Developing a Whole Plant Artemisia annua Antimalarial Therapeutic: pACT

The GRAS plant Artemisia annua L. produces the sesquiterpene lactone, artemisinin. The current therapy for malaria is artemisinin + an older drug: artemisinin combination therapy (ACT). In Plasmodium chabaudi-infected mice, dried leaves of A. annua are more potent than equal amounts of pure artemisinin and may also prevent artemisinin drug resistance from emerging. This whole plant therapy is pACT: plant-based artemisinin combination therapy. Pharmacokinetics in healthy and infected mice given either pure artemisinin or pACT is different and showed that \u3e 40 fold more artemisinin enters the blood when plant material is present; plant matrix enhanced bioavailability into serum. Dried leaves as capsules or tablets given to African malaria patients were also efficacious. Flavonoids, phenolic acids, monoterpenes and other artemisinic metabolites found in the plant have mild antimalarial activity. Some may synergize with artemisinin to enhance its efficacy. In simulated digestion studies the effects of cellulose and gelatin capsules, sucrose, 4 oils, and 3 staple grains (rice, corn, and millet) were studied to determine their effect on AN and flavonoid release into the liquid phase of the intestinal stage of digestion. Compared to pACT alone: sucrose and oil enhanced release of flavonoids by 100%, but artemisinin was unaffected; both capsule types, and corn and millet meal significantly reduced artemisinin release, but had no effect on flavonoids. From field trials in MA, it was estimated that \u3e 500,000 patients could be treated from plants grown on 1 ac of land. Analysis of 10 crops of the high artemisinin-producing WPI clone of A. annua grown under different field and lab conditions showed there was consistent production of artemisinin at about 1.4% DW. Together these results show how a simple herbal remedy could be used as an efficacious, inexpensive, controlled and sustainable orally delivered therapeutic for treating malaria and other artemisinin-susceptible diseases

AtSUC1 ROOT EXPRESSION AND SUCROSE RESPONSE LEADING TO ANTHOCYANIN ACCUMULATION

University of Minnesota Ph.D. dissertation. December 2019. Major: Plant and Microbial Biology. Advisors: John Ward, Neil Olszewski. 1 computer file (PDF); v, 112 pages.Previous research indicated that AtSUC1 root expression is controlled by intragenic sequences. The 5’ upstream region (promoter) of AtSUC1 directs pollen and trichome expression, but not root expression. However, the whole AtSUC1 gene can drive root expression and sucrose-induced root expression. Here I show that root expression of AtSUC1 is controlled by the interaction between the promoter and its two short introns. Deletion of either intron from whole-gene-GUS constructs resulted in no root expression, showing that both introns are required. The two introns in tandem, fused to GUS, produce high constitutive expression throughout the vegetative parts of the plant. When combined with the promoter, the expression driven by the introns is reduced and localized to the roots. AtSUC1 expression is also induced by exogenous sucrose, and AtSUC1 is also required for sucrose-induced anthocyanins (Sivitz et al., 2008). Anthocyanin accumulation due to high sucrose was lesser in the AtSUC1 mutant compared to Col-0 wild type. A whole-gene-GUS construct expressing a non-functional AtSUC1 (D152N) mutant, that is transport inactive, was defective in sucrose-induced AtSUC1 expression and anthocyanins accumulation when expressed in an atsuc1-null background. The results indicated that sucrose uptake via AtSUC1 is required for sucrose-induced AtSUC1 expression and anthocyanin accumulation, and that the site for sucrose detection is intracellular

Root and Shoot Elicitation in Artemisia annua: Chitosan and Salicylic Acid Effect on Artemisinin and Flavonoid Biosynthesis

Our project analyzed root and shoot elicitation of the anti-malarial drug-producing plant, Artemisia annua, using the elicitors chitosan and salicylic acid. We accomplished this by applying chitosan and salicylic acid to the roots and shoots of hydroponically grown A. annua and harvesting both the tips of the apical meristem and the third through fifth leaves from the tips. The antimalarial drug, artemisinin, and flavonoids that act synergistically with the drug were extracted and quantified using GC/MS and an aluminum chloride assay, respectively. We observed higher concentrations of artemisinin in the tips of plants treated with chitosan via the roots indicating chitosan elicits artemisinin biosynthesis and that the roots play an important role in the production of artemisinin

Ex Vitro Digestion Study of Artemisia annua as a Whole Plant Treatment for Malaria

In this study, an ex vitro digestion system was used to simulate the digestion of the whole plant Artemisia annua to gain insight into how artemisinin, the key drug component in malaria treatment, and synergistic flavonoids become bioavailable during digestion. Various delivery methods and staple foods were combined with the plant material for digestion. This study found that sucrose, canola oil, and white rice did not reduce the amount of artemisinin released in the intestinal liquid fraction while vegetarian and gelatin capsules showed a significant reduction in artemisinin release. The sucrose and canola oil A. annua delivery methods also exhibited significantly high flavonoid release. High artemisinin and flavonoid content in the intestinal liquid fraction indicates bioavailability

Worcester Rain Gardens: Developing Promotional Materials for Rain Gardens in the City of Worcester

I worked in a group in partnership with the Mayor's Office of Worcester to create promotional materials for rain gardens in the city. We researched established design principles, analyzed common practices and conducted interviews in order to produce an effective brochure, website and video. In the process we ran productive meetings with different constituents and concluded by delivering a professional presentation to various stakeholders and a substantial academic report

AutonoMS: Automated Ion Mobility Metabolomic Fingerprinting

Automation is dramatically changing the nature of laboratory life science. Robotic lab hardware that can perform manual operations with greater speed, endurance, and reproducibility opens an avenue for faster scientific discovery with less time spent on laborious repetitive tasks. A major bottleneck remains in integrating cutting-edge laboratory equipment into automated workflows, notably specialized analytical equipment, which is designed for human usage. Here we present AutonoMS, a platform for automatically running, processing, and analyzing high-throughput mass spectrometry experiments. AutonoMS is currently written around an ion mobility mass spectrometry (IM-MS) platform and can be adapted to additional analytical instruments and data processing flows. AutonoMS enables automated software agent-controlled end-to-end measurement and analysis runs from experimental specification files that can be produced by human users or upstream software processes. We demonstrate the use and abilities of AutonoMS in a high-throughput flow-injection ion mobility configuration with 5 s sample analysis time, processing robotically prepared chemical standards and cultured yeast samples in targeted and untargeted metabolomics applications. The platform exhibited consistency, reliability, and ease of use while eliminating the need for human intervention in the process of sample injection, data processing, and analysis. The platform paves the way toward a more fully automated mass spectrometry analysis and ultimately closed-loop laboratory workflows involving automated experimentation and analysis coupled to AI-driven experimentation utilizing cutting-edge analytical instrumentation. AutonoMS documentation is available at https://autonoms.readthedocs.io

AutonoMS: Automated Ion Mobility Metabolomic Fingerprinting

Automation is dramatically changing the nature of laboratory life science. Robotic lab hardware that can perform manual operations with greater speed, endurance, and reproducibility opens an avenue for faster scientific discovery with less time spent on laborious repetitive tasks. A major bottleneck remains in integrating cutting-edge laboratory equipment into automated workflows, notably specialized analytical equipment, which is designed for human usage. Here we present AutonoMS, a platform for automatically running, processing, and analyzing high-throughput mass spectrometry experiments. AutonoMS is currently written around an ion mobility mass spectrometry (IM-MS) platform and can be adapted to additional analytical instruments and data processing flows. AutonoMS enables automated software agent-controlled end-to-end measurement and analysis runs from experimental specification files that can be produced by human users or upstream software processes. We demonstrate the use and abilities of AutonoMS in a high-throughput flow-injection ion mobility configuration with 5 s sample analysis time, processing robotically prepared chemical standards and cultured yeast samples in targeted and untargeted metabolomics applications. The platform exhibited consistency, reliability, and ease of use while eliminating the need for human intervention in the process of sample injection, data processing, and analysis. The platform paves the way toward a more fully automated mass spectrometry analysis and ultimately closed-loop laboratory workflows involving automated experimentation and analysis coupled to AI-driven experimentation utilizing cutting-edge analytical instrumentation. AutonoMS documentation is available at https://autonoms.readthedocs.io