Identification of 24-O-β-d-Glycosides and 7-Deoxy-Analogues of Okadaic Acid and Dinophysistoxin-1 and -2 in Extracts from Dinophysis Blooms, Dinophysis and Prorocentrum Cultures, and Shellfish in Europe, North America and Australasia
Abstract
:1. Introduction
2. Results and Discussion
2.1. Mass Spectrometric Analysis of Glucosides 4 and 5
2.2. NMR Structure Elucidation of Glucosides 4 and 5
2.3. LC–MSn and LC–HRMS/MS Detection of DTX-1 24-O-β-d-Glucoside (6)
2.4. Detection of OA/DTX Analogues in Dinophysis and P. lima Cultures
2.5. Detection of 1–9 in Blue Mussels from Canada, Ireland, Norway, and New Zealand
3. Conclusions
4. Materials and Methods
4.1. Reagents
4.2. Shellfish Samples
4.3. Passive Sampler Extracts
4.4. Isolation of Glucosides 4 and 5
4.5. Dinophysis Cultures
4.6. Cultured P. lima Extracts
4.7. LC–MSn (Method A)
4.8. LC–HRMS (Method B)
4.9. LC–HRMS (Method C)
4.10. LC–HRMS (Method D)
4.11. LC–MS/MS (Method E)
4.12. NMR Spectroscopy
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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tRb | Ion A | Ion B | Ion C | Ion D | Ion E | Ion F | Ion G | Ion H | Ion I | Ion J | Ion K | Ion L | |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
1 | 13.42 | 255.1238 | 463.2337 | 563.2862 | 321.2071 | 723.4079 | 571.3241 | 443.2404 | 383.1829 | 483.2353 | 595.3241 | 263.1618 | 151.0730 |
2 | 14.15 | 255.1238 | 463.2337 | 563.2862 | 321.2071 | 723.4079 | 571.3241 | 429.2248 | 383.1829 | 483.2353 | 581.3085 | 263.1618 | 165.0886 |
3 | 15.75 | 255.1238 | 463.2337 | 563.2862 | 335.2228 | 737.4235 | 585.3398 | 443.2404 | 383.1829 | 483.2353 | 595.3241 | 277.1774 | 165.0886 |
4 | 9.12 | 255.1238 | 463.2337 | 725.3390 | 321.2071 | 885.4607 | 733.3770 | 605.2932 | 383.1829 | 645.2881 | 757.3770 | 263.1618 | 151.0730 |
5 | 9.48 | 255.1238 | 463.2337 | 725.3390 | 321.2071 | 885.4607 | 733.3770 | 591.2776 | 383.1829 | 645.2881 | 743.3613 | 263.1618 | 165.0886 |
6 | 11.01 | 255.1238 | 463.2337 | 725.3390 | 335.2228 | 899.4763 | 747.3926 | 605.2932 | 383.1829 | 645.2881 | 757.3770 | 277.1774 | 165.0886 |
7 | 17.68 | 239.1289 | 447.2388 | 547.2913 | 321.2071 | 707.4130 | 571.3241 | 443.2404 | 367.1880 | 467.2404 | 579.3292 | 263.1618 | 151.0730 |
8 | 18.25 | 239.1289 | 447.2388 | 547.2913 | 321.2071 | 707.4130 | 571.3241 | 429.2248 | 367.1880 | 467.2404 | 565.3136 | 263.1618 | 165.0886 |
9 | 19.76 | 239.1289 | 447.2388 | 547.2913 | 335.2228 | 721.4286 | 585.3398 | 443.2404 | 367.1880 | 467.2404 | 579.3292 | 277.1774 | 165.0886 |
OA (1) | 24-GlcOA (4) | 24-GlcDTX2 (5) | DTX2 (2) | |||||
---|---|---|---|---|---|---|---|---|
Atom | 13C | 1H | 13C | 1HG | 13C | 1H | 13C | 1H |
1 | 182.8 | 182.1 | 183.3 | 180.6 | ||||
2 | 76.3 | 76.5 | 76.2 | 76.1 | ||||
3 | 46.7 | 1.65, 1.91 | 46.8 | 1.69, 1.88 | 46.7 | 1.68, 1.89 | 45.7 | 1.65, 2.01 |
4 | 68.9 | 4.08 | 68.7 | 4.09 | 68.7 | 4.09 | 69.3 | 4.08 |
5 | 33.4 | 1.30, 1.89 | 33.6 | 1.30, 1.79 | 33.6 | 1.31, 1.81 | 33.4 | 1.34, 1.79 |
6 | 28.3 | 1.66, 1.94 | 28.4 | 1.65, 1.96 | 28.3 | 1.65, 1.97 | 28.0 | 1.65, 1.98 |
7 | 73.4 | 3.35 | 73.4 | 3.36 | 73.4 | 3.36 | 73.1 | 3.37 |
8 | 97.5 | 97.5 | 97.6 | 97.6 | ||||
9 | 123.8 | 5.26 | 123.9 | 5.28 | 123.8 | 5.28 | 123.2 | 5.29 |
10 | 139.3 | 139.2 | 139.2 | 139.9 | ||||
11 | 34.0 | 1.82, 1.98 | 34.1 | 1.83, 1.97 | 34.1 | 1.84, 1.97 | 34.1 | 1.88, 1.96 |
12 | 71.8 | 3.87 | 71.8 | 3.91 | 71.8 | 3.91 | 71.9 | 3.75 |
13 | 43.2 | 2.35 | 43.5 | 2.36 | 43.4 | 2.36 | 43.3 | 2.34 |
14 | 137.2 | 5.95 | 137.3 | 5.95 | 137.2 | 5.93 | 137.4 | 5.83 |
15 | 132.0 | 5.50 | 131.8 | 5.50 | 131.8 | 5.49 | 132.3 | 5.52 |
16 | 80.6 | 4.66 | 80.7 | 4.60 | 80.7 | 4.60 | 80.5 | 4.62 |
17 | 31.5 | 1.60, 2.21 | 31.6 | 1.61, 2.18 | 31.6 | 1.61, 2.18 | 31.5 | 1.61, 2.20 |
18 | 38.1 | 1.85, 2.00 | 38.1 | 1.89, 2.00 | 38.1 | 1.89, 2.02 | 38.0 | 1.85, 2.00 |
19 | 107.1 | 107.4 | 107.4 | 107.1 | ||||
20 | 34.2 | 1.84–1.87 | 34.3 | 1.85–1.92 | 34.3 | 1.85–1.92 | 34.1 | 1.82–1.89 |
21 | 27.7 | 1.80, 1.90 | 28.0 | 1.80, 1.91 | 28.0 | 1.81, 1.90 | 27.7 | 1.79, 1.90 |
22 | 71.4 | 3.63 | 71.6 | 3.72 | 71.6 | 3.72 | 71.4 | 3.64 |
23 | 78.3 | 3.42 | 76.9 | 3.59 | 76.8 | 3.59 | 78.1 | 3.40 |
24 | 72.1 | 4.07 | 75.9 | 4.44 | 75.5 | 4.44 | 72.2 | 4.10 |
25 | 147.1 | 143.9 | 144.1 | 147.4 | ||||
26 | 86.4 | 3.94 | 86.8 | 3.93 | 87.0 | 3.94 | 86.4 | 3.97 |
27 | 66.3 | 4.08 | 65.9 | 4.14 | 66.1 | 4.13 | 67.0 | 4.07 |
28 | 36.8 | 0.96, 1.37 | 37.0 | 0.92, 1.41 | 38.9 | 1.37, 1.42 | 38.6 | 1.37, 1.44 |
29 | 32.3 | 1.89 | 32.5 | 1.87 | 35.3 | 1.82 | 35.3 | 1.83 |
30 | 76.8 | 3.27 | 76.8 | 3.27 | 75.0 | 3.42 | 75.0 | 3.42 |
31 | 28.8 | 1.81 | 28.9 | 1.82 | 28.8 | 1.27, 1.52 | 28.7 | 1.27, 1.52 |
32 | 27.5 | 1.38, 2.00 | 27.6 | 1.38, 2.00 | 20.1 | 1.61, 1.74 | 20.1 | 1.74, 1.60 |
33 | 31.2 | 1.36, 1.59 | 31.3 | 1.32, 1.59 | 33.4 | 1.17, 1.67 | 33.4 | 1.16, 1.68 |
34 | 97.0 | 97.0 | 99.2 | 99.3 | ||||
35 | 37.0 | 1.43, 1.61 | 37.1 | 1.41, 1.61 | 37.4 | 1.62 | 37.4 | 1.61 |
36 | 19.8 | 1.53, 1.88 | 19.9 | 1.53, 1.88 | 26.9 | 1.32, 2.17 | 26.7 | 1.29, 2.17 |
37 | 26.5 | 1.49–1.55 | 26.6 | 1.49–1.56 | 20.9 | 1.28, 1.78 | 21.0 | 1.78, 1.26 |
38 | 61.3 | 3.51, 3.71 | 61.4 | 3.52, 3.71 | 61.4 | 3.51, 3.70 | 61.4 | 3.51, 3.70 |
25 =CH2 | 112.6 | 5.04, 5.38 | 114.6 | 5.11, 5.67 | 114.3 | 5.10, 5.69 | 112.1 | 5.04, 5.36 |
10-Me | 23.1 | 1.72 | 23.2 | 1.72 | 23.2 | 1.73 | 23.1 | 1.75 |
13-Me | 17.1 | 1.11 | 17.1 | 1.11 | 17.2 | 1.11 | 16.8 | 1.07 |
29-Me | 16.6 | 1.05 | 17.0 | 1.07 | 14.5 | 0.96 | 14.4 | 0.95 |
2-Me | 27.9 | 1.31 | 27.6 | 1.32 | 27.5 | 1.32 | 27.7 | 1.34 |
31-Me | 11.1 | 0.93 | 11.2 | 0.94 | ||||
35-Me | 14.6 | 1.00 | 14.7 | 0.99 |
OA 24-O-β-d-Glucoside (4) | DTX2 24-O-β-d-Glucoside (5) | |||||
---|---|---|---|---|---|---|
atom | 13C | 1H | Mult, J (Hz) | 13C | 1H | Mult, J (Hz) |
1′ | 102.5 | 4.39 | d, 7.8 a | 102.5 | 4.39 | d, 7.8 a |
2′ | 75.0 | 3.39 | dd, 7.8, 9.0 b | 75.0 | 3.39 | dd, 7.8, 9.0 b |
3′ | 78.1 | 3.35 | ~t, J = 9.0 b | 78.2 | 3.32 | ~t, 9.0 b |
4′ | 71.9 | 3.28 | ~t, 9.0 b | 71.9 | 3.29 | ~t, 9.0 b |
5′ | 78.5 | 3.20 | m | 78.4 | 3.19 | m |
6′ | 63.1 | 3.64 | dd, 12.0, 5.5 b | 63.1 | 3.64 | dd, 12.0, 5.5 b |
3.83 | dd, 12.0, 2.2 b | 3.83 | dd, 12.0, 2.2 b |
Percentage of Sum of 1–9 | Sum 1–9 | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Sample | Description | Origin b | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | ng/mL | µg/g |
Mixed standard | Std of 4, 5 and 7 | N/A | 0.4 | 0.2 | 0.07 | 21 | 72 | ND | 6.1 | ND | ND | 2100 | |
RM-Multi-toxin | Multitoxin RM | N/A | 30 | 31 | 40 | ND | ND | ND | ND | ND | ND | 750 | |
D. acuta | VGO1065/1 | ES | 71 | 28 | 0.2 | 0.3 | 0.05 | ND | 0.09 | 0.04 | ND | 38,000 | |
D. caudata (conc.) | VGO1396/2 | ES | 77 | 12 | 10 | 0.5 | ND | ND | ND | ND | ND | 5.2 | |
D. caudata (conc.) | VGO1396/4 | ES | 68 | 14 | 17 | 0.3 | ND | ND | ND | ND | ND | 2.0 | |
D. acuminata | VGO1349/3 | ES | 99 | ND | 0.01 | 0.6 | 0.01 | ND | 0.1 | ND | ND | 2000 | |
D. acuminata | VGO1349/5 | ES | 100 | ND | 0.03 | 0.07 | ND | ND | 0.1 | ND | ND | 1200 | |
D. acuminata | VGO1391/7 | ES | 100 | ND | 0.01 | 0.3 | ND | ND | 0.1 | ND | ND | 2700 | |
P. lima | CCMP2579 | RI, US | 7.3 | ND | 93 | ND | ND | ND | 0.01 | ND | 0.04 | 2300 | |
P. lima | JHPL2 | NS, CA | 7.4 | ND | 92 | ND | ND | ND | 0.01 | ND | 0.07 | 18,000 | |
P. lima | JHPL3 | NS, CA | 5.8 | ND | 94 | ND | ND | ND | 0.01 | ND | 0.07 | 12,000 | |
P. lima | KP200 | PE, CA | 27 | ND | 73 | ND | ND | ND | 0.02 | ND | 0.03 | 13,000 | |
P. lima | KP209 | NB, CA | 37 | ND | 60 | ND | ND | ND | 2.1 | ND | 0.03 | 4100 | |
P. lima | IP197 | NS, CA | 44 | ND | 45 | ND | ND | ND | 11 | ND | 0.05 | 3100 | |
P. lima | CCMI1036 | IE | 5.0 | ND | 95 | ND | ND | ND | ND | ND | 0.08 | 1900 | |
P. lima | PL20V, Galicia | ES | 81 | ND | 17 | ND | ND | ND | 0.08 | ND | 0.02 | 1800 | |
FDMT1-SPE (conc.) | CRM concentrate | IE/NO | 19 | 55 | 13 | 8.6 | 3.9 | 0.3 | 0.1 | 0.2 | 0.07 | 6500 | |
CRM FDMT1 | CRM Extract | IE/NO | 25 | 59 | 8.8 | 5.1 | 2.5 | 0.2 | ND | ND | ND | 120 | |
CRM DSP-Mus | CRM Extract | IE/NO | 34 | 30 | 32 | 1.2 | 0.9 | 0.03 | 1.3 | 0.03 | 0.02 | 330 | 1.63 |
Mussel HP | Flødevigen | NO | 14 | 20 | 57 | 1.5 | 0.4 | 6.9 | ND | ND | ND | 7.3 | 0.04 |
Whole mussel | Flødevigen | NO | 7.4 | 8.5 | 78 | 0.4 | ND | 5.4 | ND | ND | ND | 61 | 0.32 |
Whole broth | Indian Pt. | NS, CA | 0.2 | ND | 98 | ND | ND | 1.4 | ND | ND | 0.02 | 93 | 0.52 |
Whole mussel | Bonavista Bay | NF, CA | 0.2 | ND | 97 | ND | ND | 2.5 | ND | ND | 0.5 | 41 | 0.33 |
Whole mussel | Ship Harbour | NS, CA | 92 | ND | 4.2 | 3.9 | ND | 0.2 | ND | ND | ND | 50 | 0.48 |
Whole mussel c | Castlemaine Harbour | IE | 35 | 5.7 | 3.8 | 54 | 1.3 | 0.2 | ND | ND | ND | 10 | 0.09 |
Whole mussel d | Bantry Middle | IE | 67 | 8.7 | 2.0 | 21 | 0.5 | 0.1 | 0.7 | ND | ND | 36 | 0.43 |
SPATT (conc.) | Ship Harbour | NS, CA | 8.8 | ND | 91 | ND | ND | 0.2 | ND | ND | 0.1 | 37 |
Reported Results | |||||||
---|---|---|---|---|---|---|---|
Location | Date b | Species | 4 + 5 (%) | 1 | 2 | 1 Equiv. c | |
1 | Ardgroom | 02/06/20 | M. edulis | 5.5 | 2.68 | <LOD | 2.68 |
2 | Kilmakilloge | 02/06/20 | M. edulis | 6.7 | 2.61 | <LOD | 2.61 |
3 | Mine Head Ground | 03/06/20 | P. maximus | ND | 0.11 | 0.03 | 0.13 |
4 | Youghal Bay | 10/06/20 | S. solida | ND | 0.13 | <LOD | 0.13 |
5 | Valentia River | 11/06/20 | C. gigas | ND | 0.12 | <LOD | 0.12 |
6 | Castlemaine Harbour | 24/06/20 | C. edule | ND | 0.20 | <LOD | 0.20 |
7 | Tahilla | 30/06/20 | M. edulis | 20.2 | 0.69 | 0.02 | 0.70 |
8 | Bantry Middle | 30/06/20 | M. edulis | 16.8 | 1.21 | 0.08 | 1.26 |
9 | Gouleenacoush | 01/07/20 | M. edulis | 17.2 | 1.45 | <LOD | 1.45 |
10 | Adrigole | 01/07/20 | M. edulis | 14.1 | 0.66 | 0.07 | 0.70 |
11 | Tahilla | 13/07/20 | M. edulis | 39.2 | 0.33 | 0.04 | 0.35 |
12 | Castlemaine Harbour d | 13/07/20 | M. edulis | 58.0 | 0.09 | <LOQ | 0.09 |
13 | Castlemaine Harbour | 13/07/20 | M. edulis | 44.1 | 0.08 | <LOQ | 0.08 |
14 | Bantry North | 13/07/20 | M. edulis | 40.5 | 0.22 | 0.06 | 0.26 |
15 | Bantry Middle | 13/07/20 | M. edulis | 21.4 | 0.40 | 0.05 | 0.43 |
16 | South of Smalls Ground | 04/08/20 | P. maximus | ND | 0.10 | 0.03 | 0.12 |
17 | Bantry North d | 31/08/20 | M. edulis | 5.7 | 0.35 | 0.73 | 0.79 |
18 | Bantry South | 31/08/20 | M. edulis | 4.9 | 0.84 | 0.76 | 1.30 |
19 | Ardgroom | 31/08/20 | M. edulis | 26.0 | 0.07 | 0.12 | 0.14 |
Non-Hydrolyzed | Hydrolyzed | |||||||
---|---|---|---|---|---|---|---|---|
Location | Species | Date b | 1 | 3 | 4 | 1 | 3 | 4 |
Sumner | M. edulis | 04/01/17 | 0.151 | 0.008 | 0.011 | 0.226 | <LOQ | 0.013 |
Akaroa harbour | M. edulis | 05/07/17 | 0.420 | 0.010 | 0.010 | 0.826 | 0.012 | <LOQ |
Akaroa harbour | M. edulis | 12/07/17 | 0.268 | 0.005 | 0.004 | 0.590 | <LOQ | <LOQ |
Akaroa harbour | M. edulis | 31/01/17 | 0.412 | 0.018 | 0.006 | 0.833 | 0.022 | <LOQ |
Akaroa harbour | M. edulis | 07/08/17 | 0.514 | 0.025 | 0.0010 | 0.978 | 0.032 | <LOQ |
Motukiekie Beach | P. canaliculus | 25/02/19 | 0.025 | ND | ND | 0.201 | ND | ND |
Cape Foulwind | P. canaliculus | 04/03/19 | 0.021 | ND | ND | 0.191 | ND | ND |
Cape Foulwind | P. canaliculus | 18/03/19 | 0.020 | ND | ND | 0.220 | ND | ND |
Motukiekie Beach | M. edulis | 01/04/19 | 0.037 | ND | ND | 0.220 | ND | ND |
Sumner | P. subtriangulata | 02/06/20 | ND | <LOQ | ND | ND | 0.025 | ND |
Sumner | P. canaliculus | 09/06/20 | <LOQ | 0.009 | ND | 0.012 | 0.072 | ND |
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Wilkins, A.L.; Rundberget, T.; Sandvik, M.; Rise, F.; Knudsen, B.K.; Kilcoyne, J.; Reguera, B.; Rial, P.; Wright, E.J.; Giddings, S.D.; et al. Identification of 24-O-β-d-Glycosides and 7-Deoxy-Analogues of Okadaic Acid and Dinophysistoxin-1 and -2 in Extracts from Dinophysis Blooms, Dinophysis and Prorocentrum Cultures, and Shellfish in Europe, North America and Australasia. Toxins 2021, 13, 510. https://doi.org/10.3390/toxins13080510
Wilkins AL, Rundberget T, Sandvik M, Rise F, Knudsen BK, Kilcoyne J, Reguera B, Rial P, Wright EJ, Giddings SD, et al. Identification of 24-O-β-d-Glycosides and 7-Deoxy-Analogues of Okadaic Acid and Dinophysistoxin-1 and -2 in Extracts from Dinophysis Blooms, Dinophysis and Prorocentrum Cultures, and Shellfish in Europe, North America and Australasia. Toxins. 2021; 13(8):510. https://doi.org/10.3390/toxins13080510
Chicago/Turabian StyleWilkins, Alistair L., Thomas Rundberget, Morten Sandvik, Frode Rise, Brent K. Knudsen, Jane Kilcoyne, Beatriz Reguera, Pilar Rial, Elliott J. Wright, Sabrina D. Giddings, and et al. 2021. "Identification of 24-O-β-d-Glycosides and 7-Deoxy-Analogues of Okadaic Acid and Dinophysistoxin-1 and -2 in Extracts from Dinophysis Blooms, Dinophysis and Prorocentrum Cultures, and Shellfish in Europe, North America and Australasia" Toxins 13, no. 8: 510. https://doi.org/10.3390/toxins13080510
APA StyleWilkins, A. L., Rundberget, T., Sandvik, M., Rise, F., Knudsen, B. K., Kilcoyne, J., Reguera, B., Rial, P., Wright, E. J., Giddings, S. D., Boundy, M. J., Rafuse, C., & Miles, C. O. (2021). Identification of 24-O-β-d-Glycosides and 7-Deoxy-Analogues of Okadaic Acid and Dinophysistoxin-1 and -2 in Extracts from Dinophysis Blooms, Dinophysis and Prorocentrum Cultures, and Shellfish in Europe, North America and Australasia. Toxins, 13(8), 510. https://doi.org/10.3390/toxins13080510