Dissertations, Theses, and Capstone Projects

Date of Degree

9-2026

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy

Program

Biology

Advisor

Edward J. Kennelly

Committee Members

Renuka P. Sankaran

Nathalia G. Holtzman

Dake Zhao

Matthew Devany

Subject Categories

Biochemistry | Biology | Plant Sciences

Keywords

American Aconitum, diterpenoid alkaloids, UPLC-qTof-MS, Cell Painting, cardiotoxicity

Abstract

Analysis of Diterpenoid Alkaloids from American Aconitum Using LC-MS Metabolomic Methods

by

Yi Zhao

Advisor: Edward J. Kennelly

Aconitum plants have been used as a traditional medicine in Asian countries for millennia to treat pain, rheumatism, syncope, fever, and other conditions. These therapeutic effects are mainly due to the bioactive diterpenoid alkaloids accumulated in the roots. Comparing with the large records and studies of Asian Aconitum and their medicinal use and the broad bioactive compounds, American Aconitum has been rarely investigated, not to mention how American Aconitum has been used medicinally. Based on previous phylogenetic and chemotaxonomic studies of Aconitum, we hypothesis that American Aconitum also contain bioactive diterpenoid alkaloids. This project aims to investigate the chemical profile from American Aconitum and discover the bioactive diterpenoid alkaloids in them.

In the first study, the bioactivity of American Aconitum was tested using a high-throughput phenotypic screening assay (Cell Painting), and the bioactive compounds in two American species (A. columbianum and A. uncinatum) were predictively targeted based on a systematic networking strategy integrating both mass spectrometry data and biological profiles. The chemical profiles of four different plant parts of two American Aconitum species were obtained by ion mobility mass spectrometry and compared with two Asian (A. carmichaelii and A. fischeri), and one European species (A. napellus). Biological screening, image analysis and feature extraction were performed on Aconitum extracts using the Cell Painting assay. The results provided 2,090 unique morphological features per extract, which was further reduced to 429. In conjunction with 4,400 chemicals from a library with known mechanisms of action, 198 unique hierarchical clusters were established. An overall activity heuristic called CP score was calculated for each sample. After integrating CP score and spectrometric data, a network filtered for higher CP scores was constructed and the compounds with high activity were tentatively identified. The network contained mostly American Aconitum species, suggesting that these understudied plants produce useful bioactive compounds.

In the second study, the chemistry and bioactivity of American Aconitum was investigated, since they also produce diterpenoid alkaloids that possess toxicity. The chemical profiles of three American Aconitum species, A. delphinifolium, A. columbianum, and A. uncinatum, and their four individual plant parts (roots, stems, leaves, and flowers) were studied using UPLC-qTof-MSe technique, and compared to Asian and European species. Multivariate including PCA and OPLS-DA were used to analyze the chemical profile. PCA suggests that American Aconitum is more chemically similar to Asian than European species. Using OPLS-DA, 29 unique marker compounds were found in American Aconitum, but not in the Asian or European species we studied. Nineteen marker compounds were tentatively identified using Progenesis Qi and reference standards, including proved analgesic compounds such as talatisamine and 14-benzoyltalatisamine and other bioactive compounds. The results suggest that American Aconitum is a potential source for lead drugs. The cardiotoxicity of American Aconitum root extracts (200 µg/mL) was assessed using a zebrafish embryo model, and compared to the most widely used Asian species, A. carmichaelii.  Aconitum delphinifolium did not show any cardiotoxicity in this zebrafish model, as compared to A. carmichaelii and the control compound, aconitine. However, A. columbianum and A. uncinatum induced some morphological heart differences, but not as profoundly as A. carmichaelii.

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