Papain Adsorption Research Articles

Hollow spherical vaterite calcium carbonate prepared by spray-bubble template method for immobilization of papain

Calcium carbonate (CaCO3) possesses unique advantages as a carrier , however, some critical issues need to be addressed, such as low productivity, irregular structures, and weak adsorption ability. Herein, hollow spherical vaterite CaCO3 (HSVC) was synthesized in an improved spray apparatus with high mass transfer using carbon dioxide (CO2) as a template and was in turn applied for papain immobilization. Results revealed that HSVC can be prepared cleanly and efficiently with a regular spherical morphology having a surface area of 17.03 m2·g−1 and immobilization amount of 1068 mg·g−1 for papain. Interestingly, the pH stability, temperature stability and reusability of papain were dramatically improved after immobilization by HSVC. Molecular dynamics simulation (MDS) disclosed that the formation of CO32– and Ca2+ ion pairs was the key factor for the preparation of HSVC and the binding energy (BE) between amino acid molecules and vaterite (110) facilitated the ultra-high adsorption of papain. The decrease in BE of system after immobilization facilitated high catalytic capacity of the immobilized papain. The prepared HSVC exhibits promising prospects in enzymes immobilization based applications.

Chirality-specific Near-infrared Photoluminescent Responses of Single-walled Carbon Nanotubes to Detect Papain Adsorption Assisted by the Use of DNA and Frustules

Chirality-specific Near-infrared Photoluminescent Responses of Single-walled Carbon Nanotubes to Detect Papain Adsorption Assisted by the Use of DNA and Frustules

Fabrication of a novel core-shell-shell temperature-sensitive magnetic composite with excellent performance for papain adsorption.

Herein, a novel temperature-sensitive magnetic composite (Fe3O4@SiO2@P(NIPAM-co-VI)/Cu2+) with a uniform core–shell–shell structure was successfully prepared via a layer-by-layer method. The resulting magnetic composite revealed good magnetic properties and remarkable affinity to papain with a maximum adsorption capacity of 199.17 mg g−1. The adsorption equilibrium data fitted the pseudo-second-order kinetic and Freundlich models well, and the major thermodynamics parameters indicated that adsorption was an endothermic and spontaneous process. Fe3O4@SiO2@P(NIPAM-co-VI)/Cu2+ could thermally protect papain, which is attributed to the reversible hydrophilic–hydrophobic transition of the composite at temperatures below and above the lower critical solution temperature. More importantly, the magnetic composite could be recycled at least six times without a remarkable loss in its adsorption capacity, and the process of adsorption and elution had no significant effect on the activity and structure of papain. This work could provide a novel separation method for papain without loss of its activity.

Papain Adsorption on Latex Particles: Charging, Aggregation, and Enzymatic Activity.

The effect of papain adsorption on the surface charge properties and aggregation mechanism of sulfate-functionalized polystyrene latex particles was studied. The positively charged enzyme possessed a high affinity to the oppositely charged particles, giving rise to charge neutralization and charge reversal at appropriate papain concentrations. The tendency in the particle aggregation rates at different enzyme doses revealed that the colloidal stability of the samples is governed by interparticle forces of electrostatic origin. The aggregation mechanism was qualitatively described within the classical DLVO theory, and unstable dispersions were detected near the charge neutralization point, while particle aggregation was not observed at low and elevated papain concentrations. The relatively high dispersion stability of the bare latex particles was maintained upon the formation of an enzyme layer on the surface, and the obtained latex-papain composite showed notable resistance against salt-induced aggregation. Remarkable hydrolytic and antioxidant activities of the immobilized enzyme were observed in probe reactions; therefore, the obtained hybrid can be considered as a multifunctional biocatalytic system with great promise in applications in industrial manufacturing processes.

Artificial intelligence-based behavioral analysis of protein in a nanoscale cubic space on a Si substrate

Special behaviors of proteins have been exploited and used for biomedical devices and biosensors; fundamentally, the behavior of proteins on solid surfaces is fascinating. However, no comprehensive understanding has been garnered from current studies. In our study, the behavior of tetrapeptides in active sites of papain in a nanoscale cubic space on a solid surface was analyzed using a synergistic combination of artificial intelligence and computer simulations. After identifying suitable tetrapeptides, half of them were deemed not suitable for free papain adsorption. A nanoscale cubic space enabled novel functions of those proteins to be captured. Additionally, the decision tree identified attributes that governed suitability. The decision tree would be an effective tool to comprehend the big picture of the complex behavior.

Cage like ordered carboxylic acid functionalized mesoporous silica with enlarged pores for enzyme adsorption

Cubic FDU-12 type mesoporous silicas with enlarged pores and carboxylic acid (–COOH) functionality in the pore channels (denoted as LP-FTC-x) are synthesized using tetraethyl orthosilicate (TEOS) and carboxyethylsilanetriol sodium salt (CES) as silica sources, Pluronic F127 triblock copolymer as template, and trimethylbenzene (TMB) as pore expander, and utilized them as supports for enzyme immobilization. When the –COOH content is increased from 0 to 30%, the pore size of LP-FTC-x decreases from 23.6 to 11.1 nm, and its particle size decreases from around 2 μm to 600–800 nm. The material exhibits a high papain adsorption capacity (895 mg g−1) with a low leaching rate at pH 8.2 due to the well-defined surface chemistry in the pore channel. The adsorption kinetics and isotherms follow the pseudo-second-order model and the Langmuir isotherm model, respectively. The excellent structural properties of LP-FTC-x are also advantageous for enhancement in stability of enzyme toward the temperature, solution pH, and incubation time variations.

Reversible papain immobilization onto poly(AAm–MMA)-based cryogels

In the present work, new dye ligand affinity chromatographic support for papain adsorption was synthesized. For this purpose, poly(acrylamide–methyl methacrylate) [poly(AAm–MMA)] cryogels were synthesized by using the free radical cryopolymerization technique. These cryogels were then functionalized with Reactive Green 19 dye and were characterized with Fourier transform infrared, environmental scanning electron microscopy, macroporosity and energy-dispersive X-ray spectrometer analyses. Incorporation of the dye molecule onto cryogel structure was clearly demonstrated by these characterization techniques and pore diameter of the cryogel was found to be around 30–40μm. Effects of medium pH, initial papain concentration, medium temperature and ionic strength on the papain adsorption onto dye-attached cryogel were also investigated. Maximum papain adsorption was found to be 40.66 mg g−1 cryogel by using pH 5.5 acetate buffer at 25°C. Reusability profile of the cryogel was also investigated and it was found that the adsorption capacity of the cryogel decreased only about 2.13% at the end of the 10 reuses. Activity studies of papain in desorption medium were carried out and it was found that desorbed papain was active and showed 88.5% of its initial activity.

Adsorption of Papain on solid substrates of different hydrophobicity.

Adsorption properties of protein Papain at the solid|liquid (0.1 M KCl) interfaces of different hydrophobicity [highly oriented pyrolytic graphite (HOPG), bare gold, CH3, OH, and COOH-terminated self-assembled monolayers on gold] were studied by a combined quartz crystal microbalance and atomic force microscopy techniques. It was found that Papain forms an incomplete monolayer at hydrophobic interfaces (HOPG and CH3-terminated substrate), whereas on more hydrophilic ones, a complete monolayer formation was always observed with either the onset of the formation of a second layer (bare gold substrate) or adsorption in a multilayer fashion, possibly a bilayer formation (OH-terminated substrate). The surface concentration and compact monolayer film thickness was much lower on the COOH-terminated substrate compared to other surfaces studied. This result was explained by partial dissociation of the interfacial COOH groups leading to additional electrostatic interactions between the positively charged protein domains and negatively charged carboxylate anions, as well as to local pH changes promoting protein denaturation.

Adsorption and Activity of Papain on Hierarchically Nanoporous Silica

Adsorption and Activity of Papain on Hierarchically Nanoporous Silica

Usage of immobilized papain for enzymatic hydrolysis of proteins

In this presented work, magnetic poly(HEMA-GMA) nanoparticles were synthesized by using emulsion polymerization technique. These magnetic nanoparticles were then functionalized with the affinity dye ligand, Cibacron Blue F3GA. The dye attached magnetic nanoparticles were characterized by using FTIR, ESR, SEM, AFM and EDX analysis. Synthesized magnetic nanoparticles demonstrated magnetic behavior and their size was about 200nm. Dye loading on the nanoparticle was calculated as 173.96μmol/g polymer by using sulfur stoichiometry. Effects of medium pH, initial papain concentration and ionic strength on the papain adsorption onto dye attached magnetic nanoparticles were also investigated and maximum papain adsorption onto the nanoparticles was found to be 764.0mg/g polymer in pH 7.0 HEPES buffer. Kinetic constants of the free and immobilized papain were compared and their pH, thermal, operational and storage stabilities were investigated. It was demonstrated with these studies that, immobilized form of papain was much more stable than that of free form. At the end of the study, catalytic efficiency of immobilized papain was investigated toward the different type of proteins such as casein, BSA, IgG and cytochrome c and it was demonstrated that, highest catalytic efficiency was achieved with IgG.

Purification of Papain Using Reactive Green 5 Attached Supermacroporous Monolithic Cryogel

Supermacroporous poly(2-hydroxyethyl methacrylate) [poly(HEMA)] monolithic cryogel was prepared by radical cryocopolymerization of HEMA with N,N'-methylene bisacrylamide as crosslinker. Reactive Green 5 dye was immobilized to the cryogel with nucleophilic substitution reaction, and this dye attached cryogel column was used for affinity purification of papain from Carica papaya latex. Reactive Green 5-immobilized poly(HEMA) cryogel was characterized by swelling studies, Fourier transform infrared spectroscopy, scanning electron microscopy, and energy dispersive X-ray analysis. Maximum papain adsorption capacity was found to be 68.5 mg/g polymer while nonspecific papain adsorption onto plain cryogel was negligible (3.07 mg/g polymer). Papain from C. papaya was purified 42-fold in single step with dye attached cryogel, and purity of papain was shown by silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Preparation and properties of nitrogen-containing hollow carbon nanospheres by pyrolysis of polyaniline–lignosulfonate composites

Polyaniline–lignosulfonate composite hollow spheres were synthesized by using one-step unstirred polymerization of aniline in the presence of lignosulfonate. Novel nitrogen-containing hollow carbon nanospheres were prepared by direct pyrolysis of the polyaniline–lignosulfonate composite spheres at different temperatures under a nitrogen atmosphere. Thermal behavior of the polyaniline–lignosulfonate composite spheres was studied by TG–DTG, FTIR and element analyze instruments. The resultant carbon spheres were characterized by SEM, XRD and nitrogen adsorption–desorption measurement. It was found that the pyrolysis products of the polyaniline–lignosulfonate composite spheres were made up of uniform hollow carbon nanospheres with an average diameter of 135 nm. Furthermore, the hollow carbon nanospheres exhibit high BET surface area range from 381.6 m 2 g −1 to 700.2 m 2 g −1. The hollow carbon nanospheres could be used as adsorbents of papain. The papain adsorption capacity for the carbon spheres prepared at 1200 °C was up to 1161 mg g −1 at an initial papain concentration of 10 mg mL −1 at 25 °C.

Papain Adsorption on Chitosan-Coated Nylon-Based Immobilized Metal Ion (Cu2+, Ni2+, Zn2+, Co2+) Affinity Membranes

A novel immobilized metal affinity membrane was prepared for papain adsorption in this article. Higher papain adsorption capacity between 43-67 mg/g was observed and the adsorption isotherm fitted the Freundlich equation. Experimental data were analyzed using two adsorption kinetic models. The pseudo-second-order kinetic model provided better correlation to the experimental results. A significant amount of the adsorbed papain was eluted by 1.0 M NaSCN at pH 5.0 for all affinity membranes. It was concluded that the novel chitosan-coated nylon-based immobilized metal ion affinity membrane could be applied for the large-scale isolation of papain without resulting in enzyme denaturation.

Use of TOF-SIMS to Study Adsorption and Loading Behavior of Methylene Blue and Papain in a Nano-Porous Silicon Layer

TOF-SIMS was applied to study the cross-sectional distribution of methylene blue and papain in porous silicon layers. Elemental and molecular information were used to study their distributions in the porous region and the chemistry of their adsorption. Methylene blue (MW = 284 Da) penetrated to the base to the pores. Positive ions (SiCH(3)(+)) suggest methylene blue binds to the substrate via its methyl groups. Negative fragments (SiOSH(3)(-) and SiO(2)SCH(-)) also suggested chemisorption via O bridging of the substrate Si and methylene blue S. The larger Papain molecule (23,406 Da) distributed itself in a similar manner to methylene blue demonstrating larger molecules can be effectively incorporated into such pore structures.

Removal of mercury from its aqueous solution using charcoal-immobilized papain ( CIP)

In the present work mercury has been eradicated from its aqueous solution using papain, immobilized on activated charcoal by physical adsorption method. Operating parameters for adsorption of papain on activated charcoal like pH, amount of activated charcoal, initial concentration of papain in solution have been varied in a suitable manner for standardization of operating conditions for obtaining the best immobilized papain sample based on their specific enzymatic activity. The immobilized papain sample obtained at initial papain concentration 40.0 g/L, activated charcoal amount 0.5 g and pH 7 shows the best specific enzymatic activity. This sample has been designated as charcoal-immobilized papain ( CIP) and used for further studies of mercury removal. Adsorption equilibrium data fit most satisfactorily with the Langmuir isotherm model for adsorption of papain on activated charcoal. Physicochemical characterization of CIP has been done. The removal of mercury from its simulated solution of mercuric chloride using CIP has been studied in a lab-scale batch contactor. The operating parameters viz., the initial concentration of mercury in solution, amount of CIP and pH have been varied in a prescribed manner. Maximum removal achieved in the batch study was about 99.4% at pH 7, when initial metal concentration and weight of CIP were 20.0 mg/L and 0.03 g respectively. Finally, the study of desorption of mercury has been performed at different pH values for assessment of recovery process of mercury. The results thus obtained have been found to be satisfactory.

Comparison: Adsorption of papain using immobilized dye ligands on affinity membranes

Novel membranes that involve the immobilization of Reactive Red 120 or Reactive Brown 10 as dye ligands were prepared. These were used in the purification of papain from papaya powder extracts. Papain adsorption capacities for the Red 120 and Brown 10 membranes were 143.6 mg/g and 107.3 mg/g, respectively. The effectiveness of adsorption was demonstrated by Freundlich isotherm proficiency. The enzyme was eluted from the respective dye membranes using 1.0 M NaCl at pH 6.0 and yields of over 80% were found for the Red 120–CS (chitosan)–nylon membrane whereas only a 50% recovery was possible using the Brown 10–CS–nylon membranes. It is concluded that Red 120–CS–nylon membranes could play an active role in the separation and purification of papain from crude extracts. This system has the potential to be developed for the commercial isolation of the protein.

Optimization of adsorption conditions of papain on dye affinity membrane using response surface methodology

The adsorption of papain on Reactive Blue 4 dye–ligand affinity membrane was investigated in a batch system. The combined effects of operating parameters such as initial pH, temperature, and initial papain concentration on the adsorption were analyzed using response surface methodology. The optimum adsorption conditions were determined as initial pH 7.05, temperature 39 °C, and initial papain concentration 11.0 mg/ml. At optimum conditions, the adsorption capacity of dye–ligand affinity membrane for papain was found to be 27.85 mg/g after 120 min adsorption. The papain was purified 34.6-fold in a single step determined by fast protein liquid chromatography. More than 85% of the adsorbed papain was desorbed using 1.0 M NaCl at pH 9.0 as the elution agent. The purification process showed that the dye–ligand immobilized composite membrane gave good separation of papain from aqueous solution.

Influence of Pore Size on Enzymes Adsorption in Mesostructure Cellular Foams (MCFs)

Mesoporous silica materials are attractive candidates for enzymes immobilization due to their high surface area, tunable pore size, large pore volume and biocompatibility. In this work, two different enzymes, papain, a small globular enzyme with molecular diameter of 3.6nm, and catalase, a relatively larger enzyme with molecular diameter of 10.4nm were introduced into the pores of siliceous mesostructed cellular foams (MCFs) that had a large cellular pores of 29nm and a small window size of 12nm, respectively. The amount of adsorbed enzymes was found to be dependent on the molecular size of enzymes. The amount of adsorbed catalase was more than two times that of adsorbed papain, suggesting that MCFs with large pores is a suitable host for large enzymes adsorption. The blocking of pores which resulted from aggregation of enzymes in the windows of MCFs and the higher leaching of enzymes form MCFs may be the reason of lower papain adsorption capacity in MCFs.

Electrospun nylon nanofiber as affinity membrane for papain adsorption

Electrospun nylon nanofiber as affinity membrane for papain adsorption

Analysis of papain adsorption on nylon-based immobilized copper ion affinity membrane

Analysis of papain adsorption on nylon-based immobilized copper ion affinity membrane