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Diacylglycerol Acyltransferase-inhibitory Prenylated Flavonoids from Maackia amurensis Rupr
Diacylglycerol Acyltransferase-inhibitory Prenylated Flavonoids from Maackia amurensis Rupr
Bulletin of the Korean Chemical Society. 2014. Oct, 35(10): 3092-3094
Copyright © 2014, Korea Chemical Society
  • Received : April 23, 2014
  • Accepted : June 10, 2014
  • Published : October 20, 2014
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About the Authors
Na Li
College of Pharmacy, Beihua University, Jilin Province 132013, China.
Hyun Sun Lee
Targeted Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Chungbuk 363-883, Korea
Zhen Dong Tuo
College of Pharmacy, Beihua University, Jilin Province 132013, China.
Jia Lin Li
College of Pharmacy, Beihua University, Jilin Province 132013, China.
Pei Ge Du
College of Pharmacy, Beihua University, Jilin Province 132013, China.
Long Cui
College of Pharmacy, Beihua University, Jilin Province 132013, China.

Abstract
Keywords
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Experimental Section
General Experimental Procedures. UV spectra were taken in MeOH using a Shimadzu spectrophotometer. Nuclear magnetic resonance (NMR) spectra were obtained from a Varian Unity Inova 400 MHz spectrometer using acetone- d 6 as solvents, with TMS as the internal standard. All accurate mass experiments were performed on a Micromass QTOF2 (Micromass, UK) mass spectrometer. Column chromatography was conducted using silica gel 60, Sephadex LH-20 and RP-18 For thin-layer chromatography, precoated TLC silica gel 60 F 254 plates from Merck were used. HPLC runs were carried out using a Shimadzu System LC-10AD pump equipped with a model SPD-10Avp UV detector, and an Optima Pak ® C 18 column (10 × 250 mm, 10 m particle size, RS Tech Korea).
Bovine serum albumin and Trizma-base were obtained from Sigma Chemical Co (ST. Louis, MO, USA). [1- 14 C] oleoyl-CoA (250 μCi) was purchased from Amersham. For the cell culture, Dulbecco’s modified eagle medium (DMEM), ʟ-glutamine, kanamycin sulfate, and fetal bovine serum (FBS) were purchased from GIBCO-BRL (Gaithersburg, MD, USA).
Plant Material. The stem bark of M. amurensis (3.0 kg) was collected in Yanji, Jilin province, China, in May 2009. A voucher specimen of the plant (No. 2009510) was deposited at the College of Pharmacy, Beihua University, Jilin, China.
Extraction and Isolation . The stem bark (3.0 kg) of M. amurensis was extracted with MeOH at room temperature for 2 weeks and the solution was concentrated to obtain a crude extract (165.0 g, 66% inhibition at 30 μg/mL). This extract was suspended in H 2 O, partitioned successively with CHCl 3 . EtOAc and BuOH, and then the organic solvents were removed. A portion of the EtOAc-soluble fraction (10.0 g, 74% inhibition at 30 μg/mL) was chromatographed over a silica gel column using a gradient of CHCl 3 –MeOH (from 20:1, 19:1 to 0:1), and was separated into 10 fractions (Fr.1–Fr.10). Fr.5 (CHCl 3 –MeOH 10:1, 1.0 g, 77% inhibition at 30 μg/mL) was chromatographed over Silica gel, eluted with a stepwise gradient of CH 2 Cl 2 /MeOH (from 100:1, 80:1, 50:1 to 0:1) to afford 10 subfractions (Fr.5A–Fr.5J). Purification of Fr.5D (110.0 mg) by semipreparative HPLC using an gradient solvent system of 70-80% MeCN in H 2 O over 60 min to yield compounds 2 (4.4 mg) and 7 (6.4 mg). Fr.5F (123.0 mg) was purified by preparative HPLC using an isocratic solvent system of 50% MeCN in H 2 O over 30 min followed by 60% MeCN in H 2 O over 70 min to obtain compounds 5 (5.9 mg), 6 (5.1 mg), and 3 (3.1 mg). Fr.6 (84% inhibition at 30 μg/mL) was subjected to an RP-18 column and was eluted with MeOH-H 2 O (1:10, 1:2, to 10:1) to yield six fractions (F.6A-F.6F). The most active fraction, Fr.6C (171.0 mg), was further separated by a Silica gel column eluted with CHCl 3 -MeOH (40:1, 35:1, to 10:1) to yield 10 subfractions (Fr.6C1-Fr.6C10). Fr.6C8 was separated by HPLC, using a gradient of 20-30% MeCN in H 2 O as the mobile phase to produce compounds 1 (3.1 mg) and 4 (4.9 mg).
Compound 1: White amorphous powder;
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−28.10 ( c 0.1, MeOH); UV (MeOH): λ max (log ε) 224 (4.29), 289 (4.21), 326 (3.43) nm; CD ( c 0.55 MeOH): [ θ ] 329 +3.25, [ θ ] 301 −5.30, [ θ ] 215 +15.74; 1 H NMR (400 MHz, acetone- d 6 ) and 13 C NMR (100 MHz, acetone- d 6 ) spectral data see Table 1 ; HR-EI-MS m/z : 368.1261 [M] + (calcd for C 21 H 20 O 6 , 368.1260).
1H (400 MHz) and13C NMR (100 MHz) spectroscopic data of compound1(acetone-d6,δ, ppm,J/Hz)
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aChemical shifts in ppm relative to TMS; coupling constants (J) in Hz.
DGAT1 Assay. Preparation of microsomes from rat liver and measurement of the in vitro DGAT1 activity was measured as reported previously. 12
Acknowledgements
This research was supported in part by the grants from the Project Sponsored by the State Education Ministry and Key Laboratory of Natural Resources of Changbai Mountain & Functional Molecules (Yanbian University), Ministry of Education, China (No. 201305).Supporting Information.The UV, CD and NMR spectral data of compound1are available as Supporting Information.
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