Highly specific ID-UHPLC-MS/MS method for analyzing polar and non-polar steroid hormones

Area of Science:

• Endocrinology
• Analytical Chemistry
• Biomarker Discovery

Background:

• Accurate measurement of endogenous steroid hormones is crucial for diagnosing endocrine and metabolic diseases.
• Existing methods may lack the precision or range for diverse patient populations.
• Steroid hormone profiles serve as vital biomarkers in clinical care and translational research.

Purpose of the Study:

• To develop and validate a novel, highly accurate, and precise isotope-dilution liquid chromatography-mass spectrometry (LC-MS/MS) method.
• To simultaneously measure eight clinically relevant steroid hormones in a small serum volume (200 µL).
• To establish a method suitable for analyzing steroid hormone levels across the general population, including children and postmenopausal women.

Main Methods:

• Developed a sequential liquid-liquid extraction technique for isolating eight polar and non-polar steroid hormones without derivatization or hydrolysis.
• Employed isotope-dilution LC-MS/MS for simultaneous quantification of total testosterone (TT), estradiol (E2), progesterone (P4), 17-hydroxyprogesterone (17-OHP), androstenedione (AD), estrone (E1), estrone sulfate (E1S), and dehydroepiandrosterone sulfate (DHEAS).
• Validated the method's analytical measurement range, selectivity, sensitivity, imprecision (CV: 3.5–10.7%), and bias using reference materials and 268 serum samples.

Main Results:

• The developed LC-MS/MS method accurately and precisely measures eight key steroid hormones simultaneously from 200 µL of serum.
• The method exhibits a broad analytical measurement range, high sensitivity, and low imprecision, suitable for diverse populations.
• Analysis of 268 serum samples showed high success rates, with only 2% of results below the limit of detection.

Conclusions:

• This new steroid hormone panel LC-MS/MS method provides a robust tool for clinical diagnostics and translational research.
• It facilitates comprehensive investigations into the associations between steroid profiles and various health disorders.
• The method's suitability for general population analysis supports individualized health evaluations and population-level studies.