Presence Of Different Enzymes Research Articles

Washing Performance of Detergent Formulations with Sodium Oxalate as Builder in Presence of Different Enzymes

Abstract In this work, we have investigated the washing performance of sodium oxalate-based detergents containing different enzymes (protease, lipase and cellulase). The results show that sodium oxalate-based detergents with enzymes have a better washing performance than formulations containing the conventional builder STPP and zeolite 4A. For sodium oxalate-based with protease, the amount of sodium oxalate and protease have a significant effect on the detergency. Lipase can improve detergency of sodium oxalate-based detergents after “first washing” in the presence of different anionic surfactant (LAS, MES and AES). For cellulase, only sodium oxalate in the presence of LAS has a detergency increase.

Antifungal protein determination for submerged cultures of the medicinal mushroom <i>Ganoderma lucidum</i> (Ganodermataceae) with activity over the phytopathogen fungus Mycosphaerella fijiensis (Mycosphaerellaceae)

Banana is one of the most important edible crops in the world, however, it’s attacked by different pathogens, one of the most prominent of these is the fungus Mycosphaerella fijiensis, causal agent of Black Sigatoka. The environmental and economic issues related to the pesticides used for its control have encouraged the search for cleaner alternative biomolecules. Previous studies were made by the Biotechnology group from the Universidad de Antioquia searching for a biological alternative for the control of M. fijiensis; in these the antifungal capacity of the fungus Ganoderma lucidum was determined as an antagonist and then as a source of protein extracts with inhibitory activity in vitro and in greenhouse plants; these findings were the foundation of this work, which focuses in the study of the enzymatic capacity of the proteins present in the protein extracts, due to their potential ability to degrade different compounds, including polysaccharides, lipids, peptides and nucleic acids, constituents of essential parts of a living cell; therefore these extracts can act as possible antifungal agents. In this study, protein extracts of G. lucidum obtained from bioreactor cultures (BIOFLO 110 ®) were characterized in terms of their deoxyribonuclease, ribonuclease, protease, glucanase and chitinase enzymatic activities. The extracts were also fractionated and each fraction obtained was evaluated for its inhibitory capacity against the phytopathogen fungus M. fijiensis, and through mass spectrometry analysis the presence of different enzymes with antifungal potential was confirmed.

Phenolic compounds extraction in enzymatic macerations of grape skins identified as low-level extractable total anthocyanin content.

Anthocyanins in wine principally depend on grape skin extractable anthocyanin content, that is, the amount of anthocyanins present in grape skin that are released to wine during the maceration stage. This amount of extractable anthocyanins is closely linked to the cell wall degradation of skin cells. Indeed, among other methodologies, the maceration in presence of different enzymes can be used to increase cell wall degradation, and therefore, the amount of anthocyanins extracted from grape skins to wine. Vitis vinifera L. cv. Tempranillo and Syrah red grapes have been identified as samples with low anthocyanin extraction potential by near infrared hyperspectral imaging. Grape skins have been macerated in the presence of cellulase, glucosidase, and pectinase. Then, color of the supernatants and phenolic compounds extracted from grape skins (total phenols, total flavanols, and total and individual anthocyanins) has been determined. Cellulase and glucosidase have shown a positive effect in the extraction of phenolic compounds from these grapes. Macerations carried out in the presence of cellulase have produced supernatants with a more intense color (lower lightness and higher chroma values), and a higher extraction of flavanols and anthocyanins than the respective control essays. However, pectinase treatments have produced the opposite effect, which could be partially explained by an eventual interaction between the cell wall polysaccharides liberated by pectinase and the phenolic compounds extracted. Synergy effects do not appear between cellulase and glucosidase. Moreover, the negative effect of the addition of pectinase might be due to the interactions between the cell wall material liberated by pectinase and the phenolic compounds extracted. PRACTICAL APPLICATION: In the present study, grape samples with a low anthocyanin extraction potential have been identified, and these samples have been macerated in the presence of different enzymes. The applied enzymes were three of the most common enzymes that are applied in the wine industry. Individual enzymes and mixtures have been applied to Syrah and Tempranillo grape skin samples and the results have been compared to control macerations. Knowledge in this topic will help the production of quality wines.

Biodegradation of Poly(aspartamide) Based Hydrogels

Biocompatibility and biodegradability are one of the most important properties of polymer hydrogels for pharmaceutical or medical applications. Poly(aspartamide)s, due to their peptide like structure possess these properties. In this study, swelling behavior and biodegradation of poly(aspartamide) based hydrogels are reported. Degradation of poly(aspartic acid) based hydrogels was followed according to the mass and elastic modulus change of hydrogel cylinders in the presence of different enzymes and in minimal essential medium.

Cellulases Specific Activities Study of Enzymes secreted by Fusarium sp. and Phlebia sp. in Aerobic and Anaerobic Conditions

Cellulose is the most abundant organic compound on Earth but we can't use it plenty as raw material due to its molecular arrangement and composition. Cellulases are powerful enzymes secreted by some organisms that can be used to degrade cellulose into simple organic molecules like glucose or cellobiose. Although traditional filamentous fungi are known to be excellent cellulose degraders in aerobic conditions, there is little known about cellulose degradation by fungi in anaerobic condition. In this study two facultative anaerobic filamentous fungi isolated and identified using ITS were used to compare cellulase specific activities in aerobic and anaerobic conditions. One Fusarium sp. and Phlebia sp. were grown individually in Potato Dextrose agar for 7 days. Suspensions of spores were prepared for each organism and use it as inoculums. Liquid media culture was prepared and enriched with Carboxylmethylcellulose, Wheat Brand Flour and Whatman no.1 filer paper and inoculated with 1mL of 106 suspension spores of each fungi individually. Cellulase specific activities were determined using Dinitrosalicylic acid method at ph 4, 5, 6, and 7 at 30°C, 40°C, 50°C and 60°C after one month of incubation. Fusarium sp. showed a maximum of cellulase activity at 40°C and pH 5 for both aerobic and anaerobic conditions with values of 0.436 and 0.403 respectively. On the other hand Phlebia sp. showed better results in anaerobic condition with a value of 0.715 at 60°C and pH 6 against a value of 0.632 in aerobic condition at 60°C and ph 5. These results confirm different trending in cellulase specific activities in aerobic and anaerobic conditions and could suggest the presence of different enzymes secreted depending of oxygen concentrations.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Kinetics of dopamine release from poly(aspartamide)-based prodrugs

Preparation of novel biocompatible and biodegradable polymer-based prodrugs that can be applied in complex drug delivery systems is one of the most researched fields in pharmaceutics. The kinetics of the drug release strongly depends on the physicochemical parameters of prodrugs as well as environmental properties, therefore precise kinetical description is crucial to design the appropriate polymer prodrug formula. The aim of the present study was to investigate the dopamine release from different poly(aspartamide) based dopamine drug conjugates in different environments and to work out a kinetic description which can be extended to describe drug release in similar systems. Poly(aspartamide) was conjugated with different amounts of dopamine. In order to alter the solubility of the conjugates, 2-aminoethanol was also grafted to the main chain. Chemical structure as well as physical properties such as solubility, lipophilicity measurements and thermogravimetric analysis has been carried out. Kinetics of dopamine release from the macromolecular prodrugs which has good water solubility has been studied and compared in different environments (phosphate buffer, Bromelain and α-Chymotrypsin). It was found that the kinetics of release in those solutions can be satisfactorily described by first order reaction rate. For poorly-soluble conjugates, the release of dopamine was considered as a result of coupling of diffusion and chemical reaction. Besides the time dependence of dopamine cleavage, a practical quantity, the half–life of the release of loading capacity has been introduced and evaluated. It was found, that dopamine containing macromolecular prodrugs exhibit prolonged release kinetics and the quantitative description of the kinetics, including the most important physical parameters provides a solid base for future pharmaceutical and medical studies. Statement of SignificancePoly(aspartamide) based polymer-drug conjugates are promising for controlled and prolonged drug delivery due to their biocompatibility and biodegradability. In this study different poly(aspartamide) based dopamine conjugates were synthesized which can protect dopamine from deactivation in the human body. Since there is no satisfying kinetics description for drug release from covalent polymer-drug conjugates in the literature, dopamine release was investigated in different environments and a complete kinetical description was worked out. This study demonstrates that poly(aspartamide) is able to protect conjugated dopamine from deactivation and provide prolonged release in alkaline pH as well as in the presence of different enzymes. Furthermore, detailed kinetical descriptions were demonstrated which can be used in case of other covalent polymer-drug conjugates.

Biochemical and Molecular Studies of Various Enzymes Activity in Fungi

Fungi are decomposers in most ecosystems and make important contribution to the ecological balance of our world. They have great industrial importance due to the presence of different enzymes like laccase, superoxide dismutase, cellulases, amylases and catalase etc. These enzymes performed synthetic and degradative functions. They are physiologically necessary for all of the living organisms and are universally occur with wide genetic diversity in plants, animals and micro-organisms. Mostly the micro-organisms are an attractive and efficient source of various enzymes and also owing to have the limited space required for their cultivation and their ready susceptibility to genetic manipulations. Although the extensive research on various aspects of these enzymes, there is scarcity of the knowledge about the role that governed the diverse specificity of these enzymes. After deciphering the secrets about these enzymes would enable us to exploit their use in biotechnology. Fungi have vital roles in biotechnology such as production of drugs and enzymes. Fungi can be cultured easily and hence they can be used in microbiological, genetic and molecular research. It is very important to investigate genes and the role of genes that are responsible for the formation of these enzymes. In the current review described the study of production of laccase, superoxide dismutase and catalase enzyme through various fungi, the activity of enzymes and the genetic diversity of genes involved for the formation of these enzymes.

Indole-3-Acetic Acid Biosynthesis in Fusarium delphinoides Strain GPK, a Causal Agent of Wilt in Chickpea

Fusarium delphinoides (Ascomycota; Nectriaceae) is an indole-3-acetic acid (IAA) producing plant pathogen and a causal agent of wilt in chickpea. The IAA biosynthetic pathway in F. delphinoides strain GPK (FDG) was examined by analyzing metabolic intermediates and by feeding experiments. Gas chromatograph (GC) analysis of FDG culture filtrates showed the presence of metabolic intermediates of indole-3-pyruvic acid (IPyA), indole-3-acetamide (IAM), and tryptamine (TRA) pathways. The different IAA biosynthetic pathways were further confirmed by identifying the presence of different enzymes of these pathways. Substrate specificity study of aromatic amino acid aminotransferase revealed that the enzyme is highly specific for tryptophan (Trp) and α-ketoglutarate (α-kg) as amino group donor and acceptor, respectively. Furthermore, the concentration-dependent effect of exogenous IAA on fungal growth was established. Low concentration of exogenous IAA increases the fungal growth and at high concentration it decreases the growth of FDG.

Human serum albumin (HSA) nanoparticles: Reproducibility of preparation process and kinetics of enzymatic degradation

Nanoparticles prepared from human serum albumin (HSA) are versatile carrier systems for drug delivery and can be prepared by an established desolvation process. A reproducible process with a low batch-to-batch variability is required for transfer from the lab to an industrial production. In the present study the batch-to-batch variability of the starting material HSA on the preparation of nanoparticles was investigated. HSA can build dimers and higher aggregates because of a free thiol group present in the molecule. Therefore, the quality of different HSA batches was analysed by size exclusion chromatography (SEC) and analytical ultracentrifugation (AUC). The amount of dimerised HSA detected by SEC did not affect particle preparation. Higher aggregates of the protein detected in two batches by AUC disturbed nanoparticle formation at pH values below 8.0. At pH 8.0 and above monodisperse particles between 200 and 300 nm could be prepared with all batches, with higher pH values leading to smaller particles. Besides human derived albumin a particle preparation was also feasible based on recombinant human serum albumin (rHSA). Under comparable preparation conditions monodisperse nanoparticles could be achieved and the same effects of protein aggregates on particle formation were observed. For nanoparticulate drug delivery systems the enzymatic degradation is a crucial parameter for the release of an embedded drug. For this reason, besides the particle preparation process, particle degradation in the presence of different enzymes was studied. Under acidic conditions HSA as well as rHSA nanoparticles could be digested by pepsin and cathepsin B. At neutral pH trypsin, proteinase K, and protease were suitable for particle degradation. It could be shown that the kinetics of particle degradation was dependent on the degree of particle stabilisation. Therefore, the degree of particle stabilisation will influence drug release after cellular accumulation of HSA nanoparticles.

Enzyme-catalyzed desymmetrization of meso-skipped polyols to useful chiral building blocks

the biocatalytic desymmetrization, in presence of different enzymes, of two models of polyfunctionalized diols in a meso form was studied PFL was found to be the most selective enzyme with excellent results in chemical and optical yield with some of the tested substrates. The obtained chiral building block was utilized for some useful synthetic transformations toward the synthesis of natural products

Aminopyrine N-demethylase activity in human liver microsomes

Aminopyrine N-demethylase activity and the contents of cytochrome P-450, cytochrome b5, and NADPH-reductase activity in human liver microsomes from 31 different patients were studied. Our results show the existence of significant interindividual, but not sex- or age-related differences in N-demethylase activity (ranging between 0.52 and 4.42 nmol/min/mg protein), and that these differences are directly correlated with the content of cytochrome P-450 and NADPH-reductase activity, but not with that of cytochrome b5 content, in the samples. In contrast, no differences were found in the Michaelis-Menten constant values for aminopyrine (mean, 2.4 mmol/L) or NADPH (mean, 69 mumol/L), strongly suggesting that interindividual differences could be due to the occurrence of different amounts of the same enzyme, rather than the presence of different enzymes, in the population studied.

Degradation of terephthalic acid by a Bacillus species

A Gram-positive bacterium with the ability to utilize terephthalic acid as sole carbon source was isolated from soil. The strain was identified as a Bacillus sp. Protocatechuate was shown to be a key intermediate in the degradation of terephthalate. Oxygen uptake studies were carried out with the probable intermediates. The presence of different enzymes was tested for. A mechanism is proposed for the degradation of terephthalate.

Degradation of terephthalic acid by aBacillusspecies

A Gram-positive bacterium with the ability to utilize terephthalic acid as sole carbon source was isolated from soil. The strain was identified as a Bacillus sp. Protocatechuate was shown to be a key intermediate in the degradation of terephthalate. Oxygen uptake studies were carried out with the probable intermediates. The presence of different enzymes was tested for. A mechanism is proposed for the degradation of terephthalate.

Influence of fasting and refeeding on the elongation step of protein synthesis in rat liver of young (3M) and aging (18M) animals

The in vitro binding capacity for AA-tRNA of ribosomes, catalysed by EF 1 enzyme, was studied in livers of rats, which had been starved for 5 days and re-fed for 3, 9, and 24 hours. The experiments were carried out with 3 and 18-month-old rats. The results were compared to those obtained with normally fed rats of the same ages. For both age groups, there was a similar change in the binding capacity, including a return to normal values after 24 hours of refeeding. When ribosomes of old control animals were treated with different enzymes, we obtained lower binding activities for all the experimental groups than in homologous cell-free systems (ribosomes and EF 1 isolated from the same group). The ribosomes of young animals, tested in the presence of different enzymes, display an increased binding capacity, when the enzymes used were those of old (control and re-fed) animals. The recovery of the catalytic activity of EF 1 on the binding process was observed after 9 hours in old animals, whereas it was not seen for young re-fed animals at that time of refeeding. EF 1 seems to play an important role in the adaptation of older animals to fasting and refeeding.

Endogenous Role of Microsomal Epoxide Hydrolase

The specific activities of microsomal epoxide hydrolase with 16 alpha, 17 alpha-epoxyandrosten-3-one (androstene oxide) as substrate were measured in various metabolically important and in various steroidogenic organs of the male and female rat and compared with the activities of 16 alpha, 17 alpha-epoxyestratrienol (estroxide) and benzo[a]pyrene 4,5-oxide. Androstene oxide was an exceptionally good substrate. The specific activities differed widely between organs but the ratio of the activities towards these substrates was constant in all organs investigated. The ratios compared to benzo[a]pyrene 4,5-oxide were 2.5 for estroxide, and 8.6 for androstene oxide. The ontogenetic development of specific epoxide hydrolase activity in the livers of both sexes reached a maximum at about day 40 and descended to the adult enzyme level at about 45 days in males and clearly later in females. While in the livers and ovaries significant increases of the enzyme activity with increasing age took place before day 28, the specific activity remained very low in the testis until day 28 and then rose suddenly. During all these differential developments no significant changes in the ratios of activities towards the three substrates were observed. The specific activity of epoxide hydrolase towards these substrates in subcellular fractions of the rat liver was smooth endoplasmic reticulum greater than microsomes approximately equal to rough endoplasmic reticulum much greater than mitochondria, no activity was detectable in cytosol. The ratio of the activities in the different fractions was similar when measured with androstene oxide, estroxide and styrene oxide as substrates. Microsomal hydrolysis responded to pretreatment of animals with phenobarbital, 3-methylcholanthrene. Arochlor 1254 and trans-stilbene oxide in a manner which was characteristically different for the various agents but similar for the three substrates. Microsomal epoxide hydrolase which was purified to apparent homogeneity was able to hydrolyse the steroid epoxides, but the apparent purification factors were different for the different substrates: 77 for styrene oxide, 45 for estroxide, and 10 for androstene oxide. The three substrates mutually inhibited their hydrolysis by the microsomal fraction. Some differences in the extent of their effect and in the inhibition of the activities by known epoxide hydrolase inhibitors were observed. Similarly, hydrolysis of the steroid epoxides but not of styrene oxide was inhibited by nonionic detergents (Cutscum, Triton X-100 and Emulgen 911). These differences could be due to the presence of different enzymes or a single enzyme, the conformational requirements of which are much more demanding for steroid epoxides than for xenobiotic epoxides. Mono-specific antiserum precipitated epoxide hydrolase activity from solubilized microsomes with dose-response curves which were not distinguishable for androstene oxide, estroxide, benzo[a]pyrene 4,5-oxide and styrene oxide as substrates...

The nature and origin of the soluble protein in human amniotic fluid.

Summary1. Amniotic fluid surrounds the human fetus and is separated from the uterus by the amnion, chorion and placenta. The ability to obtain samples of amniotic fluid from women by a simple procedure has encouraged studies on the nature and origin of the fluid, and on its use for the diagnosis of a variety of clinical conditions. The fluid contains cells, which are of fetal origin, and can be grown in a tissue culture. Cyto‐genetic and biochemical analyses can therefore be used to detect chromosomal aberrations and inborn errors of metabolism in the fetus.2. The supernatant of amniotic fluid contains many of the solutes typical of extracellular fluid. In particular, it contains a wide range of proteins and those which are of fetal origin are likely to be of use in the prenatal diagnosis of fetal disease. This review examines the nature and origin of the soluble protein in amniotic fluid, and discusses the diagnostic uses of the proteins which are of fetal origin.3. In other mammals, the arrangement of the fetal membranes is different from that in man, and these differences are reflected by changes in the nature of the amniotic fluid. Thus data from other animals have little applicability to man.4. Electrophoresis and immunoelectrophoresis have established that the major proteins in amniotic fluid are also present in maternal and fetal sera. Their concentrations in the fluid are influenced by their molecular weight and proteins larger than about 2.5 times 106 may be excluded. Towards term, phenotyping studies show that a number of serum proteins in amniotic fluid are of maternal origin. In the case of group‐specific component (Gc) this has been shown to be so throughout pregnancy. Such proteins must enter the fluid by diffusing across either the chorion or the chorionic plate and then the amnion.5. It has been previously claimed that various serum proteins in amniotic fluid are of fetal origin. For albumin and IgG there are data that strongly support a maternal origin. The evidence on the origin of insulin is inconclusive. The concentration of β2‐microglobulin in amniotic fluid exceeds that in maternal serum and is probably too high also for fetal serum to be its major source. It has a wide tissue distribution and probably enters the fluid from surrounding structures.6. Alpha‐fetoprotein in amniotic fluid is of fetal origin as it is present in maternal serum at far lower concentrations. It is found in fetal serum, urine and yolk sac, but it is not clear how it enters the amniotic fluid of normal fetuses. The concentrations of Gc and alpha‐fetoprotein have been measured in amniotic fluid and in their sera of origin. The relative concentration of Gc in amniotic fluid was found to be much greater than that of alpha‐fetoprotein and the concentration gradients of these marker proteins can be compared with data for other proteins. In this way further evidence has been obtained that the albumin, α1,‐antitrypsin and transferrin in amniotic fluid are mainly of maternal origin throughout pregnancy.7. Immunological studies have shown that at least three proteins of non‐serum origin are present in amniotic fluid and they have also been located in the amnion and uterine decidua.8. The enzymes present in amniotic fluid are summarized. Many lysosomal enzymes are clearly of fetal origin since they show altered specific activities in the appropriate cases where the fetus is affected with an inborn error of metabolism. For other enzymes, analysis of specific activity gradients can help to decide the extent to which an enzyme is of serum origin, although this will not exclude the possibility of a maternal (uterine) contribution. The results of such analyses suggest that, relative to the serum protein in amniotic fluid, the greatest concentrations of the minor non‐serum proteins in the fluid occurs between thirteen and eighteen weeks of pregnancy and also towards term.9. Some inborn errors of metabolism may be diagnosed prenatally by measuring the specific activity of the respective enzyme in amniotic fluid. However, the presence of different enzymes with similar substrate specificities has prevented this in Pompe's disease.10. In cases where the fetus is affected with anencephaly or spina bifida there is an increase in the concentration of alpha‐fetoprotein in the amniotic fluid. This has provided a way of detecting these diseases early enough to allow termination of pregnancy.11. The discovery of new proteins in fetal serum and in the tissues surrounding the amniotic cavity would seem to provide the best chance of extending the uses of amniotic fluid into the other areas of prenatal medicine.

Effect of isomeric cis-octadecenoic acids on the growth of Leptospira interrogans serotype patoc.

Leptospira interrogans serotype patoc exhibited an increasing growth response when cultivated in media containing from 50 to 250 mug of sodium oleate per ml. Leptospiral growth in the presence of 250 mug of sodium oleate per ml was as good as that in the basal medium which contained 700 mug of oleic acid (in Tween 80) per ml. When positional isomers of oleic acid (9-octadecenoic acid) were present at a concentration of 200 mug/ml, the 2- and 8-isomers were not readily utilized, whereas the 3-, 4-, 6-, 11-, 15-, and 16-isomers gave a growth response equivalent to that of oleic acid, i.e., the 9-isomer. The 5-, 7-, 10-, 12-, 13-, 14-, and 17-isomers of octadecenoic acid induced growth responses which differed in magnitude but were intermediate to those of 2-18:1 and 3-18:1. When 200 mug of either 2- or 3-octadecenoic acid per ml was added in addition to 200 mug of 9-18:1 alone; 400 mug of 9-18:1 alone per ml inhibited growth of this organism. The growth response of leptospira to octadecenoic acids differed from that of mammalian cells, suggesting the presence of different enzymes in the two systems for the utilization of these substrates.