Enzyme Technology

Enzymes are protein-based biocatalysts that speed up biochemical reactions without affecting equilibrium, with microbial sources being the most efficient and cost-effective for large-scale production. Their dual roles as processing aids and inhibitors of spoilage make understanding their structure, function, and specific activity vital for both in vitro applications and food product development. While the development of recombinant DNA technology and the advancements achieved by microbes in food applications are more economically and practically advantageous, certain of these enzymes are plentiful enough in their natural sources to allow for large-scale synthesis. Enzymes find extensive use across many domains, including but not limited to agriculture, environment, paper and pulp, leather tanning, chemicals and pharmaceuticals, detergents, food and drink, and so on. In this review, we have focused on the molecular mechanisms of enzyme activity and the enzyme manufacturing process within the food industry. We also dedicate a section to various enzymes used in this sector, including amylase, lipase, proteases, cellulase, lactase, glucose oxidase, glucose isomerase, invertase, catalase, and rennet. Additionally, we have presented case studies showcasing the successful implementation of sustainable enzyme practices across different food sectors, highlighting both practical benefits and challenges. The review concludes with an outlook on future trends and research directions aimed at achieving greater safety and sustainability in enzyme technology within the food industry.

The contemporaneously produced alkaline protease and I±-amylase by locally isolated Bacillus methylotrophicus SCJ4 had been purified in two steps and some characteristics of the purified enzymes were studied. Crude enzymes were first salted out using ammonium sulfate, and then concentrated proteins were applied to anion column chromatography Q-sepharose (HiPrep Q FF 16/10 column). The purified alkaline protease has an estimated molecular mass of 24 kDa with maximum enzyme activity (1185U/ml/min) at pH 9.0 and 55ÂoC. This enzyme belongs to serine proteases family with remarkable stability up to 62% and 61.5% toward Sodium Dodecyl Sulfate (SDS) and Hg++ respectively. On the other hand, the purified amylase is a calcium-independent I±-amylase, showing a good stability against SDS up to 73%. The estimated molecular mass was 67.5 kDa with maximum enzyme activity (280U/ml/min) at pH 7.0 and 65ÂoC.

Pseudomonas aeruginosa BTS-2 produced lipase (total yield, 55.4 U/L) optimally on a mineral based (MB) broth, supplemented with 1% (v/v) cottonseed oil as sole carbon source, after incubation at 56°C for 48 h. The presence of 4% (w/v) yeast extract in the MB broth further increased the lipase production by ~ 3-folds. The purified lipase was efficiently (~80%) immobilized in alginate than carrageenan beads. The alginate-immobilized lipase (beads) showed optima at pH 7.5 and 55°C temperature. An increase in C-chain of alcohols increased the activity of entraped lipase. Exposure of alginate beads to decyl (C:10) and dodecyl (C:12) alcohol enhanced the lipase activity by 47 and 64%, respectively. Whereas, most alkanes decreased the activity of immobilized lipase but exposure to nonane enhanced the activity by 52%. The lipase efficiently esterified ethanol and acetic acid to ethyl acetate.

ABSTRACT Production of antimicrobial substances is often regarded a priori as an important trait in the context of bacterial fitness but also in terms of probiotic efficacy. Several probiotic bacteria produce a variety of antimicrobial compounds viz. short chain fatty acids, Hydrogen peroxide, Nitric oxide, Bacteriocins. Bacteriocins are peptide or protein complexes showing antibacterial activity against closely related species. These antimicrobial agents are gaining more and more attention as an alternative therapeutics not only in pharmaceutical but also as a preservative in food industries. In present study, bacteriocin production was carried out by Lactobacillus rennanquilfy WHL 3 isolated from whey and identified by 16s rRNA sequence analysis. This newly isolated strain showed antibacterial activity against several Gram positive and Gram negative bacteria. To obtain maximum bacteriocin yield, medium was optimized with various fermentation conditions like carbon and nitrogen sou...

The article explores how pineapple waste can redefine the soap industry by turning discarded fruit skins into natural, effective cleaning products. It addresses two urgent crises: waste management and environmental pollution. Pineapple skins contain natural acids and enzymes like bromelain that can break down grease and stains. The article highlights a Vietnamese company, Fuwa Biotech, that is turning rotting peels into raw materials, creating a potent, biodegradable cleaner that replaces chemical-laden alternatives. This concept is a textbook example of waste valorization and a move toward a circular economy

Hydroxynitrile lyase (HNL) catalyzed enantioselective CeC bond formation is an efficient approach to synthesize chiral cyanohydrins which are important building blocks in the synthesis of a number of fine chemicals, agrochemicals and pharmaceuticals. Immobilization of HNL is known to provide robustness, reusability and in some cases also enhances activity and selectivity. We optimized the preparation of immobilization of Baliospermium montanum HNL (BmHNL) by cross linking enzyme aggregate (CLEA) method and characterized it by SEM. Optimization of biocatalytic parameters was performed to obtain highest % conversion and ee of (S)-mandelonitrile from benzaldehyde using CLEA-BmHNL. The optimized reaction parameters were: 20 min of reaction time, 7 U of CLEA-BmHNL, 1.2 mM substrate, and 300 mM citrate buffer pH 4.2, that synthesized (S)-mandelonitrile in ∼99% ee and ∼60% conversion. Addition of organic solvent in CLEA-BmHNL biocatalysis did not improve in % ee or conversion of product unlike other CLEA-HNLs. CLEA-BmHNL could be successfully reused for eight consecutive cycles without loss of conversion or product formation and five cycles with a little loss in enantioselectivity. Eleven different chiral cyanohydrins were synthesized under optimal biocatalytic conditions in up to 99% ee and 59% conversion, however the % conversion and ee varied for different products. CLEA-BmHNL has improved the enantioselectivity of (S)mandelonitrile synthesis compared to the use of purified BmHNL. Nine aldehydes not tested earlier with BmHNL were converted into their corresponding (S)-cyanohydrins for the first time using CLEA-BmHNL. Among the eleven (S)-cyanohydrins syntheses reported here, eight of them have not been synthesized by any CLEA-HNL. Overall, this study showed preparation, characterization of a stable, robust and recyclable biocatalyst i.e. CLEA-BmHNL and its biocatalytic application in the synthesis of different (S)-aromatic cyanohydrins.

Indian black berry known as Jamun is a minor fruit which contains high amount of tannin. Extraction and clarification of juice is quite difficult due to its pulp nature. Tannase a membrane bound enzyme is added to clarify the Jamun juice which helps to obtain high yield. The target area of the work is to build up the procedure for streamlining of process factor to obtain Jamun juice, utilizing tannase (strain: Aspergillus ficcum). Physicochemical parameters (clarity, colour change, polyphenol, turbidity, protein, TSS and yield) were analyzed at a temperature range between (30C – 50C), with (0.01% – 0.1% w/v) concentration and time orbit of (40-120 min). Coefficient of determination (R) value more than 0.9 have been used to measure the prominent differences in the response characteristics. Clarified juice was obtained at 0.05% enzyme concentration at 40C for 80 minutes.

The present study is about a novel reaction course to produce biodiesel. The effect of sonication treatment for enhancing the activity of enzymes, particularly dealing with transesterification reaction using lipase to produce Biodiesel [Fatty Acid Methyl Ester (FAME)], has also been studied. The normal batch time for these reactions without sonication treatment was 12 hours and hence, the reaction requires intensification. Therefore, sonication treatment is tried to intensify the process, which adds to the novelty of the current scheme for biodiesel synthesis.

Cereals and legumes contain high amount of phosphorus in the form of phytic acid or phytate. Due to the absence of phytatse enzyme, monogastric animals cannot digest phytic acid nutritional constituent of animal diet. Phytic acid act as an anti metal ions like Ca, Mg, Fe, Zn. phytase producing bacteria from tr phytase production by the isolated strain of bacteria. A total of 10 samples collected from fermented idli and jalebi batter were streaked on phytase screening medium (PSM). Total 50 isola apparently different colony morphology were collected and screened based on their morphological properties. The selected 22 isolates were further evaluated among them, 4 isolates PR2, PR6, PR16, PR24 could be grown on PSM plate with 2.5% agar af shown phytase activity in shake flask experiment. Upon screening of 4 potential phytase producing strains, one isolated strain (PR6) was selected and was identified as morphologi enzymatic assay. This result was optimized the performance of this isolated at different parameters. PR6 showed best results at pH 5.5 and temp 40°C with glucose and sucrose yeast extract as nitrogen source.

Cephalosporin C acylase (CCA) is an essential enzyme for the one-step conversion of cephalosporin C into 7-aminocephalosporanic acid (7-ACA), an intermediate compound used to synthesize various semi-synthetic cephalosporin antibiotics. The industrial process prefers to use enzymes in immobilized form rather than soluble. A crosslinked enzyme aggregate (CLEAs) is a potential matrix-less enzyme immobilization technique to produce stable immobilized enzymes with high activity and low production costs. This study aimed to optimize the CCA immobilization using the CLEAs technique with Chitosan as a co-aggregate. The CCA lysate was obtained from harvesting CCA fermentation broth using a mutant strain of Escherichia coli through cell separation and lysis steps. Partially purified CCA by ammonium sulfate addition was conducted to obtain an active fraction of 20-60% saturation, followed by co-aggregation with Chitosan to form physical CCA aggregates. The aggregates were then immobilized by a...

Laccases are valuable enzymes as an excellent ecological alternative for bioremediation issues because they can oxidize persistent xenobiotic compounds. The production and characterization of extracellular laccases from saprotrophic fungi from disturbed environments have been scarcely explored, even though this could diversify their functional characteristics and expand the conditions in which they carry out their catalysis. Agrocybe pediades, isolated from a disturbed forest, produces an extracellular laccase in liquid culture. The enzyme was purified, identified and characterized. Copper and hexachlorobenzene do not function as inducers for the laccase produced. Partial amino acid sequences were obtained by LC-MS/MS that share similarity with laccases from other fungi. Purified laccase is a monomer with a molecular mass between 55-60 kDa and had an optimum activity at pH 5.0 and the optimum temperature at 45 • C using 2,6-dimethoxyphenol (2,6-DMP) as substrate. The K m and V max also determined with 2,6-DMP were 100 µM and 285 µmol•min -1 •mg -1 , respectively, showing that the laccase of A. pediades has a higher affinity for this substrate than that of other Agaricales. These features could provide a potential catalyst for different toxic substrates and in the future laccase could be used in environmental recovery processes.

An innovative enzymatic polycondensation of dicarboxylic acids and dialcohols in aqueous polymerization media using free and immobilized lipases was developed. Various parameters (type of lipases, temperature, pH, stirring type and rate, and monomer carbon chain length) of the polycondensation in an oil-in-water (o/w) miniemulsion (>80% in water) were evaluated. The best results for polycondensation were achieved with an equimolar monomer concentration (0.5 M) of octanedioic acid and 1,8-octanediol in the miniemulsion and water, both at initial pH 5.0 with immobilized Pseudozyma antarctica lipase B (PBLI). The synthesized poly(octamethylene suberate) (POS) in the miniemulsion is characterized by a molecular weight of 7800 g mol−1 and a conversion of 98% at 45 °C after 48 h of polycondensation in batch operation mode. A comparative study of polycondensation using different operation modes (batch and fed-batch), stirring type, and biocatalyst reutilization in the miniemulsion, wate...

The food industry has been undergoing a transformation in recent years, with a significant emphasis on sustainable, efficient, and health-promoting solutions. One promising avenue is bioinspired multienzyme engineering, which draws upon nature's highly efficient biochemical processes to design complex enzyme systems for various applications. This approach harnesses the advantages of enzyme cascades, multi-enzyme interactions, and synergistic effects to enhance food processing, improve product quality, and address challenges such as waste reduction, allergenicity, and shelf-life extension.This succinct review provides an overview of the recent advances in bioinspired multienzyme engineering with specific relevance to the food industry, highlighting its applications, innovations, and potential. 1

In view of the fact that temperature plays key role in the activity of fungal cellulases, this experiment was designed to assess the stability pattern of the cellulase Enzyme produced by Aspergillus flavus derived from rotten wood. The characterization of the purified cellulase enzyme of Aspergillus flavus was carried out using Carboxymethyl cellulose, Filter paper and Cotton wool assays. The maximum and optimum temperatures were 90°C and 70°C, respectively. The enzyme retained above 50% of its original activity at 100°C for 1h and up to 80% at 80°C for 1h Enzyme decay study showed that Cellulase activity remained 100 % at 0 h for both CMC and filter paper. However, as incubation period increased, the activity decreased, but the enzyme still retained above 50 % activity in both substrates at 1h, over the range of 40-100°C. The study carried out showed that the purified Aspergillus flavus cellulases demonstrated useful properties among which include; high optimum temperature, increased thermal stability. They also offer additional advantage as they are more rigid than meso enzymes and moreover, are more resistant to thermal and chemical denaturation. Hence are remarkable tools for developing commercial biotechnologies and for studying protein stability.

Agricultural wastes are generated in rural areas can be effectively utilized for production of mushroom as food supplements as well as for income generation. The present research explored some of the unexplored locally available lignocellulosic substrates such as paddy straw, reeds, banana stem, sugar cane bagasse milled and crushed, sugar cane leaves, coir pith, sorghum husk and sun flower stem for field scale production of Pleurotus florida and Calocybe indica. The field level cultivation was carried out by 137 farmers and produced totally 15.33 tonne of P. florida and C. indica with an aggregate bioefficiency of 58.52 % and 60.38 % respectively. The produced mushrooms generated an additional income and also consumed as nutritional supplement.

Cancer is caused by a disproportionate division and growth of cells that end up forming a tumour (due to changes in the expression of oncogenes and tumour suppressor genes). It goes through different stages, the last of which is metastasis. Metastasis occurs when some tumour cells are able to travel through lymphatic or blood vessels and end up spreading in another part of the body. One of the cancers with the highest mortality is pancreatic ductal adenocarcinoma (PDAC). It has been shown that the glucidic antigen sialyl Lewis x (sLe) plays an important role in the adhesion of tumour cells to the endothelium. Usually, it is overexpressed in tumour cells because of the overexpression of the sialyltransferase ST3GAL IV (which participates in the antigen biosynthesis). The silencing of the ST3GAL4 gene could decrease the metastatic capacity of PDAC cells. The CRISPR-Cas9 methodology allows gene edition of specific genome sites using the endonuclease Cas9 and an sgRNA complementary to t...

Eco-enzyme is a natural substance that is typically extracted from waste materials and citrus fruit peels, among other sources. It is a sophisticated mixture made from fermenting fresh kitchen trash, including fruit and vegetable peels. This kind of vinegar is created by fermenting food scraps and sugar to produce alcohol. Eco-enzyme is a fermented solution mostly composed of water, fruit peels, and sugar in a 1:3:10 ratio. Because of the phenolic profile and antioxidant qualities of citrus fruit extracts and flavonoids, this review focuses specifically on the citrus eco-enzyme. These compounds may have intriguing biological activities. A thorough discussion of the various inventive methods employed to create eco-enzymes from various raw materials has been held. These methods have several uses, are inexpensive, and are environmentally benign. The synthesis of eco-enzyme reduces the accumulation of organic waste and offers an alternative to synthetic chemicals that are detrimental to the environment and human health. Nonetheless, the main uses have been covered in this review, including cleaning floors, dishes, gardening, and other creative ways to recycle and reuse natural waste that can help cut down on fruit waste.

Highlight:
- Enzyme endo-xylanases are shown to reduce intestinal viscosity.
- The addition of endo-xylanases in the animal feed improves overall digestibility rates of digesta.
- Supplementation of endo-xylanase leads to balance in gut microbiota manifesting as healthy broiler chickens.
- The activities of endo-xylanases improved the growth performance of broiler chicken.

The biofilm-mediated persistence of Pseudomonas aeruginosa in the food and biomedical sectors is currently a global concern. In light of this challenge, this study investigated a preventive approach against P. aeruginosa biofilm formation using Flavourzyme, a food-grade peptidase, considering its antibiofilm potential. The results revealed that a co-culture comprising 300 µL/mL (1 × minimum inhibitory concentration [MIC]) of Flavourzyme could kill P. aeruginosa. On the MBEC™ biofilmforming device, 0.125 × MIC of Flavourzyme inhibited > 4.5 log CFU/peg of biofilm. Cell motilities and the biosynthesis of quorum sensing (QS) molecules such as N-acyl-homoserine lactones (AHLs), including C 4-HSL, decreased significantly at 0.06 × MIC of Flavourzyme and became undetectable at 0.125 × MIC. Interestingly, while 0.03 × MIC of Flavourzyme elicited diverse expressions of QS and virulence-regulating genes, ≥ 0.06 × MIC of Flavourzyme remarkably suppressed the relative genomic expressions. This study proposes Flavourzyme as a potent antibiofilm agent against P. aeruginosa biofilms, recommending specific concentrations for effective use in food preservation.

Involvement of one out of three tryptophan residues in the active site of the low-molecular-mass xylanase from Chainia has been demonstrated previously Biochim. Biophys. Acta 1041, 172-177]. The work described here aims at : (i) deducing the structure-function relationship for the tryptophan residue involved at the active site (a) by correlating the effect of N-bromosuccinimide (NBS) on the fluorescence and activity, and (b) by assessing the ability of xylan to protect against decrease in fluorescence versus activity of NBS-treated enzyme ; and (ii) probing into the environment of the tryptophan residues by studying the quenching of their fluorescence by various solute quenchers in the presence and absence of guanidine hydrochloride (Gdn.HCl). Complete inactivation of the NBStreated enzyme occurs well before the loss of fluorescence. Full protection by xylan (0.5 %) of the inactivation of enzyme by NBS compared with 30 % protection for the decrease in fluorescence confirms the participation of a single tryptophan at the substrate-binding site of the xylanase. The xylanase exhibited Abbreviations used : NBS, N-bromosuccinimide ; Gdn.HCl, guanidine hydrochloride ; K sv , Stern-Volmer quenching constant ; ANS, 1-anilinonaphthalene-8-sulphonate.

N- maleimide (DAM) reacts covalently with the thiol group of the xylanase from Chainia leading to complete inactivation in a manner similar to N-ethylmaleimide, but provides a reporter group at the active site of the enzyme. Increasing amounts of xylan offered enhanced protection against inactivation of the xylanase by DAM. Xylan (5 mg) showed complete protection, providing evidence for the presence of cysteine at the substrate-binding site of the enzyme. Kinetics of chemical modification of the xylanase by DAM indicated the involvement of l mol of cysteine residue per mol of enzyme, as reported earlier Biochim. Biophys. Acta 1041, 172-177]. The second-order rate constant for the reaction of DAM with the enzyme was Abbreviations used : DAM, N-(2,4-dinitroanilino)maleimide ; NEM, N-ethylmaleimide ; TFA, trifluoroacetic acid. * NCL communication no. 6193. † To whom correspondence should be addressed. will be useful for deciphering the structure-function relationship of the xylanase using site-specific mutagenesis.

Acetohydroxamic acid is a pharmaceutically active metal chelating agent which has various applications in the field of medicine. Current study focuses on the enzymatic synthesis of acetohydroxamic acid catalysed by thermophilic amidase from Bacillus smithii IIIMB2907. Bacterial cells were grown in 7 L fermenter for amidase production and effect of pH, temperature and substrate concentration for the biotransformation of acetamide to acetohydroxamic acid was studied. Batch reaction was also successfully optimized at bench scale with the recovery of ≈ 81% acetohydroxamic acid (purified).

From 42 different hot springs in six provinces belonging to distinct geographical regions of Turkey, 451 thermophilic bacilli were isolated and 67 isolates with a high amylase activity were selected to determine the α -glucosidase production capacities by using p NPG as a substrate. α -Glucosidase production capacities of the isolates varied within the range from 77.18 to 0.001 U/g. Eleven of our thermophilic bacilli produced α -glucosidase at significant levels comparable with that of the reference strains tested; thus, five strains, F84b (77.18 U/g), A333 (48.64 U/g), F84a (36.64 U/g), E134 (32.09 U/g), and A343 (10.79 U/g), were selected for further experiments. 16S rDNA sequence analysis revealed that these selected isolates all belonged to thermophilic bacilli 16S rDNA genetic group 5, four of them representing the genus Geobacillus , while strain A343 had an uncultured bacterium as the closest relative. Changes in α -glucosidase levels in the intracellular and extracellular fractions were determined during 48-h cultivation of A333, A343, F84a, F84b, E134, and the reference strain G. stearothermophilus ATCC 12980. According to α -glucosidase production type and enzyme levels in intracellular and extracellular fractions, Geobacillus spp. A333, F84a, and F84b were defined as extracellular enzyme producers, whereas the thermophilic bacterium A343 was found to be an intracellular α -glucosidase producer, similar to ATCC 12980 strain. Geobacillus sp. E134 differed in α -glucosidase production type from all tested isolates and the reference strain; it was described as a membrane-associated cell-bound enzyme producer. In this study, apart from screening a great number of new thermophilic bacilli from the hot springs of Turkey, which have not yet been thoroughly studied, five new thermostable α -1,4-glucosidase-producing bacilli that have biotechnological potential with α -glucosidases located at different cell positions were obtained.

Biofilm formation by Aeromonas hydrophila in the food industry poses significant challenges to food safety and quality. Therefore, this comprehensive review aimed to provide insights into the mechanisms and key factors influencing A. hydrophila biofilm formation. It explores the molecular processes involved in initial attachment, microcolony formation, and biofilm maturation; moreover, it concurrently examines the impact of intrinsic factors, including quorum sensing, cyclic-di-GMP, the efflux pump, and antibiotic resistance, as well as environmental conditions, such as temperature, nutrient availability, and osmotic pressure, on biofilm architecture and resilience. Furthermore, the article highlights the potential of bibliometric analysis as a promising method for conceptualizing the research landscape of and identifying knowledge gaps in A. hydrophila biofilm research. The findings underscore the requirement for focused interventions that prevent biofilm development and raise food sector safety. The consolidation of current information and incorporation of bibliometric analysis enhances existing understanding of A. hydrophila biofilm formation and offers insights for future research and control strategies within a food industry context.

The subject of the scientific research of the doctoral dissertation is the immobilization of two industrially important enzymes, protease from Bacillus licheniformis (commercial name alcalase) and peroxidase from horseradish, as well as their application in protein hydrolysis and removal of pollutants, such as phenols and synthetic dyes, from wastewater. The focus is on the development of techniques for obtaining hydrogel particles based on alginate and alginate/magnetite, as well as enzyme immobilization techniques that would enable obtaining biocatalysts with improved catalytic properties. Enzymatic hydrolysis of proteins and oxido-reduction of anthraquinone dyes catalyzed by enzymes from diverse groups we have chosen as two model reactions for evaluating the catalytic properties of immobilized biocatalysts, due to their industrial importance in the food industry and in the processes of treating wastewater from the textile industry. The development of immobilized enzymes enabled the application of enzymes in those areas where the application of free enzymes is limited due to low stability due to degradation by autocatalysis, as in the case of proteases in the case of protein hydrolysis, or due to inhibition by excess substrate, as in the case of peroxidases for the degradation of anthra-quinone dyes in the presence of hydrogen-peroxide. By immobilizing enzymes, we could obtain stable biocatalysts, which we separated from the reaction mixture after the end of the reaction by filtration or centrifugation and used repeatedly in batches or over a longer period in continuous processes. One of the most crucial factors in enzyme immobilization is the choice and design of the immobilization support because the physical and chemical characteristics of the support determine the activity of the immobilized enzyme. Inorganic supports such as iron oxide can have advantages compared to other mate-rials due to high thermal and mechanical resistance, reduced microbiological contamination, and magnetic properties, which is especially important for immobilized enzymes, where the main advantage compared to native enzymes has eased separation from reaction mix-tures and their reuse. However, due to the small number of functional groups on these inorganic supports, enough enzymes cannot be bound, nor can we reward a favorable micro-environment for the enzyme coupling, which would lead to the improvement of its catalytic properties. Therefore, by coating the magnetic particles with an additional coating of an organic polymer with a larger number of functional groups, such as alginate, a larger number of enzymes can potentially be bound, and we could achieve improved binding. In this dissertation, we examined the possibility of using alginate particles and magnetic particles coated with alginate as carriers for the immobilization of alcalase and horseradish peroxi-dase. The technique of obtaining particles under the influence of an electric field, which in the literature is often called electrostatic extrusion and which is based on the application of an electrostatic force to a suspension of polymers and biocatalysts that appears on the tip of a needle, in the form of drops, during which the drop splits into several small charged droplets, has a great possibility of application for obtaining particles with small diameters of uniform sizes on an industrial scale. Researchers have been extensively using this technique for the immobilization of various biocatalysts. However, this dissertation examined a new procedure for obtaining alginate submicron particles, which is based on a combination of ultrasonic dispersal, drying with hot air, and gelling in a column with wetted walls. Namely, we obtained alginate particles by the method of ultrasonic dispersion of alginate solution, whereby the air carrying the dispersed polymer-aerosol particles, depending on the tem-perature and unsaturation, to a greater or lesser extent encourages the evaporation of water from them and thus reduced their size, and thus obtained the nanoparticles gel in a column with wetted walls. We assessed the nanoparticles obtained in this way as carriers for the immobilization of alcalase and horseradish peroxidase and their catalytic properties with the immobilized systems with beads obtained by electrostatic extrusion. We used beads with the best characteristics (dimensions, sphericity, size distribution, magnetic properties, and magnetite-alginate ratio) for enzyme immobilization. Optimization of covalent immobilization of alcalase enzyme on alginate particles and magnetite/alginate particles obtained by electrostatic extrusion and ultrasonic spraying represents the second phase of the experiments. We have investigated the influence of im-mobilization conditions such as pH value, TRIS buffer molarity, amount of added alcalase, method of chemical activation of functional groups of particles and amount of chemical agent for activation, as well as the contact time of the enzyme with activated particles on activity, mass of bound enzyme and yield of activity. The amount of bound alcalase and peroxidase was determined by standard methods for protein determination, the Bradford method, while the proteolytic activity was determined spectrophotometrically by monitoring the hydrolysis of the chromogenic derivative of casein, azocasein. We analyzed the morphological characteristics of the obtained particles with and without immobilized enzymes using scanning electron microscopy (SEM). The size of the particles, as well as the surface morphology of the magnetic particles with and without alginate coating, were determined. To better understand the nature of the potentially established bonds between the enzyme and the carrier, we have characterized the particles of the carrier and the immobilizer by Infrared Spectroscopy with Fourier transformations. Next, we examined the particle size distribution and zeta potential using the dynamic light scattering method. We examined the magnetic properties of the particles and the strength of the coercive field using SQUID magnetometry (superconducting quantum interferometer). Also, we have compared the magnetic properties of the particles based on hysteresis curves for magnetic and magnetite/alginate particles at temperatures of 5 K and 300 K. The activity of free and immobilized alcalase we monitored in the reaction of protein hydrolysis from soy isolate and egg white under industrially relevant conditions, i.e., in a reactor with intensive mixing and in the presence of impurities based on the measurement of calcium hydroxide consumption to maintain a constant pH value during hydrolysis. We used trinitrobenzenesulfonic acid during hydrolysis, the so-called TNBS method. Kinetics of the reaction with native and immobilized enzyme, we have examined by monitoring the reaction rate of protein hydrolysis from soybean and egg white isolates at different initial substrate concentrations in initial conditions, while kinetic parameters were determined us-ing the Enzyme Kinetics application of the OriginPro software package. We evaluated the stability of the immobilized alcalase, the possibility of using it through several cycles of performing the protein hydrolysis reaction from soy and egg white isolates in a batch reactor with mechanical mixing, and the potential retention of activity after a certain storage time. We used the obtained results in the further development of the immobilized system with horseradish peroxidase. We used beads with the best characteristics (dimensions, sphericity, size distribution, magnetic properties, and ratio of magnetite and alginate) for horseradish peroxidase immobilization. The amount of bound peroxidase and its activity, as well as the activity of free peroxidase, were determined by the spectrophotometric method with pyrogallol as a substrate. We have examined the effects of immobilization time, mass ratio of enzyme and carrier, concentration of the activating agent on the mass of the bound enzyme, and the specific activity of the obtained biocatalyst on the activity. With the stability of the immobilized alcalase, we evaluated the possibility of using it through several cycles of oxidation for two anthraquinone dyes, Acid Violet 109 and Acid Blue 225. The effectiveness of immobilized peroxidases we compared based on the obtained kinetic parameters, using the Enzyme Kinetics application, in the oxidation reaction of AV109 and AB225. The work presented here has demonstrated a novel, simple, and fast technique for alginate submicron bead production by employing the custom-made ultrasonic spray atomi-zation system and compared it with the electrostatic extrusion in the hydrolytic reactions with standard and industrial food protein substrates. For these purposes, we have effectively covalently immobilized alcalase onto calcium-alginate microbeads and submicron beads after surface modification by carbodiimide. The optimal conditions for the alcalase immobilization on the alginate microbeads activated with EDAC were 5,32 IU of alcalase, 50 mM Tris-HCl buffer pH 8,5, 30 min of activation by EDAC/beads mass ratio (10 mg/0,5 g), and 20 h immobilization time. The highest activity was exhibited with alcalase immobi-lized onto UAD 0,4% (w/v) alginate beads, with the maximal value of 2716,1 IU/g support and the highest enzyme loading of 671,6±4 mg/g on the carrier in comparison to 2600,8 IU/g and 592,3±6,7 mg/g for EE biocatalyst made from 2% (w/v) alginate solution. The immobilized enzyme onto UAD 0,4% (w/v) alginate beads provided better performance than micro-sized alginate counterparts in terms of egg white and soy protein hydrolysis. We have reused optimal biocatalyst in seven successive soy protein hydrolysis cycles with a slight decrease in activity. Therefore, there is potential to use this type of alginate submicron beads obtained by novel ultrasonic spray atomization technique for enzyme immobilization. HR...

An innovative enzymatic polycondensation of dicarboxylic acids and dialcohols in aqueous polymerization media using free and immobilized lipases was developed. Various parameters (type of lipases, temperature, pH, stirring type and rate, and monomer carbon chain length) of the polycondensation in an oil-in-water (o/w) miniemulsion (>80% in water) were evaluated. The best results for polycondensation were achieved with an equimolar monomer concentration (0.5 M) of octanedioic acid and 1,8-octanediol in the miniemulsion and water, both at initial pH 5.0 with immobilized Pseudozyma antarctica lipase B (PBLI). The synthesized poly(octamethylene suberate) (POS) in the miniemulsion is characterized by a molecular weight of 7800 g mol−1 and a conversion of 98% at 45 °C after 48 h of polycondensation in batch operation mode. A comparative study of polycondensation using different operation modes (batch and fed-batch), stirring type, and biocatalyst reutilization in the miniemulsion, wate...

Enzymes play a vital role in baking, providing significant benefits from dough development to extending shelf life, which enhances product quality and consistency. Acting as biological catalysts, enzymes such as proteases and amylases break down proteins and starches, modifying dough rheology and improving fermentation. Lipases and oxidases further refine dough texture through emulsification and oxidation, while lipases also produce fatty acid derivatives during fermentation, contributing to the flavor and aroma of baked goods. Xylanases and cellulases optimize dough handling by altering fiber structure, and amylases help maintain moisture and texture, extending the shelf life of baked products. Ensuring regulatory compliance is essential when incorporating enzymes into baking processes, as bakers must address enzyme stability and determine appropriate dosages for reliable outcomes. Ongoing research is exploring innovative enzyme applications, including customized enzyme blends that target specific product qualities, offering new possibilities for product differentiation and innovation. In summary, enzyme-driven advancements present bakers with opportunities to improve product quality, shelf life, and consistency, while meeting industry regulations. This review emphasizes the critical impact enzymes have on dough properties and finished product characteristics, highlighting their role in driving future innovations within the baking industry.

An innovative enzymatic polycondensation of dicarboxylic acids and dialcohols in aqueous polymerization media using free and immobilized lipases was developed. Various parameters (type of lipases, temperature, pH, stirring type and rate, and monomer carbon chain length) of the polycondensation in an oil-in-water (o/w) miniemulsion (>80% in water) were evaluated. The best results for polycondensation were achieved with an equimolar monomer concentration (0.5 M) of octanedioic acid and 1,8-octanediol in the miniemulsion and water, both at initial pH 5.0 with immobilized Pseudozyma antarctica lipase B (PBLI). The synthesized poly(octamethylene suberate) (POS) in the miniemulsion is characterized by a molecular weight of 7800 g mol−1 and a conversion of 98% at 45 °C after 48 h of polycondensation in batch operation mode. A comparative study of polycondensation using different operation modes (batch and fed-batch), stirring type, and biocatalyst reutilization in the miniemulsion, wate...

The contemporaneously produced alkaline protease and α-amylase by locally isolated Bacillus methylotrophicus SCJ4 had been purified in two steps and some characteristics of the purified enzymes were studied. Crude enzymes were first salted out using ammonium sulfate, then concentrated proteins were applied to anion column chromatography Q-sepharose (HiPrep Q FF 16/10 column). The purified alkaline protease has an estimated molecular mass of 24 kDa with maximum enzyme activity (1185U/ml/min) at pH 9.0 and 55ºC. This enzyme belongs to serine proteases family with remarkable stability up to 62% and 61.5% toward Sodium Dodecyl Sulfate (SDS) and Hg ++ , respectively. On the other hand, the purified amylase is a calcium-independent α-amylase, showing a good stability against SDS up to 73%. The estimated molecular mass was 67.5 kDa with maximum enzyme activity (280U/ml/min) at pH 7.0 and 65ºC.

An innovative enzymatic polycondensation of dicarboxylic acids and dialcohols in aqueous polymerization media using free and immobilized lipases was developed. Various parameters (type of lipases, temperature, pH, stirring type and rate, and monomer carbon chain length) of the polycondensation in an oil-in-water (o/w) miniemulsion (>80% in water) were evaluated. The best results for polycondensation were achieved with an equimolar monomer concentration (0.5 M) of octanedioic acid and 1,8-octanediol in the miniemulsion and water, both at initial pH 5.0 with immobilized Pseudozyma antarctica lipase B (PBLI). The synthesized poly(octamethylene suberate) (POS) in the miniemulsion is characterized by a molecular weight of 7800 g mol-1 and a conversion of 98% at 45 • C after 48 h of polycondensation in batch operation mode. A comparative study of polycondensation using different operation modes (batch and fed-batch), stirring type, and biocatalyst reutilization in the miniemulsion, water, and an organic solvent (cyclohexane:tetrahydrofuran 5:1 v/v) was performed. Regarding the polymer molecular weight and conversion (%),batch operation mode was more appropriate for the synthesis of POS in the miniemulsion and water, and fed-batch operation mode showed better results for polycondensation in the organic solvent. The miniemulsion and water used as polymerization media showed promising potential for enzymatic polycondensation since they presented no enzyme inhibition for high monomer concentrations and excellent POS synthesis reproducibility. The PBLI biocatalyst presented high reutilization capability over seven cycles (conversion > 90%) and high stability equivalent to 72 h at 60 • C on polycondensation in the miniemulsion and water. The benefits of polycondensation in aqueous media using an o/w miniemulsion or water are the origin of the new concept strategy of the green process with a green product that constitutes the core of the new greener polymer-5B technology.

Noni is a useful tropical fruit shrub, difficult to propagate due to seeds dormancy. Hence, a two-stage experiment was conducted to develop easily applicable rapid germination method for the crop. Firstly, seeds were subjected to different pretreatments [nicking, soaking in 50% H2SO4 for 5 min., 800 ppm GA3 for 24 h, hot-water at 40 °C for 24 h and control (untreated)]. Secondly, the most suitable pre-treatment method was applied to another set of seeds and sown in six sowing media [topsoil, river-sand (reference), sawdust, topsoil+river-sand (1:1), topsoil+sawdust (1:1) and river-sand+sawdust (1:1)]. A total of 2000 and 2400 seeds were sown in first and second stages, respectively. Each set of treatments was laid in completely randomised design with four replicates. Data collected were used to determine germination percentage (GP), mean germination time (MGT), germination index (GI) and mean germination rate (MGR). The results showed that there were significant differences (p ≤ 0.05) in the effects of the treatments for all the parameters considered at each of the stages. Nicking elicited highest GP (64.8%), MGR (0.026 sprouts/day), GI (1.79) and shortest MGT (38 days) at 12 weeks after sowing. Subsequently, the results of Morinda citrifolia seeds sown in different media indicated that topsoil/river-sand mixture significantly increased GP (81.5%) and GI (3.08) while topsoil gave highest MGR (0.038 sprouts/day) and shortest MGT (26.4 days). Hence, for commercial large scale seedling production, Noni seeds should be nicked and sowed directly in topsoil or a mixture of topsoil and river-sand.

Aquesta tesi se centra en l’estudi de les possibles glicosiltransferases de Micoplasma genitalium involucrades en la sintesi de glicolipids. Aquests compostos formen part de la membrana plasmatica del microorganisme, unic envolcall que el protegeix del seu entorn ja que no disposa de paret cel•lular. La hipotesi sobre la qual s’ha desenvolupat el treball es la possibilitat que aquests glicolipids i els enzims encarregats de la seva produccio, les glicosiltransferases, siguin essencials per a la viabilitat del micoplasma i per tant, la seva inhibicio sigui una forma d’eradicar les infeccions causades pel patogen. De les tres sequencies classificades com a glicosiltrasferases en el genoma de Mycoplasma genitalium, mg025, mg060 i mg517, s’ha determinat que mg025 es l’unic dels tres gens que no es essencial per al bacteri, tot i que s’ha observat que els tres s’expressen tant a la fase exponencial com a la fase estacionaria del seu creixement. La funcio de mg025 i mg060 es encara descon...

Highlight
• Laccases can effectively decolorize textile synthetic dye effluent.
• The use of laccases as a green biocatalyst shows promise for decolorization and detoxification of textile effluent.
• Laccase-catalyzed degradation mechanisms of the synthetic dyes have the potential to degrade the toxicity of wastewater.
• Highlights the potential use of laccases as a green approach to treat textile wastewater in an environmentally friendly manner in Cambodia.

Highlight
• Enzyme endo-xylanases are shown to reduce intestinal viscosity.
• The addition of endo-xylanases in the animal feed improves overall digestibility rates of digesta.
• Supplementation of endo-xylanase leads to balance in gut microbiota manifesting as healthy broiler chickens.
• The activities of endo-xylanases improved the growth performance of broiler chicken.

The fungus Glomerella cingulata damages a wide range of crops in the tropics and subtropics. This fungus produces an extracellular enzyme, cutinase, which has been implicated in the plant disease process. A cutin monomer, 16hydroxyhexadecanoic acid was used to induce cutinolytic esterase activity during saprophytic growth of G. cingulata. Cutinolytic esterase was induced in cultures grown in cutin monomer but was repressed by glucose. The pattern of cutinolytic activity obtained from the induction and enzyme assays correlates with the results of the Northern blot analyses. The cutinolytic activity and expression from this study demonstrates that the enzyme is tightly regulated by the cutin monomer, 16hydroxyhexadecanoic acid.

Un nou mecanisme d'inhibició enzimàtica, que actua "cap per avall" © 2011 Universitat Autònoma de Barcelona -Tots els drets reservats Un nou mecanisme d'inhibició enzimàtica, que actua "cap per avall" 06/2013 -Biologia. Dos grups d'investigació de l'Institut de Biotecnologia i de Biomedicina i del Departament de Bioquímica i de Biologia Molecular de la UAB, dirigits respectivament pel Dr. Francesc X. Avilés, que fa anys que estudia les carboxipeptidases, i pel Dr. David Reverter, que lidera un grup de cristal•lografia de proteïnes, acaben de publicar en la prestigiosa revista Structure un treball en el que caracteritzen l#estructura tridimensional del complex entre una forma humana d'un enzim (que es produeix en càncer de pròstata) i un nou inhibidor proteic, procedent de l'anèl•lid marí Sabellastarte magnifica. Aquest treball ha estat fruit d#una col•laboració entre la UAB i un grup de la Universitat de l'Havana, liderat per Maday Alonso del Rivero i Maria A. Chávez, aquesta última també professora invitada de la UAB.

Novel triblock copolymers with PEG middle blocks of 1-10 kDa and poly(N-isopropylacrylamide-co-tbutylacrylamide) statistical copolymer side arms with DP n ≈88 and different compositions, were synthesized by SET-LRP. The thermogelation properties of their aqueous solutions depended on both hydrophobic monomer content of the side blocks and molecular weight (MW) of the poly(ethylene glycol) (PEG) middle block, as proven by dynamic rheometry, DSC, and tube inversion method measurements. At constant PEG chain length, increasing TBAM proportions led to a gelation process occurring at progressively lower temperatures, as well as to a lower stability of the forming hydrogels in the case of shorter-PEG-chain block copolymers. By employing longer PEG blocks (M PEG ≥6,000 Da), stable hydrogels with the gelation temperature below 37 °C could be obtained. For a constant composition of the copolyacrylamide blocks, the dependence of the phase transition temperature (T ph ) on M PEG displayed a different shape at different polymer solution concentrations, because of the stronger variation of T ph with polymer concentration as M PEG increased. Also, the viscoelastic properties of the hydrogels resulting from 20 wt.% polymer aqueous solutions at 37 °C were stronger affected by the MW of the PEG middle block than by the hydrophobic character of the thermosensitive side blocks.

This study describes the course of enzymatic hydrolysis of the native corn starches Maritena 100 and Maritena 300. Hydrolyses were carried out with glucoamylase Glm produced by Saccharomycopsis fibuligera IFO 0111, which degrades also native starch, with the purpose to substitute a two-step hydrolysis (amylase followed by glucoamylase) by a one-step process (glucoamylase only). Hydrolysis generally became more effective by adding the pullulanase Promozyme D, which cleaves a-1,6-glycosidic bonds more effectively than glucoamylase Glm does. The time course (kinetics) of hydrolysis was followed by determination of the glucose concentration and calculation of dextrose equivalents.

Background & Objectives: Thermophilic alpha-amylase can be used in different industries such as starch processing and detergents. This study was performed to isolate alpha-amylase-producing bacteria and characterization of the enzyme. Materials & Methods: After sample collection from Gorooh hot spring in Kerman province, Iran, thermophilic alpha-amylase-producing bacteria were isolated using the starch-agar medium. 16S rDNA sequencing was used to identify the bacterial strain. Characterization of the thermophilic alpha-amylase was performed in the presence of various factors such as pH, temperature, metal ions, chemical compounds, and organic solvents. Also, kinetic parameters of the enzyme were determined in different concentrations of starch. Results: Anoxybacillus gonensis AT23 was identified as the best thermophilic alpha-amylaseproducing strain. The alpha-amylase enzyme showed the optimal activity at pH 5 to 6. Sevenfold increase in the enzyme activity was observed in the presence of NaCl (3M). Mn 2+ and Zn 2+ increased the enzyme activity about 95% and 31%, respectively. Kinetic parameters including K m and V max were estimated about 1.657 mg/ml and 0.0059 mg/ml/min, respectively. Also, enzyme activity was also improved about 2 folds in the presence of organic solvents including n-butanol and 10% cyclohexane. Conclusion: Our results indicated that AT23 alpha-amylase is a halophile and organic solvent-tolerant enzyme. Therefore, it can be used in different industries.

A novel bacterial named AC21 having nitrile-converting activity was gained with acetonitrile as sole source of nitrogen, which was recognized by morphology and 16S rDNA gene sequence as Stenotrophomonas maltophilia. This intracellular nitrilase induced by acetonitrile (0.3%) catalyses the conversion of 3-cyanopyridine to nicotinic acid without noticeable production of nicotinamide. Optimal pH and temperature of Nitrilase AC21 was obtained at pH 8.0 and 45 • C. Irreversible thermo-inactivation results showed that, enzyme half-life was 6 h at 80 • C. Several process factors, including quantity of enzyme and rate of substrate feeding, were considered with the goal of reaching whole substrate transformation. In a fed batch manner at 1 l scale using hyper-induced AC21 resting cells equivalent to 10 U ml −1 nitrilase activity, a total of 945 mM of 3-cyanopyridine was formed at a rate of 3.32 g h −1 g dcw −1 , which directed AC21 has ability of application in large scale nicotinic acid production.

α-Amylase an industrially used enzyme can be obtained from Aspergillus niger and can be produced from food sources such as pigeon pea. α-Amylase was produced from Aspergillus niger isolated from pigeon pea, purified and characterized. This process was achieved using ammonium sulphate, ion exchange DEAE column and gel filtration (Sephadex A-50 and sephadex G-100) chromatography. The effect of temperature and pH was determined. Ammonium sulphate precipitation results showed that the highest specific α-amylase activity was (1.01 U/mL. mg) obtained at 11.27% saturation level, with a purity of 1.81-fold of the crude extract and yielding 1.00%. Further purification using gel filtration increased the enzyme purity and yielding 8.94-fold relative to the crude extract 3.01% and yielding Specific activity after purification was 4.99 U/mg. Having varied temperature from 30-80 o C, the optimum temperature was obtained at 50°C. The pH ranged from 3-10, at a concentration of 0.01 M which was prepared using different buffer systems, the enzyme was found stable at pH of 6. Therefore α-amylase produced from Aspergillus niger can be exploited for potential usage for industrial applications of enzymes in a wide range of production and its application in food processing.

Abstract
Conventional detergent formulations contain either petroleum-derived or
synthetic chemicals. Hence, the replacement of these chemicals with natural
products such as enzymes is considered a green technology. Among these
enzymes, protease is renowned in detergent formulations. Therefore, the
objective of this study was to explore and characterize a new protease from
Phytolacca dodecandra, because the plant is traditionally known for detergency
purposes. Enzyme extraction was performed with Glycine/NaOH buffer,
pH = 10.0. Gelatin hydrolysis and agar well diffusion tests were implemented
as qualitative assays, whereas the enzyme assay technique was applied for
the quantitative investigation of the new protease from plant extracts. The
crude enzyme was partially characterized concerning temperature and pH tolerance, washing performance, detergent compatibility, and stability. The finding
of this study revealed that Phytolacca dodecandra contains protease and is
characterized by 1418.84 U/mL of enzyme activity, where 1 unit (U) of enzyme
activity refers to the amount of enzyme required to liberate 1 μg of tyrosine/mL/
min under the defined assay conditions. The enzyme showed maximum activity
at pH 10.5 and 45
C. The enzyme was stable and maintained 99.52% activity
at 40
C and 91.73% at pH 10.0 after 2 h pre-incubation. The enzyme showed
excellent wash performance and could potentially remove blood stains
completely within 3 min. After 2 h of pre-incubation the enzyme maintained
69.06% and 54.88% of its activity in the presence of Azzy and Sunlight, respectively. The enzyme maintained 71.99% of its residual activity after 60 days of
storage. If further studies are conducted, all these properties of the newly investigated protease can make the enzyme applicable in detergent formulations.

Preparing stable protein‐based microcapsules containing functional fatty acids and oils for food applications has been a big challenge. However, recent advances with transglutaminase (TGase) enzyme as an effective protein cross‐linker could provide workable solutions for the encapsulation of omega‐3 and omega‐6 fatty acids without compromising their targeted release and their biological and physicochemical characteristics. The recent and available literature related to the microencapsulation techniques, physical and oxidative properties, and core retention and release mechanisms of TGase‐crosslinked microcapsules entrapping edible oils were reviewed. The effects of factors involved in microencapsulation processes, on the efficiency and quality of the produced innovative microcapsules were also discussed and highlighted. A brief focus has been finally addressed to new insights and additional knowledge on micro‐ and nanoencapsulation of lipophilic food‐grade ingredients by TGase‐induc...

This study targeted to optimize process factors using response surface methodology and determining their effect on delignification and cellulose recovery. Optimization of pretreatment conditions with desirability value of 0.828 was done at temperature (A) 150°C, contact time (B) 2 h, and liquid-to-solid ratio (LSR) (C) 15 ml/g. The sample was soaked in an aqueous ethanol solution (48% ethanol). Chemical composition was determined to be 1.87 ± 0.20, 7.03 ± 0.99,
28.05 ± 0.27, 39.29 ± 3.39, and 23.77 ± 3.91 wt% for ash, extractives, lignin, hemicellulose, and cellulose, respectively. Experimental results were validated by comparing actual value with predicted value. The model was verified as the best for improving delignification and cellulose yield. Coefficient of determination (R2), analysis of variance (ANOVA), response plots, and optimization nodes were used to examine the experimental results. The quadratic equation was used to test the model, and ANOVA was used to evaluate the model’s statistical performance and correctness. Temperature, interaction term (BC), and quadratic terms (A2, B2, and C2) showed significance, while contact time, LSR, and interaction terms (AC and AB) indicated insignificance for cellulose recovery. Temperature, contact time, interaction term (AB), and quadratic terms (B2 and C2) showed significance, but LSR, interaction terms (AC and BC), and quadratic term (A2) showed insignificance for delignification. Pretreatment efficacy for delignification (6.0%) and cellulose recovery (151.80%) were achieved at optimal conditions. The pretreatment method is able to produce cellulose-rich coffee husk residues.

Poly(glycidylmethacrylate-methylmethacrylate), poly(GMA-MMA) beads were prepared via suspension polymerization in the presence of ferric ions. The epoxy groups of the poly(GMA-MMA) beads were converted into amino groups during magnetization reaction, and then were grafted with methacrylic acid (MAA) via graft copolymerization. The magnetic beads were characterized by surface area measurement, swelling test, scanning electron microscope (SEM), electron spin resonance (ESR) and Mössbauer spectroscopy. The enzyme "trypsin" was immobilized on the magnetic beads via adsorption. The maximum adsorption was obtained at pH 7.0. At 2.0 mg/mL initial trypsin concentration, the maximum immobilization capacity was 123.2 mg trypsin/g beads and retained about 84.2% of its initial activity. The immobilized trypsin could not be desorbed by enzyme reaction solution in the pH range of 5.0-9.0, and could be desorbed by 1.0 M formic acid solution containing 1 M NaCl.

Manihot esculenta, Crantz (Cassava) peel was taken for the isolation of acidic amylases. The enzyme activity of 20% and 40% precipitate was found to be 0.02 and 0.18μmole/min, whereas, 50% precipitate did not show any enzyme activity. The fractionation of 40% precipitate on Sephadex G-75 column yielded two peaks with enzyme activity 0.02 and 0.08μmole/min respectively. The second peak indicated a rich source of amylase. A gradual increase in the enzyme activity was observed at 350C to 500C, along with drastic decrease at 500C onwards. A maximum activity was observed from 500C at pH 6.8 to 7.0. The activity of purified enzyme was 0.14μmole/min.Besides, substrate concentration between 1 to 3% which led to substantial increase in enzyme activity. Further, the enzyme activity of Cassava amylase against commercial starch was tested and found that, the enzyme activity increased with increase in time and remains constant after 80th min, whereas, on its own starch there was no activity till...