Rugosa Lipase Research Articles

Characteristic study of Candida rugosa lipase immobilized on lignocellulosic wastes: effect of support material.

For the first time is reported the comparison of solid biocatalysts derived from Candida rugosa lipase (CRL) immobilized on different lignocellulosic wastes (rice husk, brewer's spent grain, hemp tea waste, green tea waste, vine bark, and spent coffee grounds) focusing on the characterization of these materials and their impact on the lipase-support interaction. The wastes were subjected to meticulous characterization by ATR-FTIR, BET, and SEM analysis, besides lignin content and hydrophobicity determination. Investigating parameters influencing immobilization performance revealed the importance of morphology, textural properties, and hydrophobic interactions revealed the importance of morphology, textural properties and especially hydrophobic interactions which resulted in positive correlations between surface hydrophobicity and lipase immobilization efficiency. Hemp tea waste and spent coffee grounds demonstrated superior immobilization performances (7.20 U/g and 8.74 U/g immobilized activity, 102.3% and 33.5% efficiency, 13.4% and 15.4% recovery, respectively). Moreover, they demonstrated good temporal stability (100% and 92% residual activity after 120days, respectively) and retained 100% of their immobilized activity after five reuses in the hydrolysis of p-nitrophenyl palmitate in hexane. In addition, the study of enzymatic desorption caused by ionic strength and detergent treatments indicated mixed hydrophobic and electrostatic interactions in rice husk, vine bark, and spent coffee grounds supports, while hemp tea waste and green tea waste were dominated by hydrophobic interactions.

Enzymatic Kinetic Resolution of Racemic 1-(Isopropylamine)-3-phenoxy-2-propanol: A Building Block for β-Blockers.

This study aimed to optimize the kinetic resolution of building blocks for the synthesis of β-blockers using Candida rugosa lipases, which could be potentially used to synthesize enantiomerically pure β-blockers further. Reaction mixtures were incubated in a thermostated shaker. Qualitative and quantitative analyses of the reaction mixtures were performed using chiral stationary phases and the UPLC-IT-TOF system. Of the 24 catalytic systems prepared, a system containing lipase from Candida rugosa MY, [EMIM][BF4] and toluene as a two-phase reaction medium and isopropenyl acetate as an acetylating agent was optimal. This resulted in a product with high enantiomeric purity produced via biotransformation, whose enantioselectivity was E = 67.5. Using lipases from Candida rugosa enables the enantioselective biotransformation of the β-blockers building block. The biocatalyst used, the reaction environment, and the acetylating agent significantly influence the efficiency of performer kinetic resolutions. The studies made it possible to select an optimum system, a prerequisite for obtaining a product of high enantiomeric purity. As a result of the performed biotransformation, the (S)-enantiomer of the β-blocker derivative was obtained, which can be used to further synthesize enantiomerically pure β-blockers.

Continuous Process Verification 4.0 application in upstream: adaptiveness implementation managed by AI in the hypoxic bioprocess of the Pichia pastoris cell factory.

The experimental approach developed in this research demonstrated how the cloud, the Internet of Things (IoT), edge computing, and Artificial Intelligence (AI), considered key technologies in Industry 4.0, provide the expected horizon for adaptive vision in Continued Process Verification (CPV), the final stage of Process Validation (PV). Pichia pastoris producing Candida rugosa lipase 1 under the regulation of the constitutive GAP promoter was selected as an experimental bioprocess. The bioprocess worked under hypoxic conditions in carbon-limited fed-batch cultures through a physiological control based on the respiratory quotient (RQ). In this novel bioprocess, a digital twin (DT) was built and successfully tested. The implementation of online sensors worked as a bridge between the microorganism and AI models, to provide predictions from the edge and the cloud. AI models emulated the metabolism of Pichia based on critical process parameters and actionable factors to achieve the expected quality attributes. This innovative AI-aided Adaptive-Proportional Control strategy (AI-APC) improved the reproducibility comparing to a Manual-Heuristic Control strategy (MHC), showing better performance than the Boolean-Logic-Controller (BLC) tested. The accuracy, indicated by the Mean Relative Error (MRE), was for the AI-APC lower than 4%, better than the obtained for MHC (10%) and BLC (5%). Moreover, in terms of precision, the same trend was observed when comparing the Root Mean Square Deviation (RMSD) values, becoming lower as the complexity of the controller increases. The successful automatic real time control of the bioprocess orchestrated by AI models proved the 4.0 capabilities brought by the adaptive concept and its validity in biopharmaceutical upstream operations.

Preparation of two new chiral metal-organic frameworks for lipase immobilization and their use as biocatalysis in the enantioselective hydrolysis of racemic naproxen methyl ester

Considering the selective pharmacological activity of chiral drugs, it is important to develop new chiral materials to synthesize them. In this work, two new chiral MOFs (UiO-66@Np and UiO-66@Ib) were prepared by the covalent attachment of the chiral compounds (S-naproxen and S-ibuprofen) to the amine-functionalized Zr-MOF (UiO-66-NH2). Then, Candida rugosa lipase (CRL) was immobilized on these chiral MOFs to fabricate two new biocomposites (UiO-66@Np@CRL and UiO-66@Ib@CRL) as effective biocatalysts, which enable significant enhancement in the catalytic activity and enantioselectivity of lipase. The FTIR, SEM, EDX, TGA, and PXRD analyses were carried out to confirm the formation of the biocomposites. The catalytic performances of the biocomposites (UiO-66@Np@CRL and UiO-66@Ib@CRL) were evaluated in the typical hydrolysis of p-nitro-phenyl palmitate (p-NPP) and the catalytic enantioselective hydrolysis of (R,S)-naproxen methyl ester. Under optimal conditions, UiO-66@Np@CRL showed a higher enantiomeric excess of the substrate (ees) value and a 98 % and 50 % conversion rate, respectively, than that of UiO-66@Ib@CRL. Besides, an excellent enantioselectivity (E) value of 458 was obtained in the presence of the biocomposite (UiO-66@Np@CRL). In addition, it was observed that the catalytic activity, conversion rate and ees value of this composite (UiO-66@Np@CRL) remained almost unchanged in reuse after 6 months. The results showed that in enantioselective reactions, the highest conversion and enantioselectivity could be achieved when the chiral compound bound to the MOF and the model compound used are the same.

Enhanced catalytic performance of Candida rugosa lipase through immobilization on zirconium-2-methylimidazole: A novel biocatalyst approach

Immobilization of enzymes on suitable supports is a critical approach for enhancing enzyme stability, reusability, and overall catalytic efficiency. This study explores the immobilization of Candida rugosa lipase on zirconium-based 2-methylimidazole (ZrMI) nanoparticles, aiming to develop a stable and reusable biocatalyst. The ZrMI was produced via a solvothermal technique and analyzed using various characterization methods. Candida rugose lipase was immobilized using cross-linking agents, achieving an 87 % immobilization efficiency. The immobilized enzyme exhibited significantly enhanced thermal stability, broader pH tolerance, and increased catalytic efficiency compared to free C. rugose lipase. The ZrMI@lipase retained 69 % of its enzymatic activity following a 60-day storage period at 4 °C. Notably, it displayed significant reusability, maintaining 65 % of its original activity after undergoing 15 catalytic cycles. Examination of the kinetics revealed that the immobilized enzyme possessed a heightened substrate affinity (Km of 4.1 mM) and maximal reaction rate (Vmax of 85.7 μmol/ml/min) in comparison to the free enzyme (Km of 5.4 mM and Vmax of 69 μmol/ml/min), indicating enhanced catalytic efficiency. Validation through zeta potential and hydrodynamic size assessments verified the successful binding of the enzyme and the consistent colloidal characteristics. These results suggest that ZrMI is a promising support for C. rugose lipase immobilization, offering improved stability and reusability for various industrial applications. The study highlights the potential of ZrMI@lipase as an efficient and durable biocatalyst, contributing to advancements in enzyme immobilization technology and sustainable industrial processes.

Chemo-enzymatic, regioselective synthesis of dihydropyrimidinone-fused β-amino alcohols and their anti-inflammatory and antioxidant activity evaluation

A highly regioselective and efficient method has been developed for synthesizing novel β-amino alcohols fused with dihydropyrimidin-2-one. This method utilizes the enzyme Novozyme-435 to catalyze the reaction between epoxides and various aliphatic amines in acetonitrile. Novozyme-435 outperformed other catalysts, including Porcine Pancreatic Lipase (PPL), Pseudomonas aeruginosa lipase (PAL), and Candida rugosa lipase (CRL). This process yielded two series of β-amino alcohols (compounds 8a-h and 9a-h), whose structures were confirmed through IR, NMR (1H, 13 C), and HRMS analyses. The anti-inflammatory and antioxidant properties of these compounds were evaluated, revealing mild to moderate inhibition of TNF-α-induced ICAM-1 expression in primary human endothelial cells, with compounds 9a and 9c showing approximately 60% inhibition. Antioxidant activity, assessed using the DPPH (2,2-diphenyl-1-picrylhydrazyl) method, indicated that compounds 9a, 9b, 9c, and 9 g had the superior activity than others. This study highlights the potential of these β-amino alcohols fused with dihydropyrimidin-2-one as anti-inflammatory and antioxidant agents.

Functionalized ZnFe2O4@Mesoporous silica nano-support for lipase enzyme immobilization: Enhanced biocatalysis and antibacterial activity for food industry applications

Enzymes, as highly efficient biocatalysts, play a crucial role in diverse biotechnological applications. However, enzyme (re)purification and recovery challenges pose substantial obstacles, rendering them costly for industrial utilization. Addressing this, our study developed mesoporous zinc ferrite nanoparticles coated with amine-functionalized mesoporous silica structure (ZnFe2O4@MS) through the solvothermal method. These nanoparticles are an effective nano-support for immobilizing Candida rugosa Lipase (CRL), ensuring easy separability and recoverability through a magnetic field and providing a significant surface area for high mass transference capacity. Immobilizing the lipase on the nano-support (ZnFe2O4@MS/CRL) through covalent bonds on the surface and within the pores led to a notable increase in enzyme activity from 324 U/mg for free enzyme to 689 U/mg for immobilized enzyme. This indicates unchanged active sites post-immobilization, accompanied by enhanced catalytic activity. The immobilized lipases demonstrated prolonged stability at elevated temperatures, maintaining over 59% of initial catalytic activity through five cycles. Furthermore, ZnFe2O4@MS/CRL exhibited substantial antibacterial activity against Staphylococcus aureus, more than that observed in Gram-negative bacteria. The immobilized lipase was also effective in synthesizing banana flavor (isoamyl acetate) in n-hexane, achieving 64% esterification at 45 °C after 4 h. Overall, the study underscores the industrial promise of this immobilized enzyme system, emphasizing its potential for sustainable and cost-effective biotechnological processes.

Co-immobilization of magnesium precursor and Candida rugosa lipase on alumina via covalent bonding for biodiesel production

Catalyst development is one of the most challenging aspects in biodiesel production due to the compatibility issue with the feedstocks and catalyst poisoning upon exposure to process conditions. In the present research, chemoenzymatic catalyst was prepared by impregnation of magnesium aluminate (MA) on alumina support, modifying the support using activating agent, and immobilization of Candida rugosa lipase (CRL) onto the support. The optimum immobilization condition was resulting in 85.07 % of activity recovery of immobilized lipase, equivalent to 30.53 % of immobilization efficiency. The catalysts were characterized through XRD, FTIR, FESEM-EDX, NAA and CO2-TPD confirming the mixture of AlO(OH), magnesium aluminate (MA), (MgNO3)2 and Mg2+ on the support. CRL/L-lysine/MA showed remarkable improvement in thermal stability (40 − 70℃), solvents tolerance (ethanol, methanol, isopropanol, tert-butanol), and storage stability at 30 ℃, as compared to free CRL. CRL/L-lysine/MA successfully produced biodiesel yield up to 87.10 % at 200.71 rpm of agitation speed, 13.58 % (w/w) of water content, 10 h reaction time, 15 % (w/w) of catalyst loading, at 50 ℃, using 1:12 of oil to methanol molar ratio, and retaining 46.60 % of biodiesel yield after six cycle. The CRL/L-lysine/MA demonstrated novel catalysts characteristics comprising chemical and biological active species on a single support for biodiesel production from waste cooking oil (WCO).

Hydrophobic Residues Promote Interfacial Activation of Candida rugosa Lipase: A Study of Rotational Dynamics.

Microbial lipases constitute a class of biocatalysts with the ability to cleave ester linkages of long-chain triglycerides. This property makes them particularly attractive for industrial applications ranging from food processing to pharmaceutical preparation. Among such enzymes, Candida rugosa lipase (CRL) is one of the most frequently used in biotransformation. A notable feature of CRL, among many lipases, is its propensity for interfacial activation: these enzymes exhibit elevated catalytic rates when acting at the interface between aqueous and hydrophobic phases. Notably, this phenomenon can be attributed to the presence of a mobile lid domain, which in its closed state occludes the enzyme active site. To advance our understanding of interfacial activation, we explore the dynamics of CRL rotation at the octane-water interface in this work. To do so, we employ molecular dynamics and umbrella sampling to evaluate the free energy of rotation of the enzyme at the interface. We identify a global minimum in the rotational landscape that coincides with lid opening at the interface. Additionally, we investigate the role of surface residues outside the lid domain as they serve to instigate rotation of the lid toward the aqueous phase. In doing so, we identify a patch of leucine residues which when mutated to glycine impose a barrier to rotation that maintains the enzyme in the inactive (closed lid) state on the order of 1 μs. Importantly, this study presents a novel quantification of the rotational free energy corresponding to CRL lid opening at the octane-water interface. The accompanying mutagenesis study likewise clarifies the role of hydrophobic surface residues in the transition. As such, this work provides valuable insight into the phenomenon of interfacial activation that might open up new avenues for manipulating the microenvironment of industrially relevant lipases, affording enhanced control over the enzyme state.

Scientific prospecting of lignocellulosic and inorganic supports used to immobilize Candida rugosa lipase in the biodiesel production process

Scientific prospecting of lignocellulosic and inorganic supports used to immobilize Candida rugosa lipase in the biodiesel production process

Boosting catalytic efficiency of lipase by regulating amphiphilic microenvironment through reversible addition-fragmentation chain transfer polymerized modifications on polyacrylonitrile fiber

Lipases are increasingly attracting attention in green and sustainable biodiesel production. Currently, the research emphasis lies in immobilizing unstable lipase onto carriers to enhance its performance. Polyacrylonitrile fiber (PANF) is considered to be a promising material for lipase immobilization due to its excellent properties. In this study, functional carriers with regulated surface hydrophobicity were obtained by loading functional groups on PANF via reversible addition-fragmentation chain transfer (RAFT) polymerized modification, and Candida rugosa lipase (CRL) was covalently immobilized on the carrier with glutaraldehyde as a linker. By employing this optimized biocatalyst PANF@BMA&2VImBr-NH2-CRL in the transesterification process, the yield of biodiesel derived from soybean oil reached an impressive 92.7 %. The outstanding performance can be attributed to the activation of lipase interface induced by hydrophobic microenvironment derived from alkyl ester on the carrier skeleton. Moreover, the stability and storage performance of immobilized lipase were significantly improved. The immobilized lipase exhibited facile recovery and maintained a consistent biodiesel yield of 80.9 % even after undergoing 5 cycles of reuse, thereby highlighting its potential for sustainable production. To sum up, our research demonstrates that the designed and prepared process of PANF-supported lipase offers a promising approach for enzyme immobilization, thereby presenting extensive potential applications in the field of biotechnology.

Thermo-Switchable Enzyme@Metal-Organic Framework for Selective Biocatalysis and Biosensing.

The stimulus-responsive regulation of enzyme catalytic activity and selectivity provides a new opportunity to extend the functionality and efficiency of immobilized enzymes. This work aims to design and synthesize a thermo-switchable enzyme@MOF for size-selective biocatalysis and biosensing through the immobilization of Candida rugosa lipase (CRL) within ZIF-8 functionalized with thermally responsive polymer, poly(N-isopropylacrylamide) (PNIPAM) (CRL@ZIF-8-PNIPAM). Unlike free CRL, which does not demonstrate substrate selectivity, we can reversibly tune the pore size of the ZIF-8-PNIPAM nanostructures (open pores or blocked pores) through temperature stimulus and subsequently modulate the substrate selectivity of CRL@ZIF-8-PNIPAM. CRL@ZIF-8-PNIPAM had the highest hydrolytic activity for small molecules (12 mM p-nitrophenol/mg protein/min, 4-nitrophenyl butyrate (p-NP Be)) and the lowest hydrolytic activity for large molecules (0.16 mM p-nitrophenol/mg protein/min, 4-nitrophenyl palmitate (p-NP P)). In addition, CRL@ZIF-8-PNIPAM demonstrated thermo-switchable behavior for large molecules (p-NP P). The p-NP P hydrolytic activity of CRL@ZIF-8-PNIPAM was significantly lower at 40 °C (blocked pores) than at 27 °C (open pores). However, the transition of blocked pores and open pores is a gradual process that resulted in a delay in the "thermo-switchable" catalytic behavior of CRL@ZIF-8-PNIPAM during thermal cycling. CRL@ZIF-8-PNIPAM was also successfully used for the fabrication of electrochemical biosensors for the selective biosensing of pesticides with different molecular sizes.

Investigation on solvent-free esterification of oleic acid by hemp tea waste-immobilized Candida rugosa lipase

This study aimed at Candida rugosa lipase immobilization on a low-cost and readily available support. Among agro-industrial crops, hemp tea waste was chosen as the carrier because it provides higher immobilization performance than hemp flower and leaf wastes. Support characterization by ATR-FTIR, SEM and elemental analysis and the optimization of the adsorption immobilization process were performed. The lipase adsorption immobilization was obtained by soaking the support with hexane under mild agitation for 2 h and a successively incubating the enzyme for 1 h at room temperature without removing the solvent. The esterification of oleic acid with n-decanol was tested in a solvent-free system by studying some parameters that influence the reaction, such as the substrates molar ratio, the lipase activity/oleic acid ratio, reaction temperature and the presence/absence of molecular sieves. The biocatalyst showed the ability to bring the esterification reaction to equilibrium under 60 min and good reusability (maintaining 60 % of its original activity after three successive esterification reactions) but low conversion (21 %) at the optimized conditions (40 °C, 1:2 substrates molar ratio, 0.56 lipase/oleic acid ratio, without sieves). Comparing the results with those obtained by free lipase form at the same activity (1 U) and experimental conditions, slightly higher conversion (%) appeared for the free lipase. All this highlighted that probably the source of lipase for its carbohydrate-binding pocket and lid structure affected the esterification of oleic acid but certainly, the immobilization didn’t induce any lipase conformational change also allowing the reuse of the catalytic material.

Immobilized lipase based on SBA-15 adsorption and gel embedding for catalytic synthesis of isoamyl acetate

In this study, Candida rugosa lipase (CRL) was immobilized on the silica carrier SBA-15 by physical adsorption, then mixed with hydrophobically modified sodium alginate and dropped into calcium chloride solution. Finally, the adsorption-embedding immobilized lipase (L-SBA-15-Mgel) was successfully prepared. The conformation of the carrier before and after lipase immobilization were analyzed by scanning electron microscopy (SEM), thermogravimetric analysis (TGA), Fourier transform infrared spectroscopy (FT-IR) and other characterization techniques. The molecular docking and molecular dynamics simulation of CRL and modifier dodecyl glycidyl ether (DGE) ligand were carried out to analyze the potential influence of modifier on enzyme characteristics. The basic enzymatic properties including the activity, stability and reaction kinetic parameters of the immobilized lipase L-SBA-15-Mgel were studied. L-SBA-15-Mgel showed good storage stability and reusability. After 16 days of storage, the activity retention was 47.85%, and there was still 63.20% of the residual activity after seven cycles of reuse. The immobilized lipase was applied to the efficient synthesis of food flavor ester isoamyl acetate. Under the reaction conditions of 50 °C and 200 rpm, the highest yield of isoamyl acetate catalyzed by L-SBA-15-Mgel was 85.19%.

Functionalized Ionic Liquids-Modified Metal-Organic Framework Material Boosted the Enzymatic Performance of Lipase.

The development of immobilized enzymes with high activity and stability is critical. Metal-organic frameworks (MOFs) have attracted much academic and industrial interest in the field of enzyme immobilization due to their unique properties. In this study, the amino-functionalized ionic liquid (NIL)-modified metal-organic framework (UiO-66-NH2) was prepared to immobilize Candida rugosa lipase (CRL), using dialdehyde starch (DAS) as the cross-linker. The results of the Fourier transform infrared (FT-IR) spectra, X-ray powder diffraction (XRD), and scanning electronic microscopy (SEM) confirmed that the NIL was successfully grafted to UiO-66-NH2. The CRL immobilized on NIL-modified UiO-66-NH2 (UiO-66-NH2-NIL-DAS@CRL) exhibited satisfactory activity recovery (79.33%), stability, reusability, and excellent organic solvent tolerance. The research results indicated that ionic liquid-modified UiO-66-NH2 had practical potential for application in enzyme immobilization.

WITHDRAWN: Catalytic activity of Candida rugosa lipase immobilized on lignocellulosic wastes: Effect of support material

WITHDRAWN: Catalytic activity of Candida rugosa lipase immobilized on lignocellulosic wastes: Effect of support material

Magnetic biocatalytic nanoreactors based on graphene oxide with graded reduction degrees for the enzymatic synthesis of phytosterol esters

As we know, the carriers of enzymatic nanoreactors with different amphipathy have significant influence on the interfacial enzyme activities and its performance. In this study, the magnetic graphene oxide (GO) enzyme carriers were constructed by assembling graphene oxide with different reduction degrees (rGO(x)) and polyethyleneimine (PEI) modified nano-flowery Fe3O4 into composite materials through electrostatic interactions, and x here represents the reduction reaction hours of GO. The SEM, TEM, contact angles and BET tests qualified the ideal enzyme carrier, PEI-Fe3O4-rGO(12), by adsorption-crosslinking method, the Candida rugosa lipase (CRL) was immobilized to obtain the lipase nanoreactor, PEI-Fe3O4-rGO(12)@CRL. The nano-sheet and nano-flower structures provided a larger specific surface area and suitable pore size, and the CRL had higher protein load on the carrier material (163.11 mg/g). In addition, using hydrolyzed p-nitrophenol palmitate as the model, the PEI-Fe3O4-rGO(12)@CRL can still show 76.5 % residual activity after 8 repeated uses. At the same time, the immobilized lipase showed good catalytic performance in the biosynthesis of phytosterol esters. The optimal esterification conditions were determined by single factor experiment (isooctane, sterol concentration 150 μmol/mL, alkyd molar ratio 1:2, enzyme dosage 120 mg, 26 h, 55 °C, the esterification rate was 76.24 %). Therefore, the optimal biocatalyst PEI-Fe3O4-rGO(12)@CRL may has good performance in the biosynthesis of other novel foods.

Evaluation of Heavy Metal Concentrations in Enzyme Embedded Poly Lactic Acid (EPLA) by ICP-OES: Lipase from Candida Rugosa.

Abstract: Enzymes like lipases are essential for catalyzing important reactions in the fields of biotechnology and industrial operations. For quality control and safety evaluations, however, the quantification of these contaminants is crucial because the presence of heavy metals in enzyme preparations can negatively affect their activity and stability. Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES), a key component of this investigation, is used to measure the levels of heavy metals in lipase enzyme preparations embedded in polylactic acid (PLA). The lipase enzyme being studied comes from Candida rugosa, which is well-known for its considerable industrial applications. According to preliminary findings, the lipase enzyme is successfully protected from external heavy metal pollutants by the encapsulating technique, maintaining its enzymatic activity and stability. The lipase-PLA composite had only minimal quantities of heavy metals, according to the ICP-OES study. The research's conclusions show great potential for the creation of reliable and contaminant-free lipase enzyme preparations, which will increase their suitability for usage in a variety of industrial processes and encourage the use of biodegradable polymers, supporting environmental sustainability. By highlighting the significance of quality control and safety assessment through the determination of heavy metal concentrations, this research contributes to the larger field of enzyme biotechnology. It also highlights the potential of Candida Rugosa lipase implanted in PLA matrices for eco-friendly and sustainable biocatalytic applications in sectors ranging from food and pharmaceuticals to biofuel generation and other biodegradable polymers.

Assessment of the efficiency and stability of enzymatic membrane reaction utilizing lipase covalently immobilized on a functionalized hybrid membrane

Enzyme immobilization in membrane bioreactors has been considered as a practical approach to enhance the stability, reusability, and efficiency of enzymes. In this particular study, a new type of hybrid membrane reactor was created through the phase inversion method, utilizing hybrid of graphene oxide nanosheets (GON) and polyether sulfone (PES) in order to covalently immobilize the Candida rugosa lipase (CRL). The surface of hybrid membrane was initially modified by (3-Aminopropyl) triethoxysilane (APTES), before the use of glutaraldehyde (GLU), as a linker, through the imine bonds. The resulted enzymatic hybrid membrane reactors (EHMRs) were then thoroughly analyzed by using field-emission scanning electron microscopy (FE-SEM), contact angle goniometry, surface free energy analysis, X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, attenuated total reflection (ATR), and energy-dispersive X-ray (EDX) spectroscopy. The study also looked into the impact of factors such as initial CRL concentration, storage conditions, and immobilization time on the EHMR's performance and activity, which were subsequently optimized. The results demonstrated that the CRLs covalently immobilized on the EHMRs displayed enhanced pH and thermal stability compared to those physically immobilized or free. These covalently immobilized CRLs could maintain over 60% of their activity even after 6 reaction cycles spanning 50 days. EHMRs are valuable biocatalysts in developing various industrial, environmental, and analytical processes.

Enhanced triolein and ethyl ferulate interesterification performance by CRL-AuNPs

This study established a Candida rugosa lipase (CRL) system to catalyze triolein and ethyl ferulate interesterification. The products were identified, and the binding mode between the substrates and CRL was predicted through molecular docking. Three methods for preparing CRL-AuNPs were proposed and characterized. It was found that the addition of 40 mL of 15 nm gold nanoparticles increased the CRL activity from 3.05 U/mg to 4.75 U/mg, but the hybridization efficiency was only 32.7 %. By using 4 mL of 0.1 mg/mL chloroauric acid, the hybridization efficiency was improved to 50.7 %, but the enzyme activity was sharply decreased. However, when the molar ratio of Mb to HAuCl4 was 0.2, the hybridization efficiency increased to 71.8 %, and the CRL activity was also enhanced to 5.98 U/mg. Under optimal conditions, the enzyme activity of CRL-AuNPs③ was maintained at 95 % after 6 repetitions and 85.6 % after 30 days at room temperature.