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Analytical Methods for Melatonin Quantification: Advances, Challenges, and Clinical Applications.

Mihaela Butiulca1,2, Lenard Farczadi3, Mihaly Veres4

  • 1Department of Anesthesiology and Intensive Care Medicine, Faculty of General Medicine, George Emil Palade University of Medicine, Pharmacy, Science, and Technology of Targu Mures, 540142 Targu Mures, Romania.

Pharmaceuticals (Basel, Switzerland)
|March 28, 2026
PubMed
Summary
This summary is machine-generated.

Quantifying melatonin, vital for sleep and circadian rhythms, is difficult due to low concentrations. This review compares methods, highlighting liquid chromatography-tandem mass spectrometry (LC-MS/MS) as precise but costly, with biosensors emerging as promising alternatives.

Keywords:
LC–MS/MSanalytical techniquescircadian biomarkersmelatoninpharmacology

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Area of Science:

  • Biochemistry and Analytical Chemistry
  • Chronobiology and Sleep Medicine

Background:

  • Melatonin, an indoleamine, regulates critical physiological processes like circadian rhythms and sleep-wake cycles.
  • Its presence in biological fluids at picogram or femtogram levels presents significant analytical quantification challenges.
  • Accurate melatonin measurement is essential for clinical diagnostics, pharmacological research, and understanding circadian rhythm disorders.

Purpose of the Study:

  • To critically assess and compare current analytical methodologies for melatonin determination in diverse biological matrices.
  • To evaluate the strengths and limitations of various techniques, including immunoassays, spectrophotometry, chromatography-mass spectrometry, and biosensors.
  • To guide researchers and clinicians in selecting optimal analytical approaches for melatonin quantification.

Main Methods:

  • Narrative review synthesizing data from published literature on melatonin quantification techniques.
  • Comparative analysis of methods based on sensitivity, specificity, reproducibility, cost, and clinical applicability.
  • Discussion of established techniques (immunoassays, LC-MS/MS) and emerging technologies (biosensors, microextraction).

Main Results:

  • Immunoassays and colorimetric methods offer cost-effective, accessible options for preliminary or large-scale studies.
  • Liquid chromatography-tandem mass spectrometry (LC-MS/MS) provides high sensitivity (sub-picogram limits) and specificity, serving as a benchmark for reference analysis.
  • Emerging biosensors and sample preparation techniques (e.g., dispersive microextraction) show potential for developing portable, sustainable, and sensitive analytical platforms.

Conclusions:

  • No single method is universally optimal; the choice depends on specific application requirements (e.g., cost, throughput, required sensitivity).
  • LC-MS/MS remains the gold standard for precise melatonin quantification but faces challenges in cost and accessibility.
  • Advancements in biosensor technology and sample preparation herald a new era of more accessible and potentially point-of-care melatonin analysis.