Search results for: in vitro assay

Authors: Seok Woo Chang, Kee Yeon Kum, Kwang Shik Bae, WooCheol Lee

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Aim: The objective of this study was to compare the biocompatibilities and mineralization potential of ProRoot MTA and newly developed calcium phosphate based cement, Capseal. Materials and Methods: The biocompatibilities and mineralization-related gene expressions (Bone sialoprotein (BSP) and osteocalcin (OCN)) of ProRoot MTA and Capseal were also compared by a methylthiazol tetrazolium (MTT) assay and reverse transcription-polymerization chain reaction (RT-PCR) analysis on 1, 3, and 7 days, respectively. Empty rings were used as control group. The results were statistically analyzed by Kruskal-Wallis test with a Bonferroni correction. P-value of < 0.05 was considered significant. Results: The biocompatibilities of ProRoot MTA and Capseal were equally favorable. ProRoot MTA and Capseal affected the messenger RNA expression of osteocalcin and osteonectin. Conclusions: Based on the results, both ProRoot MTA and Capseal could be a useful biomaterial in clinical endodontics.

Keywords: biocompatibility, calcium silicate cement, MTT, RT-PCR

Procedia PDF Downloads 385

Authors: Mashayekh Amir Shahriar, Akhlaghi Morteza, Rajaee Hajar, Khani Mohammad Reza, Shokri Babak

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A new and novel approach in medicine is the use of cold plasma for various applications such as sterilization blood coagulation and cancer cell treatment. In this paper a pin-to-hole plasma jet suitable for biological applications is investigated, characterized and the possibility and feasibility of cancer cell treatment is evaluated. The characterization includes power consumption via Lissajous method, thermal behavior of plasma using Infra-red camera as a novel method, Optical Emission Spectroscopy (OES) to determine the species that are generated. Treatment of leukemia cancer cells is also implemented and MTT assay is used to evaluate viability.

Keywords: Atmospheric Pressure Plasma Jet (APPJ), Plasma Medicine, Cancer cell treatment, leukemia, Optical Emission

Procedia PDF Downloads 647

Authors: Tamar Lin, Nufar Buchshtab, Yifat Elharar, Julian Nicenboim, Rotem Edgar, Iddo Weiner, Lior Zelcbuch, Ariel Cohen, Sharon Kredo-Russo, Inbar Gahali-Sass, Naomi Zak, Sailaja Puttagunta, Merav Bassan

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Background: Atopic dermatitis (AD) is a chronic, relapsing inflammatory skin disorder that is characterized by dry skin and flares of eczematous lesions and intense pruritus. Multiple lines of evidence suggest that AD is associated with increased colonization by Staphylococcus aureus, which contributes to disease pathogenesis through the release of virulence factors that affect both keratinocytes and immune cells, leading to disruption of the skin barrier and immune cell dysfunction. The aim of the current study is to develop a bacteriophage-based product that specifically targets S. aureus. Methods: For the discovery of phage, environmental samples were screened on 118 S. aureus strains isolated from skin samples, followed by multiple enrichment steps. Natural phages were isolated, subjected to Next-generation Sequencing (NGS), and analyzed using proprietary bioinformatics tools for undesirable genes (toxins, antibiotic resistance genes, lysogeny potential), taxonomic classification, and purity. Phage host range was determined by an efficiency of plating (EOP) value above 0.1 and the ability of the cocktail to completely lyse liquid bacterial culture under different growth conditions (e.g., temperature, bacterial stage). Results: Sequencing analysis demonstrated that the 118 S. aureus clinical strains were distributed across the phylogenetic tree of all available Refseq S. aureus (~10,750 strains). Screening environmental samples on the S. aureus isolates resulted in the isolation of 50 lytic phages from different genera, including Silviavirus, Kayvirus, Podoviridae, and a novel unidentified phage. NGS sequencing confirmed the absence of toxic elements in the phages’ genomes. The host range of the individual phages, as measured by the efficiency of plating (EOP), ranged between 41% (48/118) to 79% (93/118). Host range studies in liquid culture revealed that a subset of the phages can infect a broad range of S. aureus strains in different metabolic states, including stationary state. Combining the single-phage EOP results of selected phages resulted in a broad host range cocktail which infected 92% (109/118) of the strains. When tested in vitro in a liquid infection assay, clearance was achieved in 87% (103/118) of the strains, with no evidence of phage resistance throughout the study (24 hours). A S. aureus host was identified that can be used for the production of all the phages in the cocktail at high titers suitable for large-scale manufacturing. This host was validated for the absence of contaminating prophages using advanced NGS methods combined with multiple production cycles. The phages are produced under optimized scale-up conditions and are being used for the development of a topical formulation (BX005) that may be administered to subjects with atopic dermatitis. Conclusions: A cocktail of natural phages targeting S. aureus was effective in reducing bacterial burden across multiple assays. Phage products may offer safe and effective steroid-sparing options for atopic dermatitis.

Keywords: atopic dermatitis, bacteriophage cocktail, host range, Staphylococcus aureus

Procedia PDF Downloads 147

Authors: Lin Po-Hsi, Sheu Ming-Thau

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Urea cycle disorders (UCD) are rare genetic metabolic disorders that compromise the body's urea cycle. Sodium phenylbutyrate (SPB) is a medication commonly administered in tablet or powder form to lower ammonia levels. Nonetheless, its high sodium content poses risks to sodium-sensitive UCD patients. This necessitates the creation of an alternative drug formulation to mitigate sodium load and optimize drug delivery for UCD patients. This study focused on crafting a novel oral drug formulation for UCD, leveraging choline bicarbonate and phenylbutyric acid. The active pharmaceutical ingredient-ionic liquids (API-ILs) and therapeutic deep eutectic systems (THEDES) were formed by combining these with choline chloride. These systems display characteristics like maintaining a liquid state at room temperature and exhibiting enhanced solubility. This in turn amplifies drug dissolution rate, permeability, and ultimately oral bioavailability. Incorporating choline-based phenylbutyric acid as a substitute for traditional SPB can effectively curtail the sodium load in UCD patients. Our in vitro dissolution experiments revealed that the ILs and DESs, synthesized using choline bicarbonate and choline chloride with phenylbutyric acid, surpassed commercial tablets in dissolution speed. Pharmacokinetic evaluations in SD rats indicated a notable uptick in the oral bioavailability of phenylbutyric acid, underscoring the efficacy of choline salt ILs in augmenting its bioavailability. Additional in vitro intestinal permeability tests on SD rats authenticated that the ILs, formulated with choline bicarbonate and phenylbutyric acid, demonstrate superior permeability compared to their sodium and acid counterparts. To conclude, choline salt ILs developed from choline bicarbonate and phenylbutyric acid present a promising avenue for UCD treatment, with the added benefit of reduced sodium load. They also hold merit in formulation engineering. The sustained-release capabilities of DESs position them favorably for drug delivery, while the low toxicity and cost-effectiveness of choline chloride signal potential in formulation engineering. Overall, this drug formulation heralds a prospective therapeutic avenue for UCD patients.

Keywords: phenylbutyric acid, sodium phenylbutyrate, choline salt, ionic liquids, deep eutectic systems, oral bioavailability

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Authors: Payman A. A. Zibari, Mosleh M. S. Duhoky

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These experiments were conducted at Tissue Culture Laboratory/ Faculty of Agriculture / University of Duhok during the period from January 2011 to May 2013. The objectives of this study were to study the effects of interaction between colchcine concentrations and treatment time duration of Creps capilaris (with and without 2B chromosome) on chromosome polyploidy during fifteen passages until regeneration of plants from the callus. Data showed that high percentage of chromosome polyploidy approximately can be obtained from high concentration of colchicin and long time of duration.

Keywords: polyploidy, Crepis capilaris, colchicine, B chromosome

Procedia PDF Downloads 185

Authors: Kyriaki Hatziagapiou, Konstantinos Bethanis, Olti Nikola, Elias Christoforides, Eleni Koniari, Eleni Kakouri, George Lambrou, Christina Kanaka-Gantenbein

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Glioblastoma multiforme (GBM) remains a serious health challenge, as current therapeutic modalities continue to yield unsatisfactory results, with the average survival rarely exceeding 1-2 years. Natural compounds still provide some of the most promising approaches for discovering new drugs. The non-psychotropic cannabidiol (CBD) deriving from Cannabis sativa L. provides such promise. CBD is endowed with anticancer, antioxidant, and genoprotective properties as established in vitro and in in vivo experiments. CBD’s selectivity towards cancer cells and its safe profile suggest its usage in cancer therapies. However, the bioavailability of oral CBD is low due to poor aqueous solubility, erratic gastrointestinal absorption, and significant first-pass metabolism, hampering its therapeutic potential and resulting in a variable pharmacokinetic profile. In this context, CBD can take great advantage of nanomedicine-based formulation strategies. Cyclodextrins (CDs) are cyclic oligosaccharides used in the pharmaceutical industry to incorporate apolar molecules inside their hydrophobic cavity, increasing their stability, water solubility, and bioavailability or decreasing their side effects. CBD-inclusion complexes with CDs could be a good strategy to improve its properties, like solubility and stability to harness its full therapeutic potential. The current research aims to study the potential cytotoxic effect of CBD and CBD-CDs complexes CBD-RMβCD (randomly methylated β-cyclodextrin) and CBD-HPβCD (hydroxypropyl-b-CD) on the A172 glioblastoma cell line. CBD is diluted in 10% DMSO, and CBD/CDs solutions are prepared by mixing solid CBD, solid CDs, and dH2O. For the biological assays, A172 cells are incubated at a range of concentrations of CBD, CBD-RMβCD and CBD-HPβCD, RMβCD, and HPβCD (0,03125-4 mg/ml) at 24, 48, and 72 hours. Analysis of cell viability after incubation with the compounds is performed with Alamar Blue viability assay. CBD’s dilution to DMSO 10% was inadequate, as crystals are observed; thus cytotoxicity experiments are not assessed. CBD’s solubility is enhanced in the presence of both CDs. CBD/CDs exert significant cytotoxicity in a dose and time-dependent manner (p < 0.005 for exposed cells to any concentration at 48, 72, and 96 hours versus cells not exposed); as their concentration and time of exposure increases, the reduction of resazurin to resofurin decreases, indicating a reduction in cell viability. The cytotoxic effect is more pronounced in cells exposed to CBD-HPβCD for all concentrations and time-points. RMβCD and HPβCD at the highest concentration of 4 mg/ml also exerted antitumor action per se since manifesting cell growth inhibition. The results of our study could afford the basis of research regarding the use of natural products and their inclusion complexes as anticancer agents and the shift to targeted therapy with higher efficacy and limited toxicity. Acknowledgments: The research is partly funded by ΙΚΥ (State Scholarships Foundation) – Post-doc Scholarships-Partnership Agreement 2014-2020.

Keywords: cannabidiol, cyclodextrins, glioblastoma, hydroxypropyl-b-Cyclodextrin, randomly-methylated-β-cyclodextrin

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Authors: N. P. D. Tran, H. Q. D. Nguyen, M. C. Yang

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In this study, contact lenses were prepared through the polymerization of tris-(trimethyl-silyl-propyl-methacrylate) (TRIS), N,N-dimethylacrylamide (DMA), N-vinylpyrrolidone (NVP), and cross-linked with ethylene glycol dimethylacrylate (EGDMA). The equilibrium water content (EWC), oxygen permeability (Dk), light transmittance, and in vitro cytotoxicity of TRIS-DMA-NVP with various ratios were measured. The results showed that the EWC increased while the Dk decreased with the increase of NVP content. For the sample with 25 wt% NVP, the EWC attained 53% whereas the Dk decreased to 46 barrers. All these lenses exhibited light transmittance over than 95%. In addition, all these lenses exhibited no inhibition to the growth of L292 fibroblasts. Thus, this study showed that TRIS-DMA-NVP can be applicable for making contact lens.

Keywords: DMA, TRIS, NVP, silicone hydrogel, contact lens

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Authors: Christi Jackson, Chuka Onaga

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Introduction: We report a case of delayed diagnosis of extra-pulmonary INH-mono-resistant Tuberculosis (TB) in a South African patient with drug-resistant HIV. Case Presentation: A 36-year old male was initiated on 1st line (NNRTI-based) anti-retroviral therapy (ART) in September 2009 and switched to 2nd line (PI-based) ART in 2011, according to local guidelines. He was following up at the outpatient wellness unit of a public hospital, where he was diagnosed with Protease Inhibitor resistant HIV in March 2016. He had an HIV viral load (HIVVL) of 737000 copies/mL, CD4-count of 10 cells/µL and presented with complaints of productive cough, weight loss, chronic diarrhoea and a septic buttock wound. Several investigations were done on sputum, stool and pus samples but all were negative for TB. The patient was treated with antibiotics and the cough and the buttock wound improved. He was subsequently started on a 3rd-line ART regimen of Darunavir, Ritonavir, Etravirine, Raltegravir, Tenofovir and Emtricitabine in May 2016. He continued losing weight, became too weak to stand unsupported and started complaining of abdominal pain. Further investigations were done in September 2016, including a urine specimen for Line Probe Assay (LPA), which showed M. tuberculosis sensitive to Rifampicin but resistant to INH. A lymph node biopsy also showed histological confirmation of TB. Management and outcome: He was started on Rifabutin, Pyrazinamide and Ethambutol in September 2016, and Etravirine was discontinued. After 6 months on ART and 2 months on TB treatment, his HIVVL had dropped to 286 copies/mL, CD4 improved to 179 cells/µL and he showed clinical improvement. Pharmacy supply of his individualised drugs was unreliable and presented some challenges to continuity of treatment. He successfully completed his treatment in June 2017 while still maintaining virological suppression. Discussion: Several laboratory-related factors delayed the diagnosis of TB, including the unavailability of urine-lipoarabinomannan (LAM) and urine-GeneXpert (GXP) tests at this facility. Once the diagnosis was made, it presented a treatment dilemma due to the expected drug-drug interactions between his 3rd-line ART regimen and his INH-resistant TB regimen, and specialist input was required. Conclusion: TB is more difficult to diagnose in patients with severe immunosuppression, therefore additional tests like urine-LAM and urine-GXP can be helpful in expediting the diagnosis in these cases. Patients with non-standard drug regimens should always be discussed with a specialist in order to avoid potentially harmful drug-drug interactions.

Keywords: drug-resistance, HIV, line probe assay, tuberculosis

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Authors: Praskovya Britskaya, Fatima Shaizadina, Alua Omarova, Nessipkul Alysheva

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Planned preventive vaccination, which is carried out in the Republic of Kazakhstan, promoted permanent decrease in the incidence of measles and viral hepatitis B. In the structure of VHB patients prevail people of young, working age. Monitoring of infectious incidence, monitoring of coverage of immunization of the population, random serological control over the immunity enable well-timed identification of distribution of the activator, effectiveness of the taken measures and forecasting. The serological blood analysis was conducted in indicator groups of the population of Central Kazakhstan for the purpose of identification of antibody titre for vaccine preventable infections (measles, viral hepatitis B). Measles antibodies were defined by method of enzyme-linked assay (ELA) with test-systems "VektoKor" – Ig G ('Vektor-Best' JSC). Antibodies for HBs-antigen of hepatitis B virus in blood serum was identified by method of enzyme-linked assay (ELA) with VektoHBsAg test systems – antibodies ('Vektor-Best' JSC). The result of the analysis is positive, the concentration of IgG to measles virus in the studied sample is equal to 0.18 IU/ml or more. Protective level of concentration of anti-HBsAg makes 10 mIU/ml. The results of the study of postvaccinal measles immunity showed that the share of seropositive people made 87.7% of total number of surveyed. The level of postvaccinal immunity to measles in age groups differs. So, among people older than 56 the percentage of seropositive made 95.2%. Among people aged 15-25 were registered 87.0% seropositive, at the age of 36-45 – 86.6%. In age groups of 25-35 and 36-45 the share of seropositive people was approximately at the same level – 88.5% and 88.8% respectively. The share of people seronegative to a measles virus made 12.3%. The biggest share of seronegative people was found among people aged 36-45 – 13.4% and 15-25 – 13.0%. The analysis of results of the examined people for the existence of postvaccinal immunity to viral hepatitis B showed that from all surveyed only 33.5% have the protective level of concentration of anti-HBsAg of 10 mIU/ml and more. The biggest share of people protected from VHB virus is observed in the age group of 36-45 and makes 60%. In the indicator group – above 56 – seropositive people made 4.8%. The high percentage of seronegative people has been observed in all studied age groups from 40.0% to 95.2%. The group of people which is least protected from getting VHB is people above 56 (95.2%). The probability to get VHB is also high among young people aged 25-35, the percentage of seronegative people made 80%. Thus, the results of the conducted research testify to the need for carrying out serological monitoring of postvaccinal immunity for the purpose of operational assessment of the epidemiological situation, early identification of its changes and prediction of the approaching danger.

Keywords: antibodies, blood serum, immunity, immunoglobulin

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Authors: Shashi Bala, Neha A. Chugh, Subhash C. Bansal, Mohal L. Garg, Ashwani Koul

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Purpose: The present study was designed to evaluate the radioprotective efficacy of Aloe vera from whole body X-ray exposure in rodents. Materials and Methods: For this purpose, after on week’s acclimatization, male balb/c mice procured from Central Animal House, Panjab University, Chandigarh (India), were divided into four groups: Group I mice served as control. Group II mice were orally administrated Aloe vera pulp extract (50 mg/ kg body weight) on alternate days for 30 days. Group III mice were subjected to whole body X-ray irradiation to cumulative dose of 2Gy (0.258Gy twice a day for four days in the last week). Group IV animals were pretreated with Aloe vera pulp extract on alternate days as in Group II and in the last week of the study, they were exposed to X-ray as in Group III. Results: Spleen of X-ray irradiated mice showed histopathological alterations accompanied with enhanced activity of lactate dehydrogenase (LDH) in serum. Elevated levels of reactive oxygen species (ROS), lipid peroxidation (LPO), enhanced activities in Glutathione based enzymes such as Glutathione peroxidase (GSH-Px), Glutathione reductase (GR), Catalase (CAT), Superoxide dismutase (SOD) associated with depletion in reduced Glutathione (GSH) concentration were observed after X-ray exposure in blood plasma and spleen.. Pro-inflammatory cytokines like tumor necrosis factors (TNF-α) and Inteleukin-6 (IL-6) levels were also found to be enhanced in serum of irradiated mice. Irradiation-induced significant elevation in Total leucocyte counts (TLC), neutrophil counts and decline in platelet counts, associated with unaltered levels of red blood cell counts (RBC’s) and haemoglobin (Hb) in various treatment groups. Clastogenic damage and apoptosis was also found to be increase in splenic tissue of X-ray exposed mice as assessed by micronucleus and TUNEL assay. However, X-ray irradiated animals administered with Aloe vera revealed significant improvement in levels of ROS/ LPO, LDH activity, and antioxidant mechanism. Aloe vera pretreated animals exhibited less severe damage, and early recovery in micronucleated cells, hematological parameters, apoptotic cells and inflammatory markers as compared to X-ray exposed mice. Conclusion: These results indicate that the radioprotective potential of Aloe vera against X-ray induced damage. This may be due to its free radical scavenging, antioxidant, anti-apoptotic and anti-inflammatory properties.

Keywords: aloe vera, antioxidant defense system, lactate dehydrogenase (LDH), micronucleus assay, x-ray

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Authors: İsmail Çeli̇k, Gülgün Ayhan-Kılcıgi̇l, Arzu Onay-Beşi̇kçi̇

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In this study, some 2-(2-phenyl/substitutedphenyl)- lH-benzo[d]'imidazol-l-yl)-N'-(alkylthiosemicarbazide were designed and prepared. Firstly, 2-phenyl/ suhstitutedphenyl-lH-Benzo[d]imidazole was prepared via oxidative condensation of o-phenylenediamine, benzaldehyde and sodium metabisulfite. Treatment of the benzimidazole compound with ethyl chloroacetate in KOH/DMSO gave the ester compound ethyl 2-(2-substitutedphenyl)-1H-benzo[d]imidazol-l-yl)acetate. Hydrazine hydrate and the ester in ethanol were refluxed for 4 h to give 2-(2-phenyl/substitutedphenyl)-1H-benzo[d]imidazol-l-yl)acetohydrazide. Thiosemicarbazides were obtained by condensing acyl hydrazide with the alkylisothiocyanate in ethanol. Following the structure elucidation, benzimidazole compounds were tested for their EGFR kinase inhibitory activities by using ADP-GloTM Kinase Assay.

Keywords: benzimidazole, EGFR kinase inhibitor, synthesis, thiosemicarbazide

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Authors: Nishanth Venugopal Menon, Chuah Yon Jin, Samantha Phey, Wu Yingnan, Zhang Ying, Vincent Chan, Kang Yuejun

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Cell Migration is a vital phenomenon that the cells undergo in various physiological processes like wound healing, disease progression, embryogenesis, etc. Cell migration depends primarily on the chemical and physical cues available in the cellular environment. The chemical cue involves the chemokines secreted and gradients generated in the environment while physical cues indicate the impact of matrix properties like nanotopography and stiffness on the cells. Mesenchymal Stem Cells (MSCs) have been shown to have a role wound healing in vivo and its migration to the site of the wound has been shown to have a therapeutic effect. In the field of stem cell based tissue regeneration of bones and cartilage, one approach has been to introduce scaffold laden with MSCs into the site of injury to enable tissue regeneration. In this work, we have studied the combinatorial impact of the substrate physical properties on MSC migration. A microfluidic in vitro model was created to perform the migration studies. The microfluidic model used is a three compartment device consisting of two cell seeding compartments and one migration compartment. Four different PDMS substrates with varying substrate roughness, stiffness and hydrophobicity were created. Its surface roughness and stiffness was measured using Atomic Force Microscopy (AFM) while its hydrphobicity was measured from the water contact angle using an optical tensiometer. These PDMS substrates are sealed to the microfluidic chip following which the MSCs are seeded and the cell migration is studied over the period of a week. Cell migration was quantified using fluorescence imaging of the cytoskeleton (F-actin) to find out the area covered by the cells inside the migration compartment. The impact of adhesion proteins on cell migration was also quantified using a real-time polymerase chain reaction (qRT PCR). These results suggested that the optimal substrate for cell migration would be one with an intermediate level of roughness, stiffness and hydrophobicity. A higher or lower value of these properties affected cell migration negatively. These observations have helped us in understanding that different substrate properties need to be considered in tandem, especially while designing scaffolds for tissue regeneration as cell migration is normally impacted by the combinatorial impact of the matrix. These observations may lead us to scaffold optimization in future tissue regeneration applications.

Keywords: cell migration, microfluidics, in vitro model, stem cell migration, scaffold, substrate properties

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Authors: M. Lamamra, H. Laouer, A. Adjaoud, Sahli Farida

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Essential oils can have significant antimicrobial activities and can successfully replace antibiotics that show their ineffectiveness against resistant germs. The chemical composition of the essential oil obtained by hydrodistillation from the aerial part of Daucus aristidis (Apiaceae) at the pre-flowering stage was investigated for the first time, by GC and GC-MS and evaluated for in vitro antimicrobial activity by the disk diffusion method. The Main components of D. aristidis oil were α-pinene (20.13%), cedrol (20.11%), and E- asarone (18.53%). The oil exhibited an antibacterial activity against almost strains tested except for Klebsiella pneumoniae ATCC 700603 K6 and Enterococcus faecalis ATCC 49452, the oil of D. aristidis had no activity against all fungi tested.

Keywords: α-pinene, antimicrobial activity, Daucus aridtidis, essential oil

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Authors: Shradha S. Binani, Pravin S. Bodke, Ravi V. Joat

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An efficient method has been described for the synthesis of 6-(substituted aryl)-4-(2'- hydroxy-5'-chlorophenyl)-1, 6-dihydropyrimidine-2-thiol, as a beneficial antibacterial and antifungal agents. The diketones of title compounds were synthesized in four steps and subsequently these diketones were further reacted with thiourea in the presence of DMF, which led to the formation of dihydropyrimidine derivatives 5 (a-f). Compounds 5 (a-f) were screened for their in vitro antibacterial and antifungal activity by agar well method. Compounds 5b, 5c, 5e, and 5f were exhibited significant antimicrobial potential against tested strains at 50ug/ml and 100ug/ml concentrations. Six novel dihydropyrimidine analogues have been synthesized, characterized and found to be promising antibacterial and antifungal agents.

Keywords: diketones, dihyropyrimidine, antimicrobial activity, thiol group

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Authors: Roihatul Mutiah, Sukardiman, Aty Widyawaruyanti

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Calotropis gigantiea (L.) R. Br (Apocynaceae) commonly called as “Biduri” or “giant milk weed” is a well-known weed to many cultures for treating various disorders. Several studies reported that C.gigantea roots has anticancer activity. The main aim of this research was to isolate and identify an active marker compound of C.gigantea roots for quality control purpose of its extract in the development as anticancer natural product. The isolation methods was bioactivity guided column chromatography, TLC, and HPLC. Evaluated anticancer activity of there substances using MTT assay methods. Identification structure active compound by UV, 1HNMR, 13CNMR, HMBC, HMQC spectral and other references. The result showed that the marker active compound was identical as Calotropin.

Keywords: calotropin, Calotropis gigantea, anticancer, marker active

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Authors: Yuanyuan Zhang, Yujuan Zheng, Giuseppina Barutello, Sumako Kameishi, Kungchun Chiu, Katharina Hennig, Martial Balland, Federica Cavallo, Lars Holmgren

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Angiogenesis describes that new blood vessels migrate from pre-existing ones to form 3D lumenized structure and remodeling. During directional migration toward the gradient of pro-angiogenic factors, the endothelial cells, especially the tip cells need filopodia to sense the environment and exert the pulling force. Of particular interest are the integrin proteins, which play an essential role in focal adhesion in the connection between migrating cells and extracellular matrix (ECM). Understanding how these biomechanical complexes orchestrate intrinsic and extrinsic forces is important for our understanding of the underlying mechanisms driving angiogenesis. We have previously identified Angiomotin (Amot), a member of Amot scaffold protein family, as a promoter for endothelial cell migration in vitro and zebrafish models. Hence, we established inducible endothelial-specific Amot knock-out mice to study normal retinal angiogenesis as well as tumor angiogenesis. We found that the migration ratio of the blood vessel network to the edge was significantly decreased in Amotec- retinas at postnatal day 6 (P6). While almost all the Amot defect tip cells lost migration advantages at P7. In consistence with the dramatic morphology defect of tip cells, there was a non-autonomous defect in astrocytes, as well as the disorganized fibronectin expression pattern correspondingly in migration front. Furthermore, the growth of transplanted LLC tumor was inhibited in Amot knockout mice due to fewer vasculature involved. By using MMTV-PyMT transgenic mouse model, there was a significantly longer period before tumors arised when Amot was specifically knocked out in blood vessels. In vitro evidence showed that Amot binded to beta-actin, Integrin beta 1 (ITGB1), Fibronectin, FAK, Vinculin, major focal adhesion molecules, and ITGB1 and stress fibers were distinctly induced by Amot transfection. Via traction force microscopy, the total energy (force indicater) was found significantly decreased in Amot knockdown cells. Taken together, we propose that Amot is a novel partner of the ITGB1/Fibronectin protein complex at focal adhesion and required for exerting force transition between endothelial cell and extracellular matrix.

Keywords: angiogenesis, angiomotin, endothelial cell migration, focal adhesion, integrin beta 1

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Authors: F. Bouchouka, S. Benamara

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The present work is an attempt to prepare a fresh cheese using lemon juice and lemon juice / orange juice mixture as acidifying / clotting agents. A reference cheese is obtained by acidification with commercial vinegar. The analysis performed on the final product (fat, cheese yield, sensory analysis, rheological and bacteriological properties) confirmed the technical feasibility of a natural cheese, using a lemon juice and / or lemon juice / orange juice mixture as acidifying / clotting agents. In addition, a general acceptance test allowed to select the cheese sample acidified with lemon juice as the best, compared to the two other samples (lemon juice/orange juice acidification and commercial vinegar acidification).

Keywords: clotting agent, fresh cheese, juice, lemon, orange

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Authors: Jacqueline T. Madaure, Phatu W. Mashela

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Cucurbitacin-containing phytonematicides consistently suppress root-knot (Meloidogyne species) nematode population densities. However, the impact of these products on nematode biocontrol agents is not documented. The objective of this study was to determine the influence of Nemarioc-AL and Nemafric-BL phytonematicides on growth of Trichoderma harzianum under in vitro conditions. The two phytonematicides were separately prepared to concentrations of 3% and used in poison plate assays. After exposure at different times from 0 to 72 h, there was 100% mycelial growth of T. harzianum. In conclusion, at the recommended concentrations of phytonematicides used in managing nematode population densities, there was no evidence of suppressive effects on growth of T. harzianum by the two phytonematicides.

Keywords: botanicals, crude extracts, cucumis africanus, cucumis myriocarpus, cucurbitacin a, cucurbitacin b, ethnomedicinal plants

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Authors: Tais Basaco, Stefanie Pektor, Josue A. Moreno, Matthias Miederer, Andreas Türler

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Radiopharmaceuticals based in monoclonal anti-body (mAb) via chemical linkers have become a potential tool in nuclear medicine because of their specificity and the large variability and availability of therapeutic radiometals. It is important to identify the conjugation sites and number of attached chelator to mAb to obtain radioimmunoconjugates with required immunoreactivity and radiostability. Girentuximab antibody (G250) is a potential candidate for radioimmunotherapy of clear cell carcinomas (RCCs) because it is reactive with CAIX antigen, a transmembrane glycoprotein overexpressed on the cell surface of most ( > 90%) (RCCs). G250 was conjugated with the bifunctional chelating agent DOTA (1,4,7,10-Tetraazacyclododecane-N,N’,N’’,N’’’-tetraacetic acid) via a benzyl-thiocyano group as a linker (p-SCN-Bn-DOTA). DOTA-G250 conjugates were analyzed by size exclusion chromatography (SE-HPLC) and by electrophoresis (SDS-PAGE). The potential site-specific conjugation was identified by liquid chromatography–mass spectrometry (LC/MS-MS) and the number of linkers per molecule of mAb was calculated using the molecular weight (MW) measured by matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). The average number obtained in the conjugates in non-reduced conditions was between 8-10 molecules of DOTA per molecule of mAb. The average number obtained in the conjugates in reduced conditions was between 1-2 and 3-4 molecules of DOTA per molecule of mAb in the light chain (LC) and heavy chain (HC) respectively. Potential DOTA modification sites of the chelator were identified in lysine residues. The biological activity of the conjugates was evaluated by flow cytometry (FACS) using CAIX negative (SKRC-18) and CAIX positive (SKRC-52). The DOTA-G250 conjugates were labelled with 177Lu with a radiochemical yield > 95% reaching specific activities of 12 MBq/µg. The stability in vitro of different types of radioconstructs was analyzed in human serum albumin (HSA). The radiostability of 177Lu-DOTA-G250 at high specific activity was increased by addition of sodium ascorbate after the labelling. The immunoreactivity was evaluated in vitro and in vivo. Binding to CAIX positive cells (SK-RC-52) at different specific activities was higher for conjugates with less DOTA content. Protein dose was optimized in mice with subcutaneously growing SK-RC-52 tumors using different amounts of 177Lu- DOTA-G250.

Keywords: mass spectrometry, monoclonal antibody, radiopharmaceuticals, radioimmunotheray, renal cancer

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Authors: B. T. Naveen Kumar, Anuj Tyagi, Niraj Kumar Singh, Visanu Boonyawiwat, A. H. Shanthanagouda, Orawan Boodde, K. M. Shankar, Prakash Patil, Shubhkaramjeet Kaur

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Early mortality syndrome (EMS)/Acute Hepatopancreatic Necrosis Disease (AHPND) has emerged as a major obstacle for the shrimp farming around the world. It is caused by a strain of Vibrio parahaemolyticus. The possible preventive and control measure is, early and rapid detection of the pathogen in the broodstock, post-larvae and monitoring the shrimp during the culture period. Polymerase chain reaction (PCR) based early detection methods are good, but they are costly, time taking and requires a sophisticated laboratory. The present study was conducted to develop a simple, sensitive and rapid diagnostic farmer level kit for the reliable detection of AHPND in shrimp. A panel of monoclonal antibodies (MAbs) were raised against the recombinant Pir B protein (rPirB). First, an immunodot was developed by using MAbs G3B8 and Mab G3H2 which showed specific reactivity to purified r-PirB protein with no cross-reactivity to other shrimp bacterial pathogens (AHPND free Vibrio parahaemolyticus (Indian strains), V. anguillarum, WSSV, Aeromonas hydrophila, and Aphanomyces invadans). Immunodot developed using Mab G3B8 is more sensitive than that with the Mab G3H2. However, immunodot takes almost 2.5 hours to complete with several hands-on steps. Therefore, the flow-through assay (FTA) was developed by using a plastic cassette containing the nitrocellulose membrane with absorbing pads below. The sample was dotted in the test zone on the nitrocellulose membrane followed by continuos addition of five solutions in the order of i) blocking buffer (BSA) ii) primary antibody (MAb) iii) washing Solution iv) secondary antibody and v) chromogen substrate (TMB) clear purple dots against a white background were considered as positive reactions. The FTA developed using MAbG3B8 is more sensitive than that with MAb G3H2. In FTA the two MAbs showed specific reactivity to purified r-PirB protein and not to other shrimp bacterial pathogens. The FTA is simple to farmer/field level, sensitive and rapid requiring only 8-10 min for completion. Tests can be developed to kits, which will be ideal for use in biosecurity, for the first line of screening (at the port or pond site) and during monitoring and surveillance programmes overall for the good management practices to reduce the risk of the disease.

Keywords: acute hepatopancreatic necrosis disease, AHPND, flow-through assay, FTA, farmer level, immunodot, pond site, shrimp

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Authors: Özlem Arpacı, Zeliha Soysal, Nuran Diril

Abstract:

Benzoxazoles are heterocyclic compounds that have a fused benzene and an oxazole ring. These heterocyclic compounds are reported to have antibacterial, antitubercular, antifungal, antiviral, antioxidant and anticancer activities. In this study, some benzoxazole derivatives that have broad antimicrobial and potent antitumoral activities, are investigated their mutagenic activities with using the Ames Test. The Ames test was conducted using Salmonella typhimurium TA98, TA100 and TA102 tester strains in the standard plate incorporation assay in the absence of liver S9 fraction. The results are evaluated using SPSS and none of the benzoxazole derivatives showed mutagenic activity using the Ames test in the absence of S9 liver fraction.

Keywords: benzoxazoles, ames test, mutagenic activity, antimutagenic activity, antitumoral activity

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Authors: Görkem Cemali̇, Avram Aruh, Gamze Torun Köse, Erde Can ŞAfak

Abstract:

In order to heal bone disorders, the conventional methods which involve the use of autologous and allogenous bone grafts or permanent implants have certain disadvantages such as limited supply, disease transmission, or adverse immune response. A biodegradable material that acts as structural support to the damaged bone area and serves as a scaffold that enhances bone regeneration and guides bone formation is one desirable solution. Poly(propylene fumarate) (PPF) which is an unsaturated polyester that can be copolymerized with appropriate vinyl monomers to give biodegradable network structures, is a promising candidate polymer to prepare bone tissue engineering scaffolds. In this study, hydroxyl-terminated PPF was synthesized and thermally cured with vinyl phosphonic acid (VPA) and diethyl vinyl phosphonate (VPES) in the presence of radical initiator benzoyl peroxide (BP), with changing co-monomer weight ratios (10-40wt%). In addition, the synthesized PPF was cured with VPES comonomer at body temperature (37oC) in the presence of BP initiator, N, N-Dimethyl para-toluidine catalyst and varying amounts of Beta-tricalcium phosphate (0-20 wt% ß-TCP) as filler via radical polymerization to prepare composite materials that can be used in injectable forms. Thermomechanical properties, compressive properties, hydrophilicity and biodegradability of the PPF/VPA and PPF/VPES copolymers were determined and analyzed with respect to the copolymer composition. Biocompatibility of the resulting polymers and their composites was determined by the MTS assay and osteoblast activity was explored with von kossa, alkaline phosphatase and osteocalcin activity analysis and the effects of VPA and VPES comonomer composition on these properties were investigated. Thermally cured PPF/VPA and PPF/VPES copolymers with different compositions exhibited compressive modulus and strength values in the wide range of 10–836 MPa and 14–119 MPa, respectively. MTS assay studies showed that the majority of the tested compositions were biocompatible and the overall results indicated that PPF/VPA and PPF/VPES network polymers show significant potential for applications as bone tissue engineering scaffolds where varying PPF and co-monomer ratio provides adjustable and controllable properties of the end product. The body temperature cured PPF/VPES/ß-TCP composites exhibited significantly lower compressive modulus and strength values than the thermal cured PPF/VPES copolymers and were therefore found to be useful as scaffolds for cartilage tissue engineering applications.

Keywords: biodegradable, bone tissue, copolymer, poly(propylene fumarate), scaffold

Procedia PDF Downloads 161

Authors: Rachid Kacem, Sara Talbi, Yasmina Hemissi, Sofia Bouguattoucha

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The aim of the present work is the evaluation of the antioxidant activity of the Peganum harmala (P. harmala) seeds extracts. The antioxidant activity was evaluated by applying two methods, the method of ß-carotene bleaching and DPPH (2, 2-Diphenyl-1-Picryl-Hydrazyl). Using Folin-Ciocalteu assay, these results revealed that the concentration of polyphenols in EthOH E. (122.28 ± 2.24 µg GAE/mg extract) is the highest. The antiradical activity of the P. harmala seeds extracts on DPPH was found to be dose dependent with polyphenols concentration. The E. EthOH extract showed the highest antioxidant activity (IC = 252.10 ± 11.18 μg /ml). The test of β-carotene bleaching indicates that the E. EthOH of P. harmala showed the highest percentage of the antioxidant activity (49.88 %).

Keywords: antioxidant activity, Peganum harmala, polyphenols, flavonoids

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Authors: Abeer Y. Ibrahim, Faten M. Ibrahim, Samah A. El-Newary, Saber F. Hendawy

Abstract:

Objective: Appreciation of in-vitro anti-inflammatory and antioxidant characters of Vitex agnus-castus berries alcoholic extract and fractions, as well as in-vivo antitumor ability of alcoholic extract and chloroform fraction against Ehrlich ascites carcinoma is the aim of this study. Material and methods: Antioxidant properties of crude alcoholic extract of vitex berries as well as petroleum ether, chloroform, ethyl acetate and butanol fractions were evaluated, in-vitro assessments, as compared with standard materials, l-ascorbic acid (vitamin C) and butylated hydroxyl toluene(BHT). The anti-inflammatory activity was investigated in cyclooxygenase (COX)-1 and COX-2 inhibition assays. Moreover, in-vivo antitumor effect of vitex berries alcoholic and chloroform extracts were evaluated using Ehrlich ascites carcinoma model. Data were presented as mean±SE, and data were analyzed by one-way analysis of variance test. Results and conclusion: Berries crude extract showed potent antioxidant activity followed with its fractions ethyl acetate and chloroform as compared with standard (V.C and BHT). Ethyl acetate fraction showed good reduction capability, metal ion chelation, hydrogen peroxide scavenging, nitric oxide scavenging and superoxide anion scavenging. Meanwhile, chloroform fraction produced the highest free radical scavenging activity and total antioxidant capacity. In respectable of lipid peroxidation inhibition, crude alcoholic extract and its fractions cleared weak inhibition in comparing with standard materials. Anti-inflammatory activity of V. agnus-castus berries chloroform fraction of vitex was best COX-2 inhibitor (IC₅₀, 135.41 µg/ ml) as compared to vitex alcoholic extract or ethyl acetate fraction with weak inhibitory effect on COX-1 (IC50, 778.432 µg/ ml), where the lowest effect on COX-1 was recorded with alcoholic extract. Alcoholic extract and its fractions showed weak COX-1 inhibition activity, whereas COX-2 was inhibited (100%), compared with celecoxib drug (72% at 1000ppm). The crude alcoholic and chloroform extracts of V. agnus-castus barries significantly reduced the viable Ehrlich cell count and increased nonviable count with amelioration of all hematological parameters. This amelioration was reflected on increasing median survival time and significant increase (P < 0.05) in lifespan.

Keywords: anti-inflammatory, antioxidants, ehrlich ascites carcinoma, Vitex agnus-castus

Procedia PDF Downloads 134

Authors: Hairin Taha, P. Narrima, M. A. Hapipah, A. M. Mustafa

Abstract:

The chemical constituents and antioxidant activity of the leaf essential oils of Pseuduvaria monticola and Pseuduvaria macrophylla from the Annonaceae family were investigated. GC-TOFMS analyses identified 46 compounds from Pseuduvaria monticola and 11 compounds from Pseuduvaria macrophylla. The major constituents in the leaf essential oil of Pseuduvaria monticola were a-cadinol (13.0%), calamenene-cis (6.9%), alfa copaene (4%), and epizonarene (3.8%), while in the leaf essential oil of Pseuduvaria macrophylla were caryophyllene oxide (29.7%) and elimicin (28%). The antioxidant activity of both the essential oils were determined using the 2,2'-diphenyl-1-picrylhydrazyl assay (DPPH). The present study suggests that both essential oils demonstrated good antioxidant activity.

Keywords: Pseuduvaria monticola, Pseuduvaria macrophylla, leaf essential oils, GC-MSTOF, antioxidant activity

Procedia PDF Downloads 365

Authors: Santi Maneewatchararangsri, Wanpen Chaicumpa, Patcharin Saengjaruk, Urai Chaisri

Abstract:

Leptospirosis is a globally neglected disease that continues to be a significant public health and veterinary burden, with millions of cases reported each year. Early and accurate differential diagnosis of leptospirosis from other febrile illnesses and the development of a broad spectrum of leptospirosis vaccines are needed. The LipL32 outer membrane lipoprotein is a member of Leptospira adhesive matrices and has been found to exert hemolytic activity to erythrocytes in vitro. Therefore, LipL32 is regarded as a potential target for diagnosis, broad-spectrum leptospirosis vaccines, and for passive immunotherapy. In this study, we established LipL32-specific mouse monoclonal antibodies, mAbLPF1 and mAbLPF2, and their respective mouse- and humanized-engineered single chain variable fragment (ScFv). Their antibodies’ neutralizing activities against Leptospira-mediated hemolysis in vitro, and the therapeutic efficacy of mAbs against heterologous Leptospira infected hamsters were demonstrated. The epitope peptide of mAb LPF1 was mapped to a non-contiguous carboxy-terminal β-turn and amphipathic α-helix of LipL32 structure contributing to phospholipid/host cell adhesion and membrane insertion. We found that the mAbLPF2 epitope was located on the interacting loop of peptide binding groove of the LipL32 molecule responsible for interactions with host constituents. Epitope sequences are highly conserved among Leptospira spp. and are absent from the LipL32 superfamily of other microorganisms. Both epitopes are surface-exposed, readily accessible by mAbs, and immunogenic. However, they are less dominant when revealed by LipL32-specific immunoglobulins from leptospirosis-patient sera and rabbit hyperimmune serum raised by whole Leptospira. Our study also demonstrated an adhesion inhibitory activity of LipL32 protein to host membrane components and cells mediated by mAbs as well as an anti-hemolytic activity of the respective antibodies. The therapeutic antibodies, particularly the humanized-ScFv, have a potential for further development as non-drug therapeutic agent for human leptospirosis, especially in subjects allergic to antibiotics. The epitope peptides recognized by two therapeutic mAbs have potential use as tools for structure-function studies. Finally, protective peptides may be used as a target for epitope-based vaccines for control of leptospirosis.

Keywords: leptospira lipl32-specific antibodies, therapeutic epitopes, epitopes characterization, immunotherapy

Procedia PDF Downloads 292

Authors: José Silvestre Mendoza Figueroa, Anders Kvarnheden, Jesús Méndez Lozano, Edgar Antonio Rodríguez Negrete, Manuel Soriano García

Abstract:

Agroindustrial plants such as cereals and pseudo cereals offer a substantial source of biomacromolecules, as they contain large amounts per tissue-gram of proteins, polysaccharides and lipids in comparison with other plants. In particular, Amaranthus hypochondriacus seeds have high levels of proteins in comparison with other cereal and pseudo cereal species, which makes the plant a good source of bioactive molecules such as peptides. Geminiviruses are one principal class of pathogens that causes important economic losses in crops, affecting directly the development and production of the plant. One such virus is the Tomato yellow leaf curl virus (TYLCV), which affects mainly Solanacea family plants such as tomato species. The symptoms of the disease are curling of leaves, chlorosis, dwarfing and floral abortion. The aim of this work was to get peptides derived from enzymatic hydrolysis of globulins and albumins from amaranth seeds with specific recognition of the replication origin in the TYLCV genome, and to test the antiviral activity on host plants with the idea to generate a direct control of this viral infection. Globulins and albumins from amaranth were extracted, the fraction was enzymatically digested with papain, and the aromatic peptides fraction was selected for further purification. Six peptides were tested against the replication origin (OR) using affinity assays, surface resonance plasmon and fluorescent titration, and two of these peptides showed high affinity values to the replication origin of the virus, dissociation constant values were calculated and showed specific interaction between the peptide Ampep1 and the OR. An in vitro replication test of the total TYLCV DNA was performed, in which the peptide AmPep1 was added in different concentrations to the system reaction, which resulted in a decrease of viral DNA synthesis when the peptide concentration increased. Also, we showed that the peptide can decrease the complementary DNA chain of the virus in Nicotiana benthamiana leaves, confirming that the peptide binds to the OR and that its expected mechanism of action is to decrease the replication rate of the viral genome. In an infection assay, N. benthamiana plants were agroinfected with TYLCV-Israel and TYLCV-Guasave. After confirming systemic infection, the peptide was infiltrated in new infected leaves, and the plants treated with the peptide showed a decrease of virus symptoms and viral titer. In order to confirm the antiviral activity in a commercial crop, tomato plants were infected with TYLCV. After confirming systemic infection, plants were infiltrated with peptide solution as above, and the symptom development was monitored 21 days after treatment, showing that tomato plants treated with peptides had lower symptom rates and viral titer. The peptide was also tested against other begomovirus such as Pepper huasteco yellow vein virus (PHYVV-Guasave), showing a decrease of symptoms in N. benthamiana infected plants. The model of direct biochemical control of TYLCV infection shown in this work can be extrapolated to other begomovirus infections, and the methods reported here can be used for design of antiviral agrochemicals for other plant virus infections.

Keywords: agrochemical screening, antiviral, begomovirus, geminivirus, peptides, plasmon, TYLCV

Procedia PDF Downloads 262

Authors: Patrícia Severino, Luciana Nalone, Daniele Martins, Marco Chaud, Classius Ferreira, Cristiane Bani, Ricardo Albuquerque

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Hydrogels produced with polymers have been used in the development of dressings for wound treatment and tissue revitalization. Our study on polymer nanocomposites containing silver nanoparticles shows antimicrobial activity and applications in wound healing. The effects are linked with the slow oxidation and Ag⁺ liberation to the biological environment. Furthermore, bacterial cell membrane penetration and metabolic disruption through cell cycle disarrangement also contribute to microbial cell death. The silver antimicrobial activity has been known for many years, and previous reports show that low silver concentrations are safe for human use. This work aims to develop a hydrogel using natural polymers (sodium alginate and gelatin) combined with silver nanoparticles for wound healing and with antimicrobial properties in cutaneous lesions. The hydrogel development utilized different sodium alginate and gelatin proportions (20:80, 50:50 and 80:20). The silver nanoparticles incorporation was evaluated at the concentrations of 1.0, 2.0 and 4.0 mM. The physico-chemical properties of the formulation were evaluated using ultraviolet-visible (UV-Vis) absorption spectroscopy, Fourier transform infrared (FTIR) spectroscopy, differential scanning calorimetry (DSC), and thermogravimetric (TG) analysis. The morphological characterization was made using transmission electron microscopy (TEM). Human fibroblast (L2929) viability assay was performed with a minimum inhibitory concentration (MIC) assessment as well as an in vivo cicatrizant test. The results suggested that sodium alginate and gelatin in the (80:20) proportion with 4 mM of AgNO₃ in the (UV-Vis) exhibited a better hydrogel formulation. The nanoparticle absorption spectra of this analysis showed a maximum band around 430 - 450 nm, which suggests a spheroidal form. The TG curve exhibited two weight loss events. DSC indicated one endothermic peak at 230-250 °C, due to sample fusion. The polymers acted as stabilizers of a nanoparticle, defining their size and shape. Human fibroblast viability assay L929 gave 105 % cell viability with a negative control, while gelatin presented 96% viability, alginate: gelatin (80:20) 96.66 %, and alginate 100.33 % viability. The sodium alginate:gelatin (80:20) exhibited significant antimicrobial activity, with minimal bacterial growth at a ratio of 1.06 mg.mL⁻¹ in Pseudomonas aeruginosa and 0.53 mg.mL⁻¹ in Staphylococcus aureus. The in vivo results showed a significant reduction in wound surface area. On the seventh day, the hydrogel-nanoparticle formulation reduced the total area of injury by 81.14 %, while control reached a 45.66 % reduction. The results suggest that silver-hydrogel nanoformulation exhibits potential for wound dressing therapeutics.

Keywords: nanocomposite, wound healing, hydrogel, silver nanoparticle

Procedia PDF Downloads 94

Authors: Kusmartono, Siti Chuzaemi, Hartutik dan Mashudi

Abstract:

Two studies were conducted involving an in vitro (Expt 1) and in vivo (Expt 2) measurements. Expt 1. aimed to evaluate effects of adding CT extracts on gas production and efficiency of microbial protein synthesis (EMPS), Expt 2 aimed to evaluate effects of supplementing shrub/tree leaves as CT source on feed consumption, digestibility, N retention, body weight gain and dressing percentage of growing sheep fed on elephant grass (EG) as a basal diet.Ten shrub and tree leaves used as CT sources were wild sunflower (Tithonia diversifolia), mulberry (Morus macroura), cassava (Manihot utilissima), avicienna (Avicennia marina), calliandra (Calliandra calothyrsus), sesbania (Sesbania grandiflora), acacia (acacia vilosa), glyricidia (Glyricidia sepium), jackfruit (Artocarpus heterophyllus), moringa (Moringa oleifera). The treatments applied in Expt 1 were: T1=Elephant grass (60%)+concentrate (40%); T2 = T1 + CT (3% DM); T3= T2 + PEG; T4 = T1 + CT (3.5% DM); T5 = T4 + PEG; T6 = T1 + CT (4% DM) and T7 = T6 + PEG. Data obtained were analysed using Randomized Block Design. Statistical analyses showed that treatments significanty affected (P<0.05) total gas production and EMPS. The lowest values of total gas production (45.9 ml/500 mg DM) and highest value of EMPS (64.6 g/kg BOTR) were observed in the treatment T4 (3.5% CT from cassava leave extract). Based on this result it was concluded that this treatment was the best and was chosen for further investigation using in vivo method. The treatmets applied for in vivo trial were: T1 = EG (60%) + concentrate (40%); T2 = T1 + dried cassava leave (equivalent to 3.5% CT); T3 = T2 + PEG. 18 growing sheep aging of 8-9 months and weighing of 23.67kg ± 1.23 were used in Expt 2. Results of in vivo study showed that treatments significanty affected (P<0.05) nutrients intake and digestibility (DM, OM and CP). N retention for sheep receiving treatment T2 were significantly higher (P<0.05; 15.6 g/d) than T1 (9.1 g/d) and T3 (8.53 g/d). Similar results were obtained for daily weight gain where T2 were the highest (62.79 g/d), followed by T1 (51.9 g/d) and T3 (52.85 g/d). Dressing percentage of T2 was the highest (51.54%) followed by T1 (49.61%) and T3 (49.32%). It can be concluded that adding adding dried cassava leaves did not reduce palatability due to CT, but rather increased OM digestibility and hence feed consumption was improved. N retention was increased due to the action of CT in the cassava leaves and this may have explained a higher input of N into duodenum which was further led to higer daily weight gain and dressing percentage.

Keywords: in vitro gas production, sheep, shrub and tree leaves, condensed tannin

Procedia PDF Downloads 254

Authors: Barbra Bhebhe, Melvyn Quan

Abstract:

A serological survey was done to detect antibodies against Shuni virus (SHUV) from cattle in Western Kenya. In Kenya the disease status of SHUV in cattle has never been established. It is a zoonotic virus and even though studies have been carried out as early as the 1960s, little research has been published and SHUV is still not a well-recognised Orthobunyavirus. One hundred serum samples were collected from healthy cattle in Kenya and tested for antibodies against SHUV by a serum neutralization assay. All antibody titre values were greater than 1:160, with most of the samples greater than 1:320. Of the samples tested, 87 % had titres greater than 1:320, 12% had a titre of 1:320 and 2% had a titre of 1:160. Samples were classified as positive if the antibody titre was ≥ 1:10 and negative if < 1:10. This study suggests that cattle are exposed commonly to SHUV, which may be endemic in Kenya.

Keywords: Shuni virus, Orthobunyavuruses, serum neutralization test, cell-culture

Procedia PDF Downloads 62