Valorizzazione di specie vegetali eduli spontanee e coltivate attraverso sistemi di caratterizzazione chimica e molecolare a garanzia di qualità
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1 Università degli studi di Cagliari CORSO DIDOTTORATO IN SCIENZE E TECNOLOGIE DELLA TERRA E DELL'AMBIENTE INDIRIZZO BOTANICA AMBIENTALE E APPLICATA Ciclo XXIX Valorizzazione di specie vegetali eduli spontanee e coltivate attraverso sistemi di caratterizzazione chimica e molecolare a garanzia di qualità DOTTORANDA: Dott.ssa Arianna Marengo TUTOR: Prof. Andrea Maxia CO-TUTOR: Prof.ssa Patrizia Rubiolo, Prof.ssa Cinzia Bertea
2 First PhDyear Literature research on Sardinian traditions Collection of plant species Validation of chemical and biomolecular techniques
3 Asteraceaefamily Carlininae CARDUEAE TRIBE (del Bosque et al., 2014) Carduinae Echinopsinae Sardinia~ 50 species (Pignatti, 1982) Centaureinae Cardopatiinae
4 CynaracardunculusL. Component of the mediterranean diet Nutritional and pharmacological properties Silybummarianum(L.) Gaertn. Native plant from the Mediterranean area Leaves and flowers used as vegetable Active compounds: silymarin
5 Carduus argyroabiv. Wild species Onopordum illyricum L. Ptilostemon casabonae(l.) Greuter Localtraditions Galactites tomentosa Moench.
6 AIM OF THE WORK Investigation on different edible species that are consumed in the Sardinian tradition Chemical analysis Biomolecular analysis (BARCODING) Some species has been poorly investigated
7 EXPERIMENTAL PROCEDURES Plant material Carduus cephalantus Viv. Carduus nutans subsp.macrocephalus (Desf.) Nyman Carduus pycnocephalus L. Carduus argyroa Biv. Ptilostemon casabonae (L.) Greuter
8 Absorption spectrum EXPERIMENTAL PROCEDURES 432 g/mol Chemical analysis Apigenin7-O-glucoside Mass spectrometry Photodiode detector SPD-M20A coupled to a triple quadrupole Shimadzu LCMS-8040 system and an electrosprayionization (ESI) source
9 Università degli Studi di Torino Dipartimento di Scienza e Tecnologia del farmaco
10 EXPERIMENTAL PROCEDURES Biomolecular analysis DNA extraction PCR (DNA barcoding genes) Gel electrophoresis Sequencing
11 RESULTS Species: Carduus argyroa Biv. Carduus pycnocephalusl. Carduus cephalantusviv. Carduus nutans subsp. macrocephalus(desf.) Nyman Ptilostemon casabonae(l.) Greuter Carduus Spp. Chemical analysis Biomolecular analysis (BARCODING)
12 CHEMICAL ANALYSIS Carduus argyroa mau 254nm,4nm Retention time UV spectra Mass analysis Product Ion Scan 24PM_ _PM _PM446 51PM_ _PM354 2_PM354 18_PM580 20_PM580 22_464 39_PM592 41_PM432 61_PM286 71_PM270 76_PM286 77_PM _PM min
13 CHEMICAL ANALYSIS Carduus argyroa mau 254nm,4nm Neochlorogenic acid Chlorogenic acid 18_PM580 20_PM580 Quercetin 3-O-glucoside Luteolin 3-O-glucoside 1,5 Dicaffeoylquinic acid+kaempferol 3-O-glucoside 39_PM592 Apigenin 3-O-glucoside Luteolin Apigenin Kaempferol 77_PM _PM243 51PM_432 43_PM min
14 Apigenin Apigenin 7-O-glucoside Diosmin Kaempferol Kaempferol 3-O-glucoside Kaempferol 3-O-rutinoside Luteolin Luteolin 7-O-glucoside Neochlorogenic Acid Quercetin 3-O-glucoside Rutin Chlorogenic Acid 1,5 Dicaffeoylquinic Ac. 3,5 Dicaffeoylquinic Ac. 4,5 Dicaffeoylquinic Ac. Identifiedcompounds O-glycosides of Apigenin, Kaempferol, Luteolin, and Quercetin Flavonoids CaffeoylquinicAcids derivates
15 mau 254nm,4nm Carduus argyroa Neochlorogenic acid 60 Chlorogenic acid 18_PM580 20_PM580 Quercetin 3-O-glucoside Luteolin 7-O-glucoside 1,5 Dicaff. acid+ Kaempferol 3-O-gluc. 39_PM592 Apigenin7-O-glucoside 43_PM Luteolin Apigenin Kaempferol 77_PM _PM243 51PM_ min Carduus nutans subsp. macrocephalus mau 254nm,4nm Neochlorogenic acid Chlorogenic acid 17aPM_610 18_PM580 Rutin Luteolin 7-O-glucoside Kaempferol 3-O-rutinoside 1,5 Dicaff. Acid+Kaempferol 3-O-gluc 39_PM592 Apigenin 7-O-glucoside Luteolin Apigenin Kaempferol 80_PM243 43_PM446 77_PM _PM min
16 mau 254nm,4nm Carduus cephalantus Neochlorogenic acid Chlorogenic acid 23_PM462 Luteolin 7-O-glucoside 1,5 Dicaff. Acid+Kaempferol 3-O-gluc 3,4 Dicaff. acid 46_PM608 4,5 Dicaff. acid 51_PM432 Luteolin Apigenin Kaempferol 77_PM _PM243 44_PM594 42_PM594 mau 254nm,4nm min Carduus pycnocephalus Chlorogenic acid Neochlorogenic acid 17_PM610 Rutin Quercetin 3-O-glucoside 23PM_462 Luteolin 7-O-glucoside Kaempferol 3-O-rutinoside 1,5 Dicaff. Acid+Kaempferol 3-O-gluc 3,5 Dicaff. acid 39_PM592 42_PM594 Diosmin 4,5 Dicaff acid Luteolin 51_PM432 Apigenin Kaempferol 77_PM _PM243 44_PM594 43_PM min
17 mau 254nm,4nm Ptilostemon casabonae Chlorogenic acid Neochlorogenic acid Quercetin 3-O-gl Rutin lucoside 1,5 Dicaff acid+kaempferol 3-O-gluc Kaempferol 3-O-rutinoside Apigenin 7-O-glucoside 43_PM446 4,5 Dicaff acid Luteolin Apigenin Kaempferol 80_PM243 55_PM616 58_PM716 3,5 Dicaff acid 47_PM min
18 Principalcomponentanalysis P.casabonae Carduus Species Carduus argyroa Biv. Carduus pycnocephalus L. Carduus cephalantusviv. Carduus nutans subsp. macrocephalus(desf.) Nyman Ptilostemon casabonae(l.) Greuter C.nutans C.nutans P.casabonae C.argyroa C.argyroa
19 Principalcomponentanalysis C.nutans C.argyroa Only Carduus species Species Carduus argyroa Biv. Carduus pycnocephalus L. Carduus cephalantusviv. Carduus nutans subsp. macrocephalus(desf.) Nyman C.pycnoc. C.argyroa C.pycnoc. C.nutans C.cephal. C.cephal.
20 Metabolite profile Identification of some compounds Statistical analysis differences between species Further statistical studies
21 BIOMOLECULAR ANALYSIS ITS gene Species 1. Carduus argyroa Biv. 2. Carduus pycnocephalus L. 3. Carduus cephalantusviv. 4. Carduus nutans subsp. macrocephalus(desf.) Nyman 5. Ptilostemon casabonae(l.) Greuter
22 Sequencing Electropherogram
23 Alignment of the forward and reverse sequences Alignment of the sequences belonging to each individuals CONSENSUS sequence
24 UPGMA tree Biomolecular markers
25 Sequences Discrimination between species Further genes analysis
26 CONCLUSIONS Identification of closely related thistle species in the genus Carduus as well as Ptilostemon This may suggest that both chemical and genetic differences are present between the considered species.
27 FUTURE PERSPECTIVES Run further statistical analysis and experiments Biologicalassaystest the extracts to verify the presence of biological activities
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