Echium plantagineum L. (Boraginaceae) is an annual or biennial flowering plant native to the Mediterranean region, including western and southern Europe, northern Africa, and southwestern Asia. It was introduced to Australia as an ornamental plant in the 1800s and is commonly known as Paterson's curse or Salvation Jane. Due to its adaptive ability, it has become one of Australia's most costly, noxious, and invasive weeds. It has spread widely in the eastern and south-eastern states of Australia where it is estimated to have influenced 30 million hectares of agricultural land. However, to date, there is limited natural products research related to phytochemical and biological screening of compounds isolated from E. plantagineum. This thesis outlines a comprehensive chemical investigation and subsequent in vitro cytotoxicity screening of E. plantagineum to investigate a possible medicinal use for this invasive weed (Chapter 2 to Chapter 6). To achieve this, E. plantagineum was firstly collected from farmland in Mulwala, and Young, NSW, Australia. Each plant was then separated into different parts, including leaf, flower, root, and stem, and extracted with methanol to obtain eight crude extracts. The first goal of the project was to isolate, purify and identify secondary metabolites direct from chemical profiling employing mass spectrometry. Chapter 2 describes the isolation and preliminary chemical screening of E. plantagineum methanolic extracts of each part. The chemical fingerprint of E. plantagineum methanolic extracts was conducted by LC-MS in both positive and negative modes to identify and predict different classes of compounds. Previous phytochemicals reported include pyrrolizidine alkaloids, naphthoquinones and flavonoids. Our chemical predication also indicated an abundance of polyphenols such as rosmarinic acid, globoidnan A and rabdosiin among other derivatives. Chapter 3 and 4 report the identification and structure elucidation of secondary metabolites isolated from crude extracts in E. plantagineum. The structures were determined by analysing 1D and 2D NMR (COSY, HSQC, HMBC, ROESY) spectra and high-resolution electrospray mass spectrometry. Polyphenols such as rosmarinic acid, and particularly aryl-dihydronaphthalene-type lignans, like globoidnan A, globoidnan B, a new globoidnan B diastereomer, rabdosiin, and echiumin A were isolated and characterised from the plant. Additionally, pyrrolizidine alkaloids (lycopsamine, Lactodine), nucleosides (adenosine and uridine) and amino acids were also identified. In addition, a new aryl-dihydronaphthalene-type lignan, which we have named echiumin E was characterised. The second aim of this thesis was to investigate in vitro cytotoxicity of bioactive extracts and isolated compounds as well as conduct a mechanistic investigation (Chapter 5 and Chapter 6). In Chapter 5, bioassay-guided isolation of antiproliferation against cancer cell lines (PC-3, SH-SY5Y, and Hela) and one normal human cell line (WPMY-1) as well as bacterial strains (Gram-positive and Gram-negative) are presented. Firstly, the cytotoxicity assay conditions were accessed and validated. This included cell seeding density, resazurin bioassay incubating time, DMSO vehicle and control concentration. IC50 were calculated for determining cytotoxicity. Our results showed that flower extracts collected from both locations were the most potent. In addition, Hela cells showed high sensitivity with the treatment of all extracts. Antimicrobial results revealed that flowers collected from Mulwala exhibited significant cytotoxicity against MSSA and moderate activity against MRSA. However, no activity was shown on Pseudomonas aeruginosa with the treatment of all crude extracts. The mechanistic study of the potent flower extracts is also resented in Chapter 5. Flower extracts induced significant oxidative stress, but little early apoptosis was observed, suggesting that the flower extract induced antiproliferation through another signalling pathway. After the bioassay-guided purification, the flower extract was analysed by 1D NMR. The main chemical components of these extracts were identified to be flavonoids, which are well characterised both chemically and biologically. Chapter 6 reports further cytotoxicity screening of selected compounds (elucidated in Chapter 3) rosmarinic acid, globoidnan B, rabdosiin, and echiumin E against cancer cell lines (PC-3, SH-SY5Y, Hela and WPMY-1 cells) and bacterial strains in vitro. Results indicated that all compounds exerted an inhibitory effect on all the cell lines tested, especially on Hela cells. Further mechanistic studies indicated that both globoidnan B and rabdosiin induced oxidative stress and active downstream caspase 3/7 activity and led to different stages of apoptosis, which are suggested to be ROS-mediated apoptosis. In conclusion, this project has provided a distinguishable point of view on investigating phytochemistry and cytotoxicity of an invasive plant in Australia, E. plantagineum. The determination, evaluation and mechanistic studies showed that E. plantagineum held notable bioactive metabolites, and they have induced cytotoxicity through apoptosis pathways. This study highlighted the value of Paterson's curse as a po ential source of therapeutically viable compounds for the first time. These important phytochemicals have the potential to contribute to future therapeutic applications with further high-quality studies.