Fabrication of a novel hemin-based monolithic column and its application in separation of protein from complex bio-matrix.
A novel polymer-based monolithic column was prepared via redox initiation system within the confines of a stainless steel column with 4.6mm i.d. In the processes, hemin and lauryl methacrylate were used as co-monomers; ethylene dimethacrylate as crosslinking agent; n-butyl alcohol, ethanediol, and N, N-dimethylformamide as tri-porogens; benzoyl peroxide and N, N-dimethyl aniline as redox initiation system. The resulting polymer-based monolithic columns were characterized by scanning electron microscopy, nitrogen adsorption-desorption instrument, and mercury intrusion porosimeter, respectively. The results illustrated that the improved monolith had relative uniform porous structure, good permeability, and low back pressure. Aromatic compounds were used to test the chromatographic behavior of the monolith, resulting in highest column efficiency of 19 880 plates per meter with reversed-phase mechanism. Furthermore, the homemade monolith was used as the stationary phase of high performance liquid chromatography to separate proteins from complex bio-matrix, including human plasma, egg white, and snailase. The results showed that the monolithic column occupied good separation ability with these complex bio-samples. Excellent specific character of the homemade hemin-based monolith was that it could simultaneously remove high-abundance proteins (including human serum albumin, immunoglobulin G, and human fibrinogen) from human plasma and separate other proteins to different fractions.
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China.
Xiaoya
X
Jiang
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China.
Doudou
D
Zhang
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China; Key Laboratory of Medicinal Chemistry and Molecular Diagnosis, Ministry of Education, Baoding 071002, China.
Xueying
X
Li
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China; Key Laboratory of Medicinal Chemistry and Molecular Diagnosis, Ministry of Education, Baoding 071002, China.
Xixi
X
Wang
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China; Key Laboratory of Medicinal Chemistry and Molecular Diagnosis, Ministry of Education, Baoding 071002, China. Electronic address: bailigai@163.com.
Ligai
L
Bai
College of Pharmaceutical Sciences, Hebei University, Baoding 071002, China; Key Laboratory of Medicinal Chemistry and Molecular Diagnosis, Ministry of Education, Baoding 071002, China.
Haiyan
H
Liu
Key Laboratory of Medicinal Chemistry and Molecular Diagnosis, Ministry of Education, Baoding 071002, China.
Hongyuan
H
Yan
D020001:1-Butanol / Q000737:chemistry; D000327:Adsorption; D002845:Chromatography / Q000379:methods*; D006427:Hemin / Q000737:chemistry*; D008689:Methacrylates / Q000737:chemistry; D008855:Microscopy, Electron, Scanning / Q000379:methods; D010539:Permeability; D011108:Polymers / Q000737:chemistry; D016062:Porosity; D011506:Proteins / Q000737:chemistry*
D008689:Methacrylates; D011108:Polymers; D011506:Proteins; C115846:dodecyl methacrylate; D006427:Hemin; C004919:ethylene dimethacrylate; D020001:1-Butanol
Bio-matrix; Hemin; Human plasma; Polymer monolithic columns; Protein