Determination of thiopurine S-methyltransferase (TPMT) activity by comparing various normalization factors: reference values for Estonian population using HPLC-UV assay.
J Chromatogr B Analyt Technol Biomed Life Sci
; 834(1-2): 77-83, 2006 Apr 13.
Article
em En
| MEDLINE
| ID: mdl-16517227
ABSTRACT
Thiopurine S-methyltransferase (TPMT; EC 2.1.1.67) is the key enzyme in the metabolism of thiopurine drugs. Determination of TPMT activity has been used for the individualization of thiopurine dose. We developed HPLC-UV assay for the determination of TPMT activity in human erythrocytes using 6-mercaptopurine as a substrate. Various extraction and chromatographic conditions were compared. In-house developed extraction with acetonitrile provided the lowest limit of quantification. TPMT activity was determined in 99 previously genotyped healthy Estonians. TPMT activity was expressed as the formation of 6-methylmercaptopurine ng/ml/h and normalized either to haemoglobin, haematocrit, erythrocyte count or protein content. The receiver-operating characteristic curve analysis revealed similar accuracy values for TPMT activity in predicting heterozygous and wild type individuals for each method of calculation. In healthy Estonians, TPMT activity varied from 21.5 to 129.6 ng/ml/h. For heterozygous individuals (n = 18), TPMT activity was 48.1 +/- 11.7 ng/ml/h. Wild type individuals (n = 81) revealed significantly higher TPMT activity 79.3 +/- 20.7 ng/ml/h (P < 0.001). This sensitive HPLC assay for quantitative determination of TPMT activity could easily be used in clinical settings. Under constant experimental conditions for haemolysate preparation no normalization is required.
Buscar no Google
Base de dados:
MEDLINE
Assunto principal:
Espectrofotometria Ultravioleta
/
Cromatografia Líquida de Alta Pressão
/
Genética Populacional
/
Metiltransferases
Tipo de estudo:
Prognostic_studies
Limite:
Humans
País/Região como assunto:
Europa
Idioma:
En
Ano de publicação:
2006
Tipo de documento:
Article