Metabolomic Profiles of Dinophysis acuminata and Dinophysis acuta Using Non-Targeted High-Resolution Mass Spectrometry: Effect of Nutritional Status and Prey
Abstract
:1. Introduction
2. Results
2.1. Phylogenetic Analysis
2.2. Growth Curves
2.3. Quantitative Toxin Analysis Using Low Resolution Mass Spectrometry
Esters of OA-Group Toxins
2.4. Data Treatment for Non-Targeted Liquid Chromatography High Resolution Mass Spectrometry
2.4.1. Compounds with an Already Known Physiological Meaning
Common to All Species
Compounds Exclusive to the Ciliate Mesodinium
Compounds Exclusive to Teleaulax amphioxeia
Compounds Exclusive to One of the Two Species of Dinophysis
2.4.2. Compounds First Found in Other Marine/Terrestrial Organisms
Compounds Present in Both Species
Compounds Exclusive to One of the Two Species of Dinophysis
3. Discussion
3.1. Growth Curves
3.2. Toxin Quota and Target Analysis
3.3. Semi-Quantitative Non-Targeted Analysis: Comparison of Larger Parts of Metabolomes
4. Material and Methods
4.1. Cultures
4.2. DNA Extraction, Polymerase Chain Reaction (PCR), 18S Sequencing and Phylogenetic Analysis of Mesodinium
4.3. Biovolume
4.4. Experiment 1 (Baseline Study)
4.5. Experiment 2 (Growth Curves)
4.6. Toxin Analysis
4.6.1. Extraction and Hydrolysis
4.6.2. High Resolution Mass Spectral Analysis—System A: (QTOF 6550)
4.6.3. Low Resolution Tandem Mass Spectrometry: System B (API 4000 QTrap)
4.7. Data Treatment for Non-Targeted Liquid Chromatography High Resolution Mass Spectrometry
4.7.1. Feature Identification and Variables’ Selection
5. Conclusions
Supplementary Materials
Author Contributions
Acknowledgments
Conflicts of Interest
References
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Sample | Day | Cells (N) ± Std Dev. | Pellet Biomass % Average | Volume (mL) | ||
---|---|---|---|---|---|---|
Dinophysis | M. rubrum | T. amphioxeia | ||||
T. amphioxeia | 3 | 283,333 ± 28,867 | 100 | 10 | ||
Danish M. rubrum | 7 | 89,500 ± 866 | 100 | 15 | ||
Spanish M. rubrum | 7 | 80,000 ± 8660 | 100 | 30 | ||
D. acuta + Danish M. rubrum | 7 | 40,140 ± 1247 | 93.48 | 6.52 | 90 | |
D. acuta + Spanish M. rubrum | 7 | 29,943 ± 3264 | 97.26 | 2.74 | 130 | |
D. acuminata + Danish M. rubrum | 7 | 115,470 ± 9490 | 99.40 | 0.60 | 90 | |
D. acuminata + Spanish M. rubrum | 7 | 58,413 ± 3134 | 86.62 | 10.38 | 130 | |
D. acuta + Danish M. rubrum | 15 | 21,967 ± 2627 | 81.62 | 18.38 | 100 | |
D. acuta + Spanish M. rubrum | 15 | 42,733 ± 3225 | 100 | 100 | ||
D. acuminata + Danish M. rubrum | 15 | 225,800 ± 9616 | 100 | 100 | ||
D. acuminata + Spanish M. rubrum | 15 | 58,900 ± 265 | 100 | 100 |
Sp | PO | GP/NS | Toxin Contents pb = per Biomass (fg µm−3), pc = per Cell (pg cell−1) ± Standard Deviation | |||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Free OA | Total OA | Free DTX2 | Total DTX2 | PTX2 | ||||||||
pb | pc | pb | pc | pb | pc | pb | pc | pb | pc | |||
D. acuta | ES | ME | 0.17 ± 0.03 | 9.2 ± 2.0 | 0.22 ± 0.03 | 12.2 ± 2.3 | 0.07 ± 0.01 | 3.8 ± 0.6 | 0.08 ± 0.01 | 4.4 ± 0.9 | 0.41 ± 0.14 | 22.2 ± 9.4 |
D. acuminata | ES | ME | 1.05 ± 0.34 | 21.5 ± 0.5 | 2.1 ± 0.3 | 35.2 ± 6.8 | ||||||
D. acuta | ES | Well-Fed | 0.55 ± 0.06 | 30.0 ± 3.7 | 0.75 ± 0.07 | 41.0 ± 4.9 | 0.24 ± 0.03 | 13.5 ± 1.7 | 0.32 ± 0.06 | 17.4 ± 4.1 | 0.69 ± 0.12 | 38.0 ± 8.2 |
Prey-limited | 1.49 ± 0.24 | 76.7 ± 18.6 | 1.36 ± 0.11 | 74.1 ± 8.2 | 0.56 ± 0.11 | 30.2 ± 8.5 | 0.60 ± 0.05 | 32.4 ± 3.8 | 1.29 ± 0.15 | 59.3 ± 11.8 | ||
DK | Well-Fed | 0.53 ± 0.12 | 28.7 ± 8.1 | 0.66 ± 0.11 | 35.9 ± 7.07 | 0.26 ± 0.03 | 14.2 ± 2.2 | 0.30 ± 0.01 | 16.5 ± 0.8 | 0.78 ± 0.01 | 70.0 ± 0.8 | |
Prey-limited | 0.64 ± 0.08 | 34.6 ± 5.4 | 0.71 ± 0.07 | 38.6 ± 4.5 | 0.32 ± 0.04 | 17.4 ± 2.7 | 0.35 ± 0.03 | 19.0 ± 2.3 | 0.8 ± 0.09 | 43.6 ± 6.2 | ||
D. acuminata | ES | Well-Fed | 0.37 ± 0.14 | 6.0 ± 3.0 | 0.36 ± 0.13 | 6.0 ± 2.8 | ||||||
Prey-limited | 1.02 ± 0.20 | 17.1 ± 4.6 | 1.28 ± 0.03 | 21.5 ± 0.7 | ||||||||
DK | Well-Fed | 0.49 ± 0.09 | 8.3 ± 1.5 | 0.59 ± 0.08 | 9.8 ± 1.7 | |||||||
Prey-limited | 1.44 ± 0.32 | 24.0 ± 6.5 | 1.93 ± 0.23 | 32.3 ± 4.7 |
Name | Sequence 5′–3′ | Reference |
---|---|---|
4617F | TCCTGCCAGTAGTCATATGC | [93] |
Meso580R | GACGTACAGACTACGGACG | [94] |
Meso245F | CGACTCGACGTCCCG | [94] |
UNIDEUK1416R | GTTTCAGACTTGTGTCCATACTA | [95] |
Meso580F | CGTCCGTAGTCTGTACGTC | [94] |
Meso1480R | CTAAACACTCGATCGGTAGG | [94] |
Meso1200F | ATTCCGGTAACGAACGAGAC | [94] |
Meso28S_R | AGACTTGGATGACTTTTATCACC | [94] |
ITS1 | TCCGTAGGTGAACCTGCGG | [96] |
Dir-2CR | CCTTGGTCCGTGTTTCAAGA | [97] |
Toxin | Ionization Mode | Ion | Parent Ion (m/z) | Fragment Ions (m/z) | DP (V) | CE (eV) |
---|---|---|---|---|---|---|
OA | ESI− | [M − H]− | 803.4 | 255.1 * | −170 | −62 |
113.1 | −92 | |||||
DTX2 | ESI− | [M − H]− | 803.4 | 255.1 * | −170 | −62 |
113.1 | −92 | |||||
DTX1 | ESI− | [M − H]− | 817.5 | 255.1 * | −170 | −68 |
113.1 | −92 | |||||
PTX2 | ESI+ | [M + NH4]+ | 876.6 | 823.6 * | 91 | 31 |
805.6 | 37 | |||||
213.6 | 55 | |||||
PTX2sa and 7-epi PTX2sa | ESI+ | [M + NH4]+ | 894.6 | 823.6 * | 91 | 31 |
805.6 | 37 | |||||
213.6 | 55 | |||||
PTX1, PTX4 and PTX11 | ESI+ | [M + NH4]+ | 892.6 | 839.6 * | 91 | 31 |
821.4 | 37 | |||||
213.6 | 55 | |||||
PTX3 | ESI+ | [M + NH4]+ | 890.5 | 873.6 * | 91 | 31 |
856.6 | 37 | |||||
213.6 | 55 | |||||
PTX6 and PTX7 | ESI+ | [M + NH4]+ | 906.6 | 871.6 * | 91 | 31 |
853.6 | 37 | |||||
213.6 | 55 | |||||
PTX12 and PTX14 | ESI+ | [M + NH4]+ | 874.6 | 857.6 * | 91 | 31 |
840.6 | 37 | |||||
213.6 | 55 |
© 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
García-Portela, M.; Reguera, B.; Sibat, M.; Altenburger, A.; Rodríguez, F.; Hess, P. Metabolomic Profiles of Dinophysis acuminata and Dinophysis acuta Using Non-Targeted High-Resolution Mass Spectrometry: Effect of Nutritional Status and Prey. Mar. Drugs 2018, 16, 143. https://doi.org/10.3390/md16050143
García-Portela M, Reguera B, Sibat M, Altenburger A, Rodríguez F, Hess P. Metabolomic Profiles of Dinophysis acuminata and Dinophysis acuta Using Non-Targeted High-Resolution Mass Spectrometry: Effect of Nutritional Status and Prey. Marine Drugs. 2018; 16(5):143. https://doi.org/10.3390/md16050143
Chicago/Turabian StyleGarcía-Portela, María, Beatriz Reguera, Manoella Sibat, Andreas Altenburger, Francisco Rodríguez, and Philipp Hess. 2018. "Metabolomic Profiles of Dinophysis acuminata and Dinophysis acuta Using Non-Targeted High-Resolution Mass Spectrometry: Effect of Nutritional Status and Prey" Marine Drugs 16, no. 5: 143. https://doi.org/10.3390/md16050143