Solanum chacoense Bitter is resistant to the Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say). Resistance has been associated with the presence of a rare class of glycoalkaloids, the leptines. In this study, seven tetraploid, F2 S. tuberosum x S. chacoense families were evaluated for foliar production of leptines I and II, leptinines I and II, and alpha-solanine and alpha-chaconine; and screened for resistance to CPB in the laboratory and field. Resistance was correlated with the concentrations of glycoalkaloids on a family and an individual basis. Leptine concentrations ranged from undetectable to a high of 18.0 mg/g dry weight. All of the progeny produced solanine and chaconine. Family 9623 had the highest mean leptine concentration and the lowest mean leaf disk feeding and CPB defoliation levels. Family 9616 had the lowest mean glycoalkaloid concentration and ranked as one of the most susceptible families. Regression analyses of solanine + chaconine, leptine I and II, and leptinine I and II foliar concentrations versus leaf disk consumption and field defoliation revealed that only increased foliar levels of leptines resulted in decreased CPB feeding. The regression models for leptines versus leaf disk consumption and field defoliation were highly significant, accounting for 17% and 26% of the variation in consumption and defoliation, respectively. To the best of our knowledge, this is the first work reporting the impact of leptine and leptinine concentrations on CPB feeding in tetraploid, S. tuberosum x S. chacoense potato hybrids. Results are discussed within the context of breeding for resistance to CPB.