Seasonal, Organ-, and Location-Dependent Variations in the Alkaloid Content of Pachysandra terminalis Investigated by Multivariate Data Analysis of LC-MS Profiles

<i>Pachysandra terminalis</i> (<i>P. terminalis</i>), a plant belonging to the Buxaceae family, is known as a great source of aminosteroid alkaloids. In a previous communication, we reported on the isolation of a variety of aminosteroids from <i>P. terminalis,</i> which presented interesting activity against the protozoan pathogens, <i>Trypanosoma brucei rhodesiense</i> and <i>Plasmodium falciparum</i>. In the present study, variations in the alkaloid profile of <i>P. terminalis</i> related to seasonal changes as well as differences between plant organs (leaves and twigs) and between plant populations were investigated to prioritize candidates for targeted isolation in further studies. For this purpose, sample material of <i>P. terminalis</i> was collected from the two nearby populations in monthly intervals over one year. The ethanolic (75%) extracts were analyzed using UHPLC/+ESI-QqTOF-MS/MS, and the resulting data converted to variables encoding the intensity of MS signals in particular <i>m/z</i> and retention time (<i>t</i>_R) intervals over the chromatographic runs. The very large and complex data matrix of these <<i>t</i>_R:<i>m/z</i>> variables was evaluated using multivariate data analysis, especially principal component analysis (PCA) and volcano plot analysis of <i>t</i>-test data. The results of these analyses, for the first time, allowed a holistic analysis of variation in the alkaloid profiles in <i>P. terminalis</i> organs over the vegetation period. The evaluation of the PCA scores and loadings plots of principal components 1 through 3, as well as of volcano plots, highlighted 25 different compounds, mostly identified as aminosteroid alkaloids, that were most relevant for the differences between leaves and twigs and between the two populations and mainly determined the changes in their chemical profiles over the vegetation period.