Quantitative and qualitative characterization of tree bark extracts

Tree bark contains diverse secondary metabolites with potential applications in pharmaceuticals, cosmetics, and nutraceuticals. Bark extracts have been historically used in medicine, with Betula pendula and pubescens extracts approved by the European Medicines Agency (EMA) in 2022 for epidermolysis bullosa treatment under the brand Fisuvez®, attributed to the triterpenes betulin, betulinic acid, and lupeol. Additionally, extracts from oak (Quercus spp.), pine (Pinus maritima/Pycnogenol®), and alder leaves have demonstrated anti-inflammatory and wound-healing properties, used both topically and as dietary supplements.
This study aimed to qualitatively and quantitatively characterize the secondary metabolite composition of bark extracts from six European tree species: alder, birch, beech, oak, cherry, and pine. A high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF) method was optimized to balance separation efficiency and analysis time, enabling comprehensive metabolite profiling.
Soxhlet extraction was performed on birch bark obtained from two sources (tea and horse supplement) using ethanol, methanol, and water. Qualitative analysis revealed distinct metabolic profiles between aqueous and alcoholic extracts, while no significant differences were observed between the two bark sources. Quantitative analysis using HPLC triple quadrupole mass spectrometry (HPLC-Triple Quad) identified fourteen target analytes, with flavonoids such as catechin and epicatechin present in the highest concentrations in birch bark.
To further explore the metabolite composition, MS/MS fragmentation experiments using HPLC-drift tube ion mobility quadrupole time-of-flight mass spectrometry (HPLC-DT-IM-Q-TOF-MS) were conducted, contributing to the identification of additional suspected analytes. Statistical analyses, including heatmaps and principal component analysis (PCA), confirmed species-specific metabolic fingerprints, with alder, birch, and beech exhibiting the highest metabolite diversity.
A stability study on birch bark extracts intended for topical application in the OxiWoundWood project indicated microbial contamination after five weeks of room temperature storage, suggesting the necessity for refrigerated storage.
The findings provide a refined methodological framework for tree bark extract characterization and highlight Soxhlet extraction’s suitability while underscoring the need for further optimization of the extraction solvent as well as the extraction method itself.