Analysis of Catecholamines and Metanephrines in Urine
Applications | 2017 | ShimadzuInstrumentation
Catecholamines and their metabolites, metanephrines, are critical biomarkers for disorders of the autonomic nervous system and neuroendocrine tumors such as pheochromocytoma. Precise urinary quantitation supports clinical diagnosis, therapeutic monitoring and research into neurotransmitter regulation.
The aim of this study was to develop and validate a rapid, sensitive and reproducible LC-MS/MS method using the Shimadzu LCMS-8060 for simultaneous measurement of epinephrine, norepinephrine, dopamine, metanephrine, normetanephrine and 3-methoxytyramine in urine.
The sample preparation employed automated solid-phase extraction (SPE) on a Biotage EVOLUTE EXPRESS WCX 96-well plate using a Biotage Extrahera system. Urine aliquots were spiked with isotopically labeled internal standards (except for 3-methoxytyramine) and pretreated with ammonium acetate. Interferences were washed away with ammonium acetate and isopropyl alcohol, then analytes eluted in acidified water:isopropanol.
Chromatographic separation was achieved on an ACE-Excel C18-PFP column (0.25 mM ammonium formate in water and methanol mobile phases) at 0.4 mL/min and a 7.5-minute run time. Detection utilized the Shimadzu LCMS-8060 triple quadrupole with MRM transitions optimized for each analyte.
Calibration curves for all six compounds demonstrated excellent linearity (R2 ≥ 0.99) across clinically relevant ranges: metanephrines 0.125–125 ng/mL, 3-methoxytyramine 0.25–125 ng/mL, dopamine 0.625–125 ng/mL, epinephrine 0.5–250 ng/mL and norepinephrine 2.5–250 ng/mL. Accuracy at nine concentration levels ranged from 91% to 117%. Repeatability at the LOQ yielded %RSD values below 14% for all analytes. Representative chromatograms at the LOQ confirmed well-resolved peaks and minimal matrix interference.
The validated LCMS-8060 method provides a fast, sensitive and reliable platform for quantifying catecholamines and metanephrines in urine. Its robust performance meets the demands of clinical laboratories and research facilities seeking accurate assessment of neuroendocrine biomarkers.
Shimadzu Scientific Instruments. SSI-LCMS-086: Analysis of Catecholamines and Metanephrines in Urine. June 2017.
LC/MS, LC/MS/MS, LC/QQQ
IndustriesClinical Research
ManufacturerShimadzu
Summary
Significance of the Topic
Catecholamines and their metabolites, metanephrines, are critical biomarkers for disorders of the autonomic nervous system and neuroendocrine tumors such as pheochromocytoma. Precise urinary quantitation supports clinical diagnosis, therapeutic monitoring and research into neurotransmitter regulation.
Objectives and Study Overview
The aim of this study was to develop and validate a rapid, sensitive and reproducible LC-MS/MS method using the Shimadzu LCMS-8060 for simultaneous measurement of epinephrine, norepinephrine, dopamine, metanephrine, normetanephrine and 3-methoxytyramine in urine.
Methodology and Applied Instrumentation
The sample preparation employed automated solid-phase extraction (SPE) on a Biotage EVOLUTE EXPRESS WCX 96-well plate using a Biotage Extrahera system. Urine aliquots were spiked with isotopically labeled internal standards (except for 3-methoxytyramine) and pretreated with ammonium acetate. Interferences were washed away with ammonium acetate and isopropyl alcohol, then analytes eluted in acidified water:isopropanol.
Chromatographic separation was achieved on an ACE-Excel C18-PFP column (0.25 mM ammonium formate in water and methanol mobile phases) at 0.4 mL/min and a 7.5-minute run time. Detection utilized the Shimadzu LCMS-8060 triple quadrupole with MRM transitions optimized for each analyte.
Main Results and Discussion
Calibration curves for all six compounds demonstrated excellent linearity (R2 ≥ 0.99) across clinically relevant ranges: metanephrines 0.125–125 ng/mL, 3-methoxytyramine 0.25–125 ng/mL, dopamine 0.625–125 ng/mL, epinephrine 0.5–250 ng/mL and norepinephrine 2.5–250 ng/mL. Accuracy at nine concentration levels ranged from 91% to 117%. Repeatability at the LOQ yielded %RSD values below 14% for all analytes. Representative chromatograms at the LOQ confirmed well-resolved peaks and minimal matrix interference.
Benefits and Practical Applications
- High throughput: Automated SPE in 96-well format accelerates sample processing.
- Robust quantitation: Excellent linearity and precision enable reliable clinical diagnostics.
- Versatility: Single-run analysis of multiple neurotransmitters supports broader metabolic studies.
Future Trends and Opportunities
- Microflow and multiplexed LC-MS workflows to further reduce solvent consumption and enhance sensitivity.
- Integration with high-resolution mass spectrometry for improved specificity in complex matrices.
- Expanded panels combining catecholamines with additional biomarkers for comprehensive metabolic profiling.
- Point-of-care and miniaturized devices leveraging advances in ion mobility and microfluidics.
Conclusion
The validated LCMS-8060 method provides a fast, sensitive and reliable platform for quantifying catecholamines and metanephrines in urine. Its robust performance meets the demands of clinical laboratories and research facilities seeking accurate assessment of neuroendocrine biomarkers.
Reference
Shimadzu Scientific Instruments. SSI-LCMS-086: Analysis of Catecholamines and Metanephrines in Urine. June 2017.
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