Editor-in-Chief Hatice Kübra Elçioğlu Vice Editors Levent Kabasakal Esra Tatar Online ISSN 2630-6344 Publisher Marmara University Frequency Bimonthly (Six issues / year) Abbreviation J.Res.Pharm. Former Name Marmara Pharmaceutical Journal
Journal of Research in Pharmacy 2022 , Vol 26 , Issue 1
Analysis of 3-hydroxyisovaleric acid and 3-hydroxybutyric acid in plasma samples by LC-MS/MS
Tuba REÇBER1,Ece ÖZKAN1,Emirhan NEMUTLU1,Mehmet Sinan BEKSAC3,Sedef KIR1
1Department of Analytical Chemistry, Faculty of Pharmacy, Hacettepe University, 06100 Sıhhıye, Ankara, Turkey
2Department of Analytical Chemistry, Faculty of Pharmacy, Başkent University, 06100, Ankara, Turkey
3Department of Obstetrics and Gynecology, Division of Perinatology, Faculty of Medicine, Hacettepe University, 06100 Sıhhıye, Ankara, Turkey
DOI : 10.29228/jrp.110 Down syndrome is a common genetic disorder that results from the presence of an extra chromosome in the 21st chromosome pair of humans. Metabolomics is an alternative method in discovery of new biomarkers for the screening and diagnosis of Down syndrome. In this study, quantitative analyzes of 3-hydroxybutyric acid and 3-hydroxyisovaleric acid, selected as possible markers for prenatal diagnosis of Down syndrome were performed. LC-MS/MS analyzes were performed on a Phenomenex Luna NH2 (100 x 4.6 mm, 3 μm) column using a mobile phase mixture of 0.1% formic acid and acetonitrile containing 0.1% formic acid at a flow rate of 0.35 mL/minute. The MRM transitions were 103.0→59.0 for 3-hydroxybutyric acid and 117.1→59.0 for 3-hydroxyisovaleric acid. Under these conditions, the retention times of 3-hydroxyisovaleric acid 3-hydroxybutyric acid were 2.7 and 3.1 minute, respectively. The method was found linear from 0.1 to 10.0 μg/mL for both metabolites. The limit of detection (LOD) was 0.017 μg/mL for 3-hydroxybutyric acid and 0.003 μg/mL for 3-hydroxyisovaleric acid. The lower limit quantification (LLOQ) was 0.045 μg/mL for 3-hydroxybutyric acid and 0.008 μg/mL for 3-hydroxyisovaleric acid. The method has been proven to be selective, precise, accurate, sensitive, and robust based on the validation studies results. Finally, the method was applied to plasma samples of the pregnant women with healthy fetus (n = 30) and with Down syndrome fetus (n = 17). As a result of the analysis, a statistically significant increase (p <0.01) was observed in the 3-hydroxybutyric acid level of the group with Down syndrome compared to the healthy group. This result strengthens the use of 3-hydroxybutyric acid as an important biomarker in the prenatal screening/diagnosis of Down syndrome. Keywords : Down syndrome; prenatal screening; metabolomics; liquid chromatography-tandem mass spectrometry (LC-MS/MS); 3-hydroxyisovaleric acid; 3-hydroxybutyric acid
Marmara University