Objective To investigate the effect of pregabalin combined with tramadol/acetaminophen on acute pain in patients undergoing CT‑guided puncture localization of pulmonary nodules. Methods A 2×2 factorial trial design was used, and 120 patients scheduled for CT‑guided puncture localization of pulmonary nodules were selected. According to the random number table method, the patients were divided into four groups: a control group (group C, n=30), a pregabalin plus placebo group (group PC, n=29), a tramadol/acetaminophen plus placebo group (group AC, n=30), and a pregabalin plus tramadol/acetaminophen group (group PA, n=29). Each group received the corresponding oral medication 1 h before localization. Whether there was an interaction between pregabalin and tramadol/acetaminophen would be analyzed (If there was an interaction, the individual effects of pregabalin and tramadol/acetaminophen would be analyzed. If there was no interaction, the primary effects of pregabalin and tramadol/acetaminophen would be analyzed). For analysis of the primary effect of pregabalin, the patients were divided into a pregabalin group (n=58) and a non‑pregabalin group (n=60). For analysis of the primary effect of tramadol/acetaminophen, the patients were divided into a tramadol/acetaminophen group (n=59) and a non‑tramadol/acetaminophen group (n=59). The general data and the related data of puncture localization of patients were recorded. The Numerical Rating Scale (NRS) was used to assess pain scores in all patients. Then, the systolic blood pressure, diastolic blood pressure, heart rate, and pulse oxygen saturation immediately after localization, the incidence of moderate to severe pain, the proportion of rescue analgesia, the incidence of adverse drug reactions, patient satisfaction (very satisfied, satisfied, average, dissatisfied, and very dissatisfied), and the differences in systolic blood pressure, diastolic blood pressure, and heart rate before and after localization were recorded. Results There was no statistical difference in the general data and the related data of puncture localization of patients (all P>0.05). The primary effect of pregabalin and tramadol/acetaminophen were significant (P<0.05), without obvious interaction between them (P=0.752). Compared with the non‑pregabalin group, the pregabalin group showed decreases in the NRS score, systolic blood pressure, diastolic blood pressure and heart rate immediately after localization, the incidence of moderate to severe pain, and the proportion of rescue analgesia (all P<0.05), as well as increases in pulse oxygen saturation immediately after localization and the number of people who were very satisfied (all P<0.05), without statistical differences in the incidence of adverse drug reactions (P>0.05). Compared with the non‑tramadol/acetaminophen group, the tramadol/acetaminophen group presented decreases in the NRS score, systolic blood pressure, diastolic blood pressure and heart rate immediately after localization, the incidence of moderate to severe pain, and the proportion of rescue analgesia (all P<0.05), as well as increases in pulse oxygen saturation immediately after localization and the number of people who were very satisfied (all P<0.05), without statistical differences in the incidence of adverse drug reactions (P>0.05). Compared with group C, group PC and group AC, group PA showed reduction in the NRS score, the differences in systolic blood pressure, diastolic blood pressure and heart rate before and after localization (all P<0.05). Compared with group C, group PC and group AC presented reduction in the NRS score, the differences in systolic blood pressure, diastolic blood pressure and heart rate before and after localization (all P<0.05); but there was no statistical differences between group PC and group AC (P>0.05). Compared with group C, group PA showed decrease in systolic blood pressure, diastolic blood pressure and heart rate immediately after localization, the incidence of moderate to severe pain and the proportion of rescue analgesia, as well as increase in pulse oxygen saturation immediately after localization and the number of people who were very satisfied (all P<0.05). Compared with group C, the incidence of moderate to severe pain in group AC decreased (P<0.05), with increased the number of people who were very satisfied (P<0.05). There was no statistical difference in the incidence of adverse drug reactions among group C, group PC, group AC and group PA (all P>0.05). Conclusions Pregabalin and tramadol/acetaminophen alone or both can significantly reduce the NRS score after puncture localization of pulmonary nodules. However, the combination of the two drugs has the optimal analgesic effect and significantly reduces hemodynamic fluctuations after localization, with a low incidence of adverse drug reactions.