The production of high-quality rum requires a carefully controlled fermentation process, which is determined by the physicochemical and nutritional properties of the molasses used as a substrate. The flavor, aroma, and alcohol content of the final product are heavily influenced by the composition of molasses. This paper presents the results of an extensive analysis of molasses, a byproduct of sugar production that has various applications in food and feed industries. The physicochemical properties and nutritional composition of molasses were investigated using a range of analytical techniques including Brix, moisture content, pH, total ash content, total dissolved solids, crude protein, sugar content, total nitrogen, free amino acids, amino acids, mineral content, vitamins, and organic acids. The study analyzed molasses samples obtained from three painted concrete constructed storage wells at the Bundaberg Distilling Company. The results indicated that the molasses samples possessed a substantial sugar concentration, with sucrose being the most abundant, ranging from 35.02 – 35.27% w/w. These findings support the traditional use of molasses in rum fermentation and use as an alternative feedstock for the synthesis of various value-added chemicals. The pH of the molasses was slightly acidic, and the total ash content was relatively high ranging from 8.76 – 13.55 % w/w, indicating the presence of mineral salts. Mineral content varied among the samples, but all three contained significant levels of sodium, phosphorus, potassium, calcium, and magnesium. All the samples contained essential amino acids and vitamins, but their levels varied. The molasses also contained varying levels of different organic acids, including acetic, propanoic, aconitic, formic, valeric, lactic, citric, and malic acids. The analysis revealed significant differences in ash content, nitrogen compounds, and minerals between storage wells, indicating inherent variability between batches. However, key fermentable sugars did not significantly differ between samples, and blending or standardization is recommended to minimize batch-to-batch variation prior to fermentation. This study provides valuable information on the composition and quality of molasses, which can be used to optimize its utilization in rum fermentation and various other applications.