Abstract: Reference material (RM) of nicotine purity was prepared. Six laboratories collaboratively confirmed quantitatively the purity of the prepared material by the method recommended by ISO 13276:1997"Tobacco and tobacco products-Determination of nicotine purity-Gravimetric method using tungstosilicic acid". The results showed that:1) The purity of RM of nicotine was 98.5% with the expanded uncertainty of 0.6%. 2) The prepared RM featured good homogeneity test result and higher stability during investigating period, and satisfied the technical requirements of national second grade reference material.