# Evaluation of _Sambucus nigra_ L. Biopotential as an Unused Natural Resource
## 3 Results and Discussion  
### Total Bioactive Compounds and Phenolic Components  
#### 3.1.1 Content of Total Bioactive Compounds  
The results of analysis of the total content of secondary metabolites in elderberry extracts are summarized in Figure 1.  
All examined extracts contained significant levels of bioactive components; 50% EtOH was generally more efficient solvent than water, while MAC was the most suitable technique for extraction biologically active compounds. Namely, 50% EtOH extracts produced by MAC are characterized by the highest total content of phenols, flavonoids, anthocyanins, and tannins. The amount of total phenols and anthocyanins in the 50% EtOH MAC extract was significantly different (\(p<0.05\)) from the content in MAE and UAE extracts, while the presence of total tannins did not differ significantly between MAC, MAE, and UAE extracts. Moreover, there was no statistically significant difference in the content of total phenols, anthocyanins, and tannins between MAE and UAE extracts. Regarding the content of total flavonoids, the MAE extract was the best source of total flavonoids, while somewhat lower content was observed in the MAC extract. The UAE extract was distinguished as the poorest source of total flavonoids. Regarding water extracts, the results showed that MAE was the most effective technique for the isolation of biologically valuable compounds. These outcomes, based on the usage of solvents with different polarity are in the line with previous results: 50% EtOH was demonstrated as more efficient than H\({}_{2}\)O [26].  
The presented results indicated that MAC was an extraction technique whose application resulted in the most effective isolation of biologically active molecules. The highlighting of MAC as the dominant extraction technique for the extraction of secondary metabolites could be related to the duration of the isolation process and the conditions under which the isolation of biologically active compounds was carried out. Namely, MAC lasted for 72 h, without the influence of temperature, so it can be assumed that the degradation of thermolabile phenolic compounds was avoided during this process, and, on the other hand, the length of the process of isolation resulted in the maximal release of non-phenolic structures from the plant matrix. Accordingly, the influence of temperature in MAE and the strength of ultrasound in the UAE could cause degradation of thermolabile compounds [27]. Comparing obtained results with previously reported content of phenolic compounds in the other berry fruits, it could be noticed that examined extracts have higher content than raspberry and blueberry [28], but still inferior to the phenolics amount in blackberry [29]. The diverse content of bioactive compounds in berries could be caused by different methods of sample preparation, climatic and geographic factors, as well as the composition of the soil on which these plants grew [30].  
Figure 1: Total phenols, flavonoids, anthocyanins, and tannins content. GAE: gallic acid equivalent: RE: rutin equivalent; Cy-3-G: cyanidin-3-O-glucoside equivalent. Values are reported as mean \(\pm\) SD of three parallel experiments. Means followed by different letters (a=e) differ significantly at \(p<0.05\). MAE—microwave-assisted extraction technique; UAE—ultrasound assisted extraction technique; MAC—maceration.  
The spectrophotometric methods, which have been used to estimate the total content of phenolic compounds, are not selective, so further analysis was performed by using the accurate and sensitive LC-MS/MS technique.