Spider Extracts. Subadult male and female Argiope bruennichi (Scopoli, 1772) (Araneae, Araneidae) were collected from natural meadows in Northern Germany (Buxtehude, Harmstorf, Pevestorf, Lower Saxony; Wedel, Schleswig-Holstein), between 24 June and 5 July 2019. A. bruennichi is common throughout Europe and its collection requires no permits. Spiders were transferred to the laboratory at the University of Hamburg, Germany, where they were individually housed in upturned plastic cups (250 or 500 mL depending on the spider’s size) with a hole in the bottom stuffed with cotton wool. Spiders were kept under natural light conditions at a constant temperature of 25°C and a relative humidity of 45 %. Twice a week, subadult spiders were provided with approximately 15 Drosophila spp. and adult females with three Calliphora sp. houseflies. Adult males were fed with approximately 10 Drosophila spp. once a week. All spiders were provided with water from a sprayer at least six days a week. Twenty virgin male (mean age ± SD: 13.5 ± 1.2 days) and 30 virgin female A. bruennichi (mean age ± SD: 11.8 ± 0.9 days) were used for chemical analysis.
Analysis by GC/MS. Spiders were cold anesthetized and stored at − 25° C until analysis. Cuticular extracts were prepared by individually placing females in 3 ml of dichloromethane (DCM; GC/MS grade, Merck, Darmstadt, Germany) and males in 1 ml DCM for one hour. Twenty virgin male (mean age ± SD: 13.5 ± 1.2 days) and 30 virgin female A. bruennichi (Mean age ± SD: 11.8 ± 0.9 days) were used for chemical analysis. To obtain silk samples, the females were placed into clean Perspex frames (35 x 35 x 6 cm) and allowed to build a web. After 24 h, females were removed from their webs and the silk was collected by slowly winding it around a glass Pasteur-pipet washed with ethanol. The tip of the pipet holding the silk was then snapped off into a small glass vial. Males and females were placed in clean glass vials and cold anaesthetized. All samples were stored at − 25°C until analysis. Cuticular extracts were prepared by individually placing females in 3 ml of dichloromethane (DCM; GC/MS grade, Merck, Darmstadt, Germany) and the smaller males in 1 ml DCM for one hour. Silk samples were extracted in 1 ml DCM. Extracts were concentrated by evaporation at room temperature to approximately 90 µl (females) and 50 µl (males and silk), respectively. An aliquot of 1 µl of each sample was analyzed by GC/MS on a Shimadzu GCMS-QP2010S system (Shimadzu Corporation, Kyoto, Japan). After individual analyses, the female silk and body extracts and male samples were pooled separately for further chemical identification. For the analysis of samples of individuals, the gas chromatograph was equipped with a SH-Rtx-5MS fused silica capillary column (30 x 0.25 mm ID, 0.25 µm film thickness; Shimadzu Corporation, Kyoto, Japan). The oven temperature raised from 80 to 260°C at a constant heating rate of 30°/min and from 260 to 300°C at a constant rate of 1°/min, with a 1-minute initial isothermal and a 10-minute final isothermal hold. A split-splitless injector was operated at 250°C in the splitless mode. Carrier gas was helium at a constant flow rate of 1 ml/min. The ionization voltage of the electron ionization mass spectrometer was 70 eV. Source temperature was 200°C and interface temperature was 280°C. Data acquisition and storage were performed with the software GCMSsolution (Version 4.45; Shimadzu Corporation, Kyoto, Japan). Peak areas were obtained by manual integration using the GCMSsolution software. Linear retention indices of all substances were calculated according to van den Dool and Kratz (1963). n-Alkanes were identified by comparing their mass spectra with those of authentic reference compounds. Alkenes were identified by their typical mass spectra. Methyl-branched hydrocarbons were identified by diagnostic ions resulting from their typical α-cleavage at the position of the methyl-branch and by a fragment at M-15 if the molecular ion was not detected. Moreover, their linear retention indices were compared to those published by (Carlson et al. 1998; El-Sayed 2019). Other compounds were tentatively identified by comparing their mass spectra and linear retention indices with those of a databases (NIST 08 mass spectral library 2008). Mean relative peak areas were calculated by standardizing total mean peak areas to 100%. GC/MS analysis of pooled samples were performed on a GC 7890A coupled to a MSD 5975C (Agilent Technologies, Germany). The gas chromatograph was equipped with a HP-5 MS column (Agilent, 30 m length, 0.25 mm internal diameter, 0.25 µm film thickness) with helium as the carrier gas. The combined samples were analyzed with a temperature program of 50°C for 5 min that increased by 3°C/min to 320°C with a final hold time of 10 min. Derivatized samples were analyzed with a temperature program of 50°C for 5 min, increased by 5°C/min to 320°C with a final hold time of 10 min. To compare the chemical composition between samples, relative peak areas were calculated for each of the 20 male and 30 female and web silk extracts separately by standardizing the total peak area of each extract to 100%. Values given in Table 1 are means ± standard deviation (SD). GC on chiral phases was performed using BetaDex™ 225 (Sigma, 30.0 m x 0.25 mm) or Hydrodex β-6TBDM (Machery and Nagel, 30.0 m x 0.25 mm) phases with a flow of 1.5 ml min− 1 hydrogen as the carrier gas and a flame ionization detector. Individual time programs are given at the appropriate Fig. 6 and Fig. S 5 in the Supporting Information.
Microreactions of Extracts. The extracts were derivatized in microreactions to obtain more structural information about methyl-branch positions of the acid and alcohol components of the long chain esters. Extracts were transesterified with trimethylsulfonium hydroxide (TMSH) (Müller et al. 1990) to form the corresponding methyl esters and free alcohols (Fig. 1). Subsequent transesterification of the methyl esters with sodium 3-pyridinylmethylate to form the corresponding pyridin-3-ylmethyl esters was performed (Harvey 1982) as well as esterification of the free alcohols with nicotinic acid to form the corresponding esters (Vetter and Meister 1981; Harvey 1991).
Transesterification with Trimethylsulfonium Hydroxide. Trimethylsulfonium hydroxide (TMSH, 100 µl, 0.25 m in methanol) was added to the natural sample (20 µl in dichloromethane) in a GC-vial (2 ml). The reaction mixture was placed in a heating block at 90°C for 6 h and regularly shaken vigorously. The solvents and reagents were removed with a stream of nitrogen and the residue was dissolved in DCM (20 µl) (Müller et al. 1990).
Transesterification with 3-Pyridinylmethanol. 3-Pyridinylmethanol (50 µl) was added to freshly cut sodium (0.2 mg) in a GC-vial (2 ml). The reaction mixture was heated to 80°C in a heating block until the sodium was dissolved. Sodium 3-pyridinylmethoxide (2 % in 3-pyridinylmethanol) was obtained as a syrupy yellow liquid. The thus prepared fresh sodium 3-pyridinylmethoxide (2 drops, 2 % in 3-pyridinylmethanol was added to the methyl ester sample (20 µl in dichloromethane) in a GC-vial (2 ml). The reaction was placed in a heating block at 80°C for 3 h and regularly shaken vigorously. Methanol (200 µl) and water (3 drops) was added to the solution and the mixture was extracted with pentane (3 × 200 µl). The pentane phases were combined, the solvent was removed in a stream of nitrogen and the residue was dissolved in DCM (20 µl).
Preparation of Nicotinates. DCM (50 µl), nicotinic acid (1 mg), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC, 1 mg) and 4-dimethylaminopyridine (DMAP, catalytic amount) were added to the sample of the methyl esters and free alcohols (50 µl in dichloromethane) in a GC-vial (2 ml). The reaction mixture was kept at room temperature for 2 h and regularly shaken vigorously. The solvents were removed in a stream of nitrogen and the residue was extracted with pentane (3 × 100 µl). The combined pentane phases were again concentrated in a stream of nitrogen and the residue was dissolved in DCM (50 µl).
Synthesis of 2,4-Dimethylheptadecanoic Acid. The synthesis is based on a general procedure developed by Feringa et al. for the stereoselective synthesis of homo-vicinal oligo-methyl-branched acids (Des Mazery et al. 2005; Horst et al. 2007; Ruiz et al. 2007). The synthesis of 2,4-dimethylheptadecanoic acid (9) started with compound 1 (synthesis described in the SI) which underwent an enantioselective 1,4-addition with methyl magnesium bromide using Josiphos (11) as ligand for CuBr (Fig. 2). The resulting thioester 2 was then reduced by Pd with Et3SiH as a hydrogen source, giving the corresponding aldehyde that underwent a Wittig reaction with Ph3PCHCOSEt (synthesis described in the SI) forming the α,βunsaturated thioester 3. Ester 3 was used as a substrate for the second enantioselective 1,4-addition with methylmagnesium bromide to form compound 4. Reduction with diisobutylaluminium hydride delivered the corresponding alcohol 5. Tosylate 6 was formed by reaction with tosyl chloride that undernwent substitution by C11H23MgBr to give compound 7. The cleavage of the silyl ether with tetra-n-butylammonium fluoride (TBAF) delivered alcohol 8 that was oxidized with RuO4 to finally furnish carboxylic acid 9. Esterification yielded tetradecyl (2S,4S)-2,4-dimethylheptadecanoate (10) as final product.
General Experimental Procedures. All reactions were performed in oven dried glassware under a nitrogen atmosphere. Solvents were dried according to standard procedures. Column chromatography: silica 60 (0.063–0.200 mm, 70–230 mesh ASTM). Thin layer chromatography (TLC): Polygram® SIL G/UV silica 60, 0.20 mm. Compounds were stained with potassium permanganate solution. NMR spectra were recorded either on Avance III HD 300N (1HNMR: 300 MHz, 13CNMR: 76 MHz), DRX 400 (1HNMR: 400 MHz, 13CNMR: 101 MHz), AVII 400 (1HNMR: 400 MHz, 13CNMR: 101 MHz) or AVII 600 (1HNMR: 600 MHz) instruments. Data are reported as follows: chemical shifts, multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet), coupling constants (Hz). IR spectra were measured on a Bruker Tensor 27 (diamond-ATR). Mass spectra were recorded with a combination of an Agilent Technologies 5977B gas chromatograph connected to an Agilent Technologies 8860 Series MSD. Optical rotation was determined with the help of an MCP 150 polarimeter (Anton Paar) with a cell length of 1 cm (c given in mg/mL).
Preparation of S-Ethyl (S)-4-((tert-Butyldiphenylsilyl)oxy)-3-methylbutanethioate ( 2 ). The synthesis was performed according to Horst et al. (2007) using (R)-1-[(SP)-2-(diphenylphosphino)ferrocenyl]ethyldicyclohexylphosphine (Josiphos, 11, 50 mg, 0.08 mmol, 0.012 eq.) and CuBr·SMe2 (13 mg, 0.065 mmol, 0.01 eq.), methyl tert-butyl ether (MTBE, 50 mL), MeMgBr (7.78 mmol, solution in diethyl ether), and S-ethyl (E)-4-((tert-butyldiphenylsilyl)oxy)but-2-enethioate (6.48 mmol, 2493 mg, 1 eq.) to afford 2 as a colorless oil (2199 mg, 85%). = −4.60 (10 mg/mL; CH2Cl2). FT-IR: ν / cm− 1 = 2960, 2895, 2859, 1688, 1466, 1427, 1389, 1261, 1150, 1108, 1040, 1005, 941, 821, 802, 763, 741, 703, 615. 1H-NMR: (400 MHz, CDCl3) δ / ppm = 7.71–7.66 (m, 4H), 7.47–7.37 (m, 6H), 3.53 (dddd, J = 16.2, 9.9, 5.7 Hz, 2H), 2.93–2.82 (m, 3H), 2.40 (dd, J = 14.4, 8.4 Hz, 1H), 2.36–2.25 (m, 1H), 1.26 (t, J = 7.4 Hz, 3H), 1.08 (d, J = 2.8 Hz, 9H), 0.98 (d, J = 6.6 Hz, 3H). 13C-NMR, DEPT: (101 MHz, CDCl3) δ / ppm = 199.2 (C = O), 135.7 (CHAr), 135.7 (CHAr), 133.8 (CHAr), 133.8 (CAr), 129.7 (CHAr), 127.7 (CHAr), 68.0 (CH2), 47.7 (CH2), 33.9 (CH), 27.0 (CH3), 23.4 (CH2), 19.4 (C), 16.6 (CH3), 14.9 (CH3). EI-MS (70 eV): m/z = 344 (23), 343 ([M − tBu]+, 76), 244 (26), 243 (100), 197 (13), 183 (39), 181 (22), 135 (25), 137 (23), 105 (16). The enantiomeric excess (ee) was determined by cleavage of the silyl ether using tetrabutylammonium fluoride (TBAF) (Horst et al. 2007). The formed lactone was separated by GC on a chiral phase (30.0 m x 0.25 mm, initial temp. 50°C then 10°C min− 1 to final temp. 160°C, Fig. S 5). Retention time, 13.75 min (minor), 13.85 (major) showed 94% ee (Lit.: 98% ee Horst et al. 2007).
Preparation of S-Ethyl (S,E)-6-((tert-Butyldiphenylsilyl)oxy)-5-methylhex-2-enethioate ( 3 ). The synthesis was performed as described by Horst et al. (2007) using Et3SiH (36.69 mmol, 4266 mg, 3 eq.), 2 (12.23 mmol, 4900 mg, 1 eq.) and 10% Pd/C (5 mol%, 650 mg) in CH2Cl2 (20 mL) to afford the crude aldehyde that underwent a Wittig-reaction with S-ethyl 2-(triphenyl-λ5-phosphaneylidene)ethanethioate (3.221 mmol, 1174 mg) in CH2Cl2 (40 mL) to afford 3 as a colorless oil (2010 mg, 39% over two steps). = −6.2 (10 mg/mL; CHCl3). 1H-NMR: (300 MHz, CDCl3) δ / ppm = 7.70–7.60 (m, 4H), 7.46–7.33 (m, 6H), 6.93–6.79 (m, 1H), 6.11 (d, J = 15.5 Hz, 1H), 3.49 (ddd, J = 16.3, 10.0, 5.9 Hz, 2H), 2.94 (q, J = 7.4 Hz, 2H), 2.50–2.36 (m, 1H), 2.04 (ddd, J = 15.4, 8.4, 7.3 Hz, 1H), 1.94–1.77 (m, 1H), 1.28 (dd, J = 7.7, 7.2 Hz, 3H), 1.06 (s, 9H), 0.91 (d, J = 6.8 Hz, 3H). 13C-NMR, DEPT: (76 MHz, CDCl3) δ / ppm = 190.1 (C), 144.0 (CH), 135.7 (CH), 135.7 (CH), 133.9 (CH), 133.8 (CH), 130.1, 129.8 (CH), 127.8 (CH), 68.2 (CH2), 36.1 (CH2), 35.6 (CH), 27.0 (CH3), 23.2 (CH2), 19.4 (C), 16.6 (CH3), 15.0 (CH3). EI-MS (70 eV): m/z = 283 ([M − tBu]+, 31), 200 (19), 199 (100), 181 (24), 175 (16), 139 (38), 105 (18), 83 (23), 77 (19), 41 (16).
Preparation of S-Ethyl (3R,5S)-6-((tert-Butyldiphenylsilyl)oxy)-3,5-dimethylhexanethioate ( 4 ). The synthesis was performed according to Horst et al. (2007) using 11 (75 mg, 0.12 mmol, 0.012 eq.), CuBr·SMe2 (20 mg, 0.097 mmol, 0.01 eq.) dissolved in MTBE (65 mL), MeMgBr (11.64 mmol, solution in diethyl ether), and 3 (9.70 mmol, 3730 mg, 1 eq.) to afford 4 as a colorless oil (3533 mg, 82%). = −4.3 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 2959, 2930, 2860, 1689, 1463, 1427, 1384, 1262, 1108, 1083, 1002, 821, 741, 702, 615. 1H-NMR: (300 MHz, CDCl3) δ / ppm = 7.72–7.66 (m, 4H), 7.47–7.36 (m, 6H), 3.48 (ddd, J = 24.5, 9.8, 5.9 Hz, 2H), 2.88 (q, J = 7.5 Hz, 2H), 2.53 (dd, J = 14.3, 5.0 Hz, 1H), 2.26 (dd, J = 14.3, 8.7 Hz, 1H), 2.18–2.02 (m, 1H), 1.81–1.65 (m, 1H), 1.48–1.35 (m, 1H), 1.30–1.21 (m, 3H), 1.12–1.05 (m, 9H), 1.05–0.98 (m, 1H), 0.93 (dt, J = 6.7, 4.5 Hz, 6H). 13C-NMR, DEPT: (76 MHz, CDCl3) δ / ppm = 199.3 (C = O), 135.8 (CHAr), 134.1 (CAr), 134.1 (CAr), 129.7 (CHAr), 127.7 (CHAr), 68.9 (CH2), 51.3 (CH2), 40.9 (CH2), 33.3 (CH), 28.8 (CH), 27.0 (CH3), 23.4 (CH2), 20.4 (CH3), 19.4 (Cq), 17.6 (CH3), 14.9 (CH3). EI-MS (70 eV): m/z = 386 (30), 385 ([M − tBu]+, 100), 323 (31), 243 (44), 199 (92), 183 (50), 181 (35), 135 (41), 83 (35), 55 (36).
Preparation of (3R,5S)-6-((tert-Butyldiphenylsilyl)oxy)-3,5-dimethylhexan-1-ol ( 5 ). To a solution of 4 (0.90 mmol, 400 mg, 1 eq.) in CH2Cl2 (10 mL) was added diisobutylaluminium hydride (1.17 mmol, 1.3 eq., solution in cyclohexane) at − 50°C under a nitrogen atmosphere. After stirring for 17 h the reaction mixture was allowed to warm up to room temperature. The reaction was quenched by addition of saturated Rochelle solution (10 mL) and stirred for 30 min at room temperature. The phases were separated and the aqueous phase was extracted three times with CH2Cl2 (30 mL). The combined organic phases were dried over Na2SO4 and the solvent removed under reduced pressure to yield the crude aldehyde. The reduction procedure was repeated and the obtained residue was purified by column chromatography (pentane/diethyl ether; 1:1) to afford alcohol 13 as a slightly yellow oil (267 mg, 77%). In contrast to the published procedure (Horst et al. 2007) a two-step reduction proved to be necessary to obtain good yields. = −3.7 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 2928, 2859, 1466, 1428, 1385, 1107, 1007, 821, 739, 700, 614. 1H-NMR: (300 MHz, CDCl3) δ / ppm = 7.69 (m, 4H), 7.48–7.36 (m, 6H), 3.74–3.58 (m, 2H), 3.57–3.40 (m, 2H), 1.86–1.68 (d, J = 6.6 Hz, 1H), 1.68–1.52 (m, 2H), 1.49–1.22 (m, 3H). 13C-NMR, DEPT: (76 MHz, CDCl3) δ / ppm = 135.8 (CH), 135.8 (C), 134.2 (CH), 129.6 (CH), 127.7 (CH), 68.9 (CH2), 61.2 (CH2), 41.3 (CH2), 39.9 (CH2), 33.2 (CH3), 27.1 (CH3), 27.0 (CH3), 20.4 (CH3), 19.4 (Cq), 17.8 (CH3). EI-MS (70 eV): m/z = 327 ([M − tBu]+, 2), 200 (11), 199 (63), 181 (14), 139 (11), 135 (10), 111 (37), 69 (100), 57 (10), 55 (39), 41 (19).
Preparation of (3R,5S)-6-((tert-Butyldiphenylsilyl)oxy)-3,5-dimethylhexyl 4-Methylbenzenesulfonate ( 6 ). Tosyl chloride (1.35 mmol, 257 mg, 2 eq.) was added to a solution of 5 (0.67 mmol, 259 mg, 1 eq.) and pyridine (1.35 mmol, 109 µL, 2 eq.) in CH2Cl2 (5 mL). The reaction mixture was stirred at room temperature for 19 h under a nitrogen atmosphere. The solvent was removed under reduced pressure and the resulting residue was purified by column chromatography (pentane/diethyl ether; 20:1) to afford 6 as a colorless oil (289 mg, 80%). = −4.2 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 2955, 2923, 2854, 1463, 1428, 1389, 1377, 1362, 1110, 1007, 999, 824, 795, 738, 700, 671, 665, 614, 528. 1H-NMR: (300 MHz, CDCl3) δ / ppm = 7.81–7.75 (m, 2H), 7.70–7.63 (m, 4H), 7.48–7.27 (m, 8H), 4.12–3.96 (m, 2H), 3.42 (ddd, J = 16.1, 9.8, 5.9 Hz, 2H), 2.43 (s, 3H), 1.77–1.44 (m, 3H), 1.32 (ddt, J = 13.5, 10.8, 6.6 Hz, 3H), 1.05 (s, J = 2.6 Hz, 9H), 0.99–0.81 (m, 4H), 0.77 (d, J = 6.5 Hz, 3H). 13C-NMR, DEPT: (76 MHz, CDCl3) δ / ppm = 144.7 (CAr), 135.7 (CHAr), 134.1 (CAr), 133.4 (CAr), 129.9 (CHAr), 129.7 (CHAr), 128.0 (CHAr), 127.7 (CHAr), 69.2 (CH2), 68.8 (CH2), 41.1 (CH2), 35.7 (CH2), 33.1 (CH), 27.0 (CH3), 27.0 (CH), 21.7 (CH3), 19.9 (Cq), 19.4 (CH3), 17.7 (CH3). EI-MS (70 eV): m/z = 353 (35), 293 (66), 199 (52), 181 (20), 135 (21), 111 (58), 69 (100), 91 (51), (48), 41 (28).
Preparation of tert-Butyl(((2S,4S)-2,4-dimethylheptadecyl)oxy)diphenylsilane ( 7 ). 1-Bromoundecane was added (2.13 mmol, 500 mg) to a mixture of magnesium turnings (2.55 mmol, 62 mg) in THF (10 mL). The reaction mixture was heated to refluxed for 30 min and cooled down to room temperature. Compound 6 (0.51 mmol, 273 mg, 1 eq.) and CuBr·SMe2 (0.10 mmol, 21 mg, 0.2 eq.) were dissolved in THF (6 mL). The freshly prepared Grignard solution was added at 0°C under a nitrogen atmosphere. After warming to room temperature, the reaction mixture was stirred for 1 h. After quenching with saturated NH4Cl solution (6 mL) the phases were separated and the aqueous phase was extracted three times with diethyl ether (30 mL). The combined organic phases were dried over Na2SO4 and the solvent was removed under reduced pressure. The residue was purified by column chromatography (pentane/diethyl ether; 20:1) to afford 7 as a colorless oil (113 mg, 42%). = −3.3 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 3067, 2957, 2929, 2860, 2323, 1596, 1465, 1429, 1361, 1300, 1258, 1213, 1180, 1103, 945, 892, 816, 744, 701, 663, 614, 578, 555. 1H-NMR: (300 MHz, CDCl3) δ / ppm = 7.73–7.63 (m, 4H), 7.50–7.31 (m, 6H), 3.47 (ddd, J = 16.2, 9.8, 5.9 Hz, 2H), 1.74 (dh, J = 13.3, 6.7 Hz, 1H), 1.50–1.19 (m, 25H), 1.10–1.03 (m, 9H), 0.96–0.85 (m, 7H), 0.82 (d, J = 6.4 Hz, 3H). 13C-NMR, DEPT: (76 MHz, CDCl3) δ / ppm = 135.8 (CHAr), 134.3 (CAr), 129.6 (CHAr), 127.7 (CHAr), 69.1 (CH2), 41.3 (CH2), 37.0 (CH2), 33.3 (CH), 32.1 (CH2), 30.2 (CH2), 30.2 (CH), 29.9 (CH2), 29.9 (CH2), 29.8 (CH2), 29.5 (CH2), 27.0 (CH3), 27.0 (CH2), 22.9 (CH2), 20.5 (CH3), 19.5 (Cq), 17.9 (CH3), 14.3 (CH3). EI-MS (70 eV): m/z = 465 (34), 200 (19), 199 (100), 97 (23), 83 (25), 83 (19), 69 (28), 57 (40), 55 (20), 43 (37).
Preparation of (2S,4S)-2,4-Dimethylheptadecan-1-ol ( 8 ). TBAF (0.57 mmol, 574 µL, 3 eq.) was added to a solution of 7 (0.19 mmol, 100 mg, 1 eq.) in THF (5 mL). The reaction mixture was stirred at room temperature for 24 h under a nitrogen atmosphere. The solvent was separated under reduced pressure and the residue purified by column chromatography (pentane/diethyl ether; 20:1) to afford 8 as colorless oil with traces of siloxanes as impurities (21 mg, crude). = −37.6 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 3341, 2955, 2922, 2853, 1462, 1377, 1112, 1036, 987, 865, 821, 704, 606. 1H-NMR: (600 MHz, CDCl3) δ 3.49–3.27 (m, 2H), 1.70–1.60 (m, 1H), 1.46–1.37 (m, 1H), 1.27–1.10 (m, 26H), 1.09–0.91 (m, 2H), 0.90–0.75 (m, 10H). 13C-NMR, DEPT: (151 MHz, CDCl3) δ / ppm = 68.2 (CH2), 40.9 (CH2), 36.5 (CH2), 32.9 (CH), 31.7 (CH2), 29.9 (CH), 29.8 (CH2), 29.5 (CH2), 29.5 (CH2), 29.5 (CH2), 29.2 (CH2), 26.7 (CH2), 22.5 (CH2), 20.2 (CH3), 17.1 (CH3), 13.9 (CH3). EI-MS (70 eV): m/z = 283 (< 1), 266 (< 1), 224 (10), 209 (6), 196 (7), 168 (7), 111 (22), 97 (31), 83 (100), 71 (44), 70 (39), 69 (52), 57 (97), 56 (60), 55 (85), 43 (81), 41 (61).
Preparation of (2S,4S)-2,4-Dimethylheptadecanoic acid ( 9 ). Under a nitrogen atmosphere RuCl3 (0.014 mmol, 3 mg, 0.3 eq.) and NaIO4 (0.281 mmol, 60 mg, 5 eq.) were added to a mixture of crude 8 (0.056 mmol, 16 mg, 1 eq.), H2O (1.2 mL), CH3CN (1.2 mL) and CCl4 (2.4 mL). The mixture was stirred at room temperature for 3.5 h. After addition of CH2Cl2 (4 mL) and H2O (1 mL) the phases were separated and the aqueous phase was extracted three times with CH2Cl2 (10 mL). The combined organic phases were dried over Na2SO4. The solvent was removed under reduced pressure and the residue was purified by column chromatography (pentane/ethyl acetate/acetic acid; 90:10:1) to afford acid 9 as a colorless oil (11 mg, 66%). = +7.0 (10 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 2956, 2923, 2853, 1815, 1707, 1464, 1416, 1379, 1290, 1235, 1091, 1018, 947, 810, 722, 529. 1H-NMR: (600 MHz, CDCl3) δ / ppm = 9.58 (d, J = 2.5 Hz, 1H), 2.67–2.51 (m, 1H), 1.80–1.66 (m, 1H), 1.53–1.41 (m, 1H), 1.40–1.02 (m, 31H), 0.95–0.79 (m, 7H). 13C-NMR, DEPT: (151 MHz, CDCl3) δ / ppm = 172.7 (C), 41.4 (CH2), 37.2 (CH2), 32.1 (CH2), 30.9 (CH2), 30.1 (CH), 29.9 (CH2), 29.5 (CH2), 26.9 (CH2), 22.9 (CH2), 19.7 (CH3), 18.0 (CH3), 14.27 (CH3). A small portion was converted into the respective methyl ester by treatment with trimethylsilyldiazomethane. EI-MS (70 eV): m/z = 312 ([M]+, 3), 241 (12), 129 (7), 101 (59), 88 (100), 71 (7), 69 (12), 57 (13), 55 (12), 43 (12), 41 (10).
Preparation of tetradecyl (2S,4S)-2,4-dimethylheptadecanoate ( 10 ). 1-Ttetradecanol (0.007 mmol, 0.53 mg, 1 eq., 1%wt in DCM), dicyclohexylcarbodiimide (0.007 mmol, 1.28 mg, 1 eq., 1%wt in DCM) and 4-dimethylaminopyridine (0.007 mmol, 1.38 mg, 1 eq., 1%wt in DCM) were added to a solution of (2S,4S)-2,4-dimethylheptadecanoic acid (9) (0.007 mmol, 2 mg, 1 eq.) in DCM (5 mL). After stirring for 20 h at room temperature the solvent was removed under reduced pressure and the residue was purified by column chromatography (pentane/diethyl ether; 100:1) to afford 11 as a colorless oil (2 mg, 58%).= +8.0 (2 mg/mL; CHCl3). FT-IR: ν / cm− 1 = 2923, 2855, 2319, 1736, 1461, 1372, 1172, 673, 603, 565, 546. 1H-NMR: (500 MHz, CDCl3) δ / ppm = 4.11–3.99 (m, 2H), 2.58–2.46 (m, 1H), 1.74–1.66 (m, 1H), 1.65–1.57 (m, 3H), 1.42–1.17 (m, 38H), 1.16–1.05 (m, 5H), 0.91–0.78 (m, 10H). 13C-NMR, DEPT: (126 MHz, CDCl3) δ / ppm = 177.4 (Cq), 64.4 (CH2), 41.8 (CH2), 37.7 (CH), 37.3 (CH2), 32.1 (CH2), 31.0 (CH), 30.1 (CH2), 29.9 (CH2), 29.8 (CH2), 29.8 (CH2), 29.7 (CH2), 29.5 (CH2), 29.4 (CH2), 28.9 (CH2), 27.0 (CH2), 26.1 (CH2), 22.9 (CH2), 19.7 (CH3), 18.2 (CH3), 14.3 (CH3). EI-MS (70 eV): m/z (%) = 299 (100), 241 (17), 196 (23), 111 (14), 97 (24), 87 (50), 83 (25), 75 (30), 74 (61), 71 (36), 69 (31), 57 (55), 55 (57), 43 (41), 41 (17).