This research aimed to demonstrate the feasibility of producing biobutanol by acetone-butanol-ethanol (ABE) fermentation on vine shoots at laboratory scale. In order to avoid a detoxification process prior to fermentation, an alkaline pretreatment and its posterior enzymatic hydrolysis were optimized through a response surface methodology (RSM) approach to minimize the concentration of phenolic inhibitors and maximize the amount of fermentable sugars. This vine shoots hydrolysate was directly fermented by 11 solventogenic Clostridium strains, reaching butanol concentrations over 8.0 g/L with C. beijerinckii CECT 508 and DSM 6423. The strain CECT 508 was selected to tune up a fermentation medium with the minimum possible amount of nutrients through a Plackett-Burman experimental design, thus reducing the cost of the fermentation broth while ensuring a high ABE solvent production. Finally, it was also shown that CaCO3 could be substituted by eggshell powder as buffering agent, maintaining a biobutanol production over 7 g/L. This study demonstrates, for the first time, that biobutanol production from vine shoots is possible and that cost reduction based on alternative strategies of nutrient supplementation is viable.