The significant predictors for the postoperative deterioration of separate renal function after minimally invasive stone surgery were investigated in the present prospective and observational study. A total of 117 consecutive patients who underwent retrograde intrarenal surgery or mini-percutaneous nephrolithotomy for renal calculi >10 mm were included in the present study. Perioperative changes in separate renal function were evaluated with Technetium-99m-Diethylene TriaminePenta acetic acid scan prior to intervention and at postoperative 3 months. Based on the functional differences between bilateral renal units, deterioration of separate renal function was graded into the following three groups: normal deterioration (<10%), moderate deterioration (10–20%), and severe deterioration (>20%). A total of 46 patients had a normal separate renal function, while 71 (60.7%) showed abnormal separate function in the involved side, including 29 (24.8%) moderate and 42 (35.9%) severe deterioration. Postoperatively, 48 patients (41.0%) showed aggravation or no recovery of separate renal function. Of the 46 patients with normal separate function, only 9 patients (19.5%) showed postoperative aggravation. Patients with moderate and severe deterioration showed aggravation (n = 7, 24.1%) or no recovery of separate renal function (n = 32, 76.1%, P < 0.001). Preoperative severe deterioration of separate renal function was an independent significant predictor for the postoperative deterioration of renal function (OR: 9.09, 95% CI: 4.007–20.624, P < 0.001). Lower preoperative deterioration of separate renal function showed a high probability of functional recovery. Therefore, it is hypothesized that early intervention might be necessary in cases where the patient exhibits severe aggravation of renal function.