Green and Emerging Microextraction Strategies for Bioanalytical Determination of Hormones: Trends, Challenges, and Applications

Accurate and sensitive determination of hormones in biological matrices is essential for
clinical diagnostics, therapeutic monitoring, and endocrine research. However, hormone
determination presents significant challenges due to their typically low concentrations,
complex sample matrices, and structural diversity. In recent years, microextraction techniques
have emerged as strategic tools in bioanalytical chemistry, offering advantages in
terms of miniaturization, enhanced selectivity, and compatibility with the principles of
green analytical chemistry (GAC). This review provides a comprehensive overview of
green and emerging microextraction approaches for the determination of steroidal, thyroid,
peptide, and other hormones in biological samples. Key techniques such as solid-phase microextraction
(SPME) and dispersive liquid–liquid microextraction (DLLME), followed by
high-performance liquid chromatography (HPLC) coupled to diode array detectors (DADs)
or mass spectrometry (MS), are critically discussed. Special emphasis is placed on the use of
environmentally friendly solvents, such as deep eutectic solvents (DESs), supramolecular
solvents (SUPRASs), and advanced sorbents including molecularly imprinted polymers
(MIPs) and nanostructured magnetic phases. Applications across various bioanalytical
matrices (urine, plasma, serum, saliva, tissues. . .) are examined in terms of sensitivity, selectivity,
and validation parameters. Finally, current challenges, method development gaps,
and future directions are highlighted to support the continued advancement of sustainable
hormone determination in complex biological systems.