Optimized isolation of 7,7′-biphyscion starting from Cortinarius rubrophyllus, a chemically unexplored fungal species rich in photosensitizers

Abstract

Mushrooms such as the dermocyboid Cortinarius rubrophyllus are characterized by strikingly colorful fruiting bodies. The molecular dyes responsible for such colors recently experienced a comeback as photoactive compounds with remarkable photophysical and photobiological properties. One of them—7,7′-biphyscion—is a dimeric anthraquinone that showed promising anticancer effects in the low nanomolar range under blue-light irradiation. Compared to acidic anthraquinones, 7,7′-biphyscion was more efficiently taken up by cells and induced apoptosis after photoactivation. However, seasonal collection of mushrooms producing this compound, low extraction yields, and tricky fungal identification hamper further developments to the clinics. To bypass these limitations, we demonstrate here an alternative approach utilizing a precursor of 7,7′-biphyscion, i.e., the pre-anthraquinone flavomannin-6,6′-dimethyl ether, which is abundant in many species of the subgenus Dermocybe. Controlled oxidation of the crude extract significantly increased the yield of 7,7′-biphyscion by 100%, which eased the isolation process. We also present the mycochemical and photobiological characterization of the yet chemically undescribed species, i.e. C. rubrophyllus. In total, eight pigments (1–8) were isolated, including two new glycosylated anthraquinones (1 and 2). Light-dependent generation of singlet oxygen was detected for the first time for emodin-1-O-β-d-glucopyranoside (3) [photophysical measurement: Φ∆ = 0.11 (CD3OD)]. Furthermore, emodin (7) was characterized as promising compound in the photocytotoxicity assay with EC50-values in the low micromolar range under irradiation against cells of the cancer cell lines AGS, A549, and T24. Graphical abstract

A convenient separation strategy for fungal anthraquinones by centrifugal partition chromatography

“…For the annotation of secondary metabolites, substance‐specific parameters (i.e., molecular weight, absorption maxima, and retention time) were determined and compared with reference compounds and in‐house data [ 4 , 5 ].…”

“…The dried fruiting bodies of C. malicorius (batch 2020) and C. rubrophyllus were extracted as follows: Finely ground biomaterial was extracted with methanol (V = 30 mL, n = 4) using an ultrasonic bath (5 min per extraction step) followed by filtration through a folded paper filter (particle retention: 10-20 μm). Respective filtrates were combined, solvents were removed by vacuum rotary evaporation at 40 For the annotation of secondary metabolites, substancespecific parameters (i.e., molecular weight, absorption maxima, and retention time) were determined and compared with reference compounds and in-house data [4,5].…”

“…A dimeric AQ isolated from the marsh webcap (Cortinarius uliginosus) was shown to induce blue light-dependent cell death in various cancer cell lines in Article Related Abbreviations: AQ, anthraquinone; CPC, centrifugal partition chromatography; DMSO, dimethyl sulfoxide; LP, lower phase; MeOH, methanol; MLLP, multiple liquid-liquid partition; PT, pigmentation type; SI, electronic supplementary information; UP, upper phase the nanomolar range while being inactive in the absence of light [4]. Subsequently, several other fungal AQs (e.g., monomeric AQs, AQ carboxylic acids, dimeric AQs, and even glycosylated AQs) have been identified as promising photosensitizers [5].…”

“…A dimeric AQ isolated from the marsh webcap ( Cortinarius uliginosus ) was shown to induce blue light‐dependent cell death in various cancer cell lines in the nanomolar range while being inactive in the absence of light [ 4 ]. Subsequently, several other fungal AQs (e.g., monomeric AQs, AQ carboxylic acids, dimeric AQs, and even glycosylated AQs) have been identified as promising photosensitizers [ 5 ].…”

Photochemical defense as trait of fungi from Cortinarius subgenus Dermocybe

“…Plenty of photosensitizer-producing plants are known compared to fungi, especially basidiomycetes, where only two dermocyboid Cortinarii were studied in detail (i.e., C. uliginosus [6] and C. rubrophyllus [14]). Nevertheless, photoactivity is not limited to these two species of Cortinarius; it was also detected in seven other species belonging to the colorful Cortinarius subgenus Dermocybe [5,6].…”