Neuroprotective Caffeoylquinic Acid Derivatives from the Flowers of

Mar 1, 2017 - The relative configuration of 1 was determined by the coupling ... To determine the absolute configuration of 1, the experimental and ca...
5 downloads 0 Views 1MB Size
Download PDF
Article pubs.acs.org/jnp

Neuroprotective Caffeoylquinic Acid Derivatives from the Flowers of Chrysanthemum morifolium Peng-Fei Yang, Zi-Ming Feng, Ya-Nan Yang, Jian-Shuang Jiang, and Pei-Cheng Zhang* State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, People’s Republic of China S Supporting Information *

ABSTRACT: Three new caffeoylquinic acid derivatives, chrysanthemorimic acids A−C (1−3), and 11 known compounds (4−14) were isolated and characterized from the flowers of Chrysanthemum morifolium. Their structures were confirmed by spectroscopic data as well as by comparison of the experimental and calculated electronic circular dichroism spectra. Chrysanthemorimic acids A−C possess a rare 8-oxa-bicyclo[3.2.1]oct-3-en-2-one ring that is formed through a [5+2] cycloaddition of caffeoylquinic acid with a D-glucose derivative. Compounds 1−3, 6−8, 12, and 13 displayed significant effects against hydrogen peroxide-induced neurotoxicity in SH-SY5Y cells at 10 μM.

■

C

RESULTS AND DISCUSSION Chrysanthemorimic acid A (1) was isolated as a white, amorphous powder. Its molecular formula, C31H30O15, was determined by HRESIMS (m/z 665.1477 [M + Na]+, calcd for C31H31NaO15, 665.1482), which indicated 17 indices of hydrogen deficiency. The IR spectra displayed the characteristic absorptions of hydroxy (3390 cm−1), carbonyl (1694 cm−1), and aromatic ring (1523 and 1447 cm−1) functionalities. The 1 H NMR data (Table 1) showed two ABX spin systems at δH 7.06 (1H, d, J = 2.0 Hz), 6.79 (1H, d, J = 8.0 Hz), 6.97 (1H, dd, J = 8.0, 2.0 Hz), 6.61 (1H, d, J = 2.0 Hz), 6.60 (1H, d, J = 8.0 Hz), and 6.49 (1H, dd, J = 8.0, 2.0 Hz), which were attributable to two 1,3,4-trisubstituted benzene moieties as well as two trans-olefinic protons at δH 7.60 (1H, d, J = 16.0 Hz) and 6.27 (1H, d, J = 16.0 Hz). These NMR data suggest the presence of a caffeoyl group, which was supported by the 1H−1H COSY correlation of H-7′/H-8′ and the HMBC cross-peaks (Figure 2a) from H-7′ to C-1′, C-2′, C-6′, and C-9′ and from H-8′ to C-1′ and C-9′. A quinic acid moiety was deduced based on the 1 H NMR resonances, including three oxymethine protons at δH 5.64, 5.01, and 4.32, together with two sets of sp3 methylene protons at δH 2.20−2.23 and 2.01−2.12. In the 13C NMR spectrum, the resonances at δC 70.0 (C-3) and 76.0 (C-4), 69.7 (C-5) were assigned to three oxygenated methine carbons. In addition, two sp3 methylene carbons at δC 39.6 (C-2) and 38.2 (C-6), an oxygenated tertiary carbon at δC 76.3 (C-1), and a carbonyl at δC 176.9 (C-7) were also present in the 13C NMR spectrum, confirming the presence of a quinic acid unit. The

affeoylquinic acid (CQA) esters, formed from a quinic acid and caffeic acids, are a class of phenolic compounds that can be isolated from a variety of plants and fruits such as blueberries, pears, tomatoes, and potatoes. According to the number of caffeic acid moieties, CQA derivatives can be further divided into mono-, di-, tri-, and tetra-CQAs.1−4 The pharmacological activities of CQA derivatives including antioxidant,5 anti-inflammatory,6 hepatoprotective,7 antidiabetic,8 antiviral,9 and anti-HIV effects10 have been reported. It has been demonstrated that CQA derivatives possess neuroprotective effects against Aβ-induced human SH-SY5Y cell neurotoxicity.11 In our exploration of neuroprotective agents from natural sources, the flowers of Chrysanthemum morifolium Ramat. were investigated. C. morifolium, cultivated in the Zhejiang Province of China as “Hangbaiju” (HJ), has been traditionally consumed as a medicinal remedy for approximately 2000 years. Previous chemical investigations revealed that the plant contains CQA derivatives,12 flavonoids,13 sesquiterpenoids,14 triterpenoids,15 and unsaturated fatty acids.16 The CQA derivatives and flavonoids are considered to be the main bioactive components. In the current study, three new caffeoylquinic acid derivatives (1−3) featuring an 8oxa-bicyclo[3.2.1]oct-3-en-2-one ring, along with 11 known compounds (4−14) (Figure 1), were isolated. The structures were identified via physical data analyses, and the absolute configurations were confirmed by comparison of their experimental and calculated electronic circular dichroism (ECD) spectra. Their neuroprotective properties were also assessed. © 2017 American Chemical Society and American Society of Pharmacognosy

Received: November 9, 2016 Published: March 1, 2017 1028

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033

Journal of Natural Products

Article

Figure 1. Compounds 1−14 isolated from the flowers of Chrysanthemum morifolium.

quinic acid moiety was confirmed by the 1H−1H COSY correlations of H-2ax/H-3/H-4/H-5/H-6ax and HMBC crosspeak from H-2ax to C-7 (δC 176.9). The HMBC cross-peak between H-4 (δH 5.01) and C-9′ (δC 168.3) suggested the caffeoyl group was linked to the 4-OH group of quinic acid, which was further confirmed by comparing the 1H NMR data (Figures 14 and 15, Supporting Information), optical rotation, and HPLC analysis of the acid-hydrolysis product 1b with that of the standard 4-O-caffeoylquinic acid. The presence of a dihydrocaffeoyl substituent was indicated by the remaining 1,3,4-trisubstituted phenyl unit and carbonyl moiety at δC 172.8, which is more deshielded than the typical 13C NMR chemical shift of a caffeic acid carbonyl carbon. This was confirmed by the 1H−1H COSY cross-peaks H-7″/H-8″, HMBC cross-peaks from H-7″ to C-1″, C-2″, C-6″, and C9″, and those from H-8″ to C-1′ and C-9″. In addition, the NMR data also showed an α,β-unsaturated carbonyl unit at δH 6.81 (d, 10.0 Hz) and 6.17 (dd, 1.0, 10.0 Hz) and δC 155.2, 128.5, and 196.7. The spin-coupling sequence of H-7″/H-8″/ H-10″ in the 1H−1H COSY spectrum implied the structural fragment C-7″−C-8″−C-10″. In the HMBC spectrum, crosspeaks from H-7″ to C-13″, H-8″ to C-11″, H-10″ to C-12″, and H-13″ to C-11″ permitted the establishment of a cycloheptene functionality. Among the 17 indices of hydrogen deficiency, 16 were accounted for by a caffeoyl group, a quinic acid moiety, a dihydrocaffeoyl substituent, a cycloheptene ring, and an α,βunsaturated carbonyl unit. Thus, the remaining index should represent an additional ring via an oxygen bridge between C10″ and C-14″, which was supported by the HMBC cross-peak from H-10″ to C-14″. The oxygenated methylene group was located at C-14″ based on the HMBC cross-peaks from H2-15″

to C-14″ and C-13″. The presence of the cross-peak from H-3 to C-9″ indicated that the C-9″ carbonyl (δC 172.8) was linked to OH-3 of the quinic acid moiety. Thus, the 2D structure of compound 1 was established to contain a rare 8-oxabicyclo[3.2.1]oct-3-en-2-one scaffold. The relative configuration of 1 was determined by the coupling constant and ROESY data (Figure 2b). The J7″,8″ value of 7.5 Hz in combination with the ROESY correlations from H8″ to H-2″ and H-6″ suggested that H-7″ and H-8″ were transorientated.17 The ROESY cross-peak between H-7″ and H-10″ revealed that H-7″ and H-10″ were cofacial, as is 14-CH2OH″. To determine the absolute configuration of 1, the experimental and calculated ECD data were used. Considering the numerous possible conformers for 4-O-caffeoylquinic acid, a simplified compound (1a) was obtained from acid hydrolysis of 1. According to the analysis of the relative configuration of 1, there are only two feasible stereoisomers of 1a, (7″S,8″R,10″R,14″R)-1a and its enantiomer (7″R,8″S,10″S,14″S)-1a. A systematic conformational analysis for (7″R,8″S,10″S,14″S)-1a was performed using a molecular mechanics force field calculation, and 13 conformers with Boltzmann distribution over 1% were chosen for ECD calculations (Table S1, Supporting Information). The optimized conformations of these 13 conformers were achieved using TDDFT and were carried out at the B3LYP/6311+G(d,p) level in MeOH. The overall calculated ECD spectrum (Figure 3) was created by Boltzmann weighting of their lowest energy conformers. The results showed that the calculated ECD curve of (7″S,8″R,10″R,14″R)-1a was in good agreement with the experimental ECD spectrum of 1a (Figure 1029

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033

Journal of Natural Products

Article

Table 1. NMR Spectroscopic Data for Compounds 1−3 in Methanol-d4 (500 MHz for 1H NMR, 125 MHz for 13C NMR) 1 position

δH (J, Hz)

2 δC

δH (J, Hz)

3 δC

1 2

2.20−2.23, m

76.3 39.6

2.17−2.23, m

76.4 39.5

3 4 5 6

5.64, m 5.01, dd (3.0, 10.0) 4.32, m 2.01−2.12, m

70.0 76.0 69.7 38.2

5.66, m 4.86, dd (3.0, 9.5) 4.29, m 2.04−2.16, m

69.8 76.6 69.5 38.1

7 1′ 2′ 3′ 4′ 5′ 6′ 7′ 8′ 9′ 1″ 2″ 3″ 4″ 5″ 6″ 7″ 8″ 9″ 10″ 11″ 12″ 13″ 14″ 15″

7.06, d (2.0)

6.79, 6.97, 7.60, 6.27,

d (8.0) dd (2.0, 8.0) d (16.0) d (16.0)

6.61, d (2.0)

6.60, 6.49, 3.69, 3.19,

d (8.0) dd (2.0, 8.0) d (7.5) dd (1.5, 7.5)

4.63, m 6.17, dd (1.0, 10.0) 6.81, d (10.0) α 3.66, d (12.5) β 3.76, d (12.5)

176.9 127.7 115.2 146.8 149.7 116.4 123.2 147.9 114.6 168.3 127.7 116.3 146.1 146.3 116.3 121.5 53.4 53.6 172.8 84.3 196.7 128.5 155.2 88.2 63.9

7.07, d (2.0)

6.80, 6.97, 7.58, 6.25,

d (8.0) dd (2.0, 8.0) d (16.0) d (16.0)

6.58, d (2.0)

6.57, 6.45, 3.59, 3.22,

d (8.0) dd (2.0, 8.0) d (7.5) dd (1.5, 7.5)

4.77, br s 6.18, dd (1.0, 10.0) 6.79, d (10.0) α 3.58, d (12.5) β 3.73, d (12.5)

175.5 127.7 115.3 146.8 149.7 116.5 123.3 147.9 114.7 168.3 127.8 116.3 146.1 146.3 116.2 121.6 53.9 53.3 173.0 84.7 196.7 128.6 155.3 88.3 64.0

δH (J, Hz) α 2.04, dd (10.0, 14.5) β 2.10, dd (4.0, 14.5) 4.22, m 4.96, dd (3.0, 8.5) 5.55, m α 2.26, dd (4.0, 14.5) β 1.99, dd (4.0, 14.5)

7.05, d (2.0)

6.79, 6.95, 7.48, 6.14,

d (8.5) dd (2.0, 8.5) d (16.0) d (16.0)

6.65, d (2.0)

6.61, 6.53, 3.75, 3.29,

d (8.0) dd (2.0, 8.0) d (7.5) dd (1.5, 7.5)

4.83, m 6.19, dd (1.0, 10.0) 6.80, d (10.0) α 3.82, d (12.5) β 3.72, d (12.5)

δC 75.1 41.6 65.7 77.3 69.7 36.9 178.2 127.9 115.3 146.7 149.3 116.5 123.2 147.4 115.2 168.3 128.0 116.6 146.0 146.3 116.3 121.6 53.6 53.2 173.5 84.5 196.9 128.6 155.6 88.3 63.9

Figure 2. (a) 1H−1H COSY and key HMBC correlations of compound 1. (b) Selected ROESY correlations of compound 1.

carbons. The 1H NMR resonances of 2 recorded in methanold4 were similar to those of 1, with slight differences in the chemical shift values and resonance pattern of H-10″ (δH 4.77 brs) in 2. Analysis of the 1H−1H COSY and HMBC spectra suggested that 2 shared the same 2D structure as 1. The relative configuration of 2 was established via its ROESY spectrum. After acid hydrolysis of compound 2, the hydrolysate 2a had a mirror-image-like ECD spectrum compared to 1a, which

3), implying the absolute configuration of chrysanthemorimic acid A (1) as (7″S, 8″R, 10″R, 14″R). Chrysanthemorimic acid B (2) shared the same molecular formula of C31H30O15 as 1 on the basis of the sodium-adduct HRESIMS ion (m/z 665.1477 [M + Na]+, calcd for 665.1482). The 1D NMR (Table 1) and HSQC data showed the presence of four carbonyls, 12 aromatic carbons, four olefinic carbons, three methylenes, six methines, and two oxygenated tertiary 1030

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033

Journal of Natural Products

Article

comparing the experimental ECD spectra of its acid hydrolysate 3a with the calculated ECD spectrum of (7″S,8″R,10″R,14″R)1a (Figure 3). The well-matched spectra confirmed the absolute configuration of chrysanthemorimic acid C (3) as (7″S, 8″R, 10″R, 14″R). A tentative biosynthesis pathway to the formation of chrysanthemorimic acids A−C (1−3) is proposed in Scheme 1. Keto−enol isomerization of D-glucose affords intermediate i (D-fructose), which is transformed to ii through successive dehydration of the 4- and 3-hydroxy groups. Intermediate ii is cyclized to give compound iii, which undergoes dehydration and aromatization to afford an oxypyrylium zwitterion. Chrysanthemorimic acids A−C are hypothesized to be formed via the [5+2] cycloaddition17 of the oxypyrylium zwitterion with 3,4-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid. The 11 known compounds 4-O-acetylchlorogenic acid (4),18 chlorogenic acid (5),19 1,5-di-O-caffeoylquinic acid (6),20 3,4di-O-caffeoylquinic acid (7),21 luteolin-7-O-β-D-glucopyranoside (8),22 luteolin-7-O-(6″-O-acetyl)-β-D-glucopyranoside (9),23 apigenin7-O-β-D-glucopyranoside (10),22 diosmetin-7O-β-D-glucopyranoside (11),22 luteolin-7-O-β-D-glucuronide (12),24 luteolin-7-O-rutinoside (13),25 and luteolin-7,4′-di-Oβ-D-glucopyranoside (14)26 were identified by comparing their observed and reported NMR and MS data. All isolated compounds were tested for their neuroprotective effects against hydrogen peroxide (H2O2)-, oxygen-glucose deprivation (OGD)-, and glutamate-induced cell injury in SHSY5Y cells. At 10 μM, all the compounds were inactive against OGD- and glutamate-induced cell injury in SH-SY5Y cells. However, as observed in Figure 4, compounds 1−3, 6−8, 12, and 13 at 10 μM attenuate SH-SY5Y cell damage induced by H2O2. Compounds 1−3 exhibited neuroprotective effects with cell viability of 78.4%, 76.7%, and 75.6%, respectively, compared with that of the positive control L-NBP (L-3-nbutylphthalide) with cell viability of 73.9%. Compound 5 exhibited no effect on H2O2-induced neurotoxicity, and compound 4 displayed cytotoxicity at 10 μM.

Figure 3. Calculated ECD spectra of (7″S,8″R,10″R,14″R)-1a and (7″R,8″S,10″S,14″S)-1a and the experimental ECD spectra of 1a, 2a, and 3a in MeOH.

indicated that they were enantiomers. The experimental ECD spectrum of 2a was similar to the calculated ECD curve of (7″R,8″S,10″S,14″S)-1a (Figure 3), establishing the absolute configuration of chrysanthemic acid B (2) as (7″R, 8″S, 10″S, 14″S). The molecular formula (C31H30O15) of chrysanthemorimic acid C (3) was identical to those of 1 and 2, as deduced from the protonated HRESIMS ion (m/z 643.1657 [M + H]+, calcd for C31H31O15, 643.1663). The 1H NMR and 13C NMR data (Table 1) of 3 were similar to those of 1, except for the chemical shift of the oxymethine H-3 (δH 4.22) of 3, which was shielded by −1.42 ppm, and that of H-5 (δH 5.55) was deshielded by +1.23 ppm, which implied that compound 3 was a regioisomer of 1. This was confirmed by the HMBC crosspeak between H-5 and C-9′, suggesting that the caffeoyl group was esterified to OH-5 of the quinic acid. The HMBC crosspeak from H-4 to C-9″ indicated that the carbonyl carbon at δC 173.5 (C-9″) was esterified to OH-4 of the quinic acid. The residual partial structure of 3 was identical to that of 1 based upon analysis of the 2D NMR spectra. The relative configuration was consistent with that of 1 via the ROESY spectrum. Its absolute configuration was determined by

■

EXPERIMENTAL SECTION

General Experimental Procedures. The optical rotations were measured on a Jasco P-2000 polarimeter (Jasco Inc., Easton, MD,

Scheme 1. Proposed Biosynthesis Pathway of Compounds 1−3

1031

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033

Journal of Natural Products

Article

was purified by preparative RP HPLC, using MeOH/H2O/HOAc (40:60:0.1, v/v) at 6 mL/min, to afford 6 (18 mg). Fraction D10 was further chromatographed on a Sephadex LH-20 column (150 cm × 5 cm i.d.) with MeOH/H2O (0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 85%, and 100%, v/v) to generate fractions D10-1−D10-9. Fraction D10-9 (1.56 g) was subjected to preparative RP HPLC, employing MeOH/H2O/HOAc (40:60:0.1, v/v) as the mobile phase at 6 mL/ min, to give 8 (450 mg), 12 (21 mg), 13 (26 mg), and 14 (12 mg). Fraction D10-8 (0.86 g) was purified by preparative RP HPLC, utilizing MeOH/H2O/HOAc (45:55:0.1, v/v) at 6 mL/min, to acquire 9 (18 mg), 10 (21 mg), and 11 (26 mg). Chrysanthemorimic acid A (1): white, amorphous power; [α]20D +180 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 331 (3.89), 293 (3.79) nm; ECD (c 3.1 × 10−4, MeOH) λmax (Δε) 214 (29.0), 249 (−0.85), 285 (1.9), 346 (2.4) nm; IR (KBr) νmax 3390, 1694, 1631, 1605, 1523, 1447, 1375, 1275, 980, 814 cm−1; NMR data see Table 1; HRESIMS m/z 665.1477 [M + Na] + (calcd for 665.1482, C31H30O15Na). Chrysanthemorimic acid B (2): white, amorphous power; [α]20D −285 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 332 (4.05), 295 (3.94) nm; ECD (c 3.1 × 10−4, MeOH) λmax (Δε) 214 (−35.0), 250 (1.5), 299 (−2.6), 338 (−2.9) nm; IR (KBr) νmax 3387, 1694, 1605, 1523, 1447, 1270, 1044, 980, 813 cm−1; NMR data see Table 1; HRESIMS m/z 665.1477 [M + Na] + (calcd for 665.1482, C31H30O15Na). Chrysanthemorimic acid C (3): white, amorphous power; [α]20D +162 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 330 (4.01), 295 (3.92) nm; ECD (c 2.1 × 10−4, MeOH) λmax (Δε) 214 (37.0), 250 (−1.3), 285 (2.1), 348 (3.0) nm; IR (KBr) νmax 3389, 1692, 1605, 1523, 1447, 1379, 1275, 1122, 1043, 980, 813 cm−1; NMR data see Table 1; HRESIMS m/z 643.1657 [M + H]+ (calcd for 643.1663, C31H31O15). Acid Hydrolysis of Compounds 1−3. Each compound (1, 4.4 mg, 2, 5.3 mg, 3, 6.6 mg) was dissolved in 0.5 M HCl (5 mL) and heated at 50 °C in a water bath. After 65 h, the reaction mixture was filtrered through a 0.45 μm filter and separated by RP-HPLC (MeOH/ H2O/HOAc, 15:85:0.1, v/v) to afford compounds 1a (2.2 mg), 2a (3.1 mg), and 3a (4.2 mg), respectively. Their NMR spectroscopic data are provided in the Supporting Information. Neuroprotective Activity Assay. The neuroprotective effects of compounds 1−14 against H2O2-, glutamate-, and OGD-induced neurotoxicity in SH-SY5Y cells were examined with reference to described procedures.27

Figure 4. Neuroprotective effect of selected compounds against H2O2induced toxicity in SH-SY5Y cells. USA). UV spectra were obtained on a Jasco V-650 spectrophotometer. ECD spectra were measured on a Jasco J-815 spectrometer. IR spectra were measured on a Nicolet 5700 spectrometer (Thermo Scientific, FL, USA). NMR spectra were obtained using an INOVA 500 spectrometer (Varian, Inc., Palo Alto, CA, USA). HRESIMS data were acquired on an Agilent 1100 Series LC/MSD ion trap mass spectrometer (Agilent Technologies, Waldbronn, Germany). Preparative HPLC was carried out on a Shimadzu LC-6AD instrument using a YMC-Pack ODS-column (250 mm × 20 mm, 5 μm, YMC Corp, Kyoto, Japan). Column chromatography was performed using macroporous resin (Diaion HP-20 and SP-700, Mitsubishi Chemical Corp., Tokyo, Japan) and Sephadex LH-20 (Pharmacia Fine Chemicals, Uppsala, Sweden) columns. Plant Material. The dried flowering heads of Chrysanthemum morifolium were collected in Tongxiang, Zhejiang Province, China, in September 2014. The plant was authenticated by Professor Lin Ma (Institute of Materia Medica, Chinese Academy of Medical Sciences, Beijing). A voucher specimen (ID-S-2595) was deposited at the Herbarium of the Department of Medicinal Plants, Institute of Materia Medica, Chinese Academy of Medical Sciences, Beijing, China. Extraction and Isolation. The dried capitula of C. morifolium (100 kg) were extracted with 80% EtOH (3 × 150 L) under reflux for 3 h. After the solvent was evaporated under reduced pressure, the dark brown residue (6.4 kg) was suspended in H2O (10 L) and fractionated sequentially with petroleum ether (5 × 10 L), EtOAc (5 × 10 L), and n-BuOH (6 × 10 L). The n-BuOH-soluble fraction (2355 g) was subjected to chromatography on an HP-20 macroporous absorption resin column and eluted successively with 0%, 15%, 30%, 50%, 75%, and 95% ethanol. The 30% ethanol solution (480 g) was fractionated by column chromatography on adsorptive macroporous resin SP-700, eluting with 10%, 15%, 20%, 25%, 30%, 50%, and 95% EtOH (30 L each) to afford seven fractions (fractions A−G). Fraction D (480 g) was separated on a Sephadex LH-20 column (120 cm × 10 cm i.d.), eluting with a MeOH/H2O (0−100%, with 10% stepwise increase of MeOH) mixture to give fractions D1−D10. Fraction D5 was further chromatographed on a Sephadex LH-20 column (150 cm × 5 cm i.d.) with MeOH/H2O (0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 85%, and 100%, v/v) to generate fractions D51−D5-23. Fraction D5-6 (1.2 g) was repeatedly subjected to Sephadex LH-20 CC (230 cm × 2 cm i.d.) and washed with MeOH/H2O (0:1, 1:9, 1:4, 3:7, and pure MeOH, v/v) to afford six subfractions (fractions D5-6A−D5-6F). Fraction D5-6B (120 mg) was subjected to preparative RP HPLC, using MeOH/H2O/HOAC (30:70:0.1, v/v) as the mobile phase at 6 mL/min, to obtain 1 (14 mg), 2 (23 mg), and 3 (32 mg). Fraction D5-7 (240 mg) was fractionated by preparative RP HPLC, using MeOH/H2O/HOAc (35:65:0.1, v/v) at 6 mL/min, to afford 4 (5 mg). Fraction D5-13 (636 mg) was purified by preparative RP HPLC using MeOH/H2O/HOAc (40:60:0.1, v/v) at 6 mL/min to afford 5 (46 mg) and 7 (24 mg). Fraction D5-16 (66 mg)

■

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jnatprod.6b01026. UV, IR, NMR, ECD, and HRESIMS spectra for new compounds (PDF)

■

AUTHOR INFORMATION

Corresponding Author

*E-mail (P.-C. Zhang): [email protected]. Tel: +86-163165231. ORCID

Ya-Nan Yang: 0000-0001-6367-8431 Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS This research was supported by the Chinese Academy of Medical Sciences (CAMS) Initiative for Innovative Medicine (No. 2016-I2M-1-010). 1032

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033

Journal of Natural Products

■

Article

REFERENCES

(1) Merfort, I. Phytochemistry 1992, 31, 2111−2113. (2) Islam, M. S.; Yoshimoto, M.; Yahara, S.; Okuno, S.; Ishiguro, K.; Yamakawa, O. J. Agric. Food Chem. 2002, 50, 3718−3723. (3) Agata, I.; Goto, S.; Hatano, T.; Nishibe, S.; Okuda, T. Phytochemistry 1993, 33, 508−509. (4) Scholz, E.; Heinrich, M.; Hunkler, D. Planta Med. 1994, 60, 360− 364. (5) Maruta, Y.; Kawabata, J.; Niki, R. J. Agric. Food Chem. 1995, 43, 2592−2595. (6) Peluso, G.; Feo, V. D.; Simone, F. D.; Bresciano, E.; Vuotto, M. L. J. Nat. Prod. 1995, 58, 639−646. (7) Xiang, T.; Xiong, Q. B.; Ketut, A. I.; Tezuka, Y.; Nagaoka, T.; Wu, L. J.; Kadota, S. Planta Med. 2001, 67, 322−325. (8) Xu, D. L.; Wang, Q. C.; Zhang, W. Q.; Hu, B.; Zhou, L.; Zeng, X. X.; Sun, Y. J. Agric. Food Chem. 2015, 63, 3694−3703. (9) Kurata, R.; Adachi, M.; Yamakawa, O.; Yoshimoto, M. J. Agric. Food Chem. 2007, 55, 185−190. (10) Ooi, L. S.; Wang, H.; He, Z.; Ooi, V. E. J. Ethnopharmacol. 2006, 106, 187−191. (11) Han, J.; Miyamae, Y.; Shigemori, H.; Isoda, H. Neuroscience 2010, 169, 1039−1045. (12) Kim, H. J.; Lee, Y. S. Planta Med. 2005, 71, 871−876. (13) Lee, J. S.; Kim, H. J.; Lee, Y. S. Planta Med. 2003, 69, 859−861. (14) Yoshikawa, M.; Morikawa, T.; Toguchida, I.; Harima, S.; Matsuda, H. Chem. Pharm. Bull. 2000, 48, 651−656. (15) Ukiya, M.; Akihisa, T.; Yasukawa, K.; Kasahara, Y.; Kimura, Y.; Koike, K.; Nikaido, T. J. Agric. Food Chem. 2001, 49, 3187−3197. (16) Tsao, R.; Attygalle, A. B.; Schroeder, F. C.; Marvin, C. H.; McGarvey, B. D. J. Nat. Prod. 2003, 66, 1229−1231. (17) Snider, B. B.; Grabowski, J. F. Tetrahedron 2006, 62, 5171− 5177. (18) Dawidowicz, A. L.; Tapek, R. J. Agric. Food Chem. 2012, 60, 12289−12295. (19) Cheminat, A.; Zawatzky, R.; Becker, H.; Brouillard, R. Phytochemistry 1988, 27, 2787−2794. (20) Zhu, X. F.; Zhang, H. X.; Lo, R. J. Agric. Food Chem. 2004, 52, 7272−7278. (21) Lin, L. C.; Kuo, Y. C.; Chou, C. J. J. Nat. Prod. 1999, 62, 405− 408. (22) Jia, L. Y.; Sun, Q. S.; Huang, S. W. Chin. J. Med. Chem. 2010, 27, 140−144. (23) Mizuno, M.; Kato, M.; Linuma, M.; Tanaka, T.; Sakai, H. Phytochemistry 1987, 26, 2418−2420. (24) Jin, X.; Shi, S. M.; Zhang, D. F.; Zhu, Z. Chin. Tradit. Herb. Drugs 2012, 43, 47−50. (25) Niu, B. J.; Ma, Z. K.; Liao, Z. X.; Ji, L. J.; Sun, H. F. Chin. Tradit. Herb. Drugs 2013, 44, 147−152. (26) Zhao, Q.; Qiu, L.; Bo, G. M.; Qu, G. X.; Qiu, F. J. Shengyang Pharm. Univ. 2006, 23, 151−155. (27) Huang, S. W.; Qiao, J. W.; Sun, X.; Gao, P. Y.; Li, L. Z.; Liu, Q. B.; Sun, B.; Wu, D. L.; Song, S. J. J. Funct. Foods 2016, 24, 183−195.

1033

DOI: 10.1021/acs.jnatprod.6b01026 J. Nat. Prod. 2017, 80, 1028−1033