Recent years saw an increased interest in biodiversity research regarding green spaces in urban areas around the globe. Wildlife studies of butterflies (Lepidoptera: Papilionoidea) in urban semi-natural habitats provide key information towards developing conservation strategies for such areas. This is especially true considering the charismatic aspect of these insects, as well as their ability to serve as bioindicators. The growing appreciation and interest of the general population towards biodiversity adds special importance to such studies. However, a firm understanding of the specieslevel diversity of butterflies is crucial to achieve this goal and such work is still lacking in many parts of the Neotropics. In the present study, we present an updated species checklist of butterflies for Guayaquil, Ecuador. Information of the species reported here was obtained from fieldwork, an intensive bibliographic review, and visits to entomological collections, along with information from the iNaturalist citizen science platform. A total of 557 valid records of butterflies are reported, which belong to 166 species that are represented in six families. A total of 48 species are reported for the first time for Guayaquil. Some areas, especially in parks and near protected areas are where more species can be found. This indicates that green areas in general should be preserved, managed appropriately, and increased to maintain this extraordinary urban butterfly biodiversity. We hope this new information will be utilized to increase the interest in the study of butterflies that inhabit Guayaquil. The purpose of this study is to improve our knowledge of the butterfly community, their interactions, and to design and implement adequate strategies for the conservation of the city's biodiversity.