Determination of Morphine and Its Metabolites in Human Urine by Capillary Electrophoresis with Laser Induced Fluorescence Detection Employing On-Column Labeling with a New Boronic Acid Functionalized Squarylium Cyanine Dye
Abstract
:1. Introduction
2. Materials and Methods
2.1. Reagents, Buffers, and Sample Solutions
2.2. Human Urine Sample Preparation
2.3. Instrumentation
Solvent or Buffer | SQ-BA3 Alone | SQ-BA3 with Morphine | SQ-BA3 with M3G | SQ-BA3 with M6G | |||||||
---|---|---|---|---|---|---|---|---|---|---|---|
λex (nm) | λem (nm) | λex (nm) | λem (nm) | % Fluorescence Enhancement | λex (nm) | λem (nm) | % Fluorescence Enhancement | λex (nm) | λem (nm) | % Fluorescence Enhancement | |
DMF | 631 | 661 | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A | N/A |
Ammonium Citrate (pH = 3.50) | 620 | 659 | 621 | 659 | 4.05% | 620 | 660 | 14.33% | 622 | 659 | 3.90% |
Tris-HCl (pH = 7.50) | 619 | 660 | 621 | 660 | 6.10% | 622 | 661 | 7.79% | 623 | 660 | 6.10% |
Ammonium Phosphate (pH = 9.50) | 618 | 657 | 619 | 658 | 18.20% | 621 | 659 | 23.45% | 621 | 659 | 6.20% |
Ammonium Phosphate (pH = 11.30) | 617 | 659 | 619 | 660 | 8.05% | 619 | 659 | 9.52% | 618 | 659 | 4.8% |
3. Results and Discussion
3.1. Spectral Properties of SQ-BA3 Alone and with Morphine, M3G, and M6G
3.2. Stability of SQ-BA3 Solutions
3.3. Optimization of CE-LIF Methods for Morphine and Its Metabolites
Figures of Merit | Morphine | M3G | M6G | |||
---|---|---|---|---|---|---|
Water Matrix | Diluted Urine Matrix | Water Matrix | Diluted Urine Matrix | Water Matrix | Diluted Urine Matrix | |
Linear Regression (Conc. vs. Peak Area) | y = (1 × 1010)x −1 × 106 | y = (2 × 1010)x −5 × 106 | y = (5 × 1010)x −7 × 106 | y = (3 × 1010)x −5 × 106 | y = (7 × 109)x −1 × 106 | y = (4 × 109)x −2 × 106 |
Correlation Coefficient, R2 | 0.9974 | 0.9977 | 0.9965 | 0.9962 | 0.9973 | 0.9950 |
% RSD | 5.34 | 4.47 | 5.35 | 4.61 | 4.46 | 5.89 |
Linearity range (M) | 2.50 × 10−4–2.50 × 10−3 | 3.50 × 10−4–1.75 × 10−3 | 1.50 × 10−4–1.50 × 10−3 | 2.50 × 10−4–1.25 × 10−3 | 5.00 × 10−4–2.00 × 10−3 | 1.00 × 10−3–1.75 × 10−3 |
LOD (M) | 1.20 × 10−4 | 1.10 × 10−4 | 8.70 × 10−5 | 9.30 × 10−5 | 1.10 × 10−4 | 1.80 × 10−4 |
LOQ (M) | 4.10 × 10−4 | 3.80 × 10−4 | 2.90 × 10−4 | 3.10 × 10−4 | 3.80 × 10−4 | 5.90 × 10−4 |
3.4. CE-LIF Analysis of Morphine and Its Metabolites in Human Urine Samples
4. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Sebaiy, M.M.; El-Shanawany, A.A.; Baraka, M.M.; Abdel-Aziz, L.M.; Isbell, T.A.; Colyer, C.L. Determination of Morphine and Its Metabolites in Human Urine by Capillary Electrophoresis with Laser Induced Fluorescence Detection Employing On-Column Labeling with a New Boronic Acid Functionalized Squarylium Cyanine Dye. Separations 2016, 3, 1. https://doi.org/10.3390/chromatography3010001
Sebaiy MM, El-Shanawany AA, Baraka MM, Abdel-Aziz LM, Isbell TA, Colyer CL. Determination of Morphine and Its Metabolites in Human Urine by Capillary Electrophoresis with Laser Induced Fluorescence Detection Employing On-Column Labeling with a New Boronic Acid Functionalized Squarylium Cyanine Dye. Separations. 2016; 3(1):1. https://doi.org/10.3390/chromatography3010001
Chicago/Turabian StyleSebaiy, Mahmoud M., Abdullah A. El-Shanawany, Mohamed M. Baraka, Lobna M. Abdel-Aziz, Theresa A. Isbell, and Christa L. Colyer. 2016. "Determination of Morphine and Its Metabolites in Human Urine by Capillary Electrophoresis with Laser Induced Fluorescence Detection Employing On-Column Labeling with a New Boronic Acid Functionalized Squarylium Cyanine Dye" Separations 3, no. 1: 1. https://doi.org/10.3390/chromatography3010001