Data on cardiac arrhythmia and electrolyte changes during the dialysis cycle have been limited. Fifty-two hemodialysis (HD) patients underwent 48-h Holter monitoring during early-week and mid-week HD sessions. Pre-HD and post-HD blood samples were collected in both HD sessions. The 48-h Holter data were divided into five phases: (1) 4-h during the early-week HD (HD1), (2) 12-h post-HD1, (3) 16-h period between Phases 2 and 4 (used as the patient's baseline electrocardiography [ECG]), (4) 12-h pre-HD2 phase, and (5) 4-h during the mid-week HD (HD2). The patients' mean age was 68.54 ± 13.37 years. We found that the dialysate-to-serum[K] gradient and changes of S[K] were significantly higher in HD1 than in HD2, as well as changes of S[Mg]. There were no significant ECG changes during the 4-h HD1 and HD2 when compared with the baseline ECG. Phase 2 of Holter ECG was the most common phase that showed significant changes (increased QT interval dispersion (QTD), increased ventricular events, increased number of premature ventricular contractions, ST elevation and ST depression), which was contributed from the dialysate[K] 2 mmol/L subgroup, but not the dialysate[K] 3 mmol/L subgroup. In the subgroup of patients with a high ultrafiltration rate (UFR; mean UFR ≥10 mL/kg/h), there were significantly increased ventricular events and ST-segment changes in Phase 2. In conclusion, ECG changes were associated with the dialysis cycle, significantly in the 12-h after early-week HD sessions. These may be associated with low dialysate[K] or high dialysate-to-S[K] gradient, high ultrafiltration rate and duration of the interdialytic interval.