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1.  Evaluation of Essential Oil and its Three Main Active Ingredients of Chinese Chenopodium ambrosioides (Family: Chenopodiaceae) against Blattella germanica 
Background:
The efficacy of essential oil of Chenopodium ambrosioides flowering aerial parts and its three main active ingredients was evaluated against Blattella germanica male adults.
Methods:
Composition of essential oil was determined by GC-MS. Topical application bioassay was used to evaluate contact toxicity of essential oil and three main components. Fumigant toxicity of essential oil and its main components was measured using a sealed space method.
Results:
Twenty-two components were identified in the essential oil and the main components were (Z)-ascaridole (29.7%), isoascaridole (13.0%), ρ-cymene (12.7%) and piperitone (5.0%). The essential oil and (Z)-ascaridole, isoascaridole and ρ-cymene possessed fumigant toxicity against male German cockroaches with LC50 values of 4.13, 0.55, 2.07 and 6.92 mg/L air, respectively. Topical application bioassay showed that all the three compounds were toxic to male German cockroaches and (Z)-ascaridole was the strongest with a LD50 value of 22.02 μg/adult while the crude oil with a LD50 value of 67.46 μg/adult.
Conclusion:
The essential oil from Chinese C. ambrosioides and its three main active ingredients may be explored as natural potential insecticides in the control of cockroaches.
PMCID: PMC3547299  PMID: 23378965
Blattella germanica; Chenopodium ambrosioides; Essential oil; Fumigant; Contact toxicity
2.  Antibacterial activity of leaves extracts of Trifolium alexandrinum Linn. against pathogenic bacteria causing tropical diseases 
Objective
To investigate antibacterial potential of Trifolium alexandrinum (T. alexandrinum) Linn. against seven gram positive and eleven gram negative hospital isolated human pathogenic bacterial strains responsible for many tropical diseases.
Methods
Non-polar and polar extracts of the leaves of T. alexandrinum i.e., hexane, dichloromethane (DCM), ethyl acetate (EtOAc), methanol (MeOH) and aqueous (AQ) extracts at five different concentrations (1, 2, 5, 10 and 15 mg/mL) were prepared to evaluate their antibacterial value. NCCL standards were strictly followed to perform antimicrobial disc susceptibility test using disc diffusion method.
Results
Polar extracts demonstrated significant antibacterial activity against tested pathogens. EtOAc and MeOH extracts showed maximum antibacterial activity with higher inhibition zone and were found effective against seventeen of the tested pathogens. While AQ plant extract inhibited the growth of sixteen of the test strains. EtOAc and MeOH plant extracts inhibited the growth of all seven gram positive and ten of the gram negative bacterial strains.
Conclusions
The present study strongly confirms the effectiveness of crude leaves extracts against tested human pathogenic bacterial strains causing several tropical diseases. Since Egyptian clover is used as a fodder plant, it could be helpful in controlling various infectious diseases associated with cattle as well.
doi:10.1016/S2221-1691(12)60040-9
PMCID: PMC3609279  PMID: 23569896
Trifolium alexandrinum L.; Fabaceae; Antibacterial activity; Pathogenic bacteria; Gram-positive bacteria; Gram-negative bacteria; Tropical disease; Infectious disease
3.  Identification of an anticancer compound against HT-29 cells from Phellinus linteus grown on germinated brown rice 
Objective
To isolate and identify the anticancer compound against proliferation of human colon cancer cells from ethyl acetate (EtOAC) extract of Phellinus linteus grown on germinated brown rice (PB).
Methods
EtOAC extract of PB was partitioned with n-hexane, EtOAC, and water-saturated n-butanol. Anticancer compound of n-hexane layer was isolated and identified by HPLC and NMR, respectively. Cytotoxicity against HT-29 cells was tested by SRB assay.
Results
The n-hexane layer obtained after solvent fractionation of PB EtOAC extracts showed a potent anticancer activity against the HT-29 cell line. Atractylenolide I, a eudesmane-type sesquiterpene lactone, a major anticancer substance of PB, was isolated from the n-hexane layer by silica gel column chromatography and preparative-HPLC. This structure was elucidated by one- and two-dimensional NMR spectroscopic data. Atractylenolide I has not been reported in mushrooms or rice as of yet. The isolated compound dose-dependently inhibited the growth of HT-29 human colon cancer cells.
Conclusions
Atractylenolide I might contribute to the anticancer effect of PB.
doi:10.1016/S2221-1691(13)60156-2
PMCID: PMC3761137  PMID: 24075343
Atractylenolide I; Human colon cancer cells; NMR; Phellinus linteus; Germinated brown rice
4.  The dichloromethane fraction of Stemona tuberosa Lour inhibits tumor cell growth and induces apoptosis of human medullary thyroid carcinoma cells 
Biologics : Targets & Therapy  2007;1(4):455-463.
Medullary thyroid carcinoma (MTC), a neuroendocrine tumor arising from the thyroid gland, is known to be poorly responsive to conventional chemotherapy. The root of Stemona tuberosa Lour, also called Bai Bu, is a commonly used traditional Chinese anti-tussive medicine. The present study investigated this medicinal herb for the first time with respect to its anticancer activity in human medullary thyroid carcinoma cells. Four extracts of Stemona tuberosa Lour, including the n-hexane fraction, (ST-1), dichloromethane (DCM) fraction, (ST-2), ethyl acetate (EtOAc) fraction, (ST-3), and methanol fraction, (ST-4) were examined for antiproliferative effects in two MTC cell lines. We observed that only the DCM fraction ST-2 inhibited cell growth and viability in a dose-dependent manner. Furthermore, we found that ST-2 also induced the apoptosis of MTC-SK cells. Caspase-3/7 was activated, while caspase-9 was not, implying that at least a caspase-dependent apoptotic pathway was involved in this process. In addition, the multicellular spheroids of MTC-SK were destroyed and the cell morphology was changed by ST-2. Our results show the strong apoptotic effects of the DCM fraction of Stemona tuberosa Lour on human medullary thyroid carcinomas, so suggesting a new candidate for chemotherapy of the so far chemo-resistant medullary thyroid carcinoma.
PMCID: PMC2721294  PMID: 19707315
apoptosis; chemoresistance; medullary thyroid carcinoma; plant-derived compounds; Stemona tuberosa Lour
5.  Actinidia macrosperma C. F. Liang (a Wild Kiwi): Preliminary Study of Its Antioxidant and Cytotoxic Activities 
The antioxidant potential of Actinidia macrosperma C. F. Liang (Actinidiaceae) was investigated in vitro for total phenolic content, along with total antioxidant activity (TAA), 1,1-diphenyl 2-picryl hydrazyl (DPPH), and lipid peroxidation (LP). The results indicated that different polarity extracts of A. macrosperma exhibit different biological activities, which depends mainly on the presence of phenolic compounds. The antioxidant activity was in the following decreasing order: MeOH extract > EtOAc extract > aqueous extract > CHCl3 extract > Hexane extract. Moreover, the cytotoxic activity of this plant by MTT dye assay using SMMC-7721 has been determined also. The hexane, EtOAc, and CHCl3 extracts showed cytotoxicity in a dose-dependent manner. Methanol and aqueous extracts, however, showed weak activities in this test. And a very significant cytotoxic activity, not significantly different from the positive control of quercetin, was observed in CHCl3 extract.
doi:10.1155/2012/180262
PMCID: PMC3202103  PMID: 22110544
6.  In vitro anticancer activity of Anemopsis californica 
Oncology letters  2010;1(4):711-715.
Three different extract conditions (aqueous, EtOH and EtOAc) of four different parts (bracts, leaves, roots and stems) of the plant Anemopsis californica (A. californica) were evaluated for their effect on the growth and migration of human colon cancer cells, HCT-8, and the breast cancer cell lines Hs 578T and MCF-7/AZ. Our aim was to identify potential anticancer activity in crude A. californica extracts, given that this plant is used by Native Americans to treat a variety of diseases, including cancer. Our results demonstrated that for each of the cell lines tested, the majority of ethyl acetate extracts of all the plant parts are more toxic than the aqueous and ethanol extracts. Furthermore, significant growth inhibitory activity against the three cell lines was found for the ethyl acetate extract of the roots, while the aqueous extract of the roots influenced the migratory capacity of the three cell lines. This study provides evidence for the anticancer properties of A. californica when extracted in water and ethyl acetate, and supports the importance for further purification of the crude extracts and isolation of potential new anticancer compounds through bio-guided fractionation.
doi:10.3892/ol_00000124
PMCID: PMC3176455  PMID: 21941602
Anemopsis californica; extracts; traditional medicine; cytotoxicity; growth; migration
7.  In vitro anticancer activity of Anemopsis californica 
Oncology Letters  2010;1(4):711-715.
Three different extract conditions (aqueous, EtOH and EtOAc) of four different parts (bracts, leaves, roots and stems) of the plant Anemopsis californica (A. californica) were evaluated for their effect on the growth and migration of human colon cancer cells, HCT-8, and the breast cancer cell lines Hs 578T and MCF-7/AZ. Our aim was to identify potential anticancer activity in crude A. californica extracts, given that this plant is used by Native Americans to treat a variety of diseases, including cancer. Our results demonstrated that for each of the cell lines tested, the majority of ethyl acetate extracts of all the plant parts are more toxic than the aqueous and ethanol extracts. Furthermore, significant growth inhibitory activity against the three cell lines was found for the ethyl acetate extract of the roots, while the aqueous extract of the roots influenced the migratory capacity of the three cell lines. This study provides evidence for the anticancer properties of A. californica when extracted in water and ethyl acetate, and supports the importance for further purification of the crude extracts and isolation of potential new anticancer compounds through bio-guided fractionation.
doi:10.3892/ol_00000124
PMCID: PMC3176455  PMID: 21941602
Anemopsis californica; extracts; traditional medicine; cytotoxicity; growth; migration
8.  Anti-microbial principles of selected remedial plants from Southern India 
Objective
To examine the anti-bacterial activity of leaf extracts of Morus alba L. (Moraceae) and Piper betel L. (Piperaceae), and seed extracts of Bombax ceiba L. (Borabacaceae).
Methods
We have partially purified plant extracts by solvent extraction method, and evaluated the effect of individual fractions on bacterial growth using Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) bacterial strains.
Results
Compared with Morus and Bombax fractions, Piper fractions showed significant growth inhibition on all the three types of bacteria studied. The EtOAc-hexane fractions of Piper leaves exhibited significant anti-bacterial activity with minimum inhibitory concentrations (MIC) of 50 µg/mL culture against both gram-positive and gram-negative bacteria. The EtOAc-fractions I, II, and IV inhibited bacterial colony formation on soft agar in addition to growth inhibition. A combination treatment of piper fractions with ampicillin resulted in significant growth inhibition in E. coli and P. aeruginosa, and combination with anticancer drug geldanamycin (2µg/mL) showed selective growth inhibition against P. aeruginosa and S. aureus. Three major compounds, i.e., eugenol, 3-hexene-ol and stigmasterol, were primarily identified from Piper betel leaf extractions. Among the individual compounds, eugenol treatment showed improved growth inhibition compared with stigmasterol and 3-hexene-ol.
Conclusions
We are reporting potential anti-bacterial compounds from Piper betel against both gram-positive and gram-negative bacteria either alone or in combination with drug treatment.
doi:10.1016/S2221-1691(11)60047-6
PMCID: PMC3614242  PMID: 23569779
Piper betel; Anti-microbial activity; Escherichia coli; Pseudomonas aeruginosa; Staphylococcus aureus; Morus alba; Bombax ceiba; Minimum inhibitory concentration; Growth inhibition
9.  Antioxidant Activities and Phytochemical Study of Leaf Extracts from 18 Indigenous Tree Species in Taiwan 
The objective of this study is to assess antioxidant activities of methanolic extracts from the leaves of 18 indigenous tree species in Taiwan. Results revealed that, among 18 species, Acer oliverianum exhibited the best free radical scavenging activities. The IC50 values were 5.8 and 11.8 μg/mL on DPPH radical and superoxide radical scavenging activities, respectively. In addition, A. oliverianum also exhibited the strongest ferrous ion chelating activity. Based on a bioactivity-guided isolation principle, the resulting methanolic crude extracts of A. oliverianum leaves were fractionated to yield soluble fractions of hexane, EtOAc, BuOH, and water. Of these, the EtOAc fraction had the best antioxidant activity. Furthermore, 8 specific phytochemicals were isolated and identified from the EtOAc fraction. Among them, 1,2,3,4,6-O-penta-galloyl-β-D-glucopyranose had the best free radical scavenging activity. These results demonstrate that methanolic extracts and their derived phytochemicals of A. oliverianum leaves have excellent antioxidant activities and thus they have great potential as sources for natural health products.
doi:10.1155/2012/215959
PMCID: PMC3291425  PMID: 22454657
10.  Cytotoxic and Apoptotic Effects of Different Extracts of Artemisia turanica Krasch. on K562 and HL-60 Cell Lines 
The Scientific World Journal  2013;2013:628073.
Artemisia is an important genus of Iranian flora. Cytotoxic activities for some species of the genus have already been reported. In this study, we have investigated the cytotoxic effects of n-hexane, CH2Cl2, EtOAc, EtOH, and EtOH/H2O (1 : 1) extracts of A. turanica Krasch. on two human leukemic cancer cell lines (K562 and HL-60) and J774 as normal cells using alamarBlue (resazurin) assay. PI staining of the fragmented DNA and western blot analysis were used to evaluate the possible apoptotic effect of the extract. The CH2Cl2 extract of A. turanica showed the most antiproliferative effect on cancer cells among all tested extracts with IC50 values of 69 and 104 μg/mL on K562 and HL-60 cells, respectively, whereas the normal cells were not affected significantly by this extract. Sub-G1 peak in the flow cytometry histogram of the cells treated with CH2Cl2 extract of A. turanica and cleavage of PARP protein confirmed the induction of apoptosis with CH2Cl2 extract. Taken together, the findings of the present work suggest the anticancer potential of CH2Cl2 extract of A. turanica on human leukemic cancer cell lines.
doi:10.1155/2013/628073
PMCID: PMC3830890  PMID: 24288497
11.  Larvicidal Activity against Aedes aegypti and Molluscicidal Activity against Biomphalaria glabrata of Brazilian Marine Algae 
This study investigated the biological activities of five benthic marine algae collected from Northeastern Region of Brazil. The tested activities included larvicidal activity against Aedes aegypti, molluscicidal activity against Biomphalaria glabrata, and toxicity against Artemia salina. Extracts of Ulva lactuca (Chlorophyta), Padina gymnospora, Sargassum vulgare (Phaeophyta), Hypnea musciformis, and Digenea simplex (Rhodophyta) were prepared using different solvents of increasing polarity, including dichloromethane, methanol, ethanol, and water. Of the extracts screened, the dichloromethane extracts of H. musciformis and P. gymnospora exhibited the highest activities and were subjected to bioassay-guided fractionation in hexane and chloroform. The chloroform fractions of the P. gymnospora and H. musciformis extracts showed molluscicidal activity at values below 40 μg·mL−1 (11.1460 μg·mL−1 and 25.8689 μg·mL−1, resp.), and the chloroform and hexane fractions of P. gymnospora showed larvicidal activity at values below 40 μg·mL−1 (29.018 μg·mL−1 and 17.230 μg·mL−1, resp.). The crude extracts were not toxic to A. salina, whereas the chloroform and hexane fractions of P. gymnospora (788.277 μg·mL−1 and 706.990 μg·mL−1) showed moderate toxicity, indicating that the toxic compounds present in these algae are nonpolar.
doi:10.1155/2014/501328
PMCID: PMC3943412
12.  Primary Screening of the Bioactivity of Brackishwater Cyanobacteria: Toxicity of Crude Extracts to Artemia salina Larvae and Paracentrotus lividus Embryos 
Marine Drugs  2010;8(3):471-482.
Cyanobacteria are a diverse group of Gram-negative bacteria that produce an array of secondary compounds with selective bioactivity against vertebrates, invertebrates, plants, microalgae, fungi, bacteria, viruses and cell lines. The aim of this study was to assess the toxic effects of aqueous, methanolic and hexane crude extracts of benthic and picoplanktonic cyanobacteria isolated from estuarine environments, towards the nauplii of the brine shrimp Artemia salina and embryos of the sea urchin Paracentrotus lividus. The A. salina lethality test was used as a frontline screen and then complemented by the more specific sea urchin embryo-larval assay. Eighteen cyanobacterial isolates, belonging to the genera Cyanobium, Leptolyngbya, Microcoleus, Phormidium, Nodularia, Nostoc and Synechocystis, were tested. Aqueous extracts of cyanobacteria strains showed potent toxicity against A. salina, whereas in P. lividus, methanolic and aqueous extracts showed embryo toxicity, with clear effects on development during early stages. The results suggest that the brackishwater cyanobacteria are producers of bioactive compounds with toxicological effects that may interfere with the dynamics of invertebrate populations.
doi:10.3390/md803471
PMCID: PMC2857359  PMID: 20411110
brackishwater cyanobacteria; sea urchin Paracentrotus lividus embryo larval bioassay; brine shrimp Artemia salina lethality test; benthic habitats
13.  Toxicity of Barringtonia racemosa (L.) Kernel Extract on Pomacea canaliculata (Ampullariidae) 
A number of tropical plant species have been recognised as molluscicidal plants, and Barringtonia racemosa (L.) is one of these. The toxicity effects of B. racemosa seed kernel extracts on Pomacea canaliculata were evaluated. The lethal concentration at 50% [LC50 (lower-upper limits)] values, in ppm/48 hours, were 70.71 (41.33–120.97), 94.39 (62.48–142.59), 186.84 (129.21–270.17), and 672.72 (366.57–1234.53) for the extracts withdrawn using dichloromethane (DCM), methanol (MeOH), ethyl acetate (EtOAc), heptane (hp) solvents, respectively at 95% confidence interval (C. I.). All analyses were conducted using Trimmed Spearman-Karber (TSK) program version 1.5. It is assumed that the observed biological effects of the extracts may be due to the saponins and flavonoids present in the seed. The dichloromethanic and methanolic extracts contain saponin and flavonoid substances. Therefore these extracts have shown more potent molluscicidal activity towards the tested organism compared to the remaining extracts. This observed biological activity suggests a promising role for B. racemosa in the control of P. canaliculata.
PMCID: PMC3819080  PMID: 24575198
Barringtonia racemosa; Pomacea canaliculata; Saponins; Flavonoids; Molluscicide; LC50
14.  Antimicrobial and antioxidant activity of kaempferol rhamnoside derivatives from Bryophyllum pinnatum 
BMC Research Notes  2012;5:158.
Background
Bryophyllum pinnatum (Lank.) Oken (Crassulaceae) is a perennial succulent herb widely used in traditional medicine to treat many ailments. Its wide range of uses in folk medicine justifies its being called "life plant" or "resurrection plant", prompting researchers' interest. We describe here the isolation and structure elucidation of antimicrobial and/or antioxidant components from the EtOAc extract of B. pinnatum.
Results
The methanol extract displayed both antimicrobial activities with minimum inhibitory concentration (MIC) values ranging from 32 to 512 μg/ml and antioxidant property with an IC50 value of 52.48 μg/ml. Its partition enhanced the antimicrobial activity in EtOAc extract (MIC = 16-128 μg/ml) and reduced it in hexane extract (MIC = 256-1024 μg/ml). In addition, this process reduced the antioxidant activity in EtOAc and hexane extracts with IC50 values of 78.11 and 90.04 μg/ml respectively. Fractionation of EtOAc extract gave seven kaempferol rhamnosides, including; kaempferitrin (1), kaempferol 3-O-α-L-(2-acetyl)rhamnopyranoside-7-O-α-L-rhamnopyranoside (2), kaempferol 3-O-α-L-(3-acetyl)rhamnopyranoside-7-O-α-L-rhamnopyranoside (3), kaempferol 3-O-α-L-(4-acetyl)rhamnopyranoside-7-O-α-L-rhamnopyranoside (4), kaempferol 3-O-α-D- glucopyranoside-7-O-α-L-rhamnopyranoside (5), afzelin (6) and α-rhamnoisorobin (7). All these compounds, except 6 were isolated from this plant for the first time. Compound 7 was the most active, with MIC values ranging from 1 to 2 μg/ml and its antioxidant activity (IC50 = 0.71 μg/ml) was higher than that of the reference drug (IC50 = 0.96 μg/ml).
Conclusion
These findings demonstrate that Bryophyllum pinnatum and some of its isolated compounds have interesting antimicrobial and antioxidant properties, and therefore confirming the traditional use of B. pinnatum in the treatment of infectious and free radical damages.
doi:10.1186/1756-0500-5-158
PMCID: PMC3353177  PMID: 22433844
Bryophyllum pinnatum; Crassulaceae; Kaempferol rhamnosides; Antimicrobial; Antioxidant; Minimum inhibitory concentration
15.  Antifungal activity of five species of Polygala 
Brazilian Journal of Microbiology  2011;42(3):1065-1075.
Crude extracts and fractions of five species of Polygala – P. campestris, P. cyparissias, P. paniculata, P. pulchella and P. sabulosa – were investigated for their in vitro antifungal activity against opportunistic Candida species, Cryptococcus gattii and Sporothrix schenckii with bioautographic and microdilution assays. In the bioautographic assays, the major extracts were active against the fungi tested. In the minimal concentration inhibitory (MIC) assay, the hexane extract of P. paniculata and EtOAc fraction of P. sabulosa showed the best antifungal activity, with MIC values of 60 and 30 μg/mL, respectively, against C. tropicalis, C. gattii and S. schenckii. The compounds isolated from P. sabulosa prenyloxycoumarin and 1,2,3,4,5,6-hexanehexol displayed antifungal activity against S. schenckii (with MICs of 125 μg/mL and 250 μg/mL, respectively) and C. gattii (both with MICs of 250 μg/mL). Rutin and aurapten isolated from P. paniculata showed antifungal activity against C. gattii with MIC values of 60 and 250 μg/mL, respectively. In the antifungal screening, few of the isolated compounds showed good antifungal inhibition. The compound α-spinasterol showed broad activity against the species tested, while rutin had the best activity with the lowest MIC values for the microorganisms tested. These two compounds may be chemically modified by the introduction of a substitute group that would alter several physico-chemical properties of the molecule, such as hydrophobicity, electronic density and steric strain.
doi:10.1590/S1517-838220110003000027
PMCID: PMC3768791  PMID: 24031724
Polygalaceae; Polygala species; antifungal activity; rutin; α-spinasterol
16.  Antimicrobial activity and cytotoxicity of triterpenes isolated from leaves of Maytenus undata (Celastraceae) 
Background
Plants of the genus Maytenus belong to the family Celastraceae and are widely used in folk medicine as anti-tumour, anti-asthmatic, analgesic, anti-inflammatory, antimicrobial and anti-ulcer agents, and as a treatment for stomach problems. The aim of this study was to isolate and identify active compounds with antifungal activity from Maytenus undata after a preliminary study highlighted promising activity in crude extracts.
Methods
Sequential extracts of M. undata leaves prepared using hexane, dichloromethane (DCM), acetone and methanol (MeOH) were tested for activity against Cryptococcus neoformans, a fungal organism implicated in opportunistic infections. Bioassay-guided fractionation of the hexane extract using C. neoformans as test organism was carried out to isolate antifungal compounds. The cytotoxicity of compounds isolated in sufficient quantities was evaluated using a tetrazolium-based colorimetric cellular assay (MTT) and a haemagglutination assay (HA).
Results
The hexane extract was most active with an MIC of 20 μg/ml against C. neoformans. The triterpene compounds friedelin (1), epifriedelanol (2), taraxerol (3), 3-oxo-11α-methoxyolean-12-ene-30-oic acid (4), 3-oxo-11α-hydroxyolean-12-ene-30-oic acid (5) and 3,11-dihydroxyolean-12-ene-30-oic acid (6) were isolated. Compound 6 was isolated for the first time from a plant species. The antimicrobial activity of compounds 1, 3, 5 and 6 was determined against a range of bacteria and fungi implicated in opportunistic and nosocomial infections. Compounds 5 and 6 were the most active against all the tested microorganisms with MIC values ranging between 24 and 63 μg/ml, except against Staphylococcus aureus which was relatively resistant. Compounds 1 and 3 had a low toxicity with an LC50 > 200 μg/ml towards Vero cells in the MTT assay. Compounds 5 and 6 were toxic with LC50 values of 6.03±0.02 and 2.98±0.01 μg/ml, respectively. Compounds 1 and 3 similarly were not toxic to the red blood cells (RBCs) but compounds 5 and 6 were toxic, showing HA titer values of 1.33 and 0.67 respectively.
Conclusions
Compounds 5 and 6 were the most active but were also relatively cytotoxic to monkey kidney cells and red blood cells, while the other isolated compounds were less active and less cytotoxic.
doi:10.1186/1472-6882-13-111
PMCID: PMC3711988  PMID: 23688235
Maytenus undata; Celastraceae; Antibacterial; Antifungal; Cytotoxicity; Haemagglutination assay
17.  Cytotoxicity of fucosterol containing fraction of marine algae against breast and colon carcinoma cell line 
Pharmacognosy Magazine  2012;8(29):60-64.
Context:
Marine algae produce different secondary metabolites with a wide range of biological activities. Many studies have been achieved on the screening of biological effects of marine organisms and a lot of active compounds were isolated and characterized.
Aims:
In an attempt to find cytotoxic compound of hexane fraction, isolation, identification, and cytotoxicity of active compound of this fraction were performed.
Materials and Methods:
In this study, total methanolic (70%) extract and partition fractions of hexane, chloroform (CHCl3), ethyl acetate (EtOAc), and MeOH–H2O of Sargassum angustifolium, Chondria dasyphylla, and Ulva flexuosa, collected from coastlines of the Persian Gulf in south of Iran, were studied against colon carcinoma (HT-29), colorectal adenocarcinoma (Caco-2), breast ductal carcinoma (T47D), and Swiss mouse embryo fibroblast (NIH 3T3) cell lines by MTT assay.
Statistical Analysis Used:
IC50 (median growth inhibitory concentration) values were calculated by Sigmaplot (10) software.
Results:
Hexane fraction of Chondria dasyphylla (IC50 82.26 ± 4.09 μg/ml) and MeOH-H2O fraction of Ulva flexuosa (IC50 116.92 ± 8.58 μg/ml) showed cytotoxic activity against proliferation of T47D cells. Hexane fraction of Sargassum angustifolium was also observed for cytotoxicity against T47D and HT-29 cell lines (IC50 166.42 ± 26.7 and 190.24 ± 52.8 μg/ml), respectively. An investigation of a component from the hexane fraction of Sargassum angustifolium yielded a steroidal metabolite, fucosterol, with cytotoxicity in T47D and HT29 (IC50 27.94 ± 9.3 and 70.41 ± 7.5 μg/ml).
Conclusions:
These results indicated that fucosterol, the most abundant phytosterol in brown algae, is responsible for cytotoxic effect of this extract against breast and colon carcinoma cell lines.
doi:10.4103/0973-1296.93327
PMCID: PMC3307205  PMID: 22438665
Cytotoxic activity; fucosterol; marine algae; Persian Gulf
18.  Rapid Identification of Aldose Reductase Inhibitory Compounds from Perilla frutescens 
BioMed Research International  2013;2013:679463.
The ethyl acetate (EtOAc) soluble fraction of methanol extracts of Perilla frutescens (P. frutescens) inhibits aldose reductase (AR), the key enzyme in the polyol pathway. Our investigation of inhibitory compounds from the EtOAc soluble fraction of P. frutescens was followed by identification of the inhibitory compounds by a combination of HPLC microfractionation and a 96-well enzyme assay. This allowed the biological activities to be efficiently matched with selected HPLC peaks. Structural analyses of the active compounds were performed by LC-MSn. The main AR inhibiting compounds were tentatively identified as chlorogenic acid and rosmarinic acid by LC-MSn. A two-step high speed counter current chromatography (HSCCC) isolation method was developed with a solvent system of n-hexane-ethyl acetate-methanol-water at 1.5 : 5 : 1 : 5, v/v and 3 : 7 : 5 : 5, v/v. The chemical structures of the isolated compounds were determined by 1H- and 13C-nuclear magnetic resonance spectrometry (NMR). The main compounds inhibiting AR in the EtOAc fraction of methanol extracts of P. frutescens were identified as chlorogenic acid (2) (IC50 = 3.16 μM), rosmarinic acid (4) (IC50 = 2.77 μM), luteolin (5) (IC50 = 6.34 μM), and methyl rosmarinic acid (6) (IC50 = 4.03 μM).
doi:10.1155/2013/679463
PMCID: PMC3838804  PMID: 24308003
19.  Phenolic compounds from Foeniculum vulgare (Subsp. Piperitum) (Apiaceae) herb and evaluation of hepatoprotective antioxidant activity 
Pharmacognosy Research  2012;4(2):104-108.
Objective:
The study was designed to evaluate the antioxidant and hepatoprotective activities of the 80% methanolic extract as well as the ethyl acetate (EtOAc) and butanol (BuOH) fractions of the wild fennel (Foeniculum vulgare (Subsp; Piperitum)) and cultivated fennel (F. vulgare var. azoricum). In addition, quantification of the total phenolic content in the 80% methanol extract of fennel wild and cultivated herbs is measured.
Materials and Methods:
An amount of 400 g of air dried powdered herb of wild and cultivated fennel were sonicated with aqueous methanol (80%), successively extracted with Hexane, EtOAc, and n-BuOH. The EtOAc and n-BuOH were subjected to repeated column chromatography on silica gel and Sephadex LH-20. The antioxidant effect was determined in vitro using 1,1-diphenyl-2-picrylhydrazyl (DPPH). Hepatoprotective activity was carried out using a Wistar male rat (250–300 g). Total phenolic and flavonoid contents were determined as chlorogenic acid and rutin equivalents, respectively.
Results:
Two phenolic compounds, i.e., 3,4-dihydroxy-phenethylalchohol-6-O-caffeoyl-β-D-glucopyranoside and 3΄,8΄-binaringenin were isolated from the fennel wild herb, their structures were elucidated by spectral methods including 1D NMR, 2D NMR, and UV. The EtOAc and BuOH fractions of wild fennel were found to exhibit a radical scavenging activity higher than those of cultivated fennel. An in vitro method of rat hepatocytes monolayer culture was used for the investigation of hepatotoxic effects of the 80% methanol extract on the wild and cultivated fennel, which were >1000 and 1000 μg/mL, respectively. As well as, their hepatoprotective effect against the toxic effect of paracetamol (25 mM) was exerted at 12.5 μg/mL concentration.
Conclusions:
Fennel (F. Vulgare) is a widespread plant species commonly used as a spice and flavoring. The results obtained in this study indicated that the fennel (F. vulgare) herb is a potential source of natural antioxidant. Two phenolic compounds, i.e. 3,4-dihydroxy-phenethylalchohol-6-O-caffeoyl-β-D-glucopyranoside (A) and 3΄,8΄-binaringenin (B) were isolated from the fennel wild herb for the first time.
doi:10.4103/0974-8490.94735
PMCID: PMC3326756  PMID: 22518082
Antioxidant (Apiaceae); azoricum; binaringenin; Foeniculum vulgare; hepatoprotection; phenolic compounds; piperitum
20.  The Role of the Ethylacetate Fraction from Hydnocarpi Semen in Acute Inflammation In Vitro Model 
Immune Network  2012;12(6):291-295.
We previously reported that Hydnocarpi Semen (HS) has a wound healing effect on diabetic foot ulcer lesion in mice. In this study, ethylacetate (EtOAc) fraction from HS extract were evaluated for their wound healing activity by using in vitro acute inflammation model. GC and GC/MS analysis shows that the main constituents in EtOAc fraction are chaulmoogric acid, hydnocarpic acid, and gorlic acid. EtOAc fraction activated macrophages to increase the production of TNF-α. The fraction also increased the production of TGF-β and VEGF, which induced fibroblast activation and angiogenesis. These results suggest that the mechanism that the fraction helps to enhance healing of skin wound is possibly associated with the production of TNF-α, as well as secretion of VEGF, TGF-β and HS may have a new bioactive material for the treatment of skin wound.
doi:10.4110/in.2012.12.6.291
PMCID: PMC3566425  PMID: 23396990
Hydnocarpi Semen; Ethylacetate fraction; Inflammation; Cytokines
21.  Antrodia camphorata Grown on Germinated Brown Rice Suppresses Melanoma Cell Proliferation by Inducing Apoptosis and Cell Differentiation and Tumor Growth 
Antrodia camphorata grown on germinated brown rice (CBR) was prepared to suppress melanoma development. CBR extracts were divided into hexane, EtOAc, BuOH, and water fractions. Among all the fractions, EtOAc fraction showed the best suppressive effect on B16F10 melanoma cell proliferation by CCK-8 assay. It also showed the increased cell death and the changed cellular morphology after CBR treatment. Annexin V-FITC/PI, flow cytometry, and western blotting were performed to elucidate anticancer activity of CBR. The results showed that CBR induced p53-mediated apoptotic cell death of B16F10. CBR EtOAc treatment increased melanin content and melanogenesis-related proteins of MITF and TRP-1 expressions, which supports its anticancer activity. Its potential as an anticancer agent was further investigated in tumor-xenografted mouse model. In melanoma-xenografted mouse model, melanoma tumor growth was significantly suppressed under CBR EtOAc fraction treatment. HPLC analysis of CBR extract showed peak of adenosine. In conclusion, CBR extracts notably inhibited B16F10 melanoma cell proliferation through the p53-mediated apoptosis induction and increased melanogenesis. These findings suggest that CBR EtOAc fraction can act as an effective anticancer agent to treat melanoma.
doi:10.1155/2013/321096
PMCID: PMC3596902  PMID: 23533475
22.  Antimicrobial and toxicological activities of five medicinal plant species from Cameroon Traditional Medicine 
Background
Infectious diseases caused by multiresistant microbial strains are on the increase. Fighting these diseases with natural products may be more efficacious. The aim of this study was to investigate the in vitro antimicrobial activity of methanolic, ethylacetate (EtOAc) and hexanic fractions of five Cameroonian medicinal plants (Piptadeniastum africana, Cissus aralioides, Hileria latifolia, Phyllanthus muellerianus and Gladiolus gregasius) against 10 pathogenic microorganisms of the urogenital and gastrointestinal tracts.
Methods
The fractions were screened for their chemical composition and in vivo acute toxicity was carried out on the most active extracts in order to assess their inhibitory selectivity.
The agar well-diffusion and the micro dilution methods were used for the determination of the inhibition diameters (ID) and Minimum inhibitory concentrations (MIC) respectively on 8 bacterial species including two Gram positive species (Staphylococcus aureus, Enterococcus faecalis), and six Gram negative (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Shigella flexneri, Salmonella typhi) and two fungal isolates (Candida albicans, Candida krusei). The chemical composition was done according to Harbone (1976), the acute toxicity evaluation according to WHO protocol and the hepatic as well as serum parameters measured to assess liver and kidney functions.
Results
The chemical components of each plant's extract varied according to the solvent used, and they were found to contain alkaloids, flavonoids, polyphenols, triterpens, sterols, tannins, coumarins, glycosides, cardiac glycosides and reducing sugars. The methanolic and ethylacetate extracts of Phyllanthus muellerianus and Piptadeniastum africana presented the highest antimicrobial activities against all tested microorganisms with ID varying from 8 to 26 mm and MIC from 2.5 to 0.31 mg/ml. The in vivo acute toxicity study carried out on the methanolic extracts of Phyllanthus muellerianus and Piptadeniastrum africana indicated that these two plants were not toxic. At the dose of 4 g/kg body weight, kidney and liver function tests indicated that these two medicinal plants induced no adverse effect on these organs.
Conclusion
These results showed that, all these plant's extracts can be used as antimicrobial phytomedicines which can be therapeutically used against infections caused by multiresistant agents.
Phyllanthus muellerianus, Piptadeniastum africana, antimicrobial, acute toxicity, kidney and liver function tests, Cameroon Traditional Medicine
doi:10.1186/1472-6882-11-70
PMCID: PMC3182953  PMID: 21867554
Phyllanthus muellerianus; Piptadeniastum africana; antimicrobial; acute toxicity; kidney and liver function tests; Cameroon Traditional Medicine
23.  Crude ethyl acetate extract of marine microalga, Chaetoceros calcitrans, induces Apoptosis in MDA-MB-231 breast cancer cells 
Pharmacognosy Magazine  2014;10(37):1-8.
Background:
Marine brown diatom Chaetoceros calcitrans and green microalga Nannochloropsis oculata are beneficial materials for various applications in the food, nutraceutical, pharmaceutical and cosmeceutical industries.
Objective:
This study investigated cytotoxicity of different crude solvent extracts from C. calcitrans and N. oculata against various cancer cell lines.
Materials and Methods:
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was carried out to screen the cytotoxic effects of hexane (Hex), dichloromethane (DCM), ethyl acetate, and methanol extract from C. calcitrans and N. oculata toward various cancer cell lines. Flow cytometry cell cycle was used to determine the cell cycle arrest while the mode of cell death was investigated through acridine orange/propidium iodide (AOPI) staining, Annexin V-Fluorescein Isothiocyanate (FITC) and Terminal deoxynucleotidyl transferase-mediated d-UTP Nick End Labeling (TUNEL) assays. Expression profile of apoptotic and proliferative-related genes was then determined using the multiplex gene expression profiler (GeXP).
Results:
Crude ethyl acetate (CEA) extract of C. calcitrans inhibited growth of MDA-MB-231 cells, with IC50 of 60 μg/mL after 72 h of treatment. Further studies were conducted to determine the mode of cell death at various concentrations of this extract: 30, 60 and 120 μg/mL. The mode of cell death was mainly apoptosis as shown through apoptosis determination test. The expression data from GeXP showed that caspase-4 was upregulated while B-cell leukemia/lymphoma 2(Bcl-2) was down regulated. Thus, caspase-4 induction endoplasmic reticulum death pathway is believed to be one of the mechanisms underlying the induction of apoptosis while Bcl-2 induced S and G2/M cell cycle phase arrest in MDA-MB-231 cells.
Conclusion:
CEA extract of C. calcitrans showed the highest cytotoxicity on MDA-MB-231 via apoptosis.
doi:10.4103/0973-1296.126650
PMCID: PMC3969653
Apoptosis; Chaetoceros calcitrans; crude ethyl acetate extract; gene expression profiler; MDA-MB-231
24.  Acanthopanax koreanum Fruit Waste Inhibits Lipopolysaccharide-Induced Production of Nitric Oxide and Prostaglandin E2 in RAW 264.7 Macrophages 
The Acanthopanax koreanum fruit is a popular fruit in Jeju Island, but the byproducts of the alcoholic beverage prepared using this fruit are major agricultural wastes. The fermentability of this waste causes many economic and environmental problems. Therefore, we investigated the suitability of using A. koreanum fruit waste (AFW) as a source of antiinflammatory agents. AFWs were extracted with 80% EtOH. The ethanolic extract was then successively partitioned with hexane, CH2Cl2, EtOAc, BuOH, and water. The results indicate that the CH2Cl2 fraction (100 μg/mL) of AFW inhibited the LPS-induced nitric oxide (NO) and prostaglandin E2 (PGE2) production in RAW 264.7 cells by 79.6% and 39.7%, respectively. These inhibitory effects of the CH2Cl2 fraction of AFWs were accompanied by decreases in the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins and iNOS and COX-2 mRNA in a dose-dependent pattern. The CH2Cl2 fraction of AFWs also prevented degradation of IκB-α in a dose-dependent manner. Ursolic acid was identified as major compound present in AFW, and CH2Cl2 extracts by high performance liquid chromatography (HPLC). Furthermore using pure ursolic acid as standard and by HPLC, AFW and CH2Cl2 extracts was found to contain 1.58 mg/g and 1.75 mg/g, respectively. Moreover, we tested the potential application of AFW extracts as a cosmetic material by performing human skin primary irritation tests. In these tests, AFW extracts did not induce any adverse reactions. Based on these results, we suggest that AFW extracts be considered possible anti-inflammatory candidates for topical application.
doi:10.1155/2010/715739
PMCID: PMC2846352  PMID: 20368786
25.  In vivo and in vitro antiplasmodial activities of some plants traditionally used in Guatemala against malaria. 
We present an evaluation of the antiplasmodial and cytotoxic effects of four plants commonly used in Guatemalan folk medicine against malaria. Methanol extracts of Simarouba glauca D. C., Sansevieria guineensis Willd, Croton guatemalensis Lotsy, and Neurolaena lobata (L.)R.Br. significantly reduced parasitemias in Plasmodium berghei-infected mice. Dichloromethane fractions were screened for their cytotoxicities on Artemia salina (brine shrimp) larvae, and 50% inhibitory concentrations were determined for Plasmodium falciparum in in vitro cultures. Both chloroquine-susceptible and -resistant strains of P. falciparum were significantly inhibited by these extracts. Of all dichloromethane extracts, only the S. glauca cortex extract was considered to be toxic to nauplii of A. salina in the brine shrimp test.
PMCID: PMC163947  PMID: 9210673

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