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Background: The most electronegative subfraction of low-density lipoprotein (LDL), L5, is atherogenic in vitro and in vivo, and its plasma levels are elevated in patients with high cardiovascular disease risk and ischemic stroke (Blood 2016;127:1336-45). Because L5 LDL may be a useful biomarker for predicting cardiovascular disease, we defined a reference range for L5 plasma levels in healthy adults and examined the power of L5 levels to differentiate patients with hyperlipidemia (HLP) or coronary artery disease (CAD) from healthy individuals.Methods: We enrolled 35 healthy individuals, 40 patients with stable CAD, and 35 patients with HLP without evidence of CAD. Plasma LDL was isolated from each individual, and the percentage of L5 in total LDL (L5%) was quantified by using fast-protein liquid chromatography with an anion-exchange column.Results: Plasma L5% and L5 concentration (ie, [L5]) were significantly higher in patients with HLP or CAD than in healthy controls (P<0.001). The reference ranges of L5% and [L5] in healthy adults were determined to be less than 1.6% and less than 1.7 mg/dL, respectively. In individuals with an L5% greater than 1.6%, the odds ratio was 9.636 for HLP or CAD (Figure). In individuals with an [L5] greater than 1.7 mg/dL, the odds ratio was 17.684 for HLP or CAD (Figure). The power of L5% and [L5] to differentiate patients with HLP or CAD from healthy individuals was superior to that of the LDL/HDL ratio.Conclusions: The power of L5% and [L5] in predicting cardiovascular risk is superior to that of other markers currently in use. Therefore, the reference ranges of L5% and [L5] identified in this study may be clinically useful in the prevention and treatment of cardiovascular disease. As previously reported, elevated plasma L5 levels may promote or aggravate ischemic stroke by enhancing platelet aggregation and platelet-endothelium interactions.