Exudates of Picea abies, Pinus nigra, and Larix decidua: Chromatographic Comparison and Pro-Migratory Effects on Keratinocytes In Vitro

Abstract

Balms and resins of Picea abies, Larix decidua, and Pinus nigra are traditionally used to treat wounds. Three chromatographic techniques differing in separation capacity and technical demands were employed to distinguish among these plant exudates. A TLC method was established for fingerprint comparison, providing a quick overview of a large number of samples at low cost. HPLC-DAD (RP18) and UHPSFC-DAD (Torus 2-Picolylamin), hyphenated to ESI-MS, represented orthogonal chromatographic systems with high separation performance. The developed methods allow for the separation and detection of major and minor constituents belonging to different compound classes (phenyl carboxylic acids, lignans, diterpene resin acids). The qualitative compositions of the diterpene resin acids, the main compounds in the exudates, were comparable in all three genera. Differences were detected in the distribution of hydroxylated diterpene resin acids, pinoresinol, and hydroxycinnamic acids. The three tested chromatographic systems with varying demands on lab equipment offer appropriate tools for the quality assessment of Picea abies, Larix decidua, and Pinus nigra. The extracts were furthermore tested at three different concentrations (10 µg/mL, 3 µg/mL, and 1 µg/mL) for boosted re-epithelialization, a crucial step in the wound-healing process, in an in vitro HaCaT keratinocyte-based scratch assay. Lysophosphatidic acid (LPA, 10 µM) and extracts of several medicinal plants well known for their wound-healing properties (birch, marigold, St. John’s wort, manuka honey) were used as positive controls. Picea abies and Pinus nigra showed concentration dependency; significant activity was measured for Larix decidua at 3 µg/mL.