Search results for: DHA enzyme activity; labile carbon

Authors: Ogbonnaya Nwokoro, Florence O. Anya

Abstract:

Nutritional factors relating to the production of linamarase from Lactobacillus delbrueckii NRRL B–763 were investigated. The microorganism was cultivated in a medium containing 1% linamarin. Enzyme was produced using a variety of carbon substrates but the highest enzyme activity was detected in the presence of salicin (522 U/ml) after 48 h while the lowest yield was observed with CM cellulose (38 U/ml) after 72 h. Enzyme was not produced in the presence of cellobiose. Among a variety of nitrogen substrates tested, peptone supported maximum enzyme production (412 U/ml) after 48 h. Lowest enzyme production was observed with urea (40 U/ml). Organic nitrogen substrates generally supported higher enzyme productivity than inorganic nitrogen substrates. Enzyme activity was observed in the presence of Mn2+ (% relative activity = 216) while Hg2+ was inhibitory (% relative activity = 28). Locally-formulated media were comparable to MRS broth in supporting linamarase production by the bacterium. Higher enzyme activity was produced in media with surfactant than in media without surfactant. The enzyme may be useful in enhanced degradation of cassava cyanide.

Keywords: linamarase, locally formulated media, carbon substrates, nitrogen substrates, metal ions

Procedia PDF Downloads 426

Authors: Ashish Shukla, K. P. Mishra, Pushplata Tripathi

Abstract:

This paper investigates the production and optimization of β-galactosidase enzyme using synthetic medium by isolated wild strains (S1, S2) mutated strains (M1, M2) through SSF and SmF. Among the different cell disintegration methods used, the highest specific activity was obtained when the cells were permeabilized using isoamyl alcohol. Wet lab experiments were performed to investigate the effects of carbon and nitrogen substrates present in Vogel’s medium on β-galactosidase enzyme activity using S1, S2, and M1, M2 strains through SSF. SmF experiments were performed for effects of carbon and nitrogen sources in YLK2Mg medium on β-galactosidase enzyme activity using S1, S2 and M1, M2 strains. Effect of pH on β-galactosidase enzyme production was also done using S1, S2, and M1, M2 strains. Results were found to be very appreciable in all the cases.

Keywords: β-galactosidase, cell disintegration, permeabilized, SSF, SmF

Procedia PDF Downloads 269

Authors: Furqan M. Kadhum, Asmaa A. Hussein, Maysaa Ch. Hatem

Abstract:

Hyaluronidase enzyme was purified from the clinical isolate Staphyloccus aureus in three purification steps, first by precipitation with 90% saturated ammonium sulfate, ion exchange chromatography on DEAE-Cellulose, and gel filtration chromatography throughout Sephacryl S-300. Specific activity of the purified enzyme was reached 930 U/mg protein with 7.4 folds of purification and 46.5% recovery. The enzyme has an average molecular weight of about 69 kDa, with an optimum pH of enzyme activity and stability at pH 7, also the optimum temperature for activity was 37oC. The enzyme was stable with full activity at a temperature ranged between 30-40 oC. Metal ions showed variable inhibitory degree with the strongest effect for Fe+3, however, the chelating and reducing agents had no or little effects. Cytotoxic studies for purified and crude hyaluronidase against cancer cell Hep G2 type at different enzyme concentrations and exposure times showed that the inhibition effect of both crude and purified enzyme increased by increasing the enzyme concentration with no change was observed at 24hr, while at 48 and 72 hrs the same inhibition rate were observed for purified enzyme and differ for the crude filtrate.

Keywords: hyaluronidase, S. aureus, metal ions, cytotoxicity

Procedia PDF Downloads 445

Authors: M. Umar Dahot, G. S. Mangrio

Abstract:

In this study Agro-industrial waste was used as a carbon source, which is a low cost substrate. Along with this, various sugars and molasses of 2.5% and 5% were investigated as substrate/carbon source for the growth of A.niger and Pectinase production. Different nitrogen sources were also used. An overview of results obtained show that 5% sucrose, 5% molasses and 0.4% (NH4)2SO4 were found the best carbon and nitrogen sources for the production of pectinase by A. niger. The maximum production of pectinase (26.87units/ml) was observed at pH 6.0 after 72 hrs incubation. The optimum temperature for the maximum production of pectinase was achieved at 35ºC when maximum production of pectinase was obtained as 28.25Units/ml.Pectinase enzyme was purified with ammonium sulphate precipitation and dialyzed sample was finally applied on gel filtration chromatography (Sephadex G-100) and Ion Exchange DEAE A-50. The enzyme was purified 2.5 fold by gel chromatography on Sephadex G-100 and Four fractions were obtained, Fraction 1, 2, 4 showed single band while Fraction -3 showed multiple bands on SDS Page electrophoresis. Fraction -3 was pooled, dialyzed and separated on Sephdex A-50 and two fractions 3a and 3b showed single band. The molecular weights of the purified fractions were detected in the range of 33000 ± 2000 and 38000± 2000 Daltons. The purified enzyme was specifically most active with pure pectin, while pectin, Lemon pectin and orange peel given lower activity as compared to (control). The optimum pH and temperature for pectinase activity was found between pH 5.0 and 6.0 and 40°- 50°C, respectively. The enzyme was stable over the pH range 3.0-8.0. The thermostability of was determined and it was observed that the pectinase activity is heat stable and retains activity more than 40% when incubated at 90°C for 10 minutes. The pectinase activity of F3a and F3b was increased with different metal ions. The Pectinase activity was stimulated in the presence of CaCl2 up to 10-30%. ZnSO4, MnSO4 and Mg SO4 showed higher activity in fractions F3a and F3b, which indicates that the pectinase belongs to metalo-enzymes. It is concluded that A. niger is capable to produce pH stable and thermostable pectinase, which can be used for industrial purposes.

Keywords: pectinase, a. niger, production, purification, characterization

Procedia PDF Downloads 412

Authors: Uditi Dhakad, Baldev Ram, Chaman K. Jadon, R. K. Yadav, D. L. Yadav, Pratap Singh, Shalini Meena

Abstract:

A field experiment was conducted during Kharif 2021 at Agricultural Research Station, Kota, to evaluate the effect of different post-emergence herbicides applied to soybean [Glycine max (L.) Merrill] on soil enzymes activity viz. dehydrogenase, phosphatase, and urease. The soil of the experimental site was clay loam (vertisols) in texture and slightly alkaline in reaction with 7.7 pH. The soil was low in organic carbon (0.49%), medium in available nitrogen (210 kg/ha), phosphorus (23.5 P2O5 kg/ha), and high in potassium (400 K2O kg/ha) status. The results elucidated that no significant adverse effect on soil dehydrogenase, urease, and phosphatase activity was determined with the application of post-emergence herbicides over the untreated control. Two hands weeding at 20 and 40 DAS registered maximum dehydrogenase enzyme activity (0.329 μgTPF/g soil/d) closely followed by herbicides mixtures and sole herbicide while pre-emergence application of pendimethalin + imazethapyr 960 g a.i./ha and pendimethalin 1.0 kg a.i./ha significantly reduced dehydrogenase enzyme activity compared to control. Urease enzyme activity was not much affected under different weed control treatments and weedy checks. The treatments were found statistically non-significant, and values ranged between 1.16-1.25 μgNH4N/g soil/d. Phosphatase enzyme activity was also not influenced significantly due to various weed control treatments. Though maximum phosphatase enzyme activity (30.17 μgpnp/g soil/hr) was observed under two-hand weeding, followed by fomesafen + fluazifop-p-butyl 220 g a.i./ha. Herbicidal weed control measures did not influence the total bacteria, fungi, and actinomycetes population.

Keywords: dehydrogenase, phosphatase, post-emergence, soil enzymes, urease.

Procedia PDF Downloads 103

Authors: Md. Z. Alam, Devi R. Asih, Md. N. Salleh

Abstract:

Immobilization of lipase enzyme produced from palm oil mill effluent (POME) by the activated carbon (AC) among the low cost support materials was optimized. The results indicated that immobilization of 94% was achieved by AC as the most suitable support material. A sequential optimization strategy based on a statistical experimental design, including one-factor-at-a-time (OFAT) method was used to determine the equilibrium time. Three components influencing lipase immobilization were optimized by the response surface methodology (RSM) based on the face-centered central composite design (FCCCD). On the statistical analysis of the results, the optimum enzyme concentration loading, agitation rate and carbon active dosage were found to be 30 U/ml, 300 rpm and 8 g/L respectively, with a maximum immobilization activity of 3732.9 U/g-AC after 2 hrs of immobilization. Analysis of variance (ANOVA) showed a high regression coefficient (R2) of 0.999, which indicated a satisfactory fit of the model with the experimental data. The parameters were statistically significant at p<0.05.

Keywords: activated carbon, POME based lipase, immobilization, adsorption

Procedia PDF Downloads 241

Authors: Katalin Juhos, Enikő Papdi, Flórián Kovács, Vasileios P. Vasileiadis, Andrea Veres

Abstract:

Mulching techniques can be a solution for better utilization of precipitation and irrigation water and for mitigating soil degradation and drought damages. Waste fibres as alternative biodegradable mulch materials are increasingly coming to the fore. The effect of wool mulch (WM) on water use efficiency of pepper seedlings were investigated in different soil types (sand, clay loam, peat) in a pot experiment. Two semi-field experiments were also set up to investigate the effect of WM-plant interaction on sweet pepper yield in comparison with agro-textile and straw mulches. Soil parameters (moisture, temperature, DHA, β-glucosidase enzymes, permanganate-oxidizable carbon) were measured during the growing season. The effect of WM on yield and biomass was more significant with less frequent irrigation and the greater the water capacity of soils. The microbiological activity was significantly higher in the presence of plants, because of the water retention of WM, the metabolic products of roots and the more balanced soil temperature caused by plants. On the sandy soil, the straw mulch had a significantly better effect on microbiological parameters and yields than the agro-textile and WM. WM is a sustainable practice for improving soil biological parameters and water use efficiency on soils with a higher water capacity.

Keywords: β-glucosidase, DHA enzyme activity; labile carbon, straw mulch; plastic mulch, evapotranspira-tion coefficient, soil temperature

Procedia PDF Downloads 75

Authors: S. Lekhavat, T. Kajsongkram, S. Sang-han

Abstract:

Dried ginger was extracted and investigated the effect of ethanol concentration and enzyme pre-treatment on its bioactive compounds in solvent extraction process. Sliced fresh gingers were dried by oven dryer at 70 °C for 24 hours and ground to powder using grinder which their size were controlled by passing through a 20-mesh sieve. In enzyme pre-treatment process, ginger powder was sprayed with 1 % (w/w) cellulase and then was incubated at 45 °C for 2 hours following by extraction process using ethanol at concentration of 0, 20, 40, 60 and 80 % (v/v), respectively. The ratio of ginger powder and ethanol are 1:9 and extracting conditions were controlled at 80 °C for 2 hours. Bioactive compounds extracted from ginger, either enzyme-treated or non enzyme-treated samples, such as total phenolic content (TPC), 6-Gingerol (6 G), 6-Shogaols (6 S) and antioxidant activity (IC50 using DPPH assay), were examined. Regardless of enzyme treatment, the results showed that 60 % ethanol provided the highest TPC (20.36 GAE mg /g. dried ginger), 6G (0.77%), 6S (0.036 %) and the lowest IC50 (625 μg/ml) compared to other ratios of ethanol. Considering the effect of enzyme on bioactive compounds and antioxidant activity, it was found that enzyme-treated sample has more 6G (0.17-0.77 %) and 6S (0.020-0.036 %) than non enzyme-treated samples (0.13-0.77 % 6G, 0.015-0.036 % 6S). However, the results showed that non enzyme-treated extracts provided higher TPC (6.76-20.36 GAE mg /g. dried ginger) and Lowest IC50 (625-1494 μg/ml ) than enzyme-treated extracts (TPC 5.36-17.50 GAE mg /g. dried ginger, IC50 793-2146 μg/ml).

Keywords: antioxidant activity, enzyme, extraction, ginger

Procedia PDF Downloads 254

Authors: Zohreh Bayat, Dariush Minai-Tehrani

Abstract:

Lipases constitute one of the most important groups of industrial enzymes that catalyze the hydrolysis of triacylglycerol to glycerol and fatty acids. Muscarinic antagonist relieves smooth muscle spasm of the gastrointestinal tract and effect on the cardiovascular system. In this research, the effect of a muscarinic antagonist on the lipase activity of Pseudomonas aeruginosa was studied. Lineweaver–Burk plot showed that the drug inhibited the enzyme by competitive inhibition. The IC50 value (60 uM) and Ki (30 uM) of the drug revealed the drug bound to the enzyme with high affinity. Determination of enzyme activity in various pH and temperature showed that the maximum activity of lipase was at pH 8 and 60°C both in presence and absence of the drug.

Keywords: bacteria, inhibition, kinetics, lipase

Procedia PDF Downloads 452

Authors: Munisath Khandoker, Stephan Haefele, Andy Gregory

Abstract:

Extracellular enzymes play a key role in soil organic carbon (SOC) decomposition and nutrient cycling and are known indicators for soil health; however, it is not understood how these enzymes respond to different land uses and their relationships to other soil properties have not been extensively reviewed. The relationships among the activities of three soil enzymes: β-glucosaminidase (NAG), phosphomonoesterase (PHO) and β-glucosidase (GLU), were examined. The impact of soil organic amendments, soil types and land management on soil enzyme activities were reviewed, and it was hypothesized that soils with increased SOC have increased enzyme activity. Long-term experiments at Rothamsted Research Woburn and Harpenden sites in the UK were used to evaluate how different management practices affect enzyme activity involved in carbon (C) and nitrogen (N) cycling in the soil. Samples were collected from soils with different organic treatments such as straw, farmyard manure (FYM), compost additions, cover crops and permanent grass cover to assess whether SOC can be linked with increased levels of enzymatic activity and what influence, if any, enzymatic activity has on total C and N in the soil. Investigating the interactions of important enzymes with soil characteristics and SOC can help to better understand the health of soils. Studies on long-term experiments with known histories and large datasets can better help with this. SOC tends to decrease during land use changes from natural ecosystems to agricultural systems; therefore, it is imperative that agricultural lands find ways to increase and/or maintain SOC in the soil.

Keywords: biological soil health indicators, extracellular enzymes, soil health, soil, microbiology

Procedia PDF Downloads 70

Authors: Irina E. Sukovataya, Oleg S. Sutormin, Valentina A. Kratasyuk

Abstract:

Effect of viscosity of media on kinetic parameters of the coupled enzyme system NADH:FMN-oxidoreductase–luciferase was investigated with addition of organic solvents (glycerol and sucrose), because bioluminescent enzyme systems based on bacterial luciferases offer a unique and general tool for analysis of the many analytes and enzymes in the environment, research, and clinical laboratories and other fields. The possibility of stabilization and increase of activity of the coupled enzyme system NADH:FMN-oxidoreductase–luciferase activity in vicious aqueous-organic mixtures have been shown.

Keywords: coupled enzyme system of bioluminescence bacteria NAD(P)H:FMN-oxidoreductase–luciferase, glycerol, stabilization of enzymes, sucrose

Procedia PDF Downloads 393

Authors: Dina Helmy El-Ghonemy, Thanaa Hamed Ali

Abstract:

The aim of this study was to purify and characterize a keratinolytic enzyme produced by Aspergillus sp. DHE7 cultured in basal medium containing chicken feather as substrate. The enzyme was purified through ammonium sulfate saturation of 60%, followed by gel filtration chromatography in Sephadex G-100, with a 16.4-purification fold and recovery yield of 52.2%. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the purified enzyme is a monomeric enzyme with an apparent molecular mass of 30 kDa — the purified keratinase of Aspergillus sp. DHE7 exhibited activity in a broad range of pH (7- 9) and temperature (40℃-60℃) profiles with an optimal activity at pH eight and 50℃. The keratinolytic activity was inhibited by protease inhibitors such as phenylmethylsulfonyl fluoride and ethylenediaminetetraacetate, while no reduction of activity was detected by the addition of dimethyl sulfoxide (DMSO). Bivalent cations, Ca²⁺ and Mn²⁺, were able to greatly enhance the activity of keratinase by 125.7% and 194.8%, respectively, when used at one mM final concentration. On the other hand, Cu²⁺ and Hg²⁺ inhibited the enzyme activity, which might be indicative of essential vicinal sulfhydryl groups of the enzyme for productive catalysis. Furthermore, the purified keratinase showed significant stability and compatibility against the tested commercial detergents at 37ºC. Therefore, these results suggested that the purified keratinase from Aspergillus sp. DHE7 may have potential use in the detergent industry and should be of interest in the processing of poultry feather waste.

Keywords: Aspergillus sp. DHE7, biochemical characterization, keratinase, purification, waste management

Procedia PDF Downloads 123

Authors: Supriya Chatla, Anjana Male, Srikala Kamireddy

Abstract:

L-asparaginase (E.C.3.5.1.1) is an anti-cancer enzyme that has been purified and characterized for decades to study and evaluate its anti-carcinogenic activity against Hodgkin’s lymphoma. The present investigation deals with screening, isolation and optimization of L-asparaginase giving fungal strain of soil samples from different areas of AP, India. L-Aspariginase activity was estimated on the basis of the pink color surrounding the growing colony. A total of 132 colonies were screened and isolated from different samples. Based on the zone diameter, L-asparaginase activity is determined, L- asparaginase activity is optimized at 28oc and Immobilized Aspariginase had more potency than the free enzymes.

Keywords: aspariginase, anticancer enzyme, Isolation, optimization

Procedia PDF Downloads 77

Authors: Zohreh Bayat, Dariush Minai-Tehrani

Abstract:

Pseudomonas aeruginosa is a Gram-negative, rode shape and aerobic bacterium that has shown to be resistance to many antibiotics. This resistance makes the bacterium very harmful in some diseases. It can also generate diseases in any part of the gastrointestinal tract from oropharynx to rectum. P. aeruginosa has become an important cause of infection, especially in patients with compromised host defense mechanisms. One of the most important reasons that make P. aeruginosa an emerging opportunistic pathogen in patients is its ability to use various compounds as carbon sources. Lipase is an enzyme that catalyzes the hydrolysis of lipids. Most lipases act at a specific position on the glycerol backbone of lipid substrate. Some lipases are expressed and secreted by pathogenic organisms during the infection. Muscarinic antagonist used as an antispasmodic and in urinary incontinence. The drug has little effect on glandular secretion or the cardiovascular system. It does have some local anesthetic properties and is used in gastrointestinal, biliary, and urinary tract spasms. Aim: In this study the inhibitory effect of a muscarinic antagonist on lipase of P. aeruginosa was investigated. Methods: P. aeruginosa was cultured in minimal salt medium with 1% olive oil as carbon source. The cells were harvested and the supernatant, which contained lipase, was used for enzyme assay. Results: Our results showed that the drug can inhibit P. aeruginosa lipase by competitive manner. In the presence of different concentrations of the drug, the Vmax (2 mmol/min/mg protein) of enzyme did not change, while the Km raised by increasing the drug concentration. The Ki (inhibition constant) and IC50 (the half maximal inhibitory concentration) value of drug was estimated to be about 30 uM and 60 uM which determined that the drug binds to enzyme with high affinity. Maximum activity of the enzyme was observed at pH 8 in the absence and presence of muscarinic antagonist, respectively. The maximum activity of lipase was observed at 600C and the enzyme became inactive at 900C. Conclusion: The muscarinic antagonist drug could inhibit lipase of P. aeruginosa and changed the kinetic parameters of the enzyme. The drug binded to enzyme with high affinity and did not chang the optimum pH of the enzyme. Temperature did not affect the binding of drug to musmuscarinic antagonist.

Keywords: Pseudomonas aeruginosa, drug, enzyme, inhibition

Procedia PDF Downloads 433

Authors: Misha Ali, Qayyum Husain, Nida Alam, Masood Ahmad

Abstract:

Improving the activity and stability of the enzyme is an important aspect in bioremediation processes. Immobilization of enzyme is an efficient approach to amend the properties of biocatalyst required during wastewater treatment. The present study was done to immobilize partially purified ginger peroxidase on amino functionalized silica coated titanium dioxide nanocomposite. Interestingly there was an enhancement in enzyme activity after immobilization on nanosupport which was evident from effectiveness factor (η) value of 1.76. Immobilized enzyme was characterized by transmission electron microscopy, scanning electron microscopy and Fourier transform infrared spectroscopy. Immobilized peroxidase exhibited higher activity in a broad range of pH and temperature as compared to free enzyme. Also, the thermostability of peroxidase was strikingly improved upon immobilization. After six repeated uses, the immobilized peroxidase retained around 62% of its dye decolorization activity. There was a 4 fold increase in Vmax of immobilized peroxidase as compared to free enzyme. Circular dichroism spectroscopy demonstrated conformational changes in the secondary structure of enzyme, a possible reason for the enhanced enzyme activity after immobilization. Immobilized peroxidase was highly efficient in the removal of acid yellow 42 dye in a stirred batch process. Our study shows that this bio-remediating system has remarkable potential for treatment of aromatic pollutants present in wastewater.

Keywords: acid yellow 42, decolorization, ginger peroxidase, immobilization

Procedia PDF Downloads 247

Authors: Sumayah Mohammed Asiri A., Aviva Levina B., Elizabeth New C., Peter Lay D.

Abstract:

Pt compounds have been among the most successful anticancer drugs in the last 40 years, but the development of resistance to them is an increasing problem. Cellular homeostasis of an essential metal, Cu, is known to be involved in Pt resistance, but mechanisms of this process are poorly understood. We used a novel ratiometric Cu(I)-sensitive fluorescent probeInCCu1 dye to detect Cu(I) in the mitochondria. Total Cu and labile Cu pool measured using AAS and InCCu1 dye in A2780 cells and their corresponding resistant cells A2780-cis.R cells treated with Cu and cisplatin. The main difference between both cell lines in the presence and absence of Cu(II) is that resistant cells have lower total Cu content but higher labile Cu levels than cisplatin-sensitive cells. This means that resistant cells can metabolize and export excess Cu more efficiently. Furthermore, InCCu1 has emerged not only as an indicator of labile cellular Cu levels in the mitochondria but as a potentially versatile multi-organelle probe.

Keywords: AAS and ICPMS, A2780 and its resistant cells, ratiometric fluorescent sensors, inCCu1, and total and labile Cu

Procedia PDF Downloads 211

Authors: Ashish Kumar Singh, Mehak Gulati, Neelam Verma

Abstract:

Arginine majorly acts as a substrate for the enzyme nitric oxide synthase (NOS) for the production of nitric oxide, a strong vasodilator. Current study demonstrated a novel amperometric approach for estimation of arginine using nitric oxide synthase. The enzyme was co-immobilized in carbon paste electrode with NADP+, FAD and BH4 as cofactors. The detection principle of the biosensor is enzyme NOS catalyzes the conversion of arginine into nitric oxide. The developed biosensor could able to detect up to 10-9M of arginine. The oxidation peak of NO was observed at 0.65V. The developed arginine biosensor was used to monitor arginine content in fruit juices.

Keywords: arginine, biosensor, carbon paste elctrode, nitric oxide

Procedia PDF Downloads 422

Authors: A. K. M. Kafi, S. N. Nina, Mashitah M. Yusoff

Abstract:

In this work, we have described a new 3-dimensional (3D) network of crosslinked Horseradish Peroxidase/Carbon Nanotube (HRP/CNT) on a thiol-modified Au surface in order to build up the effective electrical wiring of the enzyme units with the electrode. This was achieved by the electropolymerization of aniline-functionalized carbon nanotubes (CNTs) and 4-aminothiophenol -modified-HRP on a 4-aminothiophenol monolayer-modified Au electrode. The synthesized 3D HRP/CNT networks were characterized with cyclic voltammetry and amperometry, resulting the establishment direct electron transfer between the redox active unit of HRP and the Au surface. Electrochemical measurements reveal that the immobilized HRP exhibits high biological activity and stability and a quasi-reversible redox peak of the redox center of HRP was observed at about −0.355 and −0.275 V vs. Ag/AgCl. The electron transfer rate constant, KS and electron transfer co-efficient were found to be 0.57 s-1 and 0.42, respectively. Based on the electrocatalytic process by direct electrochemistry of HRP, a biosensor for detecting H2O2 was developed. The developed biosensor exhibits excellent electrocatalytic activity for the reduction of H2O2. The proposed biosensor modified with HRP/CNT 3D network displays a broader linear range and a lower detection limit for H2O2 determination. The linear range is from 1.0×10−7 to 1.2×10−4M with a detection limit of 2.2.0×10−8M at 3σ. Moreover, this biosensor exhibits very high sensitivity, good reproducibility and long-time stability. In summary, ease of fabrication, a low cost, fast response and high sensitivity are the main advantages of the new biosensor proposed in this study. These obvious advantages would really help for the real analytical applicability of the proposed biosensor.

Keywords: biosensor, nanomaterials, redox enzyme, thiol-modified Au surface

Procedia PDF Downloads 329

Authors: Surasak Siripornadulsil, Nutt Poomai, Wilailak Siripornadulsil

Abstract:

Due to a high ethanol demand, the approach for effective ethanol production is important and has been developed rapidly worldwide. Several agricultural wastes are highly abundant in celluloses and the effective cellulose enzymes do exist widely among microorganisms. Accordingly, the cellulose degradation using microbial cellulose to produce a low-cost substrate for ethanol production has attracted more attention. In this study, the cellulose producing bacterial strain has been isolated from rich straw and identified by 16S rDNA sequence analysis as Acinetobacter sp. KKU44. This strain is able to grow and exhibit the cellulose activity. The optimal temperature for its growth and cellulose production is 37 °C. The optimal temperature of bacterial cellulose activity is 60 °C. The cellulose enzyme from Acinetobacter sp. KKU44 is heat-tolerant enzyme. The bacterial culture of 36 h. showed highest cellulose activity at 120 U/mL when grown in LB medium containing 2% (w/v). The capability of Acinetobacter sp. KKU44 to grow in cellulosic agricultural wastes as a sole carbon source and exhibiting the high cellulose activity at high temperature suggested that this strain could be potentially developed further as a cellulose degrading strain for a production of low-cost substrate used in ethanol production.

Keywords: cellulose enzyme, bagasse, rice straw, rice husk, acinetobacter sp. KKU44

Procedia PDF Downloads 312

Authors: Khaled M. Khleifat

Abstract:

An extracellular methanol and ethanol tolerant lipase producing bacterial strain K5b4 was isolated from soil samples contaminated with hydrocarbon residues. It was identified by using morphological and biochemical characteristics and 16srRNA technique as Acinetobacter species. The immobilized lipase from Acinetobacter sp. K5b4 retained more than 98% of its residual activity after incubation with pure methanol and ethanol for 24 hours. The highest hydrolytic activity of the immobilized enzyme was obtained in the presence of 75% (v/v) methanol in the assay solution. In contrary, the enzyme was able to maintain its original activity up to only 25% (v/v) ethanol whereas at elevated concentrations of 50 and 75% (v/v) the enzyme activity was reduced to 10 and 40%, respectively. Maximum lipase activity of 31.5 mU/mL was achieved after 48 hr cultivation when the optimized medium (pH 7.0) that composed of 1.0% (w/v) olive oil, 0.2% (w/v) glycerol, 0.15% (w/v) yeast extract, and 0.05% (w/v) NaCl was inoculated with 0.4% (v/v) seed culture and incubated at 30°C and 150 rpm agitation speed. However, the presence of CaCl2 in the growth media did not show any inhibitory or stimulatory effect on the enzyme production as it compared to the control experiment. Meanwhile, the other mineral salts MgCl2, MnCl2, KCl and CoCl2 were negatively affected the production of lipase enzyme. The inhibition of lipase production from Acinetobacter sp. K5b4 in presence of glucose suggesting that lipase gene expression is prone to catabolic repression.

Keywords: K5B4, methanol and ethanol, acinetobacter, morphological

Procedia PDF Downloads 316

Authors: Shweta Sinha, Mukesh Doble, Manju S. L.

Abstract:

Pyrazole scaffold is an important group of compound in heterocyclic chemistry and is found to possess numerous uses in chemistry. Pyrazole derivatives are also known to possess important biological activities including antitumor, antimicrobial, antiviral, antifungal, anticancer and anti-inflammatory. Inflammation is associated with pain, allergy and asthma. Leukotrienes are mediators of various inflammatory and allergic disorders. 5-Lipoxygenase (5-LOX) is an important enzyme involved in the biosynthesis of leukotrienes and metabolism of arachidonic acid (AA) and thus targeted for anti-inflammation. In vitro inhibitory activity of pyrazol-3-yl thiazole 4-carboxylic acid derivatives is tested against enzyme 5-LOX. Most of these compounds exhibit good inhibitory activity against this enzyme. Binding mode study of these compounds is determined by computational tool. Further experiments are being done to understand the mechanism of action of these compounds in inhibiting this enzyme. To conclude, these compounds appear to be a promising target in drug design against 5-LOX.

Keywords: inflammation, inhibition, 5-lipoxygenase, pyrazole

Procedia PDF Downloads 240

Authors: Justina I. R. Udotong, Essien U. Essien

Abstract:

Diagnostic enzymes like aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) were determined as indices of heavy metal pollution in Tilapia guinensis. Three different sets of fishes treated with lead (Pb), iron (Fe) and copper (Cu) were used for the study while a fourth group with no heavy metal served as a control. Fishes in each of the groups were exposed to 2.65 mg/l of Pb, 0.85 mg/l of Fe and 0.35 mg/l of Cu in aerated aquaria for 96 hours. Tissue fractionation of the liver tissues was carried out and the three diagnostic enzymes (AST, ALT, and ALP) were estimated. Serum levels of the same diagnostic enzymes were also measured. The mean values of the serum enzyme activity for ALP in each experimental group were 19.5±1.62, 29.67±2.17 and 1.15±0.27 IU/L for Pb, Fe and Cu groups compared with 9.99±1.34 IU/L enzyme activity in the control. This result showed that Pb and Fe caused increased release of the enzyme into the blood circulation indicating increased tissue damage while Cu caused a reduction in the serum level as compared with the level in the control group. The mean values of enzyme activity obtained in the liver were 102.14±6.12, 140.17±2.06 and 168.23±3.52 IU/L for Pb, Fe and Cu groups, respectively compared to 91.20±9.42 IU/L enzyme activity for the control group. The serum and liver AST and ALT activities obtained in Pb, Fe, Cu and control groups are reported. It was generally noted that the presence of the heavy metal caused liver tissues damage and consequent increased level of the diagnostic enzymes in the serum.

Keywords: diagnostic enzymes, enzyme activity, heavy metals, tissues investigations

Procedia PDF Downloads 287

Authors: T. H. Derdzyan, K. A. Ghazaryan, G. A. Gevorgyan

Abstract:

Beta-glucosidase, chitinase, leucine-aminopeptidase, acid phosphomonoestearse and acetate-esterase enzyme activities in the soils under the impact of metallurgical industrial activity in Lori marz (district) were investigated. The results of the study showed that the activities of the investigated enzymes in the soils decreased with increasing distance from the Shamlugh copper mine, the Chochkan tailings storage facility and the ore transportation road. Statistical analysis revealed that the activities of the enzymes were positively correlated (significant) to each other according to the observation sites which indicated that enzyme activities were affected by the same anthropogenic factor. The investigations showed that the soils were polluted with heavy metals (Cu, Pb, As, Co, Ni, Zn) due to copper mining activity in this territory. The results of Pearson correlation analysis revealed a significant negative correlation between heavy metal pollution degree (Nemerow integrated pollution index) and soil enzyme activity. All of this indicated that copper mining activity in this territory causing the heavy metal pollution of the soils resulted in the inhabitation of the activities of the enzymes which are considered as biological catalysts to decompose organic materials and facilitate the cycling of nutrients.

Keywords: Armenia, metallurgical industrial activity, heavy metal pollutionl, soil enzyme activity

Procedia PDF Downloads 295

Authors: Khalida Boutemak, Nasssima Benali, Nadji Moulai-Mostefa

Abstract:

Effect of enzyme on the yield and chemical composition of Artemisia campestris essential oil is reported in the present study. It was demonstrated that enzyme facilitated the extraction of essential oil with increase in oil yield and did not affect any noticeable change in flavour profile of the volatile oil. Essential oil was tested for antibacterial activity using Escherichia coli; which was extremely sensitive against control with the largest inhibition (29mm), whereas Staphylococcus aureus was the most sensitive against essential oil obtained from enzymatic pre-treatment with the largest inhibition zone (25mm). The antioxidant activity of the essential oil with hemicellulase pre-treatment (EO2) and control sample (EO1) was determined through reducing power. It was significantly lower than the standard drug (vitamin C) in this order: vitamin C˃EO2˃EO1.

Keywords: Artemisia campestris, enzyme pre-treatment, hemicellulase, antibacterial activity, antioxidant activity

Procedia PDF Downloads 328

Authors: Mulanga Luscious Mulaudzi, Ignatious Ncube

Abstract:

In nature, enzymatic degradation of lignocellulose is more efficient compared to in vivo bioprocessing. Thus, a co-culture should enable production of more efficient enzyme preparations that would mimic the natural decomposition of lignocellulose. The aim of the study was to establish a microbial co-culture for the production of highly active cellulase preparations. The objectives were to determine the use of a variety of culture media to isolate cellulose degrading microorganisms from decomposing banana pseudo stem and to optimize production of cellulase by co-cultures of microorganisms producing high levels of cellulose. Screening of fungal isolates was done on carboxylmethylcellulose agar plates which were stained with Congo red to show hydrolytic activity of the isolates. Co-culture and mixed culture of these microorganisms were cultured using Mendel salts with Avicel as the carbon source. Cultures were incubated at 30 °C with shaking at 200 rpm for 240 hrs. Enzyme activity assays were performed to determine endoglycosidase and β-glucosidase. Mixed culture of fungi-dead bacterial cells showed to be the best co-culture/ mixed culture to produce higher levels of cellulase activity in submerged fermentations (SmF) using Avicel™ as a carbon source. The study concludes use microorganism 5A in co-cultures is highly recommended in order to produce high amounts of β-glucosidases, no matter the combination used.

Keywords: avicel, co-culture, submerged fermentation, pseudostem

Procedia PDF Downloads 121

Authors: A. K. M. Kafi, S. N. Nina, Mashitah M. Yusoff

Abstract:

In this work, we have described a new 3-dimensional (3D) network of cross-linked Horseradish Peroxidase/Carbon Nanotube (HRP/CNT) on a thiol-modified Au surface in order to build up the effective electrical wiring of the enzyme units with the electrode. This was achieved by the electropolymerization of aniline-functionalized carbon nanotubes (CNTs) and 4-aminothiophenol -modified-HRP on a 4-aminothiophenol monolayer-modified Au electrode. The synthesized 3D HRP/CNT networks were characterized with cyclic voltammetry and amperometry, resulting the establishment direct electron transfer between the redox active unit of HRP and the Au surface. Electrochemical measurements reveal that the immobilized HRP exhibits high biological activity and stability and a quasi-reversible redox peak of the redox center of HRP was observed at about −0.355 and −0.275 V vs. Ag/AgCl. The electron transfer rate constant, KS and electron transfer co-efficient were found to be 0.57 s-1 and 0.42, respectively. Based on the electrocatalytic process by direct electrochemistry of HRP, a biosensor for detecting H2O2 was developed. The developed biosensor exhibits excellent electrocatalytic activity for the reduction of H2O2. The proposed biosensor modified with HRP/CNT 3D network displays a broader linear range and a lower detection limit for H2O2 determination. The linear range is from 1.0×10−7 to 1.2×10−4M with a detection limit of 2.2.0×10−8M at 3σ. Moreover, this biosensor exhibits very high sensitivity, good reproducibility and long-time stability. In summary, ease of fabrication, a low cost, fast response and high sensitivity are the main advantages of the new biosensor proposed in this study. These obvious advantages would really help for the real analytical applicability of the proposed biosensor.

Keywords: redox enzyme, nanomaterials, biosensors, electrical communication

Procedia PDF Downloads 454

Authors: J. B. Minari, O. B. Oloyede, A. A. Odutuga

Abstract:

Myeloperoxidase is the most abundant enzyme found in the polymorphonuclear neutrophil and is known to play a central role in the host defense system of the leukocyte. The enzyme has been reported to interact with some drugs to generate free radical which inhibits its activity. This study investigated the effects of artesunate on the activity of the enzyme and the subsequent effect on the host immune system. In investigating the effects of the drugs on myeloperoxidase, the influence of concentration, pH, partition ratio estimation and kinetics of inhibition were studied. This study showed that artesunate is concentration-dependent inhibitor of myeloperoxidase with an IC50 of 0.078mM. Partition ratio estimation showed that 60 enzymatic turnover cycles are required for complete inhibition of myeloperoxidase in the presence of artesunate. The influence of pH on the effect of artesunate on the enzyme showed least activity of myeloperoxidase at physiological pH. The kinetic inhibition studies showed that artesunate caused a competitive inhibition with an increase in the Km value from 0.12mM to 0.26mM and no effect on the Vmax value. The Ki value was estimated to be 2.5mM. The results obtained from this study show that artesunate is a potent inhibitor of myeloperoxidase and it is capable of inactivating the enzyme. It is considered that the inhibition of myeloperoxidase in the presence of artesunate as revealed in this study may partly explain the impairment of polymorphonuclear neutrophil and consequent reduction of the strength of the host defense system against secondary infections.

Keywords: myeloperoxidase, artesunate, inhibition, nuetrophill

Procedia PDF Downloads 362

Authors: Kanchan Vishwakarma, Shivesh Sharma, Nitin Kumar

Abstract:

Soil is incoherent matter on Earth’s surface having organic and mineral content. The spatial variation of 4 soil enzyme activities and microbial biomass were assessed for two seasons’ viz. monsoon and winter along the latitudinal gradient in North-central India as the area of this study is fettered with respect to national status. The study was facilitated to encompass the effect of climate change, enzyme activity and biomass on nutrient cycling. Top soils were sampled from 4 sites in North-India. There were significant correlations found between organic C, N & P wrt to latitude gradient in two seasons. This distribution of enzyme activities and microbial biomass was consequence of alterations in temperature and moisture of soil because of which soil properties change along the latitude transect.

Keywords: latitude gradient, microbial biomass, moisture, soil, organic carbon, temperature

Procedia PDF Downloads 395

Authors: Flackson Tshuma, James Bennett, Pieter Andreas Swanepoel, Johan Labuschagne, Stephan van der Westhuizen, Francis Rayns

Abstract:

Amongst other things, tillage and synthetic agrochemicals can be effective methods of seedbed preparation and pest control. Nonetheless, frequent and intensive tillage and excessive application of synthetic agrochemicals, such as herbicides and insecticides, can reduce soil microbial enzyme activity. A decline in soil microbial enzyme activity can negatively affect nutrient cycling and crop productivity. In this study, the effects of four tillage treatments; continuous mouldboard plough; shallow tine-tillage to a depth of about 75 mm; no-tillage; and tillage rotation (involving shallow tine-tillage once every four years in rotation with three years of no-tillage), and two rates of synthetic agrochemicals (standard: with regular application of synthetic agrochemicals; and reduced: fewer synthetic agrochemicals in combination with bio-chemicals/ or bio-stimulants) on soil microbial enzyme activity were investigated between 2018 and 2020 in a typical Mediterranean climate zone in South Africa. Four different bio-stimulants applied contained: Trichoderma asperellum, fulvic acid, silicic acid, and Nereocystis luetkeana extracts, respectively. The study was laid out as a complete randomised block design with four replicated blocks. Each block had 14 plots, and each plot measured 50 m x 6 m. The study aimed to assess the combined impact of tillage practices and reduced rates of synthetic agrochemical application on soil microbial enzyme activity in a dryland cropping system. It was hypothesised that the application of bio-stimulants in combination with minimum soil disturbance will lead to a greater increase in microbial enzyme activity than the effect of applying either in isolation. Six soil cores were randomly and aseptically collected from each plot for microbial enzyme activity analysis from the 0-150 mm layer of a field trial under a dryland crop rotation system in the Swartland region. The activities of four microbial enzymes, β-glucosidase, acid phosphatase, alkaline phosphatase and urease, were assessed. The enzymes are essential for the cycling of glucose, phosphorus, and nitrogen, respectively. Microbial enzyme activity generally increased with a reduction of both tillage intensity and synthetic agrochemical application. The use of the mouldboard plough led to the least (P<0.05) microbial enzyme activity relative to the reduced tillage treatments, whereas the system with bio-stimulants (reduced synthetic agrochemicals) led to the highest (P<0.05) microbial enzyme activity relative to the standard systems. The application of bio-stimulants in combination with reduced tillage, particularly no-tillage, could be beneficial for enzyme activity in a dryland farming system.

Keywords: bio-stimulants, soil microbial enzymes, synthetic agrochemicals, tillage

Procedia PDF Downloads 80

Authors: Yves Mann Elate Lea Mbassi, Marie Solange Evehe Bebandoue, Wilfred Fon Mbacham

Abstract:

Improving the catalytic performance of enzymes has been a long-standing theme of analytical biochemistry research. Induction of peroxidase activity by metals is a common reaction in higher plants. We thought that this increase in peroxidase activity may be due, on the one hand, to the stimulation of the gene expression of these enzymes but also to a modification of their chemical reactivity following the binding of some metal ions on their active site. We tested the effect of some metal salts (MgCl₂, MnCl₂, ZnCl₂, CaCl₂ and CuSO₄) on the activity and thermostability of peroxidase A6, a thermostable peroxidase that we discovered and purified in a previous study. The chromogenic substrate used was 3,3′,5,5′-tetramethylbenzidine. Of all the metals tested for their effect on A6, only magnesium and copper had a significant effect on the activity of the enzyme at room temperature. The Mann-Whitney test shows a slight inhibitory effect of activity by the magnesium salt (P = 0.043), while the activity of the enzyme is 5 times higher in the presence of the copper salt (P = 0.002). Moreover, the thermostability of peroxidase A6 is increased when calcium and copper salts are present. The activity in the presence of CaCl₂ is 8 times higher than the residual activity of the enzyme alone after incubation at 80°C for 10 min and 35 times higher in the presence of CuSO4 under the same conditions. In addition, manganese and zinc salts slightly reduce the thermostability of the enzyme. The activity and structural stability of peroxidase A6 can clearly be activated by Cu₂+, which therefore enhance the oxidation of 3,3′,5,5′-tetramethylbenzidine, which was used in this study as a chromogenic substrate. Ca₂+ likely has a more stabilizing function for the catalytic site.

Keywords: peroxidase activity, copper ions, calcium ions, thermostability

Procedia PDF Downloads 71