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15.9 Hz, H-8), 5.34–5.30 (1H, m, H-5), 4.39–4.38 (1H, m, H-4), 4.11– evaporated and puried through column chromatography to
4.09 (1H, m, H-3), 2.20–2.16 (1H, m, H-2, 6), 2.03–2.01 (1H, m, H-2, offer pure compounds 11–14.
6), 1.92–1.89 (1H, m, H-2, 6), 1.83–1.79 (1H, m, H-2, 6), 1.40 (3H, s,
3,4-Dihydroxycinnamic acid cyclohexanol ester (11). Column
–CH3), 1.25 (3H, s, –CH3); 13C-NMR (150 MHz, DMSO-d6) d: 176.2, chromatography (eluent: DCM–CH3OH, 10 : 1), white power,
165.8, 148.5, 145.6, 145.4, 125.6, 121.5, 115.8, 114.9, 114.1, 108.1, 68% yield; 1H-NMR (600 MHz, CDCl3) d: 7.57 (d, J ¼ 15.9 Hz, 1H,
76.5, 73.2, 72.7, 70.4, 36.4, 34.8, 28.1, 26.0; HRMS (ESI): calcd for [M CH]CH), 7.11 (d, J ¼ 2.0 Hz, 1H, H-2), 7.02 (dd, J ¼ 8.2 Hz,
+ Na]+C19H22NaO9: 417.1162, found 417.1167.
2.0 Hz, 1H, H-6), 6.88 (d, J ¼ 2.0 Hz, 1H, H-5), 6.27 (d, J ¼
2.3.2. 3,4-di-O-Isopropylidene-5-(30,40-dihydroxycinnamoyl)-
15.9 Hz, 1H, CH]CH), 4.90–4.86 (m, 1H, O–CH), 1.93–1.90 (m,
1-O-quinic acid isopropylidene ester 5. To a solution of chloro- 2H, O–CH–CH2), 1.79–1.74 (m, 2H, O–CH–CH2), 1.58–1.25 (m,
genic acid (1) (0.35 g, 1 mmol) in dry tetrahydrofuran (THF) 5 mL, 6H, –(CH2)3–); 13C-NMR (150 MHz, CDCl3) d: 167.7, 146.5, 144.9,
dry acetone (0.18 g, 3 mmol) was added. Then lewis acid TMSOTf 143.9, 127.6, 122.4, 116.2, 115.6, 114.6, 73.3, 31.8, 25.5, 23.9.
(165 mL, 1 mmol) was dropped at 0 ꢀC and reacted for overnight.
Cinnamic acid cyclohexanol ester (12). Column chromatog-
When complete, sodium hydroxide (NaOH, 1.0 mol Lꢁ1) 0.18 mL raphy (eluent: PE–EA, 30 : 1), colourless oil, 53% yield; 1H-NMR
was added to quench the reaction. Then the crude product was (600 MHz, pyridine-d5) d: 7.67 (d, J ¼ 16.1 Hz, 1H, CH]CH),
extracted with ethyl acetate 20 mL, evaporated and subjected to 7.54–7.52 (m, 2H, H-2, 6), 7.38–7.37 (m, 1H, H-3, 4, 5), 6.44 (d, J
column chromatography (eluent: PE–EtOAc, 10 : 1) to gain pure ¼ 16.0 Hz, 1H, CH]CH), 4.91–4.87 (m, 1H, O–CH), 1.94–1.91
light yellow powder, 65% yield; 1H-NMR (600 MHz, DMSO-d6) d: (m, 2H, O–CH–CH2), 1.79–1.75 (m, 2H, O–CH–CH2), 1.61–1.25
9.60 (1H, s, Ph'–OH), 9.16 (1H, s, Ph'–OH), 7.51 (1H, d, J ¼ (m, 6H, –(CH2)3–); 13C-NMR (150 MHz, CDCl3) d: 166.5, 144.4,
15.8 Hz, H-9), 7.11–6.95 (2H, m, H-20, 60), 6.77 (1H, d, J ¼ 7.7 Hz, 134.6, 130.2, 129.0, 128.1, 119.0, 72.9, 31.9, 25.5, 23.9.
H-50), 6.25 (1H, d, J ¼ 15.8 Hz, H-8), 5.24–5.10 (1H, m, H-5), 4.50–
3-Hydroxycinnamic acid cyclohexanol ester (13). Column
4.36 (1H, m, H-4), 4.26–4.14 (1H, m, H-3), 2.29–2.18 (2H, m, H-2, chromatography (eluent: PE–EA, 20 : 1), white solid, 62% yield;
6), 2.14–2.04 (1H, m, H-2, 6), 1.90–1.79 (1H, m, H-2, 6), 1.57 (6H, s, 1H-NMR (600 MHz, pyridine-d5) d: 7.60 (d, J ¼ 16.0 Hz, 1H, CH]
2ꢂ –CH3), 1.41 (3H, s, –CH3), 1.27 (3H, s, –CH3); 13C-NMR (150 CH), 7.22–7.19 (m, 1H, H-5), 7.05–7.02 (m, 1H, H-4, 6), 6.91–6.89
MHz, DMSO-d6) d: 173.7, 165.9, 148.4, 145.7, 145.6, 125.5, 121.4, (m, 1H, H-2), 6.38 (d, J ¼ 16.0 Hz, 1H, CH]CH), 4.90–4.86 (m,
115.8, 114.9, 113.7, 110.4, 108.2, 78.2, 75.6, 72.5, 69.4, 35.7, 34.0, 1H, O–CH), 1.92–1.91 (m, 2H, O–CH–CH2), 1.78–1.72 (m, 2H, O–
28.1, 27.9, 27.8, 25.7; HRMS (ESI): calcd for [M + Na]+ CH–CH2), 1.57–1.23 (m, 6H, –(CH2)3–); 13C-NMR (150 MHz,
C22H26NaO9: 457.1475, found 457.1472.
CDCl3) d: 167.0, 156.9, 144.7, 135.9, 130.0, 120.3, 118.8, 117.7,
2.3.3. 3,4-di-O-Isopropylidene-5-O-(30,40-dibenzyloxycinn 114.7, 73.2, 31.8, 25.5, 23.9.
amoyl)-quinic acid benzyl ester 6. A mixture of 4 (0.05 g, 0.13
4-Hydroxycinnamic acid cyclohexanol ester (14). Column
mmol) and K2CO3 (0.09 mg, 0.6 mmol) was stirred in dry chromatography (eluent: PE–EA, 20 : 1), white solid, 64% yield;
CH3CN 20 mL, then 5 mL BnBr was dropped, and reuxed 1H-NMR (600 MHz, pyridine-d5) d: 7.61 (d, J ¼ 15.9 Hz, 1H, CH]
under N2 until completion. The mixture was added ethyl CH), 7.38–7.37 (m, 2H, H-2, 6), 6.87–6.86 (m, 2H, H-3, 5), 6.26 (d,
acetate 20 ml. The combined organic layer was washed with J ¼ 15.9 Hz, 1H, CH]CH), 4.89–4.86 (m, 1H, O–CH), 1.91–1.90
5% NaHCO3 aqueous solution, and NaCl saturated aqueous (m, 2H, O–CH–CH2), 1.79–1.72 (m, 2H, O–CH–CH2), 1.55–1.22
solution in sequence, and then dried over Na2SO4 and puri- (m, 6H, –(CH2)3–); 13C-NMR (150 MHz, CDCl3) d: 168.0, 158.5,
ed through column chromatography (eluent: PE–EtOAc, 145.0, 130.1, 126.8, 116.1, 115.6, 73.3, 31.8, 25.5, 23.9.
1 : 1) to offer pure white solid, 67% yield; 1H-NMR (600 MHz,
DMSO-d6) d: 7.55 (1H, d, J ¼ 15.7 Hz, H-9), 7.50 (1H, d, J ¼
2.4. Evaluation of the biological activity
1.7 Hz, Bn0–H), 7.47–7.44 (4H, m, Bn0–H), 7.40–7.36 (8H, m,
Bn0–H), 7.33–7.31 (3H, m, Bn0–H, H-20), 7.22 (1H, dd, J ¼
2.4.1. Cell culture. HepG2 cells were originated from
2.0 Hz, 8.3 Hz, H-60), 7.08 (1H, d, J ¼ 8.3 Hz, H-50), 6.50 (1H, d, American Type Culture Collection (ATCC) (Manassas, VA, USA)
J ¼ 15.9 Hz, H-8), 5.37–5.33 (1H, m, H-5), 5.19 (4H, s, Bn0– and obtained from the Peking Union Medical College (Beijing,
CH2), 5.15–5.09 (2H, m, Bn0–CH2), 4.41–4.39 (1H, m, H-4), China). HepG2 cells were cultured in Dulbecco's modied Eagle
4.12–4.11 (1H, m, H-3), 2.24–2.20 (1H, m, H-2, 6), 2.10–2.07 medium (DMEM) supplemented with 10% fetal bovine serum,
(1H, m, H-2, 6), 1.99–1.96 (1H, m, H-2, 6), 1.88–1.84 (1H, m, 1% penicillin and streptomycin at 37 ꢀC in 5% CO2 atmosphere.
H-2, 6), 1.41 (3H, s, –CH3), 1.26 (3H, s, –CH3); 13C-NMR (150 When grown to 70–80% conuence, cells were incubated in
MHz, DMSO-d6) d: 174.2, 166.3, 151.2, 149.0, 145.1, 137.0, serum-free DMEM containing 100 mmol Lꢁ1 oleic acid and co-
136.8, 135.2, 128.8, 128.7, 128.7, 128.3, 128.1, 128.0, 127.4, treated with 10 mmol Lꢁ1 of simvastatin or CA, compound 4–
127.3, 123.1, 115.9, 114.3, 113.9, 109.7, 77.4, 77.0, 74.0, 71.4, 6, 11–14 for 24 h respectively. Cells maintained in serum-free
71.1, 70.7, 67.9, 37.0, 34.5, 28.1, 26.0; HRMS (ESI): calcd for DMEM were used as the blank control. Compounds were dis-
[M + Na]+ C40H40NaO9: 687.2570, found 687.2573.
2.3.4. General procedure for the synthesis of compounds it was added in control group.
solved in dimethyl sulphoxide (DMSO) and the equal volume of
11–14. To a solution of cinnamic acid 7–10 (0.55 mmol) in dry
2.4.2. Oil red O staining. The cells with 70–80% conuence
1,4-dioxane (10 mL), SOCl2 (0.37 mmol) were added dropwise in 96 well plates were incubated in serum-free DMEM + OA (100
and stirred under reux for 2 h. To this mixture, n-hexanol (0.37 mmol Lꢁ1) and 10 mmol Lꢁ1 of CA, compound 4–6, 11–14
mmol) was added respectively at 0 ꢀC and the reaction mixture respectively or the positive control simvastatin (10 mmol Lꢁ1) for
was stirred until its completion during 4–6 h. The solvent was 24 h. Cells were then xed with 4% w/v paraformaldehyde
This journal is © The Royal Society of Chemistry 2019
RSC Adv., 2019, 9, 12247–12254 | 12249