Phytochemical analysis and in vitro anti-proliferative activity of Viscum album ethanolic extracts

Abstract

Background: Viscum album L. (Santalaceae), commonly known as mistletoe, is a hemiparasitic plant traditionally used in complementary cancer treatment. Its antitumor potential is mostly attributed to the presence of aqueous soluble metabolites; however, the use of ethanol as solvent also permits the extraction of pharmacological compounds with antitumor potential. The clinical efficacy of mistletoe therapy inspired the present work, which focuses on ethanolic extracts (V. album "mother tinctures", MT) prepared from different host trees. Methods: Samples from three European subspecies (album, austriacum, and abietis) were harvested, and five different V. album-MT strains were prepared. The following phytochemical analyses were performed: thin layer chromatography (TLC), high-performance liquid chromatography (HPLC) and liquid chromatography-high resolution mass spectrometry (LC-HRMS). The proliferation assay was performed with WST-1 after incubation of tumor (Yoshida and Molt-4) and fibroblast cell lines (NIH/3 T3) with different MT concentrations (0.5 to 0.05% v/v). The cell death mechanism was investigated by flow cytometry (FACS) using Annexin V-7AAD. Results: Chemical analyses of MT showed the presence of phenolic acids, flavonoids and lignans. The MT flavonoid and viscotoxin contents (mg/g fresh weight) were highest in Quercus robur (9.67 ± 0.85 mg/g) and Malus domestica (3.95 ± 0.58 mg/mg), respectively. The viscotoxin isoform proportions (% total) were also different among the VA subspecies with a higher content of A3 in V. album growing on Abies alba (60.57 ± 2.13). The phytochemical compounds as well as the viscotoxin contents are probably related to the antitumor effects of MT. The cell death mechanisms evaluated by colorimetric and FACS methodologies involved necrotic damage, which was host tree-, time-and dose-dependent, with different selectivity to tumor cells. Mother tincture from V. album ssp. abietis was the most effective at inducing in vitro cellular effects, even when incubated at the smallest concentration tested, probably because of the higher content of VT A3. Conclusion: Our results indicate the promising antitumor potential of Viscum album ethanolic extracts and the importance of botanical and phytochemical characterization for in vitro anti-proliferative effects.

Metabolomics by UHPLC-Q-TOF Reveals Host Tree-Dependent Phytochemical Variation in Viscum album L.

“…album samples ( Table 2 ). These compounds were also previously described in the Viscum album species [ 18 , 20 , 21 , 22 , 23 ]; however, they were not related to the differentiation of the three clusters that were described in the PLS-DA analysis.…”

“…Since the plant metabolism is highly age-dependent and many metabolite levels are altered during the gradual plant ageing, the harvest standardization was conducted in order to collect plant material of the same level of development, regarding general morphological characteristics. For this work, two 1-year-old leaves and one 2-year-old leaf, one stem of each age (1- and 2-year-old), and three berries, were harvested on each mistletoe plant, as previously described by Holandino and coworkers [ 21 ]. The age of the V. album bush was estimated by counting the number of nodes on the longest branch of mistletoe.…”

Thermoresponsive Hydrogel Containing Viscum album Extract for Topic and Transdermal Use: Development, Stability and Cytotoxicity Activity

“…The influence of the host tree on the cytotoxic potential of mistletoe is an exciting research topic that has been previously highlighted when five different ethanolic extracts prepared from the three European subspecies (album, austriacum, and abietis) were evaluated by an in vitro model [15]. These studies emphasize the antitumoral potential of ethanolic soluble substances from VA for cancer pharmacotherapy.…”

Chemical‐activity‐based quality marker screening strategy for Viscum articulatum

“…Further, oleanolic acid (OA), a potent drug used clinically as an anti-inflammatory agent (Zheng et al, 2015), is the only indicator. In terms of bioactivity, alcohol extracts of the plant or related species also possess some antibacterial, antioxidant, antiinflammatory, and anticancer activities (Chamlagai & Singh, 2016;de Oliveira Melo et al, 2018;Erdönmez et al, 2018;Garg et al, 2013;Holandino et al, 2020;Nag et al, 2016;Stefanucci et al, 2020;, whereas water extracts are considered to possess antioxidant, hypoglycemic, and lipidlowering activity (Mishra et al, 2018;. However, the specific active ingredients are not yet known.…”