The preparation and detection of the nuclear singlet state are valuable and prospective for many applications related with compound molecular analysis. In this work, a proton spin coupled system containing two H nuclei in tripeptide (Ala-Gly-Gly, AGG) was selected as research object, and singlets were prepared with three sets of different pulse sequences based on different principles. The lifetime of those singlets were measured, and the singlet preparation efficiencies of different sequences were compared. The results indicated that for the same spin coupling system of the same molecule, there is no significant difference in the lifetime of the singlet state prepared by different pulse sequences, while their preparation efficiencies could be relatively different.