Home About us Contact
|
Home About us Contact | ||
|
|
||
Enzyme Solution (enzyme + solution)
Selected AbstractsElectrophoretically mediated microanalysis for the evaluation of interspecies variation in cholinesterase metabolismELECTROPHORESIS, Issue 14 2010Joana Moura Abstract This study describes an electrophoretically mediated microanalysis method, suitable for the preclinical evaluation of the hydrolysis of ester drugs by the serum of different animals and for further characterization of human,animal correlation. Dog, cat, cow, horse, sheep, rat and human serum were diluted (25%) in the appropriate buffer and replaced the enzyme solution usually used in electrophoretically mediated microanalysis methods for the study of enzyme kinetics. They were then compared in terms of the ability to hydrolyze acetylthiocholine and butyrylthiocholine (0.25,mM) by in-capillary reaction. Human serum afforded the highest conversion rates (52% butyryltiocholine and 34% acetylthiocholine) followed by horse (31 and 35%), dog (26 and 24%), cat (22 and 14%), rat (11 and 15%) and sheep (8 and 8%). Hydrolysis by bovine serum was negligible. The method is fast (under 8,min including rinsing steps), sensitive (under 25,,M substrate could be quantified) and repeatable (RSD,2%), only requiring minute amounts of sample. [source] Ace2, rather than ace1, is the major acetylcholinesterase in the silkworm, Bombyx moriINSECT SCIENCE, Issue 4 2009Hui-Juan Chen Abstract, Two acetylcholinesterase (ace) genes have been reported in many insect species. In pests such as Helicoverpa assulta and Plutella xylostellas, ace1 gene encodes the predominant synaptic enzyme that is the main target of organophosphorus (OP) and carbamate pesticides. It has been reported that pesticide selection has an impact on the ace gene evolution. The domesticated silkworm, Bombyx mori, also has two ace genes. We studied ace gene expression and enzyme activities in silkworm as this has not faced pesticide selection over the past decades. The expression levels of two ace genes, Bm- ace1 and Bm- ace2, were estimated by quantitative real-time polymerase chain reaction. Bm- ace2 was expressed more highly than Bm- ace1 in all tested samples of different developmental stages or tissues, suggesting ace2, rather than ace1, is the major type of acetylcholinesterase (AChE) in Bombyx mori. This is inconsistent with the aforementioned lepidopterons agricultural pests, partly be due to the widespread use of pesticides that may induce high expression of the ace1 gene in these pests. Besides high expression in the head, Bm- ace1 also expresses highly in the silk glands and Bm- ace2 is abundant in the germline, implying both ace genes may have potential non-hydrolytic roles in development. Furthermore, we found that the mRNA levels of two ace genes and their ratios (ace2/ace1) change day to day in the first and third instars. This challenges the conventional method of estimating enzymatic activity using crude extract as an enzyme solution, as it is a mixture of AChE1 and AChE2. An efficient and simple method for separating different AChEs is necessary for reliable toxicological analyses. [source] A comparison of changes in the transformation of isoflavones in soymilk using varying concentrations of exogenous and probiotic-derived endogenous , -glucosidasesJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2007D.O. Otieno Abstract Aims:, To compare endogenous and exogenous , -glucosidases for the hydrolysis of the predominant isoflavone glucosides in soymilk in order to improve the biological activity. Methods and Results:,, -glucosidase activity of probiotic organisms, including Bifidobacterium animalis ssp. lactis Bb12, Lactobacillus acidophilus ATCC 4461 and Lactobacillus casei 2607 in soymilk, was evaluated and was related to the increase in the concentration of isoflavone aglycones during fermentation. The concentrations of isoflavone compounds in soymilk were monitored using a Varian model HPLC with an Amperometric electrochemical detector. The aglycone composition, also known as aglycone equivalent ratio, has been considered to be important for the delivery of health benefits of isoflavones, and was monitored during the fermentation of soymilk. Comparison of the hydrolytic effectiveness of both exogenous and endogenous enzyme during 4-h incubation in soymilk was conducted using the Otieno,Shah (O,S) index. Results showed that exogenous enzyme exhibited faster rate of isoflavone glucoside hydrolysis than that by endogenous enzyme. Highest O,S indices were obtained after 4, 3 and 2 h of incubation with enzyme solution having , -glucosidase activity of 0·288 U ml,1, 0·359 U ml,1 and 0·575 U ml,1, resulting into aglycone concentration increments of 5·87-, 6·07- and 5·94-fold, respectively. Conversely, aglycone concentration in the soymilk with B. animalis ssp. lactis Bb12, L. casei 2607 and L. acidophilus 4461 increased by 3·43-, 2·72- and 3·03-fold, respectively, after 4 h of fermentation at 37°C. In addition, the O,S index of endogenous enzyme was much lower than that of the exogenous enzyme over the same 4-h incubation period. Optimum aglycone equivalent ratios coincided with highest O,S indices and highest aglycone concentrations in soymilk hydrolysed with exogenous enzyme. The same correlation of O,S indices and highest aglycone concentrations occurred for endogenous enzyme during the 24 h of fermentation. Conclusions:, Obtaining highest aglycone concentration and optimum aglycone equivalent ratio could provide a critical beginning point in clinical trials for the realization of unique health benefits of soy isoflavones. Significance and Impact of the Study:, Screening for , -glucosidase activities of probiotics in soymilk and comparing their hydrolytic potentials with that of exogenous , -glucosidase could find wide applications in the development of different aglycone-rich functional soy beverages. [source] Recovery of lipase by adsorption at the n -hexadecane,water interfaceJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 11 2003Hui-Min Wang Abstract A novel separation process based on the hydrophobic adsorption at the n -hexadecane,water interface was developed for the recovery of Acinetobacter radioresistens lipase from a pre-treated fermentation broth. In a mixture containing water, lipase and n -hexadecane, a water-in-oil emulsion was formed when the n -hexadecane-to-water ratio (o/w ratio) was larger than 3, and a large amount of lipase was found to be adsorbed at the interface. Compared with the oil-in-water emulsion (occurring when o/w ratio < 3), the water-in-oil emulsion generated smaller droplets and larger interfacial area, and was more stable. The harvested emulsion phase could be centrifuged to give an aqueous, concentrated lipase solution. Adsorption of lipase at the interface could be described by the Langmuir isotherm. For lipase concentrations ranging from 8.4 to 87.2 U cm,3, a single-stage adsorption resulted in a six- to four-fold concentration and 16,45% activity recovery, where lipase concentration was the dominant factor. A method using data from a single-stage adsorption to predict multiple-stage operation was described, and the agreement between the experimental and the predicted results was good. To improve the enzyme recovery, a multiple-run adsorption process was proposed. The use of salts enhanced the hydrophobic interaction between lipase and n -hexadecane. Advantages of the proposed process include simple operation, low operational cost, environmentally friendly, no requirement for pre-concentration of the enzyme solution, and negligible enzyme denaturation. Copyright © 2003 Society of Chemical Industry [source] EFFECTS OF ENZYME-AIDED PEELING ON THE QUALITY OF LOCAL MANDARIN (CITRUS RETICULATA B.) SEGMENTSJOURNAL OF FOOD PROCESSING AND PRESERVATION, Issue 5 2004FANNY LIU ABSTRACT Pectinases are observed to selectively alter the albedo structure of citrus fruits and, hence, aid the removal of the peel and adhering albedo layer. This study was carried out to determine the optimum conditions needed to peel local mandarins using pectinases (Peelzym II, Novo Nordisk, Denmark). The experiment variables were enzyme concentration, vacuum pressure and vacuum infusion time. The mandarins were first scored from the stem end to the blossom end, followed by immersion in 1000 mL of enzyme solution at a set vacuum pressure and ambient temperature (27 ± 1C). Only one parameter was varied in any one experiment. Peelzym II at 0.4% v/w, 650 mm Hg vacuum and 16 min of vacuum time were optimal. The enzyme-peeled fruit segments were judged by the panelists using three different sensory tests to ascertain their appeal to consumers. A significant (P < 0.05) difference between enzyme-peeled and hand-peeled segments was found, with the panelists preferring the enzyme-peeled segments. [source] Enzyme degradability of benzylated sisal and its self-reinforced compositesPOLYMERS FOR ADVANCED TECHNOLOGIES, Issue 10 2003Xun Lu Abstract To produce natural polymer based composite materials, sisal fibers were slightly benzylated and then molded into sheets. Because the modified skin portions of the fibers acquired certain thermoplasticity and the unmodified core parts remain constant, the resultant composites fall into the category of self-reinforced ones. The present article is devoted to the evaluation of the materials biodegradability with the help of cellulase. It was found that the inherent biodegradability of plant fibers is still associated with the benzylated sisal and the molded composites, as characterized by structural variation, weight loss and deterioration of mechanical performance of the materials. Reaction temperature and time, pH value of the enzyme solution, and dosage of the enzyme had significant influences on the decomposition behavior of the materials. In principle, the enzymolysis of sisal and its self-reinforced composites is a diffusion-controlled process. Due to the insusceptibility of lignin to cellulase and the hindrance of it to the cellulase solution, the degradation rates of the materials are gradually slowed down with an increase in time. Copyright © 2003 John Wiley & Sons, Ltd. [source] A quick method for the assessment of activity and inhibition of fish amylasesAQUACULTURE NUTRITION, Issue 1 2001I. Fernández A sensitive and quick method was developed to determine the presence of ,-amylase in the gut of aquatic organisms, as well as its sensitivity to inhibitors. The assay is based on the utilization of Petri dishes filled with starch,agarose gel as a substrate for the enzyme solution, which is placed in small wells punched in the surface. Circular zones produced by the action of amylase remain colourless after staining with lugol. Pure commercial porcine amylase was used to fit the better conditions for developing the assay (1 g L,1 starch in the gels, 4 h of incubation). The diameter of the cleared zones were related to the activity of enzyme and the method detected linearly amylase activity in a range of 2,20 U well,1, so it was used to reveal the presence of amylase in digestive extracts obtained from different sparid fish. The method was also used to evaluate the effect produced by a specific inhibitor on fish amylases, showing a linear response when the ratio inhibitor:enzyme (in units) changed from 20:1 to 2:1. Comparison of the cleared zones produced by amylases of sparid fish in the presence or absence of inhibitor, revealed differences in their sensitivity to inhibition, which ranged from 15 to 50% of total activity. The assay is proposed for a preliminary evaluation of possible inhibitors contained in feedstuffs used in fish feeding. [source] Effects of four egg desticking procedures on hatching rate and further survival and growth of larvae in the tench (Tinca tinca L.)AQUACULTURE RESEARCH, Issue 6 2006Jose M Carral Abstract Four desticking procedures for tench eggs (A: tannic acid solution (1 g L,1) for 15 s; B: alcalase enzyme solution (8 mL L,1) for 60 s; C: alcalase enzyme solution (15 mL L,1) for 120 s; D: Woynarovich and Woynarovich (1980) solution for 58 min followed by tannic acid solution (1 g L,1) for 15 s) were tested to obtain data about influence on embryo survival to hatching stage and further survival and growth of the larvae. In the tannic acid and Woynarovich and Woynarovich (1980) treatment (A and D) few eggs stuck together and some were adhered to the incubator walls, whereas in the alcalase treatments (B and C) eggs neither stuck together nor adhered to the incubator walls. Percentages of hatched larvae did not show significant differences (mean values ranged between 47.4% in treatment A to 37.0% in treatment C). Larvae deformities observed were <0.5% in all cases. There were no significant differences among survival and growth rates of the larvae from different egg desticking origin, reaching, after 30 days, mean survival values around 90% and total length and weight of 12.5 mm and 19 mg respectively. [source] Invertase-Lipid Biocomposite Films: Preparation, Characterization, and Enzymatic ActivityBIOTECHNOLOGY PROGRESS, Issue 1 2004Sumant Phadtare The formation of biocomposite films of the industrially important enzyme invertase and fatty lipids under enzyme-friendly conditions is described. The approach involves a simple beaker-based diffusion protocol wherein invertase diffuses into the cationic lipid octadecylamine during immersion of the lipid film in the enzyme solution. Entrapment of invertase in the octadecylamine film is highly pH-dependent, underlining the role of attractive electrostatic interactions between the enzyme and the lipid in the biocomposite film formation. The kinetics of formation of the enzyme-lipid biocomposites has been studied by quartz crystal microgravimetry (QCM) measurements. The stability of the enzyme in the lipid matrix was confirmed by fluorescence spectroscopy and biocatalytic activity measurements. The biocatalytic activity of the invertase-lipid biocomposite films was comparable to that of the free enzyme in solution and showed marginally higher temperature stability. Particularly exciting was the excellent reuse characteristics of the biocomposite films, indicating potential industrial application of these films. [source] Production of a Polyester Degrading Extracellular Hydrolase from Thermomonospora fuscaBIOTECHNOLOGY PROGRESS, Issue 5 2002Mona K. Gouda The production of a polyester-degrading hydrolase from the thermophilic actinomycete Thermomonospora fusca was investigated with regard to its potential technical application. Only in the presence of a polyester (random aliphatic-aromatic copolyester from 1,4-butanediol, terephthalic acid, and adipic acid with around 40,50 mol % terephthalic acid in the acid component), the excretion of the extracellular enzyme could be achieved with an optimized synthetic medium using pectin and NH4Cl as nitrogen source. Compared to complex media, a significantly higher specific activity at comparable volumetric yields could be obtained, thus reducing the expenditure for purification. The activity profile in the medium is controlled by a complex process involving (1) induction of enzyme excretion, (2) enzyme adsorption on the hydrophobic polyester surface, (3) inhibition of enzyme generation by monomers produced by polyester cleavage, and (4) enzyme denaturation. Diafiltration with cellulose acetate membranes as the sole downstream processing step led to a product of high purity and with sufficient yield (60% of total activity). Scaling-up from shaking flasks to a fermentor scale of 100 L revealed no specific problems. However, the excretion of the hydrolase by the actinomycete turned out to be inhibited by the degradation products (monomers) of the aliphatic-aromatic copolyester used as inductor for the enzyme production. The crude enzyme exhibited generally similar properties (temperature and pH optimum) as the highly purified hydrolase described previously; however, the storage capability and thermal stability is improved when the crude enzyme solution is diafiltrated. [source] Impact of Supercritical Carbon Dioxide and High Pressure on Lipoxygenase and Peroxidase ActivityJOURNAL OF FOOD SCIENCE, Issue 8 2000W. TEDJO ABSTRACT: The effects of supercritical carbon dioxide (ScCO2) treatment and high hydrostatic pressure treatment on the activities of lipoxygenase (LOX) and peroxidase (POD) were studied. Hydrostatic pressure treatment (240 MPa, 55 °C, 15 min) of LOX and POD in 30% sucrose solutions without buffer led to approximately 80% and approximately 50% residual activity, respectively. Application of ScCO2 (35.2 MPa, 40 °C, 15 min for LOX and 62.1 MPa, 55 °C, 15 min for POD) achieved approximately 35% LOX and approximately 65% POD inactivity in 30 % sucrose solutions. Total inactivation of LOX (10.3 MPa, 50 °C and 15 min) and of POD (62.1 MPa, 55 °C and 15 min) could be achieved through ScCO2 treatment of unbuffered solution. Increasing the concentration of sucrose and buffering (pH range 4 to 9) of enzyme solutions resulted in increased resistance of the enzymes to ScCO2 treatment. [source] Methodological analysis for determination of enzymatic digestibility of cellulosic materialsBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2007Y.-H. Percival Zhang Abstract Accurate measurement of enzymatic cellulose digestibility (X) is important in evaluating the efficiency of lignocellulose pretreatment technologies, assessing the performance of reconstituted cellulase mixtures, and conducting economic analysis for biorefinery processes. We analyzed the effect of sugars contained in enzymes solutions, usually added as a preservative, and random measurement errors on the accuracy of X calculated by various methods. The analysis suggests that exogenous sugars at levels measured in several commercial enzyme preparations significantly bias the results and that this error should be minimized by accounting for these sugars in the calculation of X. Additionally, a method of calculating X equating the ratio of the soluble glucose equivalent in the liquid phase after hydrolysis to the sum of the soluble glucose equivalent in the liquid phase and the insoluble glucose equivalent in the residual solid after hydrolysis was found to be the most accurate, particularly at high conversion levels (>ca. 50%). Biotechnol. Bioeng. 2007;96: 188,194. © 2006 Wiley Periodicals, Inc. [source] Using the Aggregation of Latex Polymers in the Fabrication of Reproducible Enzyme ElectrodesELECTROANALYSIS, Issue 17 2003Wibowo Rahmat Abstract An enzyme electrode for glucose is described as a model system to demonstrate a fabrication method using latex aggregation and entrapment of enzyme. Electrosterically-stabilized latex particles synthesized by emulsion polymerization in batch from acrylic acid, methyl methacrylate and butyl acrylate, and glucose oxidase were coagulated together at pH,5.5 with ethanol. A platinum disk electrode dipped in the solution becomes coated with latex/enzyme. The relative thickness of the film and relative amount of enzyme may be controlled by the time the electrode is in contact with the solution. The enzyme was then immobilized by covalent attachment of amine groups to carboxylic moieties in the polymer using 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide hydrochloride and N -hydroxysuccinimide. Five minutes contact with the latex/enzyme solution and subsequent amide coupling, gave electrodes with a reproducibility of 5.7% RSD, a wide dynamic range (0,100,mM) and good storage properties. [source] | ||||||||||||||||