title: Capillary electrophoresis-based immobilized enzyme reactor using particle-packing technique.
abstract: A novel method using particle-packing technique to fabricate capillary electrophoresis (CE)-based immobilized enzyme reactor (IMER) was accomplished by utilizing perfusive silica single particles as the frits and large-pore beads as the enzyme supports. The fabrication procedure is rapid and simple; the length and enzyme loading amount of the CE-IMERs could be easily adjusted. Performance and feasibility of the CE-IMERs were investigated using on-line trypsin digestion as the model enzyme reaction. High reproducible on-line enzyme assay was demonstrated with RSD less than 4.1% and 3.8% for peak area and migration time of the substrate and product over 100 consecutive runs, respectively. The enzyme can still maintain the activity for at least 10 days, indicating remarkably stability of the CE-IMERs. The CE-IMERs were successfully applied for accurate analysis of trypsin inhibition as well as on-line digestion of standard proteins (myoglobin and BSA). The present method provides a new interesting alternative to open-tubular and monolithic CE-IMERs, thus expands the application of the CE technique for on-line enzyme assay and analysis and characterization of peptides and proteins.
authors.affiliation: Faculty of Chemistry, Northeast Normal University, Changchun 130024, Jilin, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of Chemistry and the MOE Key Laboratory of Spectrochemical Analysis and Instrumentation, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Faculty of Chemistry, Northeast Normal University, Changchun 130024, Jilin, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: Faculty of Chemistry, Northeast Normal University, Changchun 130024, Jilin, China.
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Shi
authors.affiliation: Faculty of Chemistry, Northeast Normal University, Changchun 130024, Jilin, China.
authors.forename: Liping
authors.initials: L
authors.lastname: Guo
authors.affiliation: Faculty of Chemistry, Northeast Normal University, Changchun 130024, Jilin, China. Electronic address: yangl330@nenu.edu.cn.
authors.forename: Li
authors.initials: L
authors.lastname: Yang
mesh_terms: D019075:Electrophoresis, Capillary / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D009211:Myoglobin / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D012822:Silicon Dioxide / Q000737:chemistry; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism
chemical_list: D004800:Enzymes, Immobilized; D009211:Myoglobin; D011506:Proteins; D012822:Silicon Dioxide; D014357:Trypsin
keywords: Capillary electrophoresis; Enzyme microreactor; Particle packing; Trypsin digestion