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Characterization of oat proteins and aggregates using asymmetric flow field-flow fractionation.

Runyon, Ray LU ; Nilsson, Lars LU ; Alftrén, Johan LU and Bergenståhl, Björn LU (2013) In Analytical and Bioanalytical Chemistry 405(21). p.6649-6655
Abstract
The soluble proteins and protein aggregates in Belinda oats were characterized using asymmetric flow field-flow fractionation (AF4) coupled with online UV-vis spectroscopy and multiangle light-scattering detection (MALS). Fractions from the AF4 separation were collected and further characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The AF4 fractogram of the oat extracts revealed three peaks which were determined to be monomeric forms of soluble proteins, globulin aggregates, and β-glucan, respectively. The early eluting monomeric proteins ranged in molar mass (MM) between 5 and 90 kg/mol and in hydrodynamic diameter (D h) from 1.6 to 13 nm. The MM at peak maximum of the globulin aggregate peak was found... (More)
The soluble proteins and protein aggregates in Belinda oats were characterized using asymmetric flow field-flow fractionation (AF4) coupled with online UV-vis spectroscopy and multiangle light-scattering detection (MALS). Fractions from the AF4 separation were collected and further characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The AF4 fractogram of the oat extracts revealed three peaks which were determined to be monomeric forms of soluble proteins, globulin aggregates, and β-glucan, respectively. The early eluting monomeric proteins ranged in molar mass (MM) between 5 and 90 kg/mol and in hydrodynamic diameter (D h) from 1.6 to 13 nm. The MM at peak maximum of the globulin aggregate peak was found to be ∼300 kg/mol and the D h was measured to be ∼20 nm. SDS-PAGE of the collected fraction across this peak revealed two bands with MM of 37 and 27 kg/mol which correspond to the α and β subunits of globulin indicating the elution of globulin aggregates. A third peak at long retention time was determined to be β-glucan through treatment of the oat extract with β-glucanase and by injection of β-glucan standards. The amount of soluble protein was measured to be 83.1 ± 2.3 wt.%, and the amount of albumin proteins was measured to be 17.6 ± 5.7 wt.% of the total protein in the oats. The results for Belinda oat extracts show that the AF4-MALS/UV platform is capable of characterizing the physicochemical properties such as MM and hydrodynamic size distribution of proteins and protein aggregates within a complicated food matrix environment and without the need to generate protein isolates. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Analytical and Bioanalytical Chemistry
volume
405
issue
21
pages
6649 - 6655
publisher
Springer
external identifiers
  • wos:000322705600005
  • pmid:23812878
  • scopus:84881367384
ISSN
1618-2642
DOI
10.1007/s00216-013-7115-7
language
English
LU publication?
yes
id
5417e380-a6dc-4eeb-90b4-f808f5a3adf7 (old id 3956336)
date added to LUP
2016-04-01 09:49:26
date last changed
2020-01-05 03:09:56
@article{5417e380-a6dc-4eeb-90b4-f808f5a3adf7,
  abstract     = {The soluble proteins and protein aggregates in Belinda oats were characterized using asymmetric flow field-flow fractionation (AF4) coupled with online UV-vis spectroscopy and multiangle light-scattering detection (MALS). Fractions from the AF4 separation were collected and further characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The AF4 fractogram of the oat extracts revealed three peaks which were determined to be monomeric forms of soluble proteins, globulin aggregates, and β-glucan, respectively. The early eluting monomeric proteins ranged in molar mass (MM) between 5 and 90 kg/mol and in hydrodynamic diameter (D h) from 1.6 to 13 nm. The MM at peak maximum of the globulin aggregate peak was found to be ∼300 kg/mol and the D h was measured to be ∼20 nm. SDS-PAGE of the collected fraction across this peak revealed two bands with MM of 37 and 27 kg/mol which correspond to the α and β subunits of globulin indicating the elution of globulin aggregates. A third peak at long retention time was determined to be β-glucan through treatment of the oat extract with β-glucanase and by injection of β-glucan standards. The amount of soluble protein was measured to be 83.1 ± 2.3 wt.%, and the amount of albumin proteins was measured to be 17.6 ± 5.7 wt.% of the total protein in the oats. The results for Belinda oat extracts show that the AF4-MALS/UV platform is capable of characterizing the physicochemical properties such as MM and hydrodynamic size distribution of proteins and protein aggregates within a complicated food matrix environment and without the need to generate protein isolates.},
  author       = {Runyon, Ray and Nilsson, Lars and Alftrén, Johan and Bergenståhl, Björn},
  issn         = {1618-2642},
  language     = {eng},
  number       = {21},
  pages        = {6649--6655},
  publisher    = {Springer},
  series       = {Analytical and Bioanalytical Chemistry},
  title        = {Characterization of oat proteins and aggregates using asymmetric flow field-flow fractionation.},
  url          = {http://dx.doi.org/10.1007/s00216-013-7115-7},
  doi          = {10.1007/s00216-013-7115-7},
  volume       = {405},
  year         = {2013},
}