Background: Dietary exposure to environmental pollutants in humans is an important public health concern. While long-term fasting interrupts the dietary exposure to these substances, fat mobilization as an energy source may also release bioaccumulated substances. This was, to our knowledge, only investigated in obese people decades ago. This study explored the effects of 10-days fasting on the excretion of heavy metals and glyphosate. Methods: Urinary levels of arsenic, chromium, cobalt, lead, nickel, mercury and glyphosate were measured before and after 10 fasting days in 109 healthy subjects. Additionally, hair analysis was done before and ten weeks after fasting in 22 subjects. Results: Fasting caused a decrease in body weight, and in urinary arsenic (by 72 %) and nickel (by 15 %) concentrations. A decrease in lead hair concentrations (by 30 %) was documented. Urinary mercury levels were unchanged for chromium, cobalt and glyphosate, which were undetectable in most of the subjects.