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.
Braddock
R J
RJ
University of Florida, Citrus Research and Education Center, 700 Experiment Station Road, Lake Alfred, Florida 33850, USA. rjbk@lal.ufl.edu
Bryan
C R
CR
Burns
J K
JK
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