Superparamagnetic poly(methyl methacrylate) beads for nattokinase purification from fermentation broth

An effective method for purification of nattokinase from fermentation broth using magnetic poly(methyl methacrylate) (PMMA) beads immobilized with p-aminobenzamidine was proposed in this study. Firstly, magnetic PMMA beads with a narrow size distribution were prepared by spraying suspension polymerization. Then, they were highly functionalized via transesterification reaction with polyethylene glycol. The surface hydroxyl-modified magnetic beads obtained were further modified with chloroethylamine to transfer the surface amino-modified magnetic functional beads. The morphology and surface functionality of the magnetic beads were examined by scanning electron microscopy and Fourier transform infrared. An affinity ligand, p-aminobenzamidine was covalently immobilized to the amino-modified magnetic beads by the glutaraldehyde method for nattokinase purification directly from the fermentation broth. The purification factor and the recovery of the enzyme activity were found to be 8.7 and 85%, respectively. The purification of nattokinase from fermentation broth by magnetic beads only took 40 min, which shows a very fast purification of nattokinase compared to traditional purification methods.     

Preparation of poly(methyl methacrylate)–grafted Hyparrhenia hirta for methyl red removal from colored solutions

Poly(methyl methacrylate)–grafted Hyparrhenia hirta (PMMA-g-Hh) biopolymer was prepared through radical polymerization using potassium persulfate (KPS) and applied in adsorption of methyl red from colored solutions. Solvent amount, initiator concentration, monomer concentration, temperature, and reaction time were the reaction parameters investigated for grafting. The biopolymer was characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and x-ray diffraction spectroscopy (XRD). The adsorption process was investigated with respect to pH, contact time, initial concentration, adsorbent dosage, and temperature. The optimum adsorption parameters were pH 6, contact time 90 min, adsorbent dosage 0.6 g, and initial concentration 50 mg/L. The Langmuir adsorption model best fitted the adsorption process, with maximum adsorption capacities of 19.95, 6.89, and 4.02 mg/g at adsorbent dosages of 0.2, 0.6, and 1.0 g, respectively. The pseudo-second-order model described the kinetics data better. The adsorption process was physical, spontaneous, and endothermic. The adsorbent was still active after 10 adsorption-desorption cycles, showing its suitability for use in colored solutions treatment.     

Sequential interpenetrating polymer networks produced from vegetable oil based polyurethane and poly(methyl methacrylate)

Sequential interpenetrating polymer networks (IPNs) were prepared using polyurethane produced from a canola oil based polyol with primary terminal functional groups and poly(methyl methacrylate) (PMMA). The properties of the material were studied and compared to the IPNs made from commercial castor oil using dynamic mechanical analysis, differential scanning calorimetry, as well as tensile measurements. The morphology of the IPNs was investigated using scanning electron microscopy and transmission electron microscopy. The chemical diversity of the starting materials allowed the evaluation of the effects of dangling chains and graftings on the properties of the IPNs. The polymerization process of canola oil based IPNs was accelerated because of the utilization of polyol with primary functional groups, which efficiently lessened the effect of dangling chains and yielded a higher degree of phase mixing. The mechanical properties of canola oil based IPNs containing more than 75 wt % PMMA were comparable to the corresponding castor oil based IPNs; both were superior to those of the constituent polymers due to the finely divided rubber and plastic combination structures in these IPNs. However, when PMMA content was less than 65 wt %, canola oil based IPNs exhibited a typical mechanical behavior of rigid plastics, whereas castor oil based IPNs showed a typical mechanical behavior of soft rubber. It is proposed that these new IPN materials with high performance prepared from alternative renewable resources can prove to be valuable substitutes for existing materials in various applications.     

Enzyme-linked immunosorbent assay of Escherichia coli O157:H7 in surface enhanced poly(methyl methacrylate) microchannels

A novel surface treatment method was developed to enhance polymer-based microchannel enzyme-linked immunosorbent assay (ELISA) for Escherichia coli O157:H7 detection. By applying an amine-bearing polymer, poly(ethyleneimine) (PEI), onto poly(methyl methacrylate) (PMMA) surface at pH higher than 11, PEI molecules were covalently attached and their amine groups were introduced to PMMA surface. Zeta potential analysis and X-ray photoelectron spectroscopy (XPS) demonstrated that the alkali condition is preferable for PEI attachment onto the PMMA surface. The amine groups on the PMMA surface were then functionalized with glutaraldehyde, whose aldehyde groups served as the active sites for binding the antibody by forming covalent bonds with the amine groups of the protein molecules. This surface modification greatly improved antibody binding efficiency and the microchannel ELISA for E. coli O157:H7 detection. Compared with untreated PMMA microchannels, approximately 45 times higher signal and 3 times higher signal/noise ratio were achieved with the PEI surface treatment, which also shortened the time required for cells to bind to the microchannel surface to approximately 2 min, much less than that usually required for the same ELISA carried out in 96-well plates. The detection in the microchannel ELISA only required 5-8 cells per sample, which is also better than 15-30 cells required in multi-well plates. With the high sensitivity, short assay time, and small reagent consumption, the microchannel ELISA can be economically used for fast detection of E. coli O157:H7.     

Functionalization of poly(methyl methacrylate) (PMMA) as a substrate for DNA microarrays

A chemical procedure was developed to functionalize poly(methyl methacrylate) (PMMA) substrates. PMMA is reacted with hexamethylene diamine to yield an aminated surface for immobilizing DNA in microarrays. The density of primary NH₂ groups was 0.29 nmol/cm². The availability of these primary amines was confirmed by the immobilization of DNA probes and hybridization with a complementary DNA strand. The hybridization signal and the hybridization efficiency of the chemically aminated PMMA slides were comparable to the hybridization signal and the hybridization efficiency obtained from differently chemically modified PMMA slides, silanized glass, commercial silylated glass and commercial plastic Euray™ slides. Immobilized and hybridized densities of 10 and 0.75 pmol/cm², respectively, were observed for microarrays on chemically aminated PMMA. The immobilized probes were heat stable since the hybridization performance of microarrays subjected to 20 PCR heat cycles was only reduced by 4%. In conclusion, this new strategy to modify PMMA provides a robust procedure to immobilize DNA, which is a very useful substrate for fabricating single use diagnostics devices with integrated functions, like sample preparation, treatment and detection using microfabrication and microelectronic techniques.     

Molecular Mobility in Poly(butyl methacrylate) and Poly(methyl methacrylate)

Molecular mobility is studied in poly(butyl methacrylate) and poly(methyl methacrylate) (PMMA) with molecular dynamics simulations in order to understand the effect of the α–β crossover on the β relaxation activation energy and co-operativity. In the high frequency range investigated, the estimated β process activation energy is decreased as compared to the low frequency value. This deviation is stronger in poly(n butyl methacrylate) (PnBMA) than in PMMA. The intra-molecular co-operativity related to the β process is also higher in PnBMA than in PMMA. These results could be related to the relative position of the simulation temperature range and of the extrapolated α–β crossover temperature.     

Biomimetic poly(methyl methacrylate)-based terpolymers: modulation of bacterial adhesion effect

Adherence of Staphylococcus aureus, responsible for major foreign body infections, was assessed onto functionalized poly(methyl methacrylate)-based terpolymers bearing sulfonate and carboxylate groups and onto poly(methyl methacrylate) as control. These terpolymers, have been synthesized by radical copolymerization of methyl methacrylate, methacrylic acid, and sodium styrene sulfonate by varying the ratio R = [COO(-)]/[COO(-) + SO(3)(-)] from 0 to 1 and keeping ionic monomer content between 7 and 18%. Adsorption of fibronectin onto poly(methyl methacrylate) was shown to dramatically promote bacterial adherence, whereas a strong inhibition of bacteria adherence was observed onto functionalized terpolymers containing both carboxylate and sulfonate groups. When terpolymers were predominantly functionalized by carboxylate groups, bacteria adherence was favored and reached values close to those obtained for poly(methyl methacrylate). These results have been related to the distribution of the anionic groups along the macromolecular chains, creating active sites responsible for specific interactions with fibronectin and inducing modifications of its conformation. The conformation of the adsorbed adhesive protein was then suggested to have an influence on the availability of its interaction sites to bacteria adhesins and therefore on modulation of bacteria adherence. Inhibition of Staphylococcus aureus adherence by functionalized poly(methyl methacrylate)-based terpolymers is of great interest in the field of biomedical implants and especially in the case of ophthalmic applications.     

Chitosan-functionalized poly(methyl methacrylate) particles by spinning disk processing for lipase immobilization

Chitosan-functionalized poly(methyl methacrylate) (PMMA-CH) particles were prepared by complexation between the negatively charged PMMA particles and the positively charged chitosan via a spinning disk processing. Processing parameters; feed rate and spinning speed, were optimized, which were traced by size distribution profiles of the formed PMMA-CH particles. Their sizes and net surface charges were found to be affected by MWs of chitosan (45, 100, and 230 kDa) used. Microscopic evidences were used to confirm their core–shell morphology. Chemical characteristics and thermal stability of such particles were determined by FTIR and TGA, respectively. Then, their ability to immobilize lipase (EC 3.1.1.3) was conducted and followed through zeta potential measurement. The percentage of lipase adsorption capacity increased with increasing lipase content, but the value decreased when the size of PMMA-CH particles increased. Also, the activity of lipase attached on PMMA-CH particles’ surface was preserved and increased with lipase loading.     

Physical properties of sequential interpenetrating polymer networks produced from canola oil-based polyurethane and poly(methyl methacrylate)

Sequential interpenetrating polymer networks (IPNs) were prepared using polyurethane (PUR) synthesized from canola oil-based polyol with terminal primary functional groups and poly(methyl methacrylate) (PMMA). The properties of the material were evaluated by dynamic mechanical analysis (DMA), differential scanning calorimetry (DSC), and modulated differential scanning calorimetry (MDSC), as well as tensile properties measurements. The morphology of the IPNs was investigated using scanning electron microscopy (SEM) and MDSC. A five-phase morphology, that is, sol phase, PUR-rich phase, PUR-rich interphase, PMMA-rich interphase, and PMMA-rich phase, was observed for all the IPNs by applying a new quantitative method based on the measurement of the differential of reversing heat capacity versus temperature from MDSC, although not confirmed by SEM, most likely due to resolution restrictions. NCO/OH molar ratios (cross-linking density) and compositional variations of PUR/PMMA both affected the thermal properties and phase behaviors of the IPNs. Higher degrees of mixing occurred for the IPN with higher NCO/OH molar ratio (2.0/1.0) at PUR concentration of 25 wt %, whereas for the IPN with lower NCO/OH molar ratio (1.6/1.0), higher degrees of mixing occurred at PUR concentration of 35 wt %. The mechanical properties of the IPNs were superior to those of the constituent polymers due to the finely divided rubber and plastic combination structures in these IPNs.     

Hydrogen bonding in stereoregular poly(methyl methacrylate)/poly(vinyl chloride) blends as studied by molecular dynamics simulations

In this work, two technically important polymer blends composed of isotactic poly (methyl methacrylate) (iPMMA) or syndiotactic poly (methyl methacrylate) (sPMMA) and isotactic poly (vinyl chloride) (iPVC) have been extensively investigated by molecular dynamics simulations. It is confirmed that sPMMA exhibits stronger interactions with iPVC than does iPMMA, and the non-conventional hydrogen bonds (HBs) occur between the two distinct components. Furthermore, the HBs in sPMMA/iPVC are more than those in iPMMA/iPVC, and the structural relaxation of HBs is closely associated with the backbone chain dynamics, which well explain the experimental trends in miscibility of the two systems and in glass transition temperature of single components. It should be noted that these results cannot be directly obtained by the experiments and single simulations, and the multiscale schemes used to prepare the initial all-atomistic configurations can play an important role. This work provides some key clues to improve the performance of polymer materials.     

Competitive cellular uptake of nanoparticles made from polystyrene, poly(methyl methacrylate), and polylactide

The uptake behavior of negatively charged fluorescent nanoparticles made from different polymers (PS, PMMA, and PLLA) is studied on HeLa cells. All particles are obtained by the miniemulsion process using sodium dodecylsulfate as anionic surfactant. The size of the particles is in the range 105-125 nm. Cell uptake is analyzed by flow cytometry and reveals a higher uptake of PLLA particles compared to PMMA and PS particles. In competitive uptake studies two different types of particles are co-incubated with the HeLa cells; the results indicate a mutual influence of the particles on their uptake behavior. A reduced internalization of PLLA particles in the presence of PS particles is observed, although neither the co-incubation of PMMA and PLLA nor of PMMA and PS shows similar effect.     

Activity,conformation and dynamics of cutinase adsorbed on poly(methyl methacrylate) latex particles

The adsorption of a recombinant cutinase from Fusarium solani pisi onto the surface of 100 nm diameter poly(methyl methacrylate) (PMMA) latex particles was evaluated. Adsorption of cutinase is a fast process since more than 70% of protein molecules are adsorbed onto PMMA at time zero of experiment, irrespective of the tested conditions. A Langmuir-type model fitted both protein and enzyme activity isotherms at 25 degrees C. Gamma(max) increased from 1.1 to 1.7 mg m(-2) and U(max) increased from 365 to 982 U m(-2) as the pH was raised from 4.5 to 9.2, respectively. A decrease (up to 50%) in specific activity retention was observed at acidic pH values (pH 4.5 and 5.2) while almost no inactivation (eta(act) congruent with 87-94%) was detected upon adsorption at pH 7.0 and 9.2. Concomitantly, far-UV circular dichroism (CD) spectra evidenced a reduction in the alpha-helical content of adsorbed protein at acidic pH values while at neutral and alkaline pH the secondary structure of adsorbed cutinase was similar to that of native protein. Fluorescence anisotropy decays showed the release of some constraints to the local motion of the Trp69 upon protein adsorption at pH 8.0, probably due to the disruption of the tryptophan-alanine hydrogen bond when the tryptophan interacts with the PMMA surface. Structural data associated with activity measurements at pH 7.0 and 9.2 showed that cutinase adsorbs onto PMMA particles in an end-on orientation with active site exposed to solvent and full integrity of cutinase secondary structure. Hydrophobic interactions are likely the major contribution to the adsorption mechanism at neutral and alkaline pH values, and a higher amount of protein is adsorbed to PMMA particles with increasing temperature at pH 9.2. The maximum adsorption increased from 88 to 140 mg cutinase per g PMMA with temperature raising from 25 to 50 degrees C, at pH 9.2.     

Design of recycling system for poly(methyl methacrylate) (PMMA). Part 1: recycling scenario analysis

Introduction

In this series of papers, we present a poly(methyl methacrylate) (PMMA) recycling system design based on environmental impacts, chemical hazards, and resource availability. We evaluated the recycling system by life cycle assessment, environment, health, and safety method, and material flow analysis.

Purpose

Previous recycling systems have not focused on highly functional plastics such as PMMA, partly because of lower available volumes of waste PMMA compared with other commodity plastics such as polyethylene or polypropylene. However, with the popularization of PMMA-containing products such as liquid crystal displays, the use of PMMA is increasing and this will result in an increase in waste PMMA in the future. The design and testing of recycling systems and technologies for treating waste PMMA is therefore a high research priority. In this study, we analyze recycling of PMMA monomers under a range of scenarios.

Methods

Based on the differences between PMMA grades and their life cycles, we developed a life cycle model and designed a range of scenarios for PMMA recycling. We obtained monomer recycling process inventory data based on the operational results of a pilot plant. Using this process inventory data, we quantified life cycle greenhouse gas (LC-GHG) emissions and fossil resource consumption, and we calculated the LIME single index.

Results and discussion

PMMA produces more than twice the amount of GHG emissions than other commodity resins. Through scenario and sensitivity analyses, we demonstrated that monomer recycling is more effective than mechanical recycling. Operational modifications in the monomer recycling process can potentially decrease LC-GHG emissions.

Conclusions

Highly functional plastics should be recycled while maintaining their key functions, such as the high transparency of PMMA. Monomer recycling has the potential to achieve a closed-loop recycling of PMMA.     

Preparation of cellulose graft poly(methyl methacrylate) copolymers by atom transfer radical polymerization in an ionic liquid

Cellulose graft poly(methyl methacrylate) copolymers were prepared by atom transfer radical polymerization (ATRP) in an ionic liquid. Cellulose chloroacetate, as a macroinitiator, was first synthesized by direct acylation of cellulose with chloroacetyl chloride without any catalysts under mild conditions in an ionic liquid, 1-allyl-3-methylimidazolium chloride (BMIMCl). Then, the macroinitiator was used for the ATRP of MMA mediated by the CuBr and 2,2′-bipyridine (bpy) catalysis system. The copolymerization was carried out in BMIMCl without homopolymer byproduct. The polymers were easily separated from the catalyst when the ionic liquid was used as reaction medium. The grafting copolymers were characterized by means of ¹H NMR, AFM and GPC. The results showed that the obtained copolymers had grafted polymer chains with well-controlled molecular weight and polydispersity, and the polymerization was a “living/controlled” system. Further, through AFM observation, it was found that the cellulose graft copolymer in solution could aggregate and self-assembly into sphere-like polymeric structure.     

Ultrathin and smooth poly(methyl methacrylate) (PMMA) films for label-free biomolecule detection with total internal reflection ellipsometry (TIRE)

Ultrathin poly(methyl methacrylate) PMMA films were prepared on gold substrates by spin coating PMMA dissolved in toluene. By varying the concentration of PMMA, spin coating speed and curing condition, we obtained very smooth and ultrathin PMMA films. The PMMA films were transformed into highly reactive film containing carboxylic functionalities using UV/O(3) irradiation. These films were shown to remain stable and reactive for at least one week. We then demonstrated the application of the UV/O(3) treated PMMA films for the detection of microRNAs using a label-free detection method called total internal reflection ellipsometry (TIRE). A limit of detection of 10 pM was established. The technique proposed here is a simple and quick method for generating carboxylic functional films for label-free bioanalytical detection techniques.     

Modeling and analysis of the compatibility of polystyrene/poly(methyl methacrylate) blends with four inducing effects

The compatibility of polystyrene (PS) and poly(methyl methacrylate) (PMMA) blends was studied over a wide range of compositions at 383, 413 and 443 K, respectively, by atomistic and mesoscopic modeling. All the calculated Flory–Huggins interaction parameters showed positive values; furthermore, they were all above the critical Flory–Huggins interaction parameter value, which means that the PS/PMMA blends were immiscible. Both the addition of a block copolymer and the introduction of a shear field influenced the phase morphologies of the blends, while the degree of influence depended on the compositions of the blends. The study of PS/PMMA blends doped with nanoparticles showed that the mesoscopic phase was influenced by not only the properties of the nanoparticles, such as their size, number and number density, but also the compositions of the blends. The effect of the surface roughness of the planes on the phase separation of the blends was also studied.     

Magnetic poly(glycidyl methacrylate) microspheres for protein capture

Immunoprecipitation of protein p53 from cell lysate on magnetic PGMA microspheres.

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Design of recycling system for poly(methyl methacrylate) (PMMA). Part 2: process hazards and material flow analysis

Introduction

In this series of papers, we present a design of poly(methyl methacrylate) (PMMA) recycling system considering environmental impacts, chemical hazards, and resource availability. We applied life cycle assessment (LCA), environment, health, and safety (EHS) assessment as well as material flow analysis to the evaluation of the recycling system.

Purpose

Recycling systems for highly functional plastics such as PMMA have not been studied sufficiently. Along with the popularization of PMMA-containing products such as liquid crystal displays (LCDs), the use of PMMA is steadily increasing, which will result in more waste of PMMA in the next decades. In this study, pyrolysis process for recycling waste PMMA into methyl methacrylate (MMA) monomer was examined, considering not only general environmental impacts quantified by life cycle assessment but also local environment, health, and safety hazards, and raw material availability.

Methods

Process EHS hazards assessment was applied to quantify the local effects of the PMMA monomer recycling process. Process hazards are strongly connected with the hazardous properties of chemical substances and stream conditions within the process. Two alternative cooling methods exist, and their difference was analyzed by LCA and EHS assessment. Besides the process hazard, the availability of waste PMMA must be an important point for the feasibility of implementing the PMMA monomer recycling process. The available amount can be quantified by analyzing the material flow of PMMA-containing products. PMMA contained in LCDs as light guide panels was selected as a feasible source of waste PMMA, and the quantity of PMMA flows in the society was evaluated.

Results and discussion

In the case of PMMA, monomer recycling has less process hazard than the production of fresh MMA from crude oil. The implementation of circulated cooling water could significantly decrease the process hazard in PMMA pyrolysis attributable to chemical hazards. Material flow analysis revealed that the availability of waste PMMA shows a fluctuating trend in the next 20 years because of the sharp peak demand for LCD television sets. The fluctuation is strongly dependent on the lifetime of LCD television sets.

Conclusions

PMMA monomer recycling has a potential to reduce environmental impacts with a less process hazards than fresh MMA production from crude oil. The availability of waste PMMA has a strong relationship with the lifetime of LCD television sets. The multiple and comprehensive assessments can reveal various aspects of a process technology.     

Modulating fibroblast cell proliferation with functionalized poly(methyl methacrylate) based copolymers: chemical composition and monomer distribution effect

Poly(methyl methacrylate)-based terpolymers bearing sulfonate and carboxylate groups have been synthesized by radical copolymerization leading to polymers with random distributions of ionic monomer units. Fibroblast cells were seeded on terpolymers of various molar compositions of ionic groups. Kinetics of the cell proliferation were examined and systematically compared to the nonfunctionalized control polymer, poly(methyl methacrylate). Modulation of cell proliferation was observed on 15% ionic monomer content copolymers of various compositions (R = COO(-)/(COO(-) + SO(3)(-)) and varies from 0 to 1). The inhibition percentage of cell proliferation calculated for each polymer by comparison to the cell proliferation on the control was plotted against R and gave a maximum value for R close to 0.55. Copolymers with ionic group contents higher or lower than 15% exhibit inhibition percentages fitting with those previously observed for the same R values, showing that the hydrophilic properties are not sufficient to explain the modulation effect of this material toward cells. Moreover, for each polymer tested, cells, even if inhibited in growth, were shown to be viable, indicating that the synthesized terpolymers exhibit cytostatic properties excluding any cytotoxic effect. Such polymers may be used for the fabrication of biocompatible intraocular lenses and prevent secondary cataract.