The use of kiwifruit juice as a source of metal-reducing and stabilizing agents for the production of gold nanoparticles (AuNPs) was investigated. The reaction was carried out in batch by mixing appropriate amounts of kiwifruit juice and gold(III) chloride solution. The formation of AuNPs was monitored by measuring the intensity of the surface plasmon resonance (SPR) band of gold. The effects of temperature (20–60 °C), pH (8–12) and gold-to-polyphenol ratio (GPR) were investigated. Characterisation of AuNPs by XRD, DLS and zeta-potential measurements showed that they were highly crystalline, with an average hydrodynamic diameter of about 50 nm and a zeta-potential ranging between –29.2 and –21.7 mV. Under the best reaction conditions (60 °C, pH 9, GPR = 5 mol Au3+/mol GAE), AuNPs with an average size of about 30 nm were produced. The results obtained suggest that kiwifruit juice is a suitable medium for the production of small-sized and stable AuNPs.