47036 (M − H)⁻, Δ −1.84 mmu]. IR absorptions indicated the existence of hydroxy (3421 cm⁻¹), ester carbonyl (1732 cm⁻¹), and amide carbonyl (1646 cm⁻¹) functionalities. The inspection of the HMQC and HMBC spectra with ¹H and ¹³C NMR data disclosed that 1 consists of ester and amide carbonyls, a triple bond, a double bond, three methyls, twenty seven methylenes, an oxymethine, and an oxygenated quaternary carbon (Table 1). Analysis of the ¹H–¹H COSY and TOCSY spectra of 1 revealed connectivities of C-4 to C-10, C-2′ to C-3′, C-6′ to C-10′, C-23′ to C-25′, and C-1′′ to C-2′′. The geometry of a double bond between C-8′ and C-9′ was assigned as Z by the vicinal coupling constant (³J_{H-8′/H-9′} = 10.3 Hz). HMBC correlations of H₂-1′′/C-1 and H₂-2/C-1 clarified that N-bearing carbon C-1′′ (δ_C 37.3) and a carbonyl-bearing carbon C-2 (δ_C 48.6) were connected via an amide bond containing a carbonyl carbon C-1 (δ_C 174.6). Linkings of C-2, a methylene carbon C-4, and a methyl carbon C-11 through an oxygenated quaternary carbon C-3 (δ_C 73.4) were inferred from HMBC correlations of H₃-11/C-2, H₂-4/C-3, and H₃-11/C-4. An HMBC correlation of H₂-2′/C-1′ revealed that a carbonyl-bearing carbon C-2′ (δ_C 36.2) was attached to a carbonyl carbon C-1′ (δ_C 174.6). Linkage of C-3′ and C-6′ by a triple bond between acetylenic carbons C-4′ (δ_C 80.2) and C-5′ (δ_C 82.2) was uncovered by HMBC correlations of H₂-3′/C-4′ and H₂-6′/C-5′. The chemical shift of a proton H-7 (δ_C 4.98) of an oxygenated carbon C-7 (δ_C 76.2) implied that C-7 was esterified to C-1′. These data and the molecular formula of 1 indicated an attachment of a sulfo group to C-2′′ and the connection of C-10′ and C-23′ by a methylene chain (Fig. 2).

Table 1 ¹H and ¹³C NMR data of taurospongins B (1) and C (2) in CD₃OD

Position	1		2
	δH^a multi (J in Hz)	δC^b	δH^a multi (J in Hz)	δC^b
a Recorded at 600 MHz.b Recorded at 150 MHz.c 2H.d 3H.e 24H.f 12C.g J-values were not determined because of overlapping with other signals.h These signals might be exchange.i These signals might be exchange.j These signals might be exchange.k These signals might be exchange.l These signals might be exchange.
1	—	174.6	—	174.5^k
2	2.40 d (14.0)	48.6	2.36^c m	48.4
	2.34 d (14.0)
3	—	73.4	—	74.2
4	1.59 m^g	43.8	1.59 m^g	43.9
	1.53 m^g		1.52 m^g
5	1.44^c m^g	21.6	1.47^c m^g	21.8
6	1.61^c m^g	36.6	1.64^c m^g	36.7
7	4.98 m	76.2	3.67 m^g	70.1
8	1.59^c m^g	38.3	1.85 m^g	45.3
			1.64 m^g
9	1.43^c m^g	20.5^h	5.15 m	71.3
10	0.96^d t (7.4)	15.1^i	1.30^d d (6.3)	20.7
11	1.25^d s	27.6	1.27^d s	27.7
1′	—	174.6	—	174.3^k
2′	2.51^c m	36.2	2.50^c m	36.2
3′	2.48^c m	16.5	2.48^c m	16.4
4′	—	80.2	—	80.1
5′	—	82.2	—	82.2
6′	2.18^c m	20.7^h	2.18^c m	20.7
7′	2.23^c m	28.8^j	2.23^c m	28.7^l
8′	5.44 dt (10.3, 5.6)	130.0	5.44 dt (10.7, 5.4)	130.0
9′	5.45 dt (10.3, 5.6)	132.8	5.46 dt (10.7, 5.4)	132.8
10′	2.09^c m	29.0^j	2.09^c q (6.4)	29.0^l
11′–22′	1.52–1.15^e m^g	31.7–31.1^f	1.52–1.15^e m	31.7–31.1^f
23′	1.33^c m^g	33.9	1.38^c m	33.9
24′	1.38^c m^g	24.5	1.33^c m	24.5
25′	0.94 t (7.1)	15.2^i	0.94 t (7.0)	15.3
1′′	3.66 t (6.5)	37.3	3.67 m^g	37.3
2′′	3.02 t (6.5)	52.2	3.03 t (6.6)	52.2

---

Fig. 2 Selected 2D NMR correlations for taurospongins B (1).

The structure of 1 elucidated from the NMR data was also confirmed by a charge-remote fragmentation pattern induced by a sulfo group observed in the FABMS/MS spectrum of 1 (Fig. 3). Thus, the gross structure of taurospongin B (1) was elucidated as shown.

Fig. 3 Fragmentation patterns observed in FABMS/MS spectrum of taurospongin B (1) [precursor ion, m/z 682 (M − H)⁻]. The m/z values were indicated in italics.

The relative stereochemistry of taurospongin B (1) was established by comparison of the NMR data of taurineamide part (4) of 1 with its two possible diastereomers (6a and 6b). The taurineamide part (4) was obtained by methanolysis of an ester linkage of 1 with an acetylenic fatty acid part (5) of 1 (Scheme 1).⁴

Scheme 1 Methanolysis of taurospongin B (1).

The taurine amides 6a and 6b were synthesized from D-mevalonolactone as follows (Schemes 2 and 3). A hydroxy group of D-mevalonolactone was protected by a TES group and then treated with DIBAL to give lactol (8), which was coupled with phosphorane by Wittig reaction. After protection of hydroxy group by a BOM group, the resulting chloro-olefin (10) was subjected to a coupling reaction with n-butylaldehyde to afford a mixture of acetylenic alcohols (11a and 11b), which was converted into an acetylenic ketone (12) by oxidation using AZADO.⁵

Scheme 2 Synthesis of 6a and 6b (part 1).

Scheme 3 Synthesis of 6a and 6b (part 2).

The acetylenic ketone (12) was derived to chiral acetylenic alcohol (11a) by a Noyori asymmetric transfer hydrogenation using Ru[(S,S)-Tsdpen](p-cymene).⁶ The absolute stereochemistry at C-7 of 11a was verified by modified Mosher's method⁷ to be R-configuration. After the hydroxy group of the 11a was protected by TBS group, deprotection of BOM group and reduction of the triple bond were accomplished by using Pearlman's catalyst.⁸ The resulting primary alcohol (14a) was oxidized to carboxylic acid (15a) by combination of Parikh–Doering oxidation⁹ and Lindgren–Kraus oxidation.¹⁰ Finally, the carboxylic acid was coupled with taurine by amidation using DMT-MM¹¹ to obtain 3S,7R-isomer (6a). The 3S,7S-isomer (6b) was prepared from the acetylenic ketone (12) in a similar manner using Ru[(R,R)-Tsdpen](p-cymene).⁶ Since the ¹H NMR spectra of taurineamide part (4) of taurospongin B (1) was coincident with that of 6a, relative relasionship for two hydroxy groups of 4 were uncoverd to be syn relationship.

The absolute configuration at C-7 of taurospongin B (1) was established by modified Mosher's method.⁷ Treatment of taurineamide part (4) with (R)-(−)- and (S)-(+)-MTPACl gave (S)- and (R)-MTPA esters of 4, respectively. Δδ values obtained from ¹H NMR data of MTPA esters of 4 (Fig. 4) indicated that 4 was an enantiomer of 6a and the absolute configuration at C-7 of 4 was R. Therefore, the absolute configurations at C-3 and C-7 of taurospongin B (1) were assigned as both R.

Fig. 4 Δδ values [Δδ (in ppm) = δ_S − δ_R] obtained for the (S)- and (R)-MTPA esters of taurineamide part (4) of taurospongin B (1). Δδ values were indicated in italic.

Taurospongin C (2) was obtained as an optically active colorless amorphous solid. The molecular formula of 2 was elucidated as C₃₈H₆₉NO₈S by HRESIMS data [m/z 698.46739 (M − H)⁻, Δ +0.28 mmu]. IR absorptions suggested that 2 possesses hydroxy (3369 cm⁻¹), ester carbonyl (1730 cm⁻¹), and amide carbonyl (1636 cm⁻¹) functionalitiess. The ¹H and ¹³C NMR data of 2 were almost superimposable to those of taurospongin A (3),^4,12,13 except for disappearance of signals derived from an acetyl group. In addition, the difference of the molecular formula between 2 and 3 implied that 2 was a 9-O-desacetyl form of 3. To verify the prediction, 2 was derived to 9-O-acetyl form by treatment of 2 with acetic anhydride and pyridine. Since the spectral data of 9-O-acetyl form of 2 was identical with those of 3, 2 was assigned as 9-O-desacetyl taurospongin A (3).

Antimicrobial assay¹⁴ of taurospongins B (1), C (2), and A (3) revealed that 2 and 3 exhibited mild and good antifungal activity, respectively, against Cryptococcus neoformans (MIC, 32 and 1 μg mL⁻¹), while 1 did not show such activity (MIC > 32.0 μg mL⁻¹). Taurospongins B (1), C (2), and A (3) were not active against other fungi Aspergillis niger, Trichophyton mentagrophytes, and Candida albicans (MIC > 32 μg mL⁻¹), and bacteria Escherichia coli, Staphylococcus aureus, Bacillus subtilis, and Micrococcus luteus (IC₅₀ > 32 μg mL⁻¹). Taurospongins B (1), C (2), and A (3) did not show cytotoxicity (IC₅₀ > 10 μg mL⁻¹) against murine lymphoma L1210 and human epidermoid carcinoma KB cells in vitro.

Conclusions

Taurospongins B (1) and C (2), two new acetylenic fatty acid derivatives possessing a taurine amide residue, have been isolated from an Okinawan marine sponge of the family Spongiidae. The absolute structures of 1 and 2 were established by combination of spectroscopic analysis and synthetic chemistry. Taurospongins C (2) and A (3) showed mild and good antifungal activity, respectively, against Cryptococcus neoformans (MIC, 32 and 1 μg mL⁻¹).

Experimental section

General experimental procedures

Optical rotations were recorded on a JASCO P-1030 polarimeter. IR spectra were recorded on JASCO FT/IR-230 spectrometer. ¹H and ¹³C NMR spectra were recorded on JEOL ECA400 and JEOL ECA500 spectrometers using 5 mm cell and a Bruker AMX-600 spectrometer using 2.5 mm micro cell. The 7.20 and 77.0 ppm resonances of residual CHCl₃ and CDCl₃, and 3.35 and 49.8 ppm resonances of residual CD₂HOD and CD₃OD were used as internal references for ¹H and ¹³C NMR spectra, respectively. EIMS spectra were recorded on a JEOL JMS-700TZ mass spectrometer. ESIMS spectra were recorded on JEOL JMS-700TZ and Thermo Scientific Exactive mass spectrometers.

Sponge description

The sponge (SS-1202, family Spongiidae) was collected at Unten Port, Okinawa, and kept frozen until used. The sponge was medium brown mound with a spiky, conulose surface, hispid with a fine adherent membrane, and alcohol brown stains. Dermis was unarmored. The sponge has a reticulate fibre skeleton with some pithing of fibres centrally. The reticulation is irregular and all fibres are uncored. Primary fibres are ∼50 μm wide. Secondary fibres are ∼40 μm wide with finer fibres between as a tertiary skeleton, 10 μm wide. The voucher specimen was deposited at Graduate School of Pharmaceutical Sciences, Hokkaido University.

Extraction and isolation

The sponge of the family Spongiidae (SS-1202, 1.25 kg, wet weight) collected at Okinawa, was extracted with MeOH (2L × 2) to afford the extract (73.3 g), which was partitioned stepwise between organic solvents [EtOAc (500 mL × 3) and n-BuOH (500 mL × 3)] and H₂O (500 mL) to give EtOAc-soluble materials (11.8 g) and n-BuOH-soluble materials (3.6 g). A part (1.0 g) of n-BuOH soluble materials was fractionated by gel filtration (Sephadex LH-20, GE Healthcare; eluent, MeOH). A fraction was purified by C₁₈ column chromatography (Cosmosil 140 C₁₈ PREP, Nakarai Tesque Inc.; eluent, MeOH–H₂O, 70 : 30 to 100 : 0) and SiO₂ column chromatography (Wakosil C-300, Wako Pure Chemical Industries, Ltd.; eluent, CH₃Cl–MeOH, 95 : 5 to 0 : 100) to afford taurospongins B (1, 1.7 mg, 0.00048%, wet weight) and C (2, 4.2 mg, 0.0012%).

Taurospongin B (1). Colorless amorphous solid; [α]²⁵_D + 3.4 (c 1.0, MeOH); IR (film) ν_max 3421, 1732, 1646, 1456, 1172, 1046 cm⁻¹; ¹H and ¹³C NMR data see Table 1; HRESIMS (neg.) m/z 682.47036 [calcd for C₃₈H₆₈NO₇S (M − H)⁻, Δ −1.84 mmu].

Taurospongin C (2). Colorless amorphous solid; [α]²⁵_D + 0.5 (c 1.0, MeOH); IR (film) ν_max 3369, 1730, 1636, 1465, 1171, 1044 cm⁻¹; ¹H and ¹³C NMR data see Table 1; HRESIMS (neg.) m/z 698.46739 [calcd for C₃₈H₆₈NO₈S (M − H)⁻, Δ +0.28 mmu].

Solvolysis of taurospongin B (1). Taurospongin B (1, 1.0 mg, 1.46 μmol) was dissolved in MeOH–1 M HCl (10 : 1, 500 μl). After stirring for 12 h at 80 °C, the mixture was concentrated by Ar blowing. The residue was extracted with CHCl₃, which was concentrated by Ar blowing to afford acetylenic fatty acid part (5, 0.5 mg, 1.28 μmol, 88%). The remaining CHCl₃ insoluble material was taurineamide part (4, 0.4 mg, 1.23 μmol, 84%).

Taurineamide part (4). ¹H NMR (600 MHz, CD₃OD) δ 3.67 (t, J = 6.6 Hz, 2H), 3.58 (m, 1H), 3.03 (t, J = 6.6 Hz, 2H), 2.43 (d, J = 14.0 Hz, 1H), 2.36 (d, J = 14.0 Hz, 1H), 1.66–1.32 (m, 10H), 1.27 (s, 3H), 0.97 (t, J = 7.1 Hz, 9H); HRESIMS (neg.) m/z 324.14876 [calcd for C₁₃H₂₆NO₆S (M − H)⁻, Δ +0.13 mmu].

(S)-MTPA ester of 4. (R)-(−)-MTPACl (1.23 μl) was added to a solution of 4 (0.4 mg, 1.23 μmol) in pyridine (50 μl). After stirring for 1 h at rt, (R)-(−)-MTPACl (1.23 μl) was added to the mixture additionally. After stirring for 1 h at rt, the reaction mixture was concentrated in vacuo. The residue was dissolved with MeOH and passed through an ion-exchange column (Amberlite IR-120 H⁺ form, eluent, MeOH) and concentrated in vacuo. Then, the residue was purified by C18 HPLC (Mightysil RP-18 GP, Kanto Chemical Co., Inc., 4.6 × 250 mm; eluent, CH₃CN–H₂O, 70–100%; flow rate, 1.0 mL min⁻¹; UV detection at 230 nm) to afford (S)-MTPA ester of 4 (0.4 mg, 0.74 mmol, 60%); ¹H NMR (600 MHz, CD₃OD) δ 7.57 (m, 1H), 7.48 (m, 1H), 5.16 (m, 1H), 3.65 (t, J = 6.6 Hz, 2H), 3.59 (s, 3H), 3.02 (t, J = 6.6 Hz, 2H), 2.30 (d, J = 14.3 Hz, 1H), 2.24 (d, J = 14.3 Hz, 1H), 1.59 (m, 2H), 1.24 (m, 2H), 1.80–1.10 (m, 6H), 1.14 (s, 3H), 0.99 (t, J = 7.3 Hz, 3H); HRESIMS (neg.) m/z 540.18929 [calcd for C₂₃H₃₃NO₈F₃S (M − H)⁻, Δ +0.84 mmu].

(R)-MTPA ester of 4. The (R)-MTPA ester of 4 (0.4 mg, 0.74 mmol) was obtained from 4 (0.4 mg, 1.23 μmol) in 60% by using (S)-(+)-MTPACl through the same procedure as described for preparation of the (S)-MTPA ester of 4.; ¹H NMR (600 MHz, CD₃OD) δ 7.57 (m, 1H), 7.47 (m, 4H), 5.17 (m, 1H), 3.66 (t, J = 6.6 Hz, 2H), 3.60 (s, 3H), 3.02 (t, J = 6.6 Hz, 2H), 2.38 (d, J = 14.3 Hz, 1H), 2.32 (d, J = 14.3 Hz, 1H), 1.69 (m, 2H), 1.42 (m, 2H), 1.80–1.10 (m, 6H), 1.22 (s, 3H), 0.90 (t, J = 7.3 Hz, 3H); HRESIMS (neg.) m/z 540.18851 [calcd for C₂₃H₃₃NO₈F₃S (M − H)⁻, Δ +0.06 mmu].

(R)-4-Methyl-4-((triethylsilyl)oxy)tetrahydro-2H-pyran-2-one (7). Imidazole (1.12 g, 16.45 mmol), DMAP (100.4 mg, 0.82 mmol), and TESCl (2.06 mL, 12.27 mmol) were added to a solution of D-mevalonolactone (1.07 g, 8.22 mmol) in DMF (20 mL) at 0 °C. After stirring for 24 h at rt, H₂O was added to the mixture, which was extracted with EtOAc. The organic layer was washed with saturated aqueous CuSO₄, H₂O, and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 85 : 15) to afford 7 (1.81 g, 7.41 mmol, 90%); yellow oil; [α]²¹_D − 28 (c 1.6, CHCl₃); IR (neat) ν_max 2957, 2877, 1738, 1458, 1223, 1005, 725 cm⁻¹; ¹H NMR (500 MHz, CDCl₃) δ 4.58 (m, 1H), 4.34 (m, 1H), 2.67 (brd, J = 17.2 Hz, 1H), 2.43 (d, J = 17.2 Hz, 1H), 1.84 (m, 2H), 1.39 (s, 3H), 0.94 (t, J = 8.0 Hz, 9H), 0.60 (q, J = 8.0 Hz, 6H); ¹³C NMR (100 MHz, CDCl₃) δ 170.2, 70.4, 66.1, 45.6, 36.9, 29.0, 6.8, 6.3; HRESIMS (pos.) m/z 267.13881 [calcd for C₁₂H₂₄O₃NaSi (M + Na)⁺, Δ +0.12 mmu].

(4R)-4-Methyl-4-((triethylsilyl)oxy)tetrahydro-2H-pyran-2-ol (8). DIBAL (1.04 M in hexane, 8.76 mL, 9.11 mmol) was added to a solution of 7 (1.85 g, 7.57 mmol) in CH₂Cl₂ (20 mL) at −78 °C. After stirring for 1 h at −78 °C, MeOH and saturated aqueous potassium sodium tartrate were added to the mixture, which was allowed to warm to rt with vigorous stirring for 1 h and extracted with EtOAc. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 85 : 15) to afford 8 (1.81 g, 7.35 mmol, 97%), which was directly used in the next step; yellow oil; HRESIMS (pos.) m/z 269.15447 [calcd for C₁₂H₂₆O₃NaSi (M + Na)⁺, Δ +0.13 mmu].

(S)-6-Chloro-3-methyl-3-((triethylsilyl)oxy)hex-5-en-1-ol (9). n-BuLi (1.60 M in hexane, 18.6 mL, 29.76 mmol) was added dropwise to a solution of (chloromethyl)triphenylphosphonium chloride (10.33 g, 29.76 mmol) in THF (40 mL) at −78 °C. After stirring for 30 min at −78 °C, a solution of 8 (1.83 g, 7.43 mmol) in THF (3 mL) was added dropwise to the mixture at −78 °C. After stirring for 39 h at −78 °C, saturated aqueous NH₄Cl was added to the mixture, which was extracted with EtOAc. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford 9 (1.29 g, 4.63 mmol, 62%), which was directly used in the next step; pale yellow oil; HRESIMS (pos.) m/z 301.13601 [calcd for C₁₃H₂₇O₂ClNaSi (M + Na) ⁺, Δ −0.10 mmu].

(S)-7-(3-Chloroallyl)-9,9-diethyl-7-methyl-1-phenyl-2,4,8-trioxa-9-silaundecane (10). N,N-Diisopropylethylamine (3.15 mL, 18.56 mmol) and benzylchloromethyl ether (1.91 mL, 13.92 mmol) were added to a solution of 9 (1.29 g, 4.63 mmol) in CH₂Cl₂ (32 mL) at 0 °C. After stirring for 12 h at 0 °C, saturated aqueous NH₄Cl was added to the mixture, which was extracted with EtOAc. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford 10 (1.56 g, 3.91 mmol, 84%), which was directly used in the next step; pale yellow oil; HRESIMS (pos.) m/z 421.19366 [calcd for C₂₁H₃₅O₃ClNaSi (M + Na)⁺, Δ +0.04 mmu].

(8S)-10-((Benzyloxy)methoxy)-8-methyl-8-((triethylsilyl)oxy)dec-5-yn-4-ol (11a and 11b). n-BuLi (1.60 M in hexane, 6.64 mL, 10.63 mmol) was added dropwise to a solution of 10 (1.41 g, 3.53 mmol) in THF (15 mL) at −78 °C. After stirring for 2 h at −78 °C, n-BuLi (1.60 M in hexane, 2.21 mL, 3.54 mmol) was added dropwise at −78 °C additionally. Then a solution of butyraldehyde (0.80 mL, 8.86 mmol) in THF (15 mL) was added dropwise to the mixture. After stirring for 1 h at −78 °C, saturated aqueous NH₄Cl was added to the mixture, which was extracted with EtOAc. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford mixture of 11a and 11b (1.21 g, 2.78 mmol, 79%), which was directly used in the next step; pale yellow oil; HRESIMS (pos.) m/z 457.27449 [calcd for C₂₅H₄₂O₄NaSi (M + Na)⁺, Δ +0.03 mmu].

(S)-10-((Benzyloxy)methoxy)-8-methyl-8-((triethylsilyl)oxy)dec-5-yn-4-one (12). Iodobenzene diacetate (1.36 g, 4.23 mmol) and AZADO (42.9 mg, 0.28 mmol) were added to a solution of 11a and 11b (1.24 g, 2.82 mmol) in CH₂Cl₂ (4 mL). After stirring for 4 h at rt, CH₂Cl₂ (35 mL) and a mixture of saturated aqueous NaHCO₃ and saturated aqueous Na₂SO₄ (1 : 1, 30 mL) were added to the mixture, which was extracted with CH₂Cl₂. The organic layer was washed with brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford 12 (1.16 g, 2.68 mmol, 95%); pale yellow oil; [α]²²_D + 5.0 (c 1.25, CHCl₃); IR (neat) ν_max 3019, 2876, 2211, 1673, 1455, 1377, 1240, 1171, 1114, 1045, 742, 698 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 7.37–7.26 (m, 5H), 4.75 (s, 2H), 4.60 (s, 2H), 3.72 (m, 2H), 2.61 (d, J = 17.1 Hz, 1H), 2.54 (d, J = 17.1 Hz, 1H), 2.50 (t, J = 7.3 Hz, 1H), 1.96 (ddd, J = 13.9, 7.3, 6.8 Hz, 1H), 1.89 (ddd, J = 13.9, 7.5, 6.4 Hz, 1H), 1.69 (qt, J = 7.3, 7.3 Hz, 2H), 1.37 (s, 3H), 0.95 (qt, J = 7.8 Hz, 9H), 0.93 (t, J = 7.3 Hz, 3H), 0.60 (q, J = 7.8 Hz, 6H); ¹³C NMR (150 MHz, CDCl₃) δ 188.1, 138.0, 128.4, 127.8, 127.7, 94.7, 91.1, 82.6, 73.9, 69.4, 64.1, 47.4, 41.6, 33.8, 28.1, 17.6, 13.5, 7.0, 6.7; HRESIMS (pos.) m/z 455.25877 [calcd for C₂₅H₄₀O₄NaSi (M + Na)⁺, Δ −0.04 mmu].

(4S,8S)-10-((Benzyloxy)methoxy)-8-methyl-8-((triethylsilyl)oxy)dec-5-yn-4-ol (11a). A solution of 12 (580 mg, 1.34 mmol) in i-PrOH (11.6 mL) was added to a solution of [(1S,2S)-N-(p-toluenesulfonyl)-1,2-duphenylethanediamine]-(p-cymene)ruthenium(II) (68.4 mg, 0.11 mmol) in i-PrOH (8.29 mL). After stirring for 3 h at rt, the reaction mixture was concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 80 : 20) to afford 11a (563 mg, 1.30 mmol, 97%, 97 : 3 dr); pale brown oil; [α]²³_D + 4.8 (c 1.23, CHCl₃); IR (neat) ν_max 3441, 2956, 2875, 1455, 1376, 1150, 1111, 1041, 733 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 7.38–7.26 (m, 5H), 4.76 (s, 2H), 4.61 (s, 2H), 4.33 (brt, J = 6.5 Hz, 1H), 3.74 (t, J = 7.4 Hz, 2H), 2.44 (dd, J = 16.4 and 1.5 Hz, 1H), 2.36 (dd, J = 16.4 and 1.8 Hz, 1H), 2.03–1.75 (m, 3H), 1.63 (m, 2H), 1.46 (m, 2H), 1.33 (s, 3H), 0.94 (t, J = 7.9 Hz, 9H), 0.93 (t, J = 7.3 Hz, 3H), 0.59 (q, J = 7.9 Hz, 6H); ¹³C NMR (100 MHz, CDCl₃) δ 137.8, 128.4, 127.9, 127.7, 94.5, 83.4, 82.3, 74.0, 69.3, 64.4, 62.4, 41.2, 40.1, 33.4, 28.0, 18.4, 13.7, 7.0, 6.7; HRESIMS (pos.) m/z 457.27512 [calcd for C₂₅H₄₂O₄NaSi (M + Na)⁺, Δ +0.66 mmu].

(S)-MTPA ester of 11a. DMAP (0.2 mg), Et₃N (0.65 μl), and (R)-(−)-MTPACl (0.33 μl) were added to a solution of 11a (0.5 mg, 1.15 mmol) in CH₂Cl₂ (30 μl). After stirring for 3 h at rt, N,N-dimethyl 1,3-propanediamine (0.64 μl) was added to the mixture. After stirring for 15 min at rt, the reaction mixture was concentrated in vacuo. The residue was purified by SiO₂ (hexane–EtOAc, 1 : 0 to 2 : 1) to afford (S)-MTPA ester of 11a (0.7 mg, 1.08 mmol, 94%); ¹H NMR (600 MHz, CDCl₃) δ 7.71–7.29 (m, 10H), 5.56 (t, J = 6.7 Hz, 1H), 4.73 (s, 2H), 4.59 (s, 2H), 3.71 (t, J = 7.3 Hz, 2H), 3.58 (s, 3H), 2.46 (d, J = 16.4 Hz, 1H), 2.38 (d, J = 16.4 Hz, 1H), 1.93 (m, 1H), 1.86 (m, 1H), 1.73 (m, 2H), 1.35 (m, 2H), 1.31 (s, 3H), 0.93 (t, J = 7.9 Hz, 9H), 0.87 (t, J = 7.3 Hz, 3H), 0.57 (q, J = 7.9 Hz, 6H); HRESIMS (pos.) m/z 673.31519 [calcd for C₃₅H₄₉O₆F₃NaSi (M + Na)⁺, Δ +0.92 mmu].

(R)-MTPA ester of 11a. The (R)-MTPA ester (0.7 mg, 1.08 mmol) of 11a was obtained from 11a (0.5 mg, 1.15 mmol) in 94% by using (S)-(−)-MTPACl through the same procedure as described for preparation of the (S)-MTPA ester of 11a; ¹H NMR (600 MHz, CDCl₃) δ 7.70–7.29 (m, 10H), 5.53 (t, J = 6.7 Hz, 1H), 4.73 (s, 2H), 4.59 (s, 2H), 3.70 (t, J = 7.1 Hz, 2H), 3.54 (s, 3H), 2.42 (d, J = 16.4 Hz, 1H), 2.35 (d, J = 16.4 Hz, 1H), 1.91 (m, 1H), 1.84 (m, 1H), 1.80 (m, 2H), 1.46 (m, 2H), 1.29 (s, 3H), 0.93 (t, J = 7.4 Hz, 3H), 0.92 (t, J = 7.8 Hz, 9H), 0.56 (q, J = 7.9 Hz, 6H); HRESIMS (pos.) m/z 673.31528 [calcd for C₃₅H₄₉O₆F₃NaSi (M + Na)⁺, Δ +1.01 mmu].

(4R,8S)-10-((Benzyloxy)methoxy)-8-methyl-8-((triethylsilyl)oxy)dec-5-yn-4-ol (11b). 11b (555 mg, 1.28 mmol) was obtained from 12 (580 mg, 1.34 mmol) in 96% (97 : 3 dr) by using [(1R,2R)-N-(p-toluenesulfonyl)-1,2-duphenylethanediamine]-(p-cymene)ruthenium(II) through the same procedure as described for preparation of 11a; pale brown oil; [α]²³_D + 8.4 (c 1.77, CHCl₃); IR (neat) ν_max 3447, 2956, 2875, 1455, 1376, 1150, 1112, 1041, 741 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 7.38–7.26 (m, 5H), 4.76 (s, 2H), 4.61 (s, 2H), 4.33 (brs, 1H), 3.74 (t, J = 7.3 Hz, 2H), 2.44 (dd, J = 16.4 and 1.8 Hz, 1H), 2.36 (dd, J = 16.4 and 1.8 Hz, 1H), 1.98 (m, 1H), 1.87 (m, 2H), 1.63 (m, 2H), 1.46 (m, 2H), 1.33 (s, 3H), 0.94 (t, J = 7.8 Hz, 9H), 0.93 (t, J = 7.4 Hz, 3H), 0.59 (q, J = 7.8 Hz, 6H); ¹³C NMR (100 MHz, CDCl₃) δ 137.8, 128.4, 127.9, 127.7, 94.5, 83.4, 82.3, 74.0, 69.2, 64.4, 62.4, 41.2, 40.1, 33.4, 28.0, 18.5, 13.7, 7.1 6.6; HRESIMS (pos.) m/z 457.27408 [calcd for C₂₅H₄₂O₄NaSi (M + Na)⁺, Δ −0.38 mmu].

(S)-MTPA ester of 11b. The (S)-MTPA ester (0.7 mg, 1.08 mmol) of 11b was obtained from 11b (0.5 mg, 1.15 mmol) in 94% through the same procedure as described for preparation of the (S)-MTPA ester of 11a; ¹H NMR (600 MHz, CDCl₃) δ 7.71–7.29 (m, 10H), 5.53 (t, J = 6.4 Hz, 1H), 4.72 (s, 2H), 4.58 (s, 2H), 3.69 (t, J = 7.3 Hz, 2H), 3.54 (s, 3H), 2.43 (d, J = 16.4 Hz, 1H), 2.35 (d, J = 16.4 Hz, 1H), 1.89 (m, 1H), 1.83 (m, 1H), 1.81 (m, 2H), 1.46 (m, 2H), 1.29 (s, 3H), 0.93 (t, J = 7.8 Hz, 9H), 0.88 (t, J = 7.5 Hz, 3H), 0.57 (q, J = 7.8 Hz, 6H); HRESIMS (pos.) m/z 673.31405 [calcd for C₃₅H₄₉O₆F₃NaSi (M + Na)⁺, Δ −0.22 mmu].

(R)-MTPA ester of 11b. The (R)-MTPA ester (0.7 mg, 1.08 mmol) of 11b was obtained from 11b (0.5 mg, 1.15 mmol) in 94% by using (S)-(−)-MTPACl through the same procedure as described for preparation of the (S)-MTPA ester of 11a; ¹H NMR (600 MHz, CDCl₃) δ 7.71–7.29 (m, 10H), 5.56 (t, J = 6.6 Hz, 1H), 4.73 (s, 2H), 4.58 (s, 2H), 3.71 (t, J = 7.3 Hz, 2H), 3.58 (s, 3H), 2.46 (d, J = 16.4 Hz, 1H), 2.39 (d, J = 16.4 Hz, 1H), 1.92 (m, 1H), 1.85 (m, 1H), 1.73 (m, 2H), 1.36 (m, 2H), 1.32 (s, 3H), 0.93 (t, J = 7.9 Hz, 9H), 0.87 (t, J = 7.3 Hz, 3H), 0.58 (q, J = 7.9 Hz, 6H); HRESIMS (pos.) m/z 673.31402 [calcd for C₃₅H₄₉O₆F₃NaSi (M + Na)⁺, Δ −0.25 mmu].

(7S,11S)-7,13,13,14,14-Pentamethyl-1-phenyl-11-propyl-7-((triethylsilyl)oxy)-2,4,12-trioxa-13-silapentadec-9-yne (13a). Imidazole (175.4 mg, 2.58 mmol) and TBSCl (290.3 mg, 1.94 mmol) were added to a solution of 11a (560 mg, 1.29 mmol) in CH₂Cl₂ (11 mL). After stirring for 3 h at rt, saturated aqueous NH₄Cl was added to the mixture, which was extracted with CH₂Cl₂. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford 13a (675 mg, 1.23 mmol, 95%); colorless oil; [α]²⁴_D − 14.6 (c 1.17, CHCl₃); IR (neat) ν_max 2956, 2876, 1456, 1251, 1111, 1041, 836, 776, 731 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 7.38–7.26 (m, 5H), 4.75 (s, 2H), 4.60 (s, 2H), 4.33 (brt, J = 6.3, 1H), 3.73 (t, J = 7.4 Hz, 2H), 2.43 (dd, J = 16.4 and 1.5 Hz, 1H), 2.34 (dd, J = 16.4 and 1.9 Hz, 1H), 1.98 (m, 1H), 1.86 (m, 1H), 1.61 (m, 2H), 1.42 (m, 2H), 1.33 (s, 3H), 0.94 (t, J = 7.9 Hz, 9H), 0.91 (t, J = 7.3 Hz, 3H), 0.90 (s, 9H), 0.58 (q, J = 7.9 Hz, 6H), 0.11 (s, 3H), 0.09 (s, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 137.9, 128.4, 127.9, 127.6, 94.6, 84.0, 81.1, 74.1, 69.2, 64.4, 62.9, 41.1, 41.1, 33.5, 28.0, 25.8, 18.6, 18.2, 13.8, 7.1, 6.7, −4.5, −5.1; HRESIMS (pos.) m/z 571.36206 [calcd for C₃₁H₅₆O₄NaSi₂ (M + Na)⁺, Δ +1.13 mmu].

(7S,11R)-7,13,13,14,14-Pentamethyl-1-phenyl-11-propyl-7-((triethylsilyl)oxy)-2,4,12-trioxa-13-silapentadec-9-yne (13b). 13b (674 mg, 1.23 mmol) was obtained from 11b (554 mg, 1.27 mmol) in 97% through the same procedure as described for preparation of 13a; colorless oil; [α]²³_D + 16.5 (c 1.52, CHCl₃); IR (neat) ν_max 2958, 2876, 1456, 1251, 1111, 1041, 836, 776, 731 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 7.38–7.26 (m, 5H), 4.75 (s, 2H), 4.60 (s, 2H), 4.33 (brt, J = 6.5 Hz, 1H), 3.73 (t, J = 7.4 Hz, 2H), 2.43 (dd, J = 16.3 and 1.8 Hz, 1H), 2.35 (dd, J = 16.3 and 1.8 Hz, 1H), 1.98 (m, 1H), 1.86 (m, 1H), 1.62 (m, 2H), 1.42 (m, 2H), 1.33 (s, 3H), 0.94 (t, J = 7.8 Hz, 9H), 0.91 (t, J = 7.1 Hz, 3H), 0.90 (s, 9H), 0.59 (q, J = 7.8 Hz, 6H), 0.12 (s, 3H), 0.10 (s, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 137.9, 128.4, 127.9, 127.6, 94.6, 84.0, 81.1, 74.1, 69.2, 64.3, 62.9, 41.1, 41.1, 33.5, 28.0, 25.8, 18.6, 18.2, 13.8, 7.1, 6.7, −4.5, −5.1; HRESIMS (pos.) m/z 571.36052 [calcd for C₃₁H₅₆O₄NaSi₂ (M + Na)⁺, Δ −0.41 mmu].

(3S,7S)-7-((tert-Butyldimethylsilyl)oxy)-3-methyl-3-((triethylsilyl)oxy)decan-1-ol (14a). 20% Pd(OH)₂/C (13.5 mg) was added to a solution of 13a (27 mg, 0.049 mmol) in EtOAc (2.5 mL) and the mixture was stirred under H₂ at rt. Additional 20% Pd(OH)₂/C (13.5 mg) and EtOAc (2.5 mL) were added to the mixture every 1 h. After stirring under H₂ for 5 h at rt, the mixture was filtered through a cotton plug, and the filtrate was concentrated in vacuo. The residue was purified by a SiO₂ column (CHCl₃) to afford 14a (10.9 mg, 0.025 mmol, 51%); colorless oil; [α]²⁴_D + 4.2 (c 1.87, CHCl₃); IR (neat) ν_max 3363, 2956, 2932, 2875, 1455, 1253, 1040, 773 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 3.78 (brq, 5.2 Hz, 2H), 3.63 (brquin, J = 5.6 Hz, 1H), 3.18 (brt, J = 4.5 Hz, 1H), 1.75 (dt, J = 14.5, 5.6 Hz, 1H), 1.70–1.56 (m, 2H), 1.53–1.15 (m, 12H), 0.95 (t, J = 7.9 Hz, 9H), 0.89 (t, J = 7.2 Hz, 3H), 0.87 (s, 9H), 0.60 (q, J = 7.9 Hz, 6H), 0.03 (s, 3H), 0.03 (s, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 77.8, 72.0, 60.0, 43.2, 42.0, 39.4, 37.6, 27.7, 25.9, 20.6, 18.5, 18.1, 14.3, 7.0, 6.7, –4.5; HRESIMS (pos.) m/z 455.33454 [calcd for C₂₃H₅₂O₃NaSi₂ (M + Na)⁺, Δ −0.18 mmu].

(3S,7R)-7-((tert-Butyldimethylsilyl)oxy)-3-methyl-3-((triethylsilyl)oxy)decan-1-ol (14b). 14b (8.4 mg, 0.019 mmol) was obtained from 13b (15 mg, 0.027 mmol) in 70% through the same procedure as described for preparation of 14a; colorless oil; [α]²⁴_D + 2.2 (c 1.39, CHCl₃); IR (neat) ν_max 3368, 2955, 2931, 2876, 1461, 1251, 1041, 773 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 3.79 (2H, brt, J = 5.0 Hz), 3.63 (brquin, J = 5.6 Hz), 3.18 (brs, 1H), 1.75 (dt, J = 14.4, 5.5 Hz, 1H),1.70–1.17 (m, 12H), 0.96 (9H, t, J = 7.9 Hz, TES-Me), 0.89 (t, J = 7.1 Hz, 3H), 0.88 (s, 9H), 0.61 (q, J = 7.9 Hz, 6H), 0.04 (s, 3H), 0.03 (s, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 77.9, 71.9, 60.0, 43.1, 41.9, 39.4, 37.7, 27.7, 25.9, 20.5, 18.6, 18.1, 14.3, 7.0, 6.8, −4.5; HRESIMS (pos.) m/z 455.33408 [calcd for C₂₃H₅₂O₃NaSi₂ (M + Na)⁺, Δ −0.64 mmu].

(3S,7S)-3,7-Dihydroxy-3-methyldecanoic acid (15a). DMSO (70.0 μl) and Et₃N (24.0 μl) were added to a solution of 14a (5.0 mg, 0.012 mmol) in CH₂Cl₂ (212.0 μl). After stirring for 5 min at 0 °C, SO₃–pyr. 19.2 mg was added to the mixture. After stirring for 1 h at rt, additional SO₃–pyr. (19.2 mg) was added to the reaction mixture. After stirring for 1 h at rt, the mixture was extracted with diethyl ether. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo to afford crude aldehyde. Then t-BuOH (0.73 mL), NaH₂PO₄ (8.2 mg in 182.0 μl of H₂O), and 2-methyl-2-butene (2 M in THF, 50.4 μl) were added to the crude aldehyde. After stirring for 5 min at rt, NaClO₂ (7.3 mg) was added to the mixture. After stirring for 15 min at 0 °C, saturated aqueous NaHSO₄ was added to the mixture, which was extracted with EtOAc. The organic layer was washed with H₂O and brine, dried with MgSO₄, and concentrated in vacuo. The residue was purified by a SiO₂ column (n-hexane–EtOAc, 100 : 0 to 90 : 10) to afford 15a (2.0 mg, 0.0092 mmol, 77%); colorless oil; [α]¹⁸_D − 5.9 (c 2.02, CHCl₃); IR (neat) ν_max 3410, 2923, 2853, 1717, 1562, 1456, 1240, 1129, 774 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 5.05 (brs, 1H), 3.64 (m, 1H), 2.56 (d, J = 15.6 Hz, 1H), 2.47 (d, J = 15.6 Hz, 1H), 1.61–1.29 (m, 10H), 1.28 (s, 3H), 0.92 (t, J = 6.8 Hz, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 175.8, 71.5, 71.4, 44.6, 41.6, 39.6, 37.2, 26.7, 19.9, 18.8, 14.1; HRESIMS (pos.) m/z 241.14118 [calcd for C₁₁H₂₂O₄Na (M + Na)⁺, Δ +0.15 mmu].

(3S,7R)-3,7-Dihydroxy-3-methyldecanoic acid (15b). 15b (1.4 mg, 0.0064 mmol) was obtained from 14b (4.0 mg, 0.0092 mmol) in 70% through the same procedure as described for preparation of 15a; colorless oil; [α]²⁰_D − 4.87 (c 1.72, CHCl₃); IR (neat) ν_max 3362, 2920, 2850, 1717, 1575, 1456, 1219, 1123, 772 cm⁻¹; ¹H NMR (400 MHz, CDCl₃) δ 5.20 (brs, 1H), 3.64 (m, 1H), 2.56 (d, J = 15.6 Hz, 1H), 2.48 (d, J = 15.6 Hz, 1H), 1.62–1.29 (m, 10H), 1.28 (s, 3H), 0.92 (t, J = 6.8 Hz, 3H); ¹³C NMR (100 MHz, CDCl₃) δ 175.9, 71.6, 71.5, 44.8, 41.6, 39.6, 37.2, 26.6, 19.9, 18.8, 14.1; HRESIMS (pos.) m/z 241.14116 [calcd for C₁₁H₂₂O₄Na (M + Na)⁺, Δ +0.13 mmu].

2-((3S,7S)-3,7-Dihydroxy-3-methyldecanamido)ethanesulfonic acid (6a). DMT-MM (95 mg) and Et₃N (400 μl) were added to a solution of 15a (25.0 mg, 0.11 mmol) in DMF (5 mL). After stirring for 10 min at rt, taurine (190 mg) was added to the mixture. After stirring for 10 h at rt, the mixture was concentrated in vacuo. The residue was dissolved with MeOH and passed through an ion-exchange column (Amberlite IR-120 H⁺ form, eluent, MeOH) and concentrated in vacuo. The residue was purified by a SiO₂ column (CHCl₃–MeOH, 100 : 0 to 50 : 50) to afford 6a (19.0 mg, 0.058 mmol, 53%); colorless oil; [α]²⁰_D + 3.5 (c 1.14, MeOH); IR (neat) ν_max 3366, 2955, 2871, 2280, 1638, 1555, 1284, 1144, 1059, 938 cm⁻¹; ¹H NMR (600 MHz, CD₃OD) δ 3.67 (brt, J = 6.7 Hz, 2H), 3.58 (m, 1H), 3.03 (t, J = 6.6 Hz, 2H), 2.43 (d, J = 14.0 Hz, 1H), 2.36 (d, J = 14.0 Hz, 1H), 1.66–1.32 (m, 10H), 1.27 (s, 3H), 0.97 (t, J = 7.1 Hz, 9H); ¹³C NMR (100 MHz, CD₃OD) δ 175.2, 73.9, 72.8, 52.3, 48.3, 44.2, 41.6, 39.6, 37.2, 27.8, 22.1, 20.8, 15.3; HRESIMS (neg.) m/z 324.14903 [calcd for C₁₃H₂₆NO₆S (M − H)⁻, Δ +0.40 mmu].

2-((3S,7R)-3,7-Dihydroxy-3-methyldecanamido)ethanesulfonic acid (6b). 6b (20.5 mg, 0.063 mmol) was obtained from 15b (24 mg, 0.11 mmol) in 57% through the same procedure as described for preparation of 6a; colorless oil; [α]²⁰_D − 0.4 (c 1.28, MeOH); IR (neat) ν_max 3439, 2957, 2857, 2279, 1644, 1565, 1282, 1147, 1069, 921 cm⁻¹; ¹H NMR (600 MHz, CD₃OD) δ 3.68 (brt, J = 6.6 Hz, 2H), 3.58 (m, 1H), 3.03 (t, J = 6.6 Hz, 2H), 2.44 (d, J = 14.0 Hz, 1H), 2.38 (d, J = 14.0 Hz, 1H), 1.66–1.32 (m, 10H), 1.29 (s, 3H), 0.97 (t, J = 7.1 Hz, 9H); ¹³C NMR (100 MHz, CD₃OD) δ 175.4, 74.0, 72.8, 52.3, 48.2, 44.2, 41.6, 39.6, 37.2, 27.8, 22.1, 20.8, 15.3; HRESIMS (neg.) m/z 324.14892 [calcd for C₁₃H₂₆NO₆S (M − H)⁻, Δ +0.29 mmu].

Antimicrobial assay

Antimicrobial assay of 1–3 against Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Aspergillus niger, Candida albicans, Cryptococcus neoformans, and Trichophyton mentagrophytes was carried out as previously described.¹⁴ Amphotericin B, micafungin, hygromycin B, and kanamycin showed antifungal activity against Cryptococcus neoformans (MIC, <0.05, <0.1, 4.0, and 8.0 μg mL⁻¹, respectively).

Cytotoxic assays

Human epidermoid carcinoma (KB) and murine leukemia L1210 cells were cultured in an incubator at 37 °C for 48 h in 100 μl of medium containing various concentrations of test compounds dissolved in 1% DMSO. The IC₅₀ values were obtained by plotting the logarithm of the concentration of the test compound versus the growth rate of the treated cells. Paclitaxel was used as positive control (IC₅₀, <0.005 and <0.1 μg mL⁻¹, respectively).

Acknowledgements

We thank Mr Z. Nagahama and Mr K. Uehara, Okinawa, for their help with sponge collection, and Ms S. Oka, Instrumental Analysis Division, Equipment Management Center, Creative Reseach Institution, Hokkaido University, for measurements of mass spectrometry. We specially thank ADEKA corporation for supplying D-mevalonolactone, and Prof. Y. Iwabuchi, Graduate School of Pharmaceutical Sciences, Tohoku University, for supplying AZADO. This work was supported by The Naito Foundation, Cooperative Research Program of Medical Mycology Research Center, Chiba University, and Grant-in-Aid from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.

Notes and references

1. J. Kobayashi, T. Madono and H. Shigemori, Tetrahedron, 1995, 51, 10867 , and reference cited therein.
2. D. Mori, Y. Kimura, S. Kitamura, Y. Sakagami, Y. Yoshioka, T. Shintani, T. Okamoto and M. Ojika, J. Org. Chem., 2007, 72, 7190.
3. A. A. Salim, J. Rae, F. Fontaine, M. M. Conte, Z. Khalil, S. Martin, R. G. Parton and R. J. Capon, Org. Biomol. Chem., 2010, 8, 3188.
4. H. Ishiyama, M. Ishibashi, A. Ogawa, S. Yoshida and J. Kobayashi, J. Org. Chem., 1997, 62, 3831.
5. M. Shibuya, M. Tomizawa, I. Suzuki and Y. Iwabuchi, J. Am. Chem. Soc., 2006, 128, 8412.
6. K. Matsumura, S. Hashiguchi, T. Ikariya and R. Noyori, J. Am. Chem. Soc., 1997, 119, 8738.
7. I. Ohtani, T. Kusumi, Y. Kashman and H. Kakisawa, J. Am. Chem. Soc., 1991, 113, 4092.
8. W. M. Pearlman, Tetrahedron Lett., 1967, 8, 1663.
9. J. R. Parikh and W. von E. Doering, J. Am. Chem. Soc., 1967, 89, 5505.
10. B. S. Bal, W. E. Childers Jr and H. W. Pinnick, Tetrahedron, 1981, 37, 2091.
11. M. Kunishima, C. Kawachi, F. Iwasaki, K. Terao and S. Tani, Tetrahedron Lett., 1999, 40, 5327.
12. Wu. Boshen, A. Mallinger and J. Robertson, Org. Lett., 2010, 12, 2818.
13. J. H. Christopher, S. Yamanoi and S. V. Ley, Org. Biomol. Chem., 2003, 10, 1664.
14. H. Nagai, Y. Mikami, K. Yazawa, T. Gonoi and T. Yasumoto, J. Antibiot., 1993, 46, 520.

---

Footnote

† Electronic supplementary information (ESI) available: NMR spectra of natural and synthetic compounds. See DOI: 10.1039/c3ra47796g

---