L5, the most electronegative subfraction of low-density lipoprotein cholesterol (LDL-C), may play a role in the pathogenesis of cerebrovascular dysfunction and neurodegeneration. We hypothesized that serum L5 is associated with cognitive impairment and investigated the association between serum L5 levels and cognitive performance in patients with mild cognitive impairment (MCI). This cross-sectional study conducted in Taiwan included 22 patients with MCI and 40 older people with normal cognition (healthy controls). All participants were assessed with the Cognitive Abilities Screening Instrument (CASI) and a CASI-estimated Mini-Mental State Examination (MMSE-CE). We compared the serum total cholesterol (TC), LDL-C, and L5 levels between the MCI and control groups and examined the association between lipid profiles and cognitive performance in these groups. The serum L5 concentration and total CASI scores were significantly negatively correlated in the MCI group. Serum L5% was negatively correlated with MMSE-CE and total CASI scores, particularly in the orientation and language subdomains. No significant correlation between the serum L5 level and cognitive performance was noted in the control group. Conclusions: Serum L5, instead of TC or total LDL-C, could be associated with cognitive impairment through a disease stage-dependent mode that occurs during neurodegeneration.