A novel matrix for hydrophobic interaction chromatography and its application in lysozyme adsorption

PubMed ID: 25298972A novel 1-naphthylamine (NA) coupled poly(2-hydroxyethyl methacrylate-co-N-methacryloyl-(L)-histidine methyl ester) [NA-PHEMAH] supermacroporous monolithic hydrophobic cryogel was prepared via covalent coupling of NA to PHEMAH for adsorption of lysozyme from aqueous solution. Firstly, PHEMAH monolithic cryogel was prepared by radical cryocopolymerization of HEMA with MAH as a functional comonomer and N,N'-methylene-bisacrylamide (MBAAm) as a crosslinker directly in a plastic syringe, and then NA molecules were covalently attached to the imidazole rings of MAH groups of the polymeric structure. The prepared, NA-PHEMAH, supermacroporous monolithic hydrophobic cryogel was characterized by scanning electron microscopy (SEM). The effects of initial lysozyme concentration, pH, salt type, temperature and flow rate on the adsorption efficiency of monolithic hydrophobic cryogel were studied in a column system. The maximum amount of lysozyme adsorption from aqueous solution in phosphate buffer was 86.1 mg/g polymer at pH 8.0 with a flow rate of 1 mL/min. It was observed that lysozyme could be repeatedly adsorbed and desorbed with the NA-PHEMAH monolithic hydrophobic cryogel without significant loss of the adsorption capacity

A novel adsorbent for protein chromatography: Supermacroporous monolithic cryogel embedded with Cu2+-attached sporopollenin particles

WOS: 000286680900015PubMed: 21176840The aim of this study is to prepare supermacroporous cryogels embedded with Cu2+-attached sporopollenin particles (Cu2+-ASP) having large surface area for high protein adsorption capacity. Super-macroporous poly(2-hydroxyethyl methacrylate) (PHEMA)-based monolithic cryogel column embedded with Cu2+-ASP was prepared by radical cryo-copolymerization of 2-hydroxyethyl methacrylate (HEMA) with N,N'-methylene-bis-acrylamide (MBAAm) as cross-linker directly in a plastic syringe for affinity purification of human serum albumin (HSA). Firstly, Cu2+ ions were attached to sporopollenin particles (SP), then the supermacroporous PHEMA cryogel with embedded Cu2+-ASP was produced by free radical polymerization using N,N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) as initiator/activator pair in an ice bath. Embedded particles (10 mg) in PHEMA-based cryogel column were used in the adsorption/desorption of HSA from aqueous solutions. Optimum conditions of adsorption experiments were performed at pH 8.0 phosphate buffer, with flow rate of 0.5 mL/min, and at 5 degrees C. The maximum amount of HSA adsorption from aqueous solution was very high (677.4 mg/g SP) with initial concentration 6 mg/mL It was observed that HSA could be repeatedly adsorbed and desorbed to the embedded Cu2+-ASP in PHEMA cryogel without significant loss of adsorption capacity. (C) 2010 Elsevier B.V. All rights reserved

Investigation of protein adsorption performance of Ni2+-attached diatomite particles embedded in composite monolithic cryogels

WOS: 000295189400027PubMed: 21739602As a low-cost natural adsorbent, diatomite (DA) (2 mu m) has several advantages including high surface area, chemical reactivity, hydrophilicity and lack of toxicity. In this study, the protein adsorption performance of supermacroporous composite cryogels embedded with Ni2+-attached DA particles (Ni2+-ADAPs) was investigated. Supermacroporous poly(2-hydroxyethyl methacrylate) (PHEMA)-based monolithic composite cryogel column embedded with Ni2+-ADAPs was prepared by radical cryo-copolymerization of 2-hydroxyethyl methacrylate (HEMA) with N,N'-methylene-bis-acrylamide (MBAAm) as crosslinker directly in a plastic syringe for affinity purification of human serum albumin (H SA) both from aqueous solutions and human serum. The chemical composition and surface area of DA was determined by XRF and BET method, respectively. The characterization of composite cryogel was investigated by SEM. The effect of pH, and embedded Ni2+-ADAPs amount, initial HSA concentration, temperature and flow rate on adsorption were studied. The maximum amount of HSA adsorption from aqueous solution at pH 8.0 phosphate buffer was very high (485.15 mg/g DA). It was observed that HSA could be repeatedly adsorbed and desorbed to the embedded Ni2+-ADAPs in poly(2-hydroxyethyl methacrylate) composite cryogel without significant loss of adsorption capacity. The efficiency of albumin adsorption from human serum before and after albumin adsorption was also investigated with SDS-PAGE analyses

Monolithic hydrophobic cryogel columns for protein separation

*Erzengin, Mahmut ( Aksaray, Yazar ) *Odabaşı, Mehmet ( Aksaray, Yazar )The present study was conducted for the synthesis of a novel supermacroporous monolithic hydrophobic adsorbent for lysozyme (Lyz) selected as a model protein from aqueous solution. After preparation of poly(2-hydroxyethyl methacrylate-co-N-methacryloyl-(l)-tyrosine methyl ester) monolithic cryogel column, 1-naphthylamine was covalently attached, and the prepared column was abbreviated as NA-Mcc. Scanning electron microscopy, Fourier transform infrared spectroscopy and Brunauer, Emmett and Teller device were utilized for the morphology, functional groups and surface area measurements of the column. Effects of several parameters including Lyz content of the adsorption solution, pH of the medium used, ambient temperature, type of salt and flow rate on the adsorption capacity of the polymeric material were examined in continuous operation. The maximum value achieved for Lyz adsorption from aqueous phase was found to be 105.8 mg/g in phosphate buffer. In addition, NA-Mcc was investigated in terms of reusability, and it was demonstrated that there is no significant change in the adsorption properties of prepared monolithic hydrophobic cryogels after 30 adsorption-desorption cycles

Is uterine blood flow influenced by hCG and mare age?

Doppler ultrasonography is a noninvasive technique which enables us to follow the physiologic and physiopathologic changes in blood flow in tissues. It is becoming an essential tool in veterinary medicine, especially in theriogenology. Twenty-seven Arabian mares were grouped by age ('young', 3-10 y, n = 15; 'old', 19-23 y, n = 12). The uterine arteries of the mares were examined using Doppler ultrasonography when an ovarian follicle >= 35 mm was visible (Day -1). After these measurements, human chorionic gonadotropin (hCG; 2500 IU) was administered to 14 mares selected randomly. One day later (Day 0) Doppler ultrasonography was repeated and then the dominant follicles were aspirated to collect follicular fluid in all groups. On the next day (Day +1), Doppler indices of the uterine artery blood flow were measured again. Blood samples were also collected just prior to ultrasonography, for measuring serum estradiol and progesterone levels. We found that preovulatory hCG administration had no significant effects on uterine artery blood flow indices, or serum or follicular fluid estradiol concentrations. The uterine artery resistance index might decrease in young mares after ovulation, possibly because of increased uterine perfusion. (C) 2014 Elsevier B.V. All rights reserved