ABSTRACT
A micellar electrokinetic capillary chromatography (MEKC) method with a mixed micelle system for separation and measurement of desmosine (DES) and isodesmosine (IDES) in vascular tissue hydrolysates is described. The mixed micelle system was composed of a zwitterionic surfactant named 3-(N,N-dimethylhexadecylammonium)propanesulfonate (PAPS) and a nonionic surfactant polyethylene glycol dodecyl ether (Brij 35). By using 50 mM Tris-H(3)PO(4) (pH 2.5) containing 40 mM PAPS and 0.5% (m/v) Brij-35 as the optimal running buffer, DES and IDES were baseline separated from each other and from other hydrolyzed components of the vascular tissue. The limit of quantitation (LOQ) for DES and IDES were 3.00 x 10(-6)mol/L and 2.75 x 10(-6)mol/L, respectively. The relative standard deviation (RSD) of migration times and corrected peak area in terms of the inter-day and the intra-day repeatability were less than 1.7%. Hydrolysate samples of vascular tissues of rats were analyzed with the MEKC method with satisfied results.
Subject(s)
Aorta, Thoracic/chemistry , Chromatography, Micellar Electrokinetic Capillary/methods , Desmosine/isolation & purification , Isodesmosine/isolation & purification , Animals , Desmosine/analysis , Isodesmosine/analysis , Rats , Rats, Inbred SHR , Rats, Inbred WKYABSTRACT
Desmosine and isodesmosine were detected in an invertebrate molluscan species, i.e. in an insoluble protein in the hinge ligament of a bivalve species, Sakhalin surf clam (Pseudocardium sachalinensis, in family Mactridae). The protein is rich in glycine and methionine S-oxide but devoid of hydroxyproline and hydroxylysine. 3,3'-Methylenebistyrosine was also detected in the HCl hydrolysate of the hinge-ligament protein, but it was found to be an artefact produced from tyrosine and formaldehyde derived from methionine S-oxide during the HCl hydrolysis of the protein.
Subject(s)
Amino Acids/isolation & purification , Bivalvia/analysis , Desmosine/isolation & purification , Isodesmosine/isolation & purification , Ligaments/analysis , Animals , Cross-Linking Reagents , Protein Conformation , Proteins/analysis , Tyrosine/analogs & derivatives , Tyrosine/isolation & purificationABSTRACT
A rapid large scale procedure was devised for the purification of desmosine and isodesmosine from ligamentum nuchae elastin. The method makes use of the hydrophilic nature of the desmosines which preferentially absorbs to cellulose fibers in mixtures of organic solvents. Resolution of the isomers was achieved on a polystyrene resin column.
