10.22376/ijpbs.2019.10.1.p1-12 Volume 3 Issue 4 2012(October - December) A rapid, reliable and sensitive LC-MS/MS method for the determination of N-acetylneuraminic acid (Neu5Ac) in human milk was developed and validated. Samples were hydrolysed with mild hydrochloric acid and analysed by LC-MS/MS on Thermo hypersilgold C-18 analytical column. The analytical procedure was found to be more selective, accurate, precise and linear. The method accuracy was 99.9 % and the mean precision (RSD) was 3.51 % and the calibration was linear from 0.1 to 10 µg mL lessThan sup greaterThan -1 lessThan /sup greaterThan (R lessThan sup greaterThan 2 lessThan /sup greaterThan greaterThan 0.99), the lowest limit of quantification (LLOQ) was 0.1 µg mL lessThan sup greaterThan -1 lessThan /sup greaterThan . N-glycolylneuraminicacid (Neu5Gc) was used as an internal standard. This method was proved as promising method for the determination of Neu5Ac in human milk samples. D.KARUNANITHI, A.RADHAKRISHNA AND V.M.BIJU N-acetylneuraminic acid (Neu5Ac). N-glycolylneuraminic acid (Neu5Gc). LC-MS/MS. Human milk 299-308