Search results for: RP- HPLC

Authors: Elena G. Salina, Vadim A. Makarov

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With the emergence of primary resistance to the current drugs and wide distribution of latent tuberculosis infection, a need for new compounds with a novel mode of action is growing steadily. Copper-mediated innate immunity and antibacterial toxicity propose novel strategies in TB drug discovery and development. Transcriptome of M. tuberculosis was obtained by RNA-seq, intracellular copper content was measured by ISP MS and complexes of 1-hydroxy-2-thiopyridines with copper were detected by HPLC.1-hydroxy-2-thiopyridine derivatives were found to be highly active in vitro against both actively growing and dormant non-culturable M. tuberculosis. Transcriptome response to 1-hydroxy-2-thiopyridines revealed signs of copper toxicity in M. tuberculosis bacilli. Indeed, Cu was found to accumulate inside cells treated with 1-hydroxy-2-thiopyridines. These compounds were found to form stable charged lipophylic complexes with Cu²⁺ ions which transport into mycobacterial cell. Subsequent metabolic destruction of the complex led to transformation of 1-hydroxy-2-thiopyridines into 2-methylmercapto-2-ethoxycarbonylpyridines, which did not possess antitubercular activity and releasing of free Cu²⁺ in the cytoplasm. 1-hydroxy-2-thiopyridines are a potent class of Cu-dependent inhibitors of M. tuberculosis which may control M. tuberculosis infection by impairment of copper homeostasis. Acknowledgment: This work was financially supported by the Ministry of Education and Science of the RussianFederation (Agreement No 14.616.21.0065; unique identifier RFMEFI61616X0065).

Keywords: copper toxicity, drug discovery, M. tuberculosis inhibitors, 2-thiopyridines

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Authors: Monica Rosa Loizzo, Tiziana Falco, Marco Bonesi, Maria Concetta Tenuta, Mariarosaria Leporini, Rosa Tundis

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Extra Virgin Olive Oil (EVOO) is a major source of fat in the Mediterranean diet and its nutritional properties are the main reason for the increment of its consumption all over the world in recent years. In terms of olive oil production, Italy ranks the second in the world. EVOO is obtained exclusively by physical methods from the fruit of Olea europea L. Frantoio cv is spread in all the Italian territory. The aim of this work is to identify the phenolic and fatty acids profile of EVOO from Frantoio cv growing in different area of Calabria (Italy). The phenolic profile was obtained by HPLC coupled to a diode array detector and mass spectrometry. Analyses revealed the presence of phenolic alcohols, phenolic acid, several secoiridoids, and two flavones as main components. Hydroxytyrosol and tyrosol are present in reasonable content. Fatty acids were monitored by gas chromatography. Oleic acid was the most abundant compounds. A moderate level of linoleic acid, in accordance with the general observations for oils derived from Mediterranean countries, was also found.

Keywords: extra virgin olive oils, frantoio cv, phenolic compounds, fatty acids

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Authors: Yi-Feng Kao, Huey-Jine Chai, Chin-I Chang, Yi-Chen Chen, June-Ru Chen

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Microglia is a macrophage that resides in brain, and overactive microglia may result in brain neuron damage or inflammation. In this study, the phospholipids was extracted from squid skin and manufactured into a liposome (SQ liposome) to mimic apoptotic body. We then evaluated anti-inflammatory effects of SQ liposome on mouse microglial cell line (BV-2) by lipopolysaccharide (LPS) induction. First, the major phospholipid constituents in the squid skin extract were including 46.2% of phosphatidylcholine, 18.4% of phosphatidylethanolamine, 7.7% of phosphatidylserine, 3.5% of phosphatidylinositol, 4.9% of Lysophosphatidylcholine and 19.3% of other phospholipids by HPLC-UV analysis. The contents of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in the squid skin extract were 11.8 and 28.7%, respectively. The microscopic images showed that microglia cells can engulf apoptotic cells or SQ-liposome. In cell based studies, there was no cytotoxicity to BV-2 as the concentration of SQ-liposome was less than 2.5 mg/mL. The LPS induced pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), were significant suppressed (P < 0.05) by pretreated 0.03~2.5mg/ml SQ liposome. Oppositely, the anti-inflammatory cytokines transforming growth factor-beta (TGF-β) and interleukin-10 (IL-10) secretion were enhanced (P < 0.05). The results suggested that SQ-liposome possess anti-inflammatory properties on BV-2 and may be a good strategy for against neuro-inflammatory disease.

Keywords: apoptotic mimicry, neuroinflammation, microglia, squid processing by-products

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Authors: Parvaneh Hajeb, Shahram Shakibazadeh, Md. Zaidul Islam Sarker

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High-quality fish oil for human consumption requires low levels of toxic elements. The aim of this study was to develop a method to extract oil from fish wastes with the least toxic elements contamination. Supercritical fluid extraction (SFE) was applied to detoxify fish oils from toxic elements. The SFE unit used consisted of an intelligent HPLC pump equipped with a cooling jacket to deliver CO2. The freeze-dried fish waste sample was extracted by heating in a column oven. Under supercritical conditions, the oil dissolved in CO2 was separated from the supercritical phase using pressure reduction. The SFE parameters (pressure, temperature, CO2 flow rate, and extraction time) were optimized using response surface methodology (RSM) to extract the highest levels of toxic elements. The results showed that toxic elements in fish oil can be reduced using supercritical CO2 at optimum pressure 40 MPa, temperature 61 ºC, CO2 flow rate 3.8 MPa, and extraction time 4.25 hr. There were significant reductions in the mercury (98.2%), cadmium (98.9%), arsenic (96%), and lead contents (99.2%) of the fish oil. The fish oil extracted using this method contained elements at levels that were much lower than the accepted limits of 0.1 μg/g. The reduction of toxic elements using the SFE method was more efficient than that of the conventional methods due to the high selectivity of supercritical CO2 for non-polar compounds.

Keywords: food safety, toxic elements, fish oil, supercritical carbon dioxide

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Authors: H. Boucheloukh, S. Rouissa, N. Aoun, M. Beloucifa, T. Sehili, F. Parrino, V. Loddo

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To optimize water purification and wastewater treatment by heterogeneous photocatalysis, we used NiFe₂O₄ as a catalyst and solar irradiation as a source of energy. In this concept, an organic substance present in many industrial effluents was chosen: naproxen ((S)-6-methoxy-α-methyl-2-naphthaleneacetic acid or 2-(6-methoxynaphthalenyl) propanoic), a non-steroidal anti-inflammatory drug. The main objective of this study is to degrade naproxen by an iron and nickel catalyst, the degradation of this organic pollutant by nickel ferrite has been studied in a heterogeneous aqueous medium, with the study of the various factors influencing photocatalysis such as the concentration of matter and the acidity of the medium. The photocatalytic activity was followed by HPLC-UV andUV-Vis spectroscopy. A first-order kinetic model appropriately fitted the experimental data. The degradation of naproxen was also studied in the presence of H₂O₂ as well as in an aqueous solution. The new hetero-system NiFe₂O₄/oxalic acid is also discussed. The fastest naproxen degradation was obtained with NiFe₂O₄/H₂O₂. In a first-place, we detailed the characteristics of the material NiFe₂O₄, which was synthesized by the sol-gel methods, using various analytical techniques: visible UV spectrophotometry, X-ray diffraction, FTIR, cyclic voltammetry, luminescent discharge optical emission spectroscopy.

Keywords: naproxen, nickelate, photocatalysis, oxalic acid

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Authors: Mulatu Kassie Birhanu

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Electro-oxidation of glycerol is an important process to convert the less price glycerol into other expensive (essential) and energy-rich chemicals. In this study, nickel was electro-deposited on laboratory-made carbon ceramic electrode (CCE) substrate using electrochemical techniques that is cyclic voltammetry (CV) to prepare an electro-catalyst (Ni/CCE) for electro-oxidation of glycerol. Carbon ceramic electrode was prepared from graphite and methyl tri-methoxy silane (MTMOS) through the processes called hydrolysis and condensation with methanol in acidic media (HCl) by a sol-gel technique. Physico-chemical characterization of bare CCE and modified (deposited) CCE (Ni/CCE) was measured and evaluated by Fourier Transform Infrared spectroscopy (FTIR), Scanning Electron Microscopy (SEM) and X-ray diffraction (XRD). Electro-oxidation of glycerol was performed in 0.1 M glycerol in alkaline media (0.5 M NaOH). High-Performance Liquid Chromatography (HPLC) technique was used to identify and determine the concentration of glycerol, reaction intermediates and oxidized products of glycerol after its electro-oxidation is performed. The conversion (%) of electro-oxidation of glycerol during 9-hour oxidation was 73% and 36% at 1.8V and 1.6V vs. RHE, respectively. Formate, oxalate, glycolate and glycerate are the main oxidation products of glycerol with selectivity (%) of 75%, 8.6%, 1.1% and 0.95 % at 1.8 V vs. RHE and 55.4%, 2.2%, 1.0% and 0.6% at 1.6 V vs. RHE respectively. The result indicates that formate is the main product in the electro-oxidation of glycerol on Ni/CCE using the indicated applied potentials.

Keywords: carbon-ceramic electrode, electrodeposition, electro-oxidation, Methyltrimethoxysilane

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Authors: Faisal A. Salih

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Verapamil hydrochloride (Ver) is a drug used in medicine for arrythmia, angina and hypertension as a calcium channel blocker. For the quantitative determination of Ver in dosage forms, the HPLC method is most often used. A convenient alternative to the chromatographic method is potentiometry using a Verselective electrode, which does not require expensive equipment, can be used without separation from the matrix components, which significantly reduces the analysis time, and does not use toxic organic solvents, being a "green", "environmentally friendly" technique. It has been established in this study that the rational choice of the membrane plasticizer and the preconditioning and measurement algorithms, which prevent nonexchangeable extraction of Ver into the membrane phase, makes it possible to achieve excellent analytical characteristics of Ver-selective electrodes based on commercially available components. In particular, an electrode with the following membrane composition: PVC (32.8 wt %), ortho-nitrophenyloctyl ether (66.6 wt %), and tetrakis-4-chlorophenylborate (0.6 wt % or 0.01 M) have the lower detection limit 4 × 10−8 M and potential reproducibility 0.15–0.22 mV. Both direct potentiometry (DP) and potentiometric titration (PT) methods can be used for the determination of Ver in tablets and injection solutions. Masses of Ver per average tablet weight determined by the methods of DP and PT for the same set of 10 tablets were (80.4±0.2 and80.7±0.2) mg, respectively. The masses of Ver in solutions for injection, determined by DP for two ampoules from one set, were (5.00±0.015 and 5.004±0.006) mg. In all cases, good reproducibility and excellent correspondence with the declared quantities were observed.

Keywords: verapamil, potentiometry, ion-selective electrode, pharmaceutical analysis

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Authors: Najwa Othman, Norhidayah Suleiman, Gun Hean Chong

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Palm fatty acid distillate (PFAD) is a by-product of palm oil refineries which contains valuable compounds such as phytosterols, squalene, polycosanol, co-enzyme Q10 and vitamin E (tocopherols and tocotrienols). Approximately 0.7-1.0% of vitamin E accumulates in PFAD, and it functions as antioxidants and anti-inflammatory. The objective of this research is to evaluate the effect of manipulated variables in supercritical carbon dioxide towards the recovery of tocotrienols in PFAD. The vitamin E concentrate isolated varies depending on the pre-treatment of sample and extraction techniques. In this research, tocotrienols in PFAD was concentrated by removing the extraneous matters, especially free fatty acid (FFA) and acylglycerols. Pre-treatment method such as enzymatic hydrolysis by using lipase from Candida rugosa as an enzyme was used to remove FFA and improve recovery of vitamin E. After that, treated PFAD was extracted by using supercritical fluid extraction in co-current glass beads packed column (22 cm x 75 cm i.d) at different temperatures (40-60°C) and pressures (100-300 bar) for 5 hours. After the extraction, the sample was analyzed by using high-pressure liquid chromatography (HPLC) system to quantify the tocotrienols. The results indicated that a combined pressure (200 bar) and temperature (60°C) was predicted to provide highest tocotrienols yield and the extraction yield obtained was 106.45%.

Keywords: enzymatic hydrolysis, palm fatty acid distillate, supercritical fluid extraction, tocotrienols

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Authors: Ikram Kamal, Mohamed Blaghen

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Petroleum hydrocarbons are serious problems and global pollutants in the environment due to their toxicity, carcinogenicity and persistent organic pollutant properties. One of the approaches to enhance biodegradation of petroleum hydrocarbons is to use biosurfactant. Biosurfactants are amphiphilic biomolecules produced as metabolic by-products from microorganisms they received considerable attention in the field of environmental remediation processes such as bioremediation. Biosurfactants have been considered as a desirable alternative to synthetic surfactants in various applications particularly in the environmental field. In comparison with their synthetic counterparts, biosurfactants have been reported to be less toxic, biodegradable and persistent. In this study we have investigated the potential of bacterial strains collected aseptically from the lagoon Marchika (water and soil) in Nador, Morocco; for the production of biosurfactants. This study also aimed to optimize the biosurfactant production process by changing the variables that influence the type and amount of biosurfactant produced by these microorganisms such as: carbon sources and also other physical and chemical parameters such as temperature and pH. Emulsification index, methylene blue test and thin layer chromatography (TLC) revealed the ability of strains used in this study to produce compounds that could emulsify gasoline. In addition, a HPLC/MS was used to separate and identify different biosurfactants purified.

Keywords: petroleum hydrocarbons, biosurfactants, biodegradation, lagoon marchika, emulsification index

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Authors: T. Ould Bellahcen, A. Amiri, I. Touam, F. Hmimid, A. El Amrani, M. Cherki

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The antioxidant activity of three extracts (water, lipidic and ethanolic) prepared from the microalgae Spirulina platensis isolated from Moroccan lake, using 2, 2 diphenyl-1-picrylhydrazyl (DPPH) and 2,2’-azino-bis ethylbenzthiazoline-6-sulfonic acid (ABTS) radical assay, was studied and compared. The obtained results revealed that the IC₅₀ found using DPPH were lower than that of ABTS for all extracts from these planktonic blue-green algae. The high levels of phenolic and flavonoid content were found in the ethanolic extract 0,33 ± 0,01 mg GAE/g dw and 0,21 ± 0,01 mg quercetin/g dw respectively. In addition, using DPPH, the highest activity with IC₅₀ = 0,449 ± 0,083 mg/ml, was found for the ethanolic extract, followed by that of lipidic extract (IC₅₀ = 0,491 ± 0,059 mg/ml). The lowest activity was for the aqueous extract (IC₅₀ = 4,148 ± 0,132 mg/ml). For ABTS, the highest activity was observed for the lipidic extract with IC₅₀ = 0,740 ± 0,012 mg/ml, while, the aqueous extract recorded the lowest activity (IC₅₀ = 6,914 ± 0, 0067 mg/ml). A moderate activity was showed for the ethanolic extract (IC₅₀ = 5,852 ± 0, 0171 mg/ml). It can be concluded from this first study that Spirulina platensis extracts show an interesting antioxidant and antiradicals properties suggesting that this alga could be used as a potential source of antioxidants. A qualitative and quantitative analysis of polyphenol and flavonoids in the extracts using HPLC is in progress so as to study the correlation between the antioxidant activity and chemical composition.

Keywords: Spirulina platensis, antioxidant, DPPH, ABTS

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Authors: Harish Kumar Sharma, Garima Sengar

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Rice bran oil is considered comparatively nutritionally superior than different fats/oils. Therefore, model blends prepared from pure rice bran oil (RBO) and sunflower oil (SFO) were explored for changes in the different physicochemical parameters. Repeated deep fat frying process was carried out by using dried potato in order to study the thermal behaviour of pure rice bran oil, sunflower oil and their model blends. Pure rice bran oil and sunflower oil had shown good thermal stability during the repeated deep fat frying cycles. Although, the model blends constituting 60% RBO + 40% SFO showed better suitability during repeated deep fat frying than the remaining blended oils. The quantification of pure rice bran oil in the blended oils, physically refined rice bran oil (PRBO): SnF (sunflower oil) was carried by different methods. The study revealed that regression equations based on the oryzanol content, palmitic acid composition and iodine value can be used for the quantification. The rice bran oil can easily be quantified in the blended oils based on the oryzanol content by HPLC even at 1% level. The palmitic acid content in blended oils can also be used as an indicator to quantify rice bran oil at or above 20% level in blended oils whereas the method based on ultrasonic velocity, acoustic impedance and relative association showed initial promise in the quantification.

Keywords: rice bran oil, sunflower oil, frying, quantification

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Authors: Eleftheria Barouni, Antonia Terpou, Maria Kanellaki, Argyro Bekatorou, Athanasios A.Koutinas

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The aim of the present work was to evaluate the production of cheese using a composite filter of tubular cellulose (TC) with [a] entrapped rennin enzyme and [b] immobilized L.casei and entrapped enzyme. Tubular cellulose from sawdust was prepared after lignin removal with 1% NaOH. The biocatalysts were thermally dried at 38oC and used for milk coagulation. The effect of temperature (5,20,37 oC) of the first dried biocatalyst on the pH kinetics of milk coagulation was examined. The optimum temperature (37oC) of the first biocatalyst was used for milk coagulation with the second biocatalyst prepared by entrapment of both rennin enzyme and probiotic lactic acid bacteria in order to introduce a sour taste in cheeses. This co-biocatalyst was used for milk coagulation. Samples were studied as regards its effect on lactic acid formation and its correlation with taste test results in cheeses. For both biocatalysts samples were analyzed for total acidity and lactic acid formation by HPLC. The quality of the produced cheeses was examined through the determination of volatile compounds by SPME GC/MS analysis. Preliminary taste tests and microbiological analysis were performed and encourage us for further research regarding scale up.

Keywords: tubular cellulose, Lactobacillus casei, rennin enzyme, cheese production

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Authors: Salem Abdalla

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Objective: Dipyrone is a widely used, well tolerated analgesic drug which, however, is compromised by agranulocytosis as an adverse effect. Subsequent to no enzymatic hydrolysis, the primary metabolic step is N-demethylation of 4-methylaminoantipyrine (4-MAA) to 4-aminoantipyrine (4-AA). The aim of the present study was to identify the cytochrome P-450 enzyme (CYP) mediating this reaction. Methods: We identified the relevant CYP using virus expressed isolated rat liver microsomes with chemical inhibition studies. The substrate of 4-methylaminantipyrine was employed at six different concentrations (25, 50, 100, 400, 800, and 1200 µmol/l) with varying concentrations of selective inhibitors of CYP1A2 (furafylline, fluvoxamine), CYP3A4 (ketoconazole), CYP2A6 (coumarin), CYP2D6 (quinidine), CYP2C19 (omeprazole, fluvoxamine, tranylcypromine), CYP2C9 (sulfaphenazole), and CYP1A1 (alpha-naphthoflavone). 4-MAA and 4-AA were analyzed by HPLC, and enzyme kinetic parameters (Km and Vmax) were determined by regression (Sigma plot 9.0). Results: The N-demethylation of 4-MAA by microsomes prepared from baculovirus-expressing human CYP was pronounced with CYP2C19. Intrinsic clearances of the most active enzymes were 0.092, 0.027, and 0.026 for the CYP enzymes 2C19, 2D6, and 1A2, respectively. Metabolism by rat liver microsomes was strongly inhibited by omeprazole (IC50 of 0.05). Conclusion: The enzyme CYP2C19 apparently has an important role in N-demethylation of 4-methylaminoantipyrine which should be further analyzed in clinical studies and which may also be interesting concerning the agranulocytosis.

Keywords: dipyrone, 4-methylaminoantipyrine (4-MAA), 4- aminoantipyrine (4-AA), metabolism, human CYP2C19

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Authors: Lyanne Rodriguez, Eduardo Fuente, Andrés Trostchansky, Ivan Palomo

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Cardiovascular diseases (CVD) are the leading cause of death in the world. The development of platelet-rich thrombi has been considered a trigger for acute cardiovascular events. A healthy diet, rich in fruit and vegetables, has been related to increased protection against cardiovascular events. Previous studies have observed that tomato pomace has a potent antiplatelet activity, due could be attributed to its high content of fatty acids (> 30%). It has been shown that unsaturated fatty acids can undergo endogenous intracellular nitration reactions during digestion after lipid consumption. Additionally, nitrated fatty acids (NO2-FA) can significantly reduce atherosclerotic lesion formation, inhibiting the expression of adhesion molecules on dysfunctional endothelium and platelet activation. In this work, we have proposed the nitration of fatty acids present in tomato pomace to improve its antiplatelet action. The gastric digestion of the tomato pomace allowed the nitration of the fatty acids, while by HPLC/MS/MS we were able to identify and quantify the nitrated fatty acids. The nitrated tomase extracts showed antiplatelet potential when platelets were stimulated with TRAP-6 and collagen. This activity was related to the presence of nitrated linoleic acid, which inhibited platelet activation by flow cytometry. The knowledge about the antiplatelet activity of nitrated fatty acids from tomato pomace will further develop new and more effective agents.

Keywords: cardiovascular, tomato extracts, nitrated fatty acids, antiplatelet activity

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Authors: Nadeem A. Siddique, Mohd Mujeeb, Kahkashan

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Introduction: The objective of the projected work is to study the occurrence of the aflatoxins B1, B2, G1and G2 in remedial plants, exclusively in Delphinium denudatum. The aflatoxins were analysed by high-performance liquid chromatography–tandem quadrupole mass spectrometry with electrospray ionization (HPLC–MS/MS) and immunoaffinity column chromatography were used for extraction and purification of aflatoxins. PDA media was selected for fungal count. Results: A good quality linear relationship was originated for AFB1, AFB2, AFG1 and AFG2 at 1–10 ppb (r > 0.9995). The analyte precision at three different spiking levels was 88.7–109.1 %, by means of low per cent relative standard deviations in each case. Within 5 to7 min aflatoxins can be separated using an Agilent XDB C18-column. We found that AFB1 and AFB2 were not found in D. denudatum. This was reliable through exceptionally low figures of fungal colonies observed after 6 hr of incubation. The developed analytical method is straightforward, be successfully used to determine the aflatoxins. Conclusion: The developed analytical method is straightforward, simple, accurate, economical and can be successfully used to find out the aflatoxins in remedial plants and consequently to have power over the quality of products. The presence of aflatoxin in the plant extracts was interrelated to the least fungal load in the remedial plants examined.

Keywords: aflatoxins, delphinium denudatum, liquid chromatography, mass spectrometry

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Authors: Jae Young Jang, Jong Hoon Ahn, Jae-Woong Lim, So Young Kang, Mi Kyeong Lee

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Rhus verniciflua is commonly known as a lacquer tree in Korea. Stem barks of R. verniciflua have been used as an immunostimulator in traditional medicine. It contains phenolic compounds and is known for diverse biological activities such as antioxidant and antimicrobial activity. However, it also causes allergic dermatitis due to urushiols derivatives. For the development of active natural resources with less toxicity, the content of phenolic compounds and urushiols of different parts of R. verniciflua such as stem barks, lignum and leaves were quantitated by colorimetric assay and HPLC analysis. The urushiols content were the highest in stem barks, and followed by leaves. The lignum contained trace amount of urushiols. The phenolic contents, however, were the most abundant in lignum, and followed by leaves and stem barks. These results clear showed that the content of urushiols and phenolic differs depending on the parts of R. verniciflua. Antimicrobial activity of different parts of R. verniciflua against fish pathogenic bacteria was also investigated using Edwardsiella tarda. Lignum of R. verniciflua was the most effective in antimicrobial activity against E. tarda and phenolic constituents are suggested to be active constituents for activity. Taken together, phenolic compounds are responsible for antimicrobial activity of R. verniciflua. The lignum of R. verniciflua contains high content of phenolic compounds with less urushiols, which suggests efficient antimicrobial activity with less toxicity. Therefore, lignum of R. verniciflua are suggested as good sources for antimicrobial activity against fish bacterial diseases.

Keywords: different parts, phenolic compounds, Rhus verniciflua, urushiols

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Authors: Mervet A. El-sherif, Ginat M El-sherif, Kadry H Tolba

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The objective of this study was to determine, identify and investigate the effects of natural antioxidants of licorice and carob. Besides, their effects as powder and antioxidant extracts addition on refined sunflower oil stability as natural antioxidants were evaluated. Total polyphenol contents as total phenols, total carotenoids and total tannins were 353.93mg/100g (gallic acid), 10.62mg/100g (carotenoids) and 83.33mg/100g (tannic acid), respectively in licorice, while in carob, it was 186.07, 18.66 and 106.67, respectively. Polyphenol compounds of the studied licorice and carob extracts were determined and identified by HPLC. The stability of refined sunflower oil (which determined by peroxide value and Rancimat) was increased with increasing the level of polyphenols extracts addition. Also, our study shows the effect of addition of these polyphenols extracts to El-mewled El-nabawy sweets fortified by full cream milk powder (sesames and folia). We found that, licorice and carob as powder and polyphenols extracts were delayed the rancidity of sesame and peanut significantly. That encourages using licorice and carob as powder and polyphenols extracts as a good natural antioxidants source instead of synthetic antioxidants.

Keywords: licorice, carob, natural antioxidants, antioxidant activity, applications

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Authors: Sanja Podunavac-Kuzmanović, Strahinja Kovačević, Lidija Jevrić, Evgenija Djurendić, Jovana Ajduković

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There are several methods for determination of the lipophilicity of biologically active compounds, however chromatography has been shown as a very suitable method for this purpose. Chromatographic (C18-RP-HPLC) analysis of a series of 24 17-picolyl and 17-picolinylidene androstane derivatives was carried out. The obtained retention indices (logk, methanol (90%) / water (10%)) were correlated with calculated physicochemical and lipophilicity descriptors. The QSRR analysis was carried out applying principal component regression (PCR) and partial least squares regression (PLS). The PCR and PLS model were selected on the basis of the highest variance and the lowest root mean square error of cross-validation. The obtained PCR and PLS model successfully correlate the calculated molecular descriptors with logk parameter indicating the significance of the lipophilicity of compounds in chromatographic process. On the basis of the obtained results it can be concluded that the obtained logk parameters of the analyzed androstane derivatives can be considered as their chromatographic lipophilicity. These results are the part of the project No. 114-451-347/2015-02, financially supported by the Provincial Secretariat for Science and Technological Development of Vojvodina and CMST COST Action CM1105.

Keywords: androstane derivatives, chromatography, molecular structure, principal component regression, partial least squares regression

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Authors: Vikas Sharma, V. K. Dixit

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Anacyclus pyrethrum (A. pyrethrum) has been used as Vajikaran Rasayana (aphrodisiac) in traditional Indian ayurvedic medicine to treat male sexual dysfunction, including infertility. Aphrodisiac activity may be due to an increase in the production or effect of androgens, so this study sought to evaluate the androgenic and spermatogenic potential of the alkylamide-rich ethanol solution extract. Male Wistar strain rats weighing between 150 and 180 g were completely randomized divided into five groups. The ethanol solution extract of A. pyrethrum was administered to groups of rats in 50, 100, and 150 mg/kg doses for a period of 28 days, and the action was compared with control and testosterone-treated rats. Thirteen N-alkylamides were detected in the extract by using HPLC/UV/electrospray ionization mass spectrometry method. Extract administration at all the doses produced significant increase in body weight, sperm count, motility, and viability along with serum testosterone, luteinizing hormone, and follicle-stimulating hormone concentrations. Histoarchitecture of testis revealed increased spermatogenic activities. Seminal fructose content was also significantly increased after 28 days of treatment. Our results suggest that the ethanol solution extract of the roots of A. pyrethrum has androgenic potential and may improve male fertility by enhancing spermatogenesis.

Keywords: N-alkylamides, testosterone, Anacyclus pyrethrum, androgen

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Authors: Syeda Kiran Shahzadi, Nasir Mahmood, Muhammad Abdul Qadir

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In the current research, three recombinant human interferon alpha-2b proteins (two modified and one normal form) were produced and Pegylated with an aim to produce more effective drugs against viral infections and cancers. The modified recombinant human interferon alpha-2b proteins were produced by site-directed modifications of interferon alpha 2b gene, targeting the amino acids at positions ‘R23’ and ‘H34’. The resulting chemically modified and unmodified forms of human interferon alpha 2b were conjugated with methoxy-polyethylene glycol propanealdehyde (400 KDa) and methoxy-polyethylene glycol succinimidyl succinate (400 KDa). Pegylation of normal and modified forms of Interferon alpha-2b prolong their release time and enhance their efficacy. The conjugation of PEG with modified and unmodified human interferon alpha 2b protein drugs was also characterized with 1H-NMR, HPLC, and SDS-PAGE. Antiproliferative assays of modified and unmodified forms of drugs were performed in cell based bioassays using MDBK cell lines. The results indicated that experimentally produced recombinant human interferon alpha-2b proteins were biologically active and resulted in significant inhibition of cell growth.

Keywords: protein refolding, antiproliferative activities, biomedical applications, human interferon alpha-2b, pegylation, mPEG-propionaldehyde, site directed mutagenesis, E. coli expression

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Authors: Merouani Djilali Redha, F. Abdelmalek, A. A. Addou

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Advanced oxidation processes (AOPs) utilizing Homogenous photocatalysis (Fenton and photo-Fenton reactions), and Heterogeneous photocatalyse (TiO2 and ZnO) were investigated for the degradation of commercial azo dye ‘Orange G’ wastewater. Fenton and photo-Fenton experimental conditions were: Hydrogen peroxide concentration (10-2 M), Ferrous ions concentration (5.10-4 M), pH (2.8 – 3), UV lamp power (6 watt). Adding more ferrous ions enhanced the oxidation rate for the H2O2/Fe2+ and UV/H2O2/Fe2+ processes. The optimum catalyst loading was found 2.0 g.L-1 in our case for both catalysts TiO2 and ZnO. A comparative study of the photocatalytic degradation showed that these two catalysts have a comparable reactivity; it follows a pseudo-first-order kinetics. The degradation trends followed the order: UV365/Fenton > UV365/TiO2 > Solar Fenton > Solar TiO2 > Fenton ~UV365/ZnO. Among AOPs, processes using Fenton type reagent are relatively cheap and easy to operate and maintain. Moreover, UV365/Fenton process has been shown as effective in the treatment of OG dye. Dye was degraded following second-order kinetics. The rate constants was 0,041 .10+6 L.M-1.min-1. The degradation was followed by spectrophotometric method, chemical oxygen demand (COD) measures and high performance liquid chromatography analyses (HPLC). Some aromatic and aliphatic degradation compounds were identified. Degradation of Orange G by UV Fenton mechanism was also proposed.

Keywords: AOPs, homogeneous catalysis, heterogeneous catalysis, acid orange 10, hydroxyl radical

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Authors: Hee-Jae Suh, Mi-Na Hong, Min-Ji Kim, Yeon-Seong Jeong, Ok-Hwan Lee, Jae-Wook Shin, Hyang-Sook Chun, Chan Lee

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Brown HT is a bis-azo dye which is permitted in EU as a food colorant. So far, many studies have focused on HPLC using diode array detection (DAD) analysis for detection of this food colorant with different columns and mobile phases. Even though these methods make it possible to detect Brown HT, low recovery, reproducibility, and linearity are still the major limitations for the application in foods. The purpose of this study was to compare various methods for the analysis of Brown HT and to develop an improved analytical methods including pretreatment. Among tested analysis methods, best resolution of Brown HT was observed when the following solvent was applied as a eluent; solvent A of mobile phase was 0.575g NH4H2PO4, and 0.7g Na2HPO4 in 500mL water added with 500mL methanol. The pH was adjusted using phosphoric acid to pH 6.9 and solvent B was methanol. Major peak for Brown HT appeared at the end of separation, 13.4min after injection. This method exhibited relatively high recovery and reproducibility compared with other methods. LOD (0.284 ppm), LOQ (0.861 ppm), resolution (6.143), and selectivity (1.3) of this method were better than those of ammonium acetate solution method which was most frequently used. Precision and accuracy were verified through inter-day test and intra-day test. Various methods for sample pretreatments were developed for different foods and relatively high recovery over 80% was observed in all case. This method exhibited high resolution and reproducibility of Brown HT compared with other previously reported official methods from FSA and, EU regulation.

Keywords: analytic method, Brown HT, food colorants, pretreatment method

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Authors: Youn Young Shim, Martin J. T. Reaney

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Flaxseed meal is rich in water-soluble gums and, as such, can improve texture in gluten-free products. Flaxseed bioactive-antioxidant peptides, linusorbs (LOs, a.k.a. cyclolinopeptides), are a class of molecules that may contribute health-promoting effects. The effects of dough preparation, baking, and storage on flaxseed-derived LOs stability in doughs and baked products are un-known. Gluten-free (GF) bread dough and bread were prepared with flaxseed meal and the LO content was determined in the flaxseed meal, bread flour containing the flaxseed meal, bread dough, and bread. The LO contents during storage (0, 1, 2, and 4 weeks) at different temperatures (−18 °C, 4 °C, and 22−23 °C) were determined by high-performance liquid chromatog-raphy-diode array detection (HPLC-DAD). The content of oxidized LOs like [1–9-NαC],[1(Rs, Ss)-MetO]-linusorb B2 (LO14) were substantially constant in flaxseed meal and flour produced from flaxseed meal under all conditions for up to 4 weeks. However, during GF-bread production LOs decreased. Due to microbial contamination dough could not be stored at either 4 or 21°C, and bread could only be stored for one week at 21°C. Up to 4 weeks storage was possible for bread and dough at −18 °C and bread at 4 °C without the loss of LOs. The LOs change mostly from processing and less so from storage. The concentration of reduced LOs in flour and meal were much higher than measured in dough and bread. There was not a corre-sponding increase in oxidized LOs. The LOs in flaxseed meal-fortified bread were stable for products stored at low temperatures. This study is the first of the impact of baking conditions on LO content and quality.

Keywords: flaxseed, stability, gluten-free, antioxidant

Procedia PDF Downloads 58

Authors: Swapna Guntupalli, Leela Madhuri Chalasani, Kshatri Jyothi, C. V. Rao, Bondili J. S.

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The study hypothesizes that enhanced biodegradation of Polycyclic Aromatic Hydrocarbons (PAHs) is achievable with an assemblage of microorganisms that are capable of producing biofilm and biosurfactants. Accordingly, PAHs degrading microorganism’s (bacteria, fungi, actinomycetes and yeast) were screened and grouped into different consortia based on their capabilities to produce biofilm and biosurfactants. Among these, Consortium BTSN09 consisting of bacterial fungal cocultures showed highest degradation due to the synergistic action between them. Degradation effiencies were evaluated using HPLC and GC-MS. Within 7days, BTSN09 showed 51% and 50.7% degradation of Phenanthrene (PHE) and Pyrene (PYR) with 200mg/L and 100 mg/L concentrations respectively in a liquid medium. In addition, several degradative enzymes like laccases, 1hydroxy-2-naphthoicacid dioxygenase, 2-carboxybenzaldehyde dehydrogenase, catechol1,2 dioxygenase and catechol2,3 dioxygenase activity was observed during degradation. Degradation metabolites were identified using GC-MS analysis and from the results it was confirmed that the metabolism of degradation proceeds via pthalic acid pathway for both PAHs. Besides, Microbial consortia also demonstrated good biosurfactant production capacity, achieving maximum oil displacement area and emulsification activity of 19.62 cm2, 65.5% in presence of PAHs as sole carbon source. Scanning Electron Microscopy analysis revealed exopolysaccharides (EPS) production, micro and macrocolonies formation with different stages of biofim development in presence of PAHs during degradation.

Keywords: PAHs, biosurfactant, biofilm, biodegradation

Procedia PDF Downloads 549

Authors: Bilal Murtaza, Xiaoyu Li, Liming Dong, Muhammad Kashif Saleemi, Gen Li, Bowen Jin, Lili Wang, Yongping Xu

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Fusarium spp. produce numerous mycotoxins, such as zearalenone (ZEN), deoxynivalenol (DON), and its acetylated compounds, 3-acetyl-deoxynivalenol (3-ADON) and 15-acetyl-deoxynivalenol (15-ADON) (15-ADON). In a co-culture system, the soil-derived Escherichia marmotae strain degrades ZEN and DON into 3-keto-DON and DOM-1 via enzymatic deep-oxidation. When pure mycotoxins were subjected to Escherichia marmotae in culture flasks, degradation, and detoxification were also attained. DON and ZEN concentrations, ambient pH, incubation temperatures, bacterium concentrations, and the impact of acid treatment on degradation were all evaluated. The results of the ELISA and high-performance liquid chromatography-electrospray ionization-high resolution mass spectrometry (HPLC-ESI-HRMS) tests demonstrated that the concentration of mycotoxins exposed to Escherichia marmotae was significantly lower than the control. ZEN levels were reduced by 43.9%, while zearalenone sulfate ([M/z 397.1052 C18H21O8S1) was discovered as a derivative of ZEN converted by microbes to a less toxic molecule. Furthermore, Escherichia marmotae appeared to metabolize DON 35.10% into less toxic derivatives (DOM-1 at m/z 281 of [DON - O]+ and 3-keto-DON at m/z 295 of [DON - 2H]+). These results show that Escherichia marmotae can reduce Fusarium mycotoxins production, degrade pure mycotoxins, and convert them to less harmful compounds, opening up new possibilities for study and innovation in mycotoxin detoxification.

Keywords: mycotoxins, zearalenone, deoxynivalenol, bacterial degradation

Procedia PDF Downloads 55

Authors: Harshani Uggallage, Kapila D. Dissanayaka

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A Sulforhodamine-B assay-guided fractionation on Nigella sativa seeds was conducted to determine the presence of cytotoxic compounds against human hepatoma (HepG2) cells. Initially, a freeze-dried sample of Nigella sativa seeds was sequentially extracted into solvents of increasing polarities. Crude extracts from the sequential extraction of Nigella sativa seeds in chloroform and ethyl acetate showed the highest cytotoxicity. The combined mixture of these two extracts was subjected to bioassay guided fractionation using a modified Kupchan method of partitioning, followed by Sephadex® LH-20 chromatography. This chromatographic separation process resulted in a column fraction with a convincing IC50 (half-maximal inhibitory concentration) value of 13.07µg/ml, which is considerable for developing therapeutic drug leads against human hepatoma. Reversed phase High-Performance Liquid Chromatography (HPLC) was finally conducted for the same column fraction, and the result indicates the presence of one or several main cytotoxic compounds against human HepG2 cells.

Keywords: cytotoxic compounds, half-maximal inhibitory concentration, high-performance liquid chromatography, human HepG2 cells, nigella sativa seeds, Sulforhodamine-B assay

Procedia PDF Downloads 357

Authors: Kyriaki Hatziagapiou, Eleni Kakouri, Konstantinos Bethanis, Alexandra Nikola, Eleni Koniari, Charalabos Kanakis, Elias Christoforides, George Lambrou, Petros Tarantilis

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Medulloblastoma is a highly invasive tumour, as it tends to disseminate throughout the central nervous system early in its course. Despite the high 5-year-survival rate, a significant number of patients demonstrate serious long- or short-term sequelae (e.g., myelosuppression, endocrine dysfunction, cardiotoxicity, neurological deficits and cognitive impairment) and higher mortality rates, unrelated to the initial malignancy itself but rather to the aggressive treatment. A strong rationale exists for the use of Crocus sativus L (saffron) and its bioactive constituents (crocin, crocetin, safranal) as pharmaceutical agents, as they exert significant health-promoting properties. Crocins are water soluble carotenoids. Unlike other carotenoids, crocins are highly water-soluble compounds, with relatively low toxicity as they are not stored in adipose and liver tissues. Crocins have attracted wide attention as promising anti-cancer agents, due to their antioxidant, anti-inflammatory, and immunomodulatory effects, interference with transduction pathways implicated in tumorigenesis, angiogenesis, and metastasis (disruption of mitotic spindle assembly, inhibition of DNA topoisomerases, cell-cycle arrest, apoptosis or cell differentiation) and sensitization of cancer cells to radiotherapy and chemotherapy. The current research aimed to study the potential cytotoxic effect of crocins on TE671 medulloblastoma cell line, which may be useful in the optimization of existing and development of new therapeutic strategies. Crocins were extracted from stigmas of saffron in ultrasonic bath, using petroleum-ether, diethylether and methanol 70%v/v as solvents and the final extract was lyophilized. Identification of crocins according to high-performance liquid chromatography (HPLC) analysis was determined comparing the UV-vis spectra and the retention time (tR) of the peaks with literature data. For the biological assays crocin was diluted to nuclease and protease free water. TE671 cells were incubated with a range of concentrations of crocins (16, 8, 4, 2, 1, 0.5 and 0.25 mg/ml) for 24, 48, 72 and 96 hours. Analysis of cell viability after incubation with crocins was performed with Alamar Blue viability assay. The active ingredient of Alamar Blue, resazurin, is a blue, nontoxic, cell permeable compound virtually nonfluorescent. Upon entering cells, resazurin is reduced to a pink and fluorescent molecule, resorufin. Viable cells continuously convert resazurin to resorufin, generating a quantitative measure of viability. The colour of resorufin was quantified by measuring the absorbance of the solution at 600 nm with a spectrophotometer. HPLC analysis indicated that the most abundant crocins in our extract were trans-crocin-4 and trans-crocin-3. Crocins exerted 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 increased, the reduction of resazurin to resofurin decreased, indicating reduction in cell viability. IC50 values for each time point were calculated ~3.738, 1.725, 0.878 and 0.7566 mg/ml at 24, 48, 72 and 96 hours, respectively. The results of our study could afford the basis of research regarding the use of natural carotenoids as anticancer agents and the shift to targeted therapy with higher efficacy and limited toxicity. Acknowledgements: The research was funded by Fellowships of Excellence for Postgraduate Studies IKY-Siemens Programme.

Keywords: crocetin, crocin, medulloblastoma, saffron

Procedia PDF Downloads 181

Authors: Kubra Sayin, Derya Arslan

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Red pepper (Capsicum annum L.) has long been recognized as a good source of antioxidants, being rich in ascorbic acid and other phytochemicals. In Turkish cuisine red pepper is sometimes consumed raw in salads and baked as a garnish, but its most wide consumption type is red pepper paste. The processing of red pepper into pepper paste includes various thermal treatment steps such as heating and pasteurizing. There are reports demonstrating an enhancement or reduction in antioxidant activity of vegetables after thermal treatment. So this study was conducted to investigate the total phenolics, ascorbic acid and total carotenoids as well as free radical scavenging activity of raw red pepper and various red pepper pastes obtainable on the market. The samples were analyzed for radical-scavenging activity (RSA) and total polyphenol (TP) content using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and Folin-Ciocalteu methods, respectively. They were also evaluated for ascorbic acid content (AsA) by HPLC. Total carotenoids content was determined spectrophotometrically. Results suggest that there is no significant (P > 0.05) difference in RSA, TP, AsA and total carotenoids content between various red pepper paste products. However, red pepper paste showed marked differences (P < 0.05) in the RSA, TP and AsA contents compared with raw red pepper. It is concluded that the red pepper paste, that has a wide range of consumption in Turkish cuisine, presents a good dose of phenolic compounds and antioxidant capacity and it should be regarded as a functional food.

Keywords: red pepper paste, antioxidant properties, total carotenoids, total phenolics

Procedia PDF Downloads 548

Authors: Fatih Özogul, Nurten Toy, Yesim Özogul

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The influences of cell-free solutions (CFSs) of lactic acid bacteria (LAB) on cadaverine and other biogenic amine production by Listeria monocytogenes and Staphylococcus aureus were investigated in lysine decarboxylase broth (LDB) using HPLC. Cell-free solutions were prepared from Lactococcus lactis subsp. lactis, Leuconostoc mesenteroides subsp. cremoris, Pediococcus acidophilus and Streptococcus thermophiles. Two different concentrations that were 50% and 25% CFS and the control without CFSs were prepared. Significant variations on biogenic amine production were observed in the presence of L. monocytogenes and S. aureus (P<0.05). The role of CFS on biogenic amine production by foodborne pathogens varied depending on strains and specific amine. Cadaverine formation in control by L. monocytogenes and S. aureus were 500.9 and 948.1 mg/L, respectively while the CFSs of LAB induced 4-fold lower cadaverine production by L. monocytogenes and 7-fold lower cadaverine production by S. aureus. CFSs resulted in strong decreases in cadaverine and putrescine production by L. monocytogenes and S. aureus, although remarkable increases were observed for histamine, spermidine, spermine, serotonin, dopamine, tyramine, and agmatine, in the presence of LAB in lysine decarboxylase broth.

Keywords: cell-free solution, lactic acid bacteria, cadaverine, food borne-pathogen

Procedia PDF Downloads 505

Authors: Taruna Katyal Arora, Geeta Kumari, Hari Shankar, Neelima Mishra

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Substandard and counterfeit antimalarials is a major problem in malaria endemic areas. The availability of counterfeit/ substandard medicines is not only decreasing the efficacy in patients, but it is also one of the contributing factors for developing antimalarial drug resistance. Owing to this, a pilot study was conducted to survey quality of drugs collected from different malaria endemic areas of India. Artesunate+Sulphadoxine-Pyrimethamine (AS+SP), Artemether-Lumefantrine (AL), Chloroquine (CQ) tablets were randomly picked from public health facilities in selected states of India. The quality of antimalarial drugs from these areas was assessed by using Global Pharma Health Fund Minilab test kit. This includes physical/visual inspection and disintegration test. Thin-layer chromatography (TLC) was carried out for semi-quantitative assessment of active pharmaceutical ingredients. A total of 45 brands, out of which 21 were for CQ, 14 for AL and 10 for AS+SP were tested from Uttar Pradesh (U.P.), Mizoram, Meghalaya and Gujrat states. One out of 45 samples showed variable disintegration and retension factor. The variable disintegration and retention factor which would have been due to substandard quality or other factors including storage. However, HPLC analysis confirms standard active pharmaceutical ingredient, but may be due to humid temperature and moisture in storage may account for the observed result.

Keywords: antimalarial medicines, counterfeit, substandard, TLC

Procedia PDF Downloads 279