Incomplete dialysis of protein samples containing 3-[(3-chlolamidopropyl)dimethylammonio]-1-propanesulfonate may lead to erroneous estimation of histidine content on amino acid analysis.
The zwitterionic detergents Chaps, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, and its 2-hydroxyl derivative Chapso have been widely used in the biomembrane field as well as other fields for their nondenaturing character. Due to their structure containing an amide bond in the molecules center they are prone to hydrolysis with 6 N HCl into cholic acid and 3-aminopropyldimethylammonio-1-propanesulfonate (APS) and its 2-hydroxyl derivative (APSO), respectively. On amino acid analyses with the ninhydrin detection system, His, APS, and APSO were eluted at 24.42, 24.55, and 24.85 min, respectively, and APS was identified as His due to their close retention times. Moreover, the mixture of His and APS coeluted as a single peak at 24.51 min leading to an erroneous His content for Chaps-contaminated protein samples. For full separation of APS, APSO, and His the elution program was improved by an additional 5-min elution with Buffer 7, i.e., 1:1 mixture of Buffers 3 and 4, before elution with Buffer 4, enabling complete separation of the 18 amino acids, APS, and APSO in a single run.
Nomura
K
K
Department of Biochemistry, Tokyo Metropolitan Institute of Gerontology, Tokyo, 173-0015, Japan.
D002793:Cholic Acids / Q000737:chemistry*; D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D003956:Dialysis / Q000379:methods*; D006639:Histidine / Q000032:analysis*; D015394:Molecular Structure
D002793:Cholic Acids; D006639:Histidine; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate