Bioactive Asarone-Derived Phenylpropanoids from the Rhizome of

Article Cite This: J. Nat. Prod. 2017, 80, 2923-2929

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Bioactive Asarone-Derived Phenylpropanoids from the Rhizome of Acorus tatarinowii Schott En Gao,†,§ Zheng-Qun Zhou,†,‡,§ Jian Zou,† Yang Yu,† Xiao-Lin Feng,† Guo-Dong Chen,† Rong-Rong He,† Xin-Sheng Yao,† and Hao Gao*,† †

Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy/Guangdong Province Key Laboratory of Pharmacodynamic Constituents of TCM and New Drugs Research, and ‡Integrated Chinese and Western Medicine Postdoctoral Research Station, Jinan University, Guangzhou 510632, People’s Republic of China S Supporting Information *

ABSTRACT: Eight new (1a/1b, 2a, 3a, 4a/4b, and 5a/5b) and seven known (2b, 3b, and 6−10) asarone-derived phenylpropanoids, a known asarone-derived lignan (12), and four known lignan analogues (11 and 13−15) were isolated from the rhizome of Acorus tatarinowii Schott. The structures were elucidated via comprehensive spectroscopic analyses, modified Mosher’s method, and quantum chemical calculations. Compounds 1−8 were present as enantiomers, and 1− 5 were successfully resolved via chiral-phase HPLC. Compounds 1a/1b were the first cases of asarone-derived phenylpropanoids with an isopropyl C-3 side-chain tethered to a benzene core from nature. Hypoglycemic, antioxidant, and AChE inhibitory activities of 1−15 were assessed by the αglucosidase inhibitory, ORAC, DPPH radical scavenging, and AChE inhibitory assays, respectively. All compounds except 3a showed α-glucosidase inhibitory activity. Compound 3b has the highest α-glucosidase inhibitory effect with an IC50 of 80.6 μM (positive drug acarbose IC50 of 442.4 μM). In the antioxidant assays, compounds 13−15 exhibited ORAC and DPPH radical scavenging activities. The results of the AChE inhibitory assay indicated that all compounds exhibited weak AChE inhibitory activities.

the investigation of the chemical constituents and bioactivities of its n-BuOH fraction led to the identification of eight new (1a/1b, 2a, 3a, 4a/4b, and 5a/5b) and seven known (2b, 3b, and 6−10) asarone-derived phenylpropanoids, a known asarone-derived lignan (12), and four known lignan analogues (11 and 13−15) (Figure 1). Compounds 1a/1b are the first cases of asarone derivatives with an isopropyl C-3 side-chain tethered to a benzene from nature. Considering the biological activities of asarone and its analogues, the α-glucosidase inhibitory, antioxidant, and acetylcholinesterase (AChE) inhibitory effects of 1−15 were assessed. Details of the isolation, structure identification, and biological effects of 1− 15 are reported in this paper.

Acorus tatarinowii Schott is an Acoraceae perennial herb growing in wetlands and is widely distributed in eastern and southern Asia.1 Its rhizome is a famous traditional Chinese medicine, and it was recorded as the top-grade medicine in the most ancient Chinese materia medica book, “Shen Nong’s Herbal Classic”.2 In China, a water-dipped solution was historically used for curing forgetfulness, dementia, and apoplexy.3 Modern pharmacological studies demonstrated that its extract showed a variety of biological activities such as hypoglycemic, antioxidant, anti-Alzheimer’s disease (AD), antimicrobial, anticonvulsive, and antiepileptic properties.3−8 Its chemical constituents mainly contain phenylpropanoids, lignans, terpenoids, and nitrogen-containing compounds.9−11 Among them, asarone-derived phenylpropanoids, represented by α-asarone and β-asarone, are the major and characteristic constituents of A. tatarinowii and exhibit a variety of biological activities,12−16 including hypoglycemic,12,15 antioxidant,13,16 anti-AD,14−16 and antimicrobial15,16 effects. Asarone-derived phenylpropanoids, characterized by the presence of a 1,2,4trimethoxybenzene moiety, are rare in nature and are mainly distributed in the Acoraceae, Aristolochiaceae, and Lauraceae families.17 The chemical constituents of the EtOAc fraction of the rhizome of A. tatarinowii were investigated by us.2,3,18 Herein, © 2017 American Chemical Society and American Society of Pharmacognosy

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RESULTS AND DISCUSSION Compound 1 was obtained as a yellow oil. The sodium adduct ion at m/z 293.1364 [M + Na]+ by HRESIMS demonstrated that the molecular formula of 1 was C14H22O5 (four indices of hydrogen deficiency). The 13C NMR spectrum of 1 showed 14 carbons. Based on data from the DEPT-135 experiment, these carbons could be classified into six aromatic or olefinic carbons Received: May 27, 2017 Published: November 8, 2017 2923

DOI: 10.1021/acs.jnatprod.7b00457 J. Nat. Prod. 2017, 80, 2923−2929

Journal of Natural Products

Article

Figure 1. Chemical structures of compounds 1−15.

Table 1. NMR Data of 1−5 (δ in ppm, J in Hz) 1a no.

δC

1 2 3 4 5 6 7 8 9 2-OCH3 4-OCH3 5-OCH3 7-OCH3 7-OCH3 7-OCH2CH3 7-OCH2CH3

123.1 151.3 98.0 147.8 143.0 112.7 107.9 35.4 15.6 56.7 56.0 56.6 54.6 54.0

2b δHc

6.51, s

6.78, 4.44, 3.41 1.21, 3.80, 3.87, 3.83, 3.35, 3.27,

s d (6.3) d (7.1) s s s s s

δC 118.9 152.0 98.2 148.8 142.8 111.4 81.4 69.7 19.1 56.4 55.7 56.2 56.2

3b δ Hc

6.68, s

6.75, 4.25, 3.65 0.82, 3.76, 3.78, 3.67, 3.08,

s d (7.1) d (6.4) s s s s

δC 118.6 151.8 98.2 148.6 142.7 112.0 80.7 68.3 18.0 56.3 55.7 56.3 56.5

4b δ Hc

6.66, s

6.81, 4.40, 3.72 0.94, 3.67, 3.78, 3.75, 3.14,

s d (4.1) d (6.5) s s s s

δC 119.7 151.8 98.2 148.7 142.8 111.6 79.2 69.6 19.1 56.4 55.7 56.2

63.5 15.3

5b δHc

6.66, s

6.78, 4.35, 3.64 0.84, 3.75, 3.78, 3.67,

s d (6.6) d (5.7) s s s

3.26, q (6.8) 1.07, t (6.8)

δC 119.5 151.6 98.2 148.6 142.7 112.1 78.6 68.4 18.2 56.3 55.7 56.3

63.8 15.3

δ Hc

6.65, s

6.85, 4.49, 3.71 0.96, 3.75, 3.78, 3.68,

s d (4.0) d (5.7) s s s

3.30, q (6.6) 1.09, t (6.6)

a c

Measured in CDCl3 (1H NMR for 400 MHz, 13C NMR for 100 MHz). bMeasured in DMSO-d6 (1H NMR for 400 MHz, 13C NMR for 100 MHz). The indiscernible signals due to overlap or having a complex multiplicity are reported without designating multiplicity.

Figure 2. Key 2D NMR correlations of compounds 1 and 2.

(including two sp2 methine carbons), two sp3 methine carbons (including a dioxygenated carbon), and six methyl carbons (including five O-methyl carbons). The 1H NMR spectrum showed that 1 had two aromatic or olefinic protons, two sp3 methine protons, and six sets of methyl protons (including five O-methyl) (Table 1). The attached proton signals were correlated with carbon atoms in the HSQC experiment. The analysis of the 1H−1H COSY experiment and the coupling values of the protons showed the presence of the spin system of H-7−H-8−H3-9 (Figure 2A). Combined with this spin system,

indices of hydrogen deficiency, and molecular formula, the HMBC correlations shown in Figure 2A deduced the 2D structure of 1, and the assignments of all proton and carbon signals are collated in Table 1. The specific rotation of 1 was diminutive. Thus, chiral-phase HPLC analysis and resolution of rac-1 were carried out on an Enantiocol OJ-3 chiral-phase liquid chromatography column using an isocratic elution of CH3CN−H2O (18:82, v/v, 1 mL/ min). The peaks of the enantiomers of 1 (1a and 1b) were observed at tR 23.1 (1a)/24.5 (1b) min, respectively, and their 2924



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Figure 3. Chiral-phase HPLC analytical chromatograms of compounds 1−5.

Figure 4. (A) Structures of (1′R) and (1′S); (B) experimental ECD spectra of 1a/1b and calculated ECD spectra of (1′R) and (1′S) (UV correction = −6 nm, bandwidth σ = 0.3 eV).

methine carbons, and five methyl carbons (including four Omethyl carbons). The 1H NMR spectrum indicated that 2 had two aromatic or olefinic protons, two sp3 methine protons, and five sets of methyl protons (including four O-methyl) (Table 1). Analysis of the 1H−1H COSY experiment and the coupling values of the protons showed the presence of the spin system of H-7−H-8−H3-9 (Figure 2B-1). Combined with this spin system, indices of hydrogen deficiency, and molecular formula, the HMBC correlations shown in Figure 2B-1 established the 2D structure of 2. The assignments of the proton and carbon signals are collated in Table 1. Considering the coupling values (J7,8 = 7.1 Hz), the NOESY correlations (Figures 2B-2/3) indicated that the relative configuration of 2 was (7R*, 8R*).19 Compound 3 was obtained as a yellow oil. The sodium adduct ion at m/z 279.1212 [M + Na]+ by HRESIMS showed that the molecular formula of 3 was also C13H20O5 (four indices of hydrogen deficiency). The NMR data of 3 (Table S3, Supporting Information) confirmed its 2D structure. Compounds 2 and 3 had the same 2D structures, but different NMR data, which indicated that 2 and 3 were a pair of diastereoisomers. The presence of two stereogenic carbons in

relative peak area ratio (49.8 for 1a and 50.2% for 1b) in the HPLC chromatogram was approximately 1:1 (Figure 3A). The absolute configurations of 1a/1b at C-8 were established by comparing the experimental and calculated electronic circular dichroism (ECD) spectra. The simplified structures (1′R) and (1′S) (Figure 4A) were used for ECD calculations. The experimental ECD curves of 1a and 1b were similar to the computed ECD curves of (1′R) and (1′S), respectively (Figure 4B), indicating that the absolute configurations of C-8 in 1a/1b were R and S, respectively. Thus, the structures of 1a and 1b were established as (R)-1-(1,1-dimethoxypropan-2-yl)-2,4,5trimethoxybenzene [(−)-R-isoacorphenylpropanoid] and (S)1-(1,1-dimethoxypropan-2-yl)-2,4,5-trimethoxybenzene [(+)-Sisoacorphenylpropanoid], respectively. Compound 2 was obtained as a yellow oil. The sodium adduct ion at m/z 279.1212 [M + Na]+ by HRESIMS displayed that the molecular formula of 2 was C13H20O5 (four indices of hydrogen deficiency). The 13C NMR spectrum of 2 showed 13 carbons. Based on data from the DEPT-135 experiment, these carbons could be classified as six aromatic or olefinic carbons (including two sp2 methine carbons), two oxygenated sp3 2925



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Figure 5. Δδ (δS − δR) values (in ppm) of key protons in pyridine-d5 obtained for (S)- and (R)-MTPA esters (2a, 3a, 4a, and 5a).

Figure 6. Putative biosynthesis pathway of 1.

Hz) demonstrated that the relative configuration of 4 was (7R*, 8R*). Compound 5 was obtained as a yellow oil. The sodium adduct ion at m/z 293.1372 [M + Na]+ by HRESIMS showed that the molecular formula of 5 was also C14H22O5 (four indices of hydrogen deficiency). The comprehensive analysis of the NMR data of 5 (Table S5, Supporting Information) confirmed its 2D structure. Compounds 4 and 5 had the same 2D structures, but different NMR data, which indicated that 4 and 5 were a pair of epimers and that the relative configuration of 5 was (7S*, 8R*). The specific rotations of 4 and 5 were diminutive. Thus, chiral-phase HPLC analysis and resolution of 4 and 5 were carried out on the Enantiocol OZ-3 chiral-phase liquid chromatography column using an isocratic elution of CH3CN−H2O (4, 23:77, v/v, 1 mL/min; 5, 17:83, v/v, 1 mL/min), respectively (Figure 3C/D). The peaks of the enantiomers of 4 (4a and 4b) and 5 (5a and 5b) were recorded at tR 19.4 (4a)/20.7 (4b) and 38.3 (5a)/40.9 (5b) min, respectively, and the relative peak area ratios of 4a/4b and 5a/ 5b (50.6% for 4a and 49.4% for 4b; 50.5% for 5a and 49.5% for 5b) in the HPLC chromatograms were approximately 1:1, respectively (Figure 3C/D). The absolute configurations of 4a and 5a at C-8 were established as R using the modified Mosher’s method (Figure 5C/D), which showed that the absolute configurations of 4a/ 4b and 5a/5b were (7R, 8R)/(7S, 8S) and (7S, 8R)/(7R, 8S), respectively. Therefore, the structures of 4a/4b and 5a/5b were established as (7R,8R)-7-ethoxy-8-hydroxydihydroasarone (entacoraminol C)/(7S,8S)-7-ethoxy-8-hydroxydihydroasarone (acoraminol C) and (7S,8R)-7-ethoxy-8-hydroxydihydroasarone (ent-acoraminol D)/(7R,8S)-7-ethoxy-8hydroxydihydroasarone (acoraminol D), respectively. The known compounds (7R*,8R*)-7,8-dihydroxydihydroasarone (6, CAS: 137361-00-3),21 (7S*,8R*)-7,8-dihydroxydihy-

the structures of 2 and 3 demonstrated that 2 and 3 were a pair of epimers, which showed that the relative configuration of 3 was (7S*, 8R*). The specific rotations of 2 and 3 were diminutive. Thus, chiral-phase HPLC analysis and resolution of 2/3 were done on the Enantiocol OZ-3 chiral-phase LC column using an isocratic elution of CH3CN−H2O (2/3, 11:89, v/v, 1 mL/min) (Figure 3B). The peaks of the enantiomers of 2 (2a and 2b) and 3 (3a and 3b) were recorded at tR 33.2 (2a)/38.1 (2b) and 45.7 (3a)/49.4 (3b) min, respectively, and the relative peak area ratios of 2a/2b and 3a/3b (17.1% for 2a and 17.0% for 2b; 33.0% for 3a and 32.9% for 3b) in the HPLC chromatogram were approximately 1:1, respectively (Figure 3B). The absolute configurations of 2a and 3a at C-8 were established using the modified Mosher’s method.20 According to the Δδ values of the (S)-MTPA and (R)-MTPA esters [(2aS) and (2aR); (3aS) and (3aR)] in pyridine-d5, the absolute configurations of 2a and 3a at C-8 were established as R (Figure 5A/B), which indicated that the absolute configurations of 2a/2b and 3a/3b were (7R, 8R)/(7S, 8S) and (7S, 8R)/(7R, 8S), respectively. Therefore, the structures of the two known compounds, 2b and 3b, were established as acoraminol A and acoraminol B, respectively, and the structures of 2a and 3a were defined as (7R,8R)-7-methoxy-8-hydroxydihydroasarone (ent-acoraminol A) and (7S,8R)-7-methoxy-8hydroxydihydroasarone (ent-acoraminol B), respectively. Compound 4 was obtained as a yellow oil. The sodium adduct ion at m/z 293.1372 [M + Na]+ by HRESIMS demonstrated that the molecular formula of 4 was C14H22O5 (four indices of hydrogen deficiency). The comprehensive analysis of the NMR data of 4 (Table S4, Supporting Information) confirmed its 2D structure. The NOESY correlations between H-8 and H-6, between H-6 and H3-9, and between H3-9 and H-7 and the coupling constant (J7,8 = 6.6 2926



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Figure 7. Hypoglycemic activities of compounds 1−15, α-asarone, and β-asarone were evaluated by an α-glucosidase inhibitory assay with acarbose as the positive control. Each value was expressed as mean ± SD, n = 3.

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droasarone (7, CAS: 137361-02-5),21 1-hydroxy-1-(2,4,5trimethoxyphenyl)propan-2-one (8, CAS: 1422453-77-7),22 1(2,4,5-trimethoxyphenyl)ethanone (9, CAS: 1818-28-6),23 asaraldehyde (10, CAS: 4460-86-0),24 3-(3,4dimethoxyphenyl)propan-1-ol (11, CAS: 3929-47-3), 25 (±)-magnosalicin (12, CAS: 93376-03-5),26 (±)-pinoresinol (13, CAS: 4263-88-1),27 (7S,8R)-9-O-β-D-glucopyranosyldihydrodehydrodiconiferyl alcohol (14, CAS: 126371-34-4),28 and (7S,8R)-9′-O-β- D -glucopyranosyldihydrodehydrodiconiferyl alcohol (15, CAS: 220745-70-0)29 were identified via comprehensive spectroscopic analyses together with the comparison of their reported spectroscopic data. Asarone-derived phenylpropanoids, a rare and unusual class of phenylpropanoids characterized by the presence of a 1,2,4trimethoxybenzene moiety, are primarily distributed in the Acoraceae, Aristolochiaceae, and Lauraceae families.17 Notably, compounds 1a/1b are the first cases of asarone-derived phenylpropanoids with an isopropyl C-3 side-chain tethered to a benzene core from nature, and a putative biosynthesis pathway of 1a/1b is proposed (Figure 6). The hypoglycemic activities of compounds 1−15, α-asarone, β-asarone, and acarbose (positive control) were evaluated via an α-glucosidase inhibitory assay. All of these compounds except 3a exhibited stronger α-glucosidase inhibitory activity than acarbose. Notably, 3b exhibited considerable α-glucosidase inhibitory activity, with an IC50 value of 80.6 μM (acarbose IC50 of 442.4 μM) (Figure 7A/B). The antioxidant activities of compounds 1−15, α-asarone, βasarone, and EGCG (positive control) were evaluated via the oxygen radical absorbance capacity (ORAC) assay. Only compounds 13−15 showed ORAC activities, and compound 15 exhibited potent antioxidant capacity with an ORAC value of 3.17 μM TE/μM (EGCG 1.97 μM TE/μM) (Table S8, Supporting Information). The antioxidant activities of compounds 1−15, α-asarone, β-asarone, and vitamin C (positive control) were also assessed via the DPPH radical scavenging assay. Only compounds 13−15 showed moderate DPPH radical scavenging activity (Table S9, Supporting Information). The above results demonstrated that the phenolic group was crucial for the antioxidant activities of phenylpropanoids and lignans in the ORAC and DPPH radical scavenging models. The AChE inhibitory effects of compounds 1−15, α-asarone, β-asarone, and huperzine A (positive control) were evaluated using the Ellman method. The compounds exhibited weak AChE inhibitory activity (Table S10, Supporting Information).

EXPERIMENTAL SECTION

General Experimental Procedures. For the specific details regarding instruments used, see the “General experimental procedures” section in the Supporting Information. Plant Material. The dried rhizome of A. tatarinowii was purchased from PuraPharm Corporation in 2011. The plant materials were authenticated by Jia-Fu Wei (the pharmacist of Guangxi Institute for Food and Drug Control). A voucher specimen (20110301) was deposited in the Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, Guangzhou, People’s Republic of China. Extraction and Isolation. The air-dried and powdered rhizome of A. tatarinowii (19.5 kg) was extracted twice with 80 L of 60% EtOH− H2O for 2 h each time to yield an extract (2.2 kg). The extract was suspended in H2O (10 L); then the suspension was partitioned with EtOAc and n-BuOH, respectively. After evaporation of the n-BuOH phase under vacuum, the n-BuOH fraction (102.3 g) was dissolved in 20% EtOH−H2O and passed through an AB-8 macroporous resin column (8.0 × 80.0 cm) using a stepwise elution of EtOH−H2O (20:80, 40:60, 60:40, 95:5, v/v), to yield fractions F1−F4. F2 (9.8 g) was subjected to MPLC on ODS CC (2.7 × 25.4 cm) using a stepwise elution of MeOH−H2O (15:85, 35:65, 55:45, 100:0, v/v) to yield fractions 2.1−2.6. Fraction 2.6 (1.6 g) was isolated using preparative HPLC (25% CH3CN−H2O, 8 mL/min) to give 4 (tR: 33.9 min, 10.7 mg) and 5 (tR: 47.3 min, 9.0 mg). Fraction 2.4 (1.5 g) was separated using preparative HPLC (20% CH3CN−H2O, 8 mL/min) to afford 2/ 3 (tR: 6.3 min, 36.7 mg), 14 (tR: 39.7 min, 17.6 mg), and 15 (tR: 44.1 min, 14.5 mg). Compounds 2/3 from fraction 2.4 were separated using the Phenomenex Lux Cellulose-2 chiral-phase column (13% CH3CN−H2O, 1 mL/min) to yield 2 (tR: 24.6 min, 7.6 mg) and 3 (tR: 29.7 min, 12.0 mg). Fraction 2.2 (340.0 mg) was separated using semipreparative HPLC (18% CH3CN−H2O, 3 mL/min) to yield 6 (tR: 31.0 min, 24.2 mg), 7 (tR: 35.7 min, 28.1 mg), 8 (tR: 57.0 min, 5.6 mg), and 11 (tR: 65.7 min, 4.9 mg). F3 (3.1 g) was subjected to MPLC on ODS CC (2.7 × 25.4 cm2) using a stepwise elution of MeOH− H2O (20:80, 40:60, 60:40, 100:0, v/v) to yield fractions 3.1−3.4. Fraction 3.2 (390.2 mg) was separated using semipreparative HPLC (35% CH3CN−H2O, 3 mL/min) to give 1 (tR: 30.5 min, 16.2 mg). Fraction 3.2 (690.5 mg) was separated using semipreparative HPLC (33% CH3CN−H2O, 3 mL/min) to afford 9 (tR: 26.2 min, 22.3 mg), 10 (tR: 30.5 min, 8.3 mg), 12 (tR: 37.7 min, 4.8 mg), and 13 (tR: 46.5 min, 7.2 mg). (−)-R-Isoacorphenylpropanoid (1a)/(+)-S-Isoacorphenylpropanoid (1b): yellow oil; 1a/1b, [α]32 D −79.2/+70.0 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 205 (3.88), 229 (3.40), 289 (3.20) nm; 1a, ECD (c 2.8 × 10−4 M, MeOH) λmax (Δε) 197 (−13.03), 209 (+0.41), 228 (−1.50) nm; 1b, ECD (c 2.8 × 10−4 M, MeOH) λmax (Δε) 197 (+15.81), 209 (−0.49), 228 (+1.82) nm; IR (KBr) νmax 2932, 2834, 1608, 1511, 1458, 1210, 1035 cm−1; ESIMS (positive) m/z 293.1 [M + Na]+; HRESIMS (positive) m/z 293.1364 [M + Na]+ (calcd for C14H22O5Na, 293.1365); 1D NMR see Table 1. ent-Acoraminol A (2a)/Acoraminol A (2b): yellow oil; 2a/2b, [α]32 D +83.4/−75.8 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 205 (3.84), 232 (3.32), 291 (3.13) nm; IR (KBr) νmax 3450, 2958, 2937, 2927



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1608, 1509, 1461, 1210, 1077 cm−1; ESIMS (positive) m/z 279.1 [M + Na]+; HRESIMS (positive) m/z 279.1212 [M + Na]+ (calcd for C13H20O5Na, 279.1208); 1D NMR see Table 1. ent-Acoraminol B (3a)/Acoraminol B (3b): yellow oil; 3a/3b, [α]32 D +90.0/−85.4 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 205 (3.80), 232 (3.33), 291 (3.16) nm; IR (KBr) νmax 3447, 2960, 2937, 1614, 1509, 1461, 1210, 1032 cm−1; ESIMS (positive) m/z 279.1 [M + Na]+; HRESIMS (positive) m/z 279.1212 [M + Na]+ (calcd for C13H20O5Na, 279.1208); 1D NMR see Table 1. ent-Acoraminol C (4a)/Acoraminol C (4b): yellow oil; 4a/4b, [α]32 D +72.0/−71.8 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 205 (3.91), 231 (3.35), 290 (3.15) nm; IR (KBr) νmax 3426, 2960, 2937, 1687, 1509, 1460, 1210, 1031 cm−1; ESIMS (positive) m/z 293.1 [M + Na]+; HRESIMS (positive) m/z 293.1372 [M + Na]+ (calcd for C14H22O5Na, 293.1365); 1D NMR see Table 1. ent-Acoraminol D (5a)/Acoraminol D (5b): yellow oil; 5a/5b, [α]32 D +90.2/−92.4 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 205 (3.83), 231 (3.23), 290 (3.04) nm; IR (KBr) νmax 3433, 2958, 2932, 1690, 1514, 1458, 1208, 1023 cm−1; ESIMS (positive) m/z 293.1 [M + Na]+; HRESIMS (positive) m/z 293.1372 [M + Na]+ (calcd. for C14H22O5Na, 293.1365); 1D NMR see Table 1. Preparation of (S)-MTPA and (R)-MTPA Esters of 2a, 3a, 4a, and 5a. A solution of 2a (0.5 mg) in pyridine-d5 (0.5 mL) was added to (R)-MTPA chloride (15 μL) in an NMR tube under a nitrogen atmosphere. The solution was churned at room temperature for 12 h to yield the (S)-MTPA ester (2aS). The (R)-MTPA ester (2aR) was prepared with (S)-MTPA chloride in the same manner. The same method was used to prepare the (S)- and (R)-MTPA esters of 3a, 4a, and 5a. α-Glucosidase Inhibitory Assay. The α-glucosidase inhibitory effect was assessed using a previously described method with slight modifications.30,31 For the experimental details about the αglucosidase inhibitory assay, see the related section in the Supporting Information. Oxygen Radical Absorbance Capacity Assay. The ORAC assay was carried out the same as with the Zhang method.32 For the experimental details of this assay, see the related section in the Supporting Information. DPPH Radical Scavenging Assay. The DPPH radical scavenging activity was evaluated on the basis of the method of Xie with slight modifications.33 For the experimental details about the DPPH radical scavenging assay, see the related section in the Supporting Information. AChE Inhibitory Assay. The AChE inhibitory activity was assessed based on the method of Ellman with minor modifications.34,35 For the experimental details about AChE inhibitory assay, see the related section in the Supporting Information. Statistical Analysis. For the details of statistical analysis, see the related section in the Supporting Information.

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ORCID

Xin-Sheng Yao: 0000-0003-1603-4873 Hao Gao: 0000-0003-1178-0121 Author Contributions §

E. Gao and Z.-Q. Zhou contributed equally to this work.

Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This study was financially supported by the National Natural Science Foundation of China (81422054 and 3171101305), the Guangdong Natural Science Funds for Distinguished Young Scholar (S2013050014287), Guangdong Special Support Program (2016TX03R280), Guangdong Province Universities and Colleges Pearl River Scholar Funded Scheme (Hao Gao, 2014), K. C. Wong Education Foundation (Hao Gao, 2016), and Guangdong Provincial Science and Technology Project (2016B090921005). We appreciate the identification of the plant material by Jia-Fu Wei, the pharmacist of Guangxi Institute for Food and Drug Control, Guangxi, China.

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ASSOCIATED CONTENT

* Supporting Information S

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jnatprod.7b00457. General experimental procedures, NMR assignments of 1−5, quantum chemical ECD calculations of (1′R) and (1′S), the experimental details and results of the αglucosidase inhibitory, ORAC, DPPH radical scavenging, and AChE inhibitory assays of 1−15, and the 1D and 2D NMR spectra of 1−5 (PDF)

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REFERENCES

AUTHOR INFORMATION

Corresponding Author

*Phone/fax: +86-20-85228369. E-mail: [email protected] (H. Gao). 2928



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