Tannic acid purification from pomegranate peel via tannic acid imprinted particle-embedded cryogel column.

Tannic acid (TA) is hydrolysable tannin found in the leaves and bark of many herbaceous and woody plants. Purification of TA is important due to its antibacterial, antihistaminic, antioxidant, antimutagenic and antitussive properties. In this study, 2-hydroxyethyl methacrylate-based TA-imprinted particle embedded cryogel (TA-MIP) was synthesized to purify TA from pomegranate peel. Furthermore, non-imprinted particle embedded cryogel (NIP) was synthesized to determine specific adsorption properties of TA-MIP, and control cryogel was synthesized without embedding procedure. The synthesized cryogel columns were characterized via scanning electron microscopy, Brunauer-Emmett-Teller surface area analysis, fourier-transform infrared spectroscopy, and swelling studies. Particle-embedding procedure resulted in a significantly higher specific surface area of particle-embedded columns (TA-MIP and NIP, 29 m2/g and 25 m2/g, respectively) than the specific surface area of control cryogel (9 m2/g). Adsorption studies were performed from aqueous solutions and maximum TA adsorption was found to be 34.4 mg/g for TA-MIP, 3.9 mg/g for NIP, and 2.8 mg/g for control cryogel. Within the scope of selectivity study, it was demonstrated that the synthesized columns have a high selectivity for TA against gallic acid (GA) and quercetin (QCT). Finally, purification of TA directly from pomegranate peel extract was studied and results were confirmed by HPLC. Furthermore, it has been proven that TA-MIP cryogel columns can be repeatedly used up to ten-times without any remarkable reduction in the TA adsorption amount.

Enhanced invertase binding from baker's yeast via cryogels included boronic acids.

Invertase, an industrially significant glycoenzyme, was purified from baker's yeast using poly (2-Hydroxyethyl methacrylate) [PHema-Pba] cryogels functionalized with boronic acid. At subzero temperatures, PHema-Pba cryogels were synthesized and characterized using swelling tests, scanning electron microscopy, and Fourier-transform infrared spectroscopy. The surface area of the PHema-Pba cryogels was 14 m2/g with a swelling ratio of 88.3% and macroporosity of 72%. The interconnected macropores of PHema-Pba cryogels were shown via scanning electron microscopy. Invertase binding capacity of PHema-Pba cryogel was evaluated by binding studies in different pH, temperature, and interaction time conditions and the maximum Invertase binding of PHema-Pba cryogel was found as 15.2 mg/g. and 23.7 fold Invertase purification was achieved from baker's yeast using PHema-Pba cryogels. The results show that PHema-Pba cryogels have high Invertase binding capacity and may be used as an alternative method for enzyme purification via boronate affinity systems.

Pterostilbene loaded poly(vinyl alcohol)-gelatin cryogels as potential bioactive wound dressing material.

Cryogels are support materials which are good at mimicking extracellular matrix due to their excellent hydrophilicity, biocompatibility, and macroporous structure, thus they are useful in facilitating cell activities during healing process. In this study, polyvinyl alcohol-gelatin (PVA-Gel) based cryogel membranes loaded with pterostilbene (trans-3,5-dimethoxy-4-hydroxystilbene; PTS) (PVA-Gel/PTS) was synthesized as wound dressing materials. PVA-Gel and PVA-Gel/PTS were synthesized with the polymerization yields of 96% ± 0.23% and 98% ± 0.18%, respectively, and characterized by swelling tests, Brunauer-Emmett-Teller (BET) and scanning electron microscopy (SEM) analysis. The swelling ratios were calculated as 98.6% ± 4.93% and 102% ± 5.1%, macroporosities were determined as 85% ± 2.13% and 88% ± 2.2%, for PVA-Gel and PVA-Gel/PTS, respectively. It was determined that PVA-Gel and PVA-Gel/PTS have 17 m2 /g ± 0.76 m2 /g and 20 m2 /g ± 0.92 m2 /g surface areas, respectively. SEM studies were demonstrated that they have ~100 µm pore sizes. According to 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), trypan blue exclusion and live-dead assay results, it was observed that cell proliferation, cell number and cell viability were higher in PVA-Gel/PTS cryogel at 24, 48, and 72 h compared to PVA-Gel. A strong and transparent fluorescent light intensity was observed indicating higher cell population in PVA-Gel/PTS in comparison with PVA-Gel, according to 4',6-diamidino-2-phenylindole (DAPI) staining. SEM, F-Actin, Giemsa staining and inverted-phase microscope image of fibroblasts in PVA-Gel/PTS cryogels revealed that dense fibroblast proliferation and spindle-shaped morphology of cells were preserved. Moreover, DNA agarose gel data demonstrated that PVA-Gel/PTS cryogels had no effect on DNA integrity. Consequently, produced PVA-Gel/PTS cryogel can be used as wound dressing material to promote wound therapies, inducing cell viability and proliferation.

Enhanced laccase separation from fermentation medium using cryogel columns.

The laccase enzyme family belongs to the oxidoreductase enzyme class and is one of the most commercially valuable enzymes that catalyzes the oxidation of one electron of a wide range of phenolic compounds. Separation and purification of laccases are crucial for industry since they play an important role in dye decolorization, biodegradation and food processing. Therefore, developing effective, high yielding and cost-effective methods for laccase production is vital. In this study, it was aimed to prepare cryogel columns for laccase purification following the bioproduction of laccase via Aspergillus niger. 2-hydroxyethyl methacrylate based cryogels were synthesized in the presence of 1-vinylimidazole as the affinity ligand and characterized by swelling tests, Brunauer-Emmett-Teller surface area measurement and scanning electron microscopy analysis. Surface area and water uptake ratio of cryogel columns were 35 m2/g and 93 %, respectively. The effect of pH, equilibrium laccase concentration, flow rate, interaction time and temperature on laccase adsorption were examined. The purification factor was calculated as 10.53 under optimum conditions and the enzyme recovery was found to be 86.7 % from fermentation medium. Current study revealed that laccase purification using cryogels following filtration of fermentation medium could be a promising candidate for industrial applications with eliminating the need for complex chromatographic steps.

AFB1 recognition from liver tissue via AFB1 imprinted magnetic nanoparticles.

Aflatoxins (AFs) are produced mainly by Aspergillus flavus and Aspergillus parasiticus and aflatoxin B1 (AFB1) is one of the most toxic aflatoxins with its carcinogenic property. AFB1 recognition from samples is very important and PHEMA based AFB1 imprinted magnetic nanoparticles (magAFB1-MIPs) were synthesized for the selective AFB1 recognition from liver tissue. The AFB1-MIPs were synthesized in different mole ratios and NIPs were synthesized for control. Characterization studies of magAFB1-MIPs and NIPs were carried out by swelling tests, surface area measurements, scanning electron microscopy and particle size analysis. The surface area was found as 117 m2/g and the size of the nanoparticles were found as 483 nm in diameter. The percentage yield of polymerization was calculated as 98 % and the template (AFB1) removal ratio from the magAFB1-MIPs was calculated as 91 %. The maximum adsorbtion capacities were calculated as 427.57 ng g-1 for magAFB1-MIPs and 44.6 ng g-1 for magNIPs. Selectivity tests showed that magAFB1-MIPs adsorb AFB1 1.74, 4.40, 2.46 times selective than that of AFB2, AFG1 and AFG2 molecules, respectively. AFB1 removal amount from AFB1 spiked liver tissue was satisfactory and recorded as 10.4 ng g-1 and 54.8 ng g-1 for 2 ng g-1 and 10 ng g-1 spiked liver tissue samples, respectively. AFB1 adsorption amount decrease was found negligible for 10 consecutive adsorption-desorption repeats in reusability study.

Synthesis of Double-Layer Imprinted Polymers: BSA Depletion.

A sensitive, rapid, and cost-effective method for quantitative analysis of proteins (e.g., detection, purification, depletion) for a wide variety of purposes is required in a number of areas, such as immunodiagnostics and biotechnology. Double-layer imprinting technique, which is carried out via polymerization of polymer solution with higher monomer concentration, covering and filling the supermacroporous structure of a pre-synthesized cryogel column with a lower monomer concentration, thus improving the surface area and adsorption capacity of final product, is a brand new approach for the application of cryogels in molecular imprinting technology. Within the scope of this chapter, BSA is selected as a model protein for the application of double-layer imprinting protocol. In this chapter, synthesis of double-layer BSA-imprinted and non-imprinted cryogel columns (BSA-DLIP and DLNIP, respectively) are described. In addition, characterization of synthesized columns and BSA depletion studies from aqueous solutions are described in detail, as well as selectivity of BSA-DLIPs for BSA, against competitors.

Determination of Neopterin as a Prognostic Indicator Using Neopterin-Imprinted Cryogel Membranes.

Neopterin (Neo) is thought of as a key biomarker for the diagnosis and prognosis of a wide variety of diseases associated with cellular immune response. Therefore, it has become a vital need to be able to specifically determine the Neo concentration in human serum. Molecularly imprinted cryogels have come into prominence among other affinity systems by combining advantages of Molecular Imprinting Technology (MIT) and cryogels. In this chapter, synthesis of novel Neopterin-imprinted cryogel membranes (Neo-mip), characterization studies of synthesized materials, and their use in the determination of Neo in human serum is described in detail. In addition, the evaluation of selective Neo adsorption properties of Neo-mip against competitors (Pterin and Glucose) is discussed. Neo-mip will come into prominence as important affinity materials for the selective Neo recognition in body fluids, prior to use in the health sector.

Determination of mold contamination using ergosterol imprinted particles.

Ergosterol is a key biochemical marker for fungal mycelial growth. In this study, molecularly ergosterol imprinted particles (Erg-MIPs) were newly synthesized for the selective detection of ergosterol in mold samples. Erg-MIPs were characterized via scanning electron microscopy, swelling studies, and surface area measurements. Maximum selective ergosterol adsorption achieved as 28.50 mg/g Erg-MIP. Selectivity studies showed that Erg-MIPs adsorbed Erg 2.01 and 3.27 times higher than that of cholesterol and stigmasterol, respectively. Erg adsorption from Aspergillus niger was found as 23.87 mg/g. Reusability of Erg-MIPs was studied and decrease in Erg adsorption capacity of the particles was negligible (3%). Erg-MIPs are good affinity materials for the selective Erg detection from food samples, prior to use in food industry.

Molecularly imprinted spongy columns for Angiotensin(II) recognition from human serum.

Angiotensin II (AngII), the effector peptide of the renin angiotensin system and has an important role in regulating cardiovascular hemodynamics and structure. AngII is an important biomarker for certain diseases that are associated with cardiovascular disorders, i.e., influenza, SARS-CoV-2, tumors, hypertension, etc. However, AngII presents in blood in very low concentrations and they are not stable due to their reactivity, therefore spontaneous detection of AngII is a big challenge. In this study, AngII-imprinted spongy columns (AngII-misc) synthesized for AngII detection from human serum, and characterized by surface area measurements (BET), swelling tests, scanning electron microscopy (SEM), FTIR studies. AngII binding studies were achieved from aqueous environment and maximum binding capacity was found as 0.667 mg/g. It was calculated that the AngII-miscs recognized AngII 8.27 and 14.25 times more selectively than competitor Angiotensin I and Vasopressin molecules. Newly produced AngII-misc binds 60.5 pg/g AngII from crude human serum selectively. It has a great potential for spontaneous detection of AngII from human serum for direct and critical measurements in serious diseases, that is, heart attacks, SARS-CoV-2, etc.