The present study aimed to explore changes in serum betatrophin concentration in women with gestational diabetes mellitus (GDM) and its relationship with clinical metabolic indicators. The research also aimed to provide a basis for the in-depth study of the pathogenesis of GDM, together with a new target for the treatment of the condition and improving the methods for its prediction and diagnosis. A total of 60 participants were included of whom 30 were enrolled in the pregnancy with normal glucose regulation (P-NGR) group, and 30 in the GDM group. The clinical metabolic indicators in the two groups were measured and body mass index (BMI) was calculated. Additionally, two models were established, i.e., homeostasis assessment of insulin resistance (HOMA-IR), and homeostasis assessment of beta-cell function (HOMA-beta). An enzyme-linked immunosorbent assay (ELISA) was used to detect the serum betatrophin concentrations in the two groups. The differences in the clinical metabolic indicators and betatrophin concentrations between the two groups were compared, and the correlation between betatrophin and clinical metabolic indicators was analyzed. The receiver operating characteristic (ROC) curve was used to predict GDM using the serum betatrophin concentration. The levels of triglyceride (TG) (P = 0.001), total cholesterol (TC) (P = 0.004), low-density lipoprotein (LDL) (P < 0.001), fasting blood glucose (FBG) (P < 0.001), fasting serum insulin (FSI) (P = 0.028), HOMA-IR (P < 0.001), HOMA-beta (P < 0.001), and the concentration of betatrophin (p < 0.001) were significantly higher in the GDM compared with the P-NGR group, and the difference was statistically significant. The concentration of betatrophin was positively correlated with BMI (P = 0.017), TG (P = 0.005), TC (P = 0.018), FSI (P = 0.023), and HOMA-IR (P = 0.022) in the GDM group. Multiple linear regression analysis showed that HOMA-IR and TG could affect the serum betatrophin concentration. The best cut-off point of the serum betatrophin concentration predicted by the ROC curve in the case of GDM was 800.1pg/ml. The serum concentration of betatrophin in the GDM group was significantly increased and positively correlated with BMI, TG, TC, FSI, and HOMA-IR. Insulin resistance in the GDM population may be a mechanism involved in the overexpression of serum betatrophin; therefore, serum betatrophin could potentially be used as a biochemical indicator for predicting GDM.