The analysis of human locomotion during activities such as pedalling [
1,
2], walking [
3,
4], race walking [
5,
6,
7], and running [
8,
9,
10,
11,
12], exhibits a repetitive and stereotypical movement pattern over time [
13,
14,
15,
16,
17,
18,
19,
20]. Numerous studies have focused on investigating the variability of the gait cycle paradigm (gait variability, GV) to better understand the bioenergetics and control of the human locomotion [
21]. This seemingly simple activity [
22,
23,
24,
25,
26,
27] involves a complex task [
28,
29,
30,
31,
32,
33,
34] requiring a precise synergy [
35,
36,
37,
38,
39,
40] between lower limbs coordination [
41,
42,
43,
44,
45,
46,
47] and muscle contractions [
48,
49,
50,
51,
52,
53,
54] in response to both natural and non-natural conditions [
55]. As a result, individuals need to continuously explore new strategies [
56,
57,
58,
59,
60,
61,
62,
63] and promptly adapt the motor task [
64,
65,
66,
67,
68,
69,
70] to the immediate environment conditions, adjusting their footstep cycle to the most appropriate one.
In walking, stride-to-stride variability [
71,
72,
73,
74,
75,
76,
77] arises due to the system’s constant need [
78,
79,
80,
81,
82] to adjust inaccurate movements [
83]. From neurophysiological point of view, higher variability was associated with poor coordination level, while lower variability indicates better coordination level [
84]. Pathological and non-pathological factors have been proven to affect the coordination level. Indeed, Parkinson’s disease, aging [
55], and individuals with a lower limb injury [
85] have been shown to exhibit high variability. Nonetheless, increased variability has been observed in healthy people due to changes in body positions during uphill running [
86], alterations in body posture [
1] and variations in walking speed [
83]. Jordan et al. [
83] showed that the better walking coordination level (i.e., lowest GV) in healthy young females occurred at walking speeds between 100 and 110% of the preferred walking speed. Even though the preferred speed favors better walking coordination [
83], most of the studies on GV has used to administer standardized speed on a treadmill [
85,
87,
88]. Anyway, it was showed a divergence about the physiological effort between preferred walking speed on a treadmill compared to the overground [
89] suggesting that on a treadmill the preferred walking speed was lower. Therefore, to obtain data that are more representative of daily activities, the GV should be studied at the preferred gait speed determined overground, as treadmill does not fully represent the ground of daily activities. This methodological approach could mitigate the influence of the neurophysiological factors on bioenergetics variables, such as kinematic, kinetic, and motor control aspects. Contrarily, the motorized treadmill [
90] provides the advantage of having long duration trials such as gait variability study needed. Indeed, 400 steps are required for an accurate estimation of the step kinematic variability [
91] at the constant speed. Certainly, treadmills offer researchers the advantage of precise control over walking speed, enabling extended trials with subjects confined to a limited motion capture space, and the option to connect onboard electronics to a stationary data acquisition system. However, also if Jordan et al. [
83] partially clarified that the coordination level trend is speed’s dependent, unfortunately the speed (m·s-1 or km·h-1) and metabolic demand (heart rate) were not reported and studied. Furthermore, in that study the preferred walking speed was assessed on the treadmill only. The critical aspect concerning walking speed lies in the fact that each participant exhibits a distinct preferred walking speed. Therefore, the preferred walking speed should be assessed on overground to be more realistic [
89], while the gait variability on a treadmill [
91] to overcome the GV setting. From this perspective, the aims of this study were: a) to assess individual preferred walking speed in overground setting, b) to determine the metabolic demand and gait variability related to the different walking speeds on a treadmill.