Trimethylamine N-oxide (TMAO), an important proatherogenic uremic toxin, is oxidized by hepatic-flavin monooxygenases from gut microbiome-generated trimethylamine (TMA). The present study aims to explore whether manipulating the gut microbiota by inulin-type fructans (ITFs) can reduce circulating TMAO levels in peritoneal dialysis patients.This is a randomized, double-blind, placebo-controlled, crossover trial with 10 g day-1 ITFs intervention for 3 months in continuous ambulatory peritoneal dialysis patients. The gut microbiome is measured, and TMA-producing gene clusters are annotated using shotgun metagenomic sequencing. Fecal and plasma TMA, plasma TMAO, and daily urine excretion and dialysis removal of TMAO are measured. Finally, 22 participants complete the trial. The daily intake of macronutrients and TMAO precursors is comparable during the prebiotics, washout, and placebo interventions. The ITFs intervention increases the Firmicutes/Bacteroidetes (F/B) ratio (p = 0.049) of gut microbiome. However, no significant influences are observed on fecal TMA content, circulating TMAO levels, or TMA-producing gene clusters, including choline TMA-lyase (CutC/D), carnitine monooxygenase (CntA/B), and betaine reductase (GrdH).Intervention with 10 g day-1 of ITFs for 3 months is not sufficient to reduce plasma TMAO levels in peritoneal dialysis patients, but it improves the gut microbiome composition.© 2023 Wiley-VCH GmbH.