Abstract: There are 4 main types of N-alkylamides in Anacyclus pyrethrum DC., namely, isobutylamide (IBA), N-Me isobutylamide (N-Me IBA), 2-phenylethylamide (2-PEA) and 4-hydroxyphenylethylamide (4-OH PEA). However, there was very few research on the fragmentation pathways of N-alkylamides by now. In order to obtain accurate and comprehensive information about the fragmentation pathways of N-alkylamides, 5 kinds of N-alkylamides were analyzed by HPLC-ESI-MSⁿ. They are identified as deca-2E, 4E-dienoicacid isobutylamideylamide, undeca-2E, 4E-diene-8, 10-diynoic acid phenylethylamide, deca-2E, 4E-dienoicacid 4-hydroxyphenylethlamide, dodeca-2E, 4E-dienoicacid 4-hydroxyphenyl-ethylamide and tetradeca-2E, 4E-dienoic acid 4-hydroxyphenylethylamide. Multi-stage mass spectrum diagrams of five N-alkylamides were obtained in positive ion mode. Simultaneously, the method of quantum computational chemistry was adopted to verify their fragmentation pathways. Characteristic fragment ions were identified and the patterns were deduced further. The fragmentation patterns and structural assignment of ‘isobutylamides (IBA)’type, ‘2-phenylethylamide(2-PEA)’ type and ‘4-hydroxyphenylethyl-amides (4-OH PEA)’ type in ESI-MSⁿ under positive ion mode were summarized. The results show that five N-alkylamides have the similar fragmentation pathway, M+H⁺ is prone to α-cracking in N-position at the first, loss of 28(CO), 18(H₂O) subsequently. These compounds have strong pyrolysis rules and the study can provide reference for N-alkylamides in structural identification and quantitative analysis.