An analytical method for biotin is described, which is based on its conversion to N-nitrosobiotin, purifications of this derivative and its colorimetric or gas chromatographic determination. Nitrosation is carried out by treatment with an excess of NaNO2 in 0.2 N H2SO4 at room temperature. Nitrosobiotin is extracted with chloroform, after salting out wiulphanilamide + HCl), in order to obtain hydrolysis, and then with the second oating conditions are:a glass column, 2 m long, packed with 3% SE-30 on Chromosorb WHP; colum temperature 190 degrees C; injector and detector temperature 260 degrees C; carrier gas: nitrogen (30 ml/min). The extraction of biotin from multivation solid preparations is performed with absolute ethanol, starting from an expected quantity of about 250 microgram. It is suggested to work at the same time with an internal standard.