A Fast and Reliable UHPLC–MS/MS-Based Method for Screening Selected Pharmacologically Significant Natural Plant Indole Alkaloids
Abstract
:1. Introduction
2. Results
2.1. Extraction and Purification Procedure
2.2. MIA Analysis by Liquid Chromatography–Mass Spectrometry
2.3. Method Validation
2.3.1. Method Calibration and Linearity
2.3.2. Limit of Detection and Quantitation
2.3.3. Analytical Accuracy and Precision
2.3.4. Matrix Effect
2.4. MIA Determination in Tobbaco
2.5. Determination of MIAs in Tribulus terrestris
2.6. MIA Determination in Peganum harmala Seeds
3. Discussion
4. Materials and Methods
4.1. Plant Materials
4.2. Chemicals
4.3. Extraction and Isolation Procedure
4.4. Indole Alkaloid Analysis by Ultra-High Performance Liquid Chromatography–Tandem Mass Spectrometry (UHPLC–MS/MS)
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Sample Availability: Samples of the compounds are available from the authors. |
Compound | Retention Time (min) | Calibration Equation | Correlation Coefficient R2 | LOD (pg) | LOQ (pg) | Dynamic Range (pmol) |
---|---|---|---|---|---|---|
Harmaline | 2.13 ± 0.01 | log y = 0.9707 (log x) − 0.9819 | 0.9958 | 0.31 | 1.04 | 5–100 |
Harmine | 2.16 ± 0.01 | log y = 0.7374 (log x) − 0.2451 | 0.9935 | 0.18 | 0.60 | 5–100 |
Yohimbine | 2.98 ± 0.01 | log y = 1.0147 (log x) − 0.3091 | 0.9910 | 0.01 | 0.05 | 5–100 |
Ajmalicine | 4.94 ± 0.01 | log y = 0.8678 (log x) − 0.2006 | 0.9985 | 0.11 | 0.36 | 5–100 |
Compound | Determined Spiked Content (A) (pmol) | Analytical Precision (A) (%) | Analytical Accuracy (A) (%) | Determinedspiked Content (B) (pmol) | Analytical Precision (B) (%) | Analytical Accuracy (B) (%) |
---|---|---|---|---|---|---|
Harmaline | 125.5 ± 0.2 | 3.5 | 100.4 | 252.5 ± 1.4 | 2.5 | 101.0 |
Harmine | 123.8 ± 0.9 | 1.5 | 99.0 | 245.5 ± 0.2 | 2.2 | 98.2 |
Yohimbine | 125.0 ± 0.3 | 2.1 | 100.3 | 255.1 ± 0.9 | 2.9 | 102.0 |
Ajmalicine | 122.5 ± 1.9 | 2.4 | 98.0 | 244.2 ± 1.2 | 2.3 | 97.7 |
Sample Weight (mg) | Harmaline (pg·mg–1) | Harmine (pg·mg–1) | Yohimbine (μg·mg–1) | Ajmalicine (pg·mg–1) |
---|---|---|---|---|
5 | ND | 32.34 ± 3.69 | 5.29 ± 0.63 | 3.46 ± 0.92 |
10 | ND | 25.93 ± 2.79 | 4.01 ± 0.34 | 7.46 ± 1.24 |
15 | ND | 13.59 ± 3.11 | 3.31 ± 0.37 | 10.51 ± 1.28 |
Sample Weight (mg) | Matrix Factor (%) | |||
---|---|---|---|---|
Harmaline | Harmine | Yohimbine | Ajmalicine | |
5 | 76.57 ± 0.50 | 85.55 ± 0.16 | 94.05 ± 1.34 | 96.27 ± 1.12 |
10 | 69.20 ± 0.33 | 82.70 ± 0.30 | 74.74 ± 0.37 | 69.27 ± 0.50 |
15 | 58.68 ± 0.46 | 70.51 ± 0.21 | 65.48 ± 1.06 | 49.75 ± 0.48 |
Tissue | Alkaloid Content (pg·mg–1) | |||
---|---|---|---|---|
Harmaline | Harmine | Yohimbine | Ajmalicine | |
Flowers | 0.45 ± 0.04 | 0.53 ± 0.11 | 14.40 ± 1.68 | 0.0062 ± 0.0011 |
1st–3rd leaf | 0.38 ± 0.05 | 0.44 ± 0.15 | 7.76 ± 1.11 | 0.0111 ± 0.0010 |
4th leaf | 0.25 ± 0.09 | 0.59 ± 0.02 | 10.02 ± 1.36 | 0.0047 ± 0.0008 |
5th leaf | 0.41 ± 0.11 | 0.76 ± 0.09 | 12.38 ± 1.33 | 0.0135 ± 0.0002 |
6th leaf | 0.36 ± 0.05 | 0.70 ± 0.10 | 11.30 ± 1.61 | 0.0065 ± 0.0015 |
7th leaf | 0.19 ± 0.02 | 0.89 ± 0.04 | 14.65 ± 1.09 | 0.0080 ± 0.0016 |
8th leaf | 0.23 ± 0.06 | 0.07 ± 0.02 | 9.53 ± 0.72 | 0.0067 ± 0.0007 |
9th leaf | 0.23 ± 0.07 | 0.09 ± 0.01 | 14.12 ± 1.71 | 0.0048 ± 0.0011 |
Stem | 0.23 ± 0.07 | 0.07 ± 0.01 | 20.28 ± 1.22 | 0.0090 ± 0.0022 |
Roots | ND | 0.07 ± 0.02 | 8.04 ± 1.07 | 0.0056 ± 0.0021 |
Tissue | Alkaloid Content (pg·mg−1) | |||
---|---|---|---|---|
Harmaline | Harmine | Yohimbine | Ajmalicine | |
Burrs + stems (Bulgaria) | 0.012 ± 0.005 | ND | 36.14 ± 2.35 | ND |
Whole plant (China) | 0.004 ± 0.002 | 1.76 ± 0.18 | 62.74 ± 5.02 | ND |
Burrs only (China) | 0.004 ± 0.001 | ND | 43.21 ± 0.77 | ND |
Flower, burrs (India) | 0.005 ± 0.001 | 0.21 ± 0.05 | 57.59 ± 3.25 | ND |
Alkaloid content (pg·mg–1) | |||
---|---|---|---|
Harmaline | Harmine | Yohimbine | Ajmalicine |
19.76 ± 0.44 | 1096.67 ± 14.77 | 7.25 ± 0.83 | 0.24 ± 0.03 |
Instrumental Method Used | Amount of Tissue | Type of Tissue | Limit of Detection | Reference | Capacity of the Method Used |
---|---|---|---|---|---|
UHPLC–(+)ESI–MS/MS | 5 mg | whole plant | 0.01–0.31 pg | method presented | 4 IAs |
HPLC–UV | n.a. | whole plant | 0.77–56 mg·g−1 | [5] | 2 IAs |
HPLC–UV | 0.1 g | roots | 6–8 µg·mL−1 | [8] | 3 IAs |
HPLTLC | n.a. | seeds | n.a. | [13] | 4 IAs |
HPLC–UV | 1 g | seeds | 0.01–0.05 µg·mL−1 | [14] | 2 IAs |
HPLC–UV | n.a. | cell culture | n.a. | [15] | 22 IAs |
UPLC/IM–QTOF–MS * | 0.1 g | yohimbe bark | n.a. | [18] | 55 IAs |
MEKC–UV/LIF | n.a. | none | n.a. | [19] | 6 IAs |
HPLC–(+)ESI–QTOF–MS/MS | 50 g | roots | n.a. | [26] | 47 IAs |
HPLC–UV–MS | 0.5 g | aphrodisiac products | 3–60 ng·mL−1 | [29] | yohimbine |
HPLC–UV | 2 g | roots | 0.05–0.39 µg·mL−1 | [30] | 5 IAs |
HPLC–UV | 5 g | roots | n.a. | [31] | 6 IAs |
Compound | RT (min) | MRM (Q) | Cone Voltage (V) | Collision Energy (V) | MRM (C) | Cone Voltage (V) | Collision Energy (V) |
---|---|---|---|---|---|---|---|
Harmaline | 2.13 | 214.89 > 199.89 | 40 | 25 | 214.89 > 172.11 | 40 | 30 |
Harmine | 2.16 | 212.94 > 198.00 | 40 | 25 | 212.94 > 169.84 | 40 | 30 |
Yohimbine | 2.98 | 355.02 > 143.82 | 40 | 30 | 355.02 > 211.93 | 40 | 25 |
Methyl-13C, D3-Yohimbine | 2.96 | 359.05 > 143.94 | 40 | 30 | 359.05 > 216.02 | 40 | 25 |
Ajmalicine | 4.94 | 352. 94 > 144.07 | 40 | 25 | 352.94 > 177.91 | 40 | 25 |
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Share and Cite
Tarkowská, D. A Fast and Reliable UHPLC–MS/MS-Based Method for Screening Selected Pharmacologically Significant Natural Plant Indole Alkaloids. Molecules 2020, 25, 3274. https://doi.org/10.3390/molecules25143274
Tarkowská D. A Fast and Reliable UHPLC–MS/MS-Based Method for Screening Selected Pharmacologically Significant Natural Plant Indole Alkaloids. Molecules. 2020; 25(14):3274. https://doi.org/10.3390/molecules25143274
Chicago/Turabian StyleTarkowská, Danuše. 2020. "A Fast and Reliable UHPLC–MS/MS-Based Method for Screening Selected Pharmacologically Significant Natural Plant Indole Alkaloids" Molecules 25, no. 14: 3274. https://doi.org/10.3390/molecules25143274