Following a request from the European Commission, the Panel on Dietetic Products, Nutrition and Allergies (NDA) derived Dietary Reference Values (DRVs) for biotin. Biotin is a water-soluble vitamin which serves as a co-factor for several carboxylases that play critical roles in the synthesis of fatty acids, the catabolism of branched-chain amino acids and gluconeogenesis. Dietary biotin deficiency is rare. Data on biomarkers of biotin intake or status are insufficient to be used in determining the requirement for biotin. Data available on biotin intakes and health consequences are very limited and cannot be used for deriving DRVs for biotin. As there is insufficient evidence available to derive an Average Requirement and a Population Reference Intake, an Adequate Intake (AI) is proposed. The setting of AIs is based on observed biotin intakes with a mixed diet and the apparent absence of signs of deficiency in the EU, suggesting that current intake levels are adequate. The AI for adults is set at 40 µg/day. The AI for adults also applies to pregnant women. For lactating women, an additional 5 µg biotin/day over and above the AI for adults is proposed, to compensate for biotin losses through breast milk. For infants over six months, an AI of 6 µg/day is proposed by extrapolating from the biotin intake of exclusively breastfed infants aged zero to six months, using allometric scaling and reference body weight for each age group, in order to account for the role of biotin in energy metabolism. The AIs for children aged 1–3 and 4–10 years are set at 20 and 25 µg/day, respectively, and for adolescents at 35 µg/day, based on observed intakes in the EU.