A UHPLC-MS/MS method for profiling multifunctional steroids in human hair

Area of Science:

• Endocrinology and Metabolism
• Analytical Chemistry
• Forensic Science

Background:

• Steroid profiling is crucial for understanding physiological and pathological processes.
• Hair analysis offers a noninvasive, time-sequential method for long-term endogenous steroid hormone measurement.
• Existing liquid chromatography-mass spectrometry (LC-MS) methods face challenges in comprehensive hair steroid profiling, particularly for estrogens.

Purpose of the Study:

• To develop and validate an ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for simultaneous profiling of multiple steroid classes in human scalp hair.
• To establish an optimized sample preparation and analysis procedure for comprehensive steroid analysis in hair.
• To investigate potential gender-specific differences and longitudinal trends in hair steroid levels.

Main Methods:

• Developed a novel UHPLC-MS/MS method for simultaneous analysis of 28 steroids (corticosteroids, estrogens, androgens, progestogens) in a single run.
• Optimized sample preparation including hexane wash, 5 mm hair fragmentation, and 18-hour methanol incubation at 25°C.
• Employed methanol extraction with Girard P and dansyl chloride derivatization, followed by 25-minute analysis using automated injection with diverter valve switch and step analysis (AIDSA).

Main Results:

• The UHPLC-MS/MS method demonstrated excellent validation parameters including linearity, LOD, LOQ, precision, accuracy, matrix effect, and recovery.
• Successfully applied the method to profile steroids in both male and female human scalp hair samples.
• Observed significant gender-specific differences in steroid profiles and a declining trend of steroid levels from proximal to distal hair segments.

Conclusions:

• The developed UHPLC-MS/MS method provides a robust and comprehensive approach for simultaneous profiling of 28 multifunctional steroids in human hair.
• The findings highlight the utility of hair analysis for noninvasive, long-term steroid monitoring and reveal important gender-based variations.
• Awareness of the declining steroid concentration along hair length is crucial for accurate interpretation of hair steroid analysis results.