Molecular markers linked to specific traits are essential for accelerating marker-assisted selection in crop improvement. Castor (Ricinus communis L.), a non-edible oilseed crop, is highly valued for its high oil content (40–55%) and exceptionally high ricinoleic acid content (>85%), making it a key resource for industrial applications. In this study, we developed and characterized functionally relevant EST-derived SSR markers associated with oil biosynthesis. A total of 62,105 expressed sequence tags (ESTs) were retrieved from the NCBI dbEST database and assembled into 13,811 unigenes using TGICL. Mining these sequences revealed 2,425 SSRs across 1,955 unigenes, averaging one SSR every 4.7 kb. Tri-nucleotide repeats were the most abundant (54.4%), followed by di-nucleotide repeats (42.0%). Functional annotation using Blast2GO identified several SSR-containing sequences involved in fatty acid metabolism pathways. From 1,555 designed primer pairs, 30 were selected for validation across 24 genetically diverse castor genotypes. Of these, 15 primers produced clear amplification, and 10 were polymorphic, with polymorphism information content (PIC) values ranging from 0.239 to 0.454 and an average of 2.5 alleles per locus. UPGMA clustering grouped the genotypes into three major clusters, revealing substantial genetic diversity. Notably, the polymorphic markers were associated with key genes involved in fatty acid biosynthesis, including oleosin 1, diacylglycerol acyltransferase, hydroxyacyl-ACP dehydrase, and 3-ketoacyl-CoA synthase. These newly developed EST-SSR markers represent the first functional SSR resource specifically targeting fatty acid metabolism in castor. They offer valuable tools for genetic diversity analysis and marker-assisted breeding, supporting efforts to enhance oil quality traits in this economically important crop.