Assisted Reductive Amination for Quantitation of Tryptophan, 5-Hydroxytryptophan, and Serotonin by Ultraperformance Liquid Chromatography Coupled with Tandem Mass Spectrometry
Abstract
:1. Introduction
2. Results
2.1. Reductive Amination of Tryptophan and Its Metabolites
2.2. MS/MS Fragmentation and MRM Transition Setting
2.3. Signal Enhancements of Modified Serotonin, TRP, and 5-HTP
2.4. Method Validation
3. Materials and Methods
3.1. Materials
3.2. Instrumentation
3.3. Preparation of Derivatized Standards and Internal Standards
3.4. Development of the Detection Method for Serotonin, 5-Hydroxytryptophan, and Tryptophan
3.5. Evulation of the Derivitized Method
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Sample Availability
References
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Name | Molecular Formula | Molecular Weight (Da) | MRM Transitions | LOD # (ng/mL) | LOQ # (ng/mL) | Coefficient of Determination (R2) | |
---|---|---|---|---|---|---|---|
Precursor Ion | Product Ions | ||||||
Serotonin | C10H12N2O | 176.22 | 177 | 115, 160 | --- | --- | --- |
h2-modified serotonin | C12H16N2O | 204.27 | 205 | 58, 160 | 2.35 | 14.54 | 0.9969 |
d2-modified serotonin | C12H12D4N2O | 208.29 | 209 | 62, 160 | --- | --- | --- |
Tryptophan | C11H12N2O2 | 204.23 | 205 | 118, 146 | --- | --- | --- |
h2-modified tryptophan | C13H16N2O2 | 232.28 | 233 | 102, 118, 146 | 1.96 | 4.36 | 0.9959 |
d2-modified tryptophan | C13H12D4N2O2 | 236.30 | 237 | 106, 118, 146 | --- | --- | --- |
5-HTP | C11H12N2O3 | 220.22 | 221 | 134, 162 | --- | --- | --- |
h2-modified 5-HTP | C13H16N2O3 | 248.28 | 249 | 134, 162 | 1.39 | 15.36 | 0.9938 |
d2-modified 5-HTP | C13H12D4N2O3 | 252.30 | 253 | 134, 162 | --- | --- | --- |
MRM/Average Peak Area (n = 4) | |||||
---|---|---|---|---|---|
Unmodified | RSD (%) | d2-Modified | RSD (%) | Enhancement | |
Serotonin | 55,030,586 | 4.2 | 65,536,481 | 4.5 | 1.19-fold |
Tryptophan | 6,765,226 | 3.8 | 127,646,745 | 1.2 | 18.87-fold |
5-HTP | 2,193,503 | 6.3 | 5,257,500 | 2.4 | 2.40-fold |
Serotonin | Tryptophan | 5-HTP | ||||
---|---|---|---|---|---|---|
Concentration (ng/mL) | Intra-Day | Inter-Day | Intra-Day | Inter-Day | Intra-Day | Inter-Day |
500 | 3.5–6.9% | 4.0% | 1.2–3.8% | 6.4% | 0.6–6.1% | 6.7% |
100 | 2.2–6.4% | 3.0% | 1.6–8.6% | 7.9% | 1.7–9.7% | 5.7% |
50 | 3.4–3.9% | 9.7% | 2.0–10.8% | 3.7% | 4.1–16.3% | 6.1% |
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Liang, S.-S.; Shen, P.-T.; Liang, Y.-Q.; Ke, Y.-W.; Cheng, C.-W.; Lin, Y.-R. Assisted Reductive Amination for Quantitation of Tryptophan, 5-Hydroxytryptophan, and Serotonin by Ultraperformance Liquid Chromatography Coupled with Tandem Mass Spectrometry. Molecules 2023, 28, 4580. https://doi.org/10.3390/molecules28124580
Liang S-S, Shen P-T, Liang Y-Q, Ke Y-W, Cheng C-W, Lin Y-R. Assisted Reductive Amination for Quantitation of Tryptophan, 5-Hydroxytryptophan, and Serotonin by Ultraperformance Liquid Chromatography Coupled with Tandem Mass Spectrometry. Molecules. 2023; 28(12):4580. https://doi.org/10.3390/molecules28124580
Chicago/Turabian StyleLiang, Shih-Shin, Po-Tsun Shen, Yu-Qing Liang, Yi-Wen Ke, Chieh-Wen Cheng, and Yi-Reng Lin. 2023. "Assisted Reductive Amination for Quantitation of Tryptophan, 5-Hydroxytryptophan, and Serotonin by Ultraperformance Liquid Chromatography Coupled with Tandem Mass Spectrometry" Molecules 28, no. 12: 4580. https://doi.org/10.3390/molecules28124580
APA StyleLiang, S. -S., Shen, P. -T., Liang, Y. -Q., Ke, Y. -W., Cheng, C. -W., & Lin, Y. -R. (2023). Assisted Reductive Amination for Quantitation of Tryptophan, 5-Hydroxytryptophan, and Serotonin by Ultraperformance Liquid Chromatography Coupled with Tandem Mass Spectrometry. Molecules, 28(12), 4580. https://doi.org/10.3390/molecules28124580