Linked Life Data

14508388

Source:http://linkedlifedata.com/resource/pubmed/id/14508388

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PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
2003-9-25
pubmed:abstractText
Genetic polymorphism of the S-methylation pathway catalyzed by thiopurine methyltransferase (TPMT) is responsible for variation in the metabolism, toxicity, and therapeutic efficacy of thiopurine drugs. This paper describe a new simple, nonradioactive HPLC method for determination of TPMT activity in isolated erythrocytes (Ery), based on the conversion of 6-mercaptopurine (pH 7.5, 37 degrees C) to 6-methylmercaptopurine (6-MMP) using S-adenosyl-l-methionine as methyl donor. The incubation step was stopped by a mixture of trichloroacetic acid/acetonitrile containing the internal standard 4-aminoacetophenone. 6-MMP was quantified by absorbance at 290 nm after chromatographic separation on a Zorbax SB-Phenyl column (5 microm, 4.6 x 250 mm) using mobile phases (flow rate 1.1 mL/min) consisting of acetonitrile, phosphate buffer pH 3.0, triethylamine, and dithiothreitol. The assay was linear up to 50 nmol/(mL Ery. h), and the detection limit was 0.3 nmol/(mL Ery. h). The extraction efficiency of 6-MMP was 95-103% (n = 3), and its analytic recovery ranged between 98.3% and 101.8% (n = 12). The within-day imprecision using pooled human erythrocytes (n = 12) was 4.4% at a TPMT activity of 14.3 nmol/(mL Ery.h) and 4.9% at 6.5 nmol/(mL Ery.h). The between-day imprecision (n = 12) was 6.8% and 7.5% nmol/(mL Ery.h), respectively. A very good agreement was found between TPMT activity determined with this method (y) and a widely used radiochemical procedure (x) (r = 0.94; n = 130; y = 0.502 + 0.946x; P < 0.05). Genotype analysis of all individuals with TPMT activity under 12.5 nmol/(mL Ery.h) revealed a genotype/phenotype concordance of 86%. The new HPLC method for determination of TPMT activity in Ery is a simple, rapid, and reliable nonradioactive procedure that can be successfully used for both research and routine clinical analysis.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0163-4356
pubmed:author
pubmed:issnType
Print
pubmed:volume
25
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
637-44
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2003
pubmed:articleTitle
Determination of thiopurine methyltransferase phenotype in isolated human erythrocytes using a new simple nonradioactive HPLC method.
pubmed:affiliation
Department of Clinical Laboratory, Medical University Sofia, Sofia, Bulgaria. dindjova@yahoo.com
pubmed:publicationType
Journal Article