Objective:To establish a simultaneous determination of linoleic acid(LA)

α-linolenic acid(ALA)

arachidonic acid(AA)and docosahexaenoic acid(DHA)in human breast milk by reverse phase high performance liquid chromatography(RP-HPLC).Methods:The method of liquid-liquid extraction and derivation from human breast milk was used in a HPLC analysis.The column was Agilent SB-C18(4.6 mm×150 mm

5 μm).The mobile phase was acetonitrile:water(83:17

v/v)at 1.0 ml/min with UV detection at 254 nm and the column temperature was 30℃.Results:The standard curves were linear over the ranges of 50 ～350 μg/ml for α-linolenicacid

200～1500μg/mlforlinoleicacid

10～80 μg/ml for docosahexaenoic acid and 20 ～160 μg/ml for arachidonic acid

respectively.The good linearity can be obtained over their concentration range(r>0.999).The minimumdetectable limits of ALA

LA and DHA were all 0.2 μg/ml

and AA was 0.4 μg/ml.The average recovery of the method was 92.65%～100.70% and the RSD was lower than 5.67%.Conclusion:The HPLC method in this assay is rapid

direct and sensitive

which can be successfully applied to quantity the detection of polyunsaturated fatty acid of different biological samples as well as in clinic.