title: High-throughput analysis of testosterone in serum samples by on-line solid phase extraction liquid chromatography-tandem mass spectrometry.
abstract: BACKGROUND
  Liquid chromatography tandem-mass spectrometer (LC-MS/MS) techniques are more and more common in the measurement of testosterone concentrations in biological samples. However, LC-MS/MS methods are more laborious than streamlined automated immunochemistry methods because of the need for tedious pre-purification of the sample before the mass spectrometric analysis. We therefore developed a robust and rapid sample clean-up method to improve the throughput of the whole LC-MS/MS analysis procedure by applying an automated on-line solid-phase extraction (SPE) technique instead of the still widely used conventional liquid-liquid extraction.
  
  
  METHODS
  Testosterone was purified by the on-line SPE-column-switching technique after rapid precipitation of serum samples by zinc sulphate/internal standard solution before LC-MS/MS analysis. The results were compared with those of our previous routine LC-MS/MS method using liquid-liquid extraction with tert-butyl methyl ether for the pre-purification of the samples.
  
  
  RESULTS
  The tested on-line SPE-LC-MS/MS method reached the specifications of the previous method with liquid-liquid extraction. The precision of the new method was notably better, especially in the lower concentration range, than with the former method; the total variation was below 10% in the whole quantitation range of 0.25-35 nmol/L. The new method liberates more than 50% of hands-on time of laboratory technicians as well as expensive instrument time for other applications compared with the older method.
  
  
  CONCLUSIONS
  The on-line SPE-pre-purification technique tested in long-term use offers a rapid and reliable technique in the LC-MS/MS analysis of serum testosterone and is a valuable tool in the improvement of efficiency in the laborious steroid analytics.
authors.affiliation: Department of Clinical Chemistry, University of Eastern Finland, Finland. kari.savolainen@islab.fi
authors.forename: Kari
authors.initials: K
authors.lastname: Savolainen
authors.forename: Raisa
authors.initials: R
authors.lastname: Kiimamaa
authors.forename: Toivo
authors.initials: T
authors.lastname: Halonen
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods*; D057166:High-Throughput Screening Assays; D006801:Humans; D059625:Liquid-Liquid Extraction; D008738:Methyl Ethers / Q000737:chemistry; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D044967:Serum / Q000737:chemistry*; D052616:Solid Phase Extraction / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013739:Testosterone / Q000097:blood*; D019287:Zinc Sulfate / Q000737:chemistry
chemical_list: D008738:Methyl Ethers; C043243:methyl tert-butyl ether; D013739:Testosterone; D019287:Zinc Sulfate