Despite the accumulating evidence from in vitro and animal experiments supporting the role of kynurenine (a tryptophan metabolite) in a number of degenerative age-related changes, the relationship between kynurenine and frailty in older adults is not well understood. We collected blood samples from 73 participants who underwent a comprehensive geriatric assessment, measuring kynurenine levels using liquid chromatography-tandem mass spectrometry. We assessed the phenotypic frailty and the deficit accumulation frailty index using widely validated approaches proposed by Fried et al. and Rockwood et al., respectively. After adjusting for sex, age, and body mass index, the frail participants presented 52.9% and 34.3% higher serum kynurenine levels than those with robustness and prefrailty, respectively (P = 0.005 and 0.014, respectively). Serum kynurenine levels were positively associated with the frailty index, time to complete 5 chair stands, and patient health questionnaire-2 score and inversely associated with grip strength and gait speed (P = 0.042 to <0.001). Furthermore, the odds ratio per increase in serum kynurenine level for phenotypic frailty was approximately 2.62 (95% confidence interval = 1.22–5.65, P = 0.014). These data provide clinical evidence that circulating kynurenine might be a potential biomarker for assessing the risk of frailty in humans.