Time-resolved measurements of intrapuff nicotine yield in mainstream smoke have been performed with a specially designed intrapuff smoking apparatus (IPSA). The IPSA-filter traversing mechanism collects mainstream particles on a rectangular filter pad which moves at a constant velocity perpendicular to the direction of smoke flow at the mouthend of the cigarette. Filter pads were assayed by two analytical techniques. Standard gas chromatographic (GC) methodology was used to quantify nicotine mass in five equal time segments per puff. A second method, using a Berthold TLC-Linear Analyser, measured total radioactivity across the pad for samples from [2'-14C]-nicotine labelled cigarettes. Intrapuff nicotine concentrations were determined from measured puff flow-rate profiles and the collected masses of nicotine on the filters. GC nicotine concentration measurements correlated well with total activity from the scanner measurements. Studies carried out with filtered full-flavour cigarettes revealed that nicotine concentrations in the smoke vary significantly during a puff. This work provides a new technique for studying time-resolved yields of mainstream smoke components. It may potentially be used to elucidate mechanisms controlling the yield of nicotine and other mainstream smoke components.