Comparative Proteomic Analysis of Neem (Azadirachta indica) Seed and Leaves

<p>Modern research has significantly contributed to creating well-validated research data for the holistic use of the Neem tree in disease management. Neem’s astonishing historical and modern relevance, along with its small-molecule composition, provide further evidence for its selection as the target of our study. The scientific name of the Neem tree is <em>Azadirachta Indica</em>, and it belongs to the Family Meliaceae. This research focused on the Proteomic characterization of proteins from Neem leaves and Seeds and evaluated their anticancer potential in breast cancer cells. Despite consistent advancements in therapies, breast cancer remains a concerning health issue and a deadly disease among women. Neem has shown a remarkable potential against cancer due to its wide range of biologically active compounds. Studies on the proteins present in Neem remain largely unexplored. The reviewed SwissProt database in UniProtKB lists only three proteins from the Neem tree. Therefore, this study has great potential to explore the global Proteomic profiling of Neem leaves and Seeds. Further analysis has also evaluated the anti-cancer potential of proteins from Neem leaves and Seeds against MDA-MB-231 and MCF-7 breast cancer cell lines. </p> <p>The neem leaves and seeds were defatted separately with <em>n</em>-hexane, and then proteins were extracted with a 20 mM Tris/HCl buffer, pH 8. Finally, proteins were precipitated with 80% ammonium sulphate(NH4)2SO4. The crude extracts were dialyzed in water using a 3.5 MWCO dialysis tubing and then lyophilized. The anticancer bioactivity of Neem leaves and Seeds Crude protein at different concentrations was evaluated against MDA-MB-231 and MCF-7 cells using the MTS assay for 24 and 72 hours, and the absorbance was measured at 490 nm. Neem seeds and leaves at different concentrations exhibited inhibitory effects on MDA-MB-231 cells and MCF-7 cells. Further purification of the Neem Seeds and Leaves crude extracts was achieved using gel filtration (SEC) on a HiLoad^TM16/600 Superdex^TM200 pg column, followed by SDS-PAGE on a 10% Tris/Glycine gel. Proteomic profiles were established by peptide mass fingerprinting using LC-MS/MS. The bioinformatic analysis, using PEAKS Studio 10.5, a UniProtKB/ SwissProt/TrEMBL/Chinaberry/Neem Viridiplantae database, with FDR ≤ 1% and at least 1 unique peptide match, identified 43 proteins using trypsin and 15 proteins using pepsin in Neem Leaves, and 209 proteins with enzyme trypsin and 26 proteins with enzyme pepsin on Neem Seeds. </p>

Name thesis:degree_name

Master of Science (MS)

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Pharmaceutical Sciences

Year dc:date.available

2026

Author dc:creator

• Kahsay, Abigail Yohannes

Contributors dc:contributor

• Dr. Aftab Ahmed
• Dr. Keykavous Parang
• Dr. Jason Yamaki

• Proteomics
• protein identification
• cytotoxicity assay
• bottom up proteomics
• top down proteomics
• Gene ontology
• Other Pharmacy and Pharmaceutical Sciences

Repository record dc:identifier

https://digitalcommons.chapman.edu/pharmaceutical_sciences_theses/42

OAI identifier oai:identifier

oai:digitalcommons.chapman.edu:pharmaceutical_sciences_theses-1043