Hydrophilic Nonwoven Nanofiber Membranes as Nanostructured Supports for Enzyme Immobilization

Abstract

The high porosity, interconnected pore structure, and high surface area-to-volume ratio make the hydrophilic nonwoven nanofiber membranes (NV-NF-Ms) promising nanostructured supports for enzyme immobilization in different biotechnological applications. In this work, NV-NF-Ms with excellent mechanical and chemical properties were designed and fabricated by electrospinning in one step without using additives or complicated crosslinking processes after electrospinning. To do so, two types of ultrahigh-molecular-weight linear copolymers with very different mechanical properties were used. Methyl methacrylate- co -hydroxyethyl methacrylate (p(MMA)- co -p(HEMA)) and methyl acrylate- co -hydroxyethyl acrylate (p(MA)- co -p(HEA)) were designed and synthesized by reverse atom transfer radical polymerization (reverse-ATRP) and copper-mediated living radical polymerization (Cu 0 -MC-LRP), respectively. The copolymers were characterized by nuclear magnetic resonance ( 1 H-NMR) spectroscopy and by triple detection gel permeation chromatography (GPC). The polarity, topology, and molecular weight of the copolymers were perfectly adjusted. The polymeric blend formed by (MMA) 1002 - co -(HEMA) 1002 ( M w = 230,855 ± 7418 Da; M n = 115,748 ± 35,567 Da; PDI = 2.00) and (MA) 11709 - co -(HEA) 7806 ( M w = 1.972 × 10 6 ± 33,729 Da; M n = 1.395 × 10 6 ± 35,019 Da; PDI = 1.41) was used to manufacture (without additives or chemical crosslinking processes) hydroxylated nonwoven nanofiber membranes (NV-NF-Ms-OH; 300 nm in fiber diameter) with excellent mechanical and chemical properties. The morphology of NV-NF-Ms-OH was studied by scanning electron microscopy (SEM). The suitability for enzyme binding was proven by designing a palette of different surface functionalization to enable both reversible and irreversible enzyme immobilization. NV-NF-Ms-OH were successfully functionalized with vinyl sulfone (281 ± 20 μmol/g), carboxyl (560 ± 50 μmol/g), and amine groups (281 ± 20 μmol/g) and applied for the immobilization of two enzymes of biotechnological interest. Galactose oxidase was immobilized on vinyl sulfone-activated materials and carboxyl-activated materials, while laccase was immobilized onto amine-activated materials. These preliminary results are a promising basis for the application of nonwoven membranes in enzyme technology.

Overcoming Biochemical Limitations of Galactose Oxidase through the Design of a Solid‐Supported Self‐Sufficient Biocatalyst

“…Moreover, yields of 96 % to DFF (2,5‐diformylfuran) were obtained with the co‐immobilized enzymes, much higher than the 53 % obtained with soluble enzymes without co‐immobilization. In the study of Medina‐Castillo et al., [26] the immobilization yields were moderate (40–70 %) by both covalent attachment and ionic adsorption; however, the effectiveness factors were significantly low (1‐10 %), showing the difficulty of successfully immobilizing this enzyme on activated carriers.…”

“…The specific technical features of the support materials used in this study can be found in the Supporting Information (Table S1). To achieve an effective immobilization of GalOx, we explored different strategies based on previous successful immobilization procedures [26] . The first strategy we tested was covalent bonding via glyoxyl groups.…”

“…In this manuscript, our primary objective was to develop an active and stable solid‐supported biocatalyst with galactose oxidase (GalOx). We have used a commercial preparation of the galactose oxidase broadly used in recent examples [26,27,41] . To achieve this, we conducted a comprehensive screening of various materials and immobilization chemistries to identify the suitable combination that would exhibit both high activity and stability.…”

Design of colorimetric nanostructured sensor phases for simple and fast quantification of low concentrations of acid vapors

“…First, the hydroxyl groups of cellulose were functionalized with vinyl sulfone groups to obtain Cellu-VS. To do so, a piece of cellulose (16 × 11 cm) was introduced into 70 mL of a solution of DVS (0.33 M) in sodium carbonate buffer (333 mM) at pH = 12.00 for 2 h. Subsequently, Cellu-VS was washed three times with distilled water for 15 min and dried at 50 °C in a vacuum oven. It is well-known that vinyl sulfone groups can react easily with amine groups in mild conditions by a Michael-type reaction [ 26 ]. Thus, in a second step, Cellu-VS (16 × 11 cm) was introduced into 70 mL of a solution of ethylene diamine (0.33 M) in phosphate buffer (100 mM) at pH = 8 for 4 h. Then, Cellu-NH 2 was washed three times with distilled water and dried at 50 °C in a vacuum oven.…”

Cellulase Immobilization on Nanostructured Supports for Biomass Waste Processing

“…Nanostructured supports are materials containing nanometer size features (normally, between 1 and 100 nm), such as nanoparticles (NPs) of different sizes and shapes including nanorods and nanofibers, materials with pores in a nanometer range, stimuli responsive nano-carriers, etc. [ 11 , 12 , 13 , 14 , 15 , 16 ]. This growing interest is explained by a possibility of nanobiocatalysts to overcome deficiencies of enzymes immobilized on traditional supports.…”