Capillary electrophoresis analysis of orange juice pectinesterases.
Pectinesterase (PE) was extracted from orange juice and pulp with 1 M NaCl, desalted, and separated using capillary electrophoresis (CE) gel procedures (CE-SDS-CGE) and isoelectric focusing (CE-IEF). PE resolved as a single peak using noncoated fused silica columns with CE-SDS-CGE. CE-IEF separation of PE required acryloylaminoethoxyethanol-coated columns, which had limited stability. Thermal stability of PE extracts before and after heating at 75 degrees C for 30 min and at 95 degrees C for 5 min established heat labile and heat stabile fractions with identical PE migration times by CE-SDS-CGE or CE-IEF. Peak magnitude decreased to a constant value as heating time increased at 75 degrees C. Regression analysis of CE-SDS-CGE peak migration times of molecular weight (MW) standards estimated both heat labile and heat stable PE at MW approximately 36 900. Traditional SDS-PAGE gel separation of MW standards and active PE isolated by IEF allowed estimation of MW approximately 36 000. CE-SDS-CGE allowed presumptive, but not quantitative, detection of active PE in fresh juice.
University of Florida, Citrus Research and Education Center, 700 Experiment Station Road, Lake Alfred, Florida 33850, USA. rjbk@lal.ufl.edu
R J
RJ
Braddock
C R
CR
Bryan
J K
JK
Burns
D001628:Beverages / Q000032:analysis*; D002265:Carboxylic Ester Hydrolases / Q000032:analysis*; D002957:Citrus / Q000201:enzymology*; D019075:Electrophoresis, Capillary / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D007202:Indicators and Reagents; D007525:Isoelectric Focusing / Q000379:methods
D007202:Indicators and Reagents; D002265:Carboxylic Ester Hydrolases; C022323:pectinesterase