Aims This study evaluated whether by‐products from industrial processing of acerola (Malpighia glabra L.; AB) and guava (Psidium guajava L.; GB) fruit may stimulate the growth and metabolism of probiotic Lactobacillus and Bifidobacterium and induce changes in human colonic microbiota. Methods and Results The ability of non‐digested and digested AB or GB to stimulate the growth ad metabolism of Lactobacillus acidophilus LA‐05, Lactobacillus casei L‐26 and Bifidobacterium animalis subsp. lactis BB‐12 was evaluated. Changes in populations of distinct bacterial groups of human colonic microbiota induced by digested AB and GB were evaluated using an in vitro colonic fermentation system. Non‐digested and digested AB and GB favoured probiotic growth. No difference among counts of probiotics in media with glucose, fructooligosaccharides and non‐digested and digested AB and GB was found during a 48‐h cultivation. Cultivation of probiotics in media with non‐digested and digested AB and GB resulted in decreased pH, increased organic acid production and sugar consumption over time. Digested AB and GB caused overall beneficial changes in abundance of Bifidobacterium spp., Lactobacillus–Enterococcus, Eubacterium rectall–Clostridium coccoides and Bacteroides–Provotella populations, besides to decrease the pH and increase the short‐chain fatty acid production during a 24‐h in vitro colonic fermentation. Conclusion AB and GB could be novel prebiotic ingredients because they can stimulate the growth and metabolism of probiotics and induce overall beneficial changes in human colonic microbiota. Significance and Impact of the Study AB and GB stimulated the growth and metabolism of probiotics, in addition to induce beneficial alterations in human colonic microbiota composition and increase short‐chain fatty acid production. These results characterize AB and GB as potential prebiotic ingredients and fruit processing by‐products as sources of added‐value compounds.

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