The longevity and excellent health status of the population of Crete has been attributed to its lifestyle and dietary habits. The impact of Greek Orthodox Christian Church fasting on these dietary habits has never been studied. One hundred and twenty Greek Orthodox Christians living in Crete participated in a 1-year prospective study. One half of the subjects, who fasted regularly (fasters), and sixty non-faster controls were followed longitudinally for the three main fasting periods over 1 year; Christmas (40 d), Lent (48 d) and the Assumption (15 d). Pre- and end-holy days measurements were performed in each fasting period including: 24 h dietary recall, blood collection and anthropometric measurements. Based on the 24 h recall, fasters as compared with controls had lower intakes of end-holy days dietary cholesterol, total fat, saturated fatty acids, trans-fatty acids and protein (P < 0.001). Fasters presented a decrease of 753 kJ (180 kcal) in end-holy days energy intake (P < 0.05) compared with an increase of 573 kJ (137 kcal) in the controls (P < 0.05). Fasters had a decrease in end-holy days Ca intake (P < 0.001) and an increase in end-holy days total dietary fibre (P < 0.001) and folate (P < 0.05), attributed to their higher consumption of fruit and vegetables in end-holy periods (P < 0.001). There were no differences for other vitamins or minerals between pre- and end-holy periods in both groups except for vitamin B2. The Orthodox Christian dietary regulations are an important component of the Mediterranean diet of Crete characterised by low levels of dietary saturated fatty acids, high levels of fibre and folate, and a high consumption of fruit, vegetables and legumes.