Fuel injection during negative valve overlap (NVO) can extend low-load gasoline HCCI operation through control of main combustion phasing. Reactions and heat release accompanying NVO fuel injection give rise to changes in temperature and composition of the charge prior to main combustion. The extent of reaction of injected NVO fuel and the relative importance of resulting thermal and chemical effects on main combustion are a current research topic.In this work, bulk temperature computations are used to quantify thermal conditions prior to main ignition for cases with and without NVO fueling. To separate measured thermal effects from chemical effects of NVO fuel reactions on the main combustion, cases without NVO fuel but with similar mixture temperatures and combustion phasing are compared. Effects of varying NVO fuel amount and injection timing on heat release, combustion phasing, bulk temperature evolution, and iso-octane ignition temperatures are analyzed. Chemical kinetic simulations using detailed and reduced mechanisms corroborate some general experimental trends, giving insight into the species and reactions causing observed effects.