This study investigated the value of C-terminal telopeptides of collagen type II (CTX-II) and YKL-40 in early diagnosis and treatment evaluation of osteoarthritis (OA). A total of 90 patients with OA diagnosed and treated in The First Affiliated Hospital, Guangzhou Medical University from March 2015 to January 2018 were selected as the study group. At the same time, 50 healthy elderly were included as the control group. The study group was divided into three subgroups including group A (29 cases, 500 mg glucosamine sulfate), group B (29 cases, 50 mg diacerein) and group C (32 cases, 500 mg glucosamine sulfate and 50 mg diacerein). Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) was used to assess the severity and treatment of arthritis. Enzyme-linked immunosorbent assay was used to measure the concentration of CTX-II and YKL-40 in serum. WOMAC scores in the study A, B and C groups were significantly higher than those in the control group (P<0.001). Serum CTX-II and YKL-40 concentrations were higher in the study group than in the control group (P<0.001). Sensitivity of serum CTX-II combined with YKL-40 in the diagnosis of OA was 90% and the specificity was 78%. CTX-II and YKL-40 levels in different Kellgren Lawrence (K-L) grades were significantly different (P<0.001), and increased with the increase of K-L grade. Concentrations of serum CTX-II and YKL-40 before treatment in the study group was positively correlated with WOMAC score (P<0.001). At 3, 6 and 9 weeks after the beginning of treatment, serum concentrations of CTX-II and YKL-40 decreased significantly (P<0.001). At 3 weeks of treatment, CTX-II was positively correlated with YKL-40 concentration and WOMAC score (r=0.406, P<0.001; r=0.430, P<0.001); CTX-II was positively correlated with YKL-40 concentration and WOMAC score at 6 weeks of treatment (r=0.350, P<0.001; r=0.358, P<0.001); CTX-II was positively correlated with YKL-40 concentration and WOMAC score at 9 weeks after treatment (r=0.370, P<0.001; r=0.394, P<0.001). Combined detection of serum CTX-II and YKL-40 can improve the sensitivity of early OA diagnosis, and it has an important diagnostic value for early OA patients. Therefore, it can be used as a biological indicator for early OA diagnosis, severity assessment, and evaluation of treatment effects.