Nicotine is a significant evaluation index of tobacco and its related products' quality, but nicotine overdose can pose serious health hazards and cause addiction and dependence, thus it can be seen that it is necessary to find suitable and efficient detection methods to precisely detect nicotine in diverse samples and complex matrices. In this review, an updated summary of the latest trends in pretreatment and analytical techniques for nicotine is provided. We reviewed various sample pretreatment methods, such as solid phase extraction, solid phase microextraction, liquid phase microextraction, QuEChERS, etc., and diverse nicotine assay methods including liquid chromatography, gas chromatography, electrochemical sensors, etc., focusing on the developments since 2015. Furthermore, the recent progress in the applications and applicability of these techniques as well as our prospects for future developments are discussed. HighlightsUpdated pretreatment and analysis methods of nicotine were systematically summarized.Microextraction and automation were main development trends of nicotine pretreatment.The introduction of novel materials added luster to nicotine pretreatment.The evolutions of ion source and mass analyzer were emphasized.