Obesity and metabolic disorders are strongly associated with high-fat diets, with palmitate playing a key role in adipocyte dysfunction and insulin resistance. However, proteomic changes during adipocyte differentiation under palmitate exposure using human cell lines like SGBS remain underexplored. This study investigates these changes by differentiating SGBS pre-adipocytes (PA) into white adipocytes (WA), with a subset treated with palmitate (WP). Proteomic profiling was conducted using micro sTRAP digestion and LC-ESI-MS/MS with data-independent acquisition. WA and WP both showed upregulation of metabolic enzymes involved in fatty acid metabolism and energy production. Notably, WP exhibited distinct stress-related signatures, including downregulation of ER and lysosomal proteins, ECM remodeling, and increased translation-related proteins, indicating disrupted differentiation and cellular stress resembling obesity conditions.