An improved anion-exchange high-performance liquid chromatography method for measuring oxidized form of LDLs in human plasma.
Ann Clin Biochem
; 47(Pt 5): 460-6, 2010 Sep.
Article
em En
| MEDLINE
| ID: mdl-20675341
ABSTRACT
BACKGROUND:
Circulating oxidized low-density lipoproteins (LDLs) (ox-LDLs) could be a sensitive marker to predict future cardiovascular events. However, a method to evaluate oxidized forms of LDLs systemically in human plasma is not yet established. In this study, we developed a novel and convenient high-performance liquid chromatography (HPLC) method for measuring ox-LDL levels in humans.METHODS:
Human plasma lipoproteins were separated by a modified HPLC method using a diethylaminoethyl-type anion-exchange gel column with stepwise elution. Ox-LDLs were detected by postcolumn reaction with a reagent containing cholesterol esterase and cholesterol oxidase. Particle size of each LDL fraction separated by HPLC was determined in 61 healthy subjects.RESULTS:
Our HPLC method separated LDLs into three fractions, which were designated as LDL-1, LDL-2 and LDL-3, on the basis of their negative charges, with LDL-3 the most strongly retained fraction migrating fastest in the anodic direction, a property that reflects the net negative charge of the molecule. Western blot analysis revealed that apolipoprotein B100 in LDL-3 fraction was the most fragmented and oxidatively modified. When LDLs were oxidized in vitro by Cu2+ or 2,2-azo-bis (2-aminopropane)-2HCl or modified by various aldehydes, all of the LDL fractions migrated at the position of LDL-3. Further, among three fractions, particle size was smallest in LDL-3 fraction.CONCLUSION:
Here, we developed a convenient HPLC method and identified LDL-3 as oxidized LDL fractions, although ox-LDLs were present in LDL-2 fraction, albeit lesser concentrations than in LDL-3 subfraction. Measuring ox-LDL levels in human plasma by this method may be useful to evaluate atherosclerotic disorders.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Análise Química do Sangue
/
Cromatografia Líquida de Alta Pressão
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Cromatografia por Troca Iônica
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Lipoproteínas LDL
Tipo de estudo:
Prognostic_studies
Limite:
Adult
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Female
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Humans
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Male
Idioma:
En
Revista:
Ann Clin Biochem
Ano de publicação:
2010
Tipo de documento:
Article