Purpose Currently, there remains a scarcity of established preoperative tests to accurately predict the isocitrate dehydrogenase (IDH) mutation status in clinical scenarios, with limited research has explored the potential synergistic diagnostic performance among metabolite, perfusion, and diffusion parameters. To address this issue, we aimed to develop an imaging protocol that integrated 2-hydroxyglutarate (2HG) magnetic resonance spectroscopy (MRS) and intravoxel incoherent motion (IVIM) by comprehensively assessing metabolic, cellular, and angiogenic changes caused by IDH mutations, and explored the diagnostic efficiency of this imaging protocol for predicting IDH mutation status in clinical scenarios. Methods Patients who met the inclusion criteria were categorized into two groups: IDH-wild type (IDH-WT) group and IDH-mutant (IDH-MT) group. Subsequently, we quantified the 2HG concentration, the relative apparent diffusion coefficient (rADC), the relative true diffusion coefficient value (rD), the relative pseudo-diffusion coefficient (rD*) and the relative perfusion fraction value (rf). Intergroup differences were estimated using t-test and Mann-Whitney U test. Finally, we performed receiver operating characteristic (ROC) curve and DeLong's test to evaluate and compare the diagnostic performance of individual parameters and their combinations. Results 64 patients (female, 21; male, 43; age, 47.0 +/- 13.7 years) were enrolled. Compared with IDH-WT gliomas, IDH-MT gliomas had higher 2HG concentration, rADC and rD (P < 0.001), and lower rD* (P = 0.013). The ROC curve demonstrated that 2HG + rD + rD* exhibited the highest areas under curve (AUC) value (0.967, 95%CI 0.889-0.996) for discriminating IDH mutation status. Compared with each individual parameter, the predictive efficiency of 2HG + rADC + rD* and 2HG + rD + rD* shows a statistically significant enhancement (DeLong's test: P < 0.05). Conclusions The integration of 2HG MRS and IVIM significantly improves the diagnostic efficiency for predicting IDH mutation status in clinical scenarios.