Search results for: spectrophotometer assay

Authors: Paul Awolade, Parvesh Singh

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The global threat of pathogenic resistance to available therapeutic agents has become a menace to clinical practice, public health and man’s existence inconsequential. This has therefore led to an exigency in the development of new molecular scaffolds with profound activity profiles. In this vein, a versatile synthetic tool for accessing new molecules by incorporating two or more pharmacophores into a single entity with the unique ability to be recognized by multiple receptors hence leading to an improved bioactivity, known as molecular hybridization, has been explored with tremendous success. Accordingly, aware of the similarity in pharmacological activity spectrum of quinoline and 1,2,3-triazole pharmacophores such as; anti-Alzheimer, anticancer, anti-HIV, antimalarial and antimicrobial to mention but a few, the present study sets out to synthesize hybrids of quinoline and 1,2,3-triazole. The hybrids were accessed via click chemistry using copper catalysed azide-alkyne 1,3-dipolar cycloaddition reaction. All synthesized compounds were evaluated for their pharmaco-potential in an antimicrobial assay out of which the 3-OH derivative emerged as the most active with MIC value of 4 μg/mL against Cryptococcus neoformans; a value superior to standard Fluconazole and comparable to Amphotericin B. Structures of synthesized hybrids were elucidated using appropriate spectroscopic techniques (1H, 13C and 2D NMR, FT-IR and HRMS).

Keywords: bioisostere, click chemistry, molecular hybridization, quinoline, 1, 2, 3-triazole

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Authors: M. N. Hanina, M. Hairul Shahril, I. Ismatul Nurul Asyikin, A. K. Abdul Jalil, M. R. Salina, M. R. Maryam, M. Rosfarizan

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The extracellular proteins secreted by bacteria may be increased in stressful surroundings, such as in the presence of antibiotics. It appears that many antibiotics, when used at low concentrations, have in common the ability to activate or repress gene transcription, which is distinct from their inhibitory effect. There have been comparatively few studies on the potential of antibiotics as a specific chemical signal that can trigger a variety of biological functions. Therefore, this study was carried out to determine the effect of Streptomycin Sulfate in regulating extracellular proteins secreted by Bacillus subtilis ATCC21332. Results of Microdilution assay showed that the Minimum Inhibition Concentration (MIC) of Streptomycin Sulfate on B. subtilis ATCC21332 was 2.5 mg/ml. The bacteria cells were then exposed to Streptomycin Sulfate at concentration of 0.01 MIC before being further incubated for 48h to 72 h. The extracellular proteins secreted were then isolated and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Proteins profile revealed that three additional bands with approximate sizes of 30 kDa, 22 kDa and 23 kDa were appeared for the treated bacteria with Streptomycin Sulfate. Thus, B. subtilis ATCC21332 in stressful condition with the presence of Streptomycin Sulfate at low concentration could induce the extracellular proteins secretion.

Keywords: Bacillus subtilis ATCC21332, streptomycin sulfate, extracellular proteins, antibiotics

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Authors: Srinivasarao Kondaparla, Utsab Debnath, Awakash Soni, Vasantha Rao Dola, Manish Sinha, Kumkum Kumkum Srivastava, Sunil K. Puri, Seturam B. Katti

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In a focused exploration, we have designed synthesized and biologically evaluated chiral conjugated new chloroquine (CQ) analogs with substituted piperazines as antimalarial agents. In vitro as well as in vivo studies revealed that compound 7c showed potent activity [for in vitro IC₅₀= 56.98nM (3D7), 97.76nM (K1); for in vivo (up to at the dose of 12.5 mg/kg); SI = 3510] as a new lead of antimalarial agent. Other compounds 6b, 6d, 7d, 7h, 8c, 8d, 9a, and 9c are also showing moderate activity against CQ-sensitive (3D7) strain and superior activity against resistant (K1) strain of P. falciparum. Furthermore, we have carried out docking and 3D-QSAR studies of all in-house data sets (168 molecules) of chiral CQ analogs to explain the structure activity relationships (SAR). Our new findings specified the significance of H-bond interaction with the side chain of heme for biological activity. In addition, the 3D-QSAR study against 3D7 strain indicated the favorable and unfavorable sites of CQ analogs for incorporating steric, hydrophobic and electropositive groups to improve the antimalarial activity.

Keywords: piperazines, CQ-sensitive strain-3D7, in-vitro and in-vivo assay, docking, 3D-QSAR

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Authors: Nur Fathin Alia Che Wahab, Thirumulu Ponnuraj Kannan, Zuliani Mahmood, Ismail Ab. Rahman, Hanafi Ismail

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Locally produced Biphasic Calcium Phosphate (BCP) consists of hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP) which is a promising material for dentin and bone regeneration as well as in tissue engineering applications. The study was carried out to investigate the mutagenic effect of locally produced BCP using Ames test. Mutagenicity was evaluated with and without the addition of metabolic activation system (S9). This study was performed on Salmonella typhimurium TA98, TA102, TA1537, and TA1538 strains using preincubation assay method. The doses tested were 5000, 2500, 1250, 625, 313 µg/plate. Negative and positive controls were also included. The bacteria were incubated for 48 hours at 37 ± 0.5 °C. Then, the revertant colonies were counted. Data obtained were evaluated using non-statistical method. The mean number of revertant colonies in strains with and without S9 mix treated with locally produced BCP was less than double when compared to negative control for all the tested concentrations. The results from this study indicate that the locally produced BCP is non-mutagenic under the present test conditions.

Keywords: ames test, biphasic calcium phosphate, dentin regeneration, mutagenicity

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Authors: N. A. Kazemi Sefat, M. M. Mohammadi, J. Hadjati, S. Talebi, M. Ajami, H. Daneshvar

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Colorectal cancer metastases result in a significant number of cancer related deaths. Histone deacetylase (HDAC) inhibitors induce growth arrest and apoptosis in a variety of human cancer cells. Sodium butyrate (SB) is a short chain fatty acid, belongs to HDAC inhibitors which is released in the colonic lumen as a consequence of fiber fermentation. In this study, we are about to assess the effect of sodium butyrate on HCT116 human colorectal carcinoma cell line. The viability of cells was measured by microscopic morphologic study and MTT assay. After 48 hours, treatments more than 10 mM lead to cell injury in HCT116 by increasing cell granulation and decreasing cell adhesion (p>0.05). After 72 hours, treatments at 10 mM and more lead to significant cell injury (p<0.05). Our results may suggest that the gene expression which is contributed in cell proliferation and apoptosis has been changed under pressure of HDAC inhibition.

Keywords: colorectal cancer, sodium butyrate, cytotoxicity, MTT

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Authors: Melasurej C. Francisco, Sophia D. Rusit

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Old age is a record of one’s own life; this is the crucial phase for most. However, there are individuals who believe that old people retain self-efficacy and self-worth throughout their existence. Geriatric institutions focus on the health of elderly, in which they have been supported with medicines and therapies by clinician thus, indicating that these may suffice physical, emotional, and mental health of the elderly. This study focuses on (1) Describing the level or degree of self-efficacy; (2) Recognizing the extent of self-worth; (3) Determining the significant relationship between self-efficacy and self-worth. It is a mixed method design. A combination of correlational research and in-depth interview. Purposive sampling technique was used to select participants, considering that this assay focused on elderly in geriatric institutions, it follows that respondents and participants are at least sixty years of age and must be living inside the institution. 121 senior citizens took part in this study. Scores from both General Self-Efficacy Scale (GSE) and Rosenberg Self-Esteem Scale (RSES) showed varying levels of self-efficacy and self-worth. SE had μ=28.099, σ=6.6262, σ²=43.9067 while; SW had μ=14.9669, σ=5.3789, σ²28.9322 which denotes that rₒbₜ (121)=0.3164 is higher than rcᵢₜ which is 0.150. Although this exhibits the positive moderate correlation between SE and SW, the relationship between variables is weak. Likewise, the pᵥₐₗᵤₑ (pᵥₐₗᵤₑ=0.000406) is lower than the significance level alpha=0.01, thus, rejecting the null hypothesis, and accepting the alternative hypothesis.

Keywords: elderly, geriatric, self-efficacy, self-worth

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Authors: Seyedeh Nasim Mirbahari, Taha Azad, Mehdi Totonchi

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Oncolytic viruses, which only replicate in tumor cells, are being extensively studied for their use in cancer therapy. A particular virus known as the vaccinia virus, a member of the poxvirus family, has demonstrated oncolytic abilities glioma. Treating Glioma with traditional methods such as chemotherapy and radiotherapy is quite challenging. Even though oncolytic viruses have shown immense potential in cancer treatment, their effectiveness in glioblastoma treatment is still low. Therefore, there is a need to improve and optimize immunotherapies for better results. In this study, we have designed oncoVV-TT, which can more effectively target tumor cells while minimizing replication in normal cells by replacing the thymidine kinase gene with a luc-p2a-GFP gene expression cassette. Human glioblastoma cell line U251 MG, rat glioblastoma cell line C6, and non-tumor cell line HFF were plated at 105 cells in a 12-well plates in 2 mL of DMEM-F2 medium with 10% FBS added to each well. Then incubated at 37°C. After 16 hours, the cells were treated with oncoVV-TT at an MOI of 0.01, 0.1 and left in the incubator for a further 24, 48, 72 and 96 hours. Viral replication assay, fluorescence imaging and viability tests, including trypan blue and crystal violet, were conducted to evaluate the cytotoxic effect of oncoVV-TT. The finding shows that oncoVV-TT had significantly higher cytotoxic activity and proliferation rates in tumor cells in a dose and time-dependent manner, with the strongest effect observed in U251 MG. To conclude, oncoVV-TT has the potential to be a promising oncolytic virus for cancer treatment, with a more cytotoxic effect in human glioblastoma cells versus rat glioma cells. To assess the effectiveness of vaccinia virus-mediated viral therapy, we have tested U251mg and C6 tumor cell lines taken from human and rat gliomas, respectively. The study evaluated oncoVV-TT's ability to replicate and lyse cells and analyzed the survival rates of the tested cell lines when treated with different doses of oncoVV-TT. Additionally, we compared the sensitivity of human and mouse glioma cell lines to the oncolytic vaccinia virus. All experiments regarding viruses were conducted under biosafety level 2. We engineered a Vaccinia-based oncolytic virus called oncoVV-TT to replicate specifically in tumor cells. To propagate the oncoVV-TT virus, HeLa cells (5 × 104/well) were plated in 24-well plates and incubated overnight to attach to the bottom of the wells. Subsequently, 10 MOI virus was added. After 48 h, cells were harvested by scraping, and viruses were collected by 3 sequential freezing and thawing cycles followed by removal of cell debris by centrifugation (1500 rpm, 5 min). The supernatant was stored at −80 ◦C for the following experiments. To measure the replication of the virus in Hela, cells (5 × 104/well) were plated in 24-well plates and incubated overnight to attach to the bottom of the wells. Subsequently, 5 MOI virus or equal dilution of PBS was added. At the treatment time of 0 h, 24 h, 48 h, 72 h and 96 h, the viral titers were determined under the fluorescence microscope (BZ-X700; Keyence, Osaka, Japan). Fluorescence intensity was quantified using the imagej software according to the manufacturer’s protocol. For the isolation of single-virus clones, HeLa cells seeded in six-well plates (5×105 cells/well). After 24 h (100% confluent), the cells were infected with a 10-fold dilution series of TianTan green fluorescent protein (GFP)virus and incubated for 4 h. To examine the cytotoxic effect of oncoVV-TT virus ofn U251mg and C6 cell, trypan blue and crystal violet assay was used.

Keywords: oncolytic virus, immune therapy, glioma, vaccinia virus

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Authors: Olga V. Chervyakova, Elmira T. Tailakova, Vitaliy M. Strochkov, Kulyaisan T. Sultankulova, Nurlan T. Sandybayev, Lev G. Nemchinov, Rosemarie W. Hammond

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Sheep pox is a highly contagious infection that OIE regards to be one of the most dangerous animal diseases. It causes enormous economic losses because of death and slaughter of infected animals, lower productivity, cost of veterinary and sanitary as well as quarantine measures. To control spread of sheep pox infection the attenuated vaccines are widely used in the Republic of Kazakhstan and other Former Soviet Union countries. In spite of high efficiency of live vaccines, the possible presence of the residual virulence, potential genetic instability restricts their use in disease-free areas that leads to necessity to exploit new approaches in vaccine development involving recombinant DNA technology. Vaccines on the basis of recombinant proteins are the newest generation of prophylactic preparations. The main advantage of these vaccines is their low reactogenicity and this fact makes them widely used in medical and veterinary practice for vaccination of humans and farm animals. The objective of the study is to produce recombinant immunogenic proteins for development of the high-performance means for sheep pox prophylaxis. The SPV proteins were chosen for their homology with the known immunogenic vaccinia virus proteins. Assay of nucleotide and amino acid sequences of the target SPV protein genes. It has been shown that four proteins SPPV060 (ortholog L1), SPPV074 (ortholog H3), SPPV122 (ortholog A33) and SPPV141 (ortholog B5) possess transmembrane domains at N- or C-terminus while in amino acid sequences of SPPV095 (ortholog А 4) and SPPV117 (ortholog А 27) proteins these domains were absent. On the basis of these findings the primers were constructed. Target genes were amplified and subsequently cloned into the expression vector рЕТ26b(+) or рЕТ28b(+). Six constructions (pSPPV060ΔТМ, pSPPV074ΔТМ, pSPPV095, pSPPV117, pSPPV122ΔТМ and pSPPV141ΔТМ) were obtained for expression of the SPV genes under control of T7 promoter in Escherichia coli. To purify and detect recombinant proteins the amino acid sequences were modified by adding six histidine molecules at C-terminus. Induction of gene expression by IPTG was resulted in production of the proteins with molecular weights corresponding to the estimated values for SPPV060, SPPV074, SPPV095, SPPV117, SPPV122 and SPPV141, i.e. 22, 30, 20, 19, 17 and 22 kDa respectively. Optimal protocol of expression for each gene that ensures high yield of the recombinant protein was identified. Assay of cellular lysates by western blotting confirmed expression of the target proteins. Recombinant proteins bind specifically with antibodies to polyhistidine. Moreover all produced proteins are specifically recognized by the serum from experimentally SPV-infected sheep. The recombinant proteins SPPV060, SPPV074, SPPV117, SPPV122 and SPPV141 were also shown to induce formation of antibodies with virus-neutralizing activity. The results of the research will help to develop a new-generation high-performance means for specific sheep pox prophylaxis that is one of key moments in animal health protection. The research was conducted under the International project ISTC # K-1704 “Development of methods to construct recombinant prophylactic means for sheep pox with use of transgenic plants” and under the Grant Project RK MES G.2015/0115RK01983 "Recombinant vaccine for sheep pox prophylaxis".

Keywords: prophylactic preparation, recombinant protein, sheep pox virus, subunit vaccine

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Authors: Aussara Panya, Nunghathai Sawasdee, Mutita Junking, Chatchawan Srisawat, Kiattawee Choowongkomon, Pa-Thai Yenchitsomanus

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Dengue virus (DENV) infection is a global public health problem with approximately 100 million infected cases a year. Presently, there is no approved vaccine or effective drug available; therefore, the development of anti-DENV drug is urgently needed. The clinical reports revealing the positive association between the disease severity and viral titer has been reported previously suggesting that the anti-DENV drug therapy can possibly ameliorate the disease severity. Although several anti-DENV agents showed inhibitory activities against DENV infection, to date none of them accomplishes clinical use in the patients. The surface envelope (E) protein of DENV is critical for the viral entry step, which includes attachment and membrane fusion; thus, the blocking of envelope protein is an attractive strategy for anti-DENV drug development. To search the safe anti-DENV agent, this study aimed to search for novel peptide inhibitors to counter DENV infection through the targeting of E protein using a structure-based in silico design. Two selected strategies has been used including to identify the peptide inhibitor which interfere the membrane fusion process whereby the hydrophobic pocket on the E protein was the target, the destabilization of virion structure organization through the disruption of the interaction between the envelope and membrane proteins, respectively. The molecular docking technique has been used in the first strategy to search for the peptide inhibitors that specifically bind to the hydrophobic pocket. The second strategy, the peptide inhibitor has been designed to mimic the ectodomain portion of membrane protein to disrupt the protein-protein interaction. The designed peptides were tested for the effects on cell viability to measure the toxic to peptide to the cells and their inhibitory assay to inhibit the DENV infection in Vero cells. Furthermore, their antiviral effects on viral replication, intracellular protein level and viral production have been observed by using the qPCR, cell-based flavivirus immunodetection and immunofluorescence assay. None of tested peptides showed the significant effect on cell viability. The small peptide inhibitors achieved from molecular docking, Glu-Phe (EF), effectively inhibited DENV infection in cell culture system. Its most potential effect was observed for DENV2 with a half maximal inhibition concentration (IC50) of 96 μM, but it partially inhibited other serotypes. Treatment of EF at 200 µM on infected cells also significantly reduced the viral genome and protein to 83.47% and 84.15%, respectively, corresponding to the reduction of infected cell numbers. An additional approach was carried out by using peptide mimicking membrane (M) protein, namely MLH40. Treatment of MLH40 caused the reduction of foci formation in four individual DENV serotype (DENV1-4) with IC50 of 24-31 μM. Further characterization suggested that the MLH40 specifically blocked viral attachment to host membrane, and treatment with 100 μM could diminish 80% of viral attachment. In summary, targeting the hydrophobic pocket and M-binding site on the E protein by using the peptide inhibitors could inhibit DENV infection. The results provide proof of-concept for the development of antiviral therapeutic peptide inhibitors to counter DENV infection through the use of a structure-based design targeting conserved viral protein.

Keywords: dengue virus, dengue virus infection, drug design, peptide inhibitor

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Authors: Patience Orobosa Olajide

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Five hydrocarbon degrading bacterial strains isolated from contaminated environment were investigated with respect to polyhydroxybutyrate (PHB) biosynthesis. Screening for bioplastic production was done on assay mineral salts agar medium containing 0.2% poly (3-hydroxybutyrate) as the sole carbon source. Two of the test bacteria were positive for PHB biosynthesis and were identified based on gram staining, biochemical tests, 16S rRNA gene sequence analysis as Pseudomonas aeruginosa and Bacillus licheniformis which grew at 37 and up to 65 °C respectively, thus suggesting the later to be thermotolerant. In this study, the effects of different carbon and nitrogen sources on PHB production in these strains were investigated. Maximum PHB production was obtained in 48 hr for the two strains and amounted to yields of 72.86 and 62.22 percentages for Bacillus licheniformis and Pseudomonas aeruginosa respectively. In these strains, glycine was the most efficient carbon sources for the production of PHB compared with other carbon (glucose, lactose, sucrose, Arabinose) and nitrogen (L- glycine, L-cysteine, DL-Tryptophan, and Potassium Nitrate) sources. The screening of microbial strains for industrial PHB production should be based on several factors including the cell’s capability to mineralize an inexpensive substrate, rate of growth and the extent of polymer accumulation.

Keywords: bacteria, poly-3-hydroxybutyrate (PHB), hydrocarbon, thermotolerant

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Authors: Mohammad Mosaddik Rahman

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This study embarks on an exploratory journey to assess the pervasive issue of heavy metal contamination in the water bodies along Chittagong Coast, Bangladesh. Situated along the mesmerizing Bay of Bengal, known for its potential as an emerging tourist haven, economic zone, ship breaking yard, confronts significant environmental hurdles. The core of these challenges lies in the contamination from heavy metals such as lead, cadmium, chromium, and mercury, which detrimentally impact both the ecological integrity and public health of the region. This contamination primarily stems from industrial activities, particularly those involving metallurgical and chemical processes, which release these metals into the environment, leading to their accumulation in soil and water bodies. The study's primary aim is to conduct a thorough assessment of heavy metal pollution levels, alongside an analysis of nutrient variations, focusing on nitrates and nitrites. Methodologically, the study leverages systematic sampling and advanced analytical tools like the Hach 3900 spectrophotometer to ensure precise and reliable data collection. The implications of heavy metal presence are multifaceted, affecting microbial and aquatic life, and posing severe health risks to the local population, including respiratory problems, neurological disorders, and an increased risk of cancer. The results of this study highlight the urgent need for effective mitigation strategies and regulatory measures to address this critical issue. By providing a comprehensive understanding of the environmental and public health implications of heavy metal contamination in Chittagong Coast, this research endeavours to serve as a catalyst for change, emphasising the need for pollution control and advancements in water management policies. It is envisioned that the outcomes of this study will guide stakeholders in collaborating to develop and implement sustainable solutions, ultimately safeguarding the region’s environment and public health.

Keywords: heavy metal, environmental health, pollution control policies, shipbreaking yard

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Authors: Zakieh Rostamzadeh Khameneh, Nariman Sepehrvand, Khalkhali-Zahra Shirmohamadi

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Background: Although the hepatitis E virus mostly causes a self-limited disease in general population, the disease is more severe in pregnant women. Hepatitis E accounts for about 10% of pregnancy-associated deaths in southern Asia. Methods: 136 pregnant women who referred to urban health centers of Urmia for pursuing pregnancy-related health services were selected randomly and enrolled in a descriptive, cross-sectional study. Each subject was tested for the presence of anti-HEV IgG antibody using an enzyme-linked immunosorbent assay (ELISA, Dia.Pro). Results: The mean age among 136 pregnant women was 25.12±4.91 years old (range of 14-39 years). Only five cases (3.6%) among all 136 subjects were demonstrated to be seropositive for anti-HEV IgG using ELISA method. There was no significant difference between age (P=0.88), income level (P=0.19) of two seropositive and seronegative groups. All seropositive cases were from urban areas. Conclusion: The seroprevalence of anti-HEV IgG is low in the population of pregnant women in Urmia, Iran. Because of limited sample size in this study, we recommend to perform further studies with larger sample size in other regions of Iran in order to be able to systematically generalize the findings of studies to the population of Iranian pregnant women.

Keywords: pregnancy, hepatitis E, women, ELISA

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Authors: Supakan Rattanakon, Jakkrapan Boonpimon, Akkaragiat Bhuangsaeng, Aphiwat Ratriphruek

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Rice bran oil (RBO) has been found to lower the level of serum cholesterol, has antioxidant and anti-carcinogenic property, and attenuate allergic inflammation. These properties of RBO are due to antioxidant compositions, especially, phenolic compounds. The higher amount of these active compounds in RBO, the greater value of RBO is. Thermal process of rice bran before solvent RBO extraction has been found to have a higher phenolic contents. Therefore, the purpose of this study is to using different heating methods on rice bran before the solvent extraction. Then, % yield of RBO, total phenolic content (TPC), and antioxidant property of two white Thai rice; KDML105 and RD6 were determined. The Folin-Ciocalteu colorimetric assay was used to determine TPC and scavenging of free radicals (DPPH) was used to determine antioxidant property expressed as EC50. The result showed that thermal process did not increase % yield of RBO but increase the TPC with 1.41 mg gallic acid equivalent (GAEmg-1). The highest TPC was found in KDML105 by using sonicator. The highest antioxidant activity was found in RD6 using autoclave. The EC50 of RBO was 0.04 mg/mL. Further study should be performed on different pretreatments to increase the TPC and antioxidant property.

Keywords: antioxidant, rice bran oil, total phenol content, white rice

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Authors: Aferdita Dinaku, Jonida Canaj

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Aflatoxins are potentially toxic metabolites produced by certain kinds of fungi (molds) that are found naturally all over the world; they can contaminate food crops and pose a serious health threat to humans by mutagenic and carcinogenic effects. Several types of aflatoxin (14 or more) occur in nature. In Albanian nutrition, cereals (especially wheat and corn) are common ingredients in some traditional meals. This study aimed to investigate the presence of aflatoxins in the flour of wheat and maize that are consumed in Albania’s markets. The samples were collected randomly in different markets in Albania and detected by the ELISA method, measured in 450 nm. The concentration of total aflatoxins was analyzed by enzyme-linked immunosorbent assay (ELISA), and they were ranged between 0.05-1.09 ppb. However, the screened mycotoxin levels in the samples were lower than the maximum permissible limits of European Commission No 1881/2006 (4 μg/kg). The linearity of calibration curves was good for total aflatoxins (B1, B2, G1, G2, M1) (R²=0.99) in the concentration range 0.005-4.05 ppb. The samples were analyzed in two replicated measurements and for each sample, the standard deviation (statistical parameter) is calculated. The results showed that the flour samples are safe, but the necessity of performing such tests is necessary.

Keywords: aflatoxins, ELISA technique, food contamination, flour

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Authors: Satya Eswari Jujjavarapu, Swasti Dhagat, Lata Upadhyay, Reecha Sahu

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Silver nanoparticles have a broad range of antimicrobial and antifungal properties ranging from soaps, pastes to sterilization and drug delivery systems. These can be synthesized by physical, chemical and biological methods; biological methods being the most popular owing to their non-toxic nature and reduced energy requirements. Microbial surfactants, produced on the microbial cell surface or excreted extracellularly are an alternative to synthetic surfactants for the production of silver nanoparticles. Hence, they are also called as green molecules. Microbial lipopeptide surfactants (biosurfactant) exhibit anti-tumor and anti-microbial properties and can be used as drug delivery agents. In this study, biosurfactant was synthesized by using a strain of acillus subtilis. The biosurfactant thus produced was analysed by emulsification assay, oil spilling test, and haemolytic test. Biosurfactant-mediated silver nanoparticles were synthesised by microwave irradiation of the culture supernatant and further characterized by UV–vis spectroscopy for a range of 400-600 nm. The UV–vis spectra showed a surface plasmon resonance vibration band at 410 nm corresponding to the peak of silver nanoparticles.

Keywords: biosurfactant, Bacillus subtilis, silver nano particle, lipopeptide

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Authors: Mahmoud Mabrok, Channarong Rodkhum

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Flavobacterium columanre is one of the devastating pathogen that causes noticeable economic losses in freshwater cultured fish. Like other filamentous bacteria, F. columanre tends to aggregate and fluctuate to all kind of media, thus revealing obstacles in recognition of its colonies. Since the molecular typing is the only fundamental tool for rapid and precise detection of this pathgen. The present study developed a species-specific PCR assay based on cslA unique gene of F. columnare. The cslA gene sequences of 13 F. columnare, strains retrieved from gene bank database, were aligned to identify a conserved homologous segment prior to primers design. The new primers yielded amplicons of 287 bp from F. columnare strains but not from relevant or other pathogens, unlike to other published set that showed no specificity and cross-reactivity with F. indicum. The primers were sensitive and detected as few as 7 CFUs of bacteria and 3 pg of gDNA template. The sensitivity was reduced ten times when using tissue samples. These primers precisely defined all field isolates in a double-blind study, proposing their applicable use for field detection.

Keywords: Columnaris infection, cslA gene, Flavobacterium columnare, PCR

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Authors: Christine O. Wangia, Jennifer A. Orwa, Francis W. Muregi, Patrick G. Kareru, Kipyegon Cheruiyot, Eric Guantai

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Ruellia (syn. Dipteracanthus) species are wild perennial creepers belonging to the Acanthaceae family. These species are reported to possess anti-inflammatory, analgesic, antioxidant, gastroprotective, anticancer, and immuno-stimulant properties. Phytochemical screening of both aqueous and methanolic extracts of Ruellia species revealed the presence of saponins. Saponins have been reported to possess anti-inflammatory, antioxidant, immuno-stimulant, antihepatotoxic, antibacterial, anticarcinogenic, and antiulcerogenic activities. The objective of this study was to quantify and analyze the Fourier transform infrared (FTIR) spectra of saponins in crude extracts of three Kenyan Ruellia species namely Ruellia prostrata (RPM), Ruellia lineari-bracteolata (RLB) and Ruellia bignoniiflora (RBK). Sequential organic extraction of the ground whole plant material was done using petroleum ether (PE), chloroform, ethyl acetate (EtOAc), and absolute methanol by cold maceration, while aqueous extraction was by hot maceration. The plant powders and extracts were mixed with spectroscopic grade KBr and compressed into a pellet. The infrared spectra were recorded using a Shimadzu FTIR spectrophotometer of 8000 series in the range of 3500 cm^-1 - 500 cm^-1. Quantitative determination of the saponins was done using standard procedures. Quantitative analysis of saponins showed that RPM had the highest quantity of crude saponins (2.05% ± 0.03), followed by RLB (1.4% ± 0.15) and RBK (1.25% ± 0.11), respectively. FTIR spectra revealed the spectral peaks characteristic for saponins in RPM, RLB, and RBK plant powders, aqueous and methanol extracts; O-H absorption (3265 - 3393 cm^-1), C-H absorption ranging from 2851 to 2924 cm^-1, C=C absorbance (1628 - 1655 cm^-1), oligosaccharide linkage (C-O-C) absorption due to sapogenins (1036 - 1042 cm^-1). The crude saponins from RPM, RLB and RBK showed similar peaks to their respective extracts. The presence of the saponins in extracts of RPM, RLB and RBK may be responsible for some of the biological activities reported in the Ruellia species.1

Keywords: Ruellia bignoniiflora, Ruellia linearibracteolata, Ruellia prostrata, Saponins

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Authors: Sana Abbasa, Shahzad Bhattiab, Nadir Khan

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Sargassum siliquastrum is brown seaweed with traditional claims for some medicinal properties. This research was done to investigate the methanol extract of S. siliquastrum for antiproliferative activity against human cervical cancer cell line, HeLa and its mode of cell death. From methylene blue assay, S. siliquastrum exhibited antiproliferative activity on HeLa cells with IC50 of 3.87 µg/ml without affecting non-malignant cells. Phase contrast microscopy indicated the confluency reduction in HeLa cells and changes on the cell shape. Nuclear staining with Hoechst 33258 displayed the formation of apoptotic bodies and fragmented nuclei. S. siliquastrum also induced early apoptosis event in HeLa cells as confirmed by FITC-Annexin V/propidium iodide staining by flow cytometry analysis. Cell cycle analysis indicated growth arrest of HeLa cells at G1/S phase. Protein study by flow cytometry indicated the increment of p53, slight increase of Bax and unchanged level of Bcl-2. In conclusion, S. siliquastrum demonstrated an antiproliferative activity in HeLa cell by inducing G1/S cell cycle arrest via p53-mediated pathway.

Keywords: sargassum siliquastrum, cervical cancer, P53, antiproleferation

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Authors: Drashti G. Daraji, Rosa E. Moo-Puc, Hitesh D. Patel

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Substituted 2-Thioimidazole analogues have been synthesized and confirmed by advanced spectroscopic techniques. Among them, ten compounds have been selected and evaluated for their in vitro anti-cancer activity at the National Cancer Institute (NCI) for testing against a panel of 60 different human tumor cell lines derived from nine neoplastic cancer types. Furthermore, synthesized compounds were tested for their in vitro antiprotozoal activity, and none of them exhibited significant potency against antiprotozoans. It was observed that the tested all compounds seem effective on the UACC-62 melanoma cancer cell line as compared to other cancer cell lines and also exhibited the least potent in the Non-Small Cell Lung Cancer cell line in one-dose screening. In silico studies of these derivatives were carried out by molecular docking techniques and Absorption, Distribution, Metabolism, and Excretion (ADME) using Schrödinger software to find potent B-Raf kinase inhibitor (PDB ID: 3OG7). All the compounds have been performed for docking study; Compound D4 has a good docking score for melanoma cancer as compared with other.

Keywords: anticancer activity, cancer cell line, 2-thio imidazole, one-dose assay, molecular docking

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Authors: G. Gevorgyan, S. Khudaverdyan, A. Vaseashta

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The Lake Sevan basin is situated in the eastern part of the Republic of Armenia (Gegharquniq marz/district). The heavy metal pollution of the some tributaries of Lake Sevan was investigated. Water sampling was performed in August and December, 2014 from the 4 observation sites: 1) Sotq river upstream (about 600 meters upstream from the Sotq gold mine); 2) Sotq river mouth; 3) Masrik river mouth; 4) Dzknaget river mouth. Heavy metal (V, Fe, Ni, Cu, As, Mo, Pb) concentrations in the water samples were determined by the standard methods using an atomic absorption spectrophotometer. The results of the study showed that heavy metal content mainly increased from the upstream of the Sotq river to the mouth of the Masrik river which may have been conditioned by the influence of gold mining activity as the Masrik and its tributary-Sotq rivers passing through the gold mining area were exposed to heavy metal pollution. The observation sites can be ranked by pollution degree as follows: №3> №2> №1> №4. The highest heavy metal pollution degree was observed in the Masrik river mouth which may have been conditioned by the direct impact of gold mining activity and the pressure of its tributary–the Sotq river which flows through the gold mining area. The lowest heavy metal pollution degree was registered in the Dzknaget river mouth which flowing through rural areas wasn’t subject to significant heavy metal pollution. According to the observation sites of the Sotq and Masrik rivers, high positive correlation was mainly observed between the concentrations of the investigated heavy metals (except nickel) which indicated that all the heavy metals except the nickel had the same anthropogenic pollution source which was the activity of the Sotq gold mine. In general, it is possible to state that the activity of the Sotq gold mine in the Lake Sevan basin caused the heavy metal pollution of the Sotq and Masrik rivers which may have posed environmental hazards. Heavy metals are nondegradable substances, and heavy metal pollution of freshwater systems may pose risks to the environment and human health through accumulation in the tissues of aquatic organisms, water-food chain as well as oral ingestion and dermal contact.

Keywords: Armenia, Lake Sevan basin, gold mining activity, river ecosystems, heavy metal pollution

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Authors: H. Aksas, S. Boughrara, K. Louhab

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The goal of this study was the use of life cycle assessment (LCA) methodology to assess the environmental impact of pharmaceutical product (four kinds of syrup form and tree kinds of pomade form), which are produced in one leader manufactory in Algeria town that is SAIDAL Company. The impacts generated have evaluated using SimpaPro7.1 with CML92 Method for syrup form and EPD 2007 for pomade form. All impacts evaluated have compared between them, with determination of the compound contributing to each impacts in each case. Data needed to conduct Life Cycle Inventory (LCI) came from this factory, by the collection of theoretical data near the responsible technicians and engineers of the company, the practical data are resulting from the assay of pharmaceutical liquid, obtained at the laboratories of the university. This data represent different raw material imported from European and Asian country necessarily to formulate the drug. Energy used is coming from Algerian resource for the input. Outputs are the result of effluent analysis of this factory with different form (liquid, solid and gas form). All this data (input and output) represent the ecobalance.

Keywords: pharmaceutical product, drug residues, LCA methodology, environmental impacts

Procedia PDF Downloads 235

Authors: Gulcin Yildiz, Hao Feng

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Soybean proteins are the most widely used and researched proteins in the food industry. Due to soy allergies among consumers, however, alternative legume proteins having similar functional properties have been studied in recent years. These alternative proteins are also expected to have a price advantage over soy proteins. One such protein that has shown good potential for food applications is pea protein. Besides the favorable functional properties of pea protein, it also contains fewer anti-nutritional substances than soy protein. However, a comparison of the physicochemical properties of pea protein isolate (PPI)-starch nanocomplexes and soy protein isolate (SPI)-starch nanocomplexes treated by ultrasound has not been well documented. This study was undertaken to investigate the effects of ultrasound treatment on the physicochemical properties of PPI-starch and SPI-starch nanocomplexes. Pea protein isolate (85% pea protein) provided by Roquette (Geneva, IL, USA) and soy protein isolate (SPI, Pro-Fam® 955) obtained from the Archer Daniels Midland Company were adjusted to different pH levels (2-12) and treated with 5 minutes of ultrasonication (100% amplitude) to form complexes with starch. The soluble protein content was determined by the Bradford method using BSA as the standard. The turbidity of the samples was measured using a spectrophotometer (Lambda 1050 UV/VIS/NIR Spectrometer, PerkinElmer, Waltham, MA, USA). The volume-weighted mean diameters (D4, 3) of the soluble proteins were determined by dynamic light scattering (DLS). The emulsifying properties of the proteins were evaluated by the emulsion stability index (ESI) and emulsion activity index (EAI). Both the soy and pea protein isolates showed a U-shaped solubility curve as a function of pH, with a high solubility above the isoelectric point and a low one below it. Increasing the pH from 2 to 12 resulted in increased solubility for both the SPI and PPI-starch complexes. The pea nanocomplexes showed greater solubility than the soy ones. The SPI-starch nanocomplexes showed better emulsifying properties determined by the emulsion stability index (ESI) and emulsion activity index (EAI) due to SPI’s high solubility and high protein content. The PPI had similar or better emulsifying properties at certain pH values than the SPI. The ultrasound treatment significantly decreased the particle sizes of both kinds of nanocomplex. For all pH levels with both proteins, the droplet sizes were found to be lower than 300 nm. The present study clearly demonstrated that applying ultrasonication under different pH conditions significantly improved the solubility and emulsify¬ing properties of the SPI and PPI. The PPI exhibited better solubility and emulsifying properties than the SPI at certain pH levels

Keywords: emulsifying properties, pea protein isolate, soy protein isolate, ultrasonication

Procedia PDF Downloads 294

Authors: Purnima Rawat, Divya Vohora, Sarika Gupta, Farhan J. Ahmad, Sushama Talegaonkar

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Nanoparticles (NPs) that target bone tissue were developed using PLGA–mPEG (poly(lactic-co-glycolic-acid)–polyethylene glycol) diblock copolymers by using pamidronate as a bone-targeting moieties. These NPs are expected to enable the transport of hydrophilic drugs. The NP was prepared by in situ polymerization method, and their in- vitro characteristics were evaluated using dynamic light scattering, transmission electron microscopy (TEM) and in phosphate-buffered solution. The bone targeting potential of the NP was also evaluated on in-vitro pre-osteoblast MCT3E1 cell line using ALP activity, degree of mineralization and RT-PCR assay. The average particle size of the NP was 101.6 ± 3.7nm, zeta potential values were negative (-25±0.34mV) of the formulations and the entrapment efficiency was 93± 3.1 % obtained. The moiety of the PLGA–mPEG–pamidronate NPs exhibited the best apatite mineral binding ability in-vitro MCT3E1 pre-osteoblast cell line. Our results suggested that the developed nanoparticles may use as a delivery system for Pamidronate in bone repair and regeneration, warranting further evaluation of the treatment of bone disease.

Keywords: nanoparticle, pamidronate, in-situ polymerization, osteoblast

Procedia PDF Downloads 471

Authors: Soumita Mitra, Santosh Kumar Sarkar

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Hooghly river estuary (HRE), situated at the north eastern part of Bay of Bengal has global significance due to its holiness. It is of immense importance to the local population as it gives perpetual water supply for various activities such as transportation, fishing, boating, bathing etc. to the local people who settled on both the banks of this estuary. This study was done to assess the dissolved and particulate trace metal in the estuary covering a stretch of about 175 Km. The water samples were collected from the surface (0-5 cm) along the salinity gradient and metal concentration were studied both in dissolved and particulate phase using Graphite Furnace Atomic Absorption Spectrophotometer (GF-AAS) along some physical characteristics such as water temperature, salinity, pH, turbidity and total dissolved solids. Although much significant spatial variation was noticed but little enrichment was found along the downstream of the estuary. The mean concentration of the metals in the dissolved and particulate phase followed the same trend and as follows: Fe>Mn>Cr>Zn>Cu>Ni>Pb. The concentration of the metals in the particulate phase were much greater than that in dissolved phase which was also depicted from the values of the partition coefficient (Kd)(ml mg-1). The Kdvalues ranged from 1.5x105 (in case of Pb) to 4.29x106 (in case of Cr). The high value of Kd for Cr denoted that the metal Cr is mostly bounded with the suspended particulate matter while the least value for Pb signified it presence more in dissolved phase. Moreover, the concentrations of all the studied metals in the dissolved phase were many folds higher than their respective permissible limits assested by WHO 2008, 2009 and 2011. On the other hand, according to Sediment Quality Guidelines (SQGs), Zn, Cu and Ni in the particulate phase lied between ERL and ERM values but Cr exceeded ERM values at all the stations confirming that the estuary is mostly contaminated with the particulate Cr and it might cause frequent adverse effects on the aquatic life. Multivariate statistics Cluster analysis was also performed which separated the stations according to the level of contamination from several point and nonpoint sources. Thus, it is found that the estuarine system is much polluted by the toxic metals and further investigation, toxicological studies should be implemented for full risk assessment of this system, better management and restoration of the water quality of this globally significant aquatic system.

Keywords: dissolved and particulate phase, Hooghly river estuary, partition coefficient, surface water, toxic metals

Procedia PDF Downloads 259

Authors: Ashish Kumar Jain, Deepak Mishra

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The objective of the present investigation was to prepare and evaluate a vesicular dual drug delivery system for effective management of mucosal ulcer. Inner encapsulating and Double liposomes were prepared by glass bead and reverse phase evaporation method respectively. The formulation consisted of inner liposomes bearing Ranitidine Bismuth Citrate (RBC) and outer liposomes encapsulating Amoxicillin trihydrate (AMOX). The optimized inner liposomes and double liposomes were extensively characterized for vesicle size, morphology, zeta potential, vesicles count, entrapment efficiency and in vitro drug release. In vitro, the double liposomes demonstrated a sustained release of AMOX and RBC viz 91.4±1.8% and 77.2±2.1% respectively at the end of 72 hr. Furthermore binding specificity and targeting propensity toward H. pylori (SKP-56) was confirmed by agglutination and in situ adherence assay. Reduction of the absolute alcohol induced ulcerogenic index from 3.01 ± 0.25 to 0.31 ± 0.09 and 100% H. pylori clearance rate was observed. These results suggested that double liposomes are potential vector for the development of dual drug delivery for effective treatment of H. pylori-associated peptic ulcer.

Keywords: double liposomes, H. pylori targeting, PE liposomes, glass-beads method, peptic ulcers

Procedia PDF Downloads 430

Authors: Izdihar Ismail, Alona C. Linatoc, Maryati Mohamed

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The tropical rainforest of Malaysia is known for its rich biological diversity and high endemicity. The potential for these forests to hold the cure for many diseases and illnesses is high and much is yet to be discovered. This study explores the richness of the tropical rainforest of Endau-Rompin National Park in Johor, Malaysia in search of plants traditionally used by the indigenous people in the treatment of malaria and malaria-like symptoms. Seven species of plants were evaluated and tested for antiplasmodial activities. Different plant parts were subjected to methanolic and aqueous extractions. A total of 24 extracts were evaluated by histidine-rich protein II (HRP2) assay against K1 strain of Plasmodium falciparum chloroquine-resistant. Ten extracts showed significant inhibition of the growth of P. falciparum. Phytochemical screening of the same extracts revealed the presence of alkaloids, flavonoids, terpenoids and anthraquinones. This study affirms that tropical rainforests may still hold undiscovered cures for many diseases and illnesses that have inflicted millions of people worldwide. The species studied herein have not known to have been studied elsewhere before.

Keywords: Endau-Rompin, malaria, Malaysia, tropical rainforest, traditional knowledge

Procedia PDF Downloads 254

Authors: Abdelkrim Cheriti, Mebarka Belboukhari, Nasser Belboukhari

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Launaea Cass. is a small genus of the family Asteraceae (tribe Lactuceae, subtribe Sonchinae), consisting of 54 species, of which 9 are presented in the flora of Algeria and is mainly distributed in the South Mediterranean, Africa and SW Asia. Plants in the Launaea genus have been used ethnobotanically as bitter stomachic, for treating diarrhea, gastrointestinal tracts, as anti-inflammatory, for skin diseases, treatment of infected wounds, hepatic pains, children fever, as soporific, lactagogue, diuretic and as insecticidal. Antioxidants are vital substances, which possess the ability to protect the body from damages caused by free radical induced oxidative stress. A variety of free radical scavenging antioxidants is found in a number of dietary sources. The main objective of this study focused on the screening of antioxidant activity of Launaea nudicaulis (Asteraceae) extracts. The in vitro antioxidant activity was investigated with DPPH radical scavenging assay. The quantitative evaluation of DPPH scavenging activity showed that n-BuOH and EtOAc extracts are the most active extracts with a percentage of antiradical activity of 89,62% and 71,57% respectively.

Keywords: Launaea, phytochemical, South Algeria, Sahara, endemic specie

Procedia PDF Downloads 427

Authors: Pakakrong Thongdeeying, Srisopa Ruangnoo, Arunporn Itharat

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The rhizomes of Smilax corbularia Kunth have long been used as common ingredients in anticancer preparations. Thus, the objective of this study is to investigate cytotoxicity of S. corbularia and its ingredients against cholangiocarcinoma cell line (KKU-M156) by SRB assay. Ethanolic and water extracts of S. corbularia rhizomes were obtained using the procedures followed by Thai traditional doctors. Bioassay guided isolation was used to isolate cytotoxic compounds. The results revealed that the ethanolic extract of S. corbularia exhibited activity against KKU-M156 cell line with an IC50 value of 84.53±1.62 µg/ml, but the water extract showed no cytotoxic activity. Three flavonoid compounds [astilbin (1), engeletin (2), and quercetin (3)] were isolated from the ethanolic extract. Compound 3 exhibited the strongest activity against KKU-M156 cell line (IC50 = 8.14 ± 1.15 µg/ml), but 1 and 2 showed no cytotoxic activity (IC50 > 100 µg/ml). In conclusion, quercetin showed the highest efficacy against cholangiocarcinoma. These results support the traditional use of this plant by Thai traditional doctors for cancer treatment.

Keywords: cholangiocarcinoma, cytotoxicity, flavonoid, Smilax corbularia

Procedia PDF Downloads 363

Authors: Banu Hatice Gürcüm, Pınar Arslan, Mahmut Yalçın

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Fashion designers create lots of dresses, suits, shoes, and other clothing and accessories, which are purchased every year by consumers. Fashion trends, sketches of designs, accessories affect the apparel goods, but colors make the finishing touches to an outfit. In all fields of apparel men's, women's, and children's wear, including casual wear, suits, sportswear, formal wear, outerwear, maternity, and intimate apparel, color sells. Thus, specialization in color in apparel is a basic concern each season. The perception of color is the key to sales for every sector in textile business. Mechanism of color perception, cognition in brain and color emotion are unique subjects, which scientists have been investigating for many years. The parameters of color may not be corresponding to visual scales since human emotions induced by color are completely subjective. However, with a very few exception each manufacturer concern their top selling colors for each season through seasonal sales reports of apparel companies. This paper examines sensory and instrumental methods for quantifying color of fabrics and investigates the relationship between fabric color and sale numbers. 5 top selling colors for each season from 10 leading apparel companies in the same segment are taken. The compilation is based according to the sales of the companies for 5 to 10 years. The research’s main concern is the corelation with the magnitude of seasonal color selling figures and the CIE chromaticity coordinates. The colors are chosen from the globally accepted Pantone Textile Color System and the three-dimentional measurement system CIE L*a*b* (CIELAB) is used, L* representing the degree of lightness of color, a* the degree of color ranging from magenta to green, and b* the degree of color ranging from blue to yellow. The objective of this paper is to demonstrate the feasibility of relating color perceptance to a laboratory instrument yielding measurements in the CIELAB system. Our approach is to obtain a total of a hundred reference fabrics to be measured on a laboratory spectrophotometer calibrated to the CIELAB color system. Relationships between the CIE tristimulus (X, Y, Z) and CIELAB (L*, a*, b*) are examined and are reported herein.

Keywords: CIELAB, CIE tristimulus, color preference, fashion

Procedia PDF Downloads 318

Authors: Manal Kamel, Sara Maher

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Development of sensitive non-invasive tests for detection of SARS-CoV-2 antigens is imperative to manage the extent of infection throughout the population, yet, it is still challenging. Here, we designed and optimized a sandwich enzyme-linked immunosorbent assay (ELISA) for SARS-CoV-2 S1 antigen detection in saliva. Both saliva samples and nasopharyngeal swapswere collected from 170 PCR-confirmed positive and negative cases. Gold nanoparticles (AuNPs) were conjugated with S1protein receptor binding domain (RBD) nanobodies. Recombinant S1 monoclonal antibodies (S1mAb) as primery antibody and gold conjugated nanobodies as secondary antibody were employed in sandwich ELISA. Our developed system were optimized to achieve 87.5 % sensitivity and 100% specificity for saliva samples compared to 89 % and 100% for nasopharyngeal swaps, respectively. This means that saliva could be a suitable replacement for nasopharyngeal swaps No cross reaction was detected with other corona virus antigens. These results revealed that our developed ELISAcould be establishedas a new, reliable, sensitive, and non-invasive test for diagnosis of SARS-CoV-2 infection, using the easily collected saliva samples.

Keywords: COVID 19, diagnosis, ELISA, nanobodies

Procedia PDF Downloads 111