Enzymatic Parameters Research Articles

ANALYSIS OF OXIDATIVE STRESS MARKERS IN CROSSBRED COWS WITH SUBCLINICAL MASTITIS AND CONCURRENT METABOLIC AND INFECTIOUS DISEASES

The aim of this study was to evaluate the oxidative status in crossbred cows with subclinical mastitis and concurrent metabolic and infectious diseases. The crossbred cows were groped asGp-C (n=6) as controls,Gp-M (n=6) with clinical mastitis ,Gp-TS consists cows affected withstenosis of teat,Gp-RA with ruminal acidosis,Gp-ND with diarrhea along with dehydration and loss of appetite, Gp-RTIwith coughing, fever, decreased appetite, varying degrees of dyspnea, GpRB with a history of failure to conceive after at least two successive inseminations. oxidative stress parameters likecatalase, SOD, GPx, GSH and malondialdehyde were estimated spectrophotometrically. The antioxidant enzymes like catalase, super oxide dismutase (SOD), Glutathione Peroxidase (GPx) levels were significantly declined and the non enzymatic parameters reduced glutathione (GSH) and malondialdehyde (MDA) are significantly elevated in affected animals than in healthy animals. The results of the oxidative stress parameters indicate imbalance of antioxidant profile and oxidative stress in the animals with various disorders and previous illness along with subclinical mastitis.

Cholinesterase from the Liver of Diodon hystrix for Detection of Metal Ions

The discharge of industrial effluents into nearby water bodies affects the inhabitants including living organisms. The presence of foreign materials such as heavy metals can be a threat to the ecosystem as they are enormously carcinogenic even though in minute concentration. Hence, an economical and time-efficient preliminary screening test is crucial to be developed for the detection of heavy metals, prior to employment of high technology instruments. In this study, cholinesterase (ChE) from Sabah porcupine fish, Diodon hystrix was purified to test for its potential as an alternative biosensor in detecting metal ions. Few enzymatic parameters including specificity of substrate, temperature and pH were applied to determine its optimal enzymatic activity. ChE enzyme was found to be more sensitive towards the presence of substrate, butyrylthiocholine iodide (BTC), in contrast to acetylthiocholine iodide (ATC) and propionylthiocholine iodide (PTC) with the effective coefficient at 7193, 3680.15 and 2965.26 Vmax/Km, respectively. Moreover, the extracted ChE enzyme showed the optimum activity at pH 9 of 0.1 M Tris-HCl and at 25°C to 30°C range of temperature. When subjected to heavy metals, ChE enzyme was significantly inhibited as the enzyme activity was reduced in the sequence of Hg > Ag > Cr > Cu > Cd > Pb ≥ Zn > As. As a conclusion, the partially purified ChE enzyme proved its sensitivity towards metal ion exposure and can be used as an alternative method in screening the level of contamination in the environment.

Responses of metabolic enzymes (GOT, GPT and LDH) in an Indian major carp Cirrhinus mrigala exposed to titanium dioxide (TiO2 ) nanorods under short-term exposure

Titanium dioxide (TiO2) nanoparticles are extensively manufactured due to their potential properties and applications invarious fields such as biomedical, electrical and environmental. These particles are likely to reach the aquatic environmentand may cause adverse effects on aquatic organisms. In this study, we investigated the effects of different concentrations (1, 50 and 100 mg L-l) of TiO2 nanorods (NRs). The enzymatic activity of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), and lactate dehydrogenase (LDH) was measured in the liver and muscle of an Indian major carp, Cirrhinus mrigala, under short-term exposure (96 h). The synthesised particles were characterized using X-ray diffraction (XRD), scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR), UV-Visspectroscopy (UV-Vis) and photoluminescence (PL) techniques before conducting the toxicity assay. The GOT and GPT activities were significantly elevated in both liver and muscle of fish treated with TiO2 nanorods (except 50 mg L-l in muscle for GPT). Similarly, the activity of LDH was also found to be elevated. The findings of the present investigation suggest that TiO2:NRs might have be absorbed, circulated, accumulated in liver and muscles of C. mrigala resulting in alterations in the enzyme activities. The results revealed that TiO2 nanorods induced alterations in GOT, GPT and LDH activities of fish at tested concentrations. The alterations of these enzymatic parameters can be useful for monitoring the environmental contamination of titanium dioxide (TiO2) nanoparticles in freshwater ecosystem.

Pharmacological impact of Agaricus bisporus extract in carbon tetrachloride-induced hepatotoxicity in rats

Drug-induced hepatotoxicity is a frequent cause of liver injury worldwide. The present study was designed to evaluate the possible hepato-protective potential Agaricus bisporus extract (ABE; a type of mushrooms) in albino rats using Carbon tetrachloride (CCl4)-model of liver injury. Forty-two albino rats were utilized in this experiment arranged randomly in seven groups, six rats each, of different treatments. He-patic injury model was induced by administration of CCl4 (25% in corn oil) at a dose of 2.5 ml/kg, interperitoneally, twice weekly for 8 weeks (+ve control); test group rats received ABE at escalating doses of 200 or 400 mg/kg, orally, daily for 8 weeks with exposure to CCl4; standard group rats received Silymarin at dose of 100 mg/kg, orally, daily for 8 weeks along with CCl4; further 2 groups of rats received only ABE at the same dose levels; while rats of -ve control group received only the vehicles of the used drugs. Blood and liver tissue sam-ples were picked out at the end of the experimental course for different assays. Biochemical analysis revealed that ABE exhibited dose-dependent hepatoprotection indicated by almost normalized biomarkers, including, enzymatic liver function parameters, namely, AST, ALT, GGT & ALP with potential % of 93.1, 58.2, 65.2, 68.9, respectively, after ABE large dose when standardized by Silymarin; non-enzymatic parameters, namely, total protein, albumin, globulins, total bilirubin, conjugated bilirubin, unconjugated bilirubin, TAGs, Cholesterol, HDL, LDL & VLDL with potential % of 59.3, 54.5, 57.3, 81.8, 81.0, 80.0, 75.5, 90.4, 80.8, 84.5 & 78.7, respectively, after ABE large dose when standardized by Silymarin. The mechanism of the obtained hepatoprotection of ABE may be based on impeding the oxidative stress mediated by the used hepatotoxin, indicated by reduced MDA (37.9 % of Silymarin), and restored SOD, Catalase & GPx in liver homogenate with potentials of 94.9, 63.0 & 88.4 % of Silymarin, respectively. Pathological findings, both macroscopic and microscopic, were supportive to the biochemical findings, where the pathological lesions caused by CCl4 as fatty degeneration of hepatocytes with vacuolated cytoplasm, proliferated fibrous connective tissue with eosinophilic edematous fluid cells plus focal and diffuse necrotic areas and hyperplastic biliary epithelium, were ameliorated dose-dependently when ABE was administered together with CCl4. Data of the present study may suggest ABE as a good natural source for promising hepatoprotective and antioxidative remedies.

Study on enzymatic hydrolysis of soybean β-conglycinin using alkaline protease from Bacillus subtilis ACCC 01746 and antigenicity of its hydrolysates

Due to its beneficial health effects, the use of soybean protein has shown a continuous increase, but concerns regarding the allergenicity of soybean antigenic protein have also increased. This study aimed to evaluate the hydrolytic effects of a non-commercial alkaline protease isolated from the Bacillus subtilis ACCC 01746 on soybean β-conglycinin and the allergenicity of its hydrolysates. Alkaline protease of the strain was separated by precipitation method of organic solvents, and the β-conglycinin was separated by alkali-solution and acid-isolation and purified by use of gel column. Using the degree of hydrolysis (DH) and inhibition rate as evaluation indexes, the enzymatic hydrolysis parameters of β-conglycinin was optimized by single factor and L9(34) orthogonal tests, so as to explore the effect of the protease on the hydrolysis degree and the antigenicity of β-conglycinin hydrolysates. The results showed that the native enzyme existed as an 18.3 kDa monomer with a 430 U/g maximum activity. The purity of β-conglycinin was 84.8%. The single-factor test results showed that DH showed the oppostie trendency with the inhibition rate, and the increase of protein concentration caused monotone increasing and monotone decreasing of the inhibition rate and the DH, and the optimal protein concentration was 30 mg/mL. The optimization results showed that pH had the largest impacts on both DH and the inhibition rate, followed by enzyme dosage, hydrolysis temperature and hydrolysis time. Under the optimum hydrolysis conditions of protein concentration 30 mg/mL, enzyme dosage 0.7%, hydrolysis time 40 min, temperature 55 °C and pH 8.5, the DH reached the highest of 76.28%, and the inhibition rate was the lowest of 27.03%, which was reduced greatly compared with that before optimization. These results suggested that alkaline protease appeared to show a relatively high effeciency in lowering soybean allergenicity, making it possible to produce low-allergenicity soybean protein.

Salicylic acid mediated up regulation of carvone biosynthesis during growth phase in cell suspension cultures of Anethum graveolens.

The study illustrates the system for enhanced production of a medicinally important unexplored compound, carvone occurring naturally in Anethum graveolens. The effect of salicylic acid (SA) on biomass yield, carvone biosynthesis, growth and major enzymatic antioxidant parameters in A. graveolens was evaluated. The effects of different combinations of benzyl adenine (BA) and 1-Naphthalene acetic acid (NAA) were tested. Murashige and Skoog (MS) medium comprising 1.76µM BA + 3.24µM NAA was the best for friable callus induction. The friable callus was used for the initiation of cell suspension culture. MS salts in combination with 4.4µM BA and 2.6µM NAA, 3% sucrose was appropriate for cell growth and bioactive compound accumulation. The cell suspension cultures were then treated with SA (0.1, 0.75 and 1.5mM) as an elicitor for four weeks. An up regulation of enzymatic antioxidants, ascorbate peroxidase (APX); superoxide dismutase (SOD) and catalase (CAT) activity with increasing concentrations of SA whereas a reduction in guaiacol peroxidase (GPX) activity was recorded at the end of the growth phase. The results also showed that higher concentrations of SA significantly increased malondialdehyde (MDA) and Proline content. Cell suspension culture was then subjected to extraction and isolation. The quantification of carvone through HPLC analysis revealed highest amount of carvone (0.063%) in cell suspension culture treated with 0.1mM concentration of SA whereas higher concentration 0.75mM SA showed reduction in amount (0.035%) of carvone. SA elicited cell suspension culture offered an effective and favorable invitro method to improve the production of carvone for its potential use in pharmaceuticals.

Pimpinella anisum Essential Oil Nanoemulsion Toxicity against Tribolium castaneum? Shedding Light on Its Interactions with Aspartate Aminotransferase and Alanine Aminotransferase by Molecular Docking.

The insecticidal activity is the result of a series of complex interactions between toxic substances as ligands and insect’s enzymes as targets. Actually, synthetic insecticides used in pest control programs are harmful to the environment and may affect non-target organisms; thus, the use of natural products as pest control agents can be very attractive. In the present work, the toxic effect of aniseed (Pimpinella anisum L.) essential oil (EO) and its nanoemulsion (NE) against the red flour beetle Tribolium castaneum, has been evaluated. To assess the EO mode of action, the impact of sub-lethal concentrations of aniseed EO and NE was evaluated on enzymatic and macromolecular parameters of the beetles, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein, total lipids and glucose. Finally, a molecular docking study was conducted to predict the mode of action of the major EO and NE components namely E-anethole, Limonene, alpha-himalachalene, trans-Verbenol and Linalool at binding site of the enzymes AST and ALT. Herein, the binding location of the main compounds in both proteins are discussed suggesting the possible interactions between the considered enzymes and ligands. The obtained results open new horizons to understand the evolution and response of insect-plant compounds interactions and their effect predicted at the molecular levels and side effects of both animal and human.

Dual electrophoretically-mediated microanalysis in multiple injection mode for the simultaneous determination of acetylcholinesterase and α-glucosidase activity applied to selected polyphenols

Acetylcholinesterase (AChE) and α-glucosidase (α-glu) are key target enzymes in the search for novel strategies in the treatment of Alzheimer's disease and type II diabetes. Therefore, methods to assess the enzyme inhibition are of great value in the research field. Here is proposed a novel a dual electrophoretically-mediated microanalysis for the simultaneous determination of both enzymes' activity. In order to do so, the various solutions required for both assays were introduced in the capillary electrophoresis system using the multiple injections approach. Enzymatic kinetic parameters were tested, Km for AChE and α-glu were 3.81 and 0.43 mmol L−1. Ki values were 4.27 μmol L−1 for neostigmine (an AChE inhibitor) and 0.40 mmol L−1 for acarbose (an α-glu inhibitor). Results of IC50 (concentration for 50% of inhibition) were 5.11 ± 0.47 μmol L−1 and 0.58 ± 0,02 mmol L−1 for neostigmine and acarbose, respectively. All parameters (except for Ki of neostigmine) were comparable with the literature, indicating a good reliability of the proposed method to evaluate these enzymes activity. Total time analysis was approximately 10 min, being possible to perform around 12 enzymatic assays per hour, with low sample and reagent consumption, thus satisfying some of the principles of green chemistry. The method was applied to evaluate 10 phenolic compounds, of wich p-coumaric acid showed the best inhibitory activity for AChE (40.14 ± 4.75% at 10 mg L−1); and quercetin for α-glu (46.53 ± 4.90% at 10 mg L−1).

Purification of sarcoplasmic reticulum vesicles from horse gluteal muscle

We have analyzed protein expression and enzyme activity of the sarcoplasmic reticulum Ca2+-transporting ATPase (SERCA) in horse gluteal muscle. Horses exhibit a high incidence of recurrent exertional rhabdomyolysis, with myosolic Ca2+ proposed, but yet to be established, as the underlying cause. To better assess Ca2+ regulatory mechanisms, we developed an improved protocol for isolating sarcoplasmic reticulum (SR) vesicles from horse skeletal muscle, based on mechanical homogenization and optimized parameters for differential centrifugation. Immunoblotting identified the peak subcellular fraction containing the SERCA1 protein (fast-twitch isoform). Gel analysis using the Stains-all dye demonstrated that calsequestrin (CASQ) and phospholipids are highly enriched in the SERCA-containing subcellular fraction isolated from horse gluteus. Immunoblotting also demonstrated that these horse SR vesicles show low content of glycogen phosphorylase (GP), which is likely an abundant contaminating protein of traditional horse SR preps. The maximal Ca2+-activated ATPase activity (Vmax) of SERCA in horse SR vesicles isolated using this protocol is 5‒25-fold greater than previously-reported SERCA activity in SR preps from horse skeletal muscle. We propose that this new protocol for isolating SR vesicles will be useful for determining enzymatic parameters of horse SERCA with high fidelity, plus assessing regulatory effect of SERCA peptide subunit(s) expressed in horse muscle.

Effects of Methyl Branching on the Properties and Performance of Furandioate-Adipate Copolyesters of Bio-Based Secondary Diols.

Furandioate-adipate copolyesters are an emerging class of bio-based biodegradable polymers with great potential to replace fossil-derived terephthalic acid-based copolyesters such as poly(butylene adipate-co-terephthalate) (PBAT). Furandioate-adipate polyesters have almost exclusively been prepared with conventional primary (1°) alcohol diols, while secondary (2°) alcohol diol monomers have largely been overlooked until now, despite preliminary observations that using methyl-branched diols increases the Tg of the resultant polyesters. Little is known of what impact the use of 2° alcohol diols has on other properties such as material strength, hydrophobicity, and rate of enzymatic hydrolysis—all key parameters for performance and end-of-life. To ascertain the effects of using 2° diols on the properties of furandioate-adipate copolyesters, a series of polymers from diethyl adipate (DEA) and 2,5-furandicarboxylic acid diethyl ester (FDEE) using different 1° and 2° alcohol diols was prepared. Longer transesterification times and greater excesses of diol (diol/diester molar ratio of 2:1) were found to be necessary to achieve Mws > 20 kDa using 2° alcohol diols. All copolyesters from 2° diols were entirely amorphous and exhibited higher Tgs than their linear equivalents from 1° diols. Compared to linear poly(1,4-butyleneadipate-co-1,4-butylenefurandioate), methyl-branched, poly(2,5-hexamethyleneadipate-co-2,5-hexamethylenefurandioate) (0:7:0.3 furandioate/adipate ratio) displayed both higher modulus (67.8 vs 19.1 MPa) and higher extension at break (89.7 vs 44.5 mm). All other methyl-branched copolyesters displayed lower modulus but retained higher extension at break compared with their linear analogues. Enzymatic hydrolysis studies using Humicola insolens cutinase revealed that copolyesters from 2° alcohol diols have significantly decreased rates of biodegradation than their linear equivalents synthesized using 1° alcohol diols, allowing for fine-tuning of polymer stability. Hydrophobicity, as revealed by water contact angles, was also found to generally increase through the introduction of methyl branching, demonstrating potential for these materials in coatings applications.

Occurrence and influence of arbuscular mycorrhizae on chemical and enzymatic parameters of soil grown with different crops

In the present system of agriculture, chemical fertilizers are used indiscriminately throughout the world. However, the cost of these fertilizers has been increasing, exponentially and at the same time polluting soil and ground water. In this scenario, the use of eco-friendly bio fertilizers, particularly arbuscular mycorrhizae (AM) is one of the promising alternatives to increase the productivity of agricultural crops. An attempt was made to study the chemical and enzymatic parameters of soil with the occurrence and prevalence of AM fungi in different agricultural crops of Andhra Pradesh. Root and rhizosphere soils of four major crops such as cotton, sugarcane, rice and black gram were collected at different growth stages of the crops and assayed for AM fungi colonization and spore density. Also, the soil chemical and enzymatic parameters such as organic carbon, alkaline phosphatase, glomalin were assayed from these soil samples. Our results showed that the four crop roots colonized by AM fungi. The percent root colonization by AM fungi was maximum in cotton (82.3%) followed by black gram (70%) and rice (35.6%). Whereas the spore number was highest in black gram (42/100g of soil) followed by cotton (39.2/100g of soil) and rice (26.6/100 g of soil). The chemical and enzymatic parameters measured such as organic carbon was maximum in rice (0.79%), followed by sugarcane and cotton (0.73%). Alkaline phosphatase was highest in sugarcane (32.73 p-nitro phenol μg/g wt. of soil/hour) followed by black gram (23.93 p-nitro phenol μg/g wt. of soil/hour) and glomalin was highest in sugarcane (7.96 mg/g) followed by black gram (5.49 mg/g). Our results showed that there was a positive correlation between chemical and enzymatic factors measured with the AM fungi prevalence in the roots and rhizosphere soils of the crops evaluated.

Soil Quality Indicators Associated With The Application Of Mycorrhizal Fungi In Coffee Plantations

Introduction: This article is the result of the research work “Evaluation of the quality of the soil associated with the use of mycorrhizae in coffee plantations in the Municipality of Sasaima, Cundinamarca” carried out duringthe years 2018 and 2019 at the Universidad Libre.
 Problem: Several investigations report the use of mycorrhizal fungi in plant development, however, there are few studies regarding the relationship between mycorrhizae and soil quality indicators.
 Methodology: Four plots were evaluated, applying: 1) Control; 2) Local native mycorrhizae isolated by the authors; 3) Commercial liquid mycorrhizae; and 4) Commercial solid mycorrhizae. The samples of the rhizosphericsoils were taken and bimonthly monitoring was applied until 5 collection cycles were obtained. The physicochemical parameters, abundance of functional groups of microorganisms, and enzymatic activities ofthe soil were determined and analyzed by means of univariate and multivariate statistical analysis.
 Conclusion: The research hypothesis was validated since the mycorrhizae inoculation significantly increased the values of the soil quality indicators, namely organic carbon (14%), moisture (19%), phosphorus (30%), abundanceof nitrogen-fixing bacteria (5.5%), phosphate-solubilizing bacteria (7.8%), cellulolytic bacteria (9.3%), and phosphate-solubilizing fungi (2.6%). In addition, the local native mycorrhizae that were isolated by theauthors showed greater benefits than those of the commercial mycorrhizae.
 Originality: This research reveals the impact of mycorrhizae on the soil quality indicators over time during thefirst year of application, especially on the microbial and enzymatic parameters.
 Limitations: Several authors have investigated the importance of the mycorrhizae on the physicochemical parameters of the plant, and not on the microbial and enzymatic parameters of the soil.

Potential environmental toxicity of sewage effluent with pharmaceuticals.

Sewage effluent effects on the biochemical parameters of Astyanax bimaculatus organs were investigateted. Treated sewage was collected in a treatment plant; 43 compounds, among them, pharmaceuticals and hormones, were investigated. Caffeine, ciprofloxacin, clindamycin, ofloxacin, oxytetracycline, paracetamol, sulfadiazine, sulfamethoxazole, sulfathiazole and tylosin waste was detected in the collected material. Fish were divided into four groups: control, TSE (treated sewage effluent), TSE + P (TSE with increased concentration of five pharmaceuticals) and PTSE (TSE + P post-treated with O3/H2O2/UV). Biochemical parameters were evaluated in different organs after 14-day exposure. TBARS levels increased significantly in the brain of animals in the TSE and TSE + P groups in comparison to the control. There was significant reduction in TBARS levels recorded for the liver, muscle and gills of animals in the PTSE group in comparison to those of animals in the other groups. AChE activity reduced in the muscle of animals in the groups showing the highest pharmaceutical concentrations. CAT activity in the liver of animals in groups exposed to pharmaceutical effluent was inhibited. GST activity increased in brain of animals in the TSE + P and PTSE groups, whereas reduced levels of this activity were observed in liver of animals in the TSE group. Increased GST activity was observed in the brain of animals in TSE + P and PTSE groups. Based on integrated biomarker response values, the TSE + P group presented greater changes in the analyzed parameters. Results point out that pharmaceutical waste can cause oxidative stress, as well as affect biochemical and enzymatic parameters in Astyanax sp. Post-treatment can also reduce damages caused to fish, even in case of the likely formation of metabolites. Based on these results, these metabolites can be less toxic than the original compounds; however, they were not able to fully degrade the pharmaceutical waste found in the sewage, which can interfere in fish metabolism.

Enzymatic Synthesis of Formate Ester through Immobilized Lipase and Its Reuse.

Octyl formate is an important substance used in the perfume industry in products such as cosmetics, perfumes, and flavoring. Octyl formate is mostly produced by chemical catalysts. However, using enzymes as catalysts has gathered increasing interest due to their environment-friendly proprieties. In the present study, we aimed to identify the optimal conditions for the synthesis of octyl formate through immobilized enzyme-mediated esterification. We investigated the effects of enzymatic reaction parameters including the type of immobilized enzyme, enzyme concentration, molar ratio of reactants, reaction temperature, and type of solvent using the optimization method of one factor at a time (OFAT). The maximum conversion achieved was 96.51% with Novozym 435 (15 g/L), a 1:7 formic acid to octanol ratio, a reaction temperature of 40 °C, and with 1,2-dichloroethane as solvent. Moreover, we demonstrated that the Novozym 435 can be reused under the optimal conditions without affecting the octyl formate yield, which could help reduce the economic burden associated with enzymatic synthesis.

Immobilisation of cellulase on vermiculite and the effects on enzymatic kinetics and thermodynamics

In this work, we attached cellulase onto vermiculite by adsorption and crosslinking via glutaraldehyde, and examined the properties of the immobilised enzyme. The results indicated that immobilisation was achived with 35% yield. Reusability of immobilised enzyme was determined as 18 times with 40% residual activity. Even after 100 min incubation with substrate at 50 °C, 6 and 9 cycles, 56% and 30% of activity of the immobilised enzyme was retained, respectively. Optimum pH for free cellulase was 5, while that for immobilised cellulase was 7, however, optimum temperature didn't change both free and immobilised cellulase. The Michaelis-Menten constant (Km) were found as 0.194 mg mL−1 and 0.025 mg mL−1 for free and vermiculite immobilised cellulase,respectively. The maximum reaction rate (Vmax) for the free and vermiculite immobilised cellulase were calculated as 7.42 U/min and 12.5 U/min,respectively. Furthermore, the other enzymatic kinetic parameters (kcat, and kcat/Km) suggested the affinity and specificity of vermiculite immobilised cellulase to the substrate was 1.64 and 1.14 times improved, respectively. The immobilised enzyme had a lower activation energy (8.73, kJ/mol) than the free enzyme (9.73,kJ/mol). Calculated thermodynamic parameters (ΔH≠, ∆G≠∆G≠E-S, ∆G≠E-T) (kJ/mol) indicated that the immobilised enzyme performs biocatalysis more conveniently than the free enzyme.

The enzymology of human eicosanoid pathways: the lipoxygenase branches.

Eicosanoids are short-lived derivatives of polyunsaturated fatty acids that serve as autocrine and paracrine signaling molecules. They are involved numerous biological processes of both the well state and disease states. A thorough understanding of the progression the disease state and homeostasis of the well state requires a complete evaluation of the systems involved. This review examines the enzymology for the enzymes involved in the production of eicosanoids along the lipoxygenase branches of the eicosanoid pathways with particular emphasis on those derived from arachidonic acid. The enzymatic parameters, protocols to measure them, and proposed catalytic mechanisms are presented in detail.

Assessment of Hepatoprotective and Antiulcer Activity of Launaea aspleniifolia Hook in Laboratory Animals

Background: Launaea aspleniifolia Hook (Family Asteraceae) is used traditionally in medicine in Indian system for the treatment of leucoderma. This study was conducted to evaluate the hepatoprotective and antiulcer effects using the methanolic extract of Launaea aspleniifolia Hook (MELA). Methods: The MELA in the dose of 200 and 400 mg/kg body weight (b.w.) was administered orally, daily for 7 days to prevent the acetaminophen-induced hepatotoxicity and peptic ulcer. In order to determine the antioxidant enzymes activity, various enzymatic parameters including alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBL), direct bilirubin (DBL), alkaline phosphatase (ALP), total protein (TP), albumin (ALB), high density lipoproteins (HDL), random glucose test (RBS), serum creatinine levels (SCL) and direct bilirubin (DBL) were determined. Further, the histopathology of tissue and various gastric secretion parameters like free acidity, total acidity ulcer score, % ulcer inhibition, gastric volume, pH, Na+ and K+ and histopathology were determined in PLinduced ulcer model. Results: MELA showed dose-dependent hepatoprotective and ulcer protective effect in acetaminopheninduced hepatotoxicity and antiulcer activity. Furthermore, tissue antioxidant parameter such as reduced Malondialdehyde (MDA), histopathology was also investigated. MELA was more potent in controlling all the serological parameter of liver like ALT, AST, TBL, DBL, ALP, HDL, RBS, SCL, TP and ALB in a dose-dependent manner (P<0.05) induced by Acetaminophen. Apart from this, antiulcer activity MELA was confirmed by the low level of ulcer index along with the reduction of free acidity, total acidity ulcer score, % ulcer inhibition, gastric volume, Na+ and an increase K+, pH of gastric content in a dose-dependent manner (P<0.05) induced PL model. Conclusion: The results indicated a significant dose-dependent effect of MELA from injuries induced in liver and stomach. These protective effects of MELA can be examined in various other toxicities as alone and in combinations.

Removal of dye using peroxidase-immobilized Buckypaper/polyvinyl alcohol membrane in a multi-stage filtration column via RSM and ANFIS.

The feasibility and performance of Jicama peroxidase (JP) immobilized Buckypaper/polyvinyl alcohol (BP/PVA) membrane for methylene blue (MB) dye removal was investigated in a customized multi-stage filtration column under batch recycle mode. The effect of independent variables, such as influent flow rate, ratio of H2O2/MB dye concentration, and contact time on the dye removal efficiency, were investigated using response surface methodology (RSM). To capture the inherent characteristics and better predict the removal efficiency, a data-driven adaptive neuro-fuzzy inference system (ANFIS) is implemented. Results indicated that the optimum dye removal efficiency of 99.7% was achieved at a flow rate of 2mL/min, 75:1 ratio of H2O2/dye concentration with contact time of 183min. The model predictions of ANFIS are significantly good compared with RSM, thus resulting in R2 values of 0.9912 and 0.9775, respectively. The enzymatic kinetic parameters, Km and Vmax, were evaluated, which are 1.98mg/L and 0.0219mg/L/min, respectively. Results showed that JP-immobilized BP/PVA nanocomposite membrane can be promising and cost-effective biotechnology for the practical application in the treatment of industrial dye effluents.

Virtual laboratory lessons in enzymology.

The recent COVID-19 pandemic has led to widespread lock-down strategies that force universities to perform all educational activities remotely. In this context, laboratory lessons pose a significant challenge. Here, I present an on-line tool that simulates the kinetics of chemical reactions. Enzymatic mechanisms can be easily modeled and followed through time. In addition, professors can customize the interface to hide the reaction mechanism. This setting will force students to design virtual experiments to uncover the mechanism and obtain the relevant enzymatic parameters. While some of the skills developed in a practical lesson cannot be simulated, this tool can be used to teach students important concepts about data acquisition and processing.

Biocatalysis of D,L-Peptide Nanofibrillar Hydrogel.

Self-assembling peptides are attracting wide interest as biodegradable building blocks to achieve functional nanomaterials that do not persist in the environment. Amongst the many applications, biocatalysis is gaining momentum, although a clear structure-to-activity relationship is still lacking. This work applied emerging design rules to the heterochiral octapeptide sequence His–Leu–DLeu–Ile–His–Leu–DLeu–Ile for self-assembly into nanofibrils that, at higher concentration, give rise to a supramolecular hydrogel for the mimicry of esterase-like activity. The peptide was synthesized by solid-phase and purified by HPLC, while its identity was confirmed by 1H-NMR and electrospray ionization (ESI)-MS. The hydrogel formed by this peptide was studied with oscillatory rheometry, and the supramolecular behavior of the peptide was investigated with transmission electron microscopy (TEM) analysis, circular dichroism (CD) spectroscopy, thioflavin T amyloid fluorescence assay, and attenuated total reflectance (ATR) Fourier-transform infrared (FT-IR) spectroscopy. The biocatalytic activity was studied by monitoring the hydrolysis of p-nitrophenyl acetate (pNPA) at neutral pH, and the reaction kinetics followed an apparent Michaelis–Menten model, for which a Lineweaver–Burk plot was produced to determine its enzymatic parameters for a comparison with the literature. Finally, LC–MS analysis was conducted on a series of experiments to evaluate the extent of, if any, undesired peptide acetylation at the N-terminus. In conclusion, we provide new insights that allow gaining a clearer picture of self-assembling peptide design rules for biocatalysis.