Vitality capacity (VC) reflects a physiological state and is a determinant domain of intrinsic capacity but has so far remained mainly theoretical. This study validates the vitality capacity domains 'energy and metabolism' and 'neuromuscular function' and examines its link to locomotor capacity and quality of life (QoL). Exploratory factor analysis (EFA) was performed on the combined dataset from the Fatigue Resistance AMErsfoort study (FRAME, n = 1000) and the Fatigue Plot study (FATPLOT,n = 620). Confirmatory factor analyses (CFA) were subsequently performed on data from the AMersfoort COhort study on functional decline, Healthy aging and Frailty (AMCOHF,n = 367) and the BrUssels sTudy on The Early pRedictors of FraiLtY (BUTTERFLY,n = 491), to validate VC in both middle-aged and older adults. Linear hierarchical regression analysis was used to investigate the relationship between VC, locomotor capacity, and QoL. EFA indicated a one-factor model and CFA validated this with good model fit in the dataset (BUTTERFLY) (Robust CFI; 0.960, SRMR: 0.040) and (AMCOHF) (Robust CFI; 0.942, SRMR: 0.055). This model validated maximal grip strength (GSmax), 30-s chair stand test (30CST), Multidimensional Fatigue Inventory (MFI-20) and Capacity to Perceived Vitality ratio physical (CPV-physical) to measure VC. Several assessments show a significant relationship with locomotor capacity and QoL. This study indicated that VC is a coherent domain and has a relationship with locomotor capacity and QoL.