In situ crosslinking polymerization of potassium sensitive hydrogels for advanced in vivo sensor applications is studied in detail.

Using colloidal polyacrylamide (PAAm) microgels as carriers, a novel strategy for covalent immobilization of enzymes maintained in hydrated microenvironment on/in a macroporous surface-functionalized hydrophobic polyvinylidene fluoride (PVDF) membrane is developed. The PAAm microgels are synthesized by inverse miniemulsion polymerization, and first the parameters are investigated which are suited to obtain particles in the desired size range, 100-200 nm, with narrow size distribution. Amino functions are then imparted to the microgels applying the Hofmann reaction. The modification is confirmed by Fourier-transform infrared spectroscopy analysis, ninhydrin test, and elemental analysis. In addition, functionalized microgels are characterized by dynamic light scattering. The amino-functionalized PAAm microgels are then immobilized on pre-modified PVDF membrane having aldehyde functionalities on the surface. Afterward, unreacted aldehyde groups still present on the membrane where quenched by ethanolamine and the enzyme lipase from Candida rugosa (LCR) is subsequently immobilized on the microgels loaded PVDF membrane via glutaraldehyde cross-linking, exploiting the free amino groups on immobilized microgels. Catalytic efficiency of LCR immobilized by this strategy is evaluated using para-nitrophenyl palmitate as substrate and compared with LCR directly immobilized on PVDF membrane without microgels. Results show that LCR immobilized by means of microgels exhibits better performance with a 2.3-fold higher specific biocatalytic activity.

This work presents new insights into material design and physicochemical interactions that are relevant for the use of glucose-responsive polymeric hydrogels in continuously operating biosensor systems. Investigated hydrogels were based on either acrylamide or N-isopropylacrylamide, covalently cross-linked by N,N′-methylenebis­(acrylamide), and 3-acrylamidophenylboronic acid and (N-(3-dimethylaminopropyl)) acrylamide were the comonomers to enable selective glucose binding at a physiological pH. A novel assay for the determination of the amount of bound glucose inside the hydrogel was developed, enabling the direct recording of these receptor effects parallel to the determination of the change of water content, i.e., free swelling. Binding isotherms, affinity constants, and maximum degree of complexation of boronic acid groups with glucose were determined. The affinity toward glucose could be increased 3-fold compared to literature values for phenylboronic acid free in solution by the use of a suitable hydrogel composition. The library of differently composed materials was then evaluated in a pressure sensor setup. Thereby, the long-term use of the hydrogels was established, and the hydrogels could be analyzed for a period of three months without the reduction of the pressure signal sensitivity. Based on all results, a composition that is suitable for efficient glucose recognition was identified, at which up to 25% water was released at 37 °C and pH 7.4 and a change of the glucose concentration from 0 to 10 mM. In the physiologically relevant range (3–10 mM), a linear dependence of the swelling pressure on the glucose concentration was found, allowing an accurate determination of glucose concentration. Overall, the obtained results provide significant progress in efforts to enable glucose detection by a robust sensor setup.

Solid–gas biocatalysis was performed in a specially designed continuous biocatalytic membrane reactor (BMR). In this work, lipase from Candida rugosa (LCR) and ethyl acetate in vapor phase were selected as model enzyme and substrate, respectively, to produce acetic acid and ethanol. LCR was immobilized on functionalized PVDF membranes by using two different kinds of chemical bond: electrostatic and covalent. Electrostatic immobilization of LCR was carried out using a membrane functionalized with amino groups, while covalent immobilization was carried out using membrane, with or without surface-immobilized polyacrylamide (PAAm) microgels, functionalized with aldehyde groups. These biocatalytic membranes were tested in a solid–gas BMR and compared in terms of enzyme specific activity, catalytic activity, and volumetric reaction rate. Results indicated that lipase covalently immobilized is more effective only when the immobilization is mediated by microgels, showing catalytic activity doubled with respect to the other system with covalently bound enzyme (4.4 vs 2.2 μmol h–1). Enzyme immobilized by ionic bond, despite a lower catalytic activity (3.5 vs 4.4 μmol h–1), showed the same specific activity (1.5 mmol·h–1·g–1 ENZ) of the system using microgels, due to a higher enzyme degree of freedom coupled with an analogously improved enzyme hydration. Using the optimized operating conditions regarding immobilized enzyme amount, ethyl acetate, and molar water flow rate, all three BMRs showed continuous catalytic activity for about 5 months. On the contrary, the free enzyme (in water/ethyl acetate emulsion) at 50 °C was completely inactive and at 30 °C (temperature optimum) has a specific activity 2 orders of magnitude lower (8.4 × 10–2 mmol h–1 g–1) than the solid–gas biocatalytic membrane reactor. To the best of our knowledge, this is the first example of solid–gas biocatalysis, working in the gaseous phase in which a biocatalytic membrane reactor, with the enzyme/substrate system lipase/ethyl acetate, was used.

Artificial model colloids are of special interest for the development of advanced sterile filters, able to efficiently separate pleomorphic, highly deformable and infectious bacteria such as mycoplasma, which, until now,...