Variability of soybean seed composition constituents (protein, oil, fatty acids, and sugars) due to environmental factors such as drought is significant, but the physiological mechanisms involved are not well known. The objective of this research was to investigate the effect of drought on seed composition.  A repeated greenhouse experiment was conducted where sets of soybean plants were subjected to different irrigation treatments. Plants were irrigated at vegetative (IV), flowering (IF), and seed-fill (IS) stages. Drought treatments were applied at vegetative (DV), flowering (DF), and seed-fill (DS). Irrigation at vegetative stage was considered as continuous irrigation and was used as a control. Irrigation was monitored using soil water potential sensors. Irrigated plants were kept between –15 to –20 kPa (this was considered field capacity). Drought plants were kept between –90 and –100 kPa (this represented a moderate to severe water stress level for soybean under greenhouse conditions). Soybean cultivar Hutcheson was used. Uniform size seedlings at about V1 stage were transplanted into 9.45 L size pots filled with field Dundee silt loam soil. A randomized complete block design with four replicates was used. Seeds from each treatment were collected at harvest maturity for seed analyses. The results showed that seed composition and ¹⁵N/¹⁴N and ¹³C/¹²C ratios were significantly (p≤ 0.05) altered in drought-stressed plants compared to irrigated plants. Seed protein and oleic acid increased and oil and linolenic acid decreased under drought conditions. Stachyose and raffinose significantly (p≤ 0.05) increased at the expense of sucrose, which decreased significantly (p≤ 0.05) under drought conditions. The increase in the ¹⁵N/¹⁴N ratio under drought conditions indicated inhibition of nitrogen fixation, and the increase in ¹³C/¹²C ratio under drought conditions indicated that the source of carbon fixation was altered. The research demonstrated that seed composition and the sources of nitrogen and carbon were altered, especially under drought conditions, indicating metabolic changes in the carbon and nitrogen fixation pathways. This research provides useful information for breeders to select for desirable seed composition traits (protein or oil) under drought conditions.