Identification of bioactive compounds in Phyllenthus emblica L. fruit and their free radical scavenging activities

Emblica has been used as an important traditional herbal medicine in southeast Asia since ancient times. In this study, the air-dried hulls of emblica fruit were extracted with 95% ethanol, and then the extract was partitioned by diethyl ether and ethyl acetate (EA). The EA fraction was purified by silica gel column and thin layer chromatography (TLC) to obtain six compounds. They were identified as cinnamic acid (C1), quercetin (C2), 5-hydroxymethylfurfural (C3), gallic acid (C4), β-daucosterol (C5) and ellagic acid (C6) using mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy. Cinnamic acid and 5-hydroxymethylfurfural were identified as components of emblica fruit for the first time. The DPPH and ABTS⁺ radical scavenging activities of components were evaluated. All the compounds showed significant DPPH and ABTS⁺ radical scavenging activity except for cinnamic acid. Gallic acid showed the highest DPPH radical scavenging activity while ellagic acid showed the highest ABTS⁺ scavenging activity amongst all the compounds tested.     

Immunomodulatory and anticancer activities of phenolics from emblica fruit (Phyllanthus emblica L.)

There is a paucity of studies on the immunomodulatory properties of fruit extracts of emblica with the emphasis on lymphocytes. Therefore, the aim of the study was to evaluate the immunomodulatory properties and anticancer potential of six phenolic compounds from emblica fruit by in vitro proliferation assay. Effects of these compounds on splenocyte proliferation and the cytotoxicity to both human breast cancer cell (MCF-7) and human embryonic lung fibroblast cell (HELF) were determined by the MTT method. Significantly stimulatory effects (P < 0.05) were found for geraniin and isocorilagin. The concentration of geraniin, quercetin 3-β-d-glucopyranoside, kaempferol 3-β-d-glucopyranoside, isocorilagin, quercetin, kaempferol and rutin to obtain 50% of stimulatory effect was 56, 123, 242, 42, 73, 93 and 92 μg/ml, respectively. The assay of anticancer activities suggested that geraniin and isocorilagin exhibited higher cytotoxicities than other compounds against MCF-7 with IC₅₀ of 13.2 and 80.9 μg/ml, respectively. Isocorilagin exhibited a strong cytotoxicity to HELF cell with IC₅₀ of 51.4 μg/ml. Geraniin, quercetin, kaempferol and their glycosides had weak cytotoxicity against HELF cells. Paclitaxel showed a strong cytotoxicity to MCF-7 and HELF with IC₅₀ of 6.8 and 14.5 μg/ml, respectively. These findings are in line with the reported potent antioxidant activity. These results suggested that the antitumour activity of these compounds might be achieved by immunomodulatory properties which could partially be attributed to their antioxidant activity.     

Isolation and identification of compounds from the ethanolic extract of flowers of the tea (Camellia sinensis) plant and their contribution to the antioxidant capacity

While beneficial health properties of tea leaves have been extensively studied, less attention has been given to that of flowers of the tea (Camellia sinensis) plant. In this work, the ethanolic extract and its ethyl acetate-soluble fraction (EEA) from the tea flowers were found to possess the potent antioxidant activity using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging assay. The compounds present in EEA had comparatively strong DPPH scavenging activity and strongly contributed to the antioxidant activity of the tea flowers. From EEA, besides eight catechins, five flavonol glycosides were isolated and their structures were elucidated on the basis of mass spectrometry and nuclear magnetic resonance spectroscopy as myricetin 3-O-β-d-galactopyranoside, quercetin 3-O-β-d-galactopyranoside, kaempferol 3-O-β-d-galactopyranoside, kaempferol 3-O-β-d-glucopyranoside, and kaempferol 3-O-[α-l-rhamnopyranosyl-(1-6)-β-d-glucopyranoside]. In addition, epigallocatechin gallate and epicatechin gallate were found as the major active components responsible for the antioxidant activity of tea flowers through the use of a combination of preparative liquid chromatography separation and DPPH assay.     

Antioxidant compounds from a South Asian beverage and medicinal plant, Cassia auriculata

Cassia auriculata (Caesalpiniaceae) is a common South Asian beverage and medicinal plant widely used in tradition medicine for treating diabetes and various other disease conditions. The alcoholic extract of the aerial part of C. auriculata displayed potent antioxidant activity when assessed by DPPH radical scavenging, lipid peroxidation and reducing power analysis. Fractionation of the crude extract using solvents of ascending polarity showed that the ethyl acetate fraction is the most active followed by the chloroform fraction while the petroleum ether, n-butanol and water fractions were less active than the crude extract. Further activity-guided fractionation studies on the active fractions resulted in the isolation of the major antioxidant constituent kaempferol-3-O-rutinoside together with kaempferol, quercetin and luteolin. The identity of the compounds was established based on extensive spectroscopic studies including 2D NMR.     

Isolation and evaluation of the radical-scavenging activity of the antioxidants in the leaves of an edible plant, Mallotus japonicus

The antioxidative properties of a hot water extract of the leaves of Mallotus japonicus were evaluated. The extract had a high phenolic content and strong antioxidative activity, compared with green tea, rooibos tea, and red wine. Six phenolic compounds were isolated as antioxidative components by HPLC. They were identified as mallotinic acid, mallotusinic acid, corilagin, geraniin, rutin, and ellagic acid. These antioxidative compounds were subjected to DPPH radical-scavenging, superoxide radical-scavenging, and hydroxyl radical-scavenging assays, and compared with other antioxidative compounds. Four of the compounds, mallotinic acid, mallotusinic acid, corilagin and geraniin, exhibited much stronger antioxidative activity than gallic acid, rutin, ellagic acid, quercetin, and chlorogenic acid, and were as active as epigallocatechin gallate (EGCG), a strong antioxidant in green tea. Mallotus japonicus leaves are an excellent source of strong natural antioxidative materials.     

Antioxidant activity and phenolic composition of sumac (Rhus coriaria L.) extracts

Antioxidant activity and phenolic compounds of sumac extracts were investigated. Sumac was extracted in methanol and subjected to solvent–solvent partitioning to yield two fractions as ethyl acetate and aqueous. Methanol extract was further fractioned over Sephadex LH-20 column. Antioxidant activity of extracts and fractions were screened using ferric thiocyanate and DPPH radical scavenging methods. Phenolic composition of active fraction(s) was determined by HPLC–MS systems. Those fractions which exhibited strong antioxidant activity were rich in anthocyanins and hydrolysable tannins. While gallic acid was the main phenolic acid in the extracts, anthocyanin fraction contained cyanidin, peonidin, pelargonidin, petunidin, and delphinidin glucosides and coumarates. Pentagalloyl glucose was abundant in the hydrolysable tannin fraction. Effective scavenging concentration (EC₅₀) on DPPH radical was 0.70 μg/mL both in ethyl acetate and tannin fractions, and 5.33 μg/mL in anthocyanin rich fraction. Same extracts and fractions showed moderate lipid peroxidation inhibition effect compared with the synthetic antioxidants. The findings demonstrate that sumac can be used as a natural antioxidant.     

Phenolics from hull of Garcinia mangostana fruit and their antioxidant activities

The air-dried fruit hulls of Garcinia mangostana Linn. were extracted with 70% MeOH, and then partitioned into the n-BuOH fractions. Furthermore, three major phenolic components related to their antioxidant activities were purified by silica gel column chromatography and Sephadex LH-20 and then identified as P₁ (1,3,6,7-tetrahydroxy-2,8-(3-methyl-2-butenyl)), P₂ [1,3,6-trihydroxy-7-methoxy-2,8-(3-methyl-2-butenyl) xanthone] and P₃ (epicatechin) using UV–visible spectrophotometry, IR spectrophotometry and NMR spectroscopy, respectively. The antioxidant activities of three major phenolics were evaluated using different tests, including the free radical scavenging capability and total antioxidant activity in a linoleic acid peroxidation. These three phenolic compounds exhibited different antioxidant activities in these antioxidant tests. The hydroxyl radical and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capabilities and the activity against linoleic acid peroxidation of P₁ were greater than those of P₂ and P₃, while the superoxide anion radical scavenging activity of P₃ was greater than that of P₁, but was close to that of P₂ or α-tocopherol. An activity-guided isolation and purification process was used to identify the components showing the strong DPPH radical scavenging activity from G. mangostana Linn.     

Evaluation of the antioxidant activity of extracts from buntan (Citrus grandis Osbeck) fruit tissues

The goal of the present work was to evaluate the antioxidant properties of buntan (Citrus grandis Osbeck) using various solvents, such as n-hexane, ethyl acetate (EtOAc), butanol and methanol. The antioxidant activities of crude extracts were evaluated by using the free radical scavenging β-carotene assay and total polyphenol. Ethyl acetate extracts of falvedo exhibited high antioxidative activities, followed by albedo and segment membrane extracts. Chromatography separation of EtOAc extract of flavedo using a silica gel column, yielded six fractions (A, B, C, D, E and F) using gradient elution with benzene and acetone (19:1, 14:1, 9:1, 5:1, 1:1 and 0:l). Among them, two fractions (C and D) showed strong antioxidant activities using the free radical scavenging activity (DPPH) antioxidant assay. These two fractions were further purified using silica gel column chromatography and preparative TLC. Their extracts could well be useful to prevent oxidation in fruit juices and essential oil food products as well as for health supplements. Identification of the responsible components is underway.     

Antioxidant activity of compounds isolated from the pyroligneous acid, Rhizophora apiculata

The polyphenols (CPAE II) was isolated from the dichloromethane extract of the pyroligneous acid, Rhizophora apiculata by simultaneous acid base and solvent extraction method. Its qualitative and quantitative composition was studied by gas chromatography mass spectroscopy (GC/MS) and out of 57 peaks, 52 compounds were identified, representing 95.47% of the total polyphenols. The CPAE II was then fractionated to four fractions (F1–F4) by means of thin layer chromatography and silica gel column chromatography with dichloromethane, dichloromethane/chloroform/ethyl acetate mixture (8:1:1; 4:3:3, v/v/v), and ethyl acetate, respectively. The antioxidant properties of the CPAE II and the fractions were evaluated. Among the four fractions, fraction 1 (F1) was the most potent in DPPH radical scavenging activity and molybdenum (VI) reducing power. It was subjected to further purification by means of silica gel column chromatography with hexane, hexane/diethyl ether mixture (9:1, 6:1, 3:1, v/v), and diethyl ether, respectively. 2,6-Dimethoxyphenol (syringol) and dihydroxybenzenes (catechol and 3-methoxycatechol) were isolated and identified by GC/MS, ¹H NMR, ¹³C NMR spectral analyses, and confirmed by GC co-injection with authentic standards. Syringol, catechol and 3-methoxycatechol constitute 39.08, 4.21 and 1.10% of F1, respectively. Their antioxidant activities were evaluated by DPPH radical scavenging activity, ABTS radical cation scavenging activity, phosphomolybdenum and ferric reducing antioxidant power (FRAP). The trend in antioxidant capacity was similar in all the four assays, with dihydroxybenzenes > 2,6-dimethoxyphenols, although discrepancies in the ranking within the dihydroxybenzenes were present. These three compounds which showed significant antioxidant activities were isolated for the first time from the pyroligneous acid, R. apiculata.     

Structural identification of isomallotusinin and other phenolics in Phyllanthus emblica L. fruit hull

The air-dried fruit hull of Phyllanthus emblica L. was extracted with 95% ethanol, and then the extract was partitioned by diethyl ether and ethyl acetate (EA). The EA fraction was then subjected to separation and purification using silica gel and Sephadex LH-20 column chromatography repeatedly to obtain five hydrolysable tannins. They were identified as mucic acid 1,4-lactone 3-o-gallate (C1), isocorilagin (C2), chebulanin (C3), chebulagic acid (C4) and isomallotusinin (C5) using mass spectrometry and nuclear magnetic resonance (NMR) spectrometry. Isomallotusinin and chebulanin were identified from emblica dried fruit hull for the first time, and isomallotusinin was the first time identified from Phyllanthus. Furthermore, the antioxidant abilities of these hydrolysable tannins were investigated using DPPH and ABTS⁺ radical scavenging systems. All hydrolysable tannins showed strong DPPH and ABTS⁺ radical scavenging activities. Isomallotusinin and chebulagic acid exhibited the highest antioxidant activity compared to other purified compounds tested.     

Free‐radical scavenging activity of wormwood (Artemisia absinthium L) extracts

In an effort to discover new antioxidant natural compounds, wormwood (Artemisia absinthium L) an aromatic‐bitter herb, was screened. The sequential extraction was realized with five solvents of different polarities (70% methanol, petroleum ether, chloroform, ethyl acetate, n‐butanol). The antioxidative activity was tested by measuring their ability to scavenge stable 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) free radical and reactive hydroxyl radical during the Fenton reaction trapped by 5,5‐dimethyl‐1‐pyrroline‐N‐oxide (DMPO), using electron spin resonance (ESR) spectroscopy. Results demonstrated that the antiradical and antioxidative activity depend on the type and concentration of applied extracts and increased in the order ethyl acetate > methanol > n‐butanol > chloroform > petroleum ether > remaining water extracts. The investigation showed that the antiradical activity increased with increasing concentration of all extracts. The high contents of total phenolic compounds (25.6 mg g^?1) and total flavonoids (13.06 mg g^?1) indicated that these compounds contribute to the antiradical and antioxidative activity. In a model system, the formation of o‐semiquinone radicals from quercetin and chlorogenic acid was obtained to prove the mechanism (hydrogen donating and/or one‐electron reduction) of free‐radical scavenging activity. Copyright © 2004 Society of Chemical Industry     

Neuroprotective potential of some terebinth coffee brands and the unprocessed fruits of Pistacia terebinthus L. and their fatty and essential oil analyses

In the current study, neuroprotective effects of the ethyl acetate and methanol extracts of four terebinth coffee brands and the fruits of Pistacia terebinthus L. were investigated through enzyme inhibition tests against acetylcholinesterase, butyrylcholinesterase, and tyrosinase as well as antioxidant test systems. Antioxidant activity was measured using radical scavenging activity tests and metal-related tests including metal-chelation capacity, ferric-reducing antioxidant power (FRAP), and phosphomolybdenum reducing power (PRAP). The fatty oils of the coffee brands and the fruits and the fruit essential oil were examined by GC–MS. Total phenol and flavonoid contents were calculated spectrophotometrically. The extracts had moderate inhibition against butyrylcholinesterase (9.78–45.74% at 200 μg mL⁻¹) and potent scavenging activity against DPPH. They exerted strong activity in FRAP and metal-chelation tests and modest activity in PRAP test. Oleic acid was identified as the major fatty acid in the fatty oils, while α-pinene (26.31%) was dominant in the essential oil.     

HPLC–MS analyses and bioactivities of novel chemicals in Devil’s club (Oplopanax horridus (Sm.) Miq.)

Devil’s club (Oplopanax horridus (Sm.) Miq.) has traditionally been used as a folk medicine by native North Americans for the treatment of various chronic diseases. A methanolic extract and its sub-fractions by liquid–liquid re-extraction, i.e., chloroform (CHCl₃), ethyl acetate (EtOAc), n-butanol (BuOH), and water fractions, were analysed by their anti-cancer activity with in vitro cell proliferative bioassays, and their antioxidant capacities in terms of the determination of total phenolic content (TPC), ORAC value, and DPPH free radical scavenging activity. The whole extract of the dried root contained high TPC and possessed strong ORAC and DPPH radical scavenging activities. In addition, the extract exhibited a strong anti-proliferative ability against HT-29 cancer cells, e.g., with an inhibitive rate of 56.5% with a 0.2-mg/mL extract. Further investigation by HPLC–UV–MS identified significant bioactive phenolic compounds in the chloroform fraction, including gallic acid, caffeic acid, 4-O-feruloylquinic acid, 5-O-feruloylquinic acid, ferulic acid, methyl feruloylquinate, methyl ferulate, and quercetin. These results suggested that the associated bioactivity of the plant might result from the contribution of phenolic compounds.     

Analysis of antioxidant activity and antioxidant constituents of Chinese toon

Antioxidant activity of the methanol and water extracts of Chinese toon (Toona sinensis) leaf was evaluated using DPPH radical scavenging and lipid peroxidation assays. Contents of four major types of antioxidants including β-carotene, ascorbate, α-tocopherol and phenolics were also quantified. Open column chromatography followed by semi-preparative HPLC were applied to separate phenolic antioxidants whose contents were subsequently determined by HPLC. The methanol extract demonstrated much higher antioxidant activity than the water extract. Contents of β-carotene, ascorbate, α-tocopherol and phenolics were 1.23 μmol g^?1, 34.2 μmol g^?1, 2.40 μmol g^?1 and 872 μmol gallic acid equivalents g^?1, respectively. Six phenols were isolated. Their structures were characterized as 5-O-galloylquinic acid, gallic acid, methyl gallate, β-1,2,6-tri-O-galloyl-d-glucose, quercetin 3-O-β-d-glucopyranoside and quercetin 3-O-(2′′-O-galloyl)-β-d-glucopyranoside, respectively. The results indicate that phenolic compounds are the dominant antioxidants in Chinese toon. The compounds β-1,2,6-tri-O-galloyl-d-glucose and quercetin 3-O-(2′′-O-galloyl)-β-d-glucopyranoside were reported for the first time in Chinese toon.     

Antioxidative principals of Jussiaea repens: an edible medicinal plant

Jussiaea repens L. (JRL) is an edible medicinal plant and is also used as a vegetable by the local people in southwestern China. The crude extract and its four fractions derived from JRL were evaluated for the 1,1‐diphenyl‐2‐picrylhydrazyl radical‐scavenging ability, hydroxyl radical‐scavenging capacity and the potassium ferricyanide reduction property. The ethyl acetate‐soluble fraction (EAF) and EAF6 (a subfraction derived from EAF) were the most valuable fraction and subfraction, respectively. Furthermore, bioactivity‐guided chromatographic fractionation revealed that three pure compounds greatly contributed to the antioxidant activities. Qualitative and quantitative analyses of the major antioxidant constituents in the extract were systematically conducted by NMR, mass spectral analyses and RP‐HPLC. The result demonstrated that rosmarinic acid (2.00 mg g^?1 JRL dry weight) quercetin 3‐O‐β‐d ‐glucopyranoside (9.88 mg g^?1 JRL dry weight), and kaempferol 3‐O‐β‐d ‐glucopyranoside (1.85 mg g^?1 JRL dry weight) were the major antioxidative constituents in JRL. These compounds are reported for the first time from this plant.     

Evaluation of antioxidant activities of various solvent extracts of Carissa opaca fruits

The chloroform and aqueous fractions of Carissa opaca fruit, a traditional medicinal fruit in Pakistan possessed a high amount of total phenolic and flavonoid contents as compare to other solvent fractions with potent antioxidant activities in scavenging DPPH, superoxides, hydroxyl, hydrogen peroxide, ABTS radicals, and had strong iron chelating activity. On the other hand, the ethyl acetate fraction showed the highest inhibition of β-carotene linoleic acid peroxidation and phosphomolybdate assay. A high correlation coefficient existed between EC₅₀ values of DPPH, superoxides, hydroxyl, hydrogen peroxide, ABTS radicals, total phenolics and flavonoids, but a non significant correlation was found in the case of iron chelaters, β-carotene and phosphomolybdate assay. This study verified that the chloroform and aqueous fractions have strong antioxidant activities which were correlated with its high level of phenolics and flavonoids. These fractions can be used as a source of potential antioxidant or functional food material.     

Rapid screening and identification of antioxidants in aqueous extracts of Houttuynia cordata using LC–ESI–MS coupled with DPPH assay

Houttuynia cordata Thunb. has been used traditionally as immune stimulant and anticancer agent. An aqueous extract of H. cordata tea showed high radical scavenging activity determined by off-line DPPH assays. Then, it was screened for its antioxidant components via an on-line DPPH radical scavenging technique coupled with a liquid chromatography–electrospray ionization mass spectrometer (LC–ESI–MS). Based on their mass spectra and fragmentation patterns; the antioxidant compounds were identified as quinic acid derivative, caffeic acid derivatives, procyanidin B, neo-chlorogenic acid, catechin, chlorogenic acid, crypto-chlorogenic acid and quercetin hexoside. LC–MS/MS in multiple reactions monitoring (MRM) mode was used to quantify these antioxidant compounds. Chlorogenic acid was found to be a major component in H. cordata tea.     

番石榴叶乙酸乙酯萃取物的体外抗氧化活性及化学成分的分离鉴定

本文研究了番石榴叶乙酸乙酯萃取物的体外抗氧化活性及活性物质基础。采用95%乙醇溶液浸泡提取、正己烷和乙酸乙酯萃取后得到粗提物,首先分析了乙酸乙酯萃取物的体外抗氧化活性,然后通过硅胶柱色谱、Sephadex LH-20凝胶柱色谱、质谱和核磁共振等技术对乙酸乙酯萃取物的化学成分进行了分离和鉴定。结果表明,乙酸乙酯萃取物对DPPH自由基、超氧阴离子、羟自由基具有较强的体外抗氧化活性,清除能力随着浓度的增大而增强,当浓度为1 mg/m L时,对DPPH自由基、超氧阴离子和羟自由基的清除率分别达94.15%、87.88%和67.3%,清除效果接近或强于同浓度的Vc。通过硅胶柱色普、Sephadex LH-20凝胶柱色谱等技术从番石榴叶乙酸乙酯萃取部位中分离纯化得到5个化合物,通过MS和NMR等波谱数据和结合文献,最终确定了4个化合物,分别为:去甲氧基荚果蕨(1)、胶藤素(2)、槲皮素(3)和异槲皮苷(4),其中化合物2为首次从番石榴属植物中分离得到的二氢黄酮类化合物。     

Isolation and Structure-Activity Relationship of the Antioxidant Chemical Constituents from the Flowers of Rosa chinensis Jacq

The objective of this study was to investigate the antioxidant activity, chemical constituents, and structure-activity relationship from the flowers of Rosa chinensis Jacq. The antioxidant activity of the different polar solvent extracts were assayed by a 1,1-diphenyl-2-picrylhydrazyl scavenging experiment. The extract with the strongest 1,1-diphenyl-2-picrylhydrazyl scavenging capacity was further isolated and purified by column chromatography. The structures of the isolated compounds were elucidated on the basis of spectral analysis and physiochemical properties. The antioxidant activity of these compounds was assayed by a 1,1-diphenyl-2-picrylhydrazyl scavenging experiment and ascorbic acid was chosen as the positive control. The ethyl acetate fraction had the strongest 1,1-diphenyl-2-picrylhydrazyl scavenging activity. Ten compounds were obtained, represented as: 3,4,8,9,10-pentahydroxydibenzo[b,d]pyran-6-one (1), quercetin (2), kaempferol (3), 3,5,7,4′-tetrahydroxy-8-methoxy-flavone (4), isoquercetrin (5), kampferol 3-O-β-D-glucoside (6), tiliroside (7), kampferol 3-O-(6′′-galloyl)-β-D-glucoside (8), kampferol 3-O-(2′′,6′′-digalloyl)-β-D-glucoside (9), quercetin 3-O-(2′′,6′′-digalloyl)-β-D-glucoside (10). The 1,1-diphenyl-2-picrylhydrazyl scavenging capacity for these compounds, in descending order, were as follows: 10 > 9 > 1 ≈ 8 > 5 > ascorbic acid ≈ 2 > 6 > 7 > 3 > 4. Compounds 1, 8, 9, and 10 have the stronger antioxidant activity. Our results suggested the antioxidant activities of these compounds might be influenced by the number and position of hydroxyl groups in their aromatic rings.     

HPLC测定余甘子茶中3种多酚成分及抗氧化活性研究

福林酚法测定五个产地的余甘子茶中总多酚(TP)的含量,高效液相色谱法(High performance liquid chromatography,HPLC)分析没食子酸(GA)、柯里拉京(CO)、鞣花酸(EA)的含量,并对其抗氧化活性进行研究。结果表明:TP含量最高的是凉山的余甘子茶,其值为228 mg/g;漳州的余甘子茶中GA、CO、EA含量最高,分别为5.00、28.76、4.30 mg/g;在抗氧化活性研究中,昆明的余甘子茶提取液总还原能力较高,但凉山的余甘子茶提取液对DPPH自由基(DPPH·)和羟自由基(·OH)清除能力较好,EC₅₀分别为7.46 μg/mL和0.44 mg/mL,其对自发性肝脂质氧化抑制能力好于其它产地。相比较而言,凉山产地的余甘子茶作为开发天然抗氧化剂更具优势。