Background/Aim: The insulin tolerance test (ITT) remains the gold standard for evaluating the pituitary function, but has potential risks when hypoglycaemia is induced. There are scarce data using short-acting insulin analogs for ITTs. This pilot study compares the effects of insulin lispro (LPI) with regular insulin (RGI) during an ITT. Methods: Patients with suspected hypopituitarism (n = 103) randomly received either LPI (n = 51) or RGI (n = 52). Results: All patients reported signs and symptoms when hypoglycaemia was induced. In the LPI group, hypoglycaemia occurred sooner (23.6 ± 1.6 vs. 28.3 ± 1.4 min, p < 0.05), and duration of hypoglycaemia (25.0 ± 1.7 vs. 31.9 ± 1.9 min, p < 0.05) and time for blood glucose levels to return to a ‘safe’ level (>3.3 mmol/l; 56.5 ± 2.3 vs. 76.0 ± 2.1 min, p < 0.001) were shorter as compared with the RGI group. No differences in peak growth hormone and cortisol levels were observed between the two groups. Conclusions: Our data suggest that despite inducing similar symptomatology, LPI exerted a quicker onset and a shorter duration of hypoglycaemia as compared with RGI. Thus, using LPI might reduce the potential risks associated with an ITT by shortening the hypoglycaemic phase of the test.