Objective Since there are significant variation of the dietary structure recent years in China,it is necessary to re-investigate the fatty acid composition of human breast milk for the presentation of the latest data of fatty acid composition in China. Methods Using a gas chromatography GC-2010,the composition of fatty acids was detected in the human colostrums and the mature breast milk(consecutively from postnatal day 1 to day 7 and from postnatal day 42)obtained from 62 healthy postpartum women in Shanghai and Chongqing,two big cities of China,from Jan to July,2008. Results The level of total fatty acid(TFA)tended to increase significantly from the colostrums to the mature breast milk. No significant difference in the level of TFA was found between two cities. The significantly higher monounsaturates(MUFA)level(44.06% vs. 33.85%,P ＜ 0.01)and lower linoleic acid(LA,C18 : 2n-6)level(18.43% vs. 27.62%,P ＜ 0.01)of the mature breast milk were observed in Chongqing women compared with those in Shanghai women. The docosahexenoic acid(DHA)level of the mature breast milk in Shanghai women was significantly higher than that in Chongqing women(0.41% vs. 0.29%,P ＜ 0.01). There was no significant difference in the level of arachidonic acid(AA,C20 : 4n-6)between two cities. Conclusions The fatty acid composition in human breast milk tends to vary with the extension of the lactation. There is significant difference in the fatty acid compositions in human breast milk between Shanghai and Chongqing owing to different dietary habits in the different regions of China.