Antimicrobial and cytotoxic properties of semisynthetic betulin derivatives

Betulin (lup-20(29)-ene-3β, 28-diol) is an abundant naturally occurring triterpene, which is the principal extractive of outer birch bark. A collection of 65 structurally diverse semisynthetic betulin derivatives was screened against six different microbes, Enterobacter aerogenes, Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa, Staphylococcus aureus and a fungal strain Candida albicans, using microdilution assays based on CLSI and EUCAST guidelines. Primary antimicrobial screening led to the identification of six compounds showing promising antimicrobial properties (inhibition > 70% against one or more microbial strains). According to the dose-response results, the betulin derivative SC047 was the most active, showing MIC of 6.25µM against two Gram-positive bacteria, E. faecalis and S. aureus. However, the activity of this compound was affected by albumin binding, which was demonstrated by the loss of activity in the host-pathogen co-culture assay as well as in the antibacterial assay in the presence of increased concentration of albumin. Furthermore, cytotoxicity assessment based on ATP measurement of human hepatocyte cell culture after 24h exposure to the compounds, showed that compounds SC013 and SC062 displayed cytotoxicity towards hepatocytes, showing IC₅₀ of 25.0µM, and 15.8µM, respectively. In addition, the IC₅₀ value of 55.9µM for compound SC047 was determined.

The current study presents an insight into using betulin scaffold for developing derivatives with antibacterial potential, and furthermore the necessity of in-depth analysis of found actives through selectivity profiling and functional characterization. Moreover, the results demonstrate the importance of taking a multidimensional approach when studying biological activities of natural compounds and their derivatives.