Enzyme Yield Research Articles

Statistical optimization and sequential scale-up of α-galactosidase production by Actinoplanes utahensis B1 from shake flask to pilot scale

α-Galactosidase (α-GAL) is a class of hydrolase that releases galactose from galacto-oligosaccharides and synthetic substrates such as pNPG. In this study, the production of α-GAL by Actinoplanes utahensis B1 in submerged fermentation was enhanced by using statistical methods. The effects of temperature, pH, and inoculum percentage on enzyme secretion were optimized using BBD of RSM. The optimized process was scaled up from the shake flask to the laboratory scale (5 L) and to pilot scale (30 L) using KLa based scale-up strategy. By using BBD, a maximum yield of 62.5 U/mL was obtained at a temperature of 28 °C, a pH of 6.9, and an inoculum of 6.4%. Scale-up was performed successfully and achieved a yield of 74.4 U/mL and 76.8 U/mL in laboratory scale and pilot scale fermenters. The TOST was performed to validate the scale-up strategy and the results showed a confidence level of 95% for both scales indicating the perfect execution of scale-up procedure. Through the implementation of BBD and scale-up strategy, the overall enzyme yield has been significantly increased to 76%. This is the first article to explore the scale-up of α-GAL from the A. utahensis B1 strain and provide valuable insights for industrial applications.

The Effects of Tillage and the Combined Application of Organic and Inorganic Fertilizers on the Antioxidant Enzyme Activity and Yield of Maize Leaves

The aim of this study was to explore the characteristics of the combined application of organic fertilizer and inorganic fertilizer using different tillage methods to delay the senescence of maize leaves. The yield and activities of GDH, CAT, APX, GR, and GSH enzymes in maize leaves were measured at different growth stages by using two tillage methods, three organic and inorganic combined applications (P1, P2, and P3), and four control treatments. (1) During the growth period, the R + S and R treatments were P1 treatments, with the highest enzyme activities noted for GDH, CAT, APX, GR, and GSH, which were 36.79–103.22% higher than those of CK. (2) The average yield of all R + S treatments was higher than that of R treatments, and the average yield of P1 treatment was the highest under R + S, which was 13,663.79 kg hm−2, which was 6.39%, 7.90%, and 14.67% higher than that of P2, P3, and CK, respectively, which was lower than that of R. The yield of P1 treatment was 2.53% higher. (3) There was a significant positive correlation between APX activity, CAT activity, GR activity, GDH activity, GSH activity, grain number per ear, ear length, and 100-grain weight of maize leaves at the grain filling stage, and a significant negative correlation between bald tip length and yield. The treatment details had the strongest enzyme activity and the highest yield when using the rotary tillage + subsoiling (R + S) P1 method, which was the most suitable tillage method and the best fertilizer ratio combination, which could be demonstrated and popularized in a large area in the dry farming area of spring maize in Shanxi Province.

Multiple strategies to improve extracellular secretion and activity of feruloyl esterase

Feruloyl esterase has a wide range of applications, but there are still problems with low enzyme yield and activity, and complex purification steps. Our previous research found Lactobacillus amylovorus feruloyl esterase could be secreted extracellular in Escherichia coli. In this study, multiple strategies were implemented to maximize the extracellular production of feruloyl esterase with improved activity in E. coli. Firstly, codon-optimized feruloyl esterase was obtained based on the preference of E. coli, resulting in 41.97 % increase in extracellular secretion. Furthermore, by cascading T7 promoters, replacing the 5′ UTR, randomly mutating the N-terminal sequence, and co-expressing secretory cofactors, the extracellular secretion was increased by 36.46 %, 31.25 %, 20.66 % and 25.75 %, respectively. Moreover, the feruloyl esterase were mutated to improve the substrate affinity and activity. The catalytic efficiency of Fae-Q134T and Fae-Q198A increased by 4.62-fold and 5.42-fold. Combining above strategies, extracellular feruloyl esterase activity was increased from 2013.70 U/L to 10,349.04 U/L. These results indicated that the activity and yield of feruloyl esterase secreted by E. coli were significantly increased, which laid a foundation for its industrial application.

INHIBIION ACTIVITY OF SOME PLANT SEEDS EXTRACTS AGAINST THE PROTEASE PRODUCED FROM LOCAL ISOLATE OF CANDIDA ALBICANS ISOLATE USING SOLID STATE FERMENTATION

This study was aimed at inhibition of purified protease produced by C. albicans using plant seeds extract. One hundred two local C. albicans isolates that were isolated and identified by microscopic and biochemical tests were submitted to primary and secondary screening techniques in order to select the qualified C. albicans isolate for protease synthesis. Among these isolates, nineteen isolates with the highest hydrolysis zone on skim milk media (primary screening) were chosen for secondary screening. It has been found that C. albicansV56 had the highest productivity of the enzyme (41.8U/mg protein). The optimum conditions of protease production by selected isolate using solid state fermentation by using wheat bran as best substrate , temperature 28 °C and pH 7, after 5 days of incubation . The enzyme was purified by concentration with sucrose, then used gel filtration chromatography using Sephadex G-75. The results show two peaks. The first peak has a purification folds 6.5 time with an enzyme yield of 13.08 %, while peak 2 has a purification folds 3.3 time with an enzyme yield of 20%. The purified enzyme exhibited maximal activity at pH 6.0 for peak 1 and 3.0 for peak 2, whereas the maximum stability was 7.0 for peak 1 and 8 for peak 2. The optimal temperature for purified enzyme activity was 40 for peak 1 and 30 for peak 2, and it was stable until 45°C. Protease activity was inhibited with local plant seed extracts. Lathyrus sativus extract inhibits approximately 50 % of protease activity.

Scalable Bioreactor Production of an O2‐Protected [FeFe]‐Hydrogenase Enables Simple Aerobic Handling for Clean Chemical Synthesis

AbstractThe enzyme CbA5H, a [FeFe]‐hydrogenase from Clostridium beijerinckii, has previously been shown to survive exposure to oxygen, making it a promising candidate for biotechnological applications. Thus far [NiFe]‐hydrogenases are typically considered for such applications, due to the superior O2‐tolerance and therefore simplified enzyme handling. However, methods for production of [FeFe]‐hydrogenases are generally more successful than for other classes of hydrogenases, therefore in this work we focus on demonstrating scalable CbA5H production, and report results with active enzyme prepared in bioreactors (up to 10 L) with >20‐fold improvement in purified enzyme yield. We then go on to confirm excellent H2/H+‐cycling activity of the air‐purified protein, highlighting that CbA5H can be prepared and isolated without the need for complex and expensive infrastructure. Next, we demonstrate good stability of the air‐purified CbA5H both in solution assays, and as a heterogenous catalyst system when immobilized on a carbon support. Finally, we successfully implement this enzyme within previously demonstrated biotechnologies for flavin and NADH recycling, highlighting its relevance in chemical synthesis, and we demonstrate production of an important API precursor, 3‐quinuclidinol at >0.4 g scale in standard benchtop hydrogenation infrastructure, with >100,000 CbA5H turnovers over 18 operational hours.

Comprehensive Analysis of Penicillium Sclerotiorum: Biology, Secondary Metabolites, and Bioactive Compound Potential─A Review.

The filamentous fungus Penicillium sclerotiorum is significant in ecological and industrial domains due to its vast supply of secondary metabolites that have a diverse array of biological functions. We have gathered the metabolic potential and biological activities associated with P. sclerotiorum metabolites of various structures, based on extensive research of the latest literature. The review incorporated literature spanning from 2000 to 2023, drawing from reputable databases including Google Scholar, ScienceDirect, Scopus, and PubMed, among others. Ranging from azaphilones, meroterpenoids, polyketides, and peptides group exhibits fascinating potential pharmacological activities such as antimicrobial, anti-inflammatory, and antitumor effects, holding promise in pharmaceutical and industrial sectors. Additionally, P. sclerotiorum showcases biotechnological potential through the production of enzymes like β-xylosidases, β-d-glucosidase, and xylanases, pivotal in various industrial processes. This review underscores the need for further exploration into its genetic foundations and cultivation conditions to optimize the yield of valuable compounds and enzymes, highlighting the unexplored potential of P. sclerotiorum in diverse applications across industries.

PURIFICATION AND CHARACTERIZATION OF AMYLASE PRODUCED FROM PROBIOTIC LACTOBACILLUS PLANTARUM CS FOR INDUSTRIAL APPLICATIONS

Previous studies have demonstrated that probiotic Lactobacillus plantarum CS was able to generate an appreciable amount of extracellular amylase, hence the need to purify and characterize it. The aim of the study was to purify and characterize crude amylase from probiotic Lactobacillus plantarum CS for its industrial applications Three purification steps including ammonium sulphate precipitation, ion exchange chromatography on carboxymethyl sephadex and gel filtration on Sephadex G-75 were utilized. The homogeneity of the purified enzyme was confirmed using sodium deodocyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The purified amylase was characterized on different parameters including substrates hydrolyses, pH and temperature activity and stability profiles. The general purification elution profile revealed two different peaks of amylase activities with outstanding one having a molecular weight of 59.7kDa. Its purification fold was 4.0 with specific activity of 16.44U/mg protein and enzyme yield of 3%. Temperature optimal activity and stability was at 400C and 7.5 for pH activity and stability. Mangenese (Mn2+) (135.17%), tween 80 (128.30%) and some food condiments garlic, thyme, ginger, and tumeric) significantly (p> 0.05) enhanced amylase activity (≥262.40%). However, selenium (Se4+) and hydrogen peroxide (H2O2) were observed to have greatest inhibiting effect (≥30.9%) on the enzyme. Substrate hydrolysis profiles showed that the amylase hydrolyzed all the test starchy substrates with the highest hydrolytic potential on indigenous sweet potato starch (Km value/ Vmax of 1.33mg/ml/ 7.89ml). The rate of hydrolysis of other test substrates had yam> rice>cassava>corn with km values ≤ 4.0mg/ml and Vmax ≤ 25ml. The obtained results gave an insight that amylase produced from Lactobacillus plantarum CS met with the possessed properties suitable for any industrial application especially in food

Effect of medium supplementations and extraction conditions on cellulase production through solid state fermentation of oil palm empty fruit bunches

In this study, cellulase enzyme was produced through solid state fermentation (SSF), employing oil palm empty fruit bunches (EFB) as the primary substrate. Two key aspects were explored to enhance crude enzyme yield, which are medium supplementation effects during SSF and cellulase recovery. During medium supplementation, glucose and/or Tween 80 were added alongside EFB substrate and other nutrients. Enzyme yield was determined using a filter paper assay and expressed as enzyme activity. The initial addition of glucose during fermentation led to increased crude enzyme activity, as measured by the filter paper assay. The peak crude enzyme activity was observed with the addition of 3 mg of glucose, with higher amounts showing no further increase in activity. Conversely, the addition of Tween 80 did not yield any significant increase in crude enzyme activity across all concentrations tested. The extraction conditions were varied to study cellulase recovery, specifically by adjusting the solid‐to‐solvent ratio and the number of extraction stages. Higher enzyme activity was achieved with lower solid‐to‐liquid ratios, as the increased solvent volume facilitated greater enzyme extraction. However, increasing the number of extraction steps did not significantly affect the resulting cellulase activity. Overall, this research underscores the need for further process optimization for cellulase production via SSF, utilizing the widely available EFB in Indonesia.

Optimization of hydrolases production by Penicillium crustosum in submerged fermentation using agro-waste residues as cosubstrate

This study aimed to enhance the production of exo-polygalacturonases (exo-PG), xylanases, and endoglucanases using Penicillium crustosum through submerged fermentation. Lemon peel (LP) and wheat bran (WB) were examined as potential cosubstrate for hydrolase production. The highest exo-PG and endoglucanase activities were observed with LP, whereas xylanase activity showed improvement with the addition of WB. To optimize exo-PG and endoglucanase production, a 23 Face-Centered Central Composite Design (FCCCD) was employed, varying crucial physicochemical factors. Similarly, a 24 FCCCD was utilized to optimize xylanase production by varying temperature, pH, MnSO4 concentration as a cofactor, and the PDB culture medium. Under optimized conditions, exo-PG production reached 1427.99 ± 27.41 U/mL, marking a 45% increase compared to initial fermentation conditions. Endoglucanase production peaked at 62.68 ± 9.65 U/mL under conditions of 45 °C, pH 5.5, and 2.5 g/L MnSO4. Xylanase production reached 271.77 ± 8.50 U/mL under optimal conditions (37.5 °C, pH 5.5, 2.5 g/L MnSO4, and 62.5% PDB). The optimized simultaneous production of the hydrolytic cocktail occurred at 45 °C, pH 5.5, and 2.5 g/L MnSO4 with LP, yielding maximum activities of 959.74 U/mL for exo-PG, 61.98 U/mL for endoglucanases, and 65.60 U/mL for xylanases as determined by FCCCD. In conclusion, Penicillium crustosum shows promise as a hydrolase producer, and the utilization of agro-wastes as cosubstrate enhances the yield of enzymes with industrial relevance.

RESPONSE MAIZE CULTIVARS (Zea mays L) TO FOLIAR APPLICATION OF KINETIN

This study aimed to investigate effect of foliar application of Kinetin on growth, yield, yield components and antioxidant affectivity of maize. A field experiment was conducted during the spring season of 2023 at the Alhamidia Research Station, College of Agriculture, University of Anbar. A randomized complete block design with split-plot arrangement with three replications was used in this study. The main plots contained the levels of kinetin (0, 25, 50, and 75 mg L-1), and the subplots contained the cultivars (Baghdad 3, Fajr 1, and Almaha). The results indicated that the maize cultivars significantly differed in most of the studied traits. The Baghdad 3 cultivar showed a significant superiority in most of the studied traits, such as leaf area (5473 cm2), highest average crop growth per unit area, and the highest grain yield (3.75 t ha-1) and biological yield. The Al-Maha cultivar showed superiority most active antioxidant enzymes, catalase and superoxide dismutase(CAT, SOD). Foliar application of kinetin improved growth traits, antioxidant enzyme activity, yield, and quality. The concentration of 75 mg L-1 recorded the best results,

Exogenous application of nano-silicon, potassium sulfate, or proline enhances physiological parameters, antioxidant enzyme activities, and agronomic traits of diverse rice genotypes under water deficit conditions

Water deficit is a critical obstacle that devastatingly impacts rice production, particularly in arid regions under current climatic fluctuations. Accordingly, it is decisive to reinforce the drought tolerance of rice by employing sustainable approaches to enhance global food security. The present study aimed at exploring the effect of exogenous application using different biostimulants on physiological, morphological, and yield attributes of diverse rice genotypes under water deficit and well-watered conditions in 2-year field trial. Three diverse rice genotypes (IRAT-112, Giza-178, and IR-64) were evaluated under well-watered (14400 m3/ha in total for the entire season) and water deficit (9170 m3/ha) conditions and were exogenously sprayed by nano-silicon, potassium sulfate, or proline. The results showed that drought stress substantially decreased all studied photosynthetic pigments, growth traits, and yield attributes compared to well-watered conditions. In contrast, antioxidant enzyme activities and osmoprotectants were considerably increased compared with those under well-watered conditions. However, the foliar application of nano-silicon, potassium sulfate, and proline substantially mitigated the deleterious effects of drought stress and markedly enhanced photosynthetic pigments, antioxidant enzyme activities, growth parameters, and yield contributing traits compared to untreated stressed control. Among the assessed treatments, foliar spray with nano-silicon or proline was more effective in promoting drought tolerance. The exogenous application of proline improved chlorophyll a, chlorophyll b, and carotenoids by 21.4, 19.6 and 21.0% followed by nano-silicon treatment, which enhanced chlorophyll a, chlorophyll b, and carotenoids by 21.1, 17.6 and 9.5% compared to untreated control. Besides, the application of proline demonstrated a superior improvement in the content of proline by 52.5% compared with the untreated control. Moreover, nano-silicon exhibited the maximum enhancement of catalase and peroxidase activity compared to the other treatments. The positive impacts of applied exogenously nano-silicon or proline significantly increased panicle length, number of panicles/plant, number of grains/panicle, fertility percentage, 1000-grain weight, panicle weight, and grain yield, compared to untreated plants under water deficit conditions. In addition, the physiological and agronomic performance of evaluated rice genotypes significantly contrasted under drought conditions. The genotype Giza-178 displayed the best performance under water deficit conditions compared with the other genotypes. Consequently, the integration of applied exogenously nano-silicon or proline with tolerant rice genotype as Giza-178 is an efficient approach to ameliorating drought tolerance and achieving agricultural sustainability under water-scarce conditions in arid environments.

The Effect of Nitrogen Reduction and Applying Bio-Organic Fertilisers on Soil Nutrients and Apple Fruit Quality and Yield

In this study, we investigated the effect of partially substituting inorganic nitrogen with bio-organic fertiliser on the ‘Tianhong2’ Fuji apple planting in Xinjiang. Bio-organic fertiliser was applied, and nitrogen was reduced by 20% (T2), 40% (T3), and 60% (T4) during the blooming and fruit expansion periods with conventionally fertilised fields used as control (T1); soil nutrient, soil enzyme activity, leaf nutrients, fruit quality, and yield were measured. The total nitrogen (TN), total phosphorus (TP), total potassium (TK), total calcium (TCa), available phosphorus (AP), available potassium (AK), and soil organic matter (SOM) contents, as well as the soil catalase (S-CAT), soil uretrase (S-UE), soil saccharase (S-SC), and soil nitrate reductase (S-NR) activities, significantly increased in the experimental soils compared with those in T1. In addition, TP, TCa, and total magnesium (TMg) content in apples significantly increased. Compared to T1, the T2 and T3 treatments significantly improved the fruit yield and quality, increasing the sugar–acid ratio, soluble protein, soluble sugar, peel carotenoid, and anthocyanin content and reducing peel chlorophyll content. The brightness (L*), red–green axis (a*), yellow–blue axis (b*), colour intensity (C), and tone (h°) values changed. The yield per hectare and nitrogen fertiliser partial productivity values were significantly increased. Overall, the T2 treatment resulted in the best outcome for the Yili area. In conclusion, partially substituting inorganic nitrogen with bio-organic fertiliser can effectively increase soil and leaf nutrient content and improve fruit yield and quality.

Effects of Defoliation at Different Fertility Stages on Material Accumulation, Physiological Indices and Yield of Cotton

In recent years, severe hailstorms have caused damage to cotton leaves and stalks. In order to identify the effects of cotton leaf damage on its dry matter accumulation, protective enzyme activity and yield in different periods, in this experiment, different intensities of hail were simulated by manual leaf cutting. In this study, the effects of leaf damage on dry matter accumulation, chlorophyll fluorescence, POD (peroxidase), SOD (superoxide dismutase) and MDA levels (malondialdehyde), and yield of cotton were studied in field experiments at three stages (bud, full bud and flower boll stages) and in sub-plots with different artificial defoliation intensities (0%, 25%, 50%, 75% and 100%). Removing the leaf sources had differently sized effects on the “sink” at each stage, and these are ordered as follows: flowering and boll stage > full bud stage > pregnancy stage. The greater the intensity of leaf removal, the greater the impact on the “sink”. Among them, after removing 50% of the leaves at the full bud stage, the total dry matter of the cotton plant increased by 12.46% compared to that of the control, and the boll formation rate per plant increased by 14.99%, resulting in overcompensation. Mo, Vj and φDo all showed a tendency to decrease and then increase with the increase in defoliation intensity at different periods of the treatment, and the lowest values of Mo, Vj and φDo, and the largest values of φpo, ψo and φEo were found in the 50% defoliation treatment at the gestational bud stage. The values of ψo and φEo were at the maximum in the 25% defoliation treatment at the full bud stage. The values of Mo and Vj in the different defoliation treatments at the bolling stage showed a tendency to increase and then decrease with the increase in defoliation intensity, with the highest values in the 25% treatment and the smallest values of φpo, ψo and φEo in the 25% defoliation treatment. The POD enzyme activity level was elevated in the defoliation treatments at the three different periods, and the highest value was observed in the 50% defoliation group at the full bud and boll stages, which is a reflection of supercompensation. The SOD enzyme activity level tended to increase with the intensity of defoliation, and defoliation at the gestational and full bud stages first enhanced and then weakened the stress on the cotton plants. The differences between treatments decreased after 12 weeks. The stress of defoliation on cotton plants was weakened at the boll stage. With the increase in defoliation intensity, the content of MDA showed a gradual increasing trend. The cotton MDA content was higher than that of the other treatments at 75% defoliation at both the post-fertilized bud and full bud stage.

Implications of carbon catabolite repression for Aspergillus-based cell factories: A review.

Carbon catabolite repression (CCR) is a global regulatory mechanism that allows organisms to preferentially utilize a preferred carbon source (usually glucose) by suppressing the expression of genes associated with the utilization of nonpreferred carbon sources. Aspergillus is a large genus of filamentous fungi, some species of which have been used as microbial cell factories for the production of organic acids, industrial enzymes, pharmaceuticals, and other fermented products due to their safety, substrate convenience, and well-established post-translational modifications. Many recent studies have verified that CCR-related genetic alterations can boost the yield of various carbohydrate-active enzymes (CAZymes), even under CCR conditions. Based on these findings, we emphasize that appropriate regulation of the CCR pathway, especially the expression of the key transcription factor CreA gene, has great potential for further expanding the application of Aspergillus cell factories to develop strains for industrial CAZymes production. Further, the genetically modified CCR strains (chassis hosts) can also be used for the production of other useful natural products and recombinant proteins, among others. We here review the regulatory mechanisms of CCR in Aspergillus and its direct application in enzyme production, as well as its potential application in organic acid and pharmaceutical production to illustrate the effects of CCR on Aspergillus cell factories.

Effect of co-cultivation of white and brown rot species on basidiome production, lignocelluloytic enzyme activity and dye decolourisation

The aim of this work was studying the impact of co-cultivating two mushroom species: a white (Pleurotus albidus CLA 45) and a brown rot one (Laetiporus sulphureus BAFC 205) in substrates based on poplar or pine sawdust, on their lignocellulolytic enzyme production, yield values and basidiomes properties. Laetiporus sulphureus only developed basidiome primordia, but P. albidus monoculture and co-culture in pine sawdust achieved biological efficiencies of up to 50–55 %. Co-cultivation on diverse substrates rendered varied enzyme titers. Laccase and Manganese peroxidase titers were highest in pine co-culture and P. albidus poplar monoculture, respectively. Enzymatic extracts obtained from spent poplar substrate of dual cultures displayed potential for treating non-sterile textile-coloured effluents, achieving 35 % decolourisation after 120 h. The knowledge available on the effects of co-culture of white and brown rot fungi is still limited. This study represents an initial exploration of the interaction between them within intensive cultivation conditions.

Production and characterization of protease enzyme from Acinetobacter pittii using peanut meal as substrate

ABSTRACT A significantly high protease enzyme yield of 617 U/ml was achieved with Acinetobacter pittii as the microorganism and peanut oil meal as the substrate. Peanut oil meal, which consists of proteins (40-60%) and carbohydrates (22-30%), serves as a sufficient source of nitrogen and carbon necessary for microbial growth and production of enzymes. Moreover, peanut meal offers the advantages of being affordable and available in large quantities, making the meal suitable for cost-effective enzyme production. In the present study, two bacterial strains and one fungal strain were selected to produce proteases utilizing peanut oil meal as the substrate. The experimental conditions during the enzyme production, including pH and temperature, were optimized. In addition, the substrate was enriched with various carbon and nitrogen sources. The microbial strains were streaked on nutritional agar (for bacteria) and potato dextrose agar (for fungus). Following an incubation period, the plates were stored at 4°C for further studies. The molecular weight of partially purified proteases of Acinetobacter pittii was found to be ≅ 95.5 kDa. Potassium nitrate was the most ideal nitrogen source (up to 411% increase in activity) and fructose was the best carbon source (425% increase). These enzymes exhibited excellent temperature tolerance and were capable of functioning over a wide pH range. Furthermore, the obtained proteases demonstrated ability to coagulate milk effectively, indicating their potential for various food-related applications.

Rice straw hydrolysis using in-situ produced enzymes: Feedstock influences fungal enzyme composition and hydrolytic efficiency

Trichoderma reesei RUT-C30 was cultivated on differentially pretreated rice straw and pure cellulose as a carbon source/inducer for cellulase production, and the enzymes were evaluated for hydrolysis of sequential acid and alkali pretreated rice straw. Growth on pretreated rice straw enhanced protein secretion and cellulase activities compared to pure cellulose as a carbon source. The yield of cellulolytic enzymes was higher for alkali pretreated rice straw (ALP-RS), while H2O2-treated (HP-RS) could not induce cellulases to a larger level compared to pure cellulose. Protein concentration was 3.5-fold higher on ALP-RS as compared to pure cellulose, with a maximum filter-paper cellulase (FPase) activity of 1.76 IU/ml and carboxy-methyl cellulase (CMCase) activity of 40.16 IU/ml (2.18 fold higher). Beta-glucosidase (BGL) activity was more or less the same with the different substrates and supplementation of heterologous BGL could result in a quantum jump in hydrolytic efficiencies, which in the case of ALP-RS induced enzymes was 34% (increased from 69.26% to 92.51%). The use of lignocellulosic biomass (LCB) itself as a substrate for the production of cellulase is advantageous not only in terms of raw material costs but also for obtaining a more suitable enzyme profile for biomass hydrolysis.

Molecular purification, and kinetic study of the enzyme glutathione peroxidase in patients with cardiovascular disease

This study was conducted to evaluate serum glutathione peroxidase activity and partial purification of the enzyme from the serum of heart attack patients using a precipitant or what is called salting out, ion exchange dialysis using DEAE-Cellulose, and gel filtration using Sephadex G100 gel. This study was conducted on 90 samples, (60) patients whose ages ranged between (35-75) years and control (30) individuals whose ages ranged between (23-55). It showed that the number of purification times was (4.64), and the enzyme yield was (40%). The specific activity was (1.3 U/mg). Likewise, kinetic studies were carried out in which values of Km and Vmax of glutathione peroxidase were (16.66 mM) of Vmax, and (0.909 IU/L) of Km, and the optimal concentration of the base material was (100 mmol/L). The optimal temperature for enzyme action was (37°C), while the optimal pH was (7.2), and the molecular weight of glutathione peroxidase was (23 kDa). The study also studied the effect of carvedilol as an inhibitor of the enzyme glutathione peroxidase, and the results showed a decrease in enzyme activity, and the type of inhibition was uncompetitive.

Partial Purification of 5'-Nucleotidase Enzyme in Patients with Renal Failure and Kinetic Study of the Purified Enzyme in Salah al-Din

This study involved separating and partially purifying the 5'-Nucleotidase enzyme isoforms from the sera of patients with renal failure. The purification process consisted of several steps, beginning with precipitation using 30% ammonium sulfate, resulting in a purification fold of 1.2625 and an enzyme yield of 65.63%. This was followed by membrane filtration using a buffer solution, achieving a purification fold of 1.651 and an enzyme yield of 49%. The protein fraction separated from the dialysis bag was then passed through a gel filtration column, reaching a purification fold of 51.88 and an enzyme yield of 43.1%. The enzyme isoforms from the separated protein fraction were further separated using gel filtration, employing ion-exchange chromatography techniques. This resulted in three isoforms of the 5'-Nucleotidase enzyme with varying degrees of purity, the highest for isoform. at 2.297-fold. The study also included kinetic studies of the enzyme purified by gel filtration, which involved determining the Km value under the influence of varying concentrations of the enzyme's substrate, 5'-AMP, for the purified enzyme from the sera of patients with renal failure, which was found to be 12.4. The effects of temperature, pH, and reaction time on enzyme activity were also studied.

Scale-up studies for production of α-amylase by A. Tamarii MTCC5152 and development of enzyme product

α-amylases are one of the most vital commercially produced starch hydrolyzing enzymes. Increasing enzyme production is becoming one of the most important goals in biotechnological processes. In this study, the production of α-amylase by A.tamarii MTCC5152 in flask level studies is experimentally validated by scale-up studies in Koji room under controlled conditions using wheat bran, a low cost agro-industrial waste as substrate. The production of amylase was increased by optimising various culture conditions such as inoculum size, incubation period and moisture content. It was found that A.tamarii MTCC5152 produced highest titre of α-amylase activity (668.01 u/g), after 4 days of incubation period at 5% inoculum concentration and 80% moisture content. The results of the comparative analysis of protease and amylase production demonstrated 15% increase in α- amylase and 20% increase in protease production compared to flask level studies. Studies conducted for the development of enzyme products as liquid enzyme concentrate and dry enzyme powder showed that addition of hydrophilic agents such as poly ethylene glycol (PEG- 6000) was found to be an effective way to increase the activity and enzyme yield in the preparation of a dry enzyme powder using A.tamarii MTCC5152. The ammonium sulfate precipitated enzyme at saturation, lyophilized by freeze drying gave the best recovery (96%) of the enzyme activity.