Human　milk　oligosaccharides　(HMOs)　are　the　third　most　abundant　solid　component　in　breast　milk,　which　is　essential　for　the　healthy　growth　of　infants.　2　＇-fucosyllactose　(2＇-FL)　and　3-fucosyllactose　(3-FL)　are　the　main　components　of　HMOs.　In　this　study,　we　developed　and　validated　a　quantitative　nuclear　magnetic　resonance　(qNMR)　method　for　determining　HMOs　in　different　complex　food　matrices.　The　spectra　of　HMOs　often　overlap　with　impurities　in　the　samples,　which　hampers　quantification　using　the　conventional　integral　method.　We　overcame　this　obstacle　by　building　the　Global　Spectrum　Deconvolution-area　coefficients　algorithm.　The　method　proved　to　be　precise,　yielding　satisfactory　results　in　terms　of　precision　(RSD%　0.5-1.9),　trueness　(bias%　1.9-8.2),　and　recovery　(90.5-106.6　%).　The　limit　of　quantification　for　2＇-FL　was　0.10　mg/mL,　and　for　3FL,　it　was　0.15　mg/mL.　A　comparison　using　Deming　regression　between　the　existing　high-performance　liquid　chromatography　method　and　the　developed　qNMR　method　for　determining　HMOs　demonstrated　the　high　accuracy　of　the　qNMR　method.　The　proposed　method　allows　for　high-throughput　measurements　of　large-volume　samples　due　to　its　simple　and　fast　sample　preparation.　Therefore,　the　method　is　an　essential　tool　for　determining　HMOs　in　various　complex　foods.