RESUMO
In this study, a series of hydrogels were synthesized from chitosan(s) that was crosslinking with glutaraldehyde at different concentrations. Ascorbic acid in an acidic medium was used to facilitate non-covalent interactions. The chitosan(s) was obtained from shrimp cytoskeleton; while ascorbic acid was extracted from xoconostle juice. The hydrogel reaction was monitored by UV-vis spectroscopy (550 nm) to determine the reaction kinetics and reaction order at 60 °C. The hydrogels structures were characterized by NMR, FT-IR, HR-MS and SEM, while the degree of cross-linking was examined with TGA-DA. The extracellular matrices were obtained as stable hydrogels where reached maximum crosslinking was of 7 %, independent of glutaraldehyde quantity added. The rheological properties showed a behavior of weak gels and a dependence of crosslinking agent concentration on strength at different temperatures. The cytotoxicity assay showed that the gels had no adverse effects on cellular growth for all concentrations of glutaraldehyde.
Assuntos
Materiais Biocompatíveis , Quitosana , Hidrogéis , Engenharia Tecidual , Hidrogéis/química , Hidrogéis/síntese química , Materiais Biocompatíveis/química , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/farmacologia , Quitosana/química , Quitosana/farmacologia , Quitosana/síntese química , Animais , Glutaral/química , Reologia , Reagentes de Ligações Cruzadas/químicaRESUMO
This work aims to improve the post stabilty of reusable potassium iodide hydrogel dosimter. A reusable and low-cost radiochromic dosimeter containing a gel matrix of polyvinyl alcohol, potassium iodide dye, froctose as reducing agent and glutaraldehyde as cross-linking agent was developed for dose calibration in radiotherapy. The gel samples were exposed to different absorbed doses using a medical linear acceleration. UV-vis Spectrophotometry was utilized to investigate the changes in optical-properties of irradiated gels with regard to peak wavelength of 353 nm. The stability of the gel (one of the most limitation of using this dosimeter) was improved significantly by the addition of certain concentrations of dimethyl sulfoxide. The two-dimensional optical imaging system of charge-coupled-device (CCD) camera with a uniform RGB light-emitting-diode (LED) array source was used for diffusion coefficient purpose using two dimensional gel template. The value of diffusion coefficient reported is significant and highly reduced compared with other dosimeters reported in the literatures. Moreover, heating the improved gels to certain temperatures results in resetting their optical properties, which makes it possible to reuse for multiple times.
Assuntos
Estudos de Viabilidade , Álcool de Polivinil , Iodeto de Potássio , Dosímetros de Radiação , Álcool de Polivinil/química , Iodeto de Potássio/química , Calibragem , Géis/química , Humanos , Hidrogéis/química , Radiometria/métodos , Radiometria/instrumentação , Dimetil Sulfóxido/química , Glutaral/química , Difusão , TemperaturaRESUMO
Bio-electrochemical Systems (BES), particularly Microbial Fuel Cells (MFC), have emerged as promising technologies in environmental biotechnology. This study focused on optimizing the anode bacterial culture immobilization process to enhance BES performance. The investigation combines and modifies two key immobilization methods: covalent bonding with glutaraldehyde and inclusion in a chitosan gel in order to meet the criteria and requirements of the bio-anodes in MFC. The performance of MFCs with immobilized and suspended cultures was compared in parallel experiments. Both types showed similar substrate utilization dynamics with slight advantage of the immobilized bio-anode considering the lower concentration of biomass. The immobilized MFC exhibited higher power generation and metabolic activity, as well. Probably, this is due to improved anodic respiration and higher coulombic efficiency of the reactor. Analysis of organic acids content supported this conclusion showing significant inhibition of the fermentation products production in the MFC reactor with immobilized anode culture.
Assuntos
Fontes de Energia Bioelétrica , Células Imobilizadas , Quitosana , Eletrodos , Fontes de Energia Bioelétrica/microbiologia , Células Imobilizadas/metabolismo , Quitosana/metabolismo , Quitosana/química , Fermentação , Reatores Biológicos/microbiologia , Biomassa , Glutaral/química , EletricidadeRESUMO
Heterogeneous biocatalysts were prepared by adsorbing T. lanuginosus lipase (TLL) onto uncalcined (SBAUC-TLL) and calcined (SBAC-TLL) SBA-15, using ammonium fluoride as a pore expander to facilitate TLL immobilization. At an enzyme load of 1 mg/g, high immobilization yields (>90 %) and recovered activities (>80 % for SBAUC-TLL and 70 % for SBAC-TLL) were achieved. When increasing the enzyme load to 5 mg/g, the immobilization yield of SBAUC-TLL was 80 %, and the recovered activity was 50 %, while SBAC-TLL had a yield of 100 % and a recovered activity of 36 %. Crosslinking with glutaraldehyde (GA) was conducted to improve stability (SBAUC-TLL-GA and SBAC-TLL-GA). Although SBAC-TLL-GA lost 25 % of initial activity after GA modifications, it exhibited the highest thermal (t1/2 = 5.7 h at 65 °C), when compared to SBAC-TLL (t1/2 = 12 min) and the soluble enzyme (t1/2 = 36 min), and operational stability (retained 100 % activity after 5 cycles). Both biocatalysts presented high storage stability since they retained 100 % of initial activity for 30 days. These results highlight SBA-15's potential as an enzyme support and the protocol's efficacy in enhancing stability, with implications for industrial applications in the food, chemical, and pharmaceutical sectors.
Assuntos
Biocatálise , Estabilidade Enzimática , Enzimas Imobilizadas , Lipase , Dióxido de Silício , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Lipase/química , Lipase/metabolismo , Dióxido de Silício/química , Porosidade , Temperatura , Adsorção , Concentração de Íons de Hidrogênio , Eurotiales/enzimologia , Cinética , Glutaral/químicaRESUMO
We present the synthesis of a cross-linking enzyme aggregate (CLEAS) of a peroxidase from Megathyrsus maximus (Guinea Grass) (GGP). The biocatalyst was produced using 50%v/v ethanol and 0.88%w/v glutaraldehyde for 1 h under stirring. The immobilization yield was 93.74% and the specific activity was 36.75 U mg-1. The biocatalyst surpassed by 61% the free enzyme activity at the optimal pH value (pH 6 for both preparations), becoming this increase in activity almost 10-fold at pH 9. GGP-CLEAS exhibited a higher thermal stability (2-4 folds) and was more stable towards hydrogen peroxide than the free enzyme (2-3 folds). GGP-CLEAS removes over 80% of 0.05 mM indigo carmine at pH 5, in the presence of 0.55 mM H2O2 after 60 min of reaction, a much higher value than when using the free enzyme. The operational stability showed a decrease of enzyme activity (over 60% in 4 cycles), very likely related to suicide inhibition.
Assuntos
Enzimas Imobilizadas , Peróxido de Hidrogênio , Índigo Carmim , Peroxidase , Índigo Carmim/química , Peroxidase/metabolismo , Peroxidase/química , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Concentração de Íons de Hidrogênio , Peróxido de Hidrogênio/química , Estabilidade Enzimática , Reagentes de Ligações Cruzadas/química , Temperatura , Glutaral/químicaRESUMO
BACKGROUND: Vinyl polyether silicone (VPES) is a novel impression biomaterial made of a combination of vinyl polysiloxane (VPS) and polyether (PE). Thus, it is significant to assess its properties and behaviour under varied disinfectant test conditions. This study aimed to assess the dimensional stability of novel VPES impression material after immersion in standard disinfectants for different time intervals. METHODS: Elastomeric impression material used -medium body regular set (Monophase) [Exa'lence GC America]. A total of 84 Specimens were fabricated using stainless steel die and ring (ADA specification 19). These samples were distributed into a control group (n=12) and a test group (n=72). The test group was divided into 3 groups, based on the type of disinfectant used - Group-A- 2% Glutaraldehyde, Group-B- 0. 5% Sodium hypochlorite and Group-C- 2% Chlorhexidine each test group was further divided into 2 subgroups (n=12/subgroup) based on time intervals for which each sample was immersed in the disinfectants - subgroup-1- 10 mins and Subgroup 2- 30 mins. After the impression material was set, it was removed from the ring and then it was washed in water for 15 seconds. Control group measurements were made immediately on a stereomicroscope and other samples were immersed in the three disinfection solutions for 10 mins and 30 mins to check the dimensional stability by measuring the distance between the lines generated by the stainless steel die on the samples using a stereomicroscope at x40 magnification. RESULTS: The distance measured in the control group was 4397.2078 µm and 4396.1571 µm; for the test group Group-A- 2% Glutaraldehyde was 4396.4075 µm and 4394.5992 µm; Group-B- 0. 5% Sodium hypochlorite was 4394.5453 µm and 4389.4711 µm Group-C- 2% Chlorhexidine was 4395.2953 µm and 4387.1703 µm respectively for 10 mins and 30 mins. Percentage dimensional change was in the range of 0.02 - 0.25 for all the groups for 10 mins and 30 mins. CONCLUSIONS: 2 % Glutaraldehyde is the most suitable disinfectant for VPES elastomeric impression material in terms of dimensional stability and shows minimum dimensional changes as compared to that of 2% Chlorhexidine and 0.5% Sodium hypochlorite.
Assuntos
Materiais para Moldagem Odontológica , Glutaral , Teste de Materiais , Polivinil , Siloxanas , Materiais para Moldagem Odontológica/química , Polivinil/química , Siloxanas/química , Fatores de Tempo , Glutaral/química , Desinfetantes de Equipamento Odontológico/química , Hipoclorito de Sódio/química , Desinfetantes/química , Clorexidina/química , Propriedades de Superfície , HumanosRESUMO
In this article, the synthesis of antioxidant peptides in the enzymatic hydrolysis of caprine casein was analyzed at three different time points (60 min, 90 min, and 120 min) using immobilized pepsin on activated and modified carbon (AC, ACF, ACG 50, ACG 100). The immobilization assays revealed a reduction in the biocatalysts' activity compared to the free enzyme. Among the modified ones, ACG 50 exhibited greater activity and better efficiency for reuse cycles, with superior values after 60 min and 90 min. Peptide synthesis was observed under all studied conditions. Analyses (DPPH, ß-carotene/linoleic acid, FRAP) confirmed the antioxidant potential of the peptides generated by the immobilized enzyme. However, the immobilized enzyme in ACG 50 and ACG 100, combined with longer hydrolysis times, allowed the formation of peptides with an antioxidant capacity greater than or equivalent to those generated by the free enzyme, despite reduced enzymatic activity.
Assuntos
Antioxidantes , Caseínas , Enzimas Imobilizadas , Glutaral , Cabras , Iridoides , Pepsina A , Peptídeos , Antioxidantes/química , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Caseínas/química , Animais , Pepsina A/metabolismo , Pepsina A/química , Glutaral/química , Peptídeos/química , Iridoides/química , Hidrólise , Carvão Vegetal/químicaRESUMO
Aspergillus oryzae ß-D-galactosidase (ß-Gal) efficiently hydrolyzes sesaminol triglucoside into sesaminol, which has higher biological activity. However, ß-Gal is difficult to be separate from the reaction mixture and limited by stability. To resolve these problems, ß-Gal was immobilized on amino-functionalized magnetic nanoparticles mesoporous silica pre-activated with glutaraldehyde (Fe3O4@mSiO2-ß-Gal), which was used for the first time to prepare sesaminol. Under the optimal conditions, the immobilization yield and recovered activity of ß-Gal were 57.9 ± 0.3 % and 46.5 ± 0.9 %, and the enzymatic loading was 843 ± 21 Uenzyme/gsupport. The construction of Fe3O4@mSiO2-ß-Gal was confirmed by various characterization methods, and the results indicated it was suitable for heterogeneous enzyme-catalyzed reactions. Fe3O4@mSiO2-ß-Gal was readily separable under magnetic action and displayed improved activity in extreme pH and temperature conditions. After 45 days of storage at 4 °C, the activity of Fe3O4@mSiO2-ß-Gal remained at 92.3 ± 2.8 %, which was 1.29 times than that of free enzyme, and its activity remained above 85 % after 10 cycles. Fe3O4@mSiO2-ß-Gal displayed higher affinity and catalytic efficiency. The half-life was 1.41 longer than free enzymes at 55.0 °C. Fe3O4@mSiO2-ß-Gal was employed as a catalyst to prepare sesaminol, achieving a 96.7 % conversion yield of sesaminol. The excellent stability and catalytic efficiency provide broad benefits and potential for biocatalytic industry applications.
Assuntos
Aspergillus oryzae , Enzimas Imobilizadas , Glutaral , Dióxido de Silício , beta-Galactosidase , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , beta-Galactosidase/química , beta-Galactosidase/metabolismo , Aspergillus oryzae/enzimologia , Dióxido de Silício/química , Glutaral/química , Dioxóis/química , Dioxóis/farmacologia , Nanopartículas de Magnetita/química , Porosidade , Temperatura , Concentração de Íons de Hidrogênio , Estabilidade Enzimática , FuranosRESUMO
Whey protein isolate (WPI) was incorporated within calcium pectinate (CPT) beads in order to boost their anionic qualities and meliorate their glutaraldehyde (GA)-polyethyleneimine (PEI) grafting process. The Box-Behnken Design (BBD) verified that WPI inclusion significantly raised the GA-PEI-CPT-WPI beads immobilized ß-D-galactosidase (iß-GLD) activity. The BBD also revealed the optimal settings for WPI concentration, PEI pH, PEI concentration, and GA concentration, which were 2.91â¯%, 10.8, 3.5â¯%, and 2.24â¯%, respectively. The GA-PEI-CPT-WPI beads grafting process was scrutinized via FTIR, EDX, and SEM. The optimal GA-PEI-CPT-WPI immobilizers provided fine ß-GLD immobilization efficiencies, which reached up to 65.28â¯%. The free and GA-PEI-CPT-WPI iß-GLDs pH and temperature profiles were scrutinized. It was also unveiled that the thermal stability of the iß-GLD surpassed that of its free compeer as it provided lesser kd and ΔS values and larger t1/2, D-values, Ed, ΔH, and ΔG values. Furthermore, the iß-GLD provided 92.00±3.39â¯% activity after 42 storage days, which denoted its fine storage stability. The iß-GLD short duration (15â¯min) operational stability was also inspected, and 82.70±0.78â¯% activity was provided during the fifteenth degradation run. Moreover, the iß-GLD long duration (24â¯h) operational stability was inspected while degrading the lactose of buffered lactose solution (BLS) and cheese whey (CW). It was unveiled that 81.86±0.96â¯% and 73.58±2.24â¯% of the initial glucose were detected during the sixth degradation runs, respectively.
Assuntos
Enzimas Imobilizadas , Polietilenoimina , Termodinâmica , Proteínas do Soro do Leite , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Proteínas do Soro do Leite/química , Cinética , Polietilenoimina/química , Concentração de Íons de Hidrogênio , Pectinas/química , Pectinas/metabolismo , beta-Galactosidase/metabolismo , beta-Galactosidase/química , Glutaral/química , Temperatura , Estabilidade EnzimáticaRESUMO
Crosslinking is usually required to improve the mechanical properties and stability of collagen-based scaffolds. Introducing exogenous crosslinks into collagen may however affect the collagen structure. Since the architecture of collagen is tied to its functionality, it is important to study the effect of crosslinking and to select a crosslinking method that preserves both the collagen structure and mechanical properties. The objective of this study is to compare the effect of various crosslinking methods on the structure and mechanical properties of bioartificial tendon-like materials (collagen multifilament bundles) fabricated by contact drawing. We examine both physical (ultraviolet light, UVC) and chemical (genipin, carbodiimide (EDC), and glutaraldehyde) crosslinking methods. The presence of collagen and the formation of well-ordered collagen structures are confirmed by attenuated total reflectance Fourier-transform infrared spectromicroscopy and wide-angle X-ray scattering for all crosslinking methods. The morphology of the collagen multifilament bundles is similar across crosslinking methods. Swelling of the multifilament bundles is dramatically reduced following crosslinking and varies by crosslinking method, with genipin- and carbodiimide-crosslinked specimens swelling the least. Ultimate tensile strength (UTS) and Young's modulus significantly improve for all crosslinked specimens compared to non-crosslinked specimens. Glutaraldehyde crosslinked collagen multifilament bundles display the highest UTS values ranging from 33.82±0.0â MPa to 45.59±0.76â MPa.
Assuntos
Colágeno , Reagentes de Ligações Cruzadas , Reagentes de Ligações Cruzadas/química , Colágeno/química , Glutaral/química , Raios Ultravioleta , Carbodi-Imidas/química , Iridoides/química , Animais , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios XRESUMO
OBJECTIVE: Aortic valve neocuspidization (AVNeo) using autologous pericardium is a promising technique. Expected advantages are reduced immune response, appropriate biomechanics and lower treatment expenses. Nevertheless, autologous pericardium can be affected by patient's age and comorbidities. Usually, glutaraldehyde (GA) - fixed bovine pericardium is the basic material for aortic valve prostheses, easy available and carefully pre-examined in a standardized fabrication process. Aim of the study is the verification of autologous pericardial tissue homogeneity by analysing tissue thickness, biomechanics and extracellular matrix (ECM) composition. METHODS: Segments of human GA-fixed pericardium selected by the surgeon based on visual criteria for cusp pre-cut and remaining after surgical AV replacement were investigated in comparison to bovine standard tissue treated equivalently. Pericardium sampling was performed at up to three positions of each sutured cusp for histological or biomechanical analysis, according to tissue availability. RESULTS AND CONCLUSIONS: Human pericardia exhibited a higher heterogeneity in collagen content, density of vessel structures and elastic moduli. Thickness, vessel density and collagen and elastin content differed significantly between the species. In contrast, significant interindividual differences were detected in most properties investigated for human pericardial samples but only for tissue thickness in bovine tissues. Higher heterogeneity of human pericardium, differing vessel and collagen content compared to bovine state-of-the-art material might be detrimental for long term AV functionality or deterioration and have to be intensely investigated in patients follow up after autologous cusp replacement.
Assuntos
Valva Aórtica , Bioprótese , Próteses Valvulares Cardíacas , Pericárdio , Bovinos , Humanos , Valva Aórtica/cirurgia , Animais , Fenômenos Biomecânicos , Masculino , Feminino , Idoso , Matriz Extracelular/química , Pessoa de Meia-Idade , Colágeno/química , Glutaral/química , Teste de Materiais , Implante de Prótese de Valva Cardíaca/métodosRESUMO
The freeze-drying is a technology that preserves biological samples in a dry state, which is beneficial for storage, transportation, and cost saving. In this study, the bovine pericardium was treated with a freeze-drying protectant composed of polyethylene glycol (PEG) and trehalose (Tre), and then freeze-dried. The results demonstrated that the mechanical properties of the pericardium treated with PEG + 10% w/v Tre were superior to those of the pericardium fixed with glutaraldehyde (GA). The wet state water content of the rehydrated pericardium, determined using the Karl Fischer method, was (74.81 ± 1.44)%, which was comparable to that of the GA-fixed pericardium. The dry state water content was significantly reduced to (8.64 ± 1.52)%, indicating effective dehydration during the freeze-drying process. Differential scanning calorimetry (DSC) testing revealed that the thermal shrinkage temperature of the pericardium was (84.96 ± 0.49) â, higher than that of the GA-fixed pericardium (83.14 ± 0.11) â, indicating greater thermal stability. Fourier transform infrared spectroscopy (FTIR) results showed no damage to the protein structure during freeze-drying. Hematoxylin and eosin (HE) staining demonstrated that the freeze-drying process reduced pore formation, prevented ice crystal growth, and resulted in a tighter arrangement of tissue fibers. The frozen-dried bovine pericardium was subjected to tests for cell viability and hemolysis rate. The results revealed a cell proliferation rate of (77.87 ± 0.49)%, corresponding to a toxicity grade of 1. Additionally, the hemolysis rate was (0.17 ± 0.02)%, which is below the standard of 5%. These findings indicated that the frozen-dried bovine pericardium exhibited satisfactory performance in terms of cytotoxicity and hemolysis, thus meeting the relevant standards. In summary, the performance of the bovine pericardium treated with PEG + 10% w/v Tre and subjected to freeze-drying could meet the required standards.
Assuntos
Liofilização , Pericárdio , Polietilenoglicóis , Trealose , Animais , Pericárdio/química , Trealose/química , Trealose/farmacologia , Bovinos , Polietilenoglicóis/química , Glutaral/química , Varredura Diferencial de CalorimetriaRESUMO
A robust biocompatible solid-phase microextraction (SPME) fiber, so-called Ti/APTS/GA/CS, was prepared by chemical bonding of cross-linked glutaraldehyde-chitosan to the surface of a titanium wire using APTS. The fiber was applied for sampling of phytohormones in plant tissues, followed by HPLC-UV analysis. The structure and morphology of the fiber coating was investigated by FT-IR, SEM, EDX, XRD, and TGA techniques. A Box-Behnken design was implemented to optimize the experimental variables. The calibration graphs were linear over a wide linear range (0.5-200 µg L-1) with LODs over the range of 0.01-0.06 µg L-1. The intra-day and inter-day precisions were found to be 1.3-6.3% and 4.3-7.3%, respectively. The matrix effect values ranged from 86.5% to 111.7%, indicating that the complex sample matrices had an insignificant effect on the determination of phytohormones. The fiber was successfully employed for the direct-immersion SPME (DI-SPME-HPLC) analysis of the phytohormones in cucumber, tomato, date palm, and calendula samples.
Assuntos
Quitosana , Glutaral , Reguladores de Crescimento de Plantas , Microextração em Fase Sólida , Titânio , Quitosana/química , Titânio/química , Glutaral/química , Reguladores de Crescimento de Plantas/química , Reguladores de Crescimento de Plantas/análise , Materiais Biocompatíveis/química , Reagentes de Ligações Cruzadas/químicaRESUMO
Mucosal tissues represent a major interface between the body and the external environment and are covered by a highly hydrated mucins gel called mucus. Mucus lubricates, protects and modulates the moisture levels of the tissue and is capitalized in transmucosal drug delivery. Pharmaceutical researchers often use freshly excised animal mucosal membranes to assess mucoadhesion and muco-penetration of pharmaceutical formulations which may struggle with limited accessibility, reproducibility, and ethical questions. Aiming to develop a platform for the rationale study of the interaction of drugs and delivery systems with mucosal tissues, in this work mucus-mimicking mucin-based hydrogels are synthesized by the tandem chemical and physical crosslinking of mucin aqueous solutions. Chemical crosslinking is achieved with glutaraldehyde (0.3% and 0.75% w/v), while physical crosslinking by one or two freeze-thawing cycles. Hydrogels after one freeze-thawing cycle show water content of 97.6-98.1%, density of 0.0529-0.0648 g cmâ»3, and storage and loss moduli of ≈40-60 and ≈3-5 Pa, respectively, that resemble the properties of native gastrointestinal mucus. The mechanical stability of the hydrogels increases over the number of freeze-thawing cycles. Overall results highlight the potential of this simple, reproducible, and scalable method to produce artificial mucus-mimicking hydrogels for different applications in pharmaceutical research.
Assuntos
Reagentes de Ligações Cruzadas , Hidrogéis , Mucinas , Muco , Hidrogéis/química , Hidrogéis/síntese química , Mucinas/química , Muco/química , Reagentes de Ligações Cruzadas/química , Animais , Glutaral/química , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologiaRESUMO
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.
Assuntos
Reatores Biológicos , Estabilidade Enzimática , Enzimas Imobilizadas , Lipase , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Lipase/metabolismo , Lipase/química , Membranas Artificiais , Grafite/química , Saccharomycetales/enzimologia , Glutaral/química , Espectroscopia de Infravermelho com Transformada de Fourier , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Temperatura , Difração de Raios XRESUMO
Immobilization of enzymes on aminated supports using the glutaraldehyde chemistry may involve three different interactions, cationic, hydrophobic, and covalent interactions. To try to understand the impact this heterofunctionality, we study the physical adsorption of the beta-galactosidase from Aspergillus niger, on aminated supports (MANAE) and aminated supports with one (MANAE-GLU) or two molecules of glutaraldehyde (MANAE-GLU-GLU). To eliminate the chemical reactivity of the glutaraldehyde, the supports were reduced using sodium borohydride. After enzyme adsorption, the release of the enzyme from the supports using different NaCl concentrations, Triton X100, ionic detergents (SDS and CTAB), or different temperatures (4 °C to 55 °C) was studied. Using MANAE support, at 0.3 M NaCl almost all the immobilized enzyme was released. Using MANAE-GLU, 0.3 M, and 0.6 M NaCl similar results were obtained. However, incubation at 1 M or 2 M NaCl, many enzyme molecules were not released from the support. For the MANAE-GLU-GLU support, none of the tested concentrations of NaCl was sufficient to release all enzyme bound to the support. Only using high temperatures, 0.6 M NaCl, and 1 % CTAB or SDS, could the totality of the proteins be released from the support. The results shown in this paper confirm the heterofunctional character of aminated supports modified with glutaraldehyde.
Assuntos
Enzimas Imobilizadas , Cloreto de Sódio , Glutaral/química , Estabilidade Enzimática , Adsorção , Cetrimônio , Enzimas Imobilizadas/químicaRESUMO
Glutaraldehyde (GA) is a protein crosslinker widely used in biochemical and pharmaceutical research because it can rapidly stabilize and immobilize substrates via amine group interactions. However, controlling GA crosslinking is challenging owing to its swift reactivity and the influence of various solution conditions, such as pH and concentrations of the substrate and crosslinker. Although extensive research has focused on GA cross-linking mechanisms, studies on quenching, which is critical for preventing non-specific aggregation during prolonged storage, remain sparse. This study examines the quenching efficiency of a combined amino acid mixture of glycine, histidine, and lysine, which are commonly used as individual quenchers. Our findings, confirmed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis, demonstrate that this amino acid blend offers superior quenching compared to single amino acids, enhancing quenching activity across a wide pH spectrum. These results provide a novel approach for mitigating the high reactivity of GA with implications for improving sample preservation and stabilization in a range of biochemical applications, including microscopy and cell fixation.
Assuntos
Histidina , Lisina , Glutaral/química , Glutaral/farmacologia , Reagentes de Ligações Cruzadas/química , GlicinaRESUMO
With the emergence of penicillin resistance, the development of novel antibiotics has become an urgent necessity. Semi-synthetic penicillin has emerged as a promising alternative to traditional penicillin. The demand for the crucial intermediate, 6-aminopicillanic acid (6-APA), is on the rise. Enzyme catalysis is the primary method employed for its production. However, due to certain limitations, the strategy of enzyme immobilization has also gained prominence. The magnetic Ni0.4Cu0.5Zn0.1Fe2O4 nanoparticles were successfully prepared by a rapid-combustion method. Sodium silicate was used to modify the surface of the Ni0.4Cu0.5Zn0.1Fe2O4 nanoparticles to obtain silica-coated nanoparticles (Ni0.4Cu0.5Zn0.1Fe2O4-SiO2). Subsequently, in order to better crosslink PGA, the nanoparticles were modified again with glutaraldehyde to obtain glutaraldehyde crosslinked Ni0.4Cu0.5Zn0.1Fe2O4-SiO2-GA nanoparticles which could immobilize the PGA. The structure of the PGA protein was analyzed by the PyMol program and the immobilization strategy was determined. The conditions of PGA immobilization were investigated, including immobilization time and PGA concentration. Finally, the enzymological properties of the immobilized and free PGA were compared. The optimum catalytic pH of immobilized and free PGA was 8.0, and the optimum catalytic temperature of immobilized PGA was 50°C, 5°C higher than that of free PGA. Immobilized PGA in a certain pH and temperature range showed better catalytic stability. Vmax and Km of immobilized PGA were 0.3727 µmol·min-1 and 0.0436 mol·L-1, and the corresponding free PGA were 0.7325 µmol·min-1 and 0.0227 mol·L-1. After five cycles, the immobilized enzyme activity was still higher than 25%.
Assuntos
Nanopartículas , Penicilina Amidase , Penicilina Amidase/química , Penicilina Amidase/metabolismo , Glutaral/química , Dióxido de Silício/química , Enzimas Imobilizadas/química , Catálise , Nanopartículas/química , Penicilinas , Fenômenos Magnéticos , Concentração de Íons de Hidrogênio , Temperatura , Estabilidade EnzimáticaRESUMO
A chitosan-alginate sponge (CAS) with multiple cross-linking networks was developed using chitosan, sodium alginate, polyvinyl alcohol, and glutaraldehyde to adsorb and enrich the anionic dyes form the food samples. The multiple networks in CAS refer to the electrostatic cross-linking network, hydrogen bonding cross-linking network, and covalent cross-linking network. Compared with pure chitosan and alginate sponges, the CAS showed better three-dimensional network structure, mechanical behavior, and stability, which is benefit by multiple cross-linking networks. The physical and chemical properties of CAS were systematically studied by a series of characterizations. The adsorption performance of CAS on anionic dyes was inspected with different dye concentration, time, temperature, and pH conditions. CAS exhibited a good and stable adsorption property to amaranth, carmine, and sunset yellow with the saturation adsorption capacity of 94.34, 111.5, and 80.05 mgâg-1, respectively. Furthermore, CAS performed outstanding selectivity to anionic dyes with the selectivity factor up to 16.99. Through electrostatic potential analysis, it is inferred that CAS mainly adsorbs anionic dyes through electrostatic interactions. CAS showed satisfactory reusability, maintaining 97 %-99 % of adsorption performance after six cycles of recycling. Finally, CAS was combined with high-performance liquid chromatography for the enrichment and detection of anionic dyes in candy and cocktail samples, achieving the enrichment factor up to 84.77.
Assuntos
Quitosana , Poluentes Químicos da Água , Quitosana/química , Corantes/química , Adsorção , Alginatos/química , Glutaral/química , Concentração de Íons de Hidrogênio , Poluentes Químicos da Água/química , CinéticaRESUMO
Currently, glutaraldehyde (GA)-crosslinked bioprosthetic heart valves (BHVs) still do not guarantee good biocompatibility and long-term effective durability for clinical application due to their subacute thrombus, inflammation, calcification, tearing and limited durability. In this study, double-modified xanthan gum (oxidized/vinylated xanthan gum (O2CXG)) was acquired from xanthan gum for subsequent double crosslinking and modification platform construction. Sulfonic acid groups with anticoagulant properties were also introduced through the free radical polymerization of vinyl sulfonate (VS) and vinyl on O2CXG. Taking advantage of the drug-loading function of xanthan gum, the treated pericardium was further loaded with inflammation-triggered dual drug-loaded nanogel (heparin (Hep) and atorvastatin (Ator)). Mechanical properties of O2CXG-crosslinked porcine pericardium (O2CXG-PP) were significantly improved via the first network formed by Schiff base bonds and the second C-C bonds network. Due to the presence of sulfonic acid groups as well as the dual drug release from nanogels under the stimulation of H2O2, the hemocompatibility, anti-inflammatory, pro-endothelialization and anti-calcification properties of the crosslinked pericardium modified with nanogels loaded with Hep and Ator (O2CXG+VS+(Hep+Ator) nanogel-PP) was significantly better than that of GA-crosslinked PP (GA-PP). The collaborative strategy of double crosslinking and sequential release of anticoagulant/endothelium-promoting drugs triggered by inflammation could effectively meet the requirement of enhanced multiple performance and long-term durability of bioprosthetic heart valves and provide a valuable pattern for multi-functionalization of blood contacting materials. STATEMENT OF SIGNIFICANCE: Currently, glutaraldehyde-crosslinked bioprosthetic heart valves (BHVs) are subject to subacute thrombus, inflammation, calcification and tearing, which would not guarantee good biocompatibility and long-term effective durability. We developed a cooperative strategy of double crosslinking and surface modification in which double-modified xanthan gum plays a cornerstone. The mechanical properties of this BHV were significantly improved via the first network formed by Schiff base bonds and the second C-C bonds network. Inflammation-triggered combination delivery of heparin and atorvastatin has been demonstrated to enhance anticoagulation, anti-inflammatory and pro-endothelialization of BHVs by utilizing local inflammatory response. The collaborative strategy could effectively meet the requirement of enhanced multiple performance and long-term durability of BHVs and provide a valuable pattern for the multi-functionalization of blood-contacting materials.