Christopher Rufus Sweeney and Faculty Mentor: Bradley Geary, Plant and Wildlife Sciences
Secondary metabolites are a result of natural evolutionary mechanism used to aid in plant development and defense, and the research of these metabolites have led to important discoveries in medicines, pesticides and even fossil fuels. In this project, we studied secondary metabolites of microorganisms in Mahonia aquifolium, otherwise known as Oregon grape. This plant is known to produce the chemical compound berberine as a secondary metabolite. Berberine has been shown to have similar effects to metformin and other diabetes treatments, as well as a host of other medicinal qualities. We looked at endophytic fungi and bacteria to see if they, too, produce berberine or similar organic compounds.
The project began with collecting Mahonia aquifolium from the Wasatch front near Nun’s Park. We then surface sterilized the plants and plated them onto water agar, then replated the fungi and bacteria onto either lysogeny agar or potato dextrose agar. We found about 13 species of fungi and 3 species of bacteria, the most notable of which was a distinctly yellow fungus which we deemed Oregon Grape 06 (OG 06). Because of its yellow color being consistent with the color of berberine, the chemical of interest, we decided to do further testing with this fungus.
We took OG 06 and put it in potato dextrose broth for 3 weeks. Then we filtered the broth to remove fungal material, after which we evaporated the broth to dryness. Then, we extracted the broth with dichloromethane to separate organic from aqueous materials. We rotary evaporated the organic mixture to dryness, then used TLC to further separate the materials. We collected analyte from the TLC, and used mass spectrometry to verify the molecular weight of chemical compound.
Additionally, we used a bioassay to test if OG 06 produces berberine. We took Staphylococcus aureus, which is susceptible to berberine, and plated it in a specially made agar with OG 06 to check for inhibition.
We hypothesized that endophytes of Oregon grape produce berberine or a berberine derivative. Since OG 06 was the designated fungus of interest because of the yellow compound it produced, we did the majority of our experimentation with OG 06. According to our results, our hypothesis could not be rejected. The TLC data showed that we had a chemical that matched quantitatively and qualitatively with our berberine standard.
We also had a mass spectrometry reading that was close but not identical to the berberine standard molecular weight.
With our bioassay, we noted that there was a zone of inhibition around OG 06 where the Staphylococcus aureus did not grow.
With all of our preliminary data, we cannot reject the hypothesis that our endophytic fungus (OG 06) is producing berberine or berberine derivative.
Based on our data, we believe that OG 06 is producing a derivative of berberine. This is not a surprising find, but it is nonetheless important. We believe that the production of barbering by both the plant and the endophytic fungus could mean that a genetic cassette was exchanged at some point in its evolutionary history together. This gene transfer is significant because, if it is happening it Oregon grape, it could be happening in other, more-difficult-to-isolate compounds, and could lead to less expensive forms of manufacturing pharmaceuticals and pesticides.
As Plant and Wildlife sciences continues to make breakthroughs in the study of plant genetics, this study has made a compelling case for looking further into the oft-overlooked world of endophytes and their secondary metabolites.