Background: Emerging evidences have indicated that the composition of gut microbiota was significantly influenced by central nervous system diseases. The digestion and metabolism disturbances of patients with amyotrophic lateral sclerosis (ALS) might be strongly associated with ALS; however, this has rarely been evaluated in these populations. This study was to evaluate bacterial and archaeal composition of gut flora and the corresponding metabolism performance of these micro-organisms in fecal samples of patients with ALS. Methods: A comparative study was performed on the intestinal microbiota from eight patients with ALS and eight healthy individuals at Huadong Hospital during November 2017 to April 2018; meanwhile, the metabolite concentrations of human endotoxin, short-chain fatty acids (SCFA), NO₂-N/NO₃-N, and γ-aminobutyric acid were also evaluated by spectrophotometry methods. The correlations between intestinal microbiota and metabolite concentration were compared between the two groups using one-way analysis of variance; the relative abundance of beneficial and harmful micro-organisms in fecal samples was also analyzed. Results: In general, the richness and evenness of bacterial and archaeal communities of healthy individuals were healthier than that of patients with ALS. The phylum Firmicutes/Bacteroidetes ratio, genus Methanobrevibacter showed an enhancive tendency in patients with ALS, whereas the relative abundance of beneficial micro-organisms (genera Faecalibacterium and Bacteroides) presented a significant decrease tendency in patients with ALS. In addition, the average concentrations of human endotoxin, SCFA, NO₂-N/NO₃-N, and γ-aminobutyric acid in patients with ALS and healthy individuals were 64.2 vs. 65.3 EU/mL, 57.5 vs. 55.3 μg/mL, 5.7 vs. 5.3 ng/mL, and 6.1 vs. 5.4 μmol/L, respectively, indicating that the digestion and metabolism functions of gastrointestinal tract of patients might decline with this disease. Conclusions The relative abundance of beneficial and harmful micro-organisms respectively showed decrease and increase tendency in patients with ALS.