Total synthesis of lycopene-5,6-diol and γ-carotene-5′,6′-diol stereoisomers and their HPLC separation

Article abstract of DOI:10.1016/j.tet.2017.02.048

Lycopene-5,6-diol stereoisomers (1a,b) and (2a,b) and γ-carotene-5′,6′-diol stereoisomers (3a,b) and (4a,b) were synthesized by a stepwise C₁₅?+?C₁₀?+?C₁₅ double Wittig reaction strategy. The key compounds erythro(anti)-C₁₅-dihydroxy aldehydes 17a,b and their threo(syn)-stereoisomers 23a,b were prepared via Sharpless asymmetric epoxidation of geraniol and nerol followed by acidic hydrolysis of the epoxides in a stereospecific manner. The enantiomerically enriched anti-isomers were obtained by way of recrystallization of 2,3-epoxygeranyl 3,5-dinitrobenzoates 9a,b, whereas syn-isomers were obtained as enantiomerically pure forms via recrystallization of dihydroxyneryl 3,5-dinitrobenzoates 21a,b. In order to determine the absolute stereochemistry of natural products, HPLC separation methods for each enantiomers 1a,b–4a,b were established by using a column carrying a chiral stationary phase.

Full text of DOI:10.1016/j.tet.2017.02.048

Accepted Manuscript
Total synthesis of lycopene-5,6-diol and γ-carotene-5′,6′-diol stereoisomers and their
HPLC separation
Yumiko Yamano, Yuya Nishiyama, Atsushi Aoki, Takashi Maoka, Akimori Wada
PII:
S0040-4020(17)30187-4
10.1016/j.tet.2017.02.048
TET 28489
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 17 January 2017
Revised Date: 19 February 2017
Accepted Date: 21 February 2017
Please cite this article as: Yamano Y, Nishiyama Y, Aoki A, Maoka T, Wada A, Total synthesis of
lycopene-5,6-diol and γ-carotene-5′,6′-diol stereoisomers and their HPLC separation, Tetrahedron
(2017), doi: 10.1016/j.tet.2017.02.048.
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O
OH
6
O
*
*
NO2
5
O
O
*
OH
four lycopene-5,6-diol stereoisomers
NO2
ACCEPTED MANUSCRIPT
OH
O
*
6'
NO2
5'
O
OH
four γ-carotene-5',6'-diol stereoisomers
NO2

ACCEPTED MANUSCRIPT
Total synthesis of lycopene-5,6-diol and γ-carotene-5’,6’-diol
stereoisomers and their HPLC separation
Yumiko Yamano,a, Yuya Nishiyama,a Atsushi Aoki,a Takashi Maoka,b and Akimori Wadaa
a Department of Organic Chemistry for Life Science, Kobe Pharmaceutical University, 4-19-1
Motoyamakita-machi, Higashinada-ku, Kobe 658-8558, Japan
b Research Institute for Production Development, 15 Shimogamo-morimoto-cho, Sakyo-ku, Kyoto 606-0805,
Japan
ABSTRACT
Lycopene-5,6-diol stereoisomers (1a,b) and (2a,b) and γ-carotene-5’,6’-diol stereoisomers (3a,b) and
(4a,b) were synthesized by a stepwise C15 + C10 + C15 double Wittig reaction strategy. The key
compounds erythro(anti)-C15-dihydroxy aldehydes 17a,b and their threo(syn)-stereoisomers 23a,b
were prepared via Sharpless asymmetric epoxidation of geraniol and nerol followed by acidic
hydrolysis of the epoxides in a stereospecific manner. The enantiomerically enriched anti-isomers
were obtained by way of recrystallization of 2,3-epoxygeranyl 3,5-dinitrobenzoates 9a,b, whereas
syn-isomers were obtained as enantiomerically pure forms via recrystallization of dihydroxyneryl
3,5-dinitrobenzoates 21a,b. In order to determine the absolute stereochemistry of natural products,
HPLC separation methods for each enantiomers 1a,b–4a,b were established by using a column
carrying a chiral stationary phase.
Keywords:
Carotenoids
Lycopene-5,6-diol
γ-Carotene-5’,6’-diol
Total synthesis
HPLC separation
Corresponding author. Tel.: +81-78-441-7562; fax: +81-78-441-7562; e-mail: y-yamano@kobepharma-u.ac.jp
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1. Introduction
Lycopene (Fig. 1), the major carotenoid found in tomatoes and tomato-based food products, is
expected to exhibit physiological activities, especially in cancer prevention. It has been reported that
lycopene exhibits excellent antioxidant activities1 and growth-inhibitory effects on several human
cancer cells.2 Furthermore, epidemiological studies have shown that the uptake of lycopene reduces
the risk of prostate cancer.3
Fig. 1. Lycopene, γ-carotene and their new oxidative metabolites.
Several oxidative metabolites of lycopene, including structurally unique compounds carrying a
five-membered ring end-group, have been isolated from human serum and tomato products.4 The
metabolites are considered to be formed via lycopene-5,6-epoxide as shown in Fig. 2. As lycopene is
shown convincingly to be beneficial in biological and nutritional activities, biological activities of its
metabolites have also attracted attention over years. In our intensive research on oxidative products of
lycopene and γ-carotene, we have isolated lycopene-5,6-diols (1), (2) and γ-carotene-5’,6’-diols (3),
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ACCEPTED MANUSCRIPT
(4) for the first time from the aril of gac Momordica cochinichinesis.5 Gac is a deciduous vine that
grows in South Asia and has been used traditionally as both food and folk medicine. Its aril contains
lycopene with a higher content and several carotenoids. The relative stereochemistries of diol moiety
in these carotenoids were estimated by NOESY and NOE difference experiments and molecular
calculation using the ab initio molecular orbital method.5 To give validation to their stereochemistries,
we have employed synthetic approach. Following general description of the total synthesis of
(5S,6R)-lycopene-5,6-diol (1b) and its (5S,6S)-isomer 2a in the previous report,5 we would like to
report here stereospecific synthesis of the all stereoisomers of lycopene-5,6-diol (1a, 1b, 2a, 2b) and
γ-carotene-5’,6’-diol (3a, 3b, 4a, 4b) and establishment of HPLC analysis method by using a column
carrying a chiral stationary phase.
Fig. 2. Proposed metabolic pathway of lycopene.
2. Results and discussion
We planned to synthesize the all stereoisomers of lycopene-5,6-diol (1a,b, 2a,b) and
γ-carotene-5’,6’-diol (3a,b, 4a,b) by Wittig reaction of known C15-phosphonium salts 56 and 67 with
C25-dihydroxy apocarotenals A, which would be also prepared by condensation between
C15-dihydroxy aldehydes B and C10-phosphonium salt 78 as shown in Scheme 1. The key
C15-dihydroxy aldehydes B having a desired stereochemistry were envisioned to prepare via Sharpless
asymmetric epoxidation of geraniol and nerol and subsequent regio- and stereoselective acidic
hydrolysis of epoxides C and D.
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Scheme 1. Synthetic plan for lycopene-5,6-diols (1a,b and 2a,b) and
γ-carotene-5’,6’-diols (3a,b and 4a,b).
Synthetic routes for erythro(anti)-lycopene-5,6-diols (1a,b) and erythro(anti)-γ-carotene-5’,6’-diols
(3a,b) are shown in Scheme 2. The Sharpless asymmetric epoxidation of geraniol afforded
(2S,3S)-epoxygeraniol 8a and (2R,3R)-8b in good yields.9–11 However, their reported enantiomeric
purities are moderate (81–91% ee).9–11 In order to improve them, these alcohols were subjected to
3,5-dinitrobenzoylation and subsequent recrystallization from methanol to give benzoates 9a and 9b as
needles with 98% ee, respectively. These enantiomeric excesses were determined by HPLC analysis
[CHIRALPAK ID; Daisel, EtOH–hexane (2:8), 30 °C].
The (2S,3S)-benzoate 9a was next treated with aqueous perchloric acid in N,N-dimethylformamide
(DMF)11–13 to provide the desired anti-diol 10a (66%), accompanied by its formate 11a (21%). Their
stereochemistries were confirmed by transformation into the known triol 12a11 by treatment with
methanol in the presence of triethylamine.14 Anti-configuration of 10a was also confirmed as
mentioned later. Attempted conversion of the formate 11a to the diol 10a was unsuccessful. Thus, the
epoxide 9a was treated with catalytic amount of p-tolutenesulfonic acid (p-TsOH) in MeCN-H2O
under heating to provide the anti-diol 10a in 74% yield, accompanied by small amount of recovered
epoxide. The diol 10a was next converted into aldehyde 14a by the sequence of acetonide-protection
of the diol moiety, alkaline hydrolysis and Dess-Martin oxidation. NOESY experiments of aldehyde
14a shown in Scheme 2 indicated that the configuration of the diol moiety in 10a is erythro(anti) form
and thus the epoxide ring of 9a is selectively opened with inversion of configuration at the tertiary
γ-position.
The C10-aldehyde 14a was then condensed with C5-phosphonate 15, the resulting C15-dienoate 16a
was subjected to LiAlH4 reduction followed by MnO2 oxidation and subsequent
acetonide-deprotection to provide an isomeric mixture (9E/9Z ~2/1)15 of dihydroxy aldehyde 17a
4

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without epimerization. Without separation of the geometric isomers, the aldehyde 17a was condensed
with C10-phosphonium salt 78 and then briefly treated with acidic water16 to afford an isomeric mixture
of C25-apocarotenal 18a. As HPLC analysis of this mixture showed that the all-E-isomer was minor,
the mixture was treated with a palladium catalyst17,18 to yield a mixture in which the all-E-isomer
constituted 51%. After purification by preparative HPLC, the resulting all-E-apocarotenal 18a was
condensed with phosphonium salt 56 and followed by isomerization to give (5R,6S)-lycopene-5,6-diol
(1a) in 70% yield, as a geometrical isomeric mixture (all-E: 51%). Crystallization from
CH2Cl2-Et2O-hexane afforded all-E-isomer of (5R,6S)-1a as a red powder, whose 1H NMR spectral
data were identical with those of natural carotenoid isolated from gac.5 Whereas the apocarotenal 18a
was condensed with phosphonium salt 67 to afford (5’R,6’S)-γ-carontene-5’,6’-diol (3a), whose 1H
NMR spectral data were good accordance with those of natural carotenoid.5
The antipodes (5S,6R)-1b and (5’S,6’S)-3b were synthesized in a similar fashion from
(2R,3R)-epoxygeranyl benzoate 9b.
Scheme 2. Synthesis of erythro-lycopene-5,6-diols (1a,b) and erythro-γ-carotene-5’,6’-diols (3a,b).
5

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Next, threo(syn)-lycopene-5,6-diols (2a,b) and threo(syn)-γ-carotene-5’,6’-diols (4a,b) were
synthesized starting from optically active 2,3-epoxynerols 19a,b10,19 as shown in Scheme 3. The
reported enantiomeric purities of 2,3-epoxynerols 19a,b prepared by Sharpless asymmetric
epoxidation are lower (62–77% ee)10,19 than those of 2,3-epoxygeraniols 8a,b (81–91% ee).9–11
Although Mori et al.19 reported that enantiomeric purities can be improved by repeated
recrystallization of their 3,5-dinitorobenzoates 20a,b and chromatographic purification of the crude
products obtained from the mother liquor, this purification is troublesome. We found that
recrystallization of 2,3-dihydroxyneryl 3,5-dinitrobenzoates 21a,b, prepared by acidic hydrolysis of
2,3-epoxyneryl benzoate 20a,b, from methanol-water (4:1) directly afford enantiomerically pure
crystals (97–99% ee). These enantiomeric excesses were determined by HPLC analysis [CHIRALPAK
IA; Daisel, 2-PrOH–hexane (3:7), 30 °C].
In the same procedure as preparation of erythro(anti)-carotenoids 1a,b and 3a,b, 2,3-dihydroxyneryl
benzoates 21a,b were transformed into threo(syn)-lycopene-5,6-diols (2a,b) and
threo(syn)-γ-carotene-5’,6’-diols (4a,b), whose 1H NMR spectral data were identical with those of
natural carotenoid isolated from gac.5 Their syn-configurations were confirmed by NOESY
experiments of aldehyde 22a shown in Scheme 3.
Scheme 3. Synthesis of threo-lycopene-5,6-diols (2a,b) and threo-γ-carotene-5’,6’-diols (4a,b).
6

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Next, HPLC conditions for identification of each enantiomers 1a,b–4a,b were established by using
a column carrying a chiral stationary phase as shown in Figs. 3 and 4. Diasteromers 1a,b and 2a,b and
also 3a,b and 4a,b can be easily separated by usual silica gel HPLC column.5 Thus combination of
two HPLC conditions enables to identify two absolute configurations, being applicable to a natural
product,20 if it is available.
In sumarry, we succeeded in the total synthesis of lycopene-5,6-diol stereoisomers (1a,b) and (2a,b)
and γ-carotene-5’,6’-diol stereoisomers (3a,b) and (4a,b) in a stereospecific manner. Relative
stereochemistries of the diol moiety of natural products could be confirmed5 by comparison of their
NMR spectral data with those of synthetic samples. HPLC separation methods for each enantiomers
were also establised toward deteremination of the absolute stereochemistries of natural products.
These synthetic carotenoids were expected to become useful authentic samples not only for the further
studies of metabolisms of lycopene and γ-carotene in various tissues but also for the investigation of
their physiological functions.
Fig. 3. HPLC elution profiles of lycopene-5,6-diol stereoisomers (1a,b) and (2a,b). Column:
CHIRALPAK AD-H (daicel) 0.46 × 25 cm; elutent: 2-PrOH-hexane (15:85); flow rate: 1.0 mL/min;
temperature: 25 °C; detection: 450 nm.
Fig. 4. HPLC elution profiles of γ-carotene-5’,6’-diol stereoisomers (3a,b) and (4a,b). Column:
CHIRALPAK AD-H (daicel) 0.46 × 25 cm; elutent: 2-PrOH-hexane (15:85); flow rate: 0.6 mL/min;
temperature: 25 °C; detection: 450 nm.
7

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3. Experimental
3.1 General
UV-VIS spectra were recorded on a JASCO V-650 instrument. IR spectra were measured on a
1
Perkin-Elmer spectrum 100 FT-IR spectrometer. H and 13C NMR spectra were determined on a
Varian Gemini-300 or a VXR-500 superconducting FT-NMR spectrometer. The chemical sifts are
expressed in ppm relative to tetramethylsilane (δ = 0) as internal standard for 1H NMR and CDCl3 (δ =
77.0) for 13C NMR. Mass spectra were taken on a Thermo Fisher Scientific Exactive spectrometer.
Optical rotations were measured on a JASCO P-2200 porarometer and CD spectra on a
Shimadzu-AVIN 62A DS circular dichroism spectrometer.
Flash column chromatography (CC) was performed on using Kanto Silica Gel 60 N. Preparative
HPLC was carried out on a Shimadzu LC-6A with a UV-VIS detector.
All operations were carried out under nitrogen or argon. Evaporation of the extract or the filtrate
was carried out under reduced pressure. In solvent extraction procedure, organic layer was dried over
anhydrous Na2SO4. Ether refers to diethyl ether, and hexane to n-hexane. NMR assighments of
C15-ester 16, C15-aldehydes 17 and 23, C25-apocarotenals 18 and 24, and carotenoids 1–4 are given
using the carotenoid numbering system.
3.1.1.
(2S,3S)-
and
(2R,3R)-3-Methyl-3-(4-methylpent-3-en-1-yl)oxiran-2-yl)methyl
3,5-dinitorobenzoate 9a,b. (2S,3S)-Epoxygeraniol 8a (84% ee) and (2R,3R)-epoxygeraniol 8b (88%
ee) were prepared according to the known Sharpless asymmetric epoxidation method of geraniol.9–11
To a stirred solution of (2S,3S)-8a (84% ee; 9.30 g, 54.7 mmol), Et3N (11.4 mL, 82 mmol) and DMAP
(133 mg, 1.1 mmol) in dry CH2Cl2 (60 mL) was added 3,5-dinitrobenzoyl chloride (13.2 g, 57.3
mmol) in some portions at 0 °C and the mixture was stirred for a further 30 min. After CH2Cl2 was
evaporated off, the resulting mixture was diluted with AcOEt and washed with water and then brine.
The organic layer was dried and evaporated to give a residue, which was purified by flash CC
(AcOEt-hexane, 1:3) to provide the benzoate 9a (19.33 g, 97%) as a pale yellow solid.
Recrystallization from MeOH (90 mL) gave 16.03 g of (2S,3S)-9a (99% ee) as a pale yellow needle.
(2R,3R)-9b was also prepared (97%) by the same procedure. The enantiomeric purities of 9a and 9b
were established by chiral HPLC [CHIRALPAK ID 0.46×25 cm (Daicel), EtOH-hexane (2:8) 1.0
mL/min, 30°C; 9a: 15.2 min, 9b: 20.5 min].
27
(2S,3S)-9a: mp 62–63 °C; [α]D –19.9 (c 1.00, CHCl3); IR (CHCl3)
ν
1737 (CO), 1549 and 1346
(NO2) cm–1; 1H NMR (300 MHz, CDCl3)
δ 1.40 (3H, s, CH3), 1.55 (1H, ddd, J=7.5, 9.5, 14 Hz) and
1.74 (1H, ddd, J=5.5, 7, 14 Hz) (CH2), 1.63 and 1.68 (each 3H, br s, gem-CH3), 2.13 (2H, br q, J= 7
Hz, CH2), 3.19 (1H, dd, J=3.5, 7.5 Hz, OCH2CH), 4.40 (1H, dd, J=7.5, 12.5 Hz) and 4.75 (1H, dd,
J=3.5, 12.5 Hz) (OCH2), 5.10 (1H, tsept, J=7, 1.5 Hz, =CH), 9.21 (2H, d, J=2 Hz, ArH), 9.25 (1H, t,
J=2 Hz, ArH); 13C NMR (75 MHz, CDCl3)
δ 16.96, 17.64, 23.55, 25.64, 38.15, 59.14, 60.82, 65.93,
122.59, 122.95, 129.53 (×2), 132.44, 133.38, 148.64 (×2), 162.41; HRMS (ESI) m/z calcd for
C17H20O7N2Na [M+Na]+ 387.1163, found 387.1161.
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(2R,3R)-9b: mp 62–64 °C; [α]D24 +17.8 (c 1.00, CHCl3); HRMS (ESI) m/z calcd for C17H20O7N2Na
[M+Na]+ 387.1163, found 387.1161.
3.1.2. Treatment of (2S,3S)-epoxy benzoate 9a with aqueous perchloic acid in DMF. An aqueous
solution of HClO4 (60%; 1.50 mL) was added slowly to a stirred solution of (2S,3S)-9a (3.64 g, 10
mmol) in DMF (20 mL) at 0 °C. After being stirred at rt for 15 h, the mixture was poured into
saturated aq. NaHCO3 and extracted with AcOEt. The extracts were washed with brine, dried and
evaporated to give a residue, which was purified by flash CC (MeOH-AcOEt-hexane, 1:35:65 to
2:40:60) to provide (2S,3R)-diol 10a (2.63 g, 66%) and (2S,3R)-formate 11a (875 mg, 21%) as pale
yellow viscous oils, respectively.
24
(2S,3R)-10a: [α]D –11.7 (c 0.98, CHCl3); IR (CHCl3)
ν
3564 (OH), 1735 (CO), 1549 and 1345
(NO2) cm–1; 1H NMR (300 MHz, CDCl3)
δ1.33 (3H, s, CH3), 1.48 and 1.75 (each 1H, m, CH2), 1.65
and 1.70 (each 3H, br s, gem-CH3), 2.14 (1H, s, OH), 2.14 (2H, m, CH2), 2.71 (1H, d, J=5 Hz, OH),
3.89 (1H, ddd, J=2.5, 5.5, 8.5 Hz, CHOH), 4.50 (1H, dd, J=8.5, 11.5 Hz) and 4.66 (1H, dd, J=2.5,
11.5 Hz) (OCH2), 5.15 (1H, tsept, J=7, 1 Hz, =CH), 9.18 (2H, d, J=2 Hz, ArH), 9.24 (1H, t, J=2 Hz,
ArH); 13C NMR (75 MHz, CDCl3)
δ17.72, 21.98, 23.41, 25.68, 37.13, 67.85, 73.83, 75.55, 122.53,
123.72, 129.52 (×2), 132.73, 133.60, 148.62 (×2), 162.90; HRMS (ESI) m/z calcd for C17H22O8N2Na
[M+Na]+ 405.1268, found 405.1265.
(2S,3R)-11a: [α]D25 –3.5 (c 1.09, CHCl3); IR (CHCl3)
ν 3598 and 3492 (OH), 1734 (CO), 1550 and
1
1345 (NO2) cm–1; H NMR (300 MHz, CDCl3)
δ
1.31 (3H, s, CH3), 1.57–1.72 (2H, m, CH2), 1.66
and 1.71 (each 3H, br s, gem-CH3), 1.92 (1H, br s, OH), 2.16 (2H, br q, J=7.5 Hz, CH2), 4.53 (1H, dd,
J=9, 12 Hz) and 4.96 (1H, dd, J=2, 12 Hz) (OCH2), 5.14 (1H, tsept, J=7, 1 Hz, =CH), 5.40 (1H, br dd,
J=2, 9 Hz, CHOCHO), 8.21 (1H, s, OCHO), 9.12 (2H, d, J=2.5 Hz, ArH), 9.22 (1H, t, J=2.5 Hz,
ArH); 13C NMR (75 MHz, CDCl3)
δ17.73, 21.91, 23.28, 25.69, 38.40, 65.31, 73.14, 75.05, 122.55,
123.23, 129.51 (×2), 133.16, 133.35, 148.66 (×2), 160.26, 162.39; HRMS (ESI) m/z calcd for
C18H22O9N2Na [M+Na]+ 433.1218, found 433.1219 .
3.1.3. Hydrolysis of epoxy benzoates 9a,b by use of p-TsOH in acetonitrile. A solution of (2S,3S)-9a
(3.30 g, 9.1 mmol) and p-TsOH•H2O (172 mg, 0.90 mmol) in MeCN (33 mL) and H2O (3.3 mL) was
heated at 50 °C for 1.5 h. After cooling, NaHCO3 (120 mg) was added to this mixture and MeCN was
evaporated off. The resulting mixture was diluted with AcOEt and washed with brine. The organic
layer was dried and evaporated to give a residue, which was purified by flash CC (AcOEt-hexane, 2:3)
to provide (2S,3R)-10a (2.58 g, 74%) as a pale yellow viscous oil. Its spectral data were identical with
those described above. (2R,3S)-10b was also prepared (74%) by the same procedure.
22
(2R,3S)-10b: [α]D +7.3 (c 1.02, CHCl3); HRMS (ESI) m/z calcd for C17H22O8N2Na [M+Na]+
405.1268, found 405.1265.
3.1.4. Treatment of (2S,3R)-dihydroxy benzoate 10a with methanol in the presence of triethylamine. To
a solution of (2S,3R)-10a (100 mg, 0.26 mmol) in MeOH (5 mL) was added Et3N (0.5 mL) and the
9

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mixture was stirred at rt for 30 min. The solvent was evaporated off and the resulting residue was
purified by flash CC (AcOEt-hexane, 1:1 to MeOH-AcOEt-hexane, 4:50:50) to provide (2S,3R)-triol
1
12a (46 mg, 92%) as a colorless oil. Its H and 13C NMR spectral data were identical with those
reported.11 [α]D23 –10.8 (c 0.92, CHCl3), lit11 [α]D25 –6.7 (c 1.05, CHCl3).
3.1.5. Treatment of (2S,3R)-formate 11a with methanol in the presence of triethylamine. In the same
manner as described above, (2S,3R)-11a (95 mg, 0.23 mmol) was treated with Et3N (0.1 mL) in
1
13
MeOH (5 mL) to provide (2S,3R)-12a (42 mg, 95%) as a colorless oil. Its H and C NMR spectral
data were identical with those reported.11 [α]D23 –10.8 (c 0.88, CHCl3).
3.1.6. (4S,5R)- and (4R,5S)-2,2,5-Trimethyl-5-(4-methylpent-3-en-1-yl)-1,3-dioxolan-4-yl)methyl
3,5-dinitrobenzoate 13a,b. A solution of (2S,3R)-10a (1.20 g, 3.14 mmol) and p-TsOH•H2O (12 mg,
0.06 mmol) in acetone (8 mL) and 2,2-dimethoxypropane (3 mL) was heated at 60 °C for 30 min.
After cooling, the mixture was poured into saturated aq. NaHCO3 and extracted with AcOEt. The
extracts were washed with brine, dried and evaporated to give a residue, which was purified by flash
CC (AcOEt-hexane, 1:4) to provide (4S,5R)-13a (1.26 g, 95%) as a pale yellow solid. (4R,5S)-13b
was also prepared (86%) by the same procedure.
(4S,5R)-13a: [α]D22 +4.0 (c 1.02, CHCl3); IR (CHCl3)
ν
1737 (CO), 1549 and 1345 (NO2) cm–1; 1H
NMR (300 MHz, CDCl3) 1.27 and 1.67 (each 1H, m, CH2), 1.39, 1.43 and 1.49 (each 3H, s,
δ
CH3×3), 1.64 and 1.70 (each 3H, br s, gem-CH3), 2.01–2.31 (2H, m, CH2), 4.17 (1H, t, J=5.5 Hz,
OCH2CH), 4.55 (2H, d, J=5.5 Hz, OCH2), 5.13 (1H, tsept, J=7, 1 Hz, =CH), 9.19 (2H, d, J=2 Hz,
ArH), 9.25 (1H, t, J=2 Hz, ArH); 13C NMR (75 MHz, CDCl3)
δ 17.59, 21.71, 23.04, 25.66, 27.08,
28.28, 35.18, 65.13, 81.39, 81.61, 108.57, 122.59, 123.80, 129.57 (×2), 132.10, 133.39, 148.63 (×2),
162.43; HRMS (ESI) m/z calcd for C20H27O8N2 [MH]+ 423.1762, found 423.1763.
(4R,5S)-13b: [α]D26 –1.1 (c 1.03, CHCl3); HRMS (ESI) m/z calcd for C20H27O8N2 [MH]+ 423.1762,
found 423.1762.
3.1.7. (4R,5R)- and (4S,5S)-2,2,5-Trimethyl-5-(4-methylpent-3-en-1-yl)-1,3-dioxolane-4-carbaldehyde
14a,b. To a solution of (4S,5R)-13a (3.40 g, 8.46 mmol) in EtOH (40 mL) was added dropwise 10%
aq. KOH (9 mL) and the mixture was stirred for a further 10 min. After EtOH was evaporated off, the
resulting mixture was poured into water and extracted with AcOEt. The extracts were washed with
brine, dried and evaporated to give a crude alcohol, which without purification was dissolved in
CH2Cl2 (40 mL). To this solution was added Dess-Martin periodinane (4.17 g, 9.88 mol) in some
portions at rt and the mixture was stirred for a further 20 min. After CH2Cl2 was evaporated off, the
resulting mixture was diluted with Et2O and filtered through a pad of Celite. The filtrate was
evaporated to afford a residue, which was purified by flash CC (AcOEt-hexane, 1:4 to 1:3) to provide
(4R,5R)-14a (1.42 g, 78% from 13a) as a colorless oil. (4S,5S)-14b was also prepared (76% from 13b)
by the same procedure.
20
1
(4R,5R)-14a: [α]D +100.6 (c 1.02, CHCl3); IR (CHCl3)
ν
1734 (CO) cm–1; H NMR (500 MHz,
10

ACCEPTED MANUSCRIPT
CDCl3)
δ
1.33 (1H, ddd, J=5, 11.5, 13.5 Hz) and 1.59 (1H, m) (CH2), 1.41 and 1.55 (each 3H, s,
gem-CH3), 1.43 (3H, s, CH3), 1.59 and 1.67 (each 3H, br s, gem-CH3), 1.99–2.15 (2H, m, CH2), 4.11
(1H, d, J=2 Hz, CHCHO), 5.06 (1H, tsept, J=7.5, 1.5 Hz, =CH), 9.75 (1H, d, J=2 Hz, CHO); 13C
NMR (125 MHz, CDCl3)
δ 17.56, 22.03, 24.59, 25.62, 27.05, 28.04, 36.59, 83.85, 87.93, 109.96,
123.59, 132.04, 200.35; HRMS (ESI) m/z calcd for C13H22O3Na [M+Na]+ 249.1461, found 249.1462.
24
(4S,5S)-14b: [α]D –103.0 (c 0.98, CHCl3); HRMS (ESI) m/z calcd for C13H22O3Na [M+Na]+
249.1461, found 249.1460.
3.1.8. Preparation of (5R,6S)- and (5S,6R)-C15-Dienoate 16a,b. To a stirred solution of phosphonate
15 (3.31 g, 12.5 mmol) and N,N’-dimethylpropyleneurea (DMPU) (3.0 mL, 25 mmol) in dry THF (25
mL) was added dropwise n-BuLi (1.63 M in hexane; 7.9 mL, 12.9 mmol) at –20 °C and the mixture
was stirred for a further 20 min. A solution of (4R,5R)-14a (1.42 g, 6.28 mmol) in dry THF (7 mL)
was then added dropwise to this solution at –20 °C. After being stirred at –20 °C for a further 20 min,
the mixture was poured into saturated aq. NH4Cl and extracted with AcOEt. The extracts were washed
with brine, dried and evaporated to give a residue, which was purified by flash CC (AcOEt-hexane,
15:85) to afford an isomeric mixture (E/Z ~1/1) (5R,6S)-16a (1.96 g, 93%) as a colorless oil.
(5S,6R)-16b was also prepared (89%) by the same procedure.
1
(5R,6S)-16a: IR (CHCl3)
ν
1704 (CO), 1642 and 1612 (C=C) cm–1; H NMR (300 MHz, CDCl3)
δ
1.20 and 1.60 (each 1H, m, 4-H2), 1.29 (3H, t, J=7 Hz, CH2CH3), 1.39, 1.40 and 1.48 (each 3H, s,
CH3×3), 1.60 and 1.68 (each 3H, br s, gem-CH3), 2.02 and 2.29 (each 3/2H, d, J=1 Hz, 9-CH3), 1.95–
2.24 (2H, m, 3-H2), 4.17 and 4.18 (each 1H, q, J=7 Hz, OCH2CH3), 4.32 and 4.36 (each 1/2H, br d,
J=7.5 Hz, 6-H), 5.09 (1H, br t, J=7 Hz, 2-H), 5.73 and 5.82 (each 1/2H, br s, 10-H), 6.03 and 6.05
(each 1/2H, dd, J=7.5, 16 Hz, 7-H), 6.39 and 7.84 (each 1/2H, d, J=16 Hz, 8-H); HRMS (ESI) m/z
calcd for C20H32O4Na [M+Na]+ 359.2193, found 359.2191.
(5S,6R)-16b: HRMS (ESI) m/z calcd for C20H32O4Na [M+Na]+ 359.2193, found 359.2191.
3.1.9. Preparation of (5R,6S)- and (5S,6R)-C15-Dienal 17a,b. A solution of (5R,6S)-16a (1.10 g, 3.27
mmol) in dry Et2O (7 mL) was added dropwise to a stirred suspension of LiAlH4 (124 mg, 3.26 mmol)
in dry Et2O (20 mL) at 0 °C. After being stirred at 0 °C for 10 min, the excess of LiAlH4 was
decomposed by dropwise addition of water. The mixture was filtered through a pad of Celite and the
filtrate was dried and evaporated. This was dissolved in Et2O (20 mL) and hexane (20 mL) and stirred
with MnO2 (7 g) at rt for 1 h. After MnO2 was filtered off, the filtrate was concentrated to give the
crude aldehyde. This was dissolved in THF (20 mL) and H2SO4 (3M in water; 4 mL), and the mixture
was heated at 80 °C for 2h. After cooling, the mixture was neutralized with saturated aq. NaHCO3 and
extracted with AcOEt. The extracts were washed with brine, dried and evaporated to give a residue,
which was purified by flash CC (acetone-hexane, 1:2) to provide an isomeric mixture (E/Z ~2/1) of
(5R,6S)-17a (498 mg, 60% from 16a) as an yellow-brown viscous oil. (5S,6R)-17b was also prepared
(60% from 16b) by the same procedure.
11

ACCEPTED MANUSCRIPT
(5R,6S)-17a: IR (CHCl3)
NMR (300 MHz, CDCl3)
ν
3611, 3566 and 3454 (OH), 1665 (CO), 1634 and 1599 (C=C) cm–1; 1H
1.25 (2H) and 1.27 (1H) (each s, 5-CH3), 1.37 and 1.62 (each 1H, m,
δ
4-H2), 1.63 and 1.69 (each 3H, br s, gem-CH3), 2.05 (2/3H) and 2.18 (1/3H) (each s, OH), 2.10 (2H, m,
3-H2), 2.10 (1H) and 2.28 (2H) (each d, J=1 Hz, 9-CH3), 4.12 (1H, m, 6-H), 5.12 (1H, br t, J=7.5 Hz,
2-H), 5.91 (1/3H) and 5.98 (2/3H) (each br d, J=8 Hz, 10-H), 6.23 (1/3H) and 6.31 (2/3H) (each dd,
J=8, 15.5 Hz, 7-H), 6.48 (2/3H) and 7.36 (1/3H) (each d, J=15.5 Hz, 8-H), 10.13 (2/3H) and 10.20
(1/3H) (each d, J=8 Hz, CHO); HRMS (ESI) m/z calcd for C15H24O3Na [M+Na]+ 275.1618, found
275.1619.
(5S,6R)-17b: HRMS (ESI) m/z calcd for C15H24O3Na [M+Na]+ 275.1618, found 275.1617.
3.1.10. Preparation of (5R,6S)- and (5S,6R)-C25-Apocarotenal 18a,b. Trimethyl orthoformate (4 mL)
and an acidic solution (1.0 mL) prepared from p-TsOH•H2O (500 mg) and H3PO4 (700 mg) in MeOH
(35 mL) were added to a solution of phosphonium salt 78 (4.70 g, 10. 5 mmol) in MeOH (20 mL) and
the mixture was stirred at rt for 2 h. The resulting solution of phosphonium salt 7’ was neutralized
with NaOMe until just before a red color of an ylide appeared and a solution of (5R,6S)-17a (E/Z ~2/1:
630 mg, 2.50 mmol) in CH2Cl2 (5 mL) and then NaOMe (650 mg, 12.0 mmol) were added to it. After
being stirred at rt for 30 min, the mixture was quenched by saturated aq. NH4Cl and extracted with
AcOEt. The extracts were shaken with 3% aq. HCl twice. The organic layer was washed with brine,
dried and evaporated to give a residue, which was purified by flash CC (acetone-CH2Cl2, 12:88 to
MeOH-acetone-CH2Cl2, 2:15:85). The resulting isomeric mixture of condensed products was
dissolved in THF (2 mL) and MeCN (60 mL), and a solution (20 mL) prepared from PdCl2(MeCN)2
(26 mg), Et3N (14 µL) and water (2.4 mL) in MeCN (17.6 mL) was added to it. After being stirred at
rt for 2 h, the solvent was evaporated off to give a residue, which was purified by flash CC
(MeOH-acetone-CH2Cl2, 2:15:85) to afford an isomeric mixture (all-E-isomer: 54% from HPLC
analysis) of 18a (751 mg, 78% from 17a). Preparative HPLC purification [Cosmosil 5SL-II (SiO2)
2×25 cm; acetone-CH2Cl2-hexane, 1:7:3] yielded all-E-isomer of (5R,6S)-18a (314 mg, 33% from
17a) and other isomeric mixture of 18a (395 mg, 41 % from 17a) as a red-orange foam, respectively.
(5S,6R)-18b was also prepared (all-E-isomer: 38% from 17b after preparative HPLC) by the same
procedure.
(5R,6S)-18a: UV-VIS (EtOH) λ 420 nm; IR (CHCl3)
ν 3601 and 3562 (OH), 1660 (CO), 1600 and
1
1551 (C=C) cm–1; H NMR (500 MHz, CDCl3)
δ 1.23 (3H, s, 5-CH3), 1.38 (1H, ddd, J=5.5, 11, 14
Hz) and 1.63 (1H, m) (4-H2), 1.62 and 1.69 (each 3H, br s, gem-CH3), 1.89 (3H, br s, 13’-CH3), 1.95
(3H, br s, 9-CH3), 2.04 (3H, br s, 13-CH3), 2.04–2.18 (2H, m, 3-H2), 2.11 (1H, br s, 5-OH), 2.20 (1H,
br d, J=4 Hz, 6-OH), 4.03 (1H, dd, J=4, 7.5 Hz, 6-H), 5.12 (1H, br t, J=6.5 Hz, 2-H), 5.81 (1H, dd,
J=7.5, 16 Hz, 7-H), 6.21 (1H, br d, J=11 Hz, 10-H), 6.32 (1H, br d, J=12 Hz, 14-H), 6.39 (1H, d, J=16
Hz, 8-H), 6.39 (1H, d, J=15.5 Hz, 12-H), 6.70 (1H, dd, J=12, 14.5 Hz, 15’-H), 6.75 (1H, dd, J=11,
15.5 Hz, 11-H), 6.96 (1H, br d, J=12 Hz, 14’-H), 7.03 (1H, dd, J=12, 14.5 Hz, 15-H), 9.46 (1H, s,
CHO); 13C NMR (125 MHz, CDCl3)
δ 9.60 (13’-CH3), 13.02 (13-CH3), 13.09 (9-CH3), 17.66
(1-CH3), 22.02 (C3), 23.40 (5-CH3), 25.70 (1-CH3), 36.55 (C4), 74.84 (C5), 79.60 (C6), 124.40 (C2),
12

ACCEPTED MANUSCRIPT
127.09 (C11), 127.41 (C7), 127.69 (C15’), 131.44 (C14), 132.00 (C1), 132.22 (C10), 136.19 (C9),
137.05 (C13’), 137.48 (C15), 137.55 (C8 and C12), 141.39 (C13), 148.82 (C14’), 194.52 (CHO);
HRMS (ESI) m/z calcd for C25H36O3Na [M+Na]+ 407.2557, found 407.2559.
(5S,6R)-18b: HRMS (ESI) m/z calcd for C25H36O3Na [M+Na]+ 407.2557, found 407.2548.
3.1.11. Preparation of (5R,6S)- and (5S,6R)-erythro-Lycopene-5,6-diol 1a,b. To a solution of
phosphonium salt 56 (2.30 g, 4.0 mmol) in CH2Cl2 (13 ml) was added NaOMe until just before a red
color of an ylide appeared and a solution and a solution of (5R,6S)-18a (250 mg, 0.65 mmol) in
CH2Cl2 (7 mL) and then NaOMe (1M in MeOH; 5.0 mL, 5.0 mmol) were added to it. After being
stirred at rt for 40 min, the mixture was quenched by saturated aq. NH4Cl and extracted with AcOEt.
The extracts were washed with brine, dried and evaporated to give a residue, which was purified by
flash CC (AcOEt-hexane, 35:65 to MeOH-AcOEt-hexane, 3:40:60). The resulting isomeric mixture of
condensed products was dissolved in THF (5 mL) and MeCN (40 mL), and a solution (8 mL) prepared
from PdCl2(MeCN)2 (26 mg), Et3N (14 µL) and water (2.4 mL) in MeCN (17.6 mL) was added to it.
After being stirred at rt for 2 h, the solvent was evaporated off to give a residue, which was purified by
flash CC (MeOH-AcOEt-hexane, 3:40:60) to afford an isomeric mixture (all-E-isomer: 51% from
HPLC analysis) of 1a (259 mg, 70% from 18a) as a red solid. Crystallization from
CH2Cl2-Et2O-hexane afforded all-E-isomer of (5R,6S)-1a (78 mg, 21% from 18a) as a red powder.
(5S,6R)-1b was also prepared (all-E-isomer: 10% from 18b after crystallization) by the same
procedure. 1H NMR spectra of 1a,b was identical with a natural specimen.5
(5R,6S)-1a: UV-VIS (EtOH) λ 280, 431, 456, 488 nm; CD (6.04×10−5 mol/L, EtOH) λ(ꢀε) 205
(−0.03), 250 (–1.09), 270 (–2.48), 272 (–2.20), 279 (–3.11), 322 (0), 331 (+0.50), 344 (+0.83), 352 (0);
1H NMR (500 MHz, CDCl3)
δ 1.22 (3H, s, 5-CH3), 1.38 (1H, ddd, J=5.5, 11, 14 Hz) and 1.63 (1H,
m) (4-H2), 1.62 and 1.69 (each 6H, br s, gem-CH3×2), 1.82 (3H, br s, 5’-CH3), 1.92 (3H, br s, 9-CH3),
1.97 (9H, br s, 9’-CH3, 13-CH3 and 13’-CH3) 2.03–2.16 (6H, m, 3-H2, 3’-H2 and 4’-H2), 2.06 (1H, s,
5-OH), 2.07 (1H, br d, J=3.5 Hz, 6-OH), 4.02 (1H, br dd, J=3.5, 7 Hz, 6-H), 5.11 and 5.12 (each 1H,
m, 2-H and 2’-H), 5.74 (1H, dd, J=7, 16 Hz, 7-H), 5.95 (1H, br d, J=11.5 Hz, 6’-H), 6.18 (1H, br d,
J=11.5 Hz, 10’-H), 6.20 (1H, br d, J=11.5 Hz, 10-H), 6.25 (1H, d, J=15.5 Hz, 8’-H), 6.25 (1H,
overlapped, 14’-H), 6.27 (1H,overlapped, 14-H), 6.35 (1H, d, J=15 Hz, 12’-H), 6.38 (2H, d, J=15.5
Hz, 8-H and 12-H), 6.50 (1H, dd, J=11.5, 15.5 Hz, 7’-H), 6.59 (1H, dd, J=11.5, 15 Hz, 11-H), 6.60–
6.68 (2H, m, 15-H and 15’-H), 6.65 (1H, dd, J=11.5, 15 Hz, 11’-H); 13C NMR (125 MHz, CDCl3)
δ
12.80 (13-CH3 and 13’-CH3), 12.92 (9’-CH3), 13.02 (9-CH3), 16.98 (5’-CH3), 17.68 and 17.71
(1-CH3 and 1’-CH3), 22.07 (C3), 23.44 (5-CH3), 25.71 and 25.72 (1-CH3 and 1’-CH3), 26.71 (C3’),
36.63 (C4), 40.25 (C4’), 74.85 (C5), 79.79 (C6), 123.96 (C2’), 124.49 and 124.51 (C2 and C11),
124.90 (C7’), 125.34 (C11’), 125.73 (C6’), 126.24 (C7), 129.88 and 130.51 (C15 and C15’), 131.50
(C10’), 131.77 and 131.95 (C1 and C1’), 132.50 (C14’), 132.87 (C10), 133.21 (C14), 134.28 (C9),
135.39 (C8’), 136.19 and 136.32 (C13 and C9’), 136.86 (C13’), 137.29 (C12’), 137.95 (C8), 138.57
(C12), 139.59 (C5’); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329, found 593.4326.
(5S,6R)-1b: CD (5.91×10−5 mol/L, EtOH) λ(ꢀε) 226 (0), 250 (–0.65), 270 (+0.96), 272 (+0.86), 279
13

ACCEPTED MANUSCRIPT
(+2.20), 289 (0), 330 (–0.90), 344 (–1.19), 382 (–0.19); HRMS (ESI) m/z calcd for C40H58O2Na
[M+Na]+ 593.4326, found 593.4319.
3.1.12. Preparation of (5’R,6’S)- and (5’S,’6R)-erythro-γ-Carotene-5’,6’-diol 3a,b. To a solution of
phosphonium salt 67 (820 dg, 2.13 mmol) in CH2Cl2 (5 ml) was added NaOMe until just before a red
color of an ylide appeared and a solution and a solution of (5R,6S)-18a (165 mg, 0.43 mmol) in
CH2Cl2 (7 mL) and then NaOMe (1M in MeOH; 2.8 mL, 2.8 mmol) were added to it. After being
stirred at rt for 20 min, the mixture was quenched by saturated aq. NH4Cl and extracted with AcOEt.
The extracts were washed with brine, dried and evaporated to give a residue, which was purified by
flash CC (AcOEt-hexane, 35:65 to MeOH-AcOEt-hexane, 2:40:60) and preparative HPLC [Cosmosil
5SL-II (SiO2) 2×25 cm; MeOH-Et2O-hexane, 0.3:35:60] to provide all-E-isomer of (5’R,6’S)-2a (314
mg, 31% from 18a) as a red power and other isomeric mixture of 3a (145 mg, 59 % from 18a). This
isomeric mixture was dissolved in THF (2 mL) and MeCN (20 mL), and a solution (6 mL) prepared
from PdCl2(MeCN)2 (26 mg), Et3N (14 µL) and water (2.4 mL) in MeCN (17.6 mL) was added to it.
After being stirred at rt for 2.5 h, the solvent was evaporated off to give a residue, which was purified
by flash CC and preparative HPLC described above to afford all-E-isomer of 3a (42 mg, 17% from
18a: combined yield, 48%). (5’S,6’R)-3b was also prepared (all-E-isomer: combined yield, 50% from
18b after preparative HPLC) by the same procedure. 1H NMR spectra of 3a,b was identical with a
natural specimen.5
(5’R,6’S)-3a: UV-VIS (EtOH) λ 267, 424, 446, 474 nm; CD (8.16×10−5 mol/L, EtOH) λ(ꢀε) 210 (–
1
1.62), 236 (+0.40), 269 (–2.54), 305 (0), 329 (+0.84), 355 (+0.03); H NMR (500 MHz, CDCl3)
δ
1.03 (6H, s, 1-gem-Me), 1.22 (3H, s, 5’-CH3), 1.38 (1H, ddd, J=5.5, 11, 14 Hz, 4’-H), 1.47 (2H, m,
2-H2), 1.59–1.65 (3H, m, 3-H2 and 4’-H), 1.62 and 1.68 (each 3H, br s, 1’-gem-Me), 1.72 (3H, br s,
5-CH3), 1.92 (3H, br s, 9’-CH3), 1.96 (3H) and 1.97 (6H) (each br s, 9-CH3, 13-CH3 and 13’-CH3) 2.02
(2H, br t, J=6.5 Hz, 4-H2), 2.07 (1H, s, 5’-OH), 2.08 (1H, d, J=4 Hz, 6’-OH), 2.05–2.16 (2H, m,
3’-H2), 4.02 (1H, dd, J=4, 7.5 Hz, 6’-H), 5.12 (1H, tsept, J=7, 1.5 Hz, 2’-H), 5.74 (1H, dd, J=7.5, 15
Hz, 7’-H), 6.13 (1H, d, J=15.5 Hz, 8-H), 6.15 (1H, br d, J=11 Hz, 10-H), 6.18 (1H, br d, J=15.5 Hz,
7-H), 6.20 (1H, br d, J=11 Hz, 10’-H), 6.25 (1H, br d, J=11 Hz, 14-H), 6.27 (1H, br d, J=11 Hz,
14’-H), 6.35 (1H, d, J=15 Hz, 12-H), 6.38 (2H, d, J=15 Hz, 8’-H and 12’-H), 6.59 (1H, dd, J=11, 15
Hz, 11’-H), 6.60–6.68 (2H, m, 15-H and 15’-H), 6.66 (1H, dd, J=11, 15 Hz, 11-H); 13C NMR (125
MHz, CDCl3)
δ 12.78, 12.80 and 12.85 (9-CH3, 13-CH3 and 13’-CH3), 13.02 (9’-CH3), 17.68 and
25.72 (1’-gem-CH3), 19.29 (C3), 21.77 (5-CH3), 22.06 (C3’), 23.44 (5’-CH3), 28.99 (1-gem-CH3),
33.14 (C4), 34.29 (C1), 36.63 (C4’), 39.67 (C2), 74.84 (C5’), 79.79 (C6’), 124.46 and 124.51 (C2’ and
C11’), 125.25 (C11), 126.23 (C7’), 126.78 (C7), 129.42 (C5), 129.80 and 130.49 (C15 and C15’),
130.81 (C10), 131.99 (C1’), 132.30 (C14), 132.86 (C10’), 133.20 (C14’), 134.25 (C9’), 136.12 and
136.19 (C9 and C13’), 136.84 (C13), 137.17 (C12), 137.75 (C8), 137.93 (C6), 137.95 (C8’), 138.57
(C12’); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329, found 593.4325.
(5’S,6’R)-3b: CD (8.09×10−5 mol/L, EtOH) λ(ꢀε) 213 (+1.38), 240 (–0.79), 269 (+1.80), 290 (0),
327 (–1.14), 354 (–0.34); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329, found 593.4332.
14

ACCEPTED MANUSCRIPT
3.1.13.
(2S,3R)-
and
(2R,3S)-3-Methyl-3-(4-methylpent-3-en-1-yl)oxiran-2-yl)methyl
3,5-dinitorobenzoate 20a,b. (2S,3R)-Epoxynerol 19a (69% ee) and (2R,3S)-epoxynerol 19b (70% ee)
were prepared according to the known Sharpless asymmetric epoxidation method of nerol.10,19 In the
same procedure described for the preparation of compound 9a, (2S,3R)-19a (69% ee; 4.90 g) and
(2R,3S)-19b (70% ee; 4.64 g) were converted into (2S,3R)-20a (10.39 g; 99%) and (2R,3S)-20b (9.88
g; 99%) as a pale yellow solid, respectively. Recrystallization of 20a (10.39 g) from MeOH (130 mL)
afforded 5.87 g (56% yield) of 20a (56% ee) as a pale yellow needle. Evaporation of the filtrate gave
4.60 g (44% yield) of crude 20a (92% ee). The enantiomeric purities of 20a and 20b were established
by chiral HPLC [CHIRALPAK ID 0.46×25 cm (Daicel), EtOH-hexane (2:8) 0.7 mL/min, 30°C; 20a:
19.9 min, 20b: 21.8 min]. Thus without recrystallization, 20a and 20b were used in the next step.
1
(2S,3R)-20a: IR (CHCl3)
ν
1735 (CO), 1549 and 1346 (NO2) cm–1; H NMR (300 MHz, CDCl3)
δ
1.41 (3H, s, CH3), 1.52–1.79 (2H, m, CH2), 1.65 and 1.71 (each 3H, br s, gem-CH3), 2.18 (2H, m,
CH2), 3.19 (1H, dd, J=3.5, 7.5 Hz, OCH2CH), 4.36 (1H, dd, J=7.5, 12 Hz) and 4.76 (1H, dd, J=3.5, 12
Hz) (OCH2), 5.13 (1H, br t, J=7 Hz, =CH), 9.20 (2H, d, J=2 Hz, ArH), 9.25 (1H, t, J=2 Hz, ArH);13C
NMR (75 MHz, CDCl3)
δ 17.64, 21.95, 24.11, 25.66, 33.28, 60.38, 61.12, 65.77, 122.59, 122.83,
129.53 (×2), 132.81, 133.39, 148.64 (×2), 162.43; HRMS (ESI) m/z calcd for C17H20O7N2Na [M+Na]+
387.1163, found 387.1162.
(2R,3S)-20b: HRMS (ESI) m/z calcd for C17H20O7N2Na [M+Na]+ 387.1163, found 387.1162.
3.1.14. (2S,3S)- and (2R,3R)-2,3-Dihydroxy-3,7-dimethyloct-6-en-1-yl 3,5-dinitrobenzoate 21a,b. In
the same procedure described for hydrolysis of compound 9a by use of p-TsOH in acetonitrile,
(2S,3R)-20a (69% ee; 7.00 g) and (2R,3S)-20b (70% ee; 7.00 g) were converted into (2S,3S)-21a (6.10
g; 83%) and (2R,3R)-21b (6.47 g; 88%) as a pale yellow solid, respectively. Twice-recrystallization of
21a,b from MeOH-H2O (4:1) afforded 2.08 g of 21a (99% ee) and 2.31 g of 21b (99% ee) as a pale
yellow needle, respectively. The enantiomeric purities of 21a and 21b were established by chiral
HPLC [CHIRALPAK IA 0.46×25 cm (Daicel), 2-PrOH-hexane (3.7) 1.0 mL/min, 30°C; 21a: 16.3 min,
21b: 19.1 min].
20
(2S,3S)-21a: mp 117–118 °C; [α]D –18.2 (c 1.00, CHCl3); IR (CHCl3)
ν
3571 and 3453 (OH),
1735 (CO), 1549 and 1345 (NO2) cm–1; 1H NMR (300 MHz, CDCl3)
δ 1.27 (3H, s, CH3), 1.57–1.74
(2H, m, CH2), 1.65 and 1.70 (each 3H, br s, gem-CH3), 2.14 (2H, br q, J=7.5 Hz, CH2), 2.18 (1H, s,
OH), 2.77 (1H, d, J=5 Hz, OH), 3.92 (1H, ddd, J=2.5, 5, 8.5 Hz, CHOH), 4.48 (1H, dd, J=8.5, 11.5
Hz) and 4.76 (1H, dd, J=2.5, 11.5 Hz)(OCH2), 5.14 (1H, br t, J=7 Hz, =CH), 9.19 (2H, d, J=2 Hz,
ArH), 9.24 (1H, t, J=2 Hz, ArH); 13C NMR (75 MHz, CDCl3)
δ 17.71, 21.89, 22.10, 25.68, 38.87,
67.78, 73.76, 74.64, 122.53, 123.62, 129.52 (×2), 132.69, 133.58, 148.63 (×2), 162.84; HRMS (ESI)
m/z calcd for C17H22O8N2Na [M+Na]+ 405.1268, found 405.1267.
21
(2R,3R)-21b: mp 115–117 °C; [α]D +16.2 (c 0.93, CHCl3); HRMS (ESI) m/z calcd for
C17H22O8N2Na [M+Na]+ 405.1268, found 405.1267.
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ACCEPTED MANUSCRIPT
3.1.15. Preparation of (5S,6S)- and (5R,6R)-threo-lycopene-5,6-diol 2a,b, and (5’S,6’S)- and
(5’R,’6R)-erythro-γ-carotene-5’,6’-diol 4a,b. In the same procedure described for the preparation of
(5R,6S)-erythro-lycopene-5,6-diol 1a and (5’R,6’S)-erythro-γ-carotene-5’,6’-diol 3a, threo-
lycopene-5,6-diols 2a,b and threo-γ-carotene-5’,6’-diols 4a,b were prepared from (2S,3S)-21a and
(2R,3R)-21b via C10-aldehydes 22a,b, C15-aldehydes 23a,b and C25-apocarotenals 24a,b.
(4R,5S)-22a (74% form 21a): [α]D27 +55.4 (c 1.24, CHCl3); IR (CHCl3)
ν
1735 (CO) cm–1; 1H NMR
(500 MHz, CDCl3) 1.18 (3H, s, CH3), 1.39 and 1.55 (each 3H, s, gem-CH3), 1.61 and 1.68 (each 3H,
δ
br s, gem-CH3), 1.66–1.71 (2H, m, CH2), 2.04–2.17 (2H, m, CH2), 4.14 (1H, d, J=2 Hz, CHCHO),
5.10 (1H, tsept, J=1.5, 7.5 Hz, =CH), 9.71 (1H, d, J=2 Hz, CHO); 13C NMR (75 MHz, CDCl3)
δ
17.64, 22.31, 22.33, 25.63, 27.15, 28.47, 40.36, 83.67, 85.82, 110.05, 123.52, 132.18, 200.73;
HRMS (ESI) m/z calcd for C13H22O3Na [M+Na]+ 249.1461, found 249.1461.
(4S,5R)-22b (73% form 21b): [α]D23 –55.2 (c 1.00, CHCl3); HRMS (ESI) m/z calcd for C13H22O3Na
[M+Na]+ 249.1461, found 249.1461.
(5S,6S)-23a (49% from 22a): IR (CHCl3)
ν 3606 and 3463 (OH), 1664 (CO), 1634 and 1600 (C=C)
1
cm–1; H NMR (300 MHz, CDCl3)
δ 1.14 (2H) and 1.15 (1H) (each s, 5-CH3), 1.54–1.62 (2H, m,
4-H2), 1.64 and 1.70 (each 3H, br s, gem-CH3), 2.05 (1/3H) and 2.07 (2/3H) (each s, OH), 2.09 (1H)
and 2.28 (2H) (each d, J=1 Hz, 9-CH3), 2.13 (2H, m, 3-H2), 2.43 (2/3H) and 2.48 (1/3H) (each d,
J=3.5 Hz, OH), 4.12 (1H, m, 6-H), 5.14 (1H, br t, J=7.5 Hz, 2-H), 5.91 (1/3H) and 5.98 (2/3H) (each
br d, J=8 Hz, 10-H), 6.19 (1/3H) and 6.27 (2/3H) (each dd, J=6.5, 15.5 Hz, 7-H), 6.47 (2/3H) and 7.36
(1/3H) (each d, J=15.5 Hz, 8-H), 10.13 (2/3H) and 10.20 (1/3H) (each d, J=8 Hz, CHO); HRMS (ESI)
m/z calcd for C15H24O3Na [M+Na]+ 275.1618, found 275.1617.
(5R,6R)-23b (53% from 22b): HRMS (ESI) m/z calcd for C15H24O3Na [M+Na]+ 275.1618, found
275.1617.
(5S,6S)-24a (all-E-isomer: 36% from 23a after preparative HPLC): UV-VIS (EtOH) λ 418 nm; IR
1
(CHCl3)
ν
3604, 3567 and 3478 (OH), 1661 (CO), 1611, 1600 and 1551 (C=C) cm–1; H NMR (500
MHz, CDCl3)
δ
1.14 (3H, s, 5-CH3), 1.54 (2H, t, J=8 Hz, 4-H2), 1.63 and 1.69 (each 3H, br s,
gem-CH3), 1.89 (3H, br s, 13’-CH3), 1.95 (3H, br s, 9-CH3), 2.04 (3H, br s, 13-CH3), 2.12 (2H, br q,
J=8 Hz, 3-H2), 2.12 (1H, br s, 5-OH), 2.26 (1H, br d, J=2.5 Hz, 6-OH), 4.05 (1H, dd, J=2.5, 7.5 Hz,
6-H), 5.13 (1H, tsept, J=7.5, 1.5 Hz, 2-H), 5.77 (1H, dd, J=7.5, 15.5 Hz, 7-H), 6.21 (1H, br d, J=11 Hz,
10-H), 6.32 (1H, br d, J=12 Hz, 14-H), 6.39 (1H, d, J=15.5 Hz, 8-H), 6.40 (1H, d, J=15 Hz, 12-H),
6.70 (1H, dd, J=11.5, 14.5 Hz, 15’-H), 6.74 (1H, dd, J=11, 15 Hz, 11-H), 6.96 (1H, br d, J=11.5 Hz,
14’-H), 7.02 (1H, dd, J=11.5, 14.5 Hz, 15-H), 9.45 (1H, s, CHO); 13C NMR (125 MHz, CDCl3)
δ
9.62 (13’-CH3), 13.05 and 13.10 (9-CH3 and 13-CH3), 17.70 (1-CH3), 21.43 (5-CH3), 22.12 (C3),
25.72 (1-CH3), 38.81 (C4), 75.03 (C5), 78.71 (C6), 124.30 (C2), 127.12 (C11), 127.47 (C7), 127.74
(C15’), 131.48 (C14), 132.07 (C1), 132.29 (C10), 136.22 (C9), 137.11 (C13’), 137.57 and 137.61
(C12 and C15), 137.73 (C8), 141.41 (C13), 148.82 (C14’), 194.53 (CHO); HRMS (ESI) m/z calcd for
C25H36O3Na [M+Na]+ 407.2557, found 407.2557.
(5R,6R)-24b (all-E-isomer: 20% from 23b after preparative HPLC): HRMS (ESI) m/z calcd for
C25H36O3Na [M+Na]+ 407.2557, found 407.2556.
16

ACCEPTED MANUSCRIPT
(5S,6S)-2a (78% from 24a for isomeric mixture, all-E-isomer: 24% from 24a after crystallization):
UV-VIS (EtOH) λ 280, 430, 455, 486 nm; CD (5.80×10−5 mol/L, EtOH) λ(ꢀε) 240 (−0.42), 271 (–
1
2.80), 280 (–4.72), 318 (0), 331 (+0.70), 344 (+1.31), 358 (0); H NMR (500 MHz, CDCl3)
δ 1.14
(3H, s, 5-CH3), 1.53 (2H, t, J=8 Hz, 4-H2), 1.62 (3H), 1.63 (3H) and 1.69 (6H) (each br s, gem-CH3×2),
1.82 (3H, br s, 5’-CH3), 1.92 (3H, br s, 9-CH3), 1.97 (9H, br s, 13-CH3, 9’-CH3 and 13’-CH3) 2.08 (1H,
s, 5-OH), 2.08–2.14 (6H, m, 3-H2, 3’-H2 and 4’-H2), 2.12 (1H, br d, J=3 Hz, 6-OH), 4.04 (1H, dd, J=3,
8 Hz, 6-H), 5.11 and 5.13 (each 1H, m, 2-H and 2’-H), 5.70 (1H, dd, J=8, 15.5 Hz, 7-H), 5.95 (1H, br
d, J=11.5 Hz, 6’-H), 6.18 (1H, br d, J=11.5 Hz, 10’-H), 6.20 (1H, br d, J=11.5 Hz, 10-H), 6.25 (1H, d,
J=15 Hz, 8’-H), 6.25 (1H, overlapped, 14’-H), 6.27 (1H, overlapped, 14-H), 6.35 (1H, d, J=15 Hz,
12’-H), 6.38 (2H, d, J=15.5 Hz, 8-H and 12-H), 6.50 (1H, dd, J=11.5, 15 Hz, 7’-H), 6.59 (1H, dd,
13
J=11.5, 15 Hz, 11-H), 6.60–6.68 (2H, m, 15-H and 15’-H), 6.65 (1H, dd, J=11.5, 15 Hz, 11’-H); C
NMR (125 MHz, CDCl3)
δ 12.80, 12.82, 12.92 and 12.99 (9-CH3, 9’-CH3, 13-CH3 and 13’-CH3),
16.98 (5’-CH3), 17.69 and 17.14 (1-CH3 and 1’-CH3), 21.43 (5-CH3), 22.12 (C3), 25.71 (1-CH3 and
1’-CH3), 26.70 (C3’), 38.80 (C4), 40.25 (C4’), 75.01 (C5), 78.86 (C6), 123.96 (C2’), 124.35 (C2),
124.48 (C11), 124.89 (C7’), 125.34 (C11’), 125.72 (C6’), 126.26 (C7), 129.87 and 130.51 (C15 and
C15’), 131.50 (C10’), 131.77 and 132.00 (C1 and C1’), 132.50 (C14’), 132.89 (C10), 133.21 (C14),
134.27 (C9), 135.38 (C8’), 136.18 and 136.32 (C9’ and C13), 136.86 (C13’), 137.29 (C12’), 138.15
(C8), 138.58 (C12), 139.59 (C5’); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329, found
593.4329.
(5R,6R)-2b (56% from 24b for isomeric mixture, all-E-isomer: 5% from 24b after crystallization):
CD (5.88×10−5 mol/L, EtOH) λ(ꢀε) 213 (0), 242 (–0.36), 270 (+1.68), 272 (+1.54), 279 (+3.18), 293
(0), 329 (–1.20), 344 (–1.73), 387 (–0.17); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329,
found 593.4329.
(5S,6S)-4a (96% from 24a for isomeric mixture, all-E-isomer: 28% from 24a after preparative
HPLC): UV-VIS (EtOH) λ 267, 423, 446, 474 nm; CD (9.48×10−5 mol/L, EtOH) λ(ꢀε) 215 (–1.85),
227 (0), 236 (+0.76), 247 (0), 273 (–3.06), 301 (0), 331 (+0.93), 359 (0); 1H NMR (500 MHz, CDCl3)
δ
1.03 (6H, s, 1-gem-Me), 1.14 (3H, s, 5’-CH3), 1.53 (2H, t, J=7.5 Hz, 4’-H2), 1.62 (2H, m, 3-H2),
1.63 and 1.69 (each 3H, br s, 1’-gem-CH3), 1.72 (3H, br s, 5-CH3), 1.92 (3H, br s, 9’-CH3), 1.97 (3H)
and 1.98 (6H) (each br s, 9-CH3, 13-CH3 and 13’-CH3) 2.02 (2H, br t, J=6.5 Hz, 4-H2), 2.09 (1H, s,
5’-OH), 2.12 (2H, br q, J=7.5 Hz, 3’-H2), 2.14 (1H, br d, J=3 Hz, 6’-OH), 4.04 (1H, dd, J=3, 7.5 Hz,
6’-H), 5.13 (1H, br t, J=7.5 Hz, 2’-H), 5.70 (1H, dd, J=7.5, 15.5 Hz, 7’-H), 6.13 (1H, d, J=15.5 Hz,
8-H), 6.15 (1H, br d, J=11.5 Hz, 10-H), 6.18 (1H, br d, J=15.5 Hz, 7-H), 6.20 (1H, br d, J=11.5 Hz,
10’-H), 6.25 (1H, overlapped, 14-H), 6.27 (1H, overlapped, 14’-H), 6.35 (1H, d, J=15 Hz, 12-H), 6.38
(2H, d, J=15 Hz, 8’-H and 12’-H), 6.59 (1H, dd, J=11.5, 15 Hz, 11’-H), 6.60–6.68 (2H, m, 15-H and
15’-H), 6.66 (1H, dd, J=11, 15 Hz, 11-H); 13C NMR (125 MHz, CDCl3)
δ 12.78, 12.80, 12.85 and
12.99 (9-CH3, 9’-CH3, 13-CH3 and 13’-CH3), 17.69 and 25.72 (1’-gem-CH3), 19.28 (C3), 21.43
(5’-CH3), 21.77 (5-CH3), 22.12 (C3’), 28.99 (1-gem-CH3), 33.14 (C4), 34.29 (C1), 38.81 (C4’), 39.67
(C2), 75.00 (C5’), 78.86 (C6’), 124.35 and 124.45 (C2’ and C11’), 125.25 (C11), 126.26 (C7’), 126.78
(C7), 129.42 (C5), 129.80 and 130.49 (C15 and C15’), 130.81 (C10), 131.99 (C1’), 132.30 (C14),
17

ACCEPTED MANUSCRIPT
132.89 (C10’), 133.20 (C14’), 134.26 (C9’), 136.12 and 136.19 (C9 and C13’), 136.84 (C13), 137.17
(C12), 137.75 (C8), 137.92 (C6), 138.14 (C8’), 138.58 (C12’); HRMS (ESI) m/z calcd for C40H58O2Na
[M+Na]+ 593.4329, found 593.4326.
(5R,6R)-4b (82% from 24b for isomeric mixture, all-E-isomer: 17% from 24b after preparative
HPLC): CD (8.72×10−5 mol/L, EtOH) λ(ꢀε) 212 (+0.75), 224 (0), 240 (–0.61), 253 (0), 269 (+1.17),
290 (0), 327 (–0.67), 365 (0); HRMS (ESI) m/z calcd for C40H58O2Na [M+Na]+ 593.4329, found
593.4328.
References and notes
1. (a) DiMascio, P.; Kaiser, S.; Sies, H. Arch. Biochem. Biophys. 1989, 274, 532–538.
(b) Cantrell, A.; MacGarvey, D. J.; Truscott, T. G.; Rancan, F.; Böhm, F. Arch. Biochem.
Biophys. 2003, 412, 47–54.
2. Nishino, H.; Tokuda, H.; Satomi, Y.; Masuda, M.; Bu, P.; Onozuka, M.; Yamaguchi, S.; Okuda,
Y.; Takayama, Y.; Tsuruta, J.; Okuda, M.; Ichiishi, E.; Murakoshi, M.; Yano, M. Pure. Appl.
Chem. 1999, 71. 2273–2278.
3. Gann, P. H.; Ma, J.; Giovannucci, E.; Willett, W.; Sacks, F. M.; Hennekens, C. H.; Stampfer. M.
J. Cancer Res. 1999, 59, 1225–1230.
4. Khachik, F.; Steck, A.; Niggli, U. A.; Pfander, H. J. Agric. Food Chem. 1998, 46, 4874–4884;
Total synthesis of racemic 2,6-cyclo-γ-carotene-1,5-diol was reported by their group: Crawford,
K.; Mazzola, E.; Khachik, F. Synthesis 2014, 46, 635–645.
5. Maoka, T.; Yamano, Y.; Wada, A.; Etho, T.; Terada, Y.; Tokuda, H.; Nishino, H. J. Agric. Food
Chem. 2015, 63, 1622–1630.
6. Märki-Fischer, E.; Marti, U.; Buchecker, R.; Eugster, C. H. Helv. Chim. Acta 1983, 66, 494–513.
7. Andrä, M. S.; Tzschucke, C. C. Eur. J. Org. Chem. 2014, 7265–7272.
8. Bernhard, K.; Kienzle, F.; Mayer, H.; Müller, R. K. Helv. Chim. Acta 1980, 63, 1473–1490.
9. Gao, Y.; Hanson, R. M.; Klunder, J. M.; Ko, S. Y.; Masamune, H.; Sharpless, K. B. J. Am.
Chem. Soc. 1987, 109, 5765–5780.
10. Nacro, K.; Baltas, M.; Escudier, J.-M.; Gorrichon, L. Tetrahedron 1996, 52, 9047–9056.
11. Li, S.; Fang, L.; Zhong, J.; Shen, J.; Xu, H.; Yang, Y.; Hou, S.; Bian, Q. Tetrahedron: Asymmetry
2014, 25, 591–595.
12. Hanson, R. M. Tetrahedron Lett. 1984, 25, 3783–3786.
13. Faraldos, J. A.; Coates, R. M.; Giner, H.-L. J. Org. Chem. 2013, 78, 10548–10554.
14. Methanolysis of dinitrobenzoate in the presence of amine will be described elsewhere.
15. The numbering system for carotenoids is used.
16. The allylic alcohol moiety of apocarotenal 18a is unstable under acidic conditions.
17. Fischli, A.; Mayer, H.; Simon, W.; Stoller, H.-J. Helv. Chim. Acta 1976, 59, 397–405.
18. Yamano, Y.; Chary, M. V.; Wada, A. Org. Lett. 2013, 15, 5310–5313.
19. Mori, N.; Kuwahara, Y.; Kurosa, K. Bioorg. Med. Chem. 1996, 4, 289–295.
20. Our attempt to apply the HPLC method to natural specimens previously isolated5 from gac was
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ACCEPTED MANUSCRIPT
unsuccessful due to its decomposition during a series of acquisition some spectral data. The fact
that carotenoids synthesized here are relatively stable by keeping in a freezer for several months
has convinced us to determine the absolute configurations of natural specimens if it is obtained
and kept in an appropriate way.
19