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1.  Polar extracts from (Tunisian) Acacia salicina Lindl. Study of the antimicrobial and antigenotoxic activities 
Methanolic, aqueous and Total Oligomer Flavonoids (TOF)-enriched extracts obtained from the leaves of Acacia salicina 'Lindl.' were investigated for antibacterial, antimutagenic and antioxidant activities.
The antimicrobial activity was tested on the Gram positive and Gram negative reference bacterial strains. The Mutagenic and antimutagenic activities against direct acting mutagens, methylmethane sulfonate (MMS) and 4-nitro-o-phenylenediamine (NOPD), and indirect acting mutagens, 2-aminoanthracene (2-AA) and benzo[a]pyrene (B(a)P) were performed with S. typhimurium TA102 and TA98 assay systems. In addition, the enzymatic and nonenzymatic methods were employed to evaluate the anti-oxidative effects of the tested extracts.
A significant effect against the Gram positive and Gram negative reference bacterial strains was observed with all the extracts. The mutagenic and antimutagenic studies revealed that all the extracts decreased the mutagenicity induced by B(a)P (7.5 μg/plate), 2-AA (5 μg/plate), MMS (1.3 mg/plate) and NOPD (10 μg/plate). Likewise, all the extracts showed an important free radical scavenging activity towards the superoxide anion generated by the xanthine/xanthine oxidase assay system, as well as high Trolox Equivalent Antioxidant Capacity (TEAC), against the 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS)+• radical. TOF-enriched extract exhibited the highest protective effect against free radicals, direct acting-mutagen and metabolically activated S9-dependent mutagens.
The present study indicates that the extracts from A. salicina leaves are a significant source of compounds with the antimutagenic and antioxidant activities, and this may be useful for developing potential chemopreventive substances.
PMCID: PMC3353866  PMID: 22490278
Acacia salicina; Antigenotoxic activity; Antioxidant activities; Ames assay
2.  Effects of Antrodia camphorata extracts on anti-oxidation, anti-mutagenesis and protection of DNA against hydroxyl radical damage 
Antrodia camphorata is a geographically special fungus and is one of the precious traditional medicines of Taiwan. A lot of reports have addressed its antioxidant activities and anticancer activities. In order to understand whether these protection effects were resulted from its ability of preventing DNA against hydroxyl radical damage, the A. camphorata extract was used to examine its antioxidant, antimutagenic and DNA-protective activities.
A. camphorata extract was prepared by extracting the lyophilized powder of A. camphorata mycelium with distilled water. The antioxidative activity of this A. camphorata extract was then evaluated by 2,2-diphenyl-1-picrylhydrozyl (DPPH) radical-scavenging assay, and the antimutagenic activities of the extract against direct mutagen 4-nitroquinoline N-oxide (4NQNO) and indirect mutagen benzo[a]pyrene (B[a]P) were evaluated by Ames test. The effects of the A. camphorata extract in terms of DNA protection against hydroxyl radical damage were also investigated.
It was found that the higher the concentration of A. camphorata extracts, the higher the DPPH radical-scavenging effect. A. camphorata extract at concentrations between 0.625 and 10 mg/ml was found to be neither toxic nor mutagenic. However, the higher A. camphorata concentration (10 mg/ml) used in the test showed higher inhibitory effects on 4NQNO in a dose-dependent manner. The A. camphorata extract also showed reducing and scavenging activities against superoxide anion radical and also exhibited protective effects on DNA against hydroxyl radical-induced damage.
Results suggested that A. camphorata is a non-toxic and novel material with antioxidant, antimutagenic and DNA-protective activities and could be developed into health foods.
PMCID: PMC4502644  PMID: 26179175
Antrodia camphorata; Submerged cultivation; Ames test; DNA damage; Antioxidant activity
3.  Phytochemical, Anti-oxidant and Anthelmintic activities of various leaf extracts of Flacourtia sepiaria Roxb 
The present study was carried out to investigate the phytochemical constituents, in vitro antioxidant potential and anthelmintic activities of Flacourtia sepiaria Roxb leaves.
The dried powdered leaves of Flacourtia sepiaria were extracted using petroleum ether, chloroform, ethyl acetate and methanol by a soxhlet extractor and preliminary phytochemical screening was performed using standard protocols. All the extract was evaluated for their potential antioxidant activities using test such as DPPH, superoxide anion radical, hydroxyl radical, nitric oxide radical scavenging abilities, ferrous chelating ability and total phenolic and flavanoid content. Anthelmintic activity of extract was screened in adult Indian earthworm model.
Preliminary screening revealed the presence of bioactive compounds especially phenolics, tannins and terpenoids in all extracts. The phenolic and flavanoid content was highest in methanolic extract and lowest in petroleum ether extract. The paralytic (9.46±0.212) and death time (31.43±0.148) of methanolic extract was found to be significant (P<0.05) when compared with paralytic (7.33±0.206) and death time (18.60±0.229) of standard piperazine citrate at 100 mg/mL concentration.
The results of the present study indicate that the leaf extracts of Flacourtia sepiaria exhibited strong antioxidant activity and possess significant anthelmintic activity and thus it is a good source of antioxidant and anthelmintic constituents.
PMCID: PMC3805094  PMID: 24093785
Antioxidant; Anthelmintic; Flacourtia sepiaria; Piperazine citrate.
4.  Phytochemical Analysis and Free Radical Scavenging Activity of Medicinal Plants Gnidia glauca and Dioscorea bulbifera 
PLoS ONE  2013;8(12):e82529.
Gnidia glauca and Dioscorea bulbifera are traditional medicinal plants that can be considered as sources of natural antioxidants. Herein we report the phytochemical analysis and free radical scavenging activity of their sequential extracts. Phenolic and flavonoid content were determined. Scavenging activity was checked against pulse radiolysis generated ABTS•+ and OH radical, in addition to DPPH, superoxide and hydroxyl radicals by biochemical methods followed by principal component analysis. G. glauca leaf extracts were rich in phenolic and flavonoid content. Ethyl acetate extract of D. bulbifera bulbs and methanol extract of G. glauca stem exhibited excellent scavenging of pulse radiolysis generated ABTS•+ radical with a second order rate constant of 2.33×106 and 1.72×106, respectively. Similarly, methanol extract of G. glauca flower and ethyl acetate extract of D. bulbifera bulb with second order rate constants of 4.48×106 and 4.46×106 were found to be potent scavengers of pulse radiolysis generated OH radical. G. glauca leaf and stem showed excellent reducing activity and free radical scavenging activity. HPTLC fingerprinting, carried out in mobile phase, chloroform: toluene: ethanol (4: 4: 1, v/v) showed presence of florescent compound at 366 nm as well as UV active compound at 254 nm. GC-TOF-MS analysis revealed the predominance of diphenyl sulfone as major compound in G. glauca. Significant levels of n-hexadecanoic acid and octadecanoic acid were also present. Diosgenin (C27H42O3) and diosgenin (3á,25R) acetate were present as major phytoconstituents in the extracts of D. bulbifera. G. glauca and D. bulbifera contain significant amounts of phytochemicals with antioxidative properties that can be exploited as a potential source for herbal remedy for oxidative stress induced diseases. These results rationalize further investigation in the potential discovery of new natural bioactive principles from these two important medicinal plants.
PMCID: PMC3867356  PMID: 24367520
5.  Antioxidant activity of Coriandrum sativum and protection against DNA damage and cancer cell migration 
Coriandrum sativum is a popular culinary and medicinal herb of the Apiaceae family. Health promoting properties of this herb have been reported in pharmacognostical, phytochemical and pharmacological studies. However, studies on C. sativum have always focused on the aerial parts of the herb and scientific investigation on the root is limited. The aim of this research was to investigate the antioxidant and anticancer activities of C. sativum root, leaf and stem, including its effect on cancer cell migration, and its protection against DNA damage, with special focus on the roots.
Powdered roots, leaves and stems of C. sativum were extracted through sequential extraction using hexane, dichloromethane, ethyl acetate, methanol and water. Total phenolic content, FRAP and DPPH radical scavenging activities were measured. Anti-proliferative activitiy on the breast cancer cell line, MCF-7, was assayed using the MTT assay. Activities of the antioxidant enzymes, catalase, superoxide dismutase, glutathione peroxidase, and of the caspases-3, -8 and -9 were assayed on treatment with the extract. Cell cycle progression was analysed using flow cytometry. The scratch motility assay was used to assess inhibition of MCF-7 cell migration. DNA damage in 3 T3-L1 fibroblasts was evaluated by the comet assay. The components in the extract were identified by HPLC and GC-MS.
The ethyl acetate extract of C. sativum roots showed the highest antiproliferative activity on MCF-7 cells (IC50 = 200.0 ± 2.6 μg/mL) and had the highest phenolic content, FRAP and DPPH scavenging activities among the extracts. C. sativum root inhibited DNA damage and prevented MCF-7 cell migration induced by H2O2, suggesting its potential in cancer prevention and inhibition of metastasis. The extract exhibited anticancer activity in MCF-7 cells by affecting antioxidant enzymes possibly leading to H2O2 accumulation, cell cycle arrest at the G2/M phase and apoptotic cell death by the death receptor and mitochondrial apoptotic pathways.
This study is the first report on the antioxidant and anticancer properties of C. sativum root. The herb shows potential in preventing oxidative stress-related diseases and would be useful as supplements used in combination with conventional drugs to enhance the treatment of diseases such as cancer.
PMCID: PMC4028854  PMID: 24517259
Coriandrum sativum; Antioxidants; Anticancer activity; MCF-7; Hydrogen peroxide; Caspase; Cell cycle; DNA damage; Cancer cell migration; Comet assay
6.  Antimutagenicity of Methanolic Extracts from Anemopsis californica in Relation to Their Antioxidant Activity 
Anemopsis californica has been used empirically to treat infectious diseases. However, there are no antimutagenic evaluation reports on this plant. The present study evaluated the antioxidant activity in relation to the mutagenic and antimutagenic activity properties of leaf (LME) and stem (SME) methanolic extracts of A. californica collected in the central Mexican state of Querétaro. Antioxidant properties and total phenols of extracts were evaluated using DPPH (1,1-diphenyl-2-picrylhydrazyl) and Folin-Ciocalteu methods, respectively. Mutagenicity was evaluated using the Ames test employing Salmonella enterica serovar Typhimurium strains (TA98, TA100, and TA102), with and without an aroclor 1254 (S9 mixture). Antimutagenesis was performed against mutations induced on the Ames test with MNNG, 2AA, or 4NQO. SME presented the highest antioxidant capacity and total phenolic content. None of the extracts exhibited mutagenicity in the Ames test. The extracts produced a significant reduction in 2AA-induced mutations in S. typhimurium TA98. In both extracts, mutagenesis induced by 4NQO or methyl-N′-nitro-N-nitrosoguanidine (MNNG) was reduced only if the exposure of strains was <10 μg/Petri dish. A. californca antioxidant properties and its capacity to reduce point mutations render it suitable to enhance medical cancer treatments. The significant effect against antimutagenic 2AA suggests that their consumption would provide protection against carcinogenic polycyclic aromatic compounds.
PMCID: PMC4135139  PMID: 25152760
7.  Piper betle shows antioxidant activities, inhibits MCF-7 cell proliferation and increases activities of catalase and superoxide dismutase 
Breast cancer is the most common form of cancer and the focus on finding chemotherapeutic agents have recently shifted to natural products. Piper betle is a medicinal plant with various biological activities. However, not much data is available on the anti-cancer effects of P. betle on breast cancer. Due to the current interest in the potential effects of antioxidants from natural products in breast cancer treatment, we investigated the antioxidant activities of the leaves of P. betle and its inhibitory effect on the proliferation of the breast cancer cell line, MCF-7.
The leaves of P. betle were extracted with solvents of varying polarities (water, methanol, ethyl acetate and hexane) and their phenolic and flavonoid content were determined using colorimetric assays. Phenolic composition was characterized using HPLC. Antioxidant activities were measured using FRAP, DPPH, superoxide anion, nitric oxide and hyroxyl radical scavenging assays. Biological activities of the extracts were analysed using MTT assay and antioxidant enzyme (catalase, superoxide dismutase, glutathione peroxidase) assays in MCF-7 cells.
Overall, the ethyl acetate extract showed the highest ferric reducing activity and radical scavenging activities against DPPH, superoxide anion and nitric oxide radicals. This extract also contained the highest phenolic content implying the potential contribution of phenolics towards the antioxidant activities. HPLC analyses revealed the presence of catechin, morin and quercetin in the leaves. The ethyl acetate extract also showed the highest inhibitory effect against the proliferation of MCF-7 cells (IC50=65 μg/ml). Treatment of MCF-7 cells with the plant extract increased activities of catalase and superoxide dismutase.
Ethyl acetate is the optimal solvent for the extraction of compounds with antioxidant and anti-proliferative activities. The increased activities of catalase and superoxide dismutase in the treated cells could alter the antioxidant defense system, potentially contributing towards the anti-proliferative effect. There is great potential for the ethyl acetate extract of P. betle leaf as a source of natural antioxidants and to be developed as therapeutics in cancer treatment.
PMCID: PMC3533855  PMID: 23153283
Piper betle; Antioxidant; Phenolic; MCF-7; Cytotoxicity; Catalase; Superoxide dismutase; HPLC
8.  Antioxidant and antigenotoxic activities of Angelica keiskei, Oenanthe javanica and Brassica oleracea in the Salmonella mutagenicity assay and in HCT116 human colon cancer cells 
BioFactors (Oxford, England)  2006;26(4):231-244.
Epidemiological studies indicate that consumption of green-yellow vegetables rich in chlorophyll, vitamin C, vitamin E, and carotenoids reduce the risk of cancer. We sought to examine the antigenotoxic and antioxidant properties of chlorophyll-rich methanol extracts of Angelica keiskei, Oenanthe javanica, and Brassica oleracea (kale). In the Salmonella mutagenicity assay, A. keiskei caused dose-dependent inhibition against three heterocyclic amine mutagens in the presence of S9, O. javanica was antimutagenic only at the highest concentration in the assay (2 mg/plate), and B. oleracea showed no consistent inhibitory activity at non-toxic levels. None of the extracts were effective against three direct-acting mutagens in the absence of S9. Extracts of A. keiskei and, to a lesser extent O. javanica, inhibited two of the major enzymes that play a role in the metabolic activation of heterocyclic amines, based on ethoxyresorufin-O-deethylase and methoxyresorufin-O-demethylase assays in vitro. All three plant extracts were highly effective in assays which measured ferric reducing/antioxidant power, oxygen radical absorbance capacity, and Fe2+/H2O2-mediated DNA nicking. Finally, using the ‘comet’ assay, all three plant extracts protected against H2O2-induced genotoxic damage in human HCT116 colon cancer cells. These findings provide support for the antigenotoxic and antioxidant properties of chlorophyll-rich extracts of A. keiskei, O. javanica, and B. oleracea, through mechanisms that include inhibition of carcinogen activation and scavenging of reactive oxygen species.
PMCID: PMC2267880  PMID: 17119270
Antimutagen; antioxidant; heterocyclic amines; phytochemical; comet assay; D NA breaks
9.  Total Phenolic Content and Antioxidant Activity of Morinda tinctoria Leaves 
The antioxidant activity and total phenolic content of Morinda tinctoria leaves was evaluated. The successively extracted leaves of Morinda tinctoria using various solvents was analyzed for their total phenolic content. The extracts were subjected to column chromatography for the isolation of bioactive molecules. In vitro antioxidant activity was evaluated by employing different assays, including 2,2-diphenyl-1-picrylhydrazyl, nitric oxide scavenging assay and phosphomolybdenum reducing power assay. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging efficacy of hexane extract is significant at higher concentration (500 μg/ml-91.2±0.05%) and the efficacy at lower concentration is more significant for ethyl acetate extract (100 μg/ml - 65.1±0.05%). The total phenolic content was highest in methanol extract (5.30±0.011 μg/mg). Cynarin, a hydroxy cinnamic acid was isolated from chloroform extract; oleuropein, a polyphenolic iridoid was isolated from methanol extract. The results obtained suggeted that Morinda tinctoria leaf extracts possessed antioxidant properties and might offer protection from free radicals. Two compounds, cynarin and oleuropein were reported for the first time from Morinda tinctoria leaves.
PMCID: PMC4442474  PMID: 26009658
DPPH activity; reducing power; nitric oxide; phenolic content; oleuropein; cynarin
10.  Isorhamnetin 3-O-robinobioside from Nitraria retusa leaves enhance antioxidant and antigenotoxic activity in human chronic myelogenous leukemia cell line K562 
In this report, the isorhamnetin 3-o-robinobioside and its original extract, the ethyl acetate extract, from Nitraria retusa leaves, were evaluated for their ability to induce antioxidant and antigenotoxic effects in human chronic myelogenous leukemia cell line.
Nitraria retusa products properties were carried out by firstly evaluating their effects against lipid peroxidation induced by H2O2, using the thiobarbituric acid reactive substances species (TBARS) assay, and proceeding to the assay of cellular antioxidant activity, then doing the comet assay.
The isorhamnetin 3-o-robinobioside showed a protective effect against lipid peroxidation induced by H2O2. The same natural compound and ethyl acetate extract inhibited oxidation induced by 2,2′-azobis (2-amidinopropane) dihydrochloride in human chronic myelogenous leukemia cells with respectively 50% inhibitory concentration values of 0.225 mg/ml and 0.31 mg/ml, reflecting a significant antioxidant potential. The same two products inhibited the genotoxicity induced by hydroxyl radicals in the same human cell line (by 77.77% at a concentration of 800 μg/ml and by 80.55% at a concentration of 1000 μg/ml respectively).
The isorhamnetin 3- o-robinobioside and its original extract, the ethyl acetate extract, from Nitraria retusa leaves, have a great antioxidant and antigenotoxic potential on human chronic myelogenous leukemia cell line K562.
PMCID: PMC3439276  PMID: 22913434
11.  Radical scavenging and singlet oxygen quenching activity of extracts from Indian seaweeds 
Free radicals and singlet oxygen are responsible for oxidative stress related diseases and many natural compounds are known to have antioxidant properties. In this study, extracts from brown and red seaweeds of Indian origin were evaluated for their ability to scavenge different radicals and quench singlet oxygen. The crude extract in methanol and its fractions in different solvents were evaluated for their activity. The methanol extract and its fractions from brown seaweed exhibited higher 2,2′-azinobis(3-ethylbenzothizoline-6-sulfonic acid) radical scavenging activity with more than 90% scavenging in butanol and ethyl acetate fractions and correlated with polyphenol content. There was a significant difference (p≤0.001) in hydroxyl radical scavenging activity between different fractions of the same seaweed. Among the crude extracts, extract from Gracilaria corticata showed the highest (14.0%) activity. Crude extract from brown seaweeds showed higher peroxyl radical scavenging activity compared to red seaweeds. In fractions from brown seaweed extracts, highest activity was observed in ethyl acetate fraction (>88%) followed by hexane fraction (>40 %). Ethyl acetate fraction from crude extract showed higher inhibitory activity against hemoglobin induced linoleic acid oxidation. Singlet oxygen quenching activity of the crude extract from brown seaweed was lower (<13%) compared to red seaweeds (16.4–20.5%).
PMCID: PMC3550984  PMID: 23572608
Seaweed; Polyphenol; Antioxidant; Free radical; Singlet oxygen
12.  Some Strychnos spinosa (Loganiaceae) leaf extracts and fractions have good antimicrobial activities and low cytotoxicities 
Strychnos spinosa Lam. is a deciduous tree used in traditional medicine to treat infectious diseases. This study is designed to determine the antimicrobial, antioxidant and cytotoxic activities of extracts and fractions from leaves of S. spinosa.
Extracts were obtained by maceration with acetone, methanol and dichloromethane/methanol (1/1) while fractions were prepared by liquid-liquid fractionation of the acetone extract. A broth serial microdilution method with tetrazolium violet as growth indicator was used to determine the minimum inhibitory concentration (MIC) against fungi, Gram-positive and Gram-negative bacteria. The antioxidant activity was determined using free-radical-scavenging assays, and the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide reduction assay was used to determine cytotoxicity.
Four extracts and five fractions had good to weak antimicrobial activity with MICs ranging from 0.04 to >1.25 mg/ml against both fungi and bacteria. The chloroform and ethyl acetate fractions had an MIC of 0.08 mg/ml against Aspergillus fumigatus. The n-butanol fraction had an MIC of 0.04 mg/ml against Cryptococcus neoformans. The hexane and chloroform fractions had an MIC of 0.08 mg/ml against Staphylococcus aureus. The antioxidant activities were much lower than that of the positive controls. Except for the alkaloid extract, all the extracts and fractions had free-radical-scavenging activity (IC50 ranging from 33.66 to 314.30 μg/ml). The cytotoxicity on Vero cells was reasonable to low with LC50 values ranging between 30.56 and 689.39 μg/ml.
The acetone extract and the chloroform fraction had the highest antibacterial activity. By solvent-solvent fractionation it was possible to increase the activity against A. fumigatus and to decrease the cytotoxicity leading to a potentially useful product to protect animals against aspergillosis. Our results therefore support the use of S. spinosa leaves in traditional medicine to treat infectious diseases.
PMCID: PMC4289352  PMID: 25428165
Extracts; Different polarity fractions; Antimicrobial; Antioxidant; Cytotoxicity; Selectivity index; Potentizing extracts
13.  Computer aided screening of Accacia nilotica phytochemicals against HCV NS3/4a 
Bioinformation  2013;9(14):710-714.
Background: HCV has become a leading cause of liver cirrhosis and hepatocellular carcinoma and is a major health concern worldwide. To date, there is no vaccine available in the market to tackle this disease, therefore there is a strong need to develop antiviral compounds that can target all genotypes of HCV with the same efficiency. Medicinal plants have low cost and are less toxic therefore, extracts of medicinal plants can serve as important antiviral agents against HCV. This study was designed to screen phytochemicals of Accacia nilotica to find a potent drug candidate that can inhibit HCV infection effectively.
Results: Docking of NS3/4A protease and Flavonoids of Accacia nilotica revealed that most of the flavonoids bound deeply with the active site of NS3/4A protease. Compound 01 showed a high ranking on docking score. All other compounds also showed reliable docking scores and had interactions with the binding cavity of NS3/4A protease, suggesting them as a potent drug candidate to block HCV replication.
Conclusion: To recognize binding interactions of Accacia nilotica phytochemicals with NS3/4A protease, molecular docking was performed to find potential inhibitor against NS3/4A protease of HCV. After post docking analysis, important interactions were found between active compounds and active site of NS3/4A protease. It can be concluded from the study that phytochemicals of Accacia nilotica may serve as a potential drug candidate with relatively simple structural changes against HCV NS3/4A protease.
PMCID: PMC3746092  PMID: 23976825
14.  Evaluation of antioxidant and anticancer properties of the seed extracts of Syzygium fruticosum Roxb. growing in Rajshahi, Bangladesh 
The use of plants and their derived substances increases day by day for the discovery of therapeutic agents owing to their versatile applications. Current research is directed towards finding naturally-occurring antioxidants having anticancer properties from plant origin since oxidants play a crucial role in developing various human diseases. The present study was designed to investigate the antioxidant and anticancer properties of Sygygium fruticosum (Roxb.) (abbreviated as SF).
The dried coarse powder of seeds of SF was exhaustively extracted with methanol and the resulting crude methanolic extract (CME) was successively fractionated with petroleum ether, chloroform and ethyl acetate to get petroleum ether (PEF), chloroform (CHF), ethyl acetate (EAF) and lastly aqueous (AQF) fraction. The antioxidant activities were determined by several assays: total antioxidant capacity assay, DPPH free radical scavenging assay, hydroxyl radical scavenging assay, ferrous reducing antioxidant capacity and lipid peroxidation inhibition assay. The in vivo anticancer activity of SF was determined on Ehrlich’s Ascite cell (EAC) induced Swiss albino mice.
All the extractives showed strong antioxidant activities related to the standard. The total antioxidant capacity (TAC) of the fractions was in the following order: EAF>AQF>CME>PEF>CHF. The TAC of EAF at 320 μg/mL was 2.60±0.005 which was significantly higher (p < 0.01) than that of standard catechin (1.37 ± 0.005). The ferrous reducing antioxidant capacity of the extracts was in the following order: EAF>AQF>CME>AA>CHF>PEF. In DPPH free radical scavenging assay, the IC50 value of EAF was 4.85 μg/mL, whereas that of BHT was 9.85 μg/mL. In hydroxyl radical scavenging assay and lipid peroxidation inhibition assay, the EAF showed the most potent inhibitory activity with IC50 of 43.3 and 68.11 μg/mL, respectively. The lipid peroxidation inhibition assay was positively correlated (p < 0 .001) with both DPPH free radical scavenging and hydroxyl radical scavenging assay. The total phenolic contents of SF were also positively correlated (p < 0 .001) with DPPH free radical scavenging, hydroxyl radical scavenging and lipid peroxidation inhibition assay. Based on antioxidant activity, EAF was selected for cytotoxic assay and it was found that EAF inhibited 67.36% (p < 0.01) cell growth at a dose of 50 mg/kg (ip) on day six of EAC cell incubation.
Our results suggest that EAF of seeds of SF possess significant antioxidant and moderate anticancer properties. Seeds of SF may therefore be a good source for natural antioxidants and a possible pharmaceutical supplement.
PMCID: PMC3691922  PMID: 23800021
Syzygium fruticosum Roxb; Myrtaceae; Free radicals; Polyphenolics; Antioxidant activity; Anticancer activity
15.  Protective effects of Quercus salicina on alloxan-induced oxidative stress in HIT-T15 pancreatic β cells 
The present study was designed to investigate the protective effect of hot water extracts from Quercus salicina leaves (QSWE) on alloxan-induced oxidative stress in HIT-T15 Syrian hamster pancreatic insulinoma cells. The HIT-T15 cells were treated with alloxan (1 mM) for 1 h and then co-incubated with the QSWE for 24 h. Alloxan significantly decreased the viability of the HIT-T15 cells (P<0.05). QSWE did not exhibit significantly cytotoxic effects and increased the viability of the HIT-T15 cells in a concentration-dependent manner. To further investigate the protective effects of QSWE on alloxan-induced oxidative stress in HIT-T15 cells, the cellular levels of reactive oxygen species (ROS), lipid peroxidation and endogenous antioxidant enzymes, including catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-px), were analyzed. QSWE decreased the intracellular levels of ROS and lipid peroxidation and increased the activity of antioxidant enzymes. These results suggest that QSWE exerted cytoprotective activity against alloxan-induced oxidative stress in HIT-T15 cells through the inhibition of lipid peroxidation, reduction of ROS levels and stimulation of antioxidant enzyme activity. In addition, QSWE also increased the insulin secretion activity of the alloxan-treated HIT-T15 cells.
PMCID: PMC3570244  PMID: 23408741
Quercus salicina; pancreatic β cells; cell viability; reactive oxygen species; insulin secretion
16.  In vitro analysis on bactericidal screening and antioxidant potentiality of leaf and root extracts of Thottea siliquosa (Lam.) Ding Hou. An ethnobotanical plant 
Natural products of plant origin are potential source of novel antimicrobial and antioxidative agents. Thottea siliquosa (Lam.) Ding Hou. (T. siliquosa). A medicinal herb used by local tribals for treating various ailments. The present study aims at the phytochemical screening, GC-MS analysis, in vitro antibacterial activity and antioxidant potentiality of root and leaf extracts of T. siliquosa.
Hot continuous Soxhlet extraction, GC-MS analysis, antibacterial analysis by disc diffusion, microdilution assay and antioxidant potentialities by hydroxyl radical and nitric oxide radical scavenging. The data was statistically analyzed.
Phytochemical screening of the ethyl acetate and methanolic extract of leaf and root revealed the presence of phenols, alkaloids, tannins and saponin. The extract revealed a pool of phytochemicals by comparison with authentic standards from spectral library. Both the extracts has shown their broad spectrum of inhibition against the selected bacteria Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumonia compared with standard antibiotic drug streptomycin. The extracts showed antioxidant activity by scavenging of free radicals such as hydroxyl and nitric oxide. The IC50 values of the ethyl acetate extracts leaf and root and standard in this assay were 167.5±0.67, 99.4±1.2, 192±2.5 µg/mL respectively. Similarly those methanolic extracts of leaf and root were 269.5±0.89 and 289.1±2.66 µg/mL respectively. Similarly, ethyl acetate and methanolic extracts also caused a moderate dose-dependent inhibition of nitric oxide with an IC50 range 65.5±1.55 to 148 ±3.09 µg/mL. The inhibitory activities were found to be dose dependent.
The present study provides evidence that ethyl acetate and methanol extract of leaf and root of T. siliquosa are potential source of natural antioxidants and bactericidal nature. It is essential that research should continue to isolate and purify the bio active components of this natural plant and use in drug discovery and development.
PMCID: PMC3793157
Antibacterial activity; Antioxidant potentiality; Phytochemicals; Soxhlet extraction; Thottea siliquosa.
17.  A Study on Antimicrobial, Antioxidant and Antimutagenic Activities of Elaeagnus Angustifolia L. Leaves 
The aim of this work was to investigate the antimicrobial, antioxidant, and antimutagenic potentials of methanol extracts from E. angustifolia.
Materials and Methods
Methanol extracts were screened for antimicrobial activity against different species of 4 Gram positive and 3 Gram negative bacteria and one fungus. These bacteria included food pathogens. The leaf extract was tested using disc diffusion assay.
The methanol extract of E. angustifolia showed maximum inhibition zone of 16 mm against Yersinia enterocolitica. Whereas, the inhibition zone was not determined by methanol extract against Escherichia coli ATCC 1122 and Candida albicans RSKK 02029. The MIC was evaluated on plant extracts as antimicrobial activity. All of bacterial strains showed the lowest sensitivity to methanol extract of E. angustifolia (3.5 mg/mL), except Yersinia enterocolitica NCTC 11174. In addition, the plant extracts were tested against the stable DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate) free-radical. Finally, the methanol extract displayed a strong antioxidant activity (Trolox equivalent = 1.49 mM). Also, E. angustifolia methanol extracts were screened for their antimutagenic activity against sodium azide by Ames test in absence of rat microsomal liver enzyme (-S9). The results showed that E. angustifolia methanol extracts can inhibit mutagenic agents of sodium azide. The plant leaf extracts with the inhibition of 36% sodium azide showed moderate potential in decreasing mutagenic agents in Salmonella typhimurium TA100.
E. angustifolia methanol extracts have antimicrobial, antioxidant and antimutagenic potential.
PMCID: PMC3957251  PMID: 24653563
Elaeagnus; antimicrobial activity; antioxidant activity; antimutagenic activity
18.  In vitro Antioxidant and Pharmacognostic Studies of Leaf Extracts of Cajanus cajan (L.) Millsp 
Cajanus cajan (L.) Millsp is one of the second most dietary legume crops. The leaf extracts may be used as a potential source of natural antioxidant. The ash values, extractive values, total phenolic and flavonoid content, in vitro antioxidant activity of various leaf extracts as well as anatomical investigation of Cajanus cajan were carried out. Physicochemical parameters such as total, acid-insoluble and water-soluble ash values and moisture content of the leaf powder of C. cajan were found to be 9.50%, 1.40 g/100 g, 4.15 g/100 g drug and 6.72%, respectively. Percent yield of acetone, aqueous, ethanol, ethyl acetate and chloroform leaf extracts were 9.0, 10.6, 13.75, 8.7 and 5.8 g/100 g, respectively. Significant amount of phenolic and flavonoid content were observed. The results of the antioxidant activity were found to be concentration-dependent. The IC50 values for DPPH assay determined for aqueous and ethanol extracts were 0.69 and 0.79 mg/ml, respectively. Reducing power is increased with increasing amount of concentration in both aqueous and ethanol leaf extracts. The highest hydroxyl radical scavenging activity reached up to 83.67% in aqueous and 78.75% in ethanol extracts and in phosphomolybdenum assay the aqueous extract showed strong antioxidant capacity up to 55.97 nM gallic acid equivalents/g. It was found that the aqueous extract possessed highest antioxidant activity in all the assays tested. The antioxidant characteristics of leaf extracts are possibly because of the presence of polyphenols. Microscopic study showed the presence of collenchyma, fibres, xylem, phloem, epidermis, trichomes, palisade tissue, basal sheath, pith and cortex in leaf, petiole and pulvinus.
PMCID: PMC4442465  PMID: 26009649
Cajanus cajan; total phenolics; flavonoid; antioxidant; free radicals; hydroxyl radical scavenging activity
19.  Antioxidant activity and mechanism of Rhizoma Cimicifugae 
As a typical Chinese herbal medicine, rhizoma Cimicifugae (RC, 升麻 in Chinese) possesses various pharmacological effects involved in antioxidant activity. However, its antioxidant activity has not been reported so far. The aim of the present study was to systematically evaluate the antioxidant ability of RC in vitro, then discuss the mechanism.
Firstly, five RC extracts (i.e. petroleum ether extract PERC, ethyl acetate extract EARC, absolute ethanol extract AERC, 95% ethanol extract 95ERC, and water extract WRC) were prepared and determined by various antioxidant methods, including anti-lipidperoxidation, protection against DNA damage, ·OH scavenging, ·O2- scavenging, DPPH· (1,1-diphenyl-2-picryl-hydrazl radical) scavenging, ABTS+· (2,2’-azino-bis (3-ethylbenzo- thiazoline-6-sulfonic acid radical ion) scavenging, Cu2+-chelating, and Fe3+ reducing assays. Subsequently, we measured the chemical contents of five RC extracts, including total phenolics, total saponins, total sugars, caffeic acid, ferulic acid and isoferulic acid. Finally, we quantitatively analyzed the correlations between antioxidant levels (1/IC50 values) and chemical contents.
In the study, the antioxidant levels and chemical contents (including total phenolics, total saponins, total sugars, caffeic acid, ferulic acid and isoferulic acid) of five RC extracts were determined by various methods. In all antioxidant assays, five RC extracts increased the antioxidant levels in a dose-dependent manner. However, their antioxidant levels (IC50 values) and chemical contents significantly differed from each other. Quantitative analysis of the correlation showed that total phenolic was of significant positive correlations (average R value was 0.56) with antioxidant levels; In contrast, total sugars and total saponins had no positive correlation with antioxidant (the average R values were −0.20 and −0.26, for total sugars and total saponins, respectively); Among total phenolics, three phenolic acids (caffeic acid, ferulic acid and isoferulic acid) also displayed positive correlations (the average R values were 0.51, 0.50, and 0.51, for caffeic acid, ferulic acid and isoferulic acid, respectively).
As an effective antioxidant, Rhizoma Cimicifugae can protect DNA and lipids against oxidative damage. Its antioxidant ability can be responsible for its various pharmacological effects and may be mainly attributed to the existence of total phenolics, among which caffeic acid, ferulic acid and isoferulic acid are regarded as main bioactive components. Rhizoma Cimicifugae exerts its antioxidant effect through metal-chelating, and radical-scavenging which is via donating hydrogen atom (H·) and donating electron (e).
PMCID: PMC3557226  PMID: 23173949
Rhizoma Cimicifugae; 升麻; Antioxidant activity; Metal chelating; Radical-scavenging; Phenolic acid
20.  Assessment of phytochemicals, antioxidant, anti-lipid peroxidation and anti-hemolytic activity of extract and various fractions of Maytenus royleanus leaves 
Maytenus royleanus is traditionally used in gastro-intestinal disorders. The aim of this study was to evaluate the methanol extract of leaves and its derived fractions for various antioxidant assays and for its potential against lipid peroxidation and hemolytic activity.
Various parameters including scavenging of free-radicals (DPPH, ABTS, hydroxyl and superoxide radical), hydrogen peroxide scavenging, Fe3+ to Fe2+ reducing capacity, total antioxidant capacity, anti-lipid peroxidation and anti-hemolytic activity were investigated. Methanol extract and its derived fractions were also subjected for chemical constituents. LC-MS was also performed on the methanol extract.
Qualitative analysis of methanol extract exhibited the presence of alkaloids, anthraquinones, cardiac glycosides, coumarins, flavonoids, saponins, phlobatannins, tannins and terpenoids. LC-MS chromatogram indicated the composition of diverse compounds including flavonoids, phenolics and phytoestrogens. Methanol extract, its ethyl acetate and n-butanol fractions constituted the highest amount of total phenolic and flavonoid contents and showed a strong correlation coefficient with the IC50 values for the scavenging of DPPH, hydrogen peroxide radicals, superoxide radicals, anti-lipid peroxidation and anti-hemolytic efficacy. Moreover, n-butanol fraction showed the highest scavenging activity for ABTS radicals and for reduction of Fe3+ to Fe2+.
Present results suggested the therapeutic potential of Maytenus royleanus leaves, in particular, methanol extract, ethyl acetate and n-butanol fraction as therapeutic agent against free-radical associated damages. The protective potential of the extract and or fraction may be attributed due to the high concentration of phenolic, flavonoid, tannins and terpenoids.
PMCID: PMC3702440  PMID: 23800043
Maytenus Royleanus; Antioxidant Activities; Phenolic Content; Solvent Extraction
21.  Cytoprotective and antioxidant effects of phenolic compounds from Haberlea rhodopensis Friv. (Gesneriaceae) 
Pharmacognosy Magazine  2013;9(36):294-301.
Haberlea rhodopensis Friv. (Gesneriaceae) is a rare poikilohydric endemic and preglacial relict growing in Balkan Peninsula. Previous investigations demonstrated strong antioxidant, antimicrobial and antimutagenic potential of alcoholic extract from the plant.
The isolation of known caffeoyl phenylethanoid glucoside – myconoside and flavone-C-glycosides hispidulin 8-C-(2-O-syringoyl-β-glucopyranoside), hispidulin 8-C-(6-O-acetyl-2-O-syringoyl-β-glucopyranoside), and hispidulin 8-C-(6-O-acetyl-β-glucopyranoside) from the leaves of H. rhodopensis was carried out. The aim of this study was to investigate cyto-protective and antioxidant effects of isolated compounds.
Materials and Methods:
Antioxidant activity of isolated substances was examined using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) radicals; ferric reducing antioxidant power (FRAP) assay and inhibition of lipid peroxidation (LPO) in linoleic acid system by ferric thyocianate method. The compounds were investigated for their possible protective and antioxidant effects against tert-butyl hydroperoxide-induced oxidative stress in isolated rat hepatocytes. The levels of thiobarbituric acid reactive substances were assayed as an index of LPO. Lactate dehydrogenase leakage, cell viability, and reduced glutathione depletion were used as signs of cytotoxicity.
Myconoside demonstrated the highest DPPH radical scavenging, ABTS, FRAP, and antioxidant activity in linoleic acid system as well as the highest and statistically most significant protection and antioxidant activity against the toxic agent.
Phenolic compounds isolated from H. rhodopensis demonstrated significant cytoprotective, radical scavenging potential, and inhibit lipid peroxidation, moreover, myconoside was found to be a new powerful natural antioxidant.
PMCID: PMC3793333  PMID: 24124280
Antioxidant activity flavone-C-glycosides; cytoprotection; Haberlea rhodopensis; myconoside
22.  Antioxidative and Antimutagenic Activities of 70% Ethanolic Extracts from Four Fungal Mycelia-Fermented Specialty Rices 
The health-promoting potential of 70% ethanolic extracts of 4 rice varieties fermented with Monascus ruber, Phellinus linteus, Cordyceps sinensis and Agaricus blazei was evaluated mainly focusing on their antioxidative and antimutagenic capacities based on the following parameters: phenolic compound and phytic acid content; inhibitory activity on lipid peroxidation; scavenging activity on DPPH radical; suppressing ability on mitomycin C-induced mutagenesis in E. coli cells; and protective effect on 4-nitroquinoline oxide-triggered DNA lesion in V79 hamster cells. The fermented rice extracts were superior in overall health-promoting parameters compared to the source material. The higher antimutagenic activity of the fermented rice extracts might be in part caused by a larger amount of antioxidant constituents such as phenolic compounds or phytic acid. Of the fungal species, Monascus ruber was found to impart a marked increase in both the antioxidative and antimutagenic abilities to the source material. The current study suggests a possibility that such fermented rice may contribute to the prevention of lifestyle-related diseases such as cancer through a daily intake of rice-based diets.
PMCID: PMC2533716  PMID: 18818745
antioxidation; antimutagenesis; anticarcinogenesis; fermented rice; fungal mycelia
23.  In vitro protection of biological macromolecules against oxidative stress and in vivo toxicity evaluation of Acacia nilotica (L.) and ethyl gallate in rats 
Recently, enormous research has been focused on natural bioactive compounds possessing potential antioxidant and anticancer properties using cell lines and animal models. Acacia nilotica (L.) is widely distributed in Asia, Africa, Australia and Kenya. The plant is traditionally used to treat mouth, ear and bone cancer. However, reports on Acacia nilotica (L.) Wild. Ex. Delile subsp. indica (Benth.) Brenan regarding its toxicity profile is limited. Hence in this study, we investigated the antioxidant capacity and acute toxicity of ethyl gallate, a phenolic antioxidant present in the A. nilotica (L.) leaf extract.
The antioxidant activity of ethyl gallate against Fenton’s system (Fe3+/H2O2/ascorbic acid) generated oxidative damage to pBR322 DNA and BSA was investigated. We also studied the interaction of ethyl gallate to CT-DNA by wave scan and FTIR analysis. The amount of ethyl gallate present in the A. nilotica (L.) leaf extract was calculated using HPLC and represented in gram equivalence of ethyl gallate. The acute toxicity profile of ethyl gallate in the A. nilotica (L.) leaf extract was analyzed in albino Wistar rats. Measurement of liver and kidney function markers, total proteins and glucose were determined in the serum. Statistical analysis was done using statistical package for social sciences (SPSS) tool version 16.0.
Ethyl gallate was found to be effective at 100 μg/mL concentration by inhibiting the free radical mediated damage to BSA and pBR322 DNA. We also found that the interaction of ethyl gallate and A. nilotica (L.) leaf extract to CT-DNA occurs through intercalation. One gram of A. nilotica (L.) leaf extract was found to be equivalent to 20 mg of ethyl gallate through HPLC analysis. Based on the acute toxicity results, A. nilotica (L.) leaf extract and ethyl gallate as well was found to be non-toxic and safe.
Results revealed no mortality or abnormal biochemical changes in vivo and the protective effect of A. nilotica (L.) leaf extract and ethyl gallate on DNA and protein against oxidative stress in vitro. Hence, A. nilotica (L.) leaf extract or ethyl gallate could be used as potential antioxidants with safe therapeutic application in cancer chemotherapy.
PMCID: PMC4223376  PMID: 25043389
Acacia nilotica (L.); Ethyl gallate; Antioxidant; Oxidative damage; Toxicity; DNA interaction
24.  Antioxidant activities of ethanol extracts and fractions of Crescentia cujete leaves and stem bark and the involvement of phenolic compounds 
Antioxidant compounds like phenols and flavonoids scavenge free radicals and thus inhibit the oxidative mechanisms that lead to control degenerative and other diseases. The aim of this study was to investigate the antioxidant activity in vitro, total phenolic and flavonoid contents in ethanol extracts and fractions of Crescentia cujete leaves and stem bark.
Crescentia cujete leaves and bark crude ethanol extract (CEE) and their partitionates petroleum ether (PEF), chloroform (CHF), ethyl acetate (EAF) and aqueous (AQF) were firstly prepared. Different established testing methods, such as 1, 1-diphenyl-2-picryl hydrazyl (DPPH) radical, ferric reducing power (FRP), and total antioxidant capacity (TAC) assays were used to detect the antioxidant activity. Further, the total yield, total phenolic (TPC) and total flavonoid contents (TFC) of CEE and all the fractions were determined. Ethanol extracts of both leaves and stem bark were also subjected to preliminary phytochemical screening to detect the presence of secondary metabolites, using standard phytochemical methods (Thin layer chromatography and spray reagents).
Phytochemical screening of crude ethanol extract of both leaves and stem bark revealed the presence of steroids, flavonoids, saponins, tannins, glycosides and terpenoids. All the fractions and CEE of leaves and bark exhibited antioxidant activities, however, EAF of leaves showing the highest antioxidant activity based on the results of DPPH, FRP and TAC assay tests. The above fraction has shown the significant DPPH scavenging activity (IC50 = 8.78 μg/ml) when compared with standard ascorbic acid (IC50 =7.68 μg/ml). The TAC and FRP activities increased with increasing crude extract/fractions content. The TPC (371.23 ± 15.77 mg GAE/g extract) and TFC (144.64 ± 5.82 mg QE/g extract) of EAF of leaves were found significantly higher as compared to other solvent fractions for both leaves and bark. TPC were highly correlated with the antioxidant activity (R2 = 0.9268 and 0.8515 in DPPH test for leaves and bark, respectively).
The results of the study show that leaves of C. cujete possesses significant free radical scavenging properties compared with stem bark and a clear correlation exists between the antioxidant activity and phenolic content.
PMCID: PMC3937116  PMID: 24495381
Calabash tree; Oxidative stress; Crude extracts; Free radicals; Anti-aging
25.  Antioxidant and Protective Effect of Ethyl Acetate Extract of Podophyllum Hexandrum Rhizome on Carbon Tetrachloride Induced Rat Liver Injury 
The antioxidant and hepatoprotective activities of ethyl acetate extract was carefully investigated by the methods of DPPH radical scavenging activity, Hydroxyl radical scavenging activity, Superoxide radical scavenging activity, Hydrogen peroxide radical scavenging activity and its Reducing power ability. All these in vitro antioxidant activities were concentration dependent which were compared with standard antioxidants such as BHT, α-tocopherol. The hepatoprotective potential of Podophyllum hexandrum extract was also evaluated in male Wistar rats against carbon tetrachloride (CCl4)-induced liver damage. Pre-treated rats were given ethyl acetate extract at 20, 30 and 50 mg/kg dose prior to CCl4 administration (1 ml/kg, 1:1 in olive oil). Rats pre-treated with Podophyllum hexandrum extract remarkably prevented the elevation of serum AST, ALT, LDH and liver lipid peroxides in CCl4-treated rats. Hepatic glutathione levels were significantly increased by the treatment with the extract in all the experimental groups. The extract at the tested doses also restored the levels of liver homogenate enzymes (glutathione peroxidase, glutathione reductase, superoxide dismutase and glutathione-S- transferase) significantly. This study suggests that ethyl acetate extract of P. hexandrum has a liver protective effect against CCl4-induced hepatotoxicity and possess in vitro antioxidant activities.
PMCID: PMC3025527  PMID: 21394192

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