Isolation and structural determination of a novel flavonol triglycoside and 7 compounds from the leaves of oriental raisin tree (Hovenia dulcis) and their antioxidative activity

The hot water and ethanol extracts of oriental raisin tree (Hovenia dulcis Thunb) leaves showed DPPH radical scavenging activities. Antioxidants were purified and isolated from hot water and ethanol extracts by various column chromatographic procedures with the guided assay of DPPH radical scavenging. The structure of a novel flavonol triglycoside was determined to kaempferol 3-O-α-l-rhamnopyranoside-7-O-[α-d-glucopyranosyl(1→3)-α-l-rhamnopyranoside] (4). In addition, 7 known compounds were identified as caffeine (1), kaempferol 3,7-O-α-l-dirhamnopyranoside (2), kaempferol 3-O-α-l-rhamnopyranosyl( 1→6)-O-β-d-glucopyranosyl(1→2)-O-β-d-glucopyranoside (3), E-3-carboxy-2-petenedioate 5-methyl ester (5), quercetin 3-O-α-l-rhamnopyranoside (6), kaempferol 3-O-α-l-rhamnopyranoside (7), and quercetin 3-O-β-d-glucopyranoside (8). Compound 1–3 and 5–8 were newly identified in this plant. Quercetin glycosides (5, 7) showed higher DPPH radical scavenging activity than other compounds.     

Phytoestrogenic activity of Aceriphyllum rossii and rapid identification of phytoestrogens by LC–NMR/MS and bioassay-guided isolation

Aceriphyllum rossii Engler (Saxifragaceae) has been used as a nutritious food in Korea, but only minimal studies on this plant have been undertaken. We have examined the estrogenic effect of A. rossii and have discovered its potent constituents. To identify the estrogenic activity of A. rossii, we measured the transactivation of the estrogen-responsive element (ERE) in MCF-7 cells transfected with an ERE-containing construct after treatment of the extract and fractions of A. rossii. A. rossii showed a potent estrogenic activity and gallic acid ethyl ester; quercetin and kaempferol were identified as major constituents of the active fractions by liquid chromatography–nuclear magnetic resonance spectroscopy/mass spectrometry. We confirmed quercetin and kaempferol are the major active constituents after activity-guided isolation of seven compounds: gallic acid ethyl ester (1), quercetin (2), kaempferol (3), quercetin-3-O-(6″-galloyl)-β-d-glucopyranoside (4), quercetin-3-O-β-d-glucopyranoside (5), kaempferol-3-O-(6″-galloyl)-β-d-glucopyranoside (6), and kaempferol-3-O-β-d-glucopyranoside (7). These results show A. rossii could be a possible candidate for the treatment of postmenopausal symptoms.     

Purification and characterization of <Emphasis Type="Italic">β</Emphasis>-glucosidase from <Emphasis Type="Italic">Oenococcus oeni</Emphasis> 31MBR

This study was designed to characterize a β-glucosidase from Oenococcus oeni 31MBR, a strain widely used in Chinese winemaking. An intracellular β-glucosidase (EC 3.2.1.21) was partially purified using a combination of ammonium sulfate precipitation and chromatographic methods. A single band was obtained in SDS-PAGE electrophoresis, indicating that the enzyme was highly purified and had an estimated molecular mass of 38.9 kDa. The enzyme exhibited highest activity at pH 4.5–5.0. The optimum temperature was 45 °C. Ethanol promoted the activity of this enzyme up to three times. Among the several metal ions assayed, only Mn²⁺ exhibited a partial promotion effect. The K _m and V _max values for p-nitrophenyl-β-d-glucopyranoside were 1.05 mmol/L and 0.957 nmol/min, respectively. Up to now, this study contains the first characterization of a native β-glucosidase purified from crude extracts of O. oeni 31MBR.     

Coumaroyl quinic acid derivatives and flavonoids from immature pear (Pyrus pyrifolia nakai) fruit

Fourteen compounds were isolated from 60% ethanol extracts of immature pear (Pyrus pyrifolia Nakai cv. Chuhwangbae) fruit using Amberlite XAD-2 column HPLC with guided DPPH radical scavenging assay. Based on MS and NMR analysis, the isolated compounds were identified as 5-O-trans-caffeoyl quinic acid methyl ester (1), malaxinic acid (2), 5-O-trans-p-coumaroyl quinic acid methyl ester (3), 5-O-cis-p-coumaroyl quinic acid methyl ester (4), 5-O-trans-p-coumaroyl quinic acid (5), trans-p-coumaric acid (6), methyl cis-p-coumarate (7), methyl trans-p-coumarate (8), 3,5-O-dicaffeoyl quinic acid (9), (-)-epicatechin (10), (S)-(+)-2-cis-abscisic acid (11), isorhamnetin 3-O-β-d-galacto-pyranoside (12), isorhamnetin-3-O-β-d-glucopyranoside (13), and isorhamnetin 3-O-α-l-rhamnopyranosyl (1→6)-O-β-d-glucopyranoside (14). Six compounds (1, 2, 6, 9, 10, and 13) were identified previously, but other compounds (3–5, 7, 8, 11, 12, and 14) were isolated for the first time from pear.     

Stability of naturally occurring 2,5-dimethyl-4-hydroxy-3 [2H]-furanone derivatives

The stability, in aqueous buffer solutions at different pH values (pH?2.0–8.0, interval: 1.5?pH units), of 2,5-dimethyl-4-hydroxy-3[2H]-furanone (Furaneol, DMHF, 1), its methoxy derivative 2,5-dimethyl-4methoxy-3[2H]-furanone (methoxyfuraneol, mesifurane, DMMF, 2 and the glycosidically bound forms DMHF β-D-glucopyranoside 3 and DMHF 6′-O-malonyl-β-Dglucopyranoside 4 was investigated over a period of 32 days at 23?°C. Only slight decomposition of 2 and 3 was observed, whereas 1 and 4 were found to be unstable at all pH values. In addition, 3 and 4 were subjected to enzymatic hydrolysis. In contrast to the rapid hydrolysis of 3, the malonylated glycoside, 4, remained unaffected by enzymatic treatment with β-glucosidase (Emulsin). Using a pectinolytic enzyme preparation (Rohapect D5L; Röhm, Darmstadt, Germany) with esterase activities, hydrolysis of 4 was achieved.     

Determination of <Emphasis Type="SmallCaps">d</Emphasis>,<Emphasis Type="SmallCaps">l</Emphasis>-Amino Acids in Collagen from Pig and Cod Skins by UPLC Using Pre-column Fluorescent Derivatization

A simple and sensitive analytical method for the quantification of d,l-amino acids by ultra-performance liquid chromatography (UPLC) with fluorescence (FL) detection is described. The reaction of the R(-)-4-(3-isothiocyanatopyrrolidin-1-yl)-7-(N,N-dimethylaminosulfonyl)-2,1,3-benzoxadiazole [R(-)-DBD-PyNCS] with d,l-hydroxyproline (Hyp), glycine (Gly), d,l-aspartic acid (Asp), and d,l-proline (Pro) effectively proceeds at 55 °C for 20 min in the presence of 3% TEA to produce the corresponding fluorescent diastereomers (excitation at 460 nm, emission at 550 nm). The mixture was simultaneously separated within 20 min on an ACQUITY UPLCTM BEH C18 (1.7 μm, 100 mm?×?2.1 mm i.d.) by gradient elutions using water–acetonitrile containing 0.2% formic acid as the mobile phase. Peak resolution was in the range of 1.62 (d,l-Asp), 2.99 (d,l-Pro), and 6.74 (d,l-Hyp). A good linearity was achieved from the calibration curves (r²?>?0.9984) in the range of 1.0~100 pmol; the limit of detection (S/N?=?3) was 42.0–250 fmol, the inter-day and intra-day assay precisions were all less than 6.23%, and the mean recoveries (%) of the d,l-amino acids spiked in the collagen from pig and dried cod skins were 87.58–107.41%. The derivatives of the free d,l-amino acids in the collagen were successfully identified by the proposed procedure. To the best of our knowledge, for the first time, these three kinds of d-amino acids, which were d-Asp, d-Pro, and d-Hyp, were detected in the collagen samples.     

Isolation of five proanthocyanidins from pear (<Emphasis Type="Italic">Pyrus pyrifolia</Emphasis> Nakai) fruit peels

Five proanthocyanidins, two B-type dimers and three A-type trimers, were purified and isolated from the fruit peels of Pyrus pyrifolia Nakai cv. Chuhwangbae. The isolated compounds were identified as (–)-epicatechin gallate-(4β → 8)-(–)-epicatechin (Hahashi et al. in Ann Biol Res 3:3200–3207, 2012), (–)-epicatechin-(4β → 8)-(–)-epicatechin (procyanidin B2) (Tanrioven and Eksi in Food Chem 93:89–93, 2005), (–)-epicatechin-(4β → 8, 2β → O-7)-(–)-epicatechin-(4β → 8)-(–)-epicatechin (cinnamtannins B1) (Salta et al. in J. Fun. Food 2: 153–157, 2010), (–)-epicatechin-(4β → 8)-(–)-epicatechin-(4β → 8, 2β → O-7)-(–)-epicatechin (aesculitannin A) (Challice and Westwood in Phytochemistry 11: 37–44, 1972), and (–)-epicatechin-(4β → 6)-(–)-epicatechin-(4β → 8, 2β → O→7)-(–)-epicatechin (Es-Safi et al. in J Agric Food Chem 54: 6969–6977, 2006). Their structures were determined by nuclear magnetic resonance and mass spectrometry. The three A-type proanthocyanidin trimers were identified for the first time from pear.     

Green Approach for Sample Preparation and Determination of Anthocyanins from <Emphasis Type="Italic">Lycium ruthenicum</Emphasis> Murr. Using a <Emphasis Type="Italic">β-</Emphasis>Cyclodextrin-Based Extraction Method Coupled with UPLC-DAD Analysis

This study aimed to develop and optimize a β-cyclodextrin (β-CD)-based technique for extracting anthocyanins from Lycium ruthenicum Murr. and to establish a ultra-high-performance liquid chromatography-diode array detector (UPLC-DAD) method for their analysis. β-CD solutions produced higher extraction yields of petunidin-3-O-(trans-p-coumaroyl)-rutinoside-5-O-glucoside and total anthocyanins from L. ruthenicum fruit than did pure water and aqueous hydroxypropyl-β-cyclodextrin (HPβ-CD) and ethanol and methanol solutions. Extraction at 50 °C for 30 min using 1.65% β-CD solution and a liquid/solid ratio of 15:1 produced the optimal extraction yield of L. ruthenicum anthocyanins. A UPLC-DAD method was developed for the determination of L. ruthenicum anthocyanins using an ethanol-based mobile phase, and the primary anthocyanins were identified using two-dimensional LC-MS/MS. Method validation showed that the developed method was accurate, stable, and reliable for the analysis of petunidin-3-O-(trans-p-coumaroyl)-rutinoside-5-O-glucoside and total anthocyanins from L. ruthenicum fruit. The present study showed that β-CD-based extraction coupled with UPLC-DAD analysis is an efficient and green method for the extraction and analysis of anthocyanins from L. ruthenicum fruit.     

C-dideoxyhexosyl flavones from the stems and leaves of Passiflora edulis Sims

The stems and leaves of Passiflora edulis Sims, are used as a folk medicine for treating both anxiety and nervousness in American countries. Phytochemical investigation of the n-butanol (n-BuOH) fraction of this plant led to the isolation of four new 2,6-dideoxyhexose-C-glycosyl flavones, including luteolin-8-C-β-digitoxopyranosyl-4′-O-β-d-glucopyranoside (1), apigenin-8-C-β-digitoxopyranoside (2), apigenin-8-C-β-boivinopyranoside (3) and luteolin-8-C-β-boivinopyranoside (4), together with five known compounds (5–9). The structures of these compounds were elucidated by extensive spectroscopic methods. All compounds were evaluated for their neurite outgrowth enhancing activities and the results indicated that luteolin (7) enhanced NGF-induced neurite outgrowth in PC12 cells at 50.0 μM.     

Two novel glycosyl cinnamic and benzoic acids from korean black raspberry (Rubus coreanus) wine

Korean black raspberry (Rubus coreanus) wine was solvent-fractionated with n-hexane, ethyl acetate, and n-butanol and the n-butanol layer was purified by various column chromatographic procedures, including Amberlite XAD-2, Sephadex LH-20, and octadecylsilane resins as well as high performance liquid chromatography. Two novel glycosyl cinnamic and benzoic acids were isolated and their structures were (E)-8-O-β-d-glucopyranosylcinnamic acid (1) and 3′-[O-β-d-glucopyranosyl)(1″→6′)-α-d-psicofuranosyl] benzoate (2) based on the spectroscopic data obtained by high resolution fast-atom bombardment mass spectroscopy and nuclear magnetic resonance analyses.     

Melanin biosynthesis inhibitory effects of calycosin-7-O-β-d-glucoside isolated from astragalus (Astragalus membranaceus)

To discover an active skin depigmentation agent, we isolated a melanin biosynthesis inhibitor from the methanol extract of astragalus (Astragalus membranaceus) using a bioactivity-guided fractionation and identified it as calycosin-7-O-β-d-glucoside via spectroscopic analysis. The active compound exhibited tyrosinase inhibitory activity with an IC₅₀ value of 68 μM. In addition, calycosin-7-O-β-d-glucoside showed a melanin biosynthesis inhibition zone in a culture plate of Streptomyces bikiniensis. Furthermore, 75.78 μM of calycosin-7-O-β-d-glucoside dramatically decreased 50% of the melanin content on Melan-a cells without any apparent cytotoxicity. Theses results indicate that the calycosin-7-O-β-d-glucoside isolated from astragalus may be a good candidate for skin-whitening agent.     

Flavonoids from Pueraria mirifica roots and quantitative analysis using HPLC

The flavonoids, wighteone (1), naringenin (2), genistein (3), isoliquiritigenin (4), daidzein (5), daidzin (6), genistein-8-C-β-d-apiofuranosyl-(1→6)-O-β-d-glucopyranoside (7), ambocin (8), and genistin (9) were isolated from roots of Pueraria mirifica. Chemical structures of the compounds were determined based on spectroscopic data analysis. Flavonoids 1, 2, 4, 7, and 8 were isolated for the first time. The contents of 6 flavonoids in P. mirifica roots was determined to be 2.5±0.01 (1), 14.8±0.09 (3), 18.6±0.18 (5), 17.3±0.11 (6), 10.4±0.05 (7), and 6.0±0.02 (9) mg/kg, respectively, using HPLC.     

Antioxidant properties and polyphenolics composition of common hedge hyssop (Gratiola officinalis L.)

The antioxidant potential of Gratiola officinalis was evaluated by the off-line and on-line HPLC/UV/DPPH radical scavenging assays, phytochemical composition was analyzed by LC/MS. On-line method was validated by using reference antioxidants and linear dependence was found between their concentration and radical scavenging peak area. Radical scavenging capacity (RSC) of methanol and acetone extracts expressed in their concentration required to scavenge 50% of DPPH^? was 0.10% and 0.13%, respectively; the RSC in ABTS^?+ assay was 1093 ± 104 and 746 ± 18 μM of trolox equivalents in 1 g, respectively. Good correlation was observed between total amount of phenolic compounds and RSC. Preliminary HPLC/UV/MS analysis revealed that the main compounds possessing antioxidant activity in the extracts might be phenylpropanoid glycosides; UPLC/UV/ESI-QTOF-MS analysis suggested 15 structures: 2,5-dihydroxy-p-benzenediacetic and caffeic acids, apigenin 6,8-di-C-β-d-glucopyranoside (vicenin-2), apigenin 8-C-α-l-arabinoside 6-C-β-d-glucoside, (shaftoside), forsythoside B, arenarioside, verbascoside (acteoside), amioside, quercetin-6-O-(2-O-acetyl-glucopyranosyl)-glucopyranoside, isoverbascoside, quercetin glucuronide, linariifolioside, methoxy luteolin-7-O-(6-O-acetyl-glucopyranosyl)-glucopyranoside, methoxy luteolin-glucuronide and luteolin glucuronide.     

A New Chemical Marker-Model Food System for Heating Pattern Determination of Microwave-Assisted Pasteurization Processes

New chemical marker-model food systems with d-ribose and NaOH precursors as color indicators and gellan gels as chemical marker carrier were explored for the assessment of the heating pattern of in packaged foods processed in microwave-assisted pasteurization system (MAPS). In determining appropriate precursor concentrations, a solution of 2% (w/w) d-ribose and 60 mM NaOH was heated at 60–90 °C for 0–20 min. The solution absorbance at 420 nm increased linearly, while the color parameters L* decreased linearly with heating time at all processing temperatures. In storage, the produced brown color was stable at 4 and 22 °C within 7 days. The new chemical marker-model foods were prepared by mixing 2% (w/w) d-ribose and 60 mM NaOH with 1% (w/v) low-acyl gellan gum and 20 mM CaCl₂·2H₂O solution. The dielectric constant of the model food samples decreased with the addition of sucrose, and the loss factors increased with the addition of salt. After processing in the pilot MAPS, the heating pattern and cold and hot spots in the new chemical marker-model food system could be clearly recognized and precisely located through a computer vision method. This is the first time that the caramelization reaction was used as a time-temperature indicator in gellan gel model food. This study shows the possibility of using the new chemical marker-model food system for heating pattern determination of the MAPS.     

Phenolic constituents from the flower buds of Lonicera japonica and their 5-lipoxygenase inhibitory activities

Thirteen phenolic constituents, luteolin (1), protocatechuic acid (2), caffeic acid (3), flavoyadorinin-B (4), 4,5-dicaffeoylquinic acid (5), luteolin 7-O-β-d-glucopyranoside (7), 3,5-dicaffeoylquinic acid methyl ester (8), methyl chlorogenate (9), quercetin 3-O-β-d-glucopyranoside (10), 3,5-dicaffeoylquinic acid (11), rhoifolin (12), chlorogenic acid (13), and a novel phenolic glucoside benzoate, vanillic acid 4-O-β-d-(6-O-benzoylglucopyranoside) (6), were isolated from the flower buds of Lonicera japonica. Flavoyadorinin-B (4) was isolated for the first time from a Caprifoliaceae plant. The structures of 1–13 were determined on the basis of chemical and spectroscopic evidence. These compounds were screened for their 5-lipoxygenase inhibitory activity. Only luteolin (1) showed significant inhibitory activity against 5-LOX-catalysed leukotriene production.     

Partial purification and characterization of polyphenol oxidase from Chinese parsley (Coriandrum sativum)

Rates of thermal degradation and isomerization of all-trans-β-carotenes in air and in triacylglycerols were determined. Degradation of carotenes in triacylglycerols was faster than that in air. The 13-cis-β-carotene level in triacylglycerols was higher than in air. Oxidized materials of triacylglycerols probably facilitated isomerization of carotenes and, thus, degradation. Amounts of all-trans-β-carotenes and all-trans-α-carotenes in pumpkin decreased with an increase in heating time. The proportion of 13-cis-β-carotene increased after heat treatment started, probably due to thermal isomerization of all-trans-β-carotenes to cis-isomers, and to decreases in amounts of all-trans-β-carotenes and all-trans-α-carotenes in pumpkin. Effects of heating methods on proportions of isomers; however, were not different.     

Anti-complementary activity of flavonoids from Gnaphalium affine D. Don

Gnaphalium affine D. Don, is used as a vegetable as well as a folk medicine in China for its anti-inflammatory, antitussive, and expectorant activities. Phytochemical investigation of the ethyl acetate (EtOAc) fraction of this plant led to isolation of three new acylated flavonol glycosides, apigenin 4′-O-β-d-(6″-E-caffeoyl)-glucopyranoside, luteolin 4′-O-β-d-(6″-E-caffeoyl)-glucopyranoside, and quercetin 4′-O-β-d-(6″-E-caffeoyl)-glucopyranoside, together with 24 known compounds. The structures of these compounds were determined by spectroscopic methods. All compounds were evaluated for their anti-complementary activity on the classical pathway of the complement system in vitro, and luteolin 4′-O-β-d-(6″-E-caffeoyl)-glucopyranoside exhibited highest anti-complementary activity with an IC₅₀ value of 0.045 ± 0.005 mg/ml. This is the first report of anti-complementary activity of G. affine D. Don, which displays the initial evidence that this plant is a potent complement inhibitor.     

An approach for extraction,purification, characterization and quantitation of acylated-anthocyanins from <Emphasis Type="Italic">Nitraria tangutorun</Emphasis> Bobr. fruit

In the present study, a systematic approach for extraction, purification and analysis of acylated-anthocyanins from Nitraria tangutorun Bobr. fruit was explored. Six acylated-anthocyanins in N. tangutorun fruit were identified by HPLC-MS/MS, and a rapid and efficient HPLC-DAD method was developed to analyze the acylated-anthocyanins. Ultrasonic-assisted extraction conditions of acylated-anthocyanins were optimized using response surface methodology, extraction at 70 °C for 32 min using 70% methanol solution (0.1% HCl, v/v) rendered an extract with 80.37?±?2.66 mg/100 g of cyanidin-3-O-(trans-p-coumaroyl)-diglucoside and 97.88?±?4.06 mg/100 g of total acylated-anthocyanins. Nine macroporous resins were investigated for preliminary purification of acylated-anthocyanins. According to the static/dynamic adsorption and desorption tests, XDA-6 macroporous resin exhibited the maximum potential for preparing acylated-anthocyanins. The purity of cyanidin-3-O-(trans-p-coumaroyl)-diglucoside (43.30 mg/g) in purified acylated-anthocyanins was 201.89 times of that of the extract (0.21 mg/g), and the purity of total acylated-anthocyanins increased from 0.36 to 56.44 mg/g. Besides, the stability (t _1/2) of cyanidin-3-O-(trans-p-coumaroyl)-diglucoside and total acylated-anthocyanins increased by more than five-fold after purification using XDA-6. The established methods of analysis, extraction and purification of acylated-anthocyanins were hopefully utilized in food industry.     

Chemical composition, mineral content and antioxidant activity of Verbena officinalis L.

Aqueous and hydroalcoholic extracts from Verbena officinalis L. were obtained and characterised. The analysis by HPLC-DAD and LC-MS allowed the detection and identification of three iridoids, fifteen flavonoids and four phenolic acid derivatives. Four flavonoids, scutellarein 7-diglucuronide (9), scutellarein 7-glucuronide (13), pedalitin 6-galactoside (15) and scutellarein 7-glucoside (19) are reported for the first time from this plant. In addition, three new flavonoids have been isolated: scutellarein 7-O-(2-O-feruloyl)-diglucuronide (5), pedalitin 6-O-diglucuronide (6) and pedalitin 6-O-(2-O-feruloyl)-diglucuronide (13). To our knowledge, these flavonoids have not been reported as natural products. Both extracts showed significant antioxidant activity using three in vitro model systems and the results have been correlated with total phenolic and total flavonoid contents. The results have allowed establishing an important relation structure-activity and significant correlations have also been found between the mineral content and the flavonoids present in both extracts.