Metabolomic fingerprinting and genetic discrimination of four Hypericum taxa from Greece Author Zeliou, Konstantina ∗ & Laboratory of Pharmacognosy and Chemistry of Natural Products, Department of Pharmacy, University of Patras, 26504 Patras, Greece Author Koui, Eirini-Maria Author Papaioannou, Charikleia Author Koulakiotis, Nikolaos Stavros Author Iatrou, Gregoris Author Tsarbopoulos, Anthony Author Papasotiropoulos, Vassilis Author Lamari, Fotini N. text Phytochemistry 2020 112290 2020-06-30 174 1 15 http://dx.doi.org/10.1016/j.phytochem.2020.112290 journal article 10.1016/j.phytochem.2020.112290 1873-3700 8295050
Table 2 Polar secondary metabolites in methanolic extracts of the four Hypericum taxa.
Peak No.RtMetaboliteMolecular FormulaObserved [Μ- Η] -Accuracy (ppm)λ max (nm)Negative Ionization m/zPositive Ionization m/z (Rel.Int.%)ID level aHypericum taxa b
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Identification was performed with two LC-MS techniques. The retention time, spectral data (λ max), the main ions after negative and positive ionization (along with their relative abundance) were obtained with UHPLC- DAD-MS (ESI-quadrupole ion trap). The observed [M-H] - and the accuracy were obtained with UPLC-MS (ESI-qTOF). a Metabolite identification level according to Metabolomics Standards Initiative recommendation (Sumner et al., 2007). b HPV– H. perforatum subsp. veronense, HP – H. perfoliatum, HT – H. triquetrifolium, HEE – H. empetrifolium subsp. empetrifolium. c Two or more different compounds with the same molecular weight are possible. d UV-vis spectra was obtained by HPLC-DAD analysis.
(Rel. Int. %)
1 2 310.4 11.2 11.9Caffeoylquinic Acid I (CQA) Flavanol I (Dimeric) Chlorogenic Acid (3–CQA)C 16H18 O 9 C 30H26 O 12 C 16H18 O 9353.0889 577.1346 353.08814.7 −6.8 2.4321 282 312353 [M-H] - (100) 191 [(M-H)- caffeoyl] - (15) 577 [M-H] - (100) 289 [(M-H)-(epi) catechin] - (20) 707 [2M-H] - (31) 353 [M-H] - (100) 191 [M-H- caffeoyl] - (6)377 [M+Na] + (100) 355 [M+H] + (29) 163[(M+H)-quinic acid] + (34) 601 [M+Na] + (25) 579 [M+H] + (100) 377 [M+Na] + (100) 355 [M+H] + (29) 163 [(M+H)-quinic acid] + (62)2 3 1HPV, HP, HT, HEE HPV HPV, HP, HT, HEE
4 5 612.4 12.6 13.2Flavanol II (Monomeric) Flavanol III (Dimeric) Caffeoylquinic Acid II (CQA)C 15H14 O 6 C 30H26 O 12 C 16H18 O 9289.0688 577.1319 353.0853−8.3 −4.7 −5.5282 425, 289 321578 [2M-H] - (10) 289 [M-H] - (100) 577 [M-H] - (100) 289[(M-H)-(epi) catechin] - (14) 707 [2M-H] - (14) 353 [M-H] - (100)579 [2M + H] + (20), 291[M+H] + (100) 601 [M+Na] + (30) 579 [M+H] + (100), 291 [(M+H)-(epi)catechin] + (18) 377 [M+Na] + (100) 355 [M+H] + (32)3 3 2HPV, HP HPV, HP, HT HP, HT, HEE
7 8 9 10 11 1213.5 13.9 14.3 14.8 15.3 15.5Flavanol IV (Monomeric) Flavanol V (Trimeric) Myricetin hexoside Quercetin–3– O –rutinoside (Rutin) Quercetin–3– O –galactoside (Hyperoside) Quercetin–3– O –glucoside (Isoquercitrin)C 15H14 O 6 C 45H38 O 18 C 21H20 O 13 C 27H30 O 16 C 21H20 O 12 C 21H20 O 12289.0692 865.1973 479.0812 609.1398 463.0852 463.0844−6.9 −0.7 −2.9 −9.5 −5.3 −7.0282 284 269, 355 265, 354 258, 355 257, 356191 [(M-H)- caffeoyl] - (4) 865 [3M-H] - (13) 578 [2M-H] - (12) 289 [M-H] - (100) 865 [M-H] - (100) 577 [(M-H)-(epi) catechin] - (83) 479 [M-H] - (100) 609 [M-H] - (100) 463 [(M-H)- rhamnose] - (7) 463 [M-H] - (100) 463 [M-H] - (100)163 [(M+H)-quinic acid] + (41) 313 [M+Na] + (31) 291 [M+H] + (100) 887 [M+Na] + (16) 867 [M+H] + (100) 503 [M+Na] + (100) 481 [M+H] + (81) 319 [(M+H)-hexose] + (70) 633 [M+Na] + (35) 611 [M+H] + (100) 465 [(M+H)-rhamnose] + (8) 303 [(M+H)-rutinose] + (8) 487 [M+Na] + (70) 465 [M+H] + (100) 303 [(M+H)-hexose] + (43) 487 [M+Na] - (59) 465 [M+H] + (100)3 3 2 1 1 1HPV, HP, HT, HEE HPV, HT HP, HEE HPV, HP, HT, HEE HPV, HP, HT, HEE HPV, HP, HT, HEE
13 14 15 16 c16.2 16.6 16.9 18.0Taxifolin 3– O –rhamnoside (Astilbin) Quercetin pentoside Quercetin 3– O –rhamnoside (Quercitrin) Quercetin 3–acetyl–galactosideC 21H22 O 11 C 20H18 O 11 C 21H20 O 11 C 23H22 O 13449.1092 433.0731 447.0887 505.09591.8 −9.2 −9.0 −4.6292 264, 355 264, 352 265, 355889 [2M-H] - (6) 449 [M-H] - (100) 433 [M-H] - (100) 447 [M-H] - (100) 505 [M-H] - (100)303 [(M+H)-hexose] + (49) 473 [M+Na] - (100) 451 [M+H] + (47) 305 [(M+H)-rhamnose] + (45) 457 [M+Na] + (100) 435 [M+H] + (87) 303 [(M+H)-pentose] + (95) 471 [M+Na] + (69) 449 [M+H] + (100) 303 [(M+H)-rhamnose] + (87) 529 [M+Na]+(72) 507 [M+H] + (100) 303 [(M+H)-acetyl–galactose] +2 2 2 2HPV HEE HPV, HP, HT, HEE HT, HEE
(11)
(Rel. Int. %)
17 c 18 1918.7 18.9 21.4Xanthone Quercetin–3– O –acetyl–arabinofuranoside QuercetinC 24H20 O 9 C 22H20 O 12 C 15H10 O 7451.1010 475.0843 301.0325−4.2 −7.1 −7.7269, 352 268,451 [M-H] - (100) 475 [M-H] - - 301 [M-H] - (100)475 [M+Na] + (53) 453 [M+H] + (100) 499 [M+Na] + (74) 477 [M+H] + (100) 303 [M+H] + (100)3 2 1HT HT, HEE HPV, HP, HT,
2023.53,8″–BiapigeninC 30H18 O 10537.0801−3.9375 269,537 [M-H] - (100)539 [M+H] + (100)2HEE HPV, HP, HT,
2123.63′,8″–Biapigenin (Amentoflavone)C 30H18 O 10537.0811−2.0332 269,537 [M-H] - (100)539 [M+H] + (100)2HEE HPV, HP, HT,
22 c 23 c 24 c 25 c 2626.4 27.1 27.8 28.4Dihydroxy–methoxyxanthone Xanthone Hyperireflexolide (A or B) Dihydroxyxanthone ProtopseudohypericinC 14H10 O 5 C 17H16 O 5 C 21H28 O 6 C 13H8 O 4 C 30H18 O 9257.0428 299.0909 375.1780 227.0339 521.0905−8.6 −3.5 −7.4 −2.4 6.2331 d 219,257 [M-H] - (100) 299 [M-H] - (100) 375 [M-H] - (100)259 [M+H] + (100) 301 [M+H] + (100) 229 [M+H] + (100)3 3 2 3 2HEE HEE HT HEE HEE HPV, HP, HT
256,
27PseudohypericinC 30H16 O 9519.07384.2541 d 218,1HPV, HP, HT,
236,HEE
28 c 29 3031.3 31.9 33.0Empetrifelixin (A or C) Empetrikathiforin EmpetriferdinolC 20H28 O 5 C 21H32 O 5 C 21H30 O 5347.1841 363.2160 361.2011−5.0 3.2 −1.1591 223, 287 223, 287 223,347 [M-H] - (100) 363 [M-H] - (100) 361 [M-H] - (100)371 [M+Na] + (71) 349 [M+H] + (100) 387 [M+Na] + (100) 365 [M+H] + (93) 359 [M+H] + (100)2 2 2HEE HEE HEE
31 32 3333.5 33.7 34.4Phloroglucinol I Phloroglucinol II 2,4,6–Trihydroxy–2–methyl–butyrophenon 4– O –3′,3′–dimethyl–allyletherC 21H30 O 6 C 21H30 O 6 C 16H22 O 4377.1933 377.1952 277.1435−8.3 −3.2 −1.8289 223, 287 223, 287 223,377 [M-H] - (100) 377 [M-H] - (100) 227 [M-H] - (100)401 [M+Na] + (100) 345 [(M+H)-86] + (95) 401 [M+Na] + (40) 379 [M+H] + (100) 315 [(M+H)-86] + (62) 229 [M+H] + (100)3 3 2HEE HEE HEE
34 3534.7 35.9Unknown Empetrikarinol AC 15H20 O 4 C 20H28 O 5263.1262 347.1848−8.1 −3.0287 224, 287 224, 286527 [2M-H] - (15) 263 [M-H] - (100) 347 [M-H] - (100)265 [M+H] + (70) 197[(M+H)-60] + (100) 371 [M+Na] + (21) 349 [M+H] + (100)4 2HEE HEE
3636.3Empetrikarinol BC 21H30 O 5361.1992−6.4225, 288361 [M-H] - (53) 227 [(M-H)-134] - (100)331 [(M+H)–H O] + (30) 2 243 [(M+H)-106] + (24) 209 [(M+H)-140] + (28) 363 [M+H] + (54) 279 [(M+H)-84] + (100) 211 [(M+H)-152] + (93)2HEE
37 c 3837.9Phloroglucinol III ProtohypericinC 23H34 O 5 C 30H18 O 8389.2314 505.0963−3.6 7.9d 220,389 [M-H] - (100)3 2HP HPV, HP, HT
254,
298,
3939.5Empetriferdinan AC 20H28 O 4331.19388.8542 228,331 [M-H] - (100)333 [M+H] + (100)2HEE
40 c 4140.8 40.8Phloroglucinol IV 1–[5,7–Dihydroxy–2–methyl–2–(4–methylpent–3–enyl)C 31H46 O 6 C 21H30 O 4513.3259 345.20538.4 −3.7291 225,513 [M-H] - (100) 345 [M-H] - (100)515 [M+H] + (100) 347 [M+H] + (100)3 2HT HEE
chroman–6–yl]–2–methylbutan–1–one292
Peak No. RtMetaboliteMolecular FormulaObserved [Μ- Η]-Accuracy (ppm)λ max (nm)Negative Ionization m/zPositive Ionization m/z (Rel.Int.%) ID level a Hypericum taxa b
(Rel. Int. %)
42HypericinC30H16O8503.0765−0.4d 219,1HPV, HP, HT,
233,HEE
43 44c 45 46 47c41.3 42.4 43.4 43.5 43.7Phloroglucinol V Phloroglucinol VI (Triquetribavarin or isomer) Furohyperforin isomer a Phloroglucinol VII Phloroglucinol VIII (Hyperforatin A or B or C or E or G or H or J or K or 33–epi–Hyperforatin E or Furohyperforin hydroperoxide)C20H28O4 C26H38O4 C35H52O5 C27H40O7 C35H52O6331.1982 413.2671 551.3691 427.2833 567.3710−8.2 −5.1 −8.2 −3.5 4.2591 226, 268331 [M-H]- (100) 827 [2M-H]- (38) 413 [M-H]- (100) 551 [M-H]- (100) 855 [2M-H]- (9) 427 [M-H]- (100) 567 [M-H]- (100) 551 [(M-H)-O]-333 [M+H]+ (100) 415 [M+H]+(100) 575 [M+Na]+ (60) 553 [M+H]+ (100) 429 [(M+H)-124]+ (70) 429 [M+H]+ (100) 341 [(M+H)-88]+ (24) 591 [M+Na]+ (100)3 3 2 3 3HT HT HPV HT HPV
48c44.2Hyperforatin (Hyperforatin A or B or C or E or G or H or J or K or 33–epi–Hyperforatin E)C35H52O6567.3633−9.3(23) 567 [M-H]- (100) 551 [(M-H)-O]- (19)591 [M+Na]+ (100) 569 [M+H]+ (59) 413 [(M+H)-156]+ (13)2HPV
49 50c 51 52c44.4 44.5 44.6 45.2Furohyperforin isomer b Phloroglucinol IX (Hyperfoliatin or isomer) Furoadhyperforin Phloroglucinol X (Triquetrijakobin B or Triquetrireboudin A or Triquetrireboudin B or Triquetriroxettin)C35H52O5 C31H46O5 C36H54O5 C31H46O5551.3685 497.3230 565.3898 497.3274−9.2 −7.4 0.9 1.4551 [M-H]- (100) 567 [M-H]- (100) 497 [M-H]- (100) 401 [(M-H)-96]-575 [M+Na]+ (59) 553 [M+H]+ (100) 499[M+H]+ (100) 325 [(M+H)-174]+ (35) 521[M+Na]+ (86) 499 [M+H]+ (100)2 3 2 3HPV HP HPV HT
53 54c 55c 56 57c 58 59 60 61 62 6345.5 46.3 46.4 46.6 46.8 47.8 49.4 49.8 51.1 52.1 52.7Phloroglucinol XI (Hyperfirin or isomer) Phloroglucinol XII (Triquetribavarin or isomer) Phloroglucinol XIII (Hyperfoliatin or isomer) Phloroglucinol XIV (Adhyperfirin or isomer) Phloroglucinol XV (Hyperibine J or Triquetriborin) Phloroglucinol XVI Furohyperforin Hyperforin Adhyperforin Phloroglucinol XVII (Adhyperfirin isomer) Phloroglucinol XVIII (Hyperibine J or Triquetriborin)C30H44O4 C26H38O4 C31H46O5 C31H46O4 C31H46O4 C32H48O4 C35H52O5 C35H52O4 C36H54O4 C31H46O4 C31H46O4467.3143 413.2675 497.3273 481.3318 481.3298 495.3429 551.3683 535.3777 549.3892 481.3316 481.3317−3.9 −4.1 1.2 −0.4 −4.2 −9.2 −9.6 −1.9 6.9 −0.4 −0.2225, 275 224, 279(36) 467 [M-H]- (100) 413 [M-H]- (100) 497 [M-H]- (100) 481 [M-H]- (100) 481 [M-H]- (100) 495 [M-H]- (100) 551 [M-H]- (100) 535 [M-H]- (100) 549 [M-H]- (100) 481 [M-H]- (100) 481 [M-H]- (100)481[(M+H)–H O]+ (44) 2 469 [M+H]+ (100) 415 [M+H]+ (100) 499 [M+H]+ (100) 325 [(M+H)-174]+ (80) 483 [M+H]+ (100) 483 [M+H]+ (100) 497 [M+H]+ (100) 325 [(M+H)-172]+ (34) 553 [M+H]+ (100) 537 [M+H]+ (100) 411 [(M+H)-126]+ (14) 551 [M+H]+ (100) 483 [(M+H)-68]+ (10) 411 [(M+H)-140]+ (20) 483 [M+H]+ (100) 483 [M+H]+ (100)3 3 3 3 3 3 2 2 2 3 3HT HT HP HP HT HP HPV HPV HPV HP HT