Chemical studies of cytotoxic natural products from toxic and edible plant sources
Doctoral thesis
Date
2024-04-19Metadata
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- Department of Chemistry [458]
Abstract
Denne doktorgradsavhandlingen fokuserer på isolering og identifikasjon av naturstoffer fra fire utvalgte planter, hvor den cytotoksiske aktiviteten til nyoppdagede og interessante naturstoffer ble testet på blodkreftceller og normale cellelinjer. I løpet av dette arbeidet var det mulig å isolere saponinene i Narthecium ossifragum, en plante assosiert med omfattende tilfeller av forgiftninger av gårdsdyr. Tre saponiner ble identifisert, i tillegg til deres sapogeninkjernestruktur sarsasapogenin aglykon. Cytotoksisiteten til disse forbindelsene ble testet mot normale celler og leukemiceller. Videre ble de fem nye naturstoffene di-C-glykosylflavonene chrysoeriol 6-C-β-arabinofuranoside-8-C-β-glukopyranoside, chrysoeriol 6-C-β-arabinopyranosyl-8-C-β-glukopyranoside, chrysoeriol 6-C-β-xylopyranosyl-8-C-β-galaktopyranoside, chrysoeriol 6-C-β-galaktopyranosyl-8-C-β-glukopyranoside og chrysoeriol 6-C-β-glukopyranosyl-8-C-β-galaktopyranoside isolert og karakterisert fra det metanoliske ekstraktet fra blomstene til N. ossifragum.
Den betydelige cytotoksiske aktiviteten til hovedsaponinet sarsasapogenin-3-O-(2'-O-β-glukopyranosyl-3'-O-α-arabinopyranosyl-β-galaktopyranoside) overfor nyreceller indikerer sterkt at dette saponinet er hovedårsaken til de observerte dødelige nyreskadene hos storfe som beiter på N. ossifragum. Studier av effekten av disse forbindelsene på biologiske membraner avdekket konsentrasjonsavhengig toksisitet, samt økt giftighet med økende antall sukkersubstituenter overfor både kreftceller og normale cellelinjer.
I tillegg ble den fytokjemiske sammensetningen av medisinplanten Osmunda regalis undersøkt, noe som førte til identifikasjon av 17 naturstoffer, hvorav 15 ble funnet for første gang i denne planten, inkludert 6 nye naturstoffer. De nye naturstoffene ble identifisert som flavonoidene kaempferol 3-O-(2''-O-(2'''-α-rhamnopyranosyl)-β-glukopyranosyl)-β-glukopyranoside, quercetin 3-O-(2''-O-(2'''-α-rhamnopyranosyl)-β-glukopyranosyl)-β-glukopyranoside, og kaempferol 3-O-(2''-O-(2'''-α-rhamnopyranosyl-6'''-O-(E)-caffeoyl-)-β-glukopyranosyl)-β-glukopyranoside, og de tre laktonene 3-metoksy-5-hydroxy-4-olid, 4-hydroxy-3-(3'-hydroxy-4'-(hydroksyetyl)-okstetrafuranon-5-metyl tetrahydropyranon, 4-O-(5-hydroxy-4-oksohexanoyl) osmundalakton. To av de isolerte laktonene, det kjente naturstoffet 5-hydroxy-2-hexen-4-oliden og det nye naturstoffet 4-O-(5-hydroxy-4-oksohexanoyl) osmundalakton) viste betydelig cytotoxisk aktivitet overfor MOLM13-celler, så vel som normale cellelinjer.
Naturproduktene i Cryptogramma crispa, også kjent som hestespreng, ble også isolert og strukturbestemt. I alt ble 15 naturstoffer isolert fra bladene til C. crispa, inkludert det nye naturstoffet 3-malonyl pteroside D. Pteroside-derivatene isolert fra dette plantematerialet viste selektiv moderat cytotoxisk aktivitet mot akutt myeloide leukemi MOLM13 cellelinjen, men ingen cytotoksisk aktivitet mot normale hjerte- og nyrecellelinjer.
I tillegg ble naturstoffene i kjørvel, vitenskapelig kjent som Anthriscus cerefolium, en urt som ofte brukes i norske storkjøkken, isolert og karakterisert. Det var mulig å isolere fire nye forbindelser fra denne plantekilden, deriblant det nye bisykliske laktonet 1,3-dikaffeoyl-5-malonyl-δ-kinidin. Denne planten er en rik kilde til fenoliske forbindelser, og det nye naturstoffet har en mild cytotoxisk aktivitet overfor normale cellelinjer og kreftcellelinjer.
En kombinasjon av avanserte NMR-spektroskopiske teknikker, CD spektroskopi og høyoppløselig massespektrometri ble brukt for strukturbestemmelsen av de 43 forbindelsene presentert i denne avhandlingen. Resultatene fra studiene av cytotoksisk aktivitet av disse naturstoffene motivere for vider og mer omfattende studier av deres cytotoksiske aktivitet og mekanismene for deres cytotoksisitet på cellulært og molekylært nivå. I motsetning til konklusjoner i eksisterende vitenskapelig litteratur viser resultatene i avhandlingen at sarsasapogenin-derivater fra N. ossifragum ikke kan brukes som virkestoffer i kreftlegemidler på grunn av signifikant ikke-selektiv cytotoksisitet overfor både kreftceller og normale cellelinjer. This PhD thesis focuses on the isolation and identification of natural compounds from four selected plant materials, where the cytotoxic activity in novel and interesting natural products was tested towards cancer and normal cell lines.
During this work, it was possible to isolate saponins from Narthecium ossifragum, a plant associated with poisoning of domesticated animals. Three saponins were identified, as well as their sapogenin core structure sarsasapogenin. The cytotoxic activities of these compounds were tested against normal cells and leukaemia cell lines. Moreover, the five previously undescribed di-C-glycosylflavones chrysoeriol 6-C-β-arabinofuranoside-8-C-β-glucopyranoside, chrysoeriol 6-C-β-arabinopyranosyl-8-C-β-glucopyranoside, chrysoeriol 6-C-β-xylopyranosyl-8-C-β-galactopyranoside, chrysoeriol 6-C-β-galactopyranosyl-8-C-β-glucopyranoside and chrysoeriol 6-C-β-glucopyranosyl-8-C-β-galactopyranoside were isolated and characterised from the methanolic extract from flowers of N. ossifragum.
The significant cytotoxicity of the main saponin sarsasapogenin-3-O-(2′-O-β-glucopyranosyl-3′-O-α-arabinopyranosyl-β-galactopyranoside) against kidney cells strongly indicate that this saponin is the primary cause of the observed lethal kidney damage in cattle that ingest N. ossifragum. The research also explores the effect of these compounds on biological membranes, revealing a concentration-dependent mechanism, as well an increasing toxicity with the increasing of the sugar moieties that leads to cell death.
In addition, the study examines the phytochemical composition of the medicinal plant Osmunda regalis, identifying 17 natural products, 15 of which were found in this species for the first time, which include 6 undescribed compounds. The new natural products identified were the flavonoids kaempferol 3-O-(2’’-O-(2’’’-α-rhamnopyranosyl)-β-glucopyranosyl)-β-glucopyranoside, quercetin 3-O-(2’’-O-(2’’’-α-rhamnopyranosyl)-β-glucopyranosyl)-β-glucopyranoside, and kaempferol 3-O-(2’’-O-(2’’’-α-rhamnopyranosyl-6’’’-O-(E)-caffeoyl-)-β-glucopyranosyl)-β-glucopyranoside, and the three lactones 3-methoxy-5-hydroxy-4-olide, 4-hydroxy-3-(3’-hydroxy-4’-(hydroxyethyl)-oxotetrafuranone-5-methyl tetrahydropyranone, 4-O-(5-hydroxy-4-oxohexanoyl) osmundalactone. Two of the isolated lactones (the known 5-hydroxy-2-hexen-4-olide, and the novel compound 4-O-(5-hydroxy-4-oxohexanoyl) osmundalactone) showed significant cytotoxic activity against MOLM13 cells, as well as normal cell lines.
This work also investigated the natural products of Cryptogramma crispa, also known as Parsley fern. The study reports the characterisation of 15 natural products isolated from the aerial parts of C. crispa, including a previously undescribed compound, 3-malonyl pteroside D. The pteroside derivatives isolated from this plant material showed selective moderate cytotoxic activity against the acute myeloid leukaemia MOLM13 cell line, but no cytotoxicity against normal heart and kidney cell lines.
Furthermore, the study Garden chervil, scientifically known as Anthriscus cerefolium, is an herb frequently used in large-scale Norwegian commercial kitchens. It was possible to isolated four new compounds from this plant source in which the previously undescribed compound 1,3-dicaffeoyl-5-malonyl-δ-quinide was isolated. This plant is a potent good source of phenolics, and the novel compounds have a mild cytotoxic activity towards normal and cancer cell lines.
A combination of advanced NMR spectroscopic techniques, CD spectroscopic and high-resolution mass spectrometry were used for the structure determination of the 43 compounds presented in this thesis. Results from the cytotoxic activity studies from these natural products motivate further and more extensive studies of their cytotoxic activity and the mechanisms of their cytotoxicity at the cellular and molecular levels. However, contrary to the conclusions that exist in the scientific literature, results presented in this work demonstrate that sarsasapogenin derivatives from N. ossifragum should not be used as anticancer agents because of their significant non-selective cytotoxicity towards cancer and normal cell lines.
Has parts
Paper I. Carpinteyro Díaz, A.E., Herfindal, L., Rathe, B.A., Sletta, K.Y., Vedeler, A., Haavik, S., & Fossen, T. (2019). Cytotoxic saponins and other natural products from flowering tops of Narthecium ossifragum L. Phytochemistry (Oxford), 164, 67–77. The article is available at: https://hdl.handle.net/1956/23761.Paper II. Carpinteyro Díaz, A.E., Haavik, S., Lunde, T.H.F., Fossen, T., Herfinal, L. (2023) Cytotoxic Saponins from the livestock-poisoning plant Narthecium ossifragum (L) – Death through destruction of biological membranes. Not available in BORA.
Paper III. Carpinteyro Díaz, A.E., Herfindal, L., Holmelid, B., Brede, C., Andersen, H.L., Vedeler, A., Fossen, T. (2023). Cytotoxic Natural Products from the Jurassic Relic Osmunda regalis L. Not available in BORA.
Paper IV. Carpinteyro Diaz, A.E., Herfindal, L, Andersen, H.L., & Fossen, T. Cytotoxic Natural Products Isolated from Cryptogramma crispa (L.) R. Br. Molecules. 2023; 28(23):7723. The article is available at: https://hdl.handle.net/11250/3124267.
Paper V. Slimestad, R., Rathe, B. A., Aesoy, R., Carpinteyro Diaz, A. E., Herfindal, L., & Fossen, T. (2022). A novel bicyclic lactone and other polyphenols from the commercially important vegetable Anthriscus cerefolium. Scientific Reports, 12(1), 7805–7805. The article is available at: https://hdl.handle.net/11250/3036765.