Search results for: enzyme hydrolysis

Authors: V. Endo, A. G. Silva Sobrinho, F. A. Almeida, N. L. L. Lima, G. M. Manzi, L. G. A. Cirne, N. M. B. L. Zeola

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We used 24 Ile de France lambs, weighing between 15 and 32 kg (BW). Treatments were supplemented with concentrate: “in nature” sugarcane (IN), sugarcane hydrolyzed using 0.6% calcium oxide (CaO) under aerobic condition (AER), and sugarcane hydrolyzed using 0.6% CaO under anaerobic condition (ANA), constituting a completely randomized design with eight repetitions per treatment. Lambs were housed in individual stalls and fed into the through, allowing 10% of leftovers. Lambs were slaughtered when body weight reached 32 kg. The following parameters were determined on Longissimus lumborum muscle of hot and cold carcasses: pH and color, 45 minutes and 24 hours after slaughtering. Qualitative analysis of the meat were performed in the loins, water-holding capacity (WHC), cooking loss (CL), and shear force (SF). We used a completely randomized design with three treatments and eight repetitions. Means were compared by Tukey test at 5% significance. A higher value for redness (a*) 45 minutes after slaughter (10.48) was found for lambs fed sugarcane hydrolyzed under anaerobic conditions. The other qualitative characteristics of meat were not affected by treatments (P >0.05). The comparison of meat quality resulting from the treatments shows that it is possible to feed in nature sugarcane to lambs, thus waiving hydrolyses process and the spending with alkalizing agent.

Keywords: oxide, hydrolysis, meat quality, pH

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Authors: Omaima A. Sharaf, Wafaa M. Abd El-Rahim, Hassan Moawad, Michael J. Sadowsky

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Textile industry is one of the important industries in the worldwide. It is known that the eco-friendly industrial and agricultural activities are significant for socio-economic stability of all countries. The absence of appropriate industrial waste water treatments is essential barrier for sustainable development in food and agricultural sectors especially in developing country like Egypt. Thus, the development of enzymatic bioremediation technology for textile dye removal will enhance the collaboration between scientists who develop the technology and industry where this technology will be implemented towards the safe disposal of the textile dye wastes. Highly efficient microorganisms are of most importance in developing and using highly effective biological treatment processes. Bacterial degradation of azo dyes is generally initiated by an enzymatic step that involves cleavage of azo linkages, usually with the aid of an azoreductase as electron donor. Thus, expanding the spectrum of microorganisms with high enzymatic activities as azoreductases and discovering novel azo-dye degrading enzymes, with enhanced stability and superior catalytic properties, are necessary for many environmental and industrial applications. Consequently, the use of molecular tools has become increasingly integrated into the understanding of enzyme properties and characterization. Researchers have utilized a gene cloning and expression methods as a tool to produce recombinant protein for decolorizing dyes more efficiently. Thus, presumptive evidence for the presence of genes encoding azoreductases in the genomes of selected local, and most potent azo-degrading strains were obtained by using specific oligonucleotides primers. These potent strains have been isolated from textile industrial wastewater in Egypt and identified using 16S rRNA sequence analysis as 'Lysinibacillus macrolidesB8, Brevibacillus parabrevisB11, Bacillus coagulansB7, and B. cereusB5'. PCR products of two full length genes designated as (AZO1;621bp and AZO2;534bp) were detected. BLASTx results indicated that AZO1 gene was corresponding to predicted azoreductase from of Bacillus sp. ABP14, complete genome, multispecies azoreductase [Bacillus], It was submitted to the gene bank by an accession no., BankIt2085371 AZO1 MG923210 (621bp; 207 amino acids). AZO1 was generated from the DNA of our identified strains Lysinibacillus macrolidesB8. On the other hand, AZO2 gene was corresponding to a predicted azoreductase from Bacillus cereus strain S2-8. Gene bank accession no. was BankIt2085839 AZO2 MG932081 (534bp;178 amino acids) and it was amplified from our Bacillus coagulansB7. Both genes were successfully cloned into pCR2.1TOPO (Invitrogen) and in pET28b+ vectors, then they transformed into E. coli DH5α and BL21(DE3) cells for heterologous expression studies. Our recombinant azoreductases (AZO1&AZO2) exhibited potential enzyme activity and efficiently decolorized an azo dye (Direct violet). They exhibited pH stability between 6 and 8 with optimum temperature up to 60°C and 37 °C after induction by 1mM and 1.5mM IPTG, for both AZO1 &AZO2, respectively. These results suggested that further optimization and purification of these recombinant proteins by using different heterologous expression systems will give great potential for the sustainable utilization of these recombinant enzymes in several industrial applications especially in wastewater treatments.

Keywords: azoreductases, decolorization, enzyme activity, gene cloning and expression

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Authors: Boris Nemzer, Tania Reyes-Izquierdo, Zbigniew Pietrzkowski

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Glucosinolate is a family of glucosides that can be found in a family of brassica vegetables. Upon the damage of the plant, glucosinolate breakdown by an internal enzyme myrosinase (thioglucosidase; EC 3.2.3.1) into isothiocyanates, such as sulforaphane. Sulforaphane is formed by glucoraphanin cleaving the sugar off by myrosinase and rearranged. Sulforaphane nitrile is formed in the same reaction as sulforaphane with the active of epithiospecifier protein (ESP). Most common food processing procedure would break the plant and mix the glucoraphanin and myrosinase together, and the formed sulforaphane would be further degraded. The purpose of this study is to understand the glucoraphanin/sulforaphane and the myrosinase activity of broccoli seeds germinated at a different time and technological processing conditions that keep the activity of the enzyme to form sulforaphane. Broccoli seeds were germinated in the house. Myrosinase activities were tested as the glucose content using glucose assay kit and measured UV-Vis spectrophotometer. Glucosinolates were measured by HPLC/DAD. Sulforaphane was measured using HPLC-DAD and GC/MS. The 6 hr germinated sprouts have a myrosinase activity 32.2 mg glucose/g, which is comparable with 12 and 24 hour germinated seeds and higher than dry seeds. The glucoraphanin content in 6 hour germinated sprouts is 13935 µg/g which is comparable to 24 hour germinated seeds and lower than the dry seeds. GC/MS results show that the amount of sulforaphane is higher than the amount of sulforaphane nitrile in seeds, 6 hour and 24 hour germinated seeds. The ratio of sulforaphane and sulforaphane nitrile is high in 6 hour germinated seeds, which indicates the inactivated ESP in the reaction. After evaluating the results, the short time germinated seeds can be used as the source of glucoraphanin and myrosinase supply to form potential higher sulforaphane content. Broccoli contains glucosinolates, glucoraphanin (4-methylsulfinylbutyl glucosinolate), which is an important metabolite with health-promoting effects. In the pilot clinical study, we observed the effects of a glucosinolates/glucoraphanin-rich extract from short time germinated broccoli seeds on blood adenosine triphosphate (ATP), reactive oxygen species (ROS) and lactate levels. A single dose of 50 mg of broccoli sprouts extract increased blood levels of ATP up to 61% (p=0.0092) during the first 2 hours after the ingestion. Interestingly, this effect was not associated with an increase in blood ROS or lactate. When compared to the placebo group, levels of lactate were reduced by 10% (p=0.006). These results indicate that broccoli germinated seed extract may positively affect the generation of ATP in humans. Due to the preliminary nature of this work and promising results, larger clinical trials are justified.

Keywords: broccoli glucosinolates, glucoraphanin, germinated seeds, myrosinase, adenosine triphosphate

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Authors: Pao-Huei Chen, Shu-Mei Lin, Yih-Ming Weng, Zer-Ran Yu, Be-Jen Wang

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Pancreatic α-amylase and intestinal α-glucosidase are the critical enzymes for the breakdown of complex carbohydrates into di- or mono-saccharide, which play an important role in modulating postprandial blood sugars. Angiotensin converting enzyme (ACE) converts inactive angiotensin-I into active angiotensin-II, which subsequently increase blood pressure through triggering vasoconstriction and aldosterone secretion. Thus, inhibition of carbohydrate-digestion enzymes and ACE will help the management of blood glucose and blood pressure, respectively. Studies showed Pleurotus citrinopileatus (PC), an edible mushroom and commonly cultured in oriental countries, exerted anticancer, immune improving, antioxidative, hypoglycemic and hypolipidemic effects. Previous studies also showed various molecular weights (MW) fractioned from extracts may affect biological activities due to varying contents of bioactive components. Thus, the objective of this study is to investigate the in vitro antioxidant, hypoglycemic and hypotenstive effects and distribution of active compounds of various MWs of cold water extract from P. citrinopileatus (CWEPC). CWEPC was fractioned into four various MW fractions, PC-I (＜1 kDa), PC-II (1-3.5 kDa), PC-III (3.5-10 kDa), and PC-IV (＞10 kDa), using an ultrafiltration system. The physiological activities, including antioxidant activities, the inhibition capabilities of pancreatic α-amylase, intestinal α-glucosidase, and hypertension-linked ACE, and the active components, including polysaccharides, protein, and phenolic contents, of CWEPC and four fractions were determined. The results showed that fractions with lower MW exerted a higher antioxidant activity (p<0.05), which was positively correlated to the levels of total phenols. In contrast, the inhibition effects on the activities of α-amylase, α-glucosidase, and ACE of PC-IV fraction were significantly higher than CWEPC and the other three low MW fractions (< 10 kDa), which was more related to protein contents. The inhibition capability of CWEPC and PC-IV on α-amylase activity was 1/13.4 to 1/2.7 relative to that of acarbose (positive control), respectively. However, the inhibitory ability of PC-IV on α-glucosidase (IC50 = 0.5 mg/mL) was significantly higher than acarbose (IC50 = 1.7 mg/mL). Kinetic data revealed that PC-IV fraction followed a non-competitive inhibition on α-glucosidase activity. In conclusion, the distribution of various bioactive components contribute to the functions of different MW fractions on oxidative stress prevention, and blood pressure and glucose modulation.

Keywords: α-Amylase, angiotensin converting enzyme, α-Glucosidase, Pleurotus citrinopileatus

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Authors: Osama Saber

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Cyanate is one such anion which is produced during protein poisoning in the body and has been studied extensively in the field of biochemistry because of its toxicity. The present work aims at confinement and storage of cyanate in the nano scale. It was achieved through the intercalation of cyanate anions into nanolayerd structures of Ni-Al LDH. In addition, the effect of aging time on the intercalation of cyanate was clarified using X-ray diffraction and scanning electron microscopy. Furthermore, the effect of cations on the affinity towards the intercalation of cyanate anions inside LDH structure was studied by replacement of tetra-valent cations Ti4+ instead of the tri-vallent cations Al3+ during the preparation of LDH structure. X-ray diffraction patterns of the Ni-Ti LDH showed that the interlayer spacing was 0.73 nm. This spacing was smaller than that of Ni-Al LDH suggesting that the interlayered anions into Ni-Ti LDH are different from those into Ni-Al LDH. Thermal analyses (TG, DTG, and DTA) and Infra-red spectra revealed the presence of only cyanate anions into Ni-Ti LDH while, in the case of Ni-Al LDH, both cyanate and carbonate anions were observed. SEM images showed plate-like morphology for both Ni-Ti and Ni-Al LDHs although the shapes of their plates are not similar. Our results suggested that the LDH structures containing titanium cations have higher affinity for cyanate anions than those containing aluminum cations. Therefore, this choice for cyanate in the interlayered spacing widens the applicability to study the effect of the confinement on the toxicity of cyanate by bio researchers.

Keywords: nanolayered structures, Ni-Al LDH, Ni-Ti LDH, intercalation of cyanate anions, urea hydrolysis

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Authors: Hamzeh Alipour, Masoumeh Bagheri, Abbasali Raz, Javad Dadgar Pakdel, Kourosh Azizi, Aboozar Soltani, Mohammad Djaefar Moemenbellah-Fard

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Background: Maggot debridement therapy is an appropriate, effective, and controlled method using sterilized larvae of Luciliasericata (L.sericata) to treat wounds. Netrin-A is an enzyme in the Laminins family which secreted from salivary gland of L.sericata with a central role in neural regeneration and angiogenesis. This study aimed to production of new recombinant Netrin-A protein of Luciliasericata larvae by baculovirus expression vector system (BEVS) in SF9. Material and methods: In the first step, gene structure was subjected to the in silico studies, which were include determination of Antibacterial activity, Prion formation risk, homology modeling, Molecular docking analysis, and Optimization of recombinant protein. In the second step, the Netrin-A gene was cloned and amplified in pTG19 vector. After digestion with BamH1 and EcoR1 restriction enzymes, it was cloned in pFastBac HTA vector. It was then transformed into DH10Bac competent cells, and the recombinant Bacmid was subsequently transfected into insect Sf9 cells. The expressed recombinant Netrin-A was thus purified in the Ni-NTA agarose. This protein evaluation was done using SDS-PAGE and western blot, respectively. Finally, its concentration was calculated with the Bradford assay method. Results: The Bacmid vector structure with Netrin-A was successfully constructed and then expressed as Netrin-A protein in the Sf9 cell lane. The molecular weight of this protein was 52 kDa with 404 amino acids. In the in silico studies, fortunately, we predicted that recombinant LSNetrin-A have Antibacterial activity and without any prion formation risk.This molecule hasa high binding affinity to the Neogenin and a lower affinity to the DCC-specific receptors. Signal peptide located between amino acids 24 and 25. The concentration of Netrin-A recombinant protein was calculated to be 48.8 μg/ml. it was confirmed that the characterized gene in our previous study codes L. sericata Netrin-A enzyme. Conclusions: Successful generation of the recombinant Netrin-A, a secreted protein in L.sericata salivary glands, and because Luciliasericata larvae are used in larval therapy. Therefore, the findings of the present study could be useful to researchers in future studies on wound healing.

Keywords: blowfly, BEVS, gene, immature insect, recombinant protein, Sf9

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Authors: Nuha Mansour Alhazmi, Magda Mohamed Aly, Fardus M. Bokhari, Ahmed Bahieldin, Sherif Edris

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Out of 30 fungal isolates, 2 new isolates were proven to degrade poly-β-hydroxybutyrate (PHB). Enzyme assay for these isolates indicated the optimal environmental conditions required for depolymerase enzyme to induce the highest level of biopolymer degradation. The two isolates were basically characterized at the morphological level as Trichoderma asperellum (isolate S1), and Aspergillus fumigates (isolate S2) using standard approaches. The aim of the present study was to characterize these two isolates at the molecular level based on the highly diverged rRNA gene(s). Within this gene, two domains of the ribosome large subunit (LSU) namely internal transcribed spacer (ITS) and 26S were utilized in the analysis. The first domain comprises the ITS1/5.8S/ITS2 regions ( > 500 bp), while the second domain comprises the D1/D2/D3 regions ( > 1200 bp). Sanger sequencing was conducted at Macrogen (Inc.) for the two isolates using primers ITS1/ITS4 for the first domain, while primers LROR/LR7 for the second domain. Sizes of the first domain ranged between 594-602 bp for S1 isolate and 581-594 bp for S2 isolate, while those of the second domain ranged between 1228-1238 bp for S1 isolate and 1156-1291 for S2 isolate. BLAST analysis indicated 99% identities of the first domain of S1 isolate with T. asperellum isolates XP22 (ID: KX664456.1), CTCCSJ-G-HB40564 (ID: KY750349.1), CTCCSJ-F-ZY40590 (ID: KY750362.1) and TV (ID: KU341015.1). BLAST of the first domain of S2 isolate indicated 100% identities with A. fumigatus isolate YNCA0338 (ID: KP068684.1) and strain MEF-Cr-6 (ID: KU597198.1), while 99% identities with A. fumigatus isolate CCA101 (ID: KT877346.1) and strain CD1621 (ID: JX092088.1). Large numbers of other T. asperellum and A. fumigatus isolates and strains showed high level of identities with S1 and S2 isolates, respectively, based on the diversity of the first domain. BLAST of the second domain of S1 isolate indicated 99 and 100% identities with only two strains of T. asperellum namely TR 3 (ID: HM466685.1) and G (ID: KF723005.1), respectively. However, other T. species (ex., atroviride, hamatum, deliquescens, harzianum, etc.) also showed high level of identities. BLAST of the second domain of S2 isolate indicated 100% identities with A. fumigatus isolate YNCA0338 (ID: KP068684.1) and strain MEF-Cr-6 (ID: KU597198.1), while 99% identities with A. fumigatus isolate CCA101 (ID: KT877346.1) and strain CD1621 (ID: JX092088.1). Large numbers of other A. fumigatus isolates and strains showed high level of identities with S2 isolate. Overall, the results of molecular characterization based on rRNA diversity for the two isolates of T. asperellum and A. fumigatus matched those obtained by morphological characterization. In addition, ITS domain proved to be more sensitive than 26S domain in diversity profiling of fungi at the species level.

Keywords: Aspergillus fumigates, Trichoderma asperellum, PHB, degradation, BLAST, ITS, 26S, rRNA

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Authors: Boukli Hacene Faiza, Soufi Wassila, Ghalem Said

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The oral antidiabetics drugs such as alpha glucosidase inhibitors present undesirable effects like acarbose. Flavonoids are class of molecules widely distributed in plants, for this reason we are interested in our work to study the inhibition in silico of alpha glucosidase by natural ligands ( flavonoids analogues) using molecular modeling methods using MOE (Molecular Operating Environment) software to predict their interaction with this enzyme with score energy, ADME /T tests and druglikeness properties experiments. Two flavonoids Beicalein and Apigenin have high binding affinity with alpha glucosidase with lower IC50 supposed potent inhibitors.

Keywords: alpha glucosidase, flavonoides analogues, drug research, molecular modeling

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Authors: Deokyeong Choe, Sung Hun Youn, Younggon Kim, Chul Soo Shin

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L-Cysteine is extensively used as a supplement of pharmaceuticals, cosmetics, food and feed additives. It has obtained industrially by hydrolysis of human hair and poultry feathers. However, there are some problems such as the restriction of using materials from animals and the intractable waste pollution. The enzymatic conversion has been regarded as an environmental-friendly method. Currently, the biggest bottle-neck of enzymatic conversion is the low yield of L-cysteine due to the low substrate solubility. In this study, the method of enhancing the solubility of the substrate D,L-2-amino-Δ2-thiazoline-4-carboxylicacid (ATC) was developed and the enzymatic reaction at high concentration levels was performed. A large amount of substrate in aqueous solutions was dissolved by pH control using salts. As the pH of the solution increased, the solubility of ATC increased. It was thought that a shift of ATC from acid form (-COOH) to dissociated carboxylic group (-COO-) would improve its hydrophilicity leading to solubility increase. The highest solubility of ATC was 610 mM at pH 10.5, whereas the maximum reaction rate was obtained at pH 8.3. As a result, a high L-cysteine yield of 250 mM was achieved at pH 9.1, which was obtained from a combination of optimum pH conditions for ATC solubility and enzymatic conversion. This yield corresponds to approximately 18 times of that in previous reports.

Keywords: D, L-2-amino-Δ2-thiazoline-4-carboxylicacid, enzymatic conversion, high-substrate solubilization, L-Cysteine

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Authors: A. Altomi, A. Alhebshi, M. Rasm, B. Osman

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This study explored the application of chitin nanocrystals (ChiNCs), derived from a renewable and environmentally friendly material, as stabilizers for oil-in-water (O/W) emulsions. O/W emulsions are commonly used in various applications but are prone to instability and degradation over time. Instability can occur due to factors such as flocculation, coalescence, and gravitational separation, including creaming and sedimentation, either independently or simultaneously. To produce ChiNCs, chitin powder underwent acid hydrolysis. Transmission electron microscopy (TEM) analysis revealed that ChiNCs exhibited a needle-like morphology, with lengths ranging from 200 to 800 nm and widths ranging from 20 to 80 nm. The surface charge of ChiNCs was negative at pH values above 7 and positive at pH values below 7. The rheological properties of O/W emulsions stabilized by ChiNCs were compared to those stabilized by synthetic surfactants, namely Tween 80 and CTAB. The emulsions stabilized by ChiNCs demonstrated higher yield stress and lower shear viscosity compared to those stabilized by synthetic surfactants. This indicates that ChiNC-stabilized emulsions are more stable and less prone to breakdown. Based on these findings, ChiNCs show promise as an alternative to synthetic surfactants for stabilizing O/W emulsions.

Keywords: chitin nanocrystals, colloidal pickering, emulsion rheology, oil-in-water, synthetic surfactant

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Authors: Laura Boschis, Elena D. Ozzello, Enzo Mastromatteo

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Point of care diagnostic finds growing interest in medicine and agri-food because of faster intervention and prevention. EliChip is a microfluidic platform to perform Point of Care immunoenzymatic assay based on ready-to-use kits and a portable instrument to manage fluidics and read reliable quantitative results. Thanks to miniaturization, analyses are faster and more sensible than conventional ELISA. EliChip is one of the crucial assets of the Europen-founded Flamingo project for in-line measuring inflammatory markers.

Keywords: lab on chip, point of care, immunoenzymatic analysis, synovial arthritis

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Authors: Pauline Nyssen, Ange Mouithys-Mickalad, Maryse Hoebeke

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Inflammation is a complex physiological phenomenon involving chemical and enzymatic mechanisms. Polymorphonuclear neutrophil leukocytes (PMNs) play an important role by producing reactive oxygen species (ROS) and releasing myeloperoxidase (MPO), a pro-oxidant enzyme. Released both in the phagolysosome and the extracellular medium, MPO produces during its peroxidase and halogenation cycles oxidant species, including hypochlorous acid, involved in the destruction of pathogen agents, like bacteria or viruses. Inflammatory pathologies, like rheumatoid arthritis, atherosclerosis induce an excessive stimulation of the PMNs and, therefore, an uncontrolled release of ROS and MPO in the extracellular medium, causing severe damages to the surrounding tissues and biomolecules such as proteins, lipids, and DNA. The treatment of chronic inflammatory pathologies remains a challenge. For many years, MPO has been used as a target for the development of effective treatments. Numerous studies have been focused on the design of new drugs presenting more efficient MPO inhibitory properties. However, some designed inhibitors can be toxic. An alternative consists of assessing the potential inhibitory action of clinically-known molecules, having antioxidant activity. Propofol, 2,6-diisopropyl phenol, which is used as an intravenous anesthetic agent, meets these requirements. Besides its anesthetic action employed to induce a sedative state during surgery or in intensive care units, propofol and its injectable form Diprivan indeed present antioxidant properties and act as ROS and free radical scavengers. A study has also evidenced the ability of propofol to inhibit the formation of the neutrophil extracellular traps fibers, which are important to trap pathogen microorganisms during the inflammation process. The aim of this study was to investigate the potential inhibitory action mechanism of propofol and Diprivan on MPO activity. To go into the anti-inflammatory action of propofol in-depth, two of its oxidative derivatives, 2,6-diisopropyl-1,4-p-benzoquinone (PPFQ) and 3,5,3’,5’-tetra isopropyl-(4,4’)-diphenoquinone (PPFDQ), were studied regarding their inhibitory action. Specific immunological extraction followed by enzyme detection (SIEFED) and molecular modeling have evidenced the low anti-catalytic action of propofol. Stopped-flow absorption spectroscopy and direct MPO activity analysis have proved that propofol acts as a reversible MPO inhibitor by interacting as a reductive substrate in the peroxidase cycle and promoting the accumulation of redox compound II. Overall, Diprivan exhibited a weaker inhibitory action than the active molecule propofol. In contrast, PPFQ seemed to bind and obstruct the enzyme active site, preventing the trigger of the MPO oxidant cycles. PPFQ induced a better chlorination cycle inhibition at basic and neutral pH in comparison to propofol. PPFDQ did not show any MPO inhibition activity. The three interest molecules have also demonstrated their inhibition ability on an important step of the inflammation pathway, the PMNs superoxide anions production, thanks to EPR spectroscopy and chemiluminescence. In conclusion, propofol presents an interesting immunomodulatory activity by acting as a reductive substrate in the peroxidase cycle of MPO, slowing down its activity, whereas PPFQ acts more as an anti-catalytic substrate. Although PPFDQ has no impact on MPO, it can act on the inflammation process by inhibiting the superoxide anions production by PMNs.

Keywords: Diprivan, inhibitor, myeloperoxidase, propofol, spectroscopy

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Authors: H. Djebar, L. Khene, M. Boucenna, M. R. Djebar, M. N. Khebbeb, M. Djekoun

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Currently in toxicology, use of alternative models permits to understand the mechanisms of toxicity at different levels of cells. Objectives of our research concern the determination of NPs ZnO, TiO2, AlO2, and FeO2 effect on ciliate protist freshwater Paramecium sp and Helix aspersa. The result obtained show that NPs increased antioxidative enzyme activity like catalase, glutathione –S-transferase and level GSH. Also, cells treated with high concentrations of NPs showed a high level of MDA. In conclusion, observations from growth and enzymatic parameters suggest on one hand that treatment with NPs provokes an oxidative stress and on the other that snale and paramecium are excellent alternatives models for ecotoxicological studies.

Keywords: NPs, GST, catalase, GSH, MDA, toxicity, snale and paramecium

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Authors: Imran Muhammad

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The main cause of infectious disease in the modern world is Mycobacterium Tuberculosis (MT). To combat tuberculosis, new and efficient drugs are an urgent need in the modern world. Schif bases are potent for their biological pharmacophore activity. Thus we selected different Vanillin-based Schiff bases for their binding activity against target enzymes of Mycobacterium tuberculosis that is (DprE1 (decaprenyl phosphoryl-β-D-ribose 2′-epimerase), and DNA gyrase subunit-A), using molecular docking. We evaluate the inhibition potential, interaction, and binding mode of these compounds with the target enzymes.

Keywords: schiff bases, tuberculosis, DNA gyrase, DprE1, docking

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Authors: Rabih O. Al-Kaysi, Lingyan Zhu, Muhannah K. Al-Muhannah, Christopher J. Bardeen

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(E)-3-(Anthracen-9-yl)acrylic acid (9-AYAA) 1 exhibits a strong photomechanical response in bulk crystals but is challenging to grow in microcrystalline form. High quality microcrystals of this molecule could not be grown using techniques like sublimation, reprecipitation, and the floating drop method. If the tertbutyl ester of 9-AYAA is used as a starting material, however, high quality, size-uniform microwires could be grown via acid catalyzed hydrolysis. 9-AYAA microwires with uniform length and thickness were produced after a suspension of (E)-tert-butyl 3-(anthracen-9-yl)acrylate ester 2 microparticles was tumble-mixed in a mixture of phosphoric acid and sodium dodecyl sulfate at 35 °C. The dependence of the results on temperature, surfactant and precursor concentration, and mixing mode was investigated. This chemical reaction-growth method was extended to grow microplates of 9-anthraldehyde 3 using the corresponding acylal 4 as the starting material. Under 475 nm irradiation, the 9-AYAA microwires undergo a photoinduced coiling–uncoiling transition, while the 9-anthraldehyde microplates undergo a folding–unfolding transition.

Keywords: photomechanical, surfactant, organic crystals, uniform

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Authors: Pedro Maldonado-Alvarado

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An important cassava quality production was detected in Ecuador. However, in this country, few products with low adding-value are produced from the tuber and none from cassava by-products. To our best knowledge, lactic acid was produced from Ecuadorian cassava bagasse starch in a biotechnological way. The objective of this contribution was to study the influence of the fermentation variables (pH and agitation) on the lactic acid production of Ecuadorian cassava varieties from bagasse starch. Enzymatic hydrolysis of cassava bagasse starch for INIAP 650 and INIAP 651 varieties spread in Ecuador was performed using α-amylase and amyloglucosidase. Then, glucose was fermented by Lactobacillus leichmannii strains in different conditions of agitation (0 and 150 rpm) and pH (4.5, 5.0, and 5.5). Significant differences in ash, fibre, protein, lipids, and amylose were found in cassava bagasse starch of INIAP 650 and INIAP 651 with 1.4 and 1.3%, 4.3 and 6%, 1.2 and 2.1%, 1.9 and 1.5%, and 24.3 and 26.5%, respectively. The determination of lactic acid was performed by potentiometric and FTIR analysis. Conversions of cassava bagasse to reduced sugars were 71.7 and 85.1% for INIAP 650 and INIAP 651, respectively. The best lactic acid concentrations were 27.6 and 33.5 g/L, obtained at agitation 150 rpm and pH 5.5 for INIAP 650 and INIAP 651. Qualitative analysis conducted by FTIR spectrophotometry confirmed the presence of lactic acid in the reacted products. This investigation could contribute to the valorisation of residues from promising cassava varieties in Ecuador and hence to increase the development of this country.

Keywords: bagasse starch, cassava, Ecuador, fermentation, lactic acid

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Authors: Siti Nur Syazni Mohd Zuki, Tan Ling Ling, Nina Suhaity Azmi, Chong Kwok Feng, Lee Yook Heng

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Nitrite (NO2-) ion is used prevalently as a preservative in processed meat. Elevated levels of nitrite also found in edible bird’s nests (EBNs). Consumption of NO2- ion at levels above the health-based risk may cause cancer in humans. Spectrophotometric Griess test is the simplest established standard method for NO2- ion detection, however, it requires careful control of pH of each reaction step and susceptible to strong oxidants and dyeing interferences. Other traditional methods rely on the use of laboratory-scale instruments such as GC-MS, HPLC and ion chromatography, which cannot give real-time response. Therefore, it is of significant need for devices capable of measuring nitrite concentration in-situ, rapidly and without reagents, sample pretreatment or extraction step. Herein, we constructed a microspheres-based microbial optode for visual quantitation of NO2- ion. Raoutella planticola, the bacterium expressing NAD(P)H nitrite reductase (NiR) enzyme has been successfully extracted by microbial technique from EBN collected from local birdhouse. The whole cells and the lipophilic Nile Blue chromoionophore were physically absorbed on the photocurable poly(n-butyl acrylate-N-acryloxysuccinimide) [poly (nBA-NAS)] microspheres, whilst the reduced coenzyme NAD(P)H was covalently immobilized on the succinimide-functionalized acrylic microspheres to produce a reagentless biosensing system. Upon the NiR enzyme catalyzes the oxidation of NAD(P)H to NAD(P)+, NO2- ion is reduced to ammonium hydroxide, and that a colour change from blue to pink of the immobilized Nile Blue chromoionophore is perceived as a result of deprotonation reaction increasing the local pH in the microspheres membrane. The microspheres-based optosensor was optimized with a reflectance spectrophotometer at 639 nm and pH 8. The resulting microbial bio-optode membrane could quantify NO2- ion at 0.1 ppm and had a linear response up to 400 ppm. Due to the large surface area to mass ratio of the acrylic microspheres, it allows efficient solid state diffusional mass transfer of the substrate to the bio-recognition phase, and achieve the steady state response as fast as 5 min. The proposed optical microbial biosensor requires no sample pre-treatment step and possesses high stability as the whole cell biocatalyst provides protection to the enzymes from interfering substances, hence it is suitable for measurements in contaminated samples.

Keywords: acrylic microspheres, microbial bio-optode, nitrite ion, reflectometric

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Authors: Hiral D. Trivedi, Dinesh S. Patel, Sachin P. Shukla

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We have immobilized alcohol dehydrogenase on k-carrageenan, which is a natural polysaccharide obtained from seaweeds by entrapment and on copolymer of acrylamide and 2-hydroxy ethylmethaacrylate by covalent coupling. We have optimized all the immobilization parameters and also carried the comparison studies of both. In case of copolymer of acrylamide and 2-hydroxy ethylmethaacrylate, we have activated both the amino and hydroxyl group individually and simultaneously using different activating agents and obtained some interesting results. We have found that covalently bound enzyme was found to be better under all tested conditions. The reaction on ethanol was carried out using these immobilized systems.

Keywords: alcohol dehydrogenase, acrylamide-co-2-hydroxy ethylmethaacrylate, ethanol, k-carrageenan

Procedia PDF Downloads 117

Authors: Friday A. Ogori, Ojotu M. Eke, Oneh J. Abu, Abraham T. Girgih

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B.aeqygtiaca del (Balanites aegyptiaca del) seed protein concentrate (APC) was hydrolyzed using different enzymes such as pepsin+pancreatin (PP), Alcalase (Alca), and Flavourzyme (Flav). The Alca has higher yield (100%) when compared to PP (83.23%) and Flav (62.90%). The hydrophobic amino acid and Sulphur containing amino acid (40.19%, 7.04%) in PP hydrolysate were higher compared to Alcalase (38.92%, 6.69%), Flavourenzyme (37.43%,6.35%), and APC (39.97%, 6.95%) samples. The PP has stronger DPPH, Hydroxyl radical quenching, Ferric reducing activity, and linoleic acid peroxidation activity, followed by the protein concentrate (APC) and Alcalase (Alca), while Flavourenzyme (Flav) derived hydrolysate was least in scavenging and inhibiting radical peroxidation properties. Flavourenzyme derived hydrolysate had stronger Ferric reducing antioxidant potential and metal chelating property. The result showed that the Alcalase hydrolysate has promising peptide yield, and PP hydrolysate had excellent amino acid residues and good in-vitro antioxidant potentials and could be a preferred ingredients in the nutraceutical and functional food emerging industries.

Keywords: balanites aegyptiaca del, protein concentrate, protein hydrolysates, enzymatic hydrolysis, antioxidants

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Authors: Khalid Elbadawi Elshafea, Mahmoud Hassan Onsa

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This paper presents the results of the experiments to measure the impact of pasteurization cows dung on important parameter of anaerobic digestion (retention time) and measure the effect in daily production rate of biogas, were used local materials in these experiments, two experiments were carried out in two bio-digesters (1 and 2) (18.0 L), volume of the mixture 16.0-litre and the mass of dry matter in the mixture 4.0 Kg of cow dung. Pasteurization process has been conducted on the mixture into the digester 2, and put two digesters under room temperature. Digester (1) produced 268.5 liter of methane in period of 49 days with daily methane production rate 1.37L/Kg/day, and digester (2) produced 302.7-liter of methane in period of 26 days with daily methane production rate 2.91 L/Kg/day. This study concluded that the use of system pasteurization cows dung speed up hydrolysis in anaerobic process, because heat to certain temperature in certain time lead to speed up chemical reactions (transfer Protein to Amino acids, Carbohydrate to Sugars and Fat to Long chain fatty acids), this lead to reduce the retention time an therefore increase the daily methane production rate with 212%.

Keywords: methane, cow dung, daily production, pasteurization, increase

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Authors: O. Mowla, E. Kennedy, M. Stockenhuber

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Finding alternative renewable resources of energy has attracted the attentions in consequence of limitation of the traditional fossil fuel resources, increasing of crude oil price and environmental concern over greenhouse gas emissions. Biodiesel (or Fatty Acid Methyl Esters (FAME)), an alternative energy source, is synthesised from renewable sources such as vegetable oils and animal fats and can be produced from waste oils. FAME can be produced via hydroesterification of oils. The process involves two stages. In the first stage of this process, fatty acids and glycerol are being obtained by hydrolysis of the feed stock oil. In the second stage, the recovered fatty acids are then esterified with an alcohol to methyl esters. The presence of a catalyst accelerates the rate of the hydroesterification reaction of oils. The overarching aim of this study is to find the effect of using zeolite as a catalyst in the heterogeneous hydroesterification of soybean oil. Both stages of the catalytic hydroesterification of soybean oil had been conducted at atmospheric and high-pressure conditions using reflux glass reactor and Parr reactor, respectively. The effect of operating parameters such as temperature and reaction time on the overall yield of biodiesel formation was also investigated.

Keywords: biodiesel, heterogeneous catalytic hydroesterification, soybean oil, zeolite

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Authors: Ž. Vaštag, Lj. Popović, S. Popović

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After cold pressing of pumpkin oil, the defatted oil cake (PUOC) was utilized as raw material for processing of bio-functional hydrolysates. In this study, the in vitro bioactivity of an alcalase (AH) and a pepsin hydrolysate (PH) prepared from the major pumpkin 12S globulin (cucurbitin) are compared. The hydrolysates were produced at optimum reaction conditions (temperature, pH) for the enzymes, during 60min. The bioactivity testing included antioxidant and angiotensin I converting enzyme inhibitory activity assays. The hydrolysates showed high potential as natural antioxidants and possibly antihypertensive agents in functional food or nutraceuticals. Additionally, preliminary studies have shown that both hydrolysates could exhibit modest α-amylase inhibitory activity, which indicates on their hypoglycemic potential.

Keywords: cucurbitin, alcalase, pepsin, protein hydrolysates, in vitro bioactivity

Procedia PDF Downloads 283

Authors: Kiran Fatima, Kashif Ali

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Staphylococcus aureus is an opportunistic human as well as animal pathogen that causes a variety of diseases. A total of 70 staphylococci isolates were obtained from soil, water, yogurt, and clinical samples. The likely staphylococci clinical isolates were identified phenotypically by different biochemical tests. Molecular identification was done by PCR using species-specific 16S rRNA primer pairs, and finally, 50 isolates were found to be positive as Staphylococcus aureus, sciuri, xylous and cohnii. Screened isolates were further analyzed by several microbiological diagnostics tests, including gram staining, coagulase, capsule, hemolysis, fermentation of glucose, lactose, maltose, and sucrose tests enzymatic reactions. It was found that 78%, 81%, and 51% of isolates were positive for gelatin hydrolysis, protease, and lipase activities, respectively. Antibiogram analysis of isolated Staphylococcus aureus strains with respect to different antimicrobial agents revealed resistance patterns ranging from 57 to 96%. Our study also shows 70% of strains to be MRSA, 54.3% as VRSA, and 54.3% as both MRSA and VRSA. All the identified isolates were subjected to detection of mecA, nuc, and hlb genes, and 70%, 84%, and 40% were found to harbour mecA, nuc, and hlb genes, respectively. The current investigation is highly important and informative for the high-level multidrug-resistant Staphylococcus aureus infections inclusive also of methicillin and vancomycin.

Keywords: MRSA, VRSA, mecA, MSSA

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Authors: Dipti Yadav, Latha Rangan, Pinakeswar Mahanta

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Recent fuel-development strategies to reduce oil dependency, mitigate greenhouse gas emissions, and utilize domestic resources have generated interest in the search for alternative sources of fuel supplies. Bioenergy production from lignocellulosic biomass has a great potential. Cellulose, hemicellulose and Lignin are main constituent of woods or agrowaste. In all the industries there are always left over or waste products mainly lignin, due to the heterogeneous nature of wood and pulp fibers and the heterogeneity that exists between individual fibers, no method is currently available for the quantitative isolation of native or residual lignin without the risk of structural changes during the isolation. The potential benefits from finding alternative uses of lignin are extensive, and with a double effect. Lignin can be used to replace fossil-based raw materials in a wide range of products, from plastics to individual chemical products, activated carbon, motor fuels and carbon fibers. Furthermore, if there is a market for lignin for such value-added products, the mills will also have an additional economic incentive to take measures for higher energy efficiency. In this study residual lignin were isolated from areca nut shells by acid hydrolysis and were analyzed and characterized by Fourier Transform Infrared (FTIR), LCMS and complexity of its structure investigated by NMR.

Keywords: Areca nut, Lignin, wood, bioenergy

Procedia PDF Downloads 450

Authors: M. Mydlárová Blaščáková, J. Poráčová, Z. Tomková, Ľ. Blaščáková, M. Nagy, M. Konečná, E. Petrejčíková, Z. Gogaľová, V. Sedlák, J. Mydlár, M. Zahatňanská, K. Hricová

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Osteoporosis is a multifactorial disease that results in reduced quality of life, causes decreased bone strength, and changes in their microarchitecture. Mostly postmenopausal women are at risk. In our study, we measured anthropometric parameters of postmenopausal women (104 women of control group – CG and 105 women of osteoporotic group - OG) and determined TSH hormone levels and PTH as well as mineral elements - Ca, P, Mg and enzyme alkaline phosphatase. Through the correlation analysis in CG, we have found association based on age and BMI, P and Ca, as well as Mg and Ca; in OG we determined interdependence based on an association of age and BMI, age and Ca. Using the Student's t test, we found significantly important differences in biochemical parameters of Mg (p ˂ 0,001) and TSH (p ˂ 0,05) between CG and OG.

Keywords: factors, bone mass density, Central Europe, biomarkers

Procedia PDF Downloads 165

Authors: N. Adjlane, N. Haddad

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The American foulbrood is one of the most serious diseases that may affect brood of larvae and pupae stages. The causative organism is a gram positive bacterium Paaenibacillus larvae. American foulbrood infected apiaries suffer from severe economic losses, resulting from significant decreases in honeybee populations and honey production. The aim of this study was to detect Paenibacillus larvae in the remains collected at the bottom of the hive from the suspected hives by direct PCR and culture growth. A total of 56 suspected beehive wax debris samples collected in 40 different apiaries located in the central region of Algeria. MYPGP the culture medium is used during all the identifications of the bacterium. After positive results on samples, biochemical confirmation tests (test of catalase, presence hydrolysis of casein) and microscopic (gram stain) are used in order to verify the accuracy of the initial results. The QIAamp DNA Mini Kit is used to identify the DNA of Paaenibacillus larvae. Paaenibacillus larvae were identified in 14 samples out of 16 by the PCR. A suspected culture-negative sample was found positive through evaluation with PCR. This research is for the bacterium Paaenibacillus larvae in the debris of the colony is an effective method for diagnosis of the pathology of American foulbrood.

Keywords: Paenibacillus larvae, honeybee, PCR, microbiological method

Procedia PDF Downloads 381

Authors: Sonia Tamanna, Akramul Hassan, Mohammad Shakil Mahmood, Farzana Ansari, Gowhar Rashid, Mir Fahim Faisal, M. Zakir Hossain Howlader

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Background: Preeclampsia (PE), a pregnancy-specific hypertensive disorder, significantly impacts maternal and fetal health. It is particularly prevalent in underdeveloped countries and is linked to preterm delivery and fetal growth. The renin-angiotensin system (RAS) plays a crucial role in ensuring a successful pregnancy outcome, with Angiotensin-Converting Enzyme 2 (ACE2) being a key component. ACE2 converts ANG II to Ang-(1-7), offering protection against ANG II-induced stress and inflammation while regulating blood pressure and osmotic balance during pregnancy. The reduced maternal plasma angiotensin-converting enzyme 2 (ACE2) seen in preeclampsia might contribute to its pathogenesis. However, there has been a dearth of comprehensive research into the association between ACE2 gene polymorphism and preeclampsia. In the South Asian population, hypertension is strongly linked to two SNPs: rs2285666 and rs879922. This genotype was therefore considered, and the possible association of maternal and fetal ACE2 gene polymorphism with preeclampsia within the Bangladeshi population was evaluated. Method: DNA was extracted from peripheral white blood cells (WBCs) using the organic method, and SNP genotyping was done via PCR-RFLP. Odds ratios (OR) with 95% confidence intervals (95% CI) were calculated using logistic regression to determine relative risk. Result: A comprehensive case-control study was conducted on 51 PE patients and their infants, along with 56 control subjects and their infants. Maternal single nuvleotide polymorphisms (SNP) (rs2285666) analysis revealed a strong association between the TT genotype and preeclampsia, with a four-fold increased risk in mothers (P=0.024, OR=4.00, 95% CI=1.36-11.37) compared to their ancestral genotype CC. However, the CT genotype (rs2285666) showed no significant difference (P=0.46, OR=1.54, 95% CI=0.57-4.14). Notably, no significant correlation was found in infants, regardless of their gender. For rs879922, no significant association was observed in both mothers and infants. This pioneering study suggests that mothers carrying the ACE2 gene variant rs2285666 (TT allele) may be at higher risk for preeclampsia, potentially influencing hypertension characteristics, whereas rs879922 does not appear to be associated with developing preeclampsia. Conclusion: This study sheds light on the role of ACE2 gene polymorphism, particularly the rs2285666 TT allele, in maternal susceptibility to preeclampsia. However, rs879922 does not appear to be linked to the risk of PE. This research contributes to our understanding of the genetic underpinnings of preeclampsia, offering insights into potential avenues for prevention and management.

Keywords: ACE2, PCR-RFLP, preeclampsia, single nuvleotide polymorphisms (SNPs)

Procedia PDF Downloads 34

Authors: P. A. Broodryk, A. F. Van Der Merwe, H. W. J. P. Neomagus

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The production of the clear liquid fertilizer ammonium polyphosphate (APP) is best achieved by the pyrolysis of urea phosphate, as it produces a product that is free from any of the impurities present in the raw phosphoric acid it was made from. This is a multiphase, multi-step reaction that produces carbon dioxide and ammonia as gasses and ammonium polyphosphate as liquid products. The polyphosphate chain length affects the solubility and thus the applicability of the product as liquid fertiliser, thus proper control of the reaction conditions is thus required for the use of this reaction in the production of fertilisers. This study investigates the reaction kinetics of the aforementioned reaction, describing a mathematical model for the kinetics of the reaction along with the accompanying rate constants. The reaction is initially exothermic, producing only carbon dioxide as a gas product and ammonium diphosphate, at higher temperatures the reaction becomes endothermic, producing ammonia gas as an additional by-product and longer chain polyphosphates, which when condensed too far becomes highly water insoluble. The aim of this study was to (i) characterise the pyrolysis reaction of urea phosphate by determining the mechanisms and the associated kinetic constants, and (ii) to determine the optimum conditions for ammonium diphosphate production. A qualitative investigation was also done to find the rate of hydrolysis of APP as this provides an estimate of the shelf life of an APP clear liquid fertiliser solution.

Keywords: ammonium polyphosphate, kinetics, pyrolysis, urea phosphate

Procedia PDF Downloads 126

Authors: Y. Kobayashi, T. Kurosaka, K. Yamamura, T. Yonezawa, K. Yamasaki

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The present work reports an effect of surface- modification of indium tin oxide (ITO) particles with chemicals on their electronic conductivity properties. Examined chemicals were polyvinyl alcohol (nonionic polymer), poly(diallyl dimethyl ammonium chloride) (cationic polymer), poly(sodium 4-styrene-sulfonate) (anionic polymer), (2-aminopropyl) trimethoxy silane (APMS) (silane coupling agent with amino group), and (3-mercaptopropyl) trimethoxy silane (MPS) (silane coupling agent with thiol group). For all the examined chemicals, volume resistivities of surface-modified ITO particles did not increase much when they were aged in air at 80 ^oC, compared to a volume resistivity of un-surface-modified ITO particles. Increases in volume resistivities of ITO particles surface-modified with the silane coupling agents were smaller than those with the polymers, since hydrolysis of the silane coupling agents and condensation of generated silanol and OH groups on ITO particles took place to provide efficient immobilization of them on particles. The APMS gave an increase in volume resistivity smaller than the MPS, since a larger solubility in water of APMS providing a larger amount of APMS immobilized on particles.

Keywords: indium tin oxide, particles, surface-modification, volume resistivity

Procedia PDF Downloads 233

Authors: Reshmi Vijayan

Abstract:

Alkaline protease is one of the most important enzymes in the biological world. Microbial production of alkaline protease is getting more attention from researchers due to its unique properties and substantial activity. Microorganisms are the most common sources of commercial enzymes due to their physiological and biochemical properties. The study was conducted on Ayiramthenghu mangrove sediments to isolate protease producing bacteria. All the isolates were screened for proteolytic activity on a skim milk agar plate at 37˚C for 48hrs. Protease activities were determined by the formation of a clear zone around the colonies on Skim milk agar medium. The activity of the enzyme was measured by the tyrosine standard curve, and it was found to be 0.186285 U/ml/min.

Keywords: protease, protease assay, skim milk agar medium, mangrove ecosystem

Procedia PDF Downloads 51