Enzyme Solution Research Articles

Isolation, Characterization and Anti-Pepsin (HIV-Protease) Activity of Vernonia amygdalina Del and Cymbopogon citratus (de Condole) Stapf

The anti-pepsin enzyme (HIV-1) activities of crude extract of Vernonia amygdalina and Cymbopogon citratus were investigated using standard methods. A portion (0.5cm3) of pepsin enzyme solution was measured and transferred into viels, 2 cm3 of hemoglobin and 1 cm3 of the extracts were mixed together. The procedure was repeated for all the different extract. A standard control was also prepared using hemoglobin, enzyme and the various concentrations of the anti-retroviral drugs. A negative control was ran using only hemoglobin and pepsin enzyme whereas positive control was ran along with the standard using hemoglobin, enzyme and various concentrations of antiretroviral drugs. The mixture was allowed to incubate at 37 ºC, after 20 mins, 700 µl of 5% TCA was added to stop the reaction. It was then centrifuged at 14000 rev/min for 5 mins and the supernatant was collected. Absorbance was recorded spectrophotometrically at 280 nm. Each sample was taken in triplicate. UV-Visible spectrophotometer was used to detect the absorbance at 280 nm. The results of the anti-pepsin enzyme (HIV-1) activities showed that petroleum ether fraction of Cymbopogon citratus exhibited wavelength of maximum absorption (גmax) ranging from 0.788 nm - 0.789 nm and its ethyl acetate fraction exhibited גmax absorption ranging from 0.785 nm- 0.788 nm. Petroleum ether fraction of Vernonia amygdalina exhibited גmax absorption ranging from 1.383 nm-1.399 nm while its ethyl acetate fraction exhibited גmax absorption ranging from 1.384 nm-1.400 nm. The result of this work shows that both Cymbopogon citratus and Vernonia amygdalina ethyl acetate fraction have higher activity with Vernonia amygdalina being the highest. Column chromatography of the ethyl acetate fraction of Vernonia amygdalina led to the isolation of the compound which shows higher activity (1.384 nm- 1.400 nm). Available spectroscopic data using 1H NMR, 13C NMR and IR showed that the compound could be β-Sitosterol. The result of this work therefore showed that the extract from the leaves of Vernonia amygdalina could be used for the treatment of HIV-1(since it has the same sequence and family with the pepsin enzyme) and other diseases caused by targeted virus.

Screening the Degradation of Polymer Microparticles on a Chip.

Enzymatic degradation of polymers has advantages over standard degradation methods, such as soil burial and weathering, which are time-consuming and cannot provide time-resolved observations. We have developed a microfluidic device to study the degradation of single microparticles. The enzymatic degradation of poly (1,4-butylene adipate-co-terephthalate) (PBAT) microparticles was studied using Novozym 51032 cutinase. PBAT microparticles were prepared via an oil-in-water emulsion solvent removal method, and their morphology and chemical composition were characterized. Then, microparticles with varying diameters of 30-60 μm were loaded into the microfluidic chip. Enzyme solutions at different concentrations were introduced to the device, and changes in the size and transparency of PBAT microparticles were observed over time. The physicochemical properties of degraded products were analyzed by FT-IR, NMR, mass spectrometry, and differential scanning calorimetry. The degradation process was also performed in bulk, and the results were compared to those of the microfluidic method. Our analysis confirms that the degradation process in both bulk and microfluidic methods was similar. In both cases, degradation takes place on aliphatic and soft segments of PBAT. Our findings serve as a proof of concept for a microfluidic method for easy and time-resolved degradation analysis, with degradation results comparable to those of conventional bulk methods.

A novel feature for monitoring the enzymatic harvesting process of adherent cell cultures based on lens-free imaging

Adherent cell cultures are often dissociated from their culture vessel (and each other) through enzymatic harvesting, where the detachment response is monitored by an operator. However, this approach is lacking standardisation and reproducibility, and prolonged exposure or too high concentrations can affect the cell’s viability and differentiation potential. Quantitative monitoring systems are required to characterise the cell detachment response and objectively determine the optimal time-point to inhibit the enzymatic reaction. State-of-the-art methodologies rely on bulky imaging systems and/or features (e.g. circularity) that lack robustness. In this study, lens-free imaging (LFI) technology was used to develop a novel cell detachment feature. Seven different donors were cultured and subsequently harvested with a (diluted) enzymatic harvesting solution after 3, 5 and 7 days of culture. Cell detachment was captured with the LFI set-up over a period of 20 min (every 20 s) and by optimising the reconstruction of the LFI intensity images, a new feature could be identified. Bright regions in the intensity image were identified as detaching cells and using image analysis, a method was developed to automatically extract this feature, defined as the percentage of detached cell regions. Next, the method was quantitatively and qualitatively validated on a diverse set of images. Average absolute error values of 1.49%, 1.34% and 1.97% were obtained for medium to high density and overconfluent cultures, respectively. The detachment response was quantified for all conditions and the optimal time for enzyme inhibition was reached when approximately 92.5% of the cells were detached. On average, inhibition times of 9.6–11.1 and 16.2–17.2 min were obtained for medium to high density and overconfluent cultures, respectively. In general, overconfluent cultures detached much slower, while their detachment rate was also decreased by the diluted harvesting solution. Moreover, several donors exhibited similar trends in cell detachment behaviour, with two clear outliers. Using the novel feature, measurements can be performed with an increased robustness, while the compact LFI design could pave the way for in situ monitoring in a variety of culture vessels, including bioreactors.

Preparation and Characterization of Flavored Sauces from Chinese Mitten Crab Processing By-Products

To achieve high-value utilization of Chinese mitten crab processing by-products, different types of sauces were prepared using crab legs (CLs) and crab bodies (CBs). Two processing methods, enzymatic hydrolysis and enzymatic hydrolysis coupled with the Maillard reaction, were employed to prepare these sauces. An electronic nose (E-nose) and electronic tongue (E-tongue) were used to measure the changes in the taste and odor, an automatic amino acid analyzer was used to measure the amino acid content, and a headspace solid-phase microextraction GC/MS (HS-SPME-GC/MS) was used to analyze the volatile compounds, qualitatively and quantitatively. The results showed that the sour, bitter, and other disagreeable odors of the enzymatic hydrolysis solution (EHS) were reduced following the Maillard reaction; meanwhile, the umami and saltiness were considerably enhanced. The quantity of free acidic amino acids with an umami and sweet taste in the CL and CB sauces after enzymatic hydrolysis and the Maillard reaction was substantially higher than that in the homogenate (HO). The Maillard reaction solution (MRS) produced more volatile compounds than the HO and EHS, such as aldehydes, pyrazines, ketones, etc. These compounds not only impart a unique flavor but also have antioxidant capabilities, making them a prototype for the high-value utilization of crab processing by-products.

Isolation, purification and PEG-mediated transient expression of mesophyll protoplasts in Camellia oleifera

BackgroundCamellia oleifera (C. oleifera) is a woody edible oil crop of great economic importance. Because of the lack of modern biotechnology research, C. oleifera faces huge challenges in both breeding and basic research. The protoplast and transient transformation system plays an important role in biological breeding, plant regeneration and somatic cell fusion. The objective of this present study was to develop a highly efficient protocol for isolating and purifying mesophyll protoplasts and transient transformation of C. oleifera. Several critical factors for mesophyll protoplast isolation from C. oleifera, including starting material (leaf age), pretreatment, enzymatic treatment (type of enzyme, concentration and digestion time), osmotic pressure and purification were optimized. Then the factors affecting the transient transformation rate of mesophyll protoplasts such as PEG molecular weights, PEG4000 concentration, plasmid concentration and incubation time were explored.ResultsThe in vitro grown seedlings of C. oleifera ‘Huashuo’ were treated in the dark for 24 h, then the 1st to 2nd true leaves were picked and vacuumed at − 0.07 MPa for 20 min. The maximum yield (3.5 × 107/g·FW) and viability (90.9%) of protoplast were reached when the 1st to 2nd true leaves were digested in the enzymatic solution containing1.5% (w/v) Cellulase R-10, 0.5% (w/v) Macerozyme R-10 and 0.25% (w/v) Snailase and 0.4 M mannitol for 10 h. Moreover, the protoplast isolation method was also applicable to the other two cultivars, the protoplast yield for ‘TXP14’ and ‘DP47’ was 1.1 × 107/g·FW and 2.6 × 107/g·FW, the protoplast viability for ‘TXP14’ and ‘DP47’ was 90.0% and 88.2%. The purification effect was the best when using W buffer as a cleaning agent by centrifugal precipitation. The maximum transfection efficiency (70.6%) was obtained with the incubation of the protoplasts with 15 µg plasmid and 40% PEG4000 for 20 min.ConclusionIn summary, a simple and efficient system for isolation and transient transformation of C. oleifera mesophyll protoplast is proposed, which is of great significance in various aspects of C. oleifera research, including the study of somatic cell fusion, genome editing, protein function, signal transduction, transcriptional regulation and multi-omics analyses.

Enzymatic Characterization of Unused Biomass Degradation Using the Clostridium cellulovorans Cellulosome.

The cellulolytic system of Clostridium cellulovorans mainly consisting of a cellulosome that synergistically collaborates with non-complexed enzymes was investigated using cellulosic biomass. The cellulosomes were isolated from the culture supernatants with shredded paper, rice straw and sugarcane bagasse using crystalline cellulose. Enzyme solutions, including the cellulosome fractions, were analyzed by SDS-PAGE and Western blot using an anti-CbpA antibody. As a result, C. cellulovorans was able to completely degrade shredded paper for 9 days and to be continuously cultivated by the addition of new culture medium containing shredded paper, indicating, through TLC analysis, that its degradative products were glucose and cellobiose. Regarding the rice straw and sugarcane bagasse, while the degradative activity of rice straw was most active using the cellulosome in the culture supernatant of rice straw medium, that of sugarcane bagasse was most active using the cellulosome from the supernatant of cellobiose medium. Based on these results, no alcohols were found when C. acetobutylicum was cultivated in the absence of C. cellulovorans as it cannot degrade the cellulose. While 1.5 mM of ethanol was produced with C. cellulovorans cultivation, both n-butanol (1.67 mM) and ethanol (1.89 mM) were detected with the cocultivation of C. cellulovorans and C. acetobutylicum. Regarding the enzymatic activity evaluation against rice straw and sugarcane bagasse, the rice straw cellulosome fraction was the most active when compared against rice straw. Furthermore, since we attempted to choose reaction conditions more efficiently for the degradation of sugarcane bagasse, a wet jet milling device together with L-cysteine as a reducing agent was used. As a result, we found that the degradation activity was almost twice as high with 10 mM L-cysteine compared with without it. These results will provide new insights for biomass utilization.

Co‐Cultivation of Aspergillus niger and Trichoderma reesei Enables Efficient Production of Enzymes for the Hydrolysis of Wheat Bran

AbstractAgro‐residues like wheat bran may be a useful carbon source for fermentation processes. However, due to their composition, hydrolysis appears complex. A co‐cultivation process was established with the two fungi A. niger and T. reesei for the production of an adapted mixture of enzymes for hydrolysis. This enzyme solution reached a higher space‐time yield of sugars during the hydrolysis of wheat bran compared to enzymes from respective monocultures, demonstrating the potential of this co‐culture for the production of complementary enzyme mixtures for the hydrolysis of wheat bran.

Synergistic Enhancement Effect of Compound Additive of Organic Alcohols and Biosurfactant on Enzymatic Hydrolysis of Lignocellulose

The insufficient of lignocellulose degradation enzymes, such as cellulase and hemicellulase, is the major obstacle that hinders the bioconversion of lignocellulosic biomass to monosaccharides, especially during the woody biomass hydrolysis process. The addition of additives has received significant attention due to their enhancement of the enzymatic degradation efficiency of lignocellulose. In the present study, a combination of organic alcohols and a biosurfactant could synergistically enhance the saccharification of the cellulose substrate of Avicel, as well as that of pretreated poplar. Results showed that compound additives can greatly improve the conversion rate of enzymatic hydrolysis. The combination of 0.1% (v/v) n-decanol and 1% (v/v) sophorolipid dramatically increased the poplar enzymatic conversion rate from 17.9% to 85%, improving it by 67.1%. Enzyme-rich Hypocrea sp. W63 was fermented to obtain beta-glucosidase (BGL) and xylanase (XYL), which were used as auxiliary enzymes during enzymatic hydrolysis. It was found that the effects of such a combination of additives improved the filter paper activity, stability, and longevity, helping in the recovery of the cellulase cocktail. The compound additives associated with the commercial cellulase and Hypocrea sp. W63 enzyme solution formed an excellent formula for improving the stability of BGL and XYL. The results provide insight into compound additives and the use of a cellulase and auxiliary enzyme cocktail to improve enzymatic hydrolysis for lignocellulose conversion into biofuels.

Kinetic and thermodynamic aspects of the process of thermal inactivition of kluyveromyces marxianus inulinase

Background: Inulinase is widely found in microorganisms and higher plants. It catalyzes the reaction of the hydrolysis of inulin, which is present in the tubers and roots of many plants, to fructose and a small amount of glucose.
 Inulin is one of several plant-based polysaccharides that contain glucose or fructose. It is used as a substrate in industrial fermentation processes and in the food industry due to its relatively cheap and abundant source for the microbiological production of high fructose juices, ethanol, acetone and butanol. Inulin-derived oligosaccharides are also used in the medical and food sectors. Inulinase which produced from the yeast K. marxianus at levels close to commercial use. This indicates that inulinase can be used in the production of fructose and fructose syrups. So our main object is to study the effect of different temperatures on the conformation of the Kluyveromyces marxianus inulinase macromolecule, to provide optimal conditions for aniline hydrolysis by inulinase.
 Materials and method: Method of isolation and purification of the inulinase enzyme from Kluyveromyces marxianus, as well as the determination of the protein content and enzyme activity as mentioned in the context of the research. Experiments were carried out to study the thermal stability of enzyme. For this, an enzyme solution at a concentration of 5•10-5 mol / l was incubated in a time interval of 10-60 min at different temperatures, followed by determination of the catalytic activity.
 It is known that different types of bonds and interactions participate in the formation of the molecular structure of enzymes, which are covalent bonds, hydrogen bonds, salt bridges, hydrophobic interactions, so our main goal is to study the effect of different temperatures on the formation of the enzyme molecule in order to provide optimal conditions for aniline hydrolysis. by enzyme.
 Results: of this study revealed that the residual activity of the enzyme after 60 minutes of incubation at 50 °C was 10% of the initial activity at 60 °C 7%. As for the Rate Constants of thermal inactivation of Inulinase at 70 °C were 3.45.
 Conclusions: Results of this study were important and useful in determining parameters of inulinase enzyme. Based on the shape of the curves of the dependence of the catalytic activity of Kluyveromyces marxianus inulinase on the time of thermal inactivation in the temperature range of 20-80 ° C.

Process Study on the Enzyme-Catalyzed Preparation of Key Chiral Intermediates for Saxagliptin

AbstractSaxagliptin is a therapeutic drug for diabetes. The key synthesis process of the drug involves catalyzing 2-(3-hydroxy-1-adamantyl)-2-oxoacetic acid (A) into (S)-3-hydroxyadamantane glycine (B), during which enzymes phenylalanine dehydrogenase mutant from Thermoactinomyces intermedius (TiPDHm) and formate dehydrogenase (FDH) were most often used for biocatalysis. However, the process was limited due to difficulty in enzyme preparation and a low conversion rate. This study focuses on co-expression of TiPDHm and FDH in recombinant Escherichia coli, cell homogenate clarification, enzyme concentration as well as the optimized conditions of enzyme-catalyzed reaction. Our data showed that the wet weight density of bacteria reached 300 g/L, and the yields of TiPDHm and FDH were 7674.24 and 2042.52 U/L, respectively. The combination of ammonium formate and polyethyleneimine favors the clarification of the bacteria homogenate. The clarified enzyme solution obtained can be concentrated by ultrafiltration and directly used in a reductive amination reaction in a high concentration of keto acid A. The reaction time was only 12 hours and the conversion rate reached 95%. Therefore, this process could provide a reference for enzyme-catalyzed preparation of saxagliptin on an industrial scale.

Corrosion resistance of stainless-steel surgical tools in enzymatic and alkaline detergent

Metallic surgical instruments are subject to corrosion, which can be accelerated when there is an attack on the passive layer formed on the stainless-steel surfaces. Corrosion can create defects on the metal surface, which can retain residues and promote microbiological growth, leading to infections in surgeries and contamination of medical devices. Since these applications require a high degree of cleaning and sterilization, such items undergo recurrent cleaning cycles using water and detergents. This study aims to test two types of detergents, alkaline and enzymatic, comparing their effects on corrosion resistance of AISI 304 L, 316 L and 420 stainless steels (SS), commonly used in surgical instruments. The electrochemical impedance spectroscopy, and cyclic polarization tests were used to evaluate the corrosion resistance of stainless steels in detergents. Scanning electron microscopy was used to study the morphology of the steel surfaces after corrosion tests. The AISI 304 L steel showed the highest corrosion resistance in the enzymatic medium among the three stainless steels tested. The AISI 316 L stainless steel had an intermediate behavior in enzymatic medium, and lower impedance when immersed in alkaline detergent, while the AISI 420 steel showed deeper corrosion marks on its surface in alkaline medium when compared to the enzymatic solution. The enzymatic detergent is the less aggressive medium to the steels, and the AISI 304 L SS was the most corrosion resistant in alkaline and enzymatic detergents. • The corrosion resistance of stainless steels used in surgical instruments was evaluated • The aggressiveness to the steel of alkaline and enzymatic detergents was studied • The electrochemical impedance spectroscopy and cyclic polarization were used • The AISI 304 steel showed the highest corrosion resistance in enzymatic medium • The enzymatic detergent is the less aggressive medium to the steels

A Review of Cross-Disciplinary Approaches for the Identification of Novel Industrially Relevant Plastic-Degrading Enzymes

The large-scale global use of plastics has led to one of the greatest environmental issues of the 21st century. The incredible durability of these polymers, whilst beneficial for a wide range of purposes, makes them hard to break down. True recycling of plastics is difficult and expensive, leading to accumulation in the environment as waste. Recently, a new field of research has developed, aiming to use natural biological processes to solve this man-made problem. Incredibly, some microorganisms are able to produce enzymes with the capacity to chemically break down plastic polymers into their monomeric building blocks. At an industrial scale, this process could allow for a circular recycling economy, whereby plastics are broken down, then built back up into novel consumer plastics. As well as providing a solution for the removal of plastics from the environment, this would also eliminate the need for the creation of virgin plastics. Analytical techniques, such as those allowing quantification of depolymerisation activity and enzyme characterization, have underpinned this field and created a strong foundation for this nascent inter-disciplinary field. Recent advances in cutting-edge ‘omics approaches such as DNA and RNA sequencing, combined with machine learning strategies, provide in-depth analysis of genomic systems involved in degradation. In particular, this can provide understanding of the specific protein sequence of the enzymes involved in the process, as well as insights into the functional and mechanistic role of the enzymes within these microorganisms, allowing for potential high-throughput discovery and subsequent exploitation of novel depolymerases. Together, these cross-disciplinary analytical techniques offer a complete pipeline for the identification, validation, and upscaling of potential enzymatic solutions for industrial deployment. In this review, we provide a summary of the research within the field to date, the analytical techniques most commonly applied for enzyme discovery and industrial upscaling, and provide recommendations for a standardised approach to allow research conducted in this field to be benchmarked to ensure focus is on the discovery and characterisation of industrially relevant enzymes.

Simulation of novel jellyfish type of process for bioremediation application

A bioinspired device was fabricated as a sustainable remedial method and its performance as a membrane-enzyme reactor with cyclic ultrafiltration was investigated. The body of the jellyfish-like device was composed of two parts: 1) Jellyfish arms: Mono and co-axial electrospinning have been utilized to synthesize the flexible parts (e.g., multilayer membrane PS-PVDF/PAN/PS-PVDF) used for immobilization of aliphatic degrading enzymes, and 2) Jellyfish tentacles: Hollow fiber membranes were selected for physical immobilization of polycyclic aromatic hydrocarbon (PAH) degrading enzymes. To study the behavior of the membrane/enzyme reactor, the hollow fiber enzyme reactor with pulsation was operated by recycling an enzyme solution to assess ultrafiltration efficiency. A mathematical model was suggested to describe the experimental data obtained in this study to predict the effectiveness of the reactor for PAH removal. When testing the performance of the jellyfish-like device, those equipped with nanofibers with an oil sorption capacity of (10. ±0.7gdilbit/gfiber) were more effective at removing oil particles before they touched the hollow fiber membrane surface. Moreover, the reaction rate measured in a free soluble enzyme and a recirculating immobilized enzyme solution exhibited a slight difference in the kinetic parameter, Km (0.03 and 0.021 mM) due to the internal diffusional resistance. Based on biodegradation studies, a synergistic effect between membrane adsorption, enzymatic degradation, and ultrafiltration was proposed for the removal of anthracene from the column of water.

AFM Investigation of the Influence of Steam Flow through a Conical Coil Heat Exchanger on Enzyme Properties.

The present study is aimed at the revelation of subtle effects of steam flow through a conical coil heat exchanger on an enzyme, incubated near the heat exchanger, at the nanoscale. For this purpose, atomic force microscopy (AFM) has been employed. In our experiments, horseradish peroxidase (HRP) was used as a model enzyme. HRP is extensively employed as a model in food science in order to determine the influence of electromagnetic fields on enzymes. Adsorption properties of HRP on mica have been studied by AFM at the level of individual enzyme macromolecules, while the enzymatic activity of HRP has been studied by spectrophotometry. The solution of HRP was incubated either near the top or at the side of the conically wound aluminium pipe, through which steam flow passed. Our AFM data indicated an increase in the enzyme aggregation on mica after its incubation at either of the two points near the heat exchanger. At the same time, in the spectrophotometry experiments, a slight change in the shape of the curves, reflecting the HRP-catalyzed kinetics of ABTS oxidation by hydrogen peroxide, has also been observed after the incubation of the enzyme solution near the heat exchanger. These effects on the enzyme adsorption and kinetics can be explained by alterations in the enzyme hydration caused by the influence of the electromagnetic field, induced triboelectrically by the flow of steam through the heat exchanger. Our findings should thus be considered in the development of equipment involving conical heat exchangers, intended for either research or industrial use (including miniaturized bioreactors and biosensors). The increased aggregation of the HRP enzyme, observed after its incubation near the heat exchanger, should also be taken into account in analysis of possible adverse effects from steam-heated industrial equipment on the human body.

Synthesis of polymer-functionalized β-cyclodextrin, Mg2+ doped, coating magnetic Fe3O4 nanoparticle carriers for penicillin G acylase immobilization

Based on the beneficial effects of magnesium ions on biomolecules, it was first introduced into enzyme immobilization carriers to promote the comprehensive performance of immobilized enzymes in repeated use. Firstly, Fe3O4 nanoparticles (NPs) were prepared by the inverse microemulsion method, then, Mg2+ was doped in the process of coating β-cyclodextrin (β-CD) on the surface of Fe3O4 NPs. Afterward, through acetal reaction, glutaraldehyde (GA) was grafted to the material’s surface to obtain Mg2+-Fe3O4 @ β-CD-g-GA NPs, and covalent bond immobilization of Penicillin G acylase (PGA) was realized by the Schiff base reaction of the carriers’ aldehyde group and PGA’s amino group. After optimization of process conditions, the results show that the performances of the immobilized PGA achieve the best, when the concentration of the enzyme solution is 3.00 vol%, pH is 8.0, the immobilization time is 30 h, and the immobilization temperature is 37 °C. Its enzyme activity recovery (EAR), enzyme activity (EA) and enzyme loading capacity (ELC) are 95.6%, 31997 U/g, 110 mg/g, respectively. Lastly, the operational stability, reusability, and storage stability of Mg2+-Fe3O4 @ β-CD-g-GA-PGA NPs were researched. Relative to free PGA, immobilized PGA has better operation stability and storage stability. After 11 repeated uses, the immobilized PGA still has 56% of the initial vitality, and the carrier recovery (Re) is 86.6%.

The response surface methodology for optimization of Halomonas sp. C2SS100 lipase immobilization onto CaCO3 for treatment of tuna wash processing wastewater

An immobilized enzyme could exhibit selectively modified physicochemical properties, and it might offer a better environment for the enzyme activity. In this study, the immobilization yield of crude Halomonas sp. lipase was optimized to improve its stability. Thanks to its high adsorption capacity, CaCO3 has been chosen as support for the immobilization process. Furthermore, response surface methodology (RSM) was used to determine optimal conditions for the immobilization of the bacterial lipase. Five tested factors (enzyme solution, support amount, time, temperature, and acetone volume) were optimized applying a central composite design of RSM. The maximum yield of lipase immobilization was improved to 96%. Furthermore, a biochemical characterization proved a significant improvement of the immobilized lipase stability. The immobilized enzyme is more stable at extreme pH values and high temperatures than the free one. We also tested the reusability of the immobilized lipase by evaluating the recovery of the support using simple filtration. Thanks to its high stability, the immobilized lipase was invested in an effective treatment of tuna wash processing wastewater. The oil biodegradation efficiency was established at 81.5% and was confirmed by Fourier transformation infrared spectrometry. Likewise, the biological oxygen demand values were reduced which makes a possible reduction of the wastewater pollution degree.

β-Glucosidase Immobilized on Magnetic Nanoparticles: Controlling Biomolecule Footprint and Particle Functional Group Density to Navigate the Activity-Stability Tradeoff.

In the present work, the immobilized footprint of β-glucosidase (BGL) on silica-coated iron oxide was explored to produce reusable catalysts with flexible active sites for high activity and heightened storage stability. Synthesized iron oxide particles were coated with silica and functionalized with various densities of (3-aminopropyl)triethoxysilane (APTES) to obtain particles with amine densities ranging from 0 to 3 × 10-5 mol/g particle. The amine-modified particles were activated with glutaraldehyde, and subsequently, BGL was immobilized using either a 0.1 or 1 mg/mL enzyme solution to produce biomolecules with a large or small footprint on the particle surface. The initial activity, activity for subsequent hydrolysis cycles, activity after extended storage, and biomolecule footprint were studied as a function of APTES density and concentration of enzyme used for immobilization. At high immobilization amounts, the specific activity and footprint were reduced, but the immobilized biomolecules were stable during storage. However, at low enzyme immobilizations, the activity of the enzymes was retained, the immobilized enzymes adopted large footprints, and the storage stability increased with APTES density relative to the free enzyme. These results highlight how controlling both the protein load and functional group density can yield immobilized enzymes possessing high activity, which are stable during storage.

Corn Stover Pretreatment with Na2CO3 Solution from Absorption of Recovered CO2

Renewable resources such as lignocellulosic biomass are effective at producing fermentable sugars during enzymatic hydrolysis when pretreated. Optimizing pretreatment methods for delignification while maintaining sustainability and low processing costs requires innovative strategies such as reusing greenhouse gas emissions for materials processing. Corn stover, an agricultural waste residue, was pretreated with 2.2 M Na2CO3 produced from CO2 captured via absorption in a 5 M NaOH solution. Composition analysis of the pretreated corn stover exhibited higher cellulose content (40.96%) and less lignin (16.50%) than the untreated biomass. Changes in the chemical structures are visible in the FTIR-ATR spectra, particularly in the cellulose and lignin-related absorption bands. The sugar release from hydrolysis was evaluated at different time intervals and by varying two enzyme ratios of CTec2-to-HTec2 (2:1 and 3:1). Enzymatic hydrolysis produced higher and more stable glucose yields for the pretreated biomass, surpassing 90% after 24 h using the 3:1 enzyme ratio. Sugar concentrations notably increased after pretreatment and even more when using the cellulase-rich enzyme solution. The maximum glucose, xylose, and arabinose recovered were 44, 19, and 2.3 g L−1. These results demonstrate the viability of capturing CO2 and converting it into an efficient Na2CO3 pretreatment for corn stover biomass. Additional processing optimizations depend on the combination of physicochemical parameters selected.

Coupling chemical degumming with enzymatic degumming of ultrasound pre-treated ramie fiber using Bacillus subtilis ABDR01

In this study, we designed, developed, and optimizedan enzymatic fiber degumming process. First, a Partially Purified Enzyme Solution (PPES) containing pectinase, cellulase and xylanase was co-produced via ultrasound-assisted fermentation of orange peels using Bacillus subtilis ABDR01. Then, degumming with PPES was optimized for pH, temperature, incubation time and PPES concentration. Finally, three sets of ultrasound-assisted degumming experiments — chemical, enzymatic, and coupled enzymatic-chemical degumming with reduced alkali concentration (0.5% NaOH, 30 min) were performed. Chemical degumming for 2 h with 1% NaOH at 98 °C resulted in residual gum content of 5.9%, whereas ultrasound-assisted (30 min, 25 kHz) chemical degumming reduced the gum content to 3.1%. At a pH of 7.5, temperature of 45 °C, 30 h incubation and 2% PPES, gum removal efficiency of >60% and fibers with <10% residual gum were produced. Coupling the ultrasound-assisted enzymatic degumming with reduced alkali chemical degumming resulted in gum content of <3%. • High frequency ultrasonicaton pre-treatment improves fiber degumming. • Combining enzymatic and chemical degumming reduced NaOH demand. • Optimized the ultrasound-assisted enzymatic degumming of Ramie fibers. • Fibers (ø 0.05 mm) containing < 3% gum and high tensile strength produced.

Fruit Enzyme Activity of Molasses Combination Against Control of S. Aureus and E. Coli Bacteria

This study aims to determine the effect of the pineapple and orange peel enzymes combined with molasses on the effectiveness of the inhibition zones of gram positive (S. Aureus) and gram negative (E. Coli) bacteria. The study was conducted using the disc method, where the bacterial samples used were at a concentration of 200 millimicrons. And the addition of enzyme solution with a concentration of 20 millimicrons. In observations using Scan 500, the results obtained in samples of pineapple peel with the highest inhibition zones for E. Coliand S. Aureusbacteria, respectively, of 11.7 mm and 12.8 mm. Meanwhile, in the test of orangepeel samples, the highest inhibition zones for E. Coliand S. Aureuswere 16.6 mm and 13.2 mm, respectively. In addition, the highest pH testing of each sample of orange peel and pineapple was obtained on the third fermentation with values of 4.63 and 4.39.