Chemical composition and immuno-stimulating properties of polysaccharide biological response modifier isolated from Radix Angelica sinensis

To investigate the immunomodulating activity of Radix Angelica sinensis, three different Radix A. sinensis polysaccharide fractions (APFs, namely APF1, APF2 and APF3) were isolated by fractionation using gel filtration and were identified as the immunomodulators of murine peritoneal macrophages. In the present study, it was found that various APFs induced a significant increase in cellular lysosomal enzyme activity, nitric oxide (NO) formation, reactive oxygen species (ROS) production and tumor necrosis factor-α (TNF-α) secretion in macrophages in vitro. Furthermore, APFs dose-dependently stimulated macrophages to produce NO through the up-regulation of inducible NO synthase (iNOS) activity and the maximal effect occurred at a concentration of 500 μg/ml by APF2, followed by APF3 and APF1 in decreasing order. Moreover, the predominant sugars in various APFs were identified as rhamnose, arabonose, glucose, galactose and the ratio of these monosaccharides differed from one polysaccharide fraction to another, which also affected the cellular effector molecule production in macrophages.     

Phenolic antioxidants from Chinese toon (fresh young leaves and shoots of Toona sinensis)

Chinese toon (the fresh young leaves and shoots of Toona sinensis), known as a tree vegetable, belongs to the family of Meliaceae. It is one of the most popular vegetables in China. The 80% acetone extract of Chinese toon exhibited considerable antioxidant activity in the 1,1-diphenyl-2-picryldydrazyl (DPPH) radical-scavenging assay. Bioassay-guided purification of the extract led to the isolation of 12 phenolic compounds (1–12), whose structures were determined by detailed spectroscopic analyses, including UV, IR, MS, 1D and 2D NMR techniques. Of them, compounds 3–8 and 11 were isolated from T. sinensis for the first time. All of the isolated compounds were examined for their antioxidant activities by the DPPH method, and the results showed that gallic acid and its derivatives, gallotannins and flavonoids were the main constituents contributing to the antioxidant activity of Chinese toon. From the health point of view, Chinese toon is an ideal dietary vegetable with natural antioxidants.     

TLC bioautography-guided isolation of antioxidants from fruit of Perilla frutescens var. acuta

Guided isolation through bioautography on TLC using 1,1-diphenyl-2-picryl-hydrazyl radical (DPPH) as a detection reagent led to the isolation of four antioxidant compounds from fruit of Perilla frutescens var. acuta. These compounds were identified as rosmarinic acid (1), luteolin (2), apigenin (3), and chrysoeriol (4), by means of UV, NMR, and ESI MS. All the compounds were isolated for the first time from the fruit of the plant. Compounds 1 and 2 showed significant DPPH scavenging capacities, with IC₅₀ values of 8.61 and 7.50 μM, respectively. Further quantitative HPLC analysis confirmed that compounds 1-4 are the predominant contributors to the free radical scavenging activity of the extract of P. frutescens var. acuta.     

Activity-guided isolation and identification of free radical-scavenging components from an aqueous extract of Coleus aromaticus

An activity-directed fractionation and purification process was used to identify the DPPH (l,l-diphenyl-2-picrylhydrazyl) free radical-scavenging components of Coleus aromaticus Benth. Fresh leaves of C. aromaticus were extracted with water and then separated into hexane, ethyl acetate, and water fractions. Among these, only the ethyl acetate phase showed strong DPPH radical-scavenging activity in vitro, when compared with water and hexane phases. The ethyl acetate fraction was then subjected to separation and purification using Sephadex LH-20 chromatography. Three compounds showing strong DPPH radical-scavenging activity were shown, by spectral methods (¹H NMR, ¹³C NMR, and MS) and by comparison with literature values, to be rosmarinic acid, chlorogenic acid and caffeic acid. In addition, HPLC identification and quantification of isolated compounds were also performed. Rosmarinic acid was found as a major component and principally responsible for the radical-scavenging activity of C. aromaticus.     

Characterisation,extraction efficiency,stability and antioxidant activity of phytonutrients in Angelica keiskei

Phytonutrients in Angelica keiskei, a dark green leafy vegetable, were characterised and their extraction efficiency by different compositions of water/ethanol as well as stability at different temperatures was determined. A range in the content of lutein (205–265 mg/kg dry wt), trans-β-carotene (103–130 mg/kg dry wt), and total phenols (8.6–9.7 g/kg) was observed amongst Angelica keiskei grown in three different conditions. LC-ESI-MS/MS analysis identified chlorogenic acid, chalcones and the glucosides of luteolin and quercetin as the major phenolic compounds in Angelica keiskei. Only 46% of lutein was extracted by 70% of ethanol and no carotenoid was detected in 40% ethanol and 100% water extracts of Angelica keiskei. Major phytonutrients in Angelica keiskei were stable at −80 °C up to 12 months whilst β-carotene was significantly degraded at room temperature and 4 °C within 2 months and lutein at room temperature within 12 months.     

In vitro evaluation of antioxidant activities of aqueous extracts from natural and cultured mycelia of Cordyceps sinensis

Cordyceps sinensis, one of the best known traditional Chinese medicines and health foods, has been highly valued for the treatment of a wide range of diseases and reported to have antioxidant properties. In the present study, the antioxidant activities of hot-water extracts from natural and cultured mycelia of C. sinensis were investigated and evaluated using six in vitro assays, including inhibition of linoleic acid peroxidation; scavenging abilities on DPPH, hydroxyl and superoxide anion radicals; the reducing power and the chelating ability on ferrous ions. Among these assays, the extracts showed the best effect on the inhibition of linoleic peroxidation with the lowest IC50 values and with an inhibition rate over 90% at concentration of 0.8-1.6 mg/ml, more stable than that of α-tocopherol, a recognised natural antioxidant. The scavenging activities on superoxide anion and hydroxyl radicals of the two extracts were slightly lower than that of butylated hydroxytoluene. DPPH scavenging activities of both extracts reached over 80% inhibition at 4-8 mg/ml. Both extracts showed moderate reducing power and ferrous ion chelating activity. The IC50 value of the extract from cultured mycelia in all the tests, except for linoleic acid peroxidation, was significantly lower than that of natural mycelia. There was no evident correlation between the antioxidant activity and the content of protein, polysaccharides and mannitol of extracts from C. sinensis; the antioxidant activity may be due to a combined effect of these or some other compounds. These results suggested that both the extracts from cultured and natural mycelia have direct and potent antioxidant activities and that the cultured mycelia of the fungus could be used for the antioxidant activity to reduce the human demands on the natural resources of the fungus, an endangered species.     

Activity guided isolation of antioxidants from the leaves of Ricinus communis L.

An activity-guided isolation and purification process was used to identify the DPPH (l,l-diphenyl-2-picrylhydrazyl) free radical scavenging components of the food plant (Ricinus communis L.) of Eri silkworm. Dry leaves of R. communis L. were extracted with different solvents and tested for their antioxidant activity against DPPH^•. The MeOH:water (8:2) extract showed strong DPPH radical-scavenging activity, and was subjected to column chromatography over silica gel. Gallic acid, quercetin, gentisic acid, rutin, epicatechin and ellagic acid were isolated as active components and characterised by different spectroscopic techniques.     

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.     

Identification of phenolic compounds and appraisal of antioxidant and antityrosinase activities from litchi (Litchi sinensis Sonn.) seeds

Antioxidant and antitryrosinase compounds from Litchi sinensis Sonn. seeds were extracted with five different types of polar solvents. The five extracts, namely ethanol extract (EE), 50% ethanol extract (50% EE), methanol extract (ME), 50% methanol extract (50% ME), and water extract (WE), were used for the evaluations of total phenolic content, antioxidant capabilities and antityrosinase activity. The 50% EE showed the highest total antioxidant capacity, scavenging the 1,1-diphenyl-2-picryl hydrazyl (DPPH) radical and inhibitory activity against lipid peroxidation, and it was comparable to the activity of the synthetic antioxidant, butylated hydroxyl toluene. Fifty percent EE showed a better antityrosinase activity compared to the other extracts. After application of reverse phase high performance liquid chromatography, coupled to a diode array detector and electrospray ionisation mass spectra, five phenolic compounds, namely, gallic acid, procyanidin B2, (−)-gallocatechin, (−)-epicatechin and (−)-epicatechin-3-gallate were identified from 50% EE. This study suggests that litchi seed can potentially be used as a readily accessible source of natural antioxidants.     

Screening of the antioxidant potentials of six Salvia species from Turkey

This study was designed to examine the in vitro antioxidant activities of the methanol extracts of six Salvia species [Salvia caespitosa Montbret & Aucher ex Bentham (ENDEMIC), Salvia hypargeia Fisch. & Mey. (ENDEMIC), Salvia euphratica subsp. euphratica Montbret & Aucher ex Bentham (ENDEMIC), Salvia sclarea L., Salvia candidissima subsp. candidissima Montbret & Aucher ex Bentham and Salvia aethiopis L.] from Turkey. The extracts were screened for their possible antioxidant activities by two complementary test systems, namely DPPH free radical-scavenging and β-carotene/linoleic acid systems. Non-polar subfractions of the methanol extracts of Salvia species studied did not show any antioxidant activity in both test systems. In the first case, the most active plant was S. euphratica subsp. euphratica, an endemic species, with an IC₅₀ value of 20.7 ± 1.22 μg/ml, followed by S. sclarea (IC₅₀ = 23.4 ± 0.97 μg/ml) among the polar subfractions. In the β-carotene/linoleic acid test system, polar extract of S. hypargeia was superior to the polar extracts of other Salvia species studied (69.2% ± 1.90%). This activity was followed by S. sclarea with 63.5% ± 4.24% inhibition rate. The inhibition rate of the synthetic antioxidant, buthylated hydroxytoluene (BHT), was also determined to be 96%. Since the polar extracts of Salvia species dealt with here exhibited excellent antioxidant activities when compared to BHT, it seems possible to keep perishable fat-containing food longer by direct addition of an extract of sage.     

Structural characterisation and immunomodulatory property of an acidic polysaccharide from mycelial culture of Cordyceps sinensis fungus Cs-HK1

An acidic polysaccharide, designated AEPS-1, was fractionated from the exopolysaccharide (EPS) produced by a medicinal fungus Cordyceps sinensis Cs-HK1 in mycelial culture. The molecular structure of AEPS-1 was characterised and elucidated by spectral and chromatographic analyses, and through derivatization by periodate oxidation, Smith degradation and methylation. AEPS-1 was composed of glucopyranose (Glcp) and pyrano-glucuronic acid (GlcUp) in an 8:1 M ratio plus a trace amount of mannose, having an average molecular weight of about 36 kDa. AEPS-1 had a linear backbone of (1→3)-linked α-d-Glcp residues with two branches, α-d-Glcp and α-d-GlcUp, attached to the main chain by (1→6) glycosidic bonds at every seventh α-d-Glcp unit. Atomic force microscopy revealed that AEPS-1 formed large networks in water that are connected primarily with triple helical strands. In Raw264.7 macrophage cell cultures, AEPS-1, at suitable doses between 25 and 250 μg/ml, significantly stimulated the release of several major cytokines, demonstrating an immunomodulatory property.     

Antioxidant activity of Camellia sinensis leaves and tea from a lowland plantation in Malaysia

Methanol extracts of fresh tea leaves from a lowland plantation in Malaysia were screened for total phenolic content (TPC) and antioxidant activity (AOA). AOA evaluation included 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical-scavenging ability, ferric-reducing antioxidant power (FRAP), and ferrous-ion chelating (FIC) ability. Ranking, based on TPC and AOA, was as follows: shoots > young leaves > mature leaves. TPC and AOA of lowland leaves were comparable to those of highland plants. A green tea produced by drying young leaves in a household microwave oven for 4 min showed significantly higher TPC and AOA than did four commercial brands of green and black tea.     

Anti-inflammatory effects of compounds from Kaempferia parviflora and Boesenbergia pandurata

Kaempferia parviflora and Boesenbergia pandurata are perennial herbs in the Zingiberaceae family. The rhizomes of these two plants have been used as food ingredients and in Thai traditional medicine for treatment of several inflammatory-related diseases, such as gout, allergy, apthous ulcer and peptic ulcer. The compounds isolated from the rhizomes of K. parviflora and B. pandurata were, therefore, examined for their inhibitory activities against nitric oxide (NO) production. For K. parviflora, compound 5 (5-hydroxy-3,7,3′,4′-tetramethoxyflavone) exhibited the highest activity against the NO inhibitory effect, with an IC₅₀ value of 16.1 μM, followed by 4 (IC₅₀ = 24.5 μM) and 3 (IC₅₀ = 30.6 μM). Regarding the NO inhibitory activity of B. pandurata, compound 2 (panduratin A) displayed the most potent effect with an IC₅₀ value of 5.3 μM, followed by 3 (hydroxypanduratin A, IC₅₀ = 13.3 μM) and 7 (cardamonin, IC₅₀ = 24.7 μM), respectively. The 5-hydroxy-3,7,3′,4′-tetramethoxyflavone (5), panduratin A (2) and hydroxypanduratin A (3), were also tested on prostaglandin E₂ (PGE₂) and tumour necrosis factor-alpha (TNF-α) production. 5-Hydroxy-3,7,3′,4′-tetramethoxyflavone (5) exhibited a potent inhibitory effect on PGE₂ production (IC₅₀ = 16.3 μM), but a mild effect on TNF-α (IC₅₀ > 100 μM). Panduratin A and hydroxypanduratin A showed strong activity against PGE₂ with IC₅₀ values of 10.5 and 12.3 μM, respectively, and a moderate effect on TNF-α (IC₅₀ = 60.3 and 57.3 μM, respectively). This study indicated that compound 5 (5-hydroxy-3,7,3′,4′-tetramethoxyflavone) is responsible for anti-inflammatory activity of K. parviflora, while 2 (panduratin A) and 3 (hydroxypanduratin A), the prenylated chalcones, are responsible for that of B. pandurata.     

Simultaneous determination of free ergosterol and ergosteryl esters in Cordyceps sinensis by HPLC

The caterpillar-shaped Chinese medicinal mushroom Cordyceps sinensis consists of the fruiting body (CsA) and the host caterpillar (CsB). Ergosterol is a principal sterol in fungi and can indicate the level of mycelia of C. sinensis. Ergosterol is present in two forms, as free ergosterol and esterified ergosterol, which have different physiological functions. The relative abundances of free to esterified ergosterol are different among the various species. In the present study, a gradient reversed-phase high-performance liquid chromatography (HPLC) method was developed for the simultaneous determination of free and esterified ergosterol in CsA and CsB of C. sinensis. The results showed that CsA and CsB had similar ergosterol compositions, but the level of ergosteryl esters in CsB was much higher than that in CsA, indicating that CsA and CsB might be in different growth phase or have different physiological functions for the growth and multiplication of C. sinensis.     

Extraction of active ingredients from green tea (Camellia sinensis): Extraction efficiency of major catechins and caffeine

The effect of different extraction set-ups that influence the extraction efficiency of catechins and caffeine from green tea leaves (variety Fanning Belas, China) were studied using different aqueous and pure solvents (acetone, ethanol, methanol, acetonitrile, water), different temperatures (60, 80, 95 and 100 °C) and times (5–240 min). Raw extracts were analysed for contents of major catechins (EC, EGC, ECG, EGCG), caffeine, proanthocyanidins and flavonols (myricetin, caempherol, quercetin). Starting material was found to contain 191 g major catechins/kg material, 36 g caffeine/kg material and 5.2 g flavonols/kg material on a dry mass basis. The content of major catechins in green tea extracts varied from approximately 280–580 g/kg dry extract, with extraction efficiencies of major catechins varying from 61% to almost 100%. Content of caffeine in extract was in the range of 75 g/kg, where its extraction efficiency varied from 62% to 76%. Average extraction yield was 30% with exceptions when using pure acetone and acetonitrile, where extraction yield was about 3%. Contents of flavonols and proanthocyanidins were in the ranges 6–20 and 12–19 g/kg, respectively. Different extraction procedures with water were also investigated and optimal conditions determined: maximum achieved extraction efficiency of catechins with water was obtained at 80 °C after 20 min (97%) and at 95 °C after 10 min of extraction (90%). Degradation of catechins was observed at higher extraction temperatures and with prolonged extraction times. Using a lower ratio of solvent to material, extraction efficiencies were increased by applying a multi-step extraction procedure. Optimal extraction procedure was then performed using decaffeinated green tea leaves, which were obtained by high-pressure extraction with CO₂, when 98% of caffeine was selectively isolated without significant impact on valuable catechins.     

Isolation of some active compounds from Origanum vulgare L. ssp. vulgare and determination of their genotoxic potentials

Origanum vulgare L. (oregano) is a flavoring herb widely used around the world. In the present study, crude extract of O. vulgare L. (oregano) and its petroleum ether, chloroform, ethyl acetate, n-butanol, water fractions were prepared in order to isolate and investigate antimutagenic compounds from the active extract through the bacterial genotoxicity assay guided fractionation procedures. The methanol extract and its n-butanol fraction showed significant antimutagenic activity. In the end of sub-fractionation process of the n-butanol extract, luteolin 7-O-glucuronide and luteolin 7-O-xyloside were isolated. These compounds showed significant antimutagenic activity against 9-Aminoacridine and N-Methyl-N′-nitro-N-nitrosoguanidine mutagenicity. The inhibition rates ranged from 22.1% (luteolin 7-O-xyloside: Salmonella typhimurium TA1537 – 0.4 mM/plate) to 67.8% (luteolin 7-O-glucuronide: S. typhimurium TA1537 – 0.8 μM/plate). In conclusion, the results revealed that luteolin 7-O-glucuronide and luteolin 7-O-xyloside are two of the antimutagenic compounds of O. vulgare L. ssp. vulgare.