Gel Entrapment Research Articles

Entrapment of enzymes in silica aerogels

Aerogels, the world's lightest solids, possess extraordinary traits such as very low density, very high surface area, very high porosity and ultra-low heat conductivity. These traits made aerogels favorable in various applications, including high-performance thermal insulators, catalyst supports, electrode materials, random laser matrices, cosmic dust collectors and more. Of the many potential applications of aerogels, one of the most challenging has been the development of a general procedure for bioactive aerogels by the entrapment of enzymes within these air-light materials. The difficulty in reaching this “holy-grail” was dual: The special procedures for obtaining the unique structure of aerogel are destructive to enzymes; and the aerogels are extremely sensitive to any procedural modification. Thus, the use of pure silica aerogel for the entrapment of enzymes was not known. Here we present a generalized, bio-friendly procedure for the entrapment of enzymes in silica aerogel, retaining both the enzymatic activity and the air-light structure of the aerogel. All of the aerogel synthesis steps were modified and optimized for reducing the risk of enzyme denaturation, while preserving the aerogel characteristic structure of the composite. The entrapment of three enzymes of different types was demonstrated: glucose oxidase, acid phosphatase and xylanase. All aerogel-entrapped enzymes showed superior activity over the common method of sol–gel entrapment in xerogels, due to the much wider and open pore network of the former. Michaelis-Menten kinetics was observed for the entrapped enzymes, indicating that the enzymes are highly accessible and diffusional limitations are negligible. The Michaelis-Menten constant, K m , has remained at the same level, indicating that enzyme-substrate affinity was not affected. Thermal stabilization was observed for entrapped acid phosphatase reaching peak activity at 70 °C. Large molecular weight substrates such as xylan for xylanase, are no obstacle for the aerogel matrix, while completely inapplicable for the xerogel. All of these properties are highly relevant for biotechnological applications.

Amylases immobilization by sol–gel entrapment: application for starch hydrolysis

α-amylase from Bacillus subtilis, α-amylase from Aspergillus oryzae, and β-amylase from barley were encapsulated within silica-based matrix by acid-catalyzed sol–gel process. The resulting systems were characterized by Fourier transform infrared spectroscopy (FT-IR) in the attenuated total reflection (ATR) mode, nitrogen adsorption (BET and BJH methods), scanning electron microscopy (SEM), and small-angle X-ray scattering (SAXS). The products from enzymatic hydrolysis of starch were analyzed by the DNS method (reducing sugars) and high-performance liquid chromatography (HPLC). The biocatalytic activity of the immobilized systems (reducing sugars in the range of 0–30.5 μmol/mL) was compared with that of the free systems (reducing sugars in the range of 11.7–33.7 μmol/mL). The porosity analysis showed that xerogels with a high surface area (above 300 m2/g) were obtained. The morphological analyses carried out by microscopy demonstrated the existence of predominantly granular (relatively spherical particles) structures. HPLC results show large differences in the musts obtained from the free enzymes and the corresponding immobilized systems. The encapsulated systems demonstrated high activity and differentiated form of saccharifying the starch.

High Sensitivity Au-based Kretschmann Surface Plasmon Resonance Sensor for Urea Detection

Kretschmann-based Surface Plasmon Resonance (SPR) optical sensor was applied to detect the presence of kidney wastes such as urea in solutions. To enhance the sensitivity of the SPR sensor, nanolaminated gold film (thickness of 50 nm) was used. In this work, the SPR response to urea in various concentrations were measured and investigated using optical wavelengths of 670 nm and 785 nm. The signals were compared between pure urea solution versus mixed solution in the presence of the 0.227 μmol urease enzyme. The proposed mixed solution is to eliminate doping and gel entrapment process for enzyme immobilization in conventional method. Angular interrogation technique was used to measure the sensor performance in urea detection using pure and mixed solutions with urea concentration of 0 - 800 mM. Upon exposure to mixed solution using 785 nm optical wavelength, the nanolaminated gold film exhibited higher SPR sensitivity as much as 7.8 M-1 than a pure urea solution (1.4 M-1). The coupling activity between urea and urease molecules in the mixed solution near the nanolaminated gold film surface lead to sensitivity enhancement. Angle shifting of mixed solution on 50 nm-thick nanolaminated gold film using 670 nm optical wavelength was greater by ~50% compared to 785 nm. Sensorgram data shows a steady and linear increment in SPR incident angle shifting when urea concentration increased. To the best of our knowledge, this is the first time that Kretschmann-based SPR has been used for urea sensing at 670 nm and 785 nm optical wavelengths.

Spore toxin complex recovery from solid-state fermentation of some mosquitocidal Bacilli

ABSTRACTThe efficiency of different methods for recovery of spore toxin complex (STC) from solid-state fermentation (SSF) extracts of Bacillus thuringiensis israelensis (Bti) and Lysinibacillus sphaericus 14N1(Ls14N1) was studied. Drying at 30–50°C, lyophilisation, flocculation by ferric chloride at 0.1–0.2% and encapsulation by entrapment in calcium alginate beads, agar, or polyacrylamide gel were efficient methods for recovery of STC of Ls14N1 and Bti from SSF extracts. Also, STC was precipitated with remained the mosquitocidal activity through adjusting the pH of the SSF liquid extract before centrifugation at pH 2–5 or after addition of acetone (200–400%), ethanol (400%) or acetic acid (2.5–4%) to SSF liquid extract of Ls14N1. On the other hand, STC of Bti showed a good recovery by centrifugation after adjusting SSF liquid extract at pH 4 followed by pH 3 and pH 5 or after the addition of ethanol (300%) or acetone (200–300%). This study contains simple and multiple methods for efficient recovery of STC of Bti and Ls14N1 grown under SSF. The choice of the best method depends on the availability of the necessary equipment as well as the cost of the treatment.

Fermentative Hydrogen Production and Bacterial Community Analysis of Immobilized Sewage Sludge

A novel anaerobic batch fermentation seeded with immobilized sludge was developed for enhanced fermentative hydrogen production using pretreated sweet sorghum substrate as carbon source. Municipal sewage sludge was immobilized to produce hydrogen gas under anaerobic conditions. Cell immobilization was essentially achieved by gel entrapment approach. Hydrogen production was more efficient with the immobilized-cell system than with the free sludge. When the heat treated and acclimatized sludge's were immobilized, the cumulative hydrogen production enhanced. The batch fermentation was operated at a hydraulic retention time (HRT) of 24 h and an influent substrate concentration of 10–40 g/L. With highest concentration of substrate, the acclimatized sludge produced 15.98 mL of H2/g of substrate. In all the treatments, maximum hydrogen yield was obtained at the substrate concentration of 40 g/L, inoculum volume of 10 g/L, at room temperature and HRT of 24 h. The immobilized beads retained 60% of their activity up to three cycles. The best fermentative hydrogen production performance was eventually dominated by presumably enhanced hydrogen-producing bacterial species identified as Escherichia coli.

FORMULATION AND EVALUATION OF ITRACONAZOLE NIOSOMAL GEL

The preparation of niosomes. The main objective of the study was to enhance the antifungal activity of the formulation. Itraconazole is a broad spectrum Imidazole derivative useful in the treatment of superfacial and systemic fungal infection. Various surfactants used were span 40 and span 60. Niosomes were prepared by using Rotary Vaccum evaporation method. Niosomes were prepared using different ratio of drug: surfactant: cholesterol (1:1:1, 1:2:1, 1:3:1).Evaluation of the Niosomal gel was done by determination of drug content 52.81-56.12, Entrapment Efficiency 42.20-45.20,Niosomal gel was prepared using Carbopol 940 (1.5%), Glycerol (10%), Triethanolamine (q.s.) and distilled water up to 15ml. Viscosity was determined by Brookfield programmable ultra-viscometer and the ranges 8173 centipoise. The drug content of the Itraconazole niosomal gel was determined at 262 nm against blank by using UV/visible spectrophotometer and found to be 56.12%. The percentage of drug entrapment in niosomal gel was calculated to be 45.20% .The in-vitro drug release study was carried out using phosphate buffer saline pH 7.4 and was found to be 41.18 ± 1.53% for 12 hours. It can be concluded that the gel formulation containing niosomes loaded with Itraconazole showed prolonged action than formulation containing Itraconazole in non-niosomal form and it can be developed successfully to improve the anti-fungal activity.

Trace determination of Ag(I) after reduction to Ag nanoparticles and sol–gel entrapment by alginic acid hydrogel

The main goal of this work is development of an effective analytical method for trace determination of Ag(I). The novelty of the Ag(I) preconcentration–determination method is mainly referred to the material and the enrichment–detection process (alginic acid gel phase was employed in a pH-switched sol–gel entrapment/Ag fluorescence detection). Ag(I) was reduced to Ag nanoparticles (AgNPs) and then was enriched by the alginic acid hydrogel phase. Then, the formed gel phase (containing AgNPs) was dissolved in alkali solution prior to Ag detection. The enrichment method was highly compatible with spectrofluorimetry, electrothermal atomic absorption spectrometry (ETAAS) and spectrophotometry. Optimization of the reduction-enrichment procedure was performed employing spectrofluorimetry. The linear working range (LWR) and limit of detection (LOD) for Ag(I) determination were found as 0.1–25 and 0.017 µmol L− 1, respectively. The effects of various anions, cations and organic chemicals on Ag(I) determination were spectrofluorimetrically studied. The applied enrichment-ETAAS Ag(I) determination method showed the LWR and LOD as 0.2–6.9 and 0.05 nmol L− 1, respectively. Also, an enrichment factor equal to 30.3 was obtained for the preconcentration method. The method was successfully applied to determine Ag in different water samples, jewels, antimicrobial suspensions and waste X-ray films. X-ray diffraction, transmission electron microscopy, field emission-scanning electron microscopy and EDX-mapping were also employed to characterize the entrapped AgNPs in the alginic acid gel phase. The complimentary experiments showed the alginic acid gel was also applicable for quantitative recovery of silver from the waste radiographic films.

Immobilization of Azospira sp. strain I13 by gel entrapment for mitigation of N2O from biological wastewater treatment plants: Biokinetic characterization and modeling

Development of a strategy to mitigate nitrous oxide (N2O) emitted from biological sources is important in the nexus of wastewater treatment and greenhouse gas emission. To this end, immobilization of N2O-reducing bacteria as a biofilm has the potential to ameliorate oxygen (O2) inhibition of the metabolic activity of the bacteria. We demonstrated the effectiveness of calcium alginate gel entrapment of the nosZ clade II type N2O-reducing bacterium, Azospira sp. strain I13, in reducing levels of N2O, irrespective of the presence of O2. Azospira sp. strain I13cells in the gel exhibited N2O reduction up to a maximum dissolved oxygen concentration of 100μM in the bulk liquid. The maximum apparent N2O uptake rate, [Formula: see text] , by gel immobilization did not appreciably decrease, retaining 72% of the N2O reduction rate of the cell suspension of Azospira sp. strain I13. Whereas gel immobilization increased the apparent half-saturation constant for N2O, [Formula: see text] , and the apparent O2 inhibition constant, [Formula: see text] , representing the degree of O2 resistance, correspondingly increased. A mechanistic model introducing diffusion and the reactions of N2O consumption was used to describe the experimental observations. Incorporating Thieles modulus into the model determined an appropriate gel size to achieve N2O reduction even under aerobic conditions.

Solution-mediated growth of NBA-ZSM-5 crystals retarded by gel entrapment

The synthesis of flat tablet-shaped ZSM-5 crystals from a gel using metakaolin as aluminosilicate source and n-butyl amine as structure directing agent was investigated. The evolution inside the solid phase was characterized by X-ray diffraction, scanning electron microscopy, energy dispersive spectroscopy, thermogravimetry and mass spectrometry. A kinetic study indicated that the nucleation of the majority crystals occurred concurrently with the formation of the gel upon heating the starting liquid suspension. Microstructural evidences undeniably showed that the gel precipitated on ZSM-5 crystals and mineral impurities originating from kaolin. As a result, crystal growth was retarded by gel entrapment, as indicated by the configuration and morphology of the embedded crystals. The results presented herein are harmonized with a solution-mediated nucleation and growth mechanism. Our observations differ from the autocatalytic model that suggests that the nuclei rest inside the gel until released when the gel is consumed. Our results show instead that it is crystals that formed in an early stage before entrapment inside the gel that rest inside the gel until exposed at the gel surface. These results illustrate the limitation of the classical method used in the field to determine nucleation profiles when the crystals become trapped inside the gel.

Immobilization of Anammox biomass in sodium alginate

Anaerobic ammonium oxidation (anammox) is a process of ammonium and nitrite conversion into nitrogen gas. Nowadays, anammox is applied into many wastewater treatment plants worldwide. However, anammox bacteria are characterized by a slow growth rate, which may cause problems in maintaining the biomass in the system. The promising technique which can help to maintain the biomass in the reactor and effectively prevent loss of anammox bacteria from a system is immobilization. Selection and optimization of the appropriate immobilization technique for investigated biomass is crucial for conducting an effective process. One of the ways for bacteria immobilization is gel entrapment. The main goal of the study was to test sodium alginate as an immobilization medium for anammox biomass. In the present study procedure of immobilization in sodium alginate was optimised, then the mechanical and chemical properties of the obtained pellets were investigated. Series of batch experiments revealed that immobilized anammox biomass was able to remove ammonia and nitrite nitrogen effectively. The calculated specific anammox activity (SAA) for immobilized anammox biomass was 0.18 g N·gVSS-1·d-1, while for non-immobilized biomass was 0.36 g N·gVSS-1·d-1.

Potential application of immobilized streptokinase extracted from Streptococcus equinus VIT_VB2

ABSTRACTStreptokinase purified from Streptococcus equinus VIT_VB2 isolated from bovine milk sample was immobilized in various solid supports namely entrapment in agarose gel, calcium alginate beads and gelatin gel by cross-linking with formaldehyde. Immobilization of streptokinase in calcium alginate beads showed maximum efficiency (81.8 ± 1.06%) when compared with entrapment with agarose gel (55.6 ± 2.17%) and cross-linked gelatin formaldehyde gel (71.0 ± 1.54%). The purified SK activity was expressed maximum in calcium alginate (1%) and gelatin gel (0.25%) with 1292.68 ± 1.33 and 1121.9 ± 1.2 U mL−1, respectively. Similarly, SK entrapped in gelatin gel and calcium alginate showed maximum in vitro blood clot lysis activity with 77.67 ± 2.64% and 76.16 ± 2.72%, respectively. The immobilized SK in gelatin gel showed complete clot lysis within 15 min; hence, this application of the study could be used in the treatment of superficial thrombophlebitis, phlebitis, and venous thrombosis. These beads were used for three repeated cycles to check the conversion of substrates into their products, and we concluded that SK can be immobilized in the suitable matrices. Therefore, this helps in the drug-delivery strategies in highly efficient way, moreover, economically competent process in the pharmaceutics.

Degradation of urea by entrapped cross-linked urease aggregates: a combinatorial approach to urease stabilization for environmental and industrial applications

Urease which catalyzes biodegradation of urea to ammonia and carbon dioxide has attracted a great deal of attention as a catalyst in food industry, agriculture and artificial kidneys. Immobilization is a unique approach for enhancement of enzyme stability before enzyme industrialization. In the present work, crude sword bean urease was immobilized in the form of cross-linked enzyme aggregate prior to entrapment in polyacrylamide gel or chitosan beads. No significant change was observed in the kinetic constant value (k m ) of urease upon formation of cross-linked enzyme aggregate. Entrapment of urease cross-linked aggregates in chitosan beads resulted in complete loss of activity while polyacrylamide-embedded cross-linked urease aggregates were enzymatically active. The optimum temperature for urease was 60 °C (soluble and cross-linked aggregate form) and 70 °C (polyacrylamide-embedded cross-linked aggregate). The optimal pH for urease was 7.0 (soluble and cross-linked aggregate form) and 5.0 (polyacrylamide-embedded cross-linked aggregate). In the other words, immobilization in the form of cross-linked aggregate and entrapment in polyacrylamide matrix could be resulted in the stabilization of urease over a wider pH range. Urease cross-linked aggregate showed improved stability at 60 °C, and its half-life increased about two times. Operational stability of cross-linked urease aggregate was also noticeably improved upon entrapment in polyacrylamide gel. In conclusion, entrapment of carrier-free cross-linked urease aggregate in a polyacrylamide matrix could result in a great improvement in both the storage and operational stabilities of urease.

First full-scale nitritation-anammox plant using gel entrapment technology for ammonia plant effluent

The first full-scale nitritation-anammox process using gel entrapment technology was evaluated for nitrogen removal. Ammonia plant effluents contained not only ammonium but also methanol, an anammox inhibitor. The nitrogen concentration and loading design were 690mg/L and 400kg-N/d, respectively. A nitritation reactor (170m3) filled with nitrification gel carriers was started up on day 20, with a resulting nitrification efficiency of 58%. A stable nitritation performance was observed without significant nitrate production for >1year. A nitrogen conversion rate of 3.2kg-N/m3/d was observed on the anammox reactor (100m3) filled with anammox gel carrier on day 69. The full-scale anammox reactor could be started up in approximately 2 months. Subsequently, stable nitrogen removal performance was observed for >1year. The average nitrogen loading, nitrogen conversion rate, and total nitrogen removal on the anammox reactor were 3.6kg-N/m3/d, 3.0kg-N/m3/d and 330kg-N/d, respectively. The nitrogen removal performance obtained in this study will facilitate further application of the anammox processes to many types of industrial wastewater treatment.

Reverse Phase Compatible TLC-Bioautography for Detection of Tyrosinase Inhibitors.

Reverse phase chromatography and bioautographic assays are key tools for natural product bioguided isolation; however, their direct coupling has not been fully achieved. To develop a bioautographic assay to detect tyrosinase inhibitors present in complex matrices sorbed on reverse phase (RP) TLC-plates that can be used for bioguided isolation of bioactive compounds. Enzyme gel entrapment with an amphiphilic copolymer was used for assay development. The gel turns into a brown "skin like" colour due to tyrosinase catalysed oxidation of l-tyrosine. The inhibitors are visualised as clear spots against a brown coloured background. The assay was able to localise cinnamaldehyde in Cinnamomum cassia essential oil, as its main constituent with known tyrosinase inhibition properties. The assay allowed the detection of 0.03% (w/w) of kojic acid co-spotted with a methanolic extract of Sphaeralcea bonariensis and chromatographed on RP-TLC. The developed assay is able to detect, with high sensitivity, tyrosinase inhibitors present in complex matrices that were chromatographed in RP-TLC. Results can be easily read by colour change, inhibitors appear as clear spots in a darker background. Copyright © 2016 John Wiley & Sons, Ltd.

Fiber Optic SPR-Based Uric Acid Biosensor Using Uricase Entrapped Polyacrylamide Gel

In this letter, we report a fiber optic biosensor based on surface plasmon resonance (SPR) for the detection of uric acid in aqueous samples. The sensing probe is prepared by successive coatings of silver and silicon layers over a small unclad length of the plastic clad silica optical fiber in the middle. Thereafter, the probe is dipped in a polyacrylamide gel containing enzyme uricase to immobilize it using a gel entrapment method. The sensor is characterized using a wavelength interrogation method. The SPR spectra corresponding to different concentrations of uric acid in the range of 0-0.9 mM are recorded. A red shift in the resonance wavelength is observed with an increase in the concentration of uric acid around the probe. In addition, the sensitivity of the proposed biosensor is studied in relation to parameters, such as concentration and pH of the uric acid solution. Finally, the sensor is highly selective and possesses better limit of detection and limit of quantification.

Enhancing Transdermal Delivery of Glimepiride Via Entrapment in Proniosomal Gel

Objective: The aim of this study is to formulate and evaluate proniosomal gel formulations as transdermal delivery systems of glimepiride (GM) to improve its therapeutic efficacy. Methods: Proniosomal formulations have been prepared using different types of non-ionic surfactants with cholesterol in different molar ratios. Proniosomal gel; PN 16 (Span 60, Tween 60, Cholesterol; 35:35:30 molar ratio) that exhibits maximum EE % (94.01 ± 0.88) and most prolonged release was chosen for ex-vivo skin permeation and in-vivo hypoglycemic activity studies. Results: Proniosomal gel produced better permeation through rabbit skin than HPMC niosomal gel and HPMC gel. The pharmacokinetic parameters; time of maximum response (T max ), % reduction in blood glucose concentration and area above the blood glucose levels-time curve (AAC) of proniosomal gel were studied. Proniosomal gel showed a controlled release behavior and a significantly higher hypoglycemic activity (65.34 ± 6.54%). Conclusion: It is evident from this study that proniosomal gel can act as an alternative approach for enhancing transdermal delivery of GM. Key words: Glimepiride, Hypoglycemic activity, Nonionic surfactants, Proniosomal gel, Area above the blood glucose levels-time curve.

Sol–gel entrapment of bromothymol blue (BTB) indicator in the presence of cationic 16E1Q and 16E1QS surfactants

Sol–gel entrapment method was used to entrap bromothymol blue (BTB) pH indicator into mesoporous silica materials in the presence of cationic surfactants: 2-(hexadecyloxy)-2-oxoethyl-N,N,N-triethyl ammonium iodide (16E1Q) and 2-(hexadecyloxy)-2-oxoethanaminium-p-toluene sulfonate (16E1QS). The use of this new family of surfactants has modified the morphology and porosity of silica host matrix for better sensing capability and for faster response toward pH change. The physical interactions between BTB molecules and host mesoporous silica network were based on the nature of the surfactant. The presence of surfactants has shifted pKa values of more basic in comparison with that of BTB-entrapped silica system.

Biosorption of hexavalent chromium from paint industrial effluent by Saraca indica leaves using with and without gel entrapment method

Raw agricultural wastes are affordable adsorbents for the removal of heavy metals from aqueous solutions. In this research, the use of Saraca indica leaves (as an eco-friendly and low cost adsorbent) having the ability to remove toxic hexavalent chromium (Cr(VI)) from paint industrial effluent has been investigated. Parameters such as biosorbent quantity, initial metal-ion concentration, and contact time in with and without immobilised batch experiments have been studied. The results revealed that Saraca indica leaves absorb over 86% of Cr (VI) ions from solutions with increasing adsorbent dosage. Adsorption isotherms were compared with Langmuir and Freundlich adsorption models and experimental data were found to fit the models. Also adsorption kinetic models were studied and found to best fit the pseudo second order model. The present study showed that such a low cost material could be used efficiently for the removal of Cr(VI) from paint industrial effluent.

MUCOADHESIVE GEL WITH IMMOBILIZED LYSOZYME AND QUERCETIN

Aim. To develop mucoadhesive gel with jointly immobilized lysozyme and quercetin and to investigate biochemical, physical, physico-chemical and biological properties of the complex preparation. Methods. The bacteriolytic activity of lysozyme was determined turbidimetrically, using Micrococcus lysodeikticus 2665 as substrate. Protein content was determined by Lowry-Hartree and that of quercetin – with usage of zirconium (IV) chloride, spectrophotometrically. Immobilization was conducted by gel entrapment method. Therapeutic and preventive action of complex gel was investigated on rats, with gingivitis, which was modelled using indomethacin. Results. Using the carboxymethyl cellulose sodium salt as support, the joint immobilization of lysozyme and quercetin was conducted by gel entrapment method. It was shown, that at mass ratio lysozyme: quercetin: matrix (1: 0.4: 6), the complete degree of lysozyme and quercetin inclusion is observed, with preservation of bacteriolytic enzyme activity. The obtained gel possesses high mucoadhesive properties (adhesive force equals 6000 Pa), prolonged action: the time of complete release of enzyme and quercetin was 180 and 75 min, respectively. The immobilized lysozyme is active in the wide range of pH-values, stable in acidic medium and during storage. The usage of oral gel with lysozyme and quercetin leads the indices of inflammation to the norm. Conclusions. The joint immobilization of lysozyme and quercetin in sodium carboxymethyl cellulose does not influence the biochemical properties of enzyme. The obtaining mucoadhesive gel with immobilized lysozyme and quercetin is the preparation with complex biological activity, antimicrobial and anti-inflammatory effect.

Recyclable solid-phase biocatalyst with improved stability by sol–gel entrapment of β-d-galactosidase

β-d-Galactosidase from Kluyveromyces lactis was for the first time immobilized by entrapment in hybrid organic-inorganic sol–gel materials with microporous structure, obtained from alkoxy silanes and alkyl substituted alkoxy silanes, in different combinations. The immobilization matrix was tailored by fine tuning of several parameters, such as: nature of alkyl group of silane precursors, molar ratio of silane precursors, nature of additives, protein concentration. Unlike other enzymes, β-d-galactosidase showed the best catalytic activity at low alkyl group content in the sol–gel matrix, at a molar ratio of 7:1 between the tetraalkoxysilane alkyl-trialkoxysilane precursor. The immobilized enzyme demonstrated enhanced storage, pH and thermal stability compared to the soluble enzyme. The composite sol–gel materials were characterized by transmission electron microscopy, scanning electron microscopy, fluorescence confocal microscopy, and porosity measurement. The biocatalyst was successfully reused in five reaction cycles, maintaining more than 60% of the initial activity.