Besides injury prevention, mouthguards can also be employed to improve physical performance. The effects of personalization of mouthguards have rarely been investigated. This 3-armed, randomized, controlled crossover trial investigated the difference of wearing (a) personalized or custom-made (CM, e.g., bite-aligned), (b) standard (BB, boil and bite), and (c) no (CON) mouthguards on general fitness parameters in experienced collegiate football players. A group of 10 upperclassmen (age, 19-22 years; mean ± SD: age = 20.7 ± 0.8 years; body mass = 83 ± 7.4 kg; height = 179.1 ± 5.2 cm; body mass index = 25.9 ± 2.2 kg·cm), National Collegiate Athletic Association Division II football players with at least 2 years of playing experience, were randomly assigned to the 3 mouthguard conditions: a randomized, within-subjects repeated-measures design was applied. All participants were randomly tested on strength and endurance performance V[Combining Dot Above]O2max testing, with Bruce treadmill protocol including (a) time to fatigue, (b) blood lactate concentration in millimoles per liter at stage 2 and (c) at peak fatigue, (d) flexibility, (e) reaction time, (f) squat vertical jump, (g) countermovement vertical jump, and (h) 1 repetition maximum bench press. Repeated-measures analysis of variance showed no significant differences between the 3 conditions for each outcome variable (0.23 < p < 0.94; 0.007 < (Equation is included in full-text article.)< 0.15). These data indicate that CM mouthguards did not superiorly affect general fitness parameters compared with BB and CON. In turn, protective BB or CM mouthpieces did not appear to impair general fitness performance vs. CON. The recommendation of a custom bite-aligning mouthguards for performance enhancement in young Division II football players is questioned. Further studies with larger sample sizes, gender comparison, and (sport) discipline-specific performance testing are needed.