Effect of cultivar, maturation stage, and year on sugar and phenolic composition of elderberries.

BACKGROUND: The chemical composition, phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO) and peroxidase (POD) activity of the three main Portuguese elderberry cultivars were determined for the first time through five stages of maturation, in different harvesting years, to gain a deeper understanding of the effect of climatic conditions and enzymatic activity involved in the synthesis and degradation of phenolic compounds on the final quality of elderberries. RESULTS: Simple sugar and anthocyanin content increased with maturation but total acidity and flavonoids content decreased, and cinnamic acids did not show a clear trend. Climatic conditions seem to have a decisive influence on the elderberry maturation, namely the total number of hot (>30 °C) days. The PAL, PPO, and POD activity can explain the differences observed in elderberry phenolic content. CONCLUSION: These results highlighted the influence of climatic conditions in each harvesting season on elderberry development and quality. © 2022 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Elderberry (Sambucus nigra L.) extracts promote anti-inflammatory and cellular antioxidant activity.

Despite the high value of Portuguese elderberries, recognized for decades by European markets, only a few studies address their beneficial effects at cellular level. Aiming to explore the anti-inflammatory and the cellular antioxidant potential characterized extracts from the three main Portuguese elderberry cultivars (Sabugueiro, Sabugueira, Bastardeira) were used. Lipopolysaccharide-stimulated RAW 264.7 cells pre-exposed to elderberry extracts exhibited dose-dependent inhibition of nitric oxide release, evidencing anti-inflammatory activity. Concerning cellular antioxidant protection, HepG2 and Caco-2 cells pre-exposure to elderberry extracts (50 µg/mL) prevented up-to 90 % of tert-butyl hydroperoxide (t-BOOH)-induced toxicity. In Caco-2 cells, elderberry extracts prevented glutathione depletion, reactive oxygen species production, abnormal morphological changes and DNA fragmentation, in response to t-BOOH oxidative insult. Results demonstrated that elderberries have high potential in reducing cellular oxidative stress as well as in preventing inflammatory processes. Thus, elderberries have high potential as health promoters, acting as functional foods or as sources of nutraceuticals.

Major anthocyanins in elderberry effectively trap methylglyoxal and reduce cytotoxicity of methylglyoxal in HepG2 cell line.

The accumulation of advanced glycation end-products (AGEs) in the body is implicated in numerous diseases, being methylglyoxal (MGO) one of the main precursors. One of the strategies to reduce AGEs accumulation might be acting in an early stage of glycation by trapping MGO. Thus, this work aimed to evaluate, for the first time, the potential of elderberries polyphenols to trap MGO, access the formation of MGO adducts, and evaluate the cytoprotection effect in HepG2 and Caco-2 cells. The results demonstrated that monoglycosylated anthocyanins (cyanidin-3-glucoside and cyanidin-3-sambubioside) are very efficient in trapping MGO, forming mono- and di-adducts. Quercetin-3-glucoside and quercetin-3-rutinoside reacted slowly, while diglycosylated anthocyanins did not react. The trapping of MGO by elderberry monoglycosylated anthocyanins significantly decreased the MGO cytotoxicity in HepG2 cells (∼70 % of cell viability), while the effect in Caco-2 cells was lower (∼50 %). Thus, elderberry phenolics present antiglycation potential by trapping MGO.

Molecular Physicochemical Properties of Selected Pesticides as Predictive Factors for Oxidative Stress and Apoptosis-Dependent Cell Death in Caco-2 and HepG2 Cells.

In this work, three pesticides of different physicochemical properties: glyphosate (GLY, herbicide), imidacloprid (IMD, insecticide), and imazalil (IMZ, fungicide), were selected to assess their cytotoxicity against Caco-2 and HepG2 cells. Cell viability was assessed by the Alamar Blue assay, after 24 and 48 h exposure to different concentrations, and IC50 values were calculated. The mechanisms underlying toxicity, namely cellular reactive oxygen species (ROS), glutathione (GSH) content, lipid peroxidation, loss of mitochondrial membrane potential (MMP), and apoptosis/necrosis induction were assessed by flow cytometry. Cytotoxic profiles were further correlated with the molecular physicochemical parameters of pesticides, namely: water solubility, partition coefficient in an n-octanol/water (Log Pow) system, topological polar surface area (TPSA), the number of hydrogen-bonds (donor/acceptor), and rotatable bonds. In vitro outputs resulted in the following toxicity level: IMZ (Caco-2: IC50 = 253.5 ± 3.37 µM, and HepG2: IC50 = 94 ± 12 µM) > IMD (Caco-2: IC50 > 1 mM and HepG2: IC50 = 624 ± 24 µM) > GLY (IC50 >>1 mM, both cell lines), after 24 h treatment, being toxicity time-dependent (lower IC50 values at 48 h). Toxicity is explained by oxidative stress, as IMZ induced a higher intracellular ROS increase and lipid peroxidation, followed by IMD, while GLY did not change these markers. However, the three pesticides induced loss of MMP in HepG2 cells while in Caco-2 cells only IMZ produced significant MMP loss. Increased ROS and loss of MMP promoted apoptosis in Caco-2 cells subjected to IMZ, and in HepG2 cells exposed to IMD and IMZ, as assessed by Annexin-V/PI. The toxicity profile of pesticides is directly correlated with their Log Pow, as affinity for the lipophilic environment favours interaction with cell membranes governs, and is inversely correlated with their TPSA; however, membrane permeation is favoured by lower TPSA. IMZ presents the best molecular properties for membrane interaction and cell permeation, i.e., higher Log Pow, lower TPSA and lower hydrogen-bond (H-bond) donor/acceptor correlating with its higher toxicity. In conclusion, molecular physicochemical factors such as Log Pow, TPSA, and H-bond are likely to be directly correlated with pesticide-induced toxicity, thus they are key factors to potentially predict the toxicity of other compounds.

Effect of harvesting year and elderberry cultivar on the chemical composition and potential bioactivity: A three-year study.

Sambucus nigra is one of the richest sources of anthocyanins and other polyphenols being used industrially as a source of antioxidants, colorants, and bioactives. Although cultivars can influence elderberry composition, no study has addressed the effect of harvesting year on elderberries composition and bioactivity. The composition of the main Portuguese cultivars, "Sabugueiro", "Sabugueira" and "Bastardeira", were evaluated during three consecutive years. Harvesting year had a stronger influence on the chemical composition than cultivars, including total sugars, anthocyanins, and phenolic compounds, being related to the different climatic conditions, especially water status. "Bastardeira" was the best cultivar concerning total soluble solids, anthocyanins, polyphenols, and antioxidant activity, but containing a lower total free sugar content compared to "Sabugueiro". The results obtained in this study provide novel information from a nutritional perspective and for breeding programs aiming to select cultivars with enhanced levels of health-promoting compounds or for other industrial applications of elderberries.