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Structures, Antioxidant and Intestinal Disaccharidase Inhibitory Activities of A-type Proanthocyanidins from Peanut Skin Huiwen Zhang, Rigui Ye, Yumei Yang, and Chaomei Ma J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/jf402518k • Publication Date (Web): 26 Aug 2013 Downloaded from http://pubs.acs.org on August 31, 2013

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Journal of Agricultural and Food Chemistry

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Structures, Antioxidant and Intestinal Disaccharidase Inhibitory Activities of

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A-type Proanthocyanidins from Peanut Skin

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Huiwen Zhang,†,‡ Rigui Ye,† Yumei Yang,‡ and Chaomei Ma*,†

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†

School of Life Sciences, Inner Mongolian University, Huhhot, China ‡

Baotou Medical College, Baotou, Inner Mongolia, China

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Corresponding Author

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*Telephone: +86 471 4992435. Fax: +86 471 4992435. E-mail: [email protected]

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1

ACS Paragon Plus Environment


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ABSTRACT: Nine compounds including a new A-type proanthocyanidin trimer,

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epicatechin-(2β→O→7,4β→8)-[catechin-(6→4β)]-epicatechin (8), and a known

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trimer,

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reported for peanut skin for the first time, were isolated and purified. Their structures

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were determined by spectroscopic methods and by degradation reactions with

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L-cysteine in acidic conditions. The DPPH radical scavenging activity and the

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inhibitory activity on maltase and sucrase of the isolated compounds were

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investigated. All compounds showed strong DPPH scavenging activities (EC50 < 20

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µg/ml). Compound 8 showed the strongest inhibitory activity on maltase with an

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IC50 value of 0.088 mg/ml, while compound 9 exhibited the strongest inhibition on

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sucrase with an IC50 value of 0.091 mg/ml.

epicatechin-(4β→8)-epicatechin-(2β→O→7,4β→8)-catechin

(9),

being

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Keywords: peanut skin, condensed tannin, proanthocyanidin trimer, inhibition on

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intestinal disaccharidases

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INTRODUCTION Peanut (Arachis hypogaea L. Fabaceae), the fourth oleaginous plant in the world, is

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known to be one of the most important economical crop owing to its wide distribution,

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nutritional characteristics and its great application in the food industry for centuries.

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China, being the largest producer and exporter of peanut, has a production of

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14,385,000 tons every year and almost 60% of the peanut is consumed as feedstock

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for oil, which is one of the main vegetal oils in Chinese daily life. However, over the

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years, the skin of peanut is generally neglected and considered as useless byproduct

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and, for the yearly generated peanut skin, only small part of it is used in traditional

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Chinese medicine (TCM) to treat illness such as chronic hemorrhage and

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bronchitis,etc.1 Early investigations have shown that the most important chemical

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constituents of peanut skin are proanthocyanidins, particularly the A-type, in which

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the subunits are connected by a 4→8 or 4→6 carbon bond and a 2→O→7 ether

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bond.2-3

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Proanthocyanidins in general have shown notable potentials of health benefits, e.g.,

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antioxidant4-5 and insulin action potentiation6 activities. Moreover, consumption of

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plant extract containing A-type proanthocyanidins has been shown to increase sugar

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tolerance in type II diabetes patients.7 Disaccharidase (or α-glucosidase) inhibitors

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could control postprandial hyperglycemia by delaying the absorption of intestinal

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carbohydrates as does the prototype clinically used anti-diabetic α-glucosidase

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inhibitor drug, acarbose.8

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In our further systematic search for proanthocyanidins, 9 compounds were isolated

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from the water-soluble fraction of peanut skin, including a novel trimer (compound 8)

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and another trimer (compound 9) which is being reported from peanut skin for the

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first time. Bioassays were carried out to test the antioxidant and intestinal

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disaccharidase inhibitory activities of the 9 compounds from peanut skin. This paper

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describes the isolation, structure elucidation, antioxidant activity and intestinal

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disaccharidase inhibitory activity of these proanthocyanidins to provide scientifically

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valuable information for better utilization of peanut skin.

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MATERIALS AND METHODS

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Materials and Apparatus. Solvents used for the extraction and isolation were of

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analytical grade. L-Cysteine hydrochloride was pursed from Sigm-Aldrich

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Corporation (St. Louis, MO, USA). High performance liquid chromatography (HPLC)

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grade solvents used for UPLC were purchased from Fisher scientific company (New

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Jersey, USA). The sources of other materials are: octadecylsilane (ODS, 38-63µm,

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wako Pure Chemcial Industries, td., Osaka, Japan), Sephadex LH-20 (GE Healthcare

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Bio-Sciences AB, Uppsala, Sweden) and high porous polymer from Mitsubishi

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Chemical Corporation (MCI CHP 20, Mitsubishi Chemical Corporation, Tokyo,

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Japan). TLC was performed on silica gel plates (GF254, 0.2mm, Rushan Taiyang

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Desiccant co., Itd, China). NMR spectra were obtained from a Bruker Avance III 500

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spectrometer with tetramethylsilane as internal standard. UPLC-DAD-ESI-MS

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experiments were carried out on an Agilent 1290 infinity UPLC-DAD system [Agilent

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Technologies Singapore (International) Pte. Ltd., Singapore] with an auto-sampler and

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a photo-diode array detector (DAD) coupled with an Agilent 6340 triple Quad MS.

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Fast atom bombardment mass spectrometry (FAB-MS) were measured with a Xevo

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G2 Q-TOF mass spectrometer (Waters). Circular dichroism (CD) was recorded with

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JASCO-J-18 spectro-polarimeter (Waters). UV and IR. Spectra were taken using a

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Shimadzu UV 240 spectrophotometer and a Shimadzu IR spectrophotometer,

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respectively.

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Plant Materials. Peanut skin used in this research was purchased from Weikang

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drug store, Huhhot, Inner Mongolia, China and identified by the authors to be the seed

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skin of Arachis hypogaea L. Fabaceae.

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Extraction and Purification. Peanut skin (2kg) was soaked with 70% acetone

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(32L) for 24h, and extracted with ultrasonication for 30 minutes. After filtration, the

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residue was extracted once again by repeating the above procedure. The solvent was

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evaporated under reduced pressure and the ensuing extract (276 g) fractionated by

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chromatography over Sephadex LH-20 with ethanol (7500 ml) to give 8 fractions

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(E1-E8). TLC and LC-MS analysis revealed that E2 contained proanthocyanidin

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monomers, while E3 and E5 contained dimeric and trimeric proanthocyanidins

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respectively. E2 was subjected to further chromatography on ODS, eluted with a

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0-100% methanol/water gradient to give compounds 1 (122 mg) and 2 (63 mg).

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Similarly E3 (27 g) was subjected to further chromatography on ODS, eluted with a

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0-100% methanol/water gradient. The 20%-30% gradient fraction contained

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polyphenols as indicated by TLC, and was further purified by repeated

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chromatography on MCI gel (CHP20/P120) and Sephadex LH-20 with

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methanol/water gradients to afford compounds 3-6. Compound 3 (780 mg) was

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recrystallized from water as colorless needles, while 4 (56 mg), 5 (25 mg) and 6 (68

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mg) were obtained as amorphous powders. Furthermore, E5 (20 g) was purified in the

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same way as described for E3 to obtain compounds 7 (145 mg), 8 (54 mg) and 9 (72

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mg) as amorphous powders.

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Compound 1-2: amorphous powder, MS (ESI) m/z 289 ([M-H]-, 100). Compound

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3-6: colorless needles (3) or amorphous powder (4-6), MS (ESI) m/z 575 ([M-H]-,

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100), NMR see supporting material. Compound 7-9: amorphous powder, MS (ESI)

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m/z 863 ([M-H]-, 100), NMR see supporting material.

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Degradation of Proanthocyanidin Trimers. L-cysteine-induced degradations

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of the trimeric proanthocyanidins were carried out according to the reported

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methods9,10 with modification. The trimer and cysteine•HCl (1:2) were dissolved in

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methanol in a centrifuge tube. The tube was tightly sealed and heated at 65 ºC for 24 h.

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The reaction mixture was filtered though a 0.22 µm microfilter and analyzed by

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UPLC-DAD-EIS-MS in comparison with standard 3 (procyanidins A1), and standards

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catechin-cystein and epicatechin-cystein which were recently prepared in our earlier

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work.11

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DPPH Radical Scavenging Assay. We examined the DPPH radical scavenging

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activity with the method described by Ma et al.12 The antioxidant activities were

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examined at four different concentration (2.5, 5, 10 and 20 µg/ml) in 96-well plates.

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10 µl of a compound solution (in DMSO) and 190 µl of DPPH solution

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(1,1-diphenyl-2- picrylhydrazyl radical in ethanol, 0.1 mM) were added to each well.

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For the color control, 10 µl of compound solution and 190 µl of ethanol were mixed in

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each well. In the control wells, 10 µL DMSO and 190 µL DPPH were added. After 20

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min at room temperature in the dark, the absorbance (A) was measured at 520 nm, and

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the activity was calculated as follow:

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Effect %=100×[Acontrol - (Acompound -Acolor)] /Acontrol

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IC50, the concentration of a sample that scavenged 50% of DPPH radical was obtained

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from a curve of effect % versus sample concentration.

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Determination of Inhibitory Activity against Maltase and Sucrase. To

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determine the maltase and sucrase inhibitory activities of samples, we prepared the

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stocking intestinal disaccharidases as reported before.11 The assay was carried out as

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following: 3 µl of sample (dissolved by DMSO), 7µl of disaccharidase [the stocking

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enzyme was diluted 10 times by phosphate buffer (pH 7)] and 20 µl maltose solution

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(2 mg/ml) were added in each well of a 96-well plate to test the inhibition activity on

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maltase. 3 µl DMSO was added instead of samples in the control wells. After 20

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minutes of incubation at 37 ºC, 10 µl of DMSO and the glucose detecting reagents of

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an assay kit from Nanjing Jiancheng Bio Company (Nan Jing, China) were added and

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the absorbances (A) were measured at 520 nm with a plate reader. The activity was

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calculated as follows:

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Inhibition %=100 × [(Acontrol-Asample)] ⁄Acontrol

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The inhibition activity on sucrase was tested in the same procedure as above,

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except that 20 µl of sucrose instead of maltose was used as the substrate and the

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concentration of the working enzyme solution was trice of that for the maltase assay.

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RESULTS AND DISCUSSION

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Proanthocyanidins in Peanut Skin. Peanut skin was extracted with 70 % (v/v)

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acetone and the extract was fractionated with Sephadex LH-20 column

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chromatogaphy. The fraction obtained from 95% ethanol eluted part was then

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subjected to repeated chromatography on ODS and MCI eluted with aqueous

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methanol. Compounds 1-9 were obtained and their structures were determined by

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interpretation of their spectral data and by L-cysteine-induced degradations (Figure 1).

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Compounds 1 and 2 were identified as catechin and epicatechin by comparison

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with authentic standards in UPLC-DAD-ESI-MS. Compounds 3~6 were identified as

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procyanidins A1, A2, epicatechin-(2β→O→7,4β→8)-ent-epicatechin, and

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epicatechin-(2β→O→7,4β→6)-catechin by comparison of their spectral data with

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those reported in literature.3

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Structural Elucidation of Compound 7. Compound 7 [circular dichroism (CD):

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∆ε272.6 -3.93, ∆ε231 +21.57] was obtained as a white amorphous powderIt showed a

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molecular formula of C45H36O18 from the negative FAB-MS (m/z [M-1]- 863.1823,

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which revealed that 7 was a trimeric proanthocyanidin. Degradation of 7 (Scheme 1)

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produced catechin and another compound with the poseudo-molecular weight of 694

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in negative ESIMS, suggesting it is composed by an A-type procyanidins and a

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catechin. By comparison of its 1H and 13C NMR spectral data with those preciously

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reported, 10-11 compound 7 was confirmed as cinnamtannin D-1.14

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Structural Elucidation of Compound 8. Compound 8 [UV (MeOH) λmax ( log

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ε): 280 (4.13) nm; IR (ν, KBr, cm-1): 3350, 1610, 1520, 1450, 1290, 1150, 1100;

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CD:∆ε273.8 -2.58, ∆ε239.4 +16.07] was obtained as a white amorphous powder. It

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showed a molecular formula of C45H36O18 from the negative HRFAB-MS (m/z

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[M-1]- 863.1823), which revealed that 8 was a trimeric proanthocyanidin. The

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presence of three flavanyl units was also indicated by 13C resonances at δ 100.88

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(C-2), 67.49 (C-3), 29.46 (C-4), 84.21 (C-2′), 68.02 (C-3′), 28.58 (C-4′), 77.21

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(C-2″), 67.49 (C-3″), 37.06 (C-4″) (Table 1), arising from the heterocyclic rings

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(rings C, I and F). The existence of an A-type proanthocyanidin unit was supported

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by the downfielded C-2 signal at δ 100.88 (Table 1) and an isolated AB coupling

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system at δ 4.20 (br.s, H-3) and 4.35 (br.s, H-4) (C-ring) in 1H NMR spectrum. The

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two flavan-3-ol units of the A-type entity in 8 were deduced to be linked through C4

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(C ring) and C-8′ (D ring) based on the long-range correlations of H-4 (C-ring) at δ

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4.35 with C-7′ at δ 150.1, C-8′ at δ 107.33, and C-9′ at δ 154.44 in the Heteronuclear

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multiple-bond correlation (HMBC) spectrum. The long-range correlations of H-4″

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(I-ring) at δ 4.63 with C-5′ at δ 149.29 , C-6′ at δ 120.08, and C-7′ at δ 150.1 in the

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HMBC indicated the C-4″→C-6′ linkage (Figure 2). The C-4″→C-6′ linkage was

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further confirmed by its complex NMR spectra arisen from conformational isomers

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due to the steric hindrance for the free rotation of the epicatechin unit.

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The terminal flavanol unit was determined to be 2,3-cis (I ring) from the singlet

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at δ 5.33 (1H, s, H-2″) and the corresponding carbon at δ 77.21(C-2″), 4 while the

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middle unit (the doubly-substituted terminal unit ) was deduced to have a 2,3-trans

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configuration (F ring) from the proton signal at δ 4.79 (1H, d, J=6.5, H-2′) and the

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corresponding carbon at δ 84.21 (C-2′) in 13C NMR.4

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Treatment of 8 with cysteine•HCl in methanol (Scheme 2) yielded compound 3,

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which was identified to be procyanidin A1 by direct comparison of its 1H and 13C

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NMR spectral data with those preciously reported.4 The cystein conjugate derived

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from the upper flavan-3-ol unit in the degradation of 8, was identified as

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epicatechin-cystein by direct comparison by UHPLC-MS with a standard compound

9



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which had been prepared and reported in our laboratory recently11 (Figure 3). Of the three trimeric proanthocyanidins (7-9) with the same molecular weight (864),

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the 1HNMR spectrum of 8 was observed to be relatively complicated compared to

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those of 7 and 9. This relative complication is alludable to the presence of conformers

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in 8, due to hindered free rotation across its 4″→6′ linkage, which the structures of 7

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and 9 conspicuously lack.

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The diagnostic high amplitude positive cotton effect at 239 nm wavelength of the

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CD spectrum of 8 informed the β-orientation assignment of the interflavonoid

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bonds.15 Consequently, the structure of 8 was determined to be epicatechin-

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(2β→O→7,4β→8)-[catechin-(6→4β)]-epicatechin. This is a branched A-type

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proanthocyanidin with C4-C6, C4-C8 linkages to the same A ring of a flavanyl unit.

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Structural Elucidation of Compound 9. Compound 9 (CD: ∆ε277.8 -4.67, ∆ε240

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+5.39, ∆ε228.2 +2.86) was obtained as a white amorphous powder. It showed a

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molecular formula of C45H36O18 from the negative FAB-MS (m/z [M-1]- 863.1823,

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which revealed that 9 was a trimeric proanthocyanidin. Treatment of 9 with

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cysteine•HCl in methanol yielded procyanidin A1 and epicatechin-cystein (Scheme 3,

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Figure 3), the same results as for the degradation of compound 8. The result indicated

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that compound 9 also composed of proanthocynidin A1 and epicatechin with

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epicatechin being its upper flavan-3-ol unit. It was thus deduced that compounds 8

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and 9 possess different linking positions between procyanidin A1 and epicatechin. The

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epicatechin was determined to be linked through C4→C8 with proanthocynidin A1,

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by direct comparison of the 1H and 13C NMR spectral data of 9 with references.16-17

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The high amplitude positive Cotton effect at a wavelength of 240 nm in the CD

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spectrum of 9 confirmed its β-orientated interflavonoid bond. Compound 9 was thus

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identified as epicatechin-(4β→8)-epicatechin-(2β→O→7,4β→8)-catechin, which is

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reported from peanut skin for the first time.

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Antioxidant activity. As shown in Table 2, all the isolated compounds, 1−9

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demonstrated strong antioxidant activities. They scavenged 50% of the DPPH radical

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at a concentration less than or near to10 µg/ml. The dimeric and trimeric

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proanthocyanidins showed radical scavenging activity with the potencies similar to

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those of the monomers, catechin and epicatechin, which are well known potent

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anti-oxidants.

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Inhibitory Activity Against Maltase and Sucrase. As shown in Table 3, five

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compounds showed maltase inhibitory activity (>50% inhibition), and four

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compounds showed sucrase inhibitory activity at concentrations of 1 mg/ml.

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Compound 8, epicatechin-(2β→O→7,4β→8)-[catechin-(6→4β)]-epicatechin, showed

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the strongest inhibition against maltase, with an IC50 value of 0.088 mg/ml. The

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lowest IC50 value (0.091 mg/ml) observed in the sucrase inhibition experiment was

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exhibited by 9, epicatechin-(4β→8)-epicatechin-(2β→O→7,4β→8)-catechin, making

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it the strongest sucrase inhibitor of all the samples. It can be deduced from the results

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that 8 and 9 have notable degrees of specificity for maltase and sucrase respectively.

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As the component units of 8 and 9 are essentially the same, the variation in their

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biological activities as by their observed differing enzyme specificities is alludable to

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their different unit-connectivity, impacting chirality (though of same configuration) at

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different positions. All the 3 trimeric proanthocyanidins showed inhibitory activities

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on maltase and sucrase. Their inhibitory activities were greater than those of their

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component dimeric proanthocyanidins. Further investigations are required to elucidate

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the detail structure-activity relationships of A-type proanthocyanidins on intestinal

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disaccharidases.

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Degradation of the trimeric proanthocyanidins was carried out with a mild reagent,

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L-cysteine, which also is relatively safe compared to the conventionally used thiolytic

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degradation reagent, benzyl mercaptan with a strong irritating smell. LC-MS analysis

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of the products of L-cysteine-induced degradation clearly indicated the component

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units and their linking order in the structures of pranthocyanidin polymers. Some

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L-cysteine conjugates of catechin/epicatechin and low-molecular-weight B-type

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proanthocyanidins have been reported to have antioxidant,10 glucosidase inhibitory,11

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and neuroprotective activities.18 However, no literature could be found for any A-type

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proanthocyanidin-cysteine conjugate. In the present research, we detected an A-type

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proanthocyanidin-cysteine conjugate for the first time from the degradation product of

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cinnamtannin D-1 (7). Preparation, purification and bioactivity evaluation of novel

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A-type proanthocyanidin-cysteine conjugates are planed to be carried out in our group

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in the near future.

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Proanthocyanidins are rich in hydrophobic aromatic rings and hydrophilic hydroxyl

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groups, which could interact with biological molecules, especially proteins. It is

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generally believed that the activity of proanthocyanidins becomes stronger as the

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number of flavanol units increases. The result of our study confirmed that

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proanthocyanidins have strong antioxidant activities, which could provide protection

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against oxidative stress and thereby prevent a variety of human diseases so

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associated.19-21

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In summary, we have isolated nine compounds, including one novel trimeric A-type

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proanthocyanidin (8), and a known trimeric A-type proanthocyanidin (9) that was

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reported for the first time for peanut skin. All these compounds showed antioxidant

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activity and the 2 trimeric A-type proanthocyanidins, 8 and 9 showed considerable

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inhibitory activities on the intestinal disaccharidases, maltase and sucrase.

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ACKNOWLEDGMENTS

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We are grateful to the Government of Inner Mongolia for financial support and to Mr.

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Menghe (Inner Mongolian University, Huhhot, China) for measurement of NMR

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spectra. We thank Dr.. Lijun (School of Pharmaceutical Sciences, Peking University,

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Beijing, China) for performing the HRESI-MS.

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ASSOCIATED CONTENT

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Supporting information

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Additional figures. This material is available free of charge via the Internet at

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http://pubs.acs.org.

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REFERENCES

282

(1) Jiansu Xin Medical College, 1977. Dictionary of Chinese MateriaMedica

283

Shanghai Science and Technology Publisher (JapaneseTranslation), pp. 2648−2649.

284

(2) Tatsuno, T.; Jinno, M.; Arima,Y.; Kawabata, T.; Hasegawa, T.; Yahagi, N.; Takano,

285

F.; Ohta, T. Anti-inflammatory and Anti-melanogenic Proanthocyanidin Oligomers

286

from Peanut Skin. Bio. Pharm .Bull. 2012, 35, 909−916.

287

(3) Lou, H.; Yamazaki, Y.; Sasaki, T.; Uchida, M.; Tanaka, H.; Oka, S. A-type

288

proanthocyanidins from peanut skins. Phytochemistry. 1999, 51, 297−308.

289

(4) Lou, H. X.; Yuan, H. Q.; Ma, B.; Ren, D. M.; Ji. M.; Oka, S. Polyphenols from

290

peanut skins and their free radical-scavenging effects. Phytochemistry. 2004, 65,

291

2391−2399.

292

(5) Monagas, M.; Garrido, I.; Lebron-Aguilar, R.; Gomez-Cordoves, M. C.;

293

Rybarczyk, A.; Amarowicz, R.; Bartolome, B. Comparative flavan-3-ol profile and

294

antioxidant capacity of roasted peanut, hazelnut, and almond skins. J. Agric. Food

295

Chem. 2009, 57, 10590−10599.

296

(6) Anderson, R.; Broadhurst, C.; Polansky, M.; Schemidt, W.; Khan, A.; Flanagan, V.;

297

Schoene, N.; Graves, D. Isolation and characterization of polyphenol type-A polymers

298

from Cinnamon with insulin-like biological activity. J. Agric. Food Chem. 2004, 52,

299

65-70.

300

(7) Wilson, T.; Luebke, J.; Morcomb, E.; Carrell, E.; Leveranz, M.; Kobs, L.; Schmidt,

301

T.; Limburg, P.; Vorsa, N.; Singh, A. Glycemic responses to sweetened dried and raw

302

cranberries in humans with type 2 diabetes. J. Agric. Food Chem. 2004, 52, 65-70.

14


Page 15 of 30


303

(8) Dimitriadis, G.; Tessari, P.; Go, V.; Gerich, J. Effects of the disaccharidase

304

inhibitor acarbose on meal and intravenous glucose tolerance in normal man.

305

Metabolism 1982, 31, 841-843.

306

(9) Torres, J. L.; Lozano, C.; Julia, L.; Sa′ nchez-Baeza, F. J.; Anglada, J. M.;

307

Centellesc J. J.; Cascante, M. Cysteinyl-flavan-3-ol conjugates from grape

308

procyanidins. Antioxidant and antiproliferative properties. Bioorg. Med. Chem. 2002,

309

10, 2497−2509.

310

(10) Fujii, H.; Nakagawa,T.; Nishioka,H.; Sato, E.; Hirose, A.; Ueno, Y.; Sun, B. X.;

311

Yokozawa, T.; Nonaka. G. I. Preparation, characterization, and antioxidative effects of

312

oligomeric proanthocyanidin-L-cysteine complexes. J. Agric. Food Chem. 2007, 55,

313

1525−1531.

314

(11) Meng, H.C.; Ma, C. M. Flavan-3-ol-cysteine and acetylcysteine conjugates from

315

edible reagents and the stems of Cynomorium songaricum as potent antioxidants.

316

Food Chem. 2013, 141, 2691-2696.

317

(12) Ma, J. N.; Wang, S. L.; Zhang, K.; Wu, Z. G.; Hattori, M.; Chen, G. L.; Ma. C. M.

318

Chemical components and antioxidant activity of the peels of commercial

319

apple-shaped pear (fruit of pyrus pyrifolia cv. pingguoli). J. Food Sci. 2012, 10,

320

1097−1102.

321

(13) Zhang, C. F.; Sun, Q.; Wang, Z. T.; Hattori, M. One new A-type

322

proanthocyanidin trimer from Lindera aggregata (Sims) Kosterm. Chin. Chem. Lett.

323

2003, 14, 1033−1036.

324

(14) Killday, K. B.; Davey, M. H.; Glinski, J. A.; Duan, P.G.; Veluri, R.; Proni, G.;

15



Page 16 of 30

325

Daugherty, F. J.; Tempesta, M. S. Bioactive A-type proanthocyanidins from

326

Cinnamomum cassia. J. Nat. Prod. 2011, 74, 1833−1841.

327

(15) Barrett, M.; Klyne, W.; Scopes, P.; Fletcher, A.; Porter, L.; Haslam, E．Plant

328

proanthocyanidins. Part 6. Chiroptical studies. Part 95. Circular dichroism of

329

procyanidins. J.Chem. Soc., Perkin Trans. I, 1979, 2375－2377．

330

(16) Morimoto, S.; Nonaka, G.; Nishioka, I. Tannins and related compounds. LX.

331

Isolation and characterization of proanthocyanidins with a doubly-linked unit from

332

Vaccinium vitis-idaea L. Chem. Pharm. Bull. 1988, 36, 33−38.

333

(17) Ho, K. Y.; Tsai, C. C.; Huang, J. S.; Chen, C. P.; Lin. T. C.; Lin, C. C.

334

Antimicrobial activity of tannin components from Vaccinium vitis-idaea L. J. Pharma.

335

Pharmacol. 2001, 53, 187−191.

336

(18) Torres, J. L.; Lozano, C;Maher, P. Conjugation of catechins with cysteine

337

generates antioxidant compounds with enhanced neuroprotective activity.

338

Phytochemistry, 2005, 66, 2032−2037.

339

(19) Shahidi, F. Natural antioxidants-and overview. In F. Shahidi (Ed.), Natural

340

antioxidants: Chemistry, health effects, and application. Champaign, IL: The

341

American Oil Chemists’ Society. 1997, pp. 1−10

342

(20) Shahidi, F. Beneficial health effects and drawbacks of antinutrients and

343

phytochemicals in foods. In F. Shahidi (Ed.), Antinutrients and phytochemicals in

344

food (ACS symposium series 662). Washington, DC: American ChemicalSociety.

345

1997, pp. 1−9

346

(21) Ren, Y.; Chen, X. Distribution, bioactivities and therapeutical potentials of

16


Page 17 of 30


347

pentagalloylglucopyranose. Curr. Bioact. Compd. 2007, 3, 81−89.

348

17



349

Page 18 of 30

Figure Captions

350 351

Figure 1. Structures of compounds 1‒9 from peanut skin.

352

Scheme 1. Degradation of compound 7 with L-cysteine HCl. Figure 2. Key correlations observed in the HMBC and 1H-1H COSY spectra of 8. ) 1H-1H COSY (

353

HMBC (

)

354

Scheme 2. Degradation of compound 8 with L-cysteine HCl.

355


356

Figure 3. Extracted ion chromatograms of the LC-MS of cysteine-induced

357

degradation of 7 (A), 8 (B), and 9 (C), in comparison with catechin (D), procyanidin

358

A1 (E) and epicatechin-cysteine (F).

359

Table 1. NMR spectral Data of Compound 8 in CD3OD.

360

Table 2. DPPH Radical Scavenging Activities of Proanthocyanidins from Peanut

361

Skin.

362

Table 3. The Maltase and Sucrase Inhibitory Activities of Compounds from Peanut

363

Skin.

364

18


Page 19 of 30


OH 13 8

HO

9

7

OH

14

12 11

O

15

2 16

OH 13 8

HO

O

9

7

OH OH

14

12 11

13

15

2

8

HO

16 10

5

7'

10

5

OH

OH

4

12'

13'

OH

10' 4'

2'

11'

3' 14'

OH

OH

16'

HO

15'

2

1

5'

O HO

6'

8' 9'

15

3

6

OH

4

2

O

OH

4

OH

16

3

6

O

9

7

10

5

OH

14

12 11

3

6

3 OH

OH 13

HO

8

7

O

9

13

15

8

HO

3

6 10

5

4

13'

12'

11'

HO

14'

15

4

8' 9'

5'

HO

O

10'

5'

2'

4'

OH

10'

7'

6'

OH 6'

11' 2'

O

OH

14'

4'

OH

OH

HO

8 7

O

9

11

2

13

OH

14

12

6 10

5

HO

15

OH

4

OH

O HO

13'

12'

HO

7'

11'

4'

8''

OH O

HO

13''

12''

11''

2''

16''

8'

OH O HO

13'

11''

12''

12'

11'

10' 4'

2'

OH

14'

16'

OH

9'

O

12'

2'

13' 14'

OH

3'

6' 5'

15''

10'

4'

HO

13''

12''

3''

14''

HO

8

16''

OH

10''

11'' 2''

OH

15'

7'' 6'' 5''

9''

O

15'

OHO 8''

OH

OH

OH

11' 16'

13''

3'

HO

8'

14''

OH 16''

5''

OH

2''

OH

4

3''

6' 5'

9'

4''

OH

7'

O

10'' 4''

10

OH HO 7'

9''

15''

7

365 366

4

HO O

15

3 5

8''

5''

2

6

7''

15

OH

6''

10'' 3''

14''

HO

11

2 3

10

5

7''

9''

15'

O

9

6

OH OH

2'

8

7

6'' 16

5' 10'

16'

HO

6'

OH

O

9

7

11 16

OH

14

12

9'

3' 14'

13

O

8'

OH

8

OH

14

12

16 3

OH

OH

6

5 13

15' 14'

13'

15'

4

16'

12'

OH

OH

16'

HO

15'

11'

3'

3'

OH

16'

13'

12'

10

7'

O

4'

9

11

2 3

5

9'

OH

10'

2'

7

O

6

O

8'

OH

3'

HO

OH

4

5'

9'

HO

10

5 6'

O

8

16

15

3

6

7'

8'

OH

11

2

HO

16

O

OH

O

9

7

16

OH

OH

14

12

14

12

11

2

OH

14

12

13

OH

4''

OH

15''

9

Figure 1. Structures of compounds 1-9 from peanut skin.

19



Page 20 of 30

367 OH

OH

OH

OH HO

HO

O OH

O

OH

O

L-cysteine HCl

OH OH

HO

O

O

OH

HO

HO

OH

HO

OH O

OH

HO O

OH

O

+

OH

OH

OH S OH H2N

OH

catechin COOH

HO OH

HO

7

368 369


20


Page 21 of 30


370 OH OH OH HO

B

O

A

C 4

OH

G HO

3

O

OH

8'

F

H

E

H OH

D

O 9'

2"

4" 6'

H

HO

I O

10'

OH

H OH

OH

2'

OH

HO

Figure 2. Key correlations observed in the HMBC and 1H-1H COSY spectra of 8. 371

HMBC (

) 1H-1H COSY (

)

372

21



Page 22 of 30

373 OH

OH

OH OH

HO

OH

O

HO OH

HO O

OH

OH

OH O

HO

L-cysteine HCl

OH OH

HO

O

OH

OH O

OH

OH

S

HO H2N

OH HO

COOH

OH

procyanidins A1

374 375

O

+

OH

OH

HO

OH

O

epicatechin-cysteine


22


Page 23 of 30


376 OH OH OH

HO

OH

OH OH

OH

HO

O OH

L-cysteine HCl

O

OH

O O

OH

HO OH

HO

OH

O OH

+

HO

HO HO

O

OH

OH

377

OH

O

OH

OH HO

OH

procyanidins A1

OH

S

H2N

COOH

epicatechin-cysteine


378

23



Page 24 of 30

379 380

Figure 3. Extracted ion chromatograms of the LC-MS of cysteine-induced

381

degradation of 7 (A), 8 (B), and 9 (C), in comparison with catechin (D), procyanidin

382

A1 (E) and epicatechin-cysteine (F).

383 384 385

24


Page 25 of 30


386

25



387

Page 26 of 30

Table 1. NMR spectral Data of Compound 8 in CD3OD Ring

No.

13

1

C

2 3 4

100.88 67.14 29.46

—— 4.20, br.s 4.35, br.s

A

5 6 7 8 9 10 11 12 13 14 15 16

156.54 96.05 157.82 98.35 156.54 104.07 131.86 116.14 146.09 146.94 116.01 120.19

D —— 6.10, br.s —— 5.96, br.s —— —— E —— 7.18, br.s —— —— 6.82, m 7.06, d (7.8)

B

C

H

Ring No . F 2′ 3′ 4′ 5′ 6′ 7′ 8′ 9′ 10′ 11′ 12′ 13′ 14′ 15′ 16′

13

1

Ring No.

13

1

84.21 67.49 28.58

4.79, d (6.5) 4.15, m 2.76, dd (16.1, 7.5) 2.50, dd (16.1, 7.5) —— —— —— —— —— —— —— 6.87, br.s —— —— 6.10, br.s 6.77, m

I

2″ 3″ 4″

77.21 73.34 37.06

5.33, br.s 4.09, m 4.63, br.s

G

5″ 6″ 7″ 8″ 9″ 10″ 11″ 12″ 13″ 14″ 15″ 16″

159.21 96.45 154.55 99.18 158.18 99.18 132.93 115.7 145.66 146.45 119.22 120.19

—— 6.04, br.s —— 5.93, br.s —— —— —— 6.94, br.s —— —— 6.78, m 6.74, m

C

149.29 109.35 150.1 107.33 154.44 103.93 130.43 115.26 145.91 146.85 120.59 120.08

H

H

C

H

388 389

26


Page 27 of 30


390

27



Page 28 of 30

391 392

Table 2. DPPH Radical Scavenging Activities of Proanthocyanidins from Peanut Skin sample

393 394 395

Effect

EC50 (µg/ml)

%

20 (µg/ml)

10 (µg/ml)

5 (µg/ml)

2.5 (µg/ml)

1 2 3 4

83.01 94.52 94.02 86.91

61.69 74.33 63.14 58.93

37.42 42.07 29.93 28.80

31.88 34.59 26.28 20.00

8.21 6.12 8.55 9.71

5

79.43

47.29

30.06

15.28

11.36

6 7

93.39 93.33

67.29 54.21

47.10 26.79

38.61 19.43

5.68 9.74

8

84.21

53.01

37.61

34.02

8.61

89.37 85.66 48.49 20.56 9 Catechin (1), a known anti-oxidant was used as a positive control.

6.32

28


Page 29 of 30


396

Table 3. The Maltase and Sucrase Inhibitory Activities of Compounds from Peanut

397

Skin Inhibition% on IC50 on Inhibition% on IC50 on Sucrase Maltase Maltase Sucrase (mg/ml) at 1 mg/ml (mg/ml) at 1 mg/ml >1 1 44 45 >1 0.088 2 60 54 0.57 >1 3 >1 47 44 >1 4 >1 44 31 >1 >1 5 47 33 0.61 >1 6 67 42 0.78 0.66 7 62 55 0.088 0.41 8 83 61 0.54 0.091 9 79 64 0.0054 0.0074 Acarbose* nt nt Acarbose*, a known α–glucosidase inhibitor, was used as a positive control. nt: not tested, as this concentration is too high for the this very potent α–glucosidase inhibitor, acarbose. Sample

398 399 400 401

29



402 403

Page 30 of 30

Graphic Abstract

404 405 406 407 408 409

30


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