A comparison of methods for determining total body protein.
The aim of the study was to find the optimal method (with respect to convenience and accuracy) for determining total protein in whole-body homogenates of rats. Three different protein extraction methods and five different protein concentration methods were assessed. The results were compared against a reference value measured by complete amino acid analysis after acid hydrolysis. The data demonstrated that extraction with 5% (w/v) sodium dodecylsulfate (SDS) in 0.5 N NaOH was far superior to that with water alone or to 6 N guanidine-HCl. A comparison of the Biuret, Bradford, and bicinchoninic acid methods on the SDS-NaOH-extracted samples showed that the Biuret method was optimal, giving a value that was 90% of the reference value with a small variation (2.4% of the mean). The Kjeldahl method gave the correct protein concentration only when a nitrogen factor of 5.51 +/- 0.03 (N = 5) was applied. The results suggest that extraction with SDS-NaOH followed by the Biuret procedure is a good method for measuring protein concentrations in whole body rat homogenates.
Brooks
S P
SP
Nutrition Research Division, Food Directorate Health Canada, Tunney's Pasture, Ottawa, Ontario.
Lampi
B J
BJ
Sarwar
G
G
Botting
H G
HG
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001738:Biuret Reaction; D001824:Body Constitution; D002364:Caseins / Q000032:analysis; D002457:Cell Extracts / Q000737:chemistry; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D019791:Guanidine; D006146:Guanidines; D008297:Male; D009555:Ninhydrin; D011506:Proteins / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000032:analysis; D012967:Sodium Dodecyl Sulfate
D000596:Amino Acids; D002364:Caseins; D002457:Cell Extracts; D006146:Guanidines; D011506:Proteins; D012710:Serum Albumin, Bovine; D012967:Sodium Dodecyl Sulfate; D009555:Ninhydrin; D019791:Guanidine