Enumeration of human colonic bacteria producing phenolic and indolic compounds: effects of pH, carbohydrate availability and retention time on dissimilatory aromatic amino acid metabolism

Abstract

Concentrations of phenolic compounds in human gut contents were more than fourfold higher in the distal colon (6.2 mmol kg-1) compared to the proximal bowel (1.4 mmol kg-1). Tryptophan metabolites were never found in more than trace amounts in large intestinal contents and phenol substituted fatty acids were the major products of aromatic amino acid fermentation that accumulated in the proximal colon, whereas phenol and p-cresol were more important in the distal gut, accounting for 70% of all products of dissimilatory aromatic amino acid metabolism. In vitro incubations of colonic material showed that phenol was produced most rapidly (1.0 mumol g-1 h-1), whereas indole was formed comparatively slowly (0.06 mumol g-1 h-1). Most probable number (MPN) estimations demonstrated that large populations of phenol and indole producing bacteria occur in the large intestine (range log10 9.8-11.5 (g dry wt faeces)-1, mean 10.6, N = 7). With respect to phenolic compounds, phenylacetate and phenylpropionate producers predominated, while indoleacetate-forming bacteria were the major tryptophan-utilizing organisms. Quantitation of products of dissimilatory aromatic amino acid metabolism in MPN tubes showed that phenol and phenylpropionate mainly accumulated at low sample dilutions, whereas phenylacetate, p-cresol, indoleacetate and indolepropionate were formed in greatest amounts at high sample dilutions. The significance of pH and carbohydrate availability with respect to aromatic amino acid metabolism was shown in batch culture fermentation studies, where net production of phenolic compounds by mixed populations of intestinal bacteria was reduced by approximately 33% during growth at pH 5.5 compared to pH 6.8, and by 60% in the presence of a fermentable carbohydrate. Experiments with 16 species of intestinal bacteria belonging to six different genera showed that environmental factors such as low pH and high carbohydrate availability markedly reduced dissimilatory aromatic amino acid metabolism in some organisms, but stimulated this process in others. A three-stage continuous culture model of the colon was used to investigate the effect of system retention time (27.1 or 66.7 h) on aromatic amino acid fermentation. Qualitative and quantitative increases in phenol production occurred from vessel 1 to vessel 3 in this model. Concentrations of phenolic compounds in vessel 3 were three times greater at R = 66.7 h compared to R = 27.1 h. Phenol and p-cresol were not detected in vessel 1, though formation of these metabolites increased from vessel 2 to vessel 3, in a pattern similar to that observed in the distal colon.

Gut microbiome development along the colorectal adenoma–carcinoma sequence

“…Even though putrefactive bacteria such as Alistipes and Bacteroides could produce short chain fatty acid from amino acids, carbohydrate fermentation is still preferred 24,39 , which might explain protective roles of fruits and vegetables. In some Fusobacterium species, however, transport of sugar depends on amino acid fermentation (Glu, Lys, His or Ser) 40,41 , suggesting that they only thrive in the presence of an ample supply of amino acids.…”

“…The lactic acid produced might help lower the pH and inhibit amino acid degradation in the colon 24,39 . Lactobacillus and Bifidobacterium have been found to stimulate NADPH oxidase 1-dependent ROS generation and intestinal stem cell proliferation 47 , and lactate was reported to accelerate colon epithelial cell turnover in starvation-refed mice 48 .…”

“…In some Fusobacterium species, however, transport of sugar depends on amino acid fermentation (Glu, Lys, His or Ser) 40,41 , suggesting that they only thrive in the presence of an ample supply of amino acids. Phenolic compounds are produced from fermentation of the aromatic amino acids phenylalanine and tyrosine 39 , which might increase DNA damage in the colon. Bile acid metabolism by Bacteroides species and B. wadsworthia would also affect gut microbial composition and impact host physiology 42,43 .…”

Hypothesis: Phenol and hydroquinone derived mainly from diet and gastrointestinal flora activity are causal factors in leukemia

“…A major source of phenol stems from catabolism of protein and other compounds by gut bacteria, which appears to depend highly on the metabolic activity of the intestinal bacterial microflora. 57,62 A potentially significant source of hydroquinone comes from ingestion of foods containing arbutin, a naturally occurring plant product that is converted to hydroquinone by stomach acids. The wide inter-individual variability in urinary levels of phenol noted above may reflect the wide range of direct intake of phenolic-containing foods and differences among individuals in the composition and chemistry of the gut flora.…”

“…High background concentrations of phenol, hydroquinone, catechol and 1,2,4-benzenetriol have been measured in the blood, urine and intestines of presumably unexposed humans [48][49][50][51][52][53][54][55][56][57][58][59][60][61][62] and in the blood and urine of rodents. [63][64][65] Representing measurements of hundreds of individuals in total, these studies showed that mean urinary phenol and hydroquinone levels were about 10 p.p.m.…”