Search results for: in vitro anticancer activity

Authors: Jin Boo Jeong

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In this study, we elucidated anti-cancer activity and potential molecular mechanism of DKC against human colorectal cancer cells. DKC-E70 suppressed the proliferation of human colorectal cancer cell lines such as HCT116, SW480, LoVo and HT-29. Although DKC-E70 decreased cyclin D1 expression in protein and mRNA level, decreased level of cyclin D1 protein by DKC-E70 occurred at the earlier time than that of cyclin D1 mRNA, which indicates that DKC-E70-mediated downregulation of cyclin D1 protein may be a consequence of the induction of degradation and transcriptional inhibition of cyclin D1. In cyclin D1 degradation, we found that cyclin D1 downregulation by DKC-E70 was attenuated in presence of MG132. In addition, DKC-E70 phosphorylated threonine-286 (T286) of cyclin D1 and T286A abolished cyclin D1 downregulation by DKC-E70. We also observed that DKC-E70-mediated T286 phosphorylation and subsequent cyclin D1 degradation was blocked in presence of the inhibitors of ERK1/2, p38 or GSK3β. In cyclin D1 transcriptional inhibition, DKC-E70 inhibited the expression of β-catenin and TCF4, and β–catenin/TCF-dependent luciferase activity. Our results suggest that DKC-E70 may downregulate cyclin D1 as one of the potential anti-cancer targets through cyclin D1 degradation by T286 phosphorylation dependent on ERK1/2, p38 or GSK3β, and cyclin D1 transcriptional inhibition through Wnt signaling. From these findings, DKC-E70 has potential to be a candidate for the development of chemoprevention or therapeutic agents for human colorectal cancer. This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2016R1D1A3B03931713).

Keywords: anticancer, calyx of persimmon, cyclin D1, Diospyros kaki Thunb., human colorectal cancer

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Authors: Sylwia K. Naliwajko, Renata Markiewicz-Zukowska, Justyna Moskwa, Krystyna Gromkowska-Kepka, Konrad Mielcarek, Patryk Nowakowski, Katarzyna Socha, Anna Puscion-Jakubik, Maria H. Borawska

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Glioblastoma (grade IV WHO) is a rapidly progressive brain tumor with very high morbidity and mortality. The vast malignant gliomas are not curable despite the therapy (surgical, radiotherapy, chemotherapy) and patients seek alternative or complementary treatments. Patients often use cannabidiol (CBD) oil as an alternative therapy of glioblastoma. CBD is one of the cannabinoids, an active component of Cannabis sativa. THC (Δ9-tetrahydrocannabinol) can be addictive, and in many countries CBD oil without THC ( < 0,2%) is available. Propolis produced by bees from the resin collected from trees has antiglioma properties in vitro and can be used as a supplement in complementary therapy of gliomas. The aim of this study was to examine the influence of extract from CBD oil in combination with propolis extract on two glioblastoma cell lines. The MTT (Thiazolyl Blue Tetrazolium Bromide) test was used to determine the influence of CBD oil extract and polish propolis extract (PPE) on the viability of glioblastoma cell lines – U87MG and LN18. The cells were incubated (24, 48 and 72 h) with CBD oil extract and PPE. CBD extract was used in concentration 1, 1.5 and 3 µM and PPE in 30 µg/mL. The data were presented compared to the control. The statistical analysis was performed using Statistica v. 13.0 software. CBD oil extract in concentrations 1, 1.5 and 3 µM did not inhibit the viability of U87MG and LN18 cells (viability more than 90% cells compared to the control). There was no dose-response viability, and IC50 value was not recognized. PPE in the concentration of 30 µg/mL time-dependently inhibited the viability of U87MG and LN18 cell line (after 48 h the viability as a percent of the control was 59,7±6% and 57,8±7%, respectively). In a combination of CBD with PPE, the viability of the treated cells was similar to PPE used alone (58,2±7% and 56,5±9%, respectively). CBD oil extract did not show anti-glioma activity and in combination with PPE did not change the activity of PPE.

Keywords: anticancer, cannabidiol, cell line, glioblastoma

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Authors: Indu Barwal, Shloka Thakur, Subhash C. Yadav

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The plant virus capsid protein based nanoparticles are extensively studied for their application in biomedical research for development of nanomedicines and drug delivery systems. We have developed a chimeric drug delivery system by controlled in vitro assembly of separately bioconjugated fluorescent dye (as reporting molecule), folic acid (as receptor binding biomolecule for targeted delivery) and doxorubicin (as anticancer drug) using modified EDC NHS chemistry on heterologously overexpressed (E. coli) capsid proteins of cowpea chlorotic mottle virus (CCMV). This chimeric vehicle was further encapsidated on gold nanoparticles (20nm) coated with 5≠ thiolated DNA probe to neutralize the positive charge of capsid proteins. This facilitates the in vitro assembly of modified capsid subunits on the gold nanoparticles to develop chimeric GNPs encapsidated targeted drug delivery system. The bioconjugation of functionalities, number of functionality on capsid subunits as well as virus like nanoparticles, structural stability and in vitro assembly were confirmed by SDS PAGE, relative absorbance, MALDI TOF, ESI-MS, Circular dichroism, intrinsic tryptophan fluorescence, zeta particle size analyzer and TEM imaging. This vehicle was stable at pH 4.0 to 8.0 suitable for many organelles targeting. This in vitro assembled chimeric plant virus like particles could be suitable for ideal drug delivery vehicles for subcutaneous cancer treatment and could be further modified for other type of cancer treatment by conjugating other functionalities (targeting, drug) on capsids.

Keywords: chimeric drug delivery vehicles, bioconjugated plant, virus, capsid

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Authors: Steven Arcidiacono, Robert Stote, Erin Anderson, Molly Richards

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There are numerous standard test methods for antimicrobial textiles that measure activity against specific microorganisms. However, many times these results do not translate to the performance of treated textiles when worn by individuals. Standard test methods apply a single target organism grown under optimal conditions to a textile, then recover the organism to quantitate and determine activity; this does not reflect the actual performance environment that consists of polymicrobial communities in less than optimal conditions or interaction of the textile with the skin substrate. Here we propose the development of in vitro skin model method to bridge the gap between lab testing and wear studies. The model will consist of a defined polymicrobial community of 5-7 commensal microbes simulating the skin microbiome, seeded onto a solid tissue platform to represent the skin. The protocol would entail adding a non-commensal test organism of interest to the defined community and applying a textile sample to the solid substrate. Following incubation, the textile would be removed and the organisms recovered, which would then be quantitated to determine antimicrobial activity. Important parameters to consider include identification and assembly of the defined polymicrobial community, growth conditions to allow the establishment of a stable community, and choice of skin surrogate. This model could answer the following questions: 1) is the treated textile effective against the target organism? 2) How is the defined community affected? And 3) does the textile cause unwanted effects toward the skin simulant? The proposed model would determine activity under conditions comparable to the intended application and provide expanded knowledge relative to current test methods.

Keywords: antimicrobial textiles, defined polymicrobial community, in vitro skin model, skin microbiome

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Authors: Marek Halenár, Eva Tvrdá, Simona Baldovská, Ľubomír Ondruška, Peter Massányi, Adriana Kolesárová

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This study aimed to assess the in vitro effects of different concentrations of the Viscum album extract on the motility, viability, and reactive oxygen species (ROS) production by rabbit spermatozoa during different time periods (0, 2, and 8h). Spermatozoa motility was assessed by using the CASA (Computer aided sperm analysis) system. Cell viability was evaluated by using the metabolic activity MTT assay, and the luminol-based luminometry was applied to quantify the ROS formation. The CASA analysis revealed that low Viscum concentrations were able to prevent a rapid decline of spermatozoa motility, especially in the case of concentrations ranging between 1 and 5 µg/mL (P<0.05 with respect to time 8h). At the same time, concentrations ranging between 1 and 100 µg/mL of the extract led to a significant preservation of the cell viability (P<0.05 in case of 5, 50 and 100 µg/mL; P<0.01 with respect to 1 and 10 µg/mL, time 8h). 1 and 5 µg/mL of the extract exhibited antioxidant characteristics, translated into a significant reduction of the ROS production, particularly notable at time 8h (P<0.01). The results indicate that the Viscum extract is capable of delaying the damage inflicted to the spermatozoon by the in vitro environment.

Keywords: CASA, mistletoe, mitochondrial activity, motility, reactive oxygen species, rabbits, spermatozoa, Viscum album

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Authors: Maria Borawska, Patryk Nowakowski, Sylwia K. Naliwajko, Renata Markiewicz-Zukowska, Anna Puscion-Jakubik, Krystyna Gromkowska-Kepka, Justyna Moskwa

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Coprinus comatus (O. F. Müll.) Pers.) should not be confused with the common Ink Cap, which contains coprine and can induce coprine poisoning. We study the possibility of applying coprinus mushroom (Coprinus comatus), available in Poland, as food product supporting the treatment of human glioblastoma cells. The U87MG and T98 glioblastoma cell lines were exposed to water (CW) or ethanol 95° (CE) Cantharellus extracts (50-500 μg/ml), with or without temozolomide (TMZ) during 24, 48 or 72 hours. The cell division was examined by the H³-thymidine incorporation. The statistical analysis was performed using Statistica v. 13.0 software. Significant differences were assumed for p < 0.05. We found that both, CW and CE, administrated alone, had inhibitory effect on cell lines growth, but the CE extract had a higher degree of growth inhibition. The anti-tumor effect of TMZ (50 μM) on U87MG was enhanced by mushroom extracts, and the effect was lower to the effect after using Coprinus comatus extracts (CW and CE) alone. A significant decrease (p < 0.05) in pro-MMP2 (82.61 ± 6.3% of control) secretion in U87MG cells was observed after treated with CE (250 μg/ml). We conclude that extracts of Coprinus comatus, edible mushroom, present cytotoxic properties on U87MG and T98 cell lines and may cooperate with TMZ synergistically enhancing its growth inhibiting activity against glioblastoma U87MG cell line.

Keywords: anticancer, glioma, mushroom, temozolomide

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Authors: Kabange Kasumbwe, Viresh Mohanlall, Bharti Odhav, Venu Narayanaswamy

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Coumarins are naturally occurring plant metabolites known for their pharmacological properties such as anticoagulant, antimicrobial, anticancer, antioxidant, anti-inflammatory and antiviral properties. The pharmacological and biochemical properties and curative applications of coumarins depend on the substitution around the coumarin core structure. In the present study, seven halogenated coumarins CMRN1-CMRN7 were synthesized and evaluated for their anticancer activity. The cytotoxicity potential of the test compounds was evaluated against UACC62 (Melanoma), MCF-7 (Breast cancer) and PBM (Peripheral Blood Mononuclear) cell lines using MTT assay keeping doxorubicin as standard drug. The apoptotic potential of the coumarin compounds was evaluated against UACC62 (Melanoma) cell by assessing their morphological changes, membrane change, mitochondria membrane potential; pro-apoptotic changes were investigated using the AnnexinV-PI staining, JC-1, caspase-3 enzyme kits respectively on flow cytometer. The synthetic coumarin has strongly suppressed the cell proliferation of UACC-62 (Melanoma) and MCF-7 (Breast) Cancer cells, the higher toxicity of these compounds against UACC-62 (Melanoma) and MCF-7 (Breast) were CMRN3, CMRN4, CMRN5, CMRN6. However, compounds CMRN1, CMRN2, and CMRN7 had no significant inhibitory effect. Furthermore the active compounds CMRN3, CMRN4, CMRN5, CMRN6 exerted antiproliferative effects through apoptosis induction against UACC-62 (Melanoma), suggesting their potential could be considered as attractive lead molecules in the future for the development of potential anticancer agents since one of the important criteria in the development of therapeutic drugs for cancer treatment is to have high selectivity and less or no side-effects on normal cells and these compounds had no inhibitory effect against the PBMC cells.

Keywords: coumarin, MTT, apoptosis, cytotoxicity

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Authors: Khadidja Bouzid, Fouzia Benali Toumi, Mohamed Bouzouina

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We made a comparison between the total phenolic content, concentrations of flavonoids and antioxidant activity of four different extracts (butanol, ethyl acetate, chloroform, water) of Arbutus unedo L. fruit (Ericacea) of El Marsa and Terni area. The total phenolic content in the extracts was determined using the Folin-Ciocalteu reagent and it ranged between 26.57 and 48.23 gallic acid equivalents mg/g of dry weight of extract. The concentrations of flavonoids in plant extracts varied from 17.98 to 56.84 catechin equivalents mg/g. The antioxidant activity was analyzed in vitro using the DPPH reagent; among all extracts, ethyl acetate fraction from El Marsa area showed the highest antioxidant activity.

Keywords: antioxidant activity, Arbutus unedo L., fruit flavonoids, phenols, Western Algeria

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Authors: Tooba N. Shamsi, Romana Perveen, Sadaf Fatima

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Protease Inhibitors (PIs) are widespread in nature, produced by animals, plants and microorganisms. They play vital role in various biological activities by keeping a check on activity of proteases. Present study aims to investigate antioxidant and anti-inflammatory properties of PPI from Cajanus cajan (CCTI) and Phaseolus limensis (LBTI). PPI was purified from C. cajan (PUSA-992) by ammonium sulfate precipitation followed by ion exchange chromatography. The anti-oxidant activity was analyzed by two most common radical scavenging assays of FRAP (ferric reducing antioxidant power) and DPPH (1,1- diphenyl-2-picrylhydrazyl). Also, in-vitro anti-inflammatory activity was evaluated using albumin denaturation assay and membrane stabilization assay at different concentrations. Ascorbic acid and aspirin were used as a standards for antioxidant and anti-inflammatory assays respectively. The PPIs were also checked for antimicrobial activity against a number of bacterial strains. The CCTI and LBTI showed DPPH radical scavenging activity in a concentration–dependent manner with IC50 values 544 µg/ml and 506 µg/ml respectively comparative to ascorbic acid which was 258 µg/ml. Following FRAP assay, it was evaluated that LBTI had 87.5% and CCTI showed 84.4% antioxidant activity, taking value of standard ascorbic acid to be 100%. The PPIs also showed in-vitro anti‐inflammatory activity by inhibiting the heat induced albumin denaturation with IC50 values of 686 µg/ml and 615 µg/ml for CCTI and LBTI respectively compared to the standard (aspirin) which was 70.8 µg/ml. Red blood cells membrane stabilization with IC50 values of 641 µg/ml and 587 µg/ml for CCTI and LBTI respectively against aspirin which showed IC50 value of 70.4 µg/ml. PPIs showed antibacterial activity against 7 known strains while there was apparently no action against fungi.

Keywords: Cajanus cajan, Phaseolus limensis, Lima beans, protein protease inhibitor, antioxidant, anti-inflammatory, antimicrobial activity

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Authors: Olaniyi Fawole, Umezuruike Opara

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Co-products obtained from pomegranate juice processing contain high levels of polyphenols with potential high added values. From value-addition viewpoint, the aim of this study was to evaluate the stability of polyphenolic concentrations in pomegranate fruit co-products in different solvent extracts and assess the effect on the total antioxidant capacity using the FRAP, DPPH˙ and ABTS˙+ assays during simulated in vitro digestion. Pomegranate juice, marc and peel were extracted in water, 50% ethanol (50%EtOH) and absolute ethanol (100%EtOH) and analysed for total phenolic concentration (TPC), total flavonoids concentration (TFC) and total antioxidant capacity in DPPH˙, ABST˙+ and FRAP assays before and after in vitro digestion. Total phenolic concentration (TPC) and total flavonoid concentration (TFC) were in the order of peel > marc > juice throughout the in vitro digestion irrespective of the extraction solvents used. However, 50% ethanol extracted 1.1 to 12-fold more polyphenols than water and ethanol solvents depending on co-products. TPC and TFC increased significantly in gastric digests. In contrast, after the duodenal, polyphenolic concentrations decreased significantly (p < 0.05) compared to those obtained in gastric digests. Undigested samples and gastric digests showed strong and positive relationships between polyphenols and the antioxidant activities measured in DPPH, ABTS and FRAP assays, with correlation coefficients (r2) ranging between 0.930 – 0.990 whereas, the correlation between polyphenols (TPC and TFC) and radical cation scavenging activity (in ABTS) were moderately positive in duodenal digests. Findings from this study also showed that the concentration of pomegranate polyphenols and antioxidant thereof during in vitro gastro-intestinal digestion may not reflect the pre-digested phenolic concentration. Thus, this study highlights the need to provide biologically relevant information on antioxidants by providing data reflecting their stability and activity after in vitro digestion.

Keywords: by-product, DPPH, polyphenols, value addition

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Authors: Qaiser Akram, Ahsan Naeem, Hafiz Muhammad Rizwan, Waqas Ahmad, Rubeena Yasmeen

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Aspergillus has good secretory potential for phytases. Morphologically and microscopically identified Aspergillus terreus (A. terreus) (n=20) were screened for phytase production and non-toxicity. Phytases produced by non-toxigenic A. terreus under optimum conditions were quantified. Phytases of highest producer A. terreus were evaluated for stability after exposure to temperature (35, 55, 75 and 95ºC) and pH (2, 4, 6 and 8). Effect of metal ions (Fe⁺³, Ba⁺², Ca⁺², Cu⁺², Mg⁺², Mn⁺², K⁺¹ and Na⁺¹) was assessed on phytase activity. Log reduction in phytase activity was calculated. The highest activity units of phytase produced by A. terreus were 271.49 ± 8.14 phytase unit / mL (FTU/ mL). The lowest reduction in phytase activity was 50.20 ± 7.36 (18.5%) and 68.22 ± 10.3 FTU/mL (25.13%) at 35ºC and pH 6, respectively for 15 minutes. The highest reduction 259 ± 0.84 (95.5%) and 211.99 ± 4.39 FTU/mL (78.1%) was recorded at 95ºC for 60 minutes and pH 2.0 for 45 minutes exposure, respectively. All metal ions negatively affected phytase activity. Phytase activity was inhibited minimum (45.32 ± 28.54 FTU/mL, 16.69%) by K⁺¹(1 mM) and maximum (231.48 ± 3.68 FTU/mL, 80.8%) by Cu⁺² (10 mM). It was concluded that A. terreus phytase stability and activity was dependent on physio-chemical factors.

Keywords: stability, phytase, aspergillus terreus, physio-chemical factors and metal ions

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Authors: Alaa M. M. Badr Eldin, Marwa I. Ezzat, Mohammed S. Sedeek, Manal S. Afifi, Omar M. Sabry

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Cytotoxic activity of the total methanol extract against breast cancer cell line MCF-7 was amazing IC50 at 54 ug/ml. 130 polyphenolic compounds were tentatively identified in pomegranate peel (Punica granatum L.) methanol extract using HPLC-MS/MS technique. The antiestrogenic activity of the polyphenolic constituents found in pomegranate extract was confirmed experimentally in-vitro and by the in-silico molecular docking using gallagic acid, ellagic acid, and Punicalagin as these are considered model compounds confirmed in pomegranate peel extract. The methanolic extract was found to suppress ER, TGF-β, and NF-kB in-vitro gene expression strongly, and that was verified by qPCR and Western Blot gel electrophoresis techniques.

Keywords: HPLC-MS/MS, pomegranate, breast cancer, ovarian cancer, ER, TGF-β, NF-kB

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Authors: Narin Ozturk, Xiao-Mei Li, Sylvie Giachetti, Francis Levi, Alper Okyar

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P-glycoprotein (P-gp, MDR1, ABCB1) is a transmembrane protein acting as an ATP-dependent efflux pump and functions as a biological barrier by extruding drugs and xenobiotics out of cells in healthy tissues especially in intestines, liver and brain as well as in tumor cells. The circadian timing system controls a variety of biological functions in mammals including xenobiotic metabolism and detoxification, proliferation and cell cycle events, and may affect pharmacokinetics, toxicity and efficacy of drugs. Selective mTOR (mammalian target of rapamycin) inhibitor everolimus is an immunosuppressant and anticancer drug that is active against many cancers, and its pharmacokinetics depend on P-gp. The aim of this study was to investigate the dosing time-dependent toxicity of everolimus with respect to the intestinal P-gp expression rhythms in mdr1a::Luc mice using Real Time-Biolumicorder (RT-BIO) System. Mdr1a::Luc male mice were synchronized with 12 h of Light and 12 h of Dark (LD12:12, with Zeitgeber Time 0 – ZT0 – corresponding Light onset). After 1-week baseline recordings, everolimus (5 mg/kg/day x 14 days) was administered orally at ZT1-resting period- and ZT13-activity period- to mdr1a::Luc mice singly housed in an innovative monitoring device, Real Time-Biolumicorder units which let us monitor real-time and long-term gene expression in freely moving mice. D-luciferin (1.5 mg/mL) was dissolved in drinking water. Mouse intestinal mdr1a::Luc oscillation profile reflecting P-gp gene expression and locomotor activity pattern were recorded every minute with the photomultiplier tube and infrared sensor respectively. General behavior and clinical signs were monitored, and body weight was measured every day as an index of toxicity. Drug-induced body weight change was expressed relative to body weight on the initial treatment day. Statistical significance of differences between groups was validated with ANOVA. Circadian rhythms were validated with Cosinor Analysis. Everolimus toxicity changed as a function of drug timing, which was least following dosing at ZT13, near the onset of the activity span in male mice. Mean body weight loss was nearly twice as large in mice treated with 5 mg/kg everolimus at ZT1 as compared to ZT13 (8.9% vs. 5.4%; ANOVA, p < 0.001). Based on the body weight loss and clinical signs upon everolimus treatment, tolerability for the drug was best following dosing at ZT13. Both rest-activity and mdr1a::Luc expression displayed stable 24-h periodic rhythms before everolimus and in both vehicle-treated controls. Real-time bioluminescence pattern of mdr1a revealed a circadian rhythm with a 24-h period with an acrophase at ZT16 (Cosinor, p < 0.001). Mdr1a expression remained rhythmic in everolimus-treated mice, whereas down-regulation was observed in P-gp expression in 2 of 4 mice. The study identified the circadian pattern of intestinal P-gp expression with an unprecedented precision. The circadian timing depending on the P-gp expression rhythms may play a crucial role in the tolerability/toxicity of everolimus. The circadian changes in mdr1a genes deserve further studies regarding their relevance for in vitro and in vivo chronotolerance of mdr1a-transported anticancer drugs. Chronotherapy with P-gp-effluxed anticancer drugs could then be applied according to their rhythmic patterns in host and tumor to jointly maximize treatment efficacy and minimize toxicity.

Keywords: circadian rhythm, chronotoxicity, everolimus, mdr1a::Luc mice, p-glycoprotein

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Authors: Asghar Abbas, Rao Zahid Abbas, Muhammad Asif Raza, Kashif Hussain

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In the current experiment, in vitro anticoccidial potential of Vitis venifera (grape seed) extract was evaluated. For this purpose, an in vitro sporulation inhibition assay was used. Collected oocysts of different Eimeria species of chicken were exposed to six different concentrations (w/v) of Vitis venifera extract (TAE) in 10% dimethylsulphoxide solution (DMSO). Dimethylsulphoxide (DMSO) and potassium dichromate solution (K₂Cr₂O₇) served as control groups. Results of the study revealed that Vitis venifera extract (TAE) showed an inhibitory effect on sporulation (%) and damage (%) of Eimeria oocysts in a dose-dependent manner as compared to both control groups. Vitis venifera extract also damaged the morphology of oocysts in terms of shape, size, and number of sporocysts.

Keywords: Vitis venifera, in vitro, Eimeria, oocysts

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Authors: Ravi J. Shah

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The present study deals with the synthesis of novel spiro[azetidine-2, 3’-indole]-2’, 4(1’H)-dione derivative from the reactions of 3-(phenylimino)-1,3-dihydro-2H-indol-2-one derivatives with chloracetyl chloride in presence of triethyl amine (TEA). All the compounds were characterized using IR, 1H NMR, MS and elemental analysis. They were screened for their antibacterial and antifungal activities. Results revealed that, compounds (7a), (7b), (7c), (7d) and (7e) showed very good activity with MIC value of 6.25-12.5 μg/ml against three evaluated bacterial strains and the remaining compounds showed good to moderate activity comparable to standard drugs as antibacterial agents. Compounds (7c) and (7h) displayed equipotent antifungal activity in comparison to standard drugs. Structure-activity relationship study of the compounds showed that the presence of electron withdrawing group substitution at 5’ and 7’ positions of indoline ring and on ortho or para position of phenyl ring increases both antibacterial and antifungal activity of the compound. Henceforth, our findings will have a good impact on chemists and biochemists for further investigations in search of bromine containing spiro fused antimicrobial agents.

Keywords: antibacterial activity, antifungal activity, 2-Azetidinone, indoline

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Authors: Kun Chun, Jin-Chun Heo, Sang-Han Lee

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Asthma is a chronic airway inflammatory disease characterized by the infiltration of inflammatory cells and tissue remodeling. In this study, we examined the anti-asthmatic activity of a derivative of L-allo threonine by in vitro and in vivo anti-asthmatic assays. Ovalbumin (OVA)-induced C57BL/6 mice were used to analyze lung inflammation and cytokine expressions for exhibiting anti-atopic activity of the derivative. LX519290, a derivative of L-allo threonine, induced an increased IFN-γ and a decreased IL-10 mRNA level. This compound exhibited potent anti-asthmatic activity by decreasing immune cell infiltration in the lung, and IL-4 and IL-13 cytokine levels in the serum of OVA-induced mice. These results indicated that chronic airway injury was decreased by LX519290. We also assessed that LX519290 inhibits infiltration of immune cell, mucus release and cytokine expression in an in vivo model. Our results collectively suggest that the L-allo threonine is effective in alleviating asthmatic symptoms by treating inflammatory factors in the lung.

Keywords: asthma, L -allo threonine, LX519290, mice

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Authors: Eva Tvrdá, Boris Botman, Marek Halenár, Tomáš Slanina, Norbert Lukáč

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In vitro storage and processing of animal semen represents a risk factor to spermatozoa vitality, potentially leading to reduced fertility. A variety of substances isolated from natural sources may exhibit protective or antioxidant properties on the spermatozoon, thus extending the lifespan of stored ejaculates. This study compared the ability of different concentrations of the Salvia officinalis extract on the motility, mitochondrial activity, viability and reactive oxygen species (ROS) production by bovine spermatozoa during different time periods (0, 2, 6 and 24 h) of in vitro culture. Spermatozoa motility was assessed using the Computer-assisted sperm analysis (CASA) system. Cell viability was examined using the metabolic activity MTT assay, the eosin-nigrosin staining technique was used to evaluate the sperm viability and ROS generation was quantified using luminometry. The CASA analysis revealed that the motility in the experimental groups supplemented with 0.5-2 µg/mL Salvia extract was significantly lower in comparison with the control (P<0.05; Time 24 h). At the same time, a long-term exposure of spermatozoa to concentrations ranging between 0.05 µg/mL and 2 µg/mL had a negative impact on the mitochondrial metabolism (P<0.05; Time 24 h). The viability staining revealed that 0.001-1 µg/mL Salvia extract had no effects on bovine male gametes, however 2 µg/mL Salvia had a persisting negative effect on spermatozoa (P<0.05). Furthermore 0.05-2 µg/mL Salvia exhibited an immediate ROS-promoting effect on the sperm culture (P>0.05; Time 0 h and 2 h), which remained significant throughout the entire in vitro culture (P<0.05; Time 24 h). Our results point out to the necessity to examine specific effects the biomolecules present in Salvia officinalis may have individually or collectively on the in vitro sperm vitality and oxidative profile.

Keywords: bulls, CASA, MTT test, reactive oxygen species, sage, Salvia officinalis, spermatozoa

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Authors: Jamila Fliou, Federica Spinola, Ouassima Riffi, Asmaa Zriouel, Ali Amechrouq, Luca Nalbone, Alessandro Giuffrida, Filippo Giarratana

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This study performed a preliminary evaluation of the phytochemical composition and in vitro antibacterial activity of ethanolic extracts of Lavandula x intermedia leaves and flowers collected in the Fez-Meknes region of Morocco. Phytochemical analyses comprised qualitative colourimetric determinations of alkaloids, anthraquinones, and terpenes and quantitative analysis of total polyphenols, flavonoids, and condensed tannins by UV spectrophotometer. Antibacterial activity was evaluated by determining minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values against different ATCC bacterial strains. The phytochemical analysis showed a high amount of total polyphenols, flavonoids, and tannins in the leaf extract and a higher amount of terpenes based on colourimetric reaction than the flower extract. A positive colourimetric reaction for alkaloids and anthraquinones was detected for both extracts. The antibacterial activity of leaves and flower extract was not different against Gram-positive and Gram-negative strains (p<0.05). The results of the present study suggest the possible use of ethanolic extracts of L. x intermedia collected in the Fez-Meknes region of Morocco as a natural agent against bacterial pathogens.

Keywords: antimicrobial activity, Lavandula spp., lavender, lavandin, UV spectrophotometric analysis

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Authors: Mohamed E. Khalifa, Adil A. Gobouri

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Potential synthesis of a series of 3-amino-4-arylazothiophene derivatives from reaction of 2-cyano-2-phenylthiocarbamoyl acetamide and the appropriate α-halogenated reagents, followed by coupling with different aryl diazonium salts (Japp-Klingemann reaction), and another series of 5-arylazo-thiazol-2-ylcarbamoyl-thiophene derivatives from base-catalyzed intramolecular condensation of 5-arylazo-2-(N-chloroacetyl)amino-thiazole with selected B-keto compounds (Thorpe-Ziegler reaction) was performed. The biological activity of the two series was studied in vitro. Their versatility for pharmaceutical purposes was reported, where they displayed remarkable activities against selected pathogenic microorganisms; Bacillus subtilize, Staphylococcus aureus (Gram positive bacteria), Escherichia coli, Pseudomonas aeruginosa (Gram negative bacteria) and Aspergillus flavus, Candida albicans (fungi) with various degrees related to their chemical structures.

Keywords: thiophene, 2-aminothiazole, compounds, antioxidant, antitumor, antimicrobial

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Authors: A. M. Plante, F. O’Halloran, A. L. McCarthy

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By 2050 it is projected that 2 billion of the global population will be more than 60 years old. Older adults have unique dietary requirements and aging is associated with physiological changes that affect appetite, sensory perception, metabolism, and digestion. Therefore, it is essential that foods recommended and designed for older adults promote healthy aging. To assess cheese as a functional food for the elderly, a range of commercial cheese products were selected and compared for their antioxidant properties. Cheese from various milk sources (bovine, goats, sheep) with different textures and fat content, including cheddar, feta, goats, brie, roquefort, halloumi, wensleydale and gouda, were initially digested with two different simulated in vitro gastrointestinal digestion (SGID) models. One SGID model represented a validated in vitro adult digestion system and the second model, an elderly SGID, was designed to consider the physiological changes associated with aging. The antioxidant potential of all cheese digestates was investigated using in vitro chemical-based antioxidant assays, (2,2-Diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferric reducing antioxidant power (FRAP) and total phenolic content (TPC)). All adult model digestates had high antioxidant activity across both DPPH ( > 70%) and FRAP ( > 700 µM Fe²⁺/kg.fw) assays. Following in vitro digestion using the elderly SGID model, full-fat red cheddar, low-fat white cheddar, roquefort, halloumi, wensleydale, and gouda digestates had significantly lower (p ≤ 0.05) DPPH radical scavenging properties compared to the adult model digestates. Full-fat white cheddar had higher DPPH radical scavenging activity following elderly SGID digestion compared to the adult model digestate, but the difference was not significant. All other cheese digestates from the elderly model were comparable to the digestates from the adult model in terms of radical scavenging activity. The FRAP of all elderly digestates were significantly lower (p ≤ 0.05) compared to the adult digestates. Goats cheese was significantly higher (p ≤ 0.05) in FRAP (718 µM Fe²/kg.fw) compared to all other digestates in the elderly model. TPC levels in the soft cheeses (feta, goats) and low-fat cheeses (red cheddar, white cheddar) were significantly lower (p ≤ 0.05) in the elderly digestates compared to the adult digestates. There was no significant difference in TPC levels, between the elderly and adult model for full-fat cheddar (red, white), roquefort, wensleydale, gouda, and brie digestates. Halloumi cheese was the only cheese that was significantly higher in TPC levels following elderly digestion compared to adult digestates. Low fat red cheddar had significantly higher (p ≤ 0.05) TPC levels compared to all other digestates for both adult and elderly digestive systems. Findings from this study demonstrate that aging has an impact on the bioactivity of cheese, as antioxidant activity and TPC levels were lower, following in vitro elderly digestion compared to the adult model. For older adults, soft cheese, particularly goats cheese, was associated with high radical scavenging and reducing power, while roquefort cheese had low antioxidant activity. Also, elderly digestates of halloumi and low-fat red cheddar were associated with high TPC levels. Cheese has potential as a functional food for the elderly, however, bioactivity can vary depending on the cheese matrix. Funding for this research was provided by the RISAM Scholarship Scheme, Cork Institute of Technology, Ireland.

Keywords: antioxidants, cheese, in-vitro digestion, older adults

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Authors: Yuva Bellik, Selles Mohamed Amar

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The beneficial health effects including antioxidant properties of mint (Mentha spicata) and honey bees (Apis mellifera) have been extensively studied. However, there is no data about the effects of their associated use. In this study the total phenolic and flavonoid contents for individual extracts of mint and honey and their combination were determined. The antioxidant activity was investigated by using reducing power, 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2´- azinobis-(3-ethylbenzothiazoline-6-sulphonic acid diamonium salt (ABTS), and chelating power methods. The results showed that individual extracts contained important quantity of phenolics and flavonoids and their combination was found to produce best antioxidant activity. A significant linear correlation between the phenolic/flavonoid contents and antioxidant activity, especially with reducing power and free radical scavenging abilities, was observed.

Keywords: honey, mint, synergy, antioxidant activity

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Authors: Awatif Boumaza, Abderraouf Hilali, Hayat Talbi, Houda Sbayou

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The methanolic extract of Zygophyllum cornutun coss, an Algerian medicinal plant, was screened to the presence of mutagenic activity and genotoxic effect using the Ames test (Salmonella/microsome) and the micronucleus assay respectively. Positive results were obtained with both tests. The Ames test showed mutagenic activity in the presence of microsomal activation, while negative result was observed without microsomal activation. In the micronucleus test, two parameters were evaluated: the frequency of the micronucleus that increased in a dose dependent way and the proliferation index that decreased according to the micronucleus frequency. Even that further studies must be carried out, the mutagenic activity and the genotoxic effect of Zygophyllum cornutum should be taken in consideration when used as therapeutic plant.

Keywords: ames test, micronucleus test, mutagenic activity, genotoxicity, Zygophyllum cornutum

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Authors: Meenu Mehta, Munish Garg

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The use of oral anticancer drugs from AYUSH system of medicine is widely increased among the society due to their low cost, enhanced efficacy, increased patient preference, lack of inconveniences related to infusion and they provide an opportunity to develop chronic treatment regimens. However, oral delivery of these drugs usually laid down by the limited bioavailability of the drug, which is associated with a wide variation. As most of the cytotoxic agents have a narrow therapeutic window and are dosed at or near the maximum tolerated dose, a wide variability in the bioavailability can negatively affect treatment result. It is estimated that 40% of active substances are poorly soluble in water. The improvement of bio-availability of drugs with such properties presents one of the greatest challenges in drug formulations. There are several techniques reported in literature. Among all these Self Emulsifying Drug Delivery System (SEDDS) has gained more attention due to enhanced oral bio-availability enabling a reduction in dose. Thus, SEDDS anticancer drugs will have the increased bioavailability and efficacy. These dosage form will provide societal benefit in a cost-effective manner as compared to other oral dosage forms. Present study reflects on the formulation strategies as SEDDS for oral anticancer agents of AYUSH system for enhanced bioavailability with proven efficacy by cancer cell lines.

Keywords: anticancer agents, AYUSH system, bioavailability, SEDDS

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Authors: O. Kahraman, M. S. Alatas, O. B. Citil

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In determination of values of feeds which, are used in ruminant nutrition, different methods are used like in vivo, in vitro, in situ or in sacco. Generally, the most reliable results are taken from the in vivo studies. But because of the disadvantages like being hard, laborious and expensive, time consuming, being hard to keep the experiment conditions under control and too much samples are needed, the in vitro techniques are more preferred. The most widely used in vitro techniques are two-staged digestion technique and gas production technique. In vitro gas production technique is based on the measurement of the CO2 which is released as a result of microbial fermentation of the feeds. In this review, the factors affecting the results obtained from in vitro gas production technique (Hohenheim Feed Test) were discussed. Some factors must be taken into consideration when interpreting the findings obtained in these studies and also comparing the findings reported by different researchers for the same feeds. These factors were discussed in 3 groups: factors related to animal, factors related to feeds and factors related with differences in the application of method. These factors and their effects on the results were explained. Also it can be concluded that the use of in vitro gas production technique in feed evaluation routinely can be contributed to the comprehensive feed evaluation, but standardization is needed in this technique to attain more reliable results.

Keywords: In vitro, gas production technique, Hohenheim feed test, standardization

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Authors: Kinza Khan, Dad Muhmmad, Asif Saleem, Nadia Mukhtar, Tahir Yaqub

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Ethomedicines are more commonly used in underdeveloped and developing countries. These medicines are sometimes more potent in controlling microbial infections than conventional medicines. Medicinal plants have been common practice to cure many diseases for centuries. Murraya koenigii is one of these plants and is commonly used in South Asian countries as a flavoring agent in food. To evaluate its anti-microbial activity, six different bacterial strains (Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Salmonella typhi, Bacillus cereus and Klebsiella pneumonia were used. N-hexane extract of Murraya koenigii leaves shows maximum activity against Bacillus cereus. Acetone extract of Murraya koenigii shoots showed more efficient activity against Pseudomonas aeruginosa Dichloromethane extracts showed maximum activity against Bacillus cereus. Ethanol extract exhibited maximum activity against Pseudomonas aeruginosa and Klebsiella pneumoniae. The methanol extract of Murraya koenigii shoots displayed maximum antibacterial activity against Bacillus cereus. Antifungal activity Ethanol extract was more effective against Candida albicans.

Keywords: ethnomedicines, bacteria, fungi, murraya koenigii, antimicrobial activity

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Authors: Sylwia K. Naliwajko, Justyna Moskwa, Patryk Nowakowski, Renata Markiewicz-Zukowska, Krystyna Gromkowska-Kepka, Anna Puscion-Jakubik, Maria H. Borawska

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Amygdalin is found in many fruit seeds, including apricot, peach, quince, apples, and almonds. Amygdalin (also named vitamin B17), as well as its sources, are commonly used as an alternative therapy or prevention of cancer. The potential activity of amygdalin is related to its enzymatic degradation to the hydrogen cyanide. Hydrogen cyanide is a toxic substance that causes liver and nerves damage, fever, coma or even death. Amygdalin is much better tolerated after intravenous than oral administration. The aim of this study was to examine the influence of amygdalin on glioblastoma multiforme cell lines. Three glioblastoma multiforme cell lines – U87MG, T98, LN18 were incubated (48 h) with amygdalin in concentrations 100, 250, 500, 1000 and 2000 µg/mL. The MTT (Thiazolyl Blue Tetrazolium Bromide) test and DNA binding test by [3H]-thymidine incorporation were used to determine the anti-proliferative activity of amygdalin. The secretion of metalloproteinases (MMP2 and MMP-9) from U87MG cells was estimated by gelatin zymography. The statistical analysis was performed using Statistica v. 13.0 software. The data was presented as a % of control. Amygdalin did not show significant inhibition of viability of all the glioblastoma cells in concentrations 100, 250, 500, 1000 µg/mL. In 2000 µg/mL there were significant differences compared to the control, but inhibition of viability was less than 20% (more than 80% of control). The average viability of U87MG cells was 92,0±4%, T98G: 85,8±3% and LN18: 94,7±2% of the control. There was no dose-response viability, and IC50 value was not recognized. DNA binding in U87MG cells was not inhibited (109,0±3 % of control). After treatment with amygdalin, we observed significantly increased secretion of MMP2 and MMP9 in U87MG cells (130,3±14% and 112,0±5% of control, respectively). Our results suggest that amygdalin has no anticancer activity in glioblastoma cell lines.

Keywords: amygdalin, anticancer, cell line, glioblastoma

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Authors: Alaa M. Badr Eldin, Mayar M. Shahen, Mohammed S. Sedeek, Marwa I. Ezzat, Sawsan M. ElSonbaty, Muhammed A. Saad, Manal S. Afifi, Omar M. Sabry

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Using HPLC-MS/MS technique, 133 polyphenolic compounds were identified in the methanol extract of pomegranate rind (Punica granatum L.). In-vitro cytotoxic activity against breast cancer cell line MCF-7 was investigated, with an IC50 of 54 ug/ml. In-silico molecular docking using ellagic acid, gallagic acid, and Punicalagin as model compounds identified in pomegranate rind extract confirmed the intriguing anti-estrogenic action of the key polyphenolic components in pomegranate rind extract. Surprisingly, taxol showed low activity compared to pomegranate compounds as ERα antagonist and ERβ agonist. Pomegranate rind extract enhanced apoptosis of breast cancer cells through upregulation of the caspase-3 expression and downregulation of NF-κB transcription factor.

Keywords: HPLC-MS/MS, pomegranate rind, cytotoxicity, MCF-7, ER, caspase-3, NF-kB

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Authors: Rajesh Kumar, Anil Kamboj

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Inhibition of α- amylase play a vital role in the clinical management of postprandial hyperglycemia. Although, powerful synthetic inhibitors are available, natural inhibitors are potentially safer. The present study was carried out to evaluate α- amylase inhibition activity from hydroalcoholic extracts from aerial parts of Cestrum nocturnum. Hydroalcoholic extract was prepared by Soxhletation Method. The extract showed strong inhibition towards α- amylase activity and IC50 value were 45.9 µg. This In vitro studies indicate the potential of C. nocturnum in the development of effective anti-diabetic agents.

Keywords: α- amylase, cestrum nocturnum, hyperglycemia, hydroalcoholic extracts, diabetes

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Authors: Omar M. Al Aufi, Abdulwahab Noorwali, Ahmed Al Abd, Safia Alattas, Fathya Zahran, Fahd Almutairi

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Cisplatin (CDDP) is a potent anticancer agent used for several tumor types. Thymoquinone (TQ) is a naturally occurring compound drawing great attention as an anticancer and chemomodulator for chemotherapies. Herein, we studied the potential cytotoxicity of thymoquinone, CDDP and their combination against human oral squamous cell carcinoma cells in contrast to normal oral epithelial cells. CDDP similarly killed both head and neck squamous cell carcinoma cells (UMSCC-14C) and normal oral epithelial cells (OEC). TQ alone exerted considerable cytotoxicity against UMSCC-14C cells, while it induced a weaker killing effect against normal oral epithelial cells (OEC). The equitoxic combination of TQ and CDDP showed additive to synergistic interaction against both UMSCC-14C and OEC cells. TQ alone increased apoptotic cell fraction in UMSCC-14C cells as early as after 6 hours. In addition, prolonged exposure of UMSCC-14C to TQ alone resulted in 96.7±1.6% total apoptosis, which was increased after combination with CDDP to 99.3±1.2% in UMSCC-14C cells. On the other hand, TQ induced a marginal increase in the apoptosis in OEC and even decreased the apoptosis induced by CDDP alone. Finally, apoptosis induction results were confirmed by the change in the expression levels of p53, Bcl-2 and Caspase-9 proteins in both UMSCC-14c and OEC cells.

Keywords: thymoquinone, cisplatin, apoptosis, oral squamous cell carcinoma, P53, Caspase-9, Bcl-2

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Authors: Zeinab Hajifathali, Moghan Amirhosseinian

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This study had two main aims: firstly, to determine how the individual substitution of CaO/MgO and CaO/SrO can affect the in vitro bioactivity of sol-gel derived substituted 58S bioactive glass (BG) and secondly to introduce a composition in the 60SiO₂–(36-x)CaO–4P₂O₅–(x)MgO and 60SiO₂–(36-x)CaO–4P₂O₅–(x)SrO quaternary systems (where x= 0, 5, 10 mol.%) with enhanced biocompatibility, alkaline phosphatase (ALP) activity, and more efficient antibacterial activity against MRSA bacteria. Results showed that both magnesium-substituted bioactive glasses (M-BGs) and strontium- substituted bioactive glasses (S-BGs) retarded the Hydroxyapatite (HA) formation. Meanwhile, magnesium had more pronounced effect. The 3-(4, 5dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and ALP assays revealed that the presence of moderate amount (5 mol%) of Mg and Sr had a stimulating effect on increasing of both proliferation and differentiation of MC3T3-E1 cells. Live dead and Dapi/actin staining revealed both substitution of CaO/MgO and CaO/SrO resulted in more biocompatibility and stimulation potential of the MC3T3 cells compared with control. Taken together, among all of the synthesized magnesium substituted (MBGs) and strontium substituted (SBGs), the sample 58- BG with 5 mol% CaO/MgO substitution (BG-5M) was considered as a multifunctional biomaterial in bone tissue regeneration field with enhanced biocompatibility, ALP activity as well as the highest antibacterial efficiency against methicillin-resistant Staphylococcus aureus (MRSA) bacteria.

Keywords: apatite, alkaline earth, bioactivity, biomedical applications, Sol-gel

Procedia PDF Downloads 158