Analytical approach for the determination of steroid profile of humans by gas chromatography isotope ratio mass spectrometry aimed at distinguishing between endogenous and exogenous steroids.

The contamination of commonly used supplements by unknown steroids as well as their metabolites (parent compounds) become a challenge for the analytical laboratories. Although the determination of steroids profile is not trivial because of the complex matrix and low concentration of single compound, one of the most difficult current problem is to distinguish, during analytical procedure, endogenous androgens such as testosterone, dehydrotestosterone or dehydroepiandrosterone from their synthetic equivalents. The aim of this work was to develop and validate an analytical procedure for determination of the steroid profile in human urine by gas chromatography-combustion-isotope ratio mass spectrometry (GC/C/IRMS) toward distinguishing between endogenous and exogenous steroids. Beside the optimization of the experimental parameters for gas chromatography separation and mass spectrometry, attention was focused on urine sample preparation. Using an optimized sample preparation protocol it was possible to achieve better chromatographic resolutions and better sensitivity enabling the determination of 5 steroids, androsterone, etiocholanolone, testosterone, 5-androstandiol, 11-hydroxyandrdostane, pregnandiol, with the expanded uncertainty (k=2) below 1‰. This enable to evaluate the significant shift of the δ(13)C/(12)C [‰] values for each of examined steroids (excluding ERC). The analytical protocol described in this work was successfully used for the confirmation of positive founding urine by evaluation T/E ratio after GC/C/IRMS analysis.

Detection of TT virus (TTV) in children and youth with chronic viral hepatitis B and C.

BACKGROUND: TTV is a newly discovered virus and little is known about the frequency of TTV infection in children in Poland. The aim of our study was to investigate the frequency of TTV infection in children with chronic viral hepatitis B and C. MATERIAL/METHODS: Two patient groups were tested: 74 patients with chronic hepatitis B aged 4 to 20 years, and 13 patients with chronic hepatitis C aged 10 to 18 years. Nucleic acids were extracted from serum using the spin column technique (QIAGEN(r), Hilden, Germany), and TTV DNA sequences were amplified by nested PCR. RESULTS: TTV DNA was found in 47.3% of patients with chronic hepatitis B and in 53.8% of patients with hepatitis C. Among those patients with chronic B hepatitis there was no statistical difference between the frequency of coinfection with TTV and clinical-histopathological diagnosis, activity of aminotransaminases, frequency of seroconversion of HBeAg to antiHBe, or interferon alpha therapy. CONCLUSIONS: In Poland, TTV viremia is frequent in patients with chronic viral hepatitis B and C. TTV coinfection did not modify the course and activity of chronic hepatitis B or influence the outcome of interferon therapy.