The effect of drinking water pH on the human gut microbiota and glucose regulation: results of a randomized controlled cross-over intervention

Abstract

Studies in rodent models have shown that alterations in drinking water pH affect both the composition of the gut microbiota and host glucose regulation. To explore a potential impact of electrochemically reduced alkaline (pH ≈ 9) versus neutral (pH ≈ 7) drinking water (2 L/day) on human intestinal microbiota and host glucose metabolism we conducted a randomized, non-blinded, cross-over study (two 2-week intervention periods, separated by a 3-week wash-out) in 29 healthy, non-smoking Danish men, aged 18 to 35 years, with a body mass index between 20.0 to 27.0 kg m-2. Volunteers were ineligible if they had previously had abdominal surgery, had not been weight stabile for at least two months, had received antibiotic treatment within 2 months, or had a habitual consumption of caloric or artificially sweetened beverages in excess of 1 L/week or an average intake of alcohol in excess of 7 units/week. Microbial DNA was extracted from faecal samples collected at four time points, before and after each intervention, and subjected to 16S rRNA gene amplicon sequencing (Illumina MiSeq, V4 region). Glycaemic regulation was evaluated by means of an oral glucose tolerance test.No differential effect of alkaline versus neutral drinking water was observed for the primary outcome, overall gut microbiota diversity as represented by Shannon's index. Similarly, neither a differential effect on microbiota richness or community structure was observed. Nor did we observe a differential effect on the abundance of individual operational taxonomic units (OTUs) or genera. However, analyses of within period effects revealed a significant (false discovery rate ≤5%) increase in the relative abundance of 9 OTUs assigned to order Clostridiales, family Ruminococcaceae, genus Bacteroides, and species Prevotella copri, indicating a potential effect of quantitative or qualitative changes in habitual drinking habits. An increase in the concentration of plasma glucose at 30 minutes and the incremental area under the curve of plasma glucose from 0 30 and 0 120 minutes, respectively, was observed when comparing the alkaline to the neutral intervention. However, results did not withstand correction for multiplicity. In contrast to what has been reported in rodents, a change in drinking water pH had no impact on the composition of the gut microbiota or glucose regulation in young male adults. The study is registered at www.clinicaltrials.gov (NCT02917616).

Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1

Flowchart.

Figure 2

Principal coordinate plots of faecal microbiota community structure. Principal coordinate plots (axes 1 and 2) of treatment effect on community structure based on unweighted UniFrac (A,D,G), weighted UniFrac (B,E,H), and Bray-Curtis distances (C,F,I). Samples from the alkaline (A–C) and neutral water intervention (D–F), as well as post-intervention samples from both periods (G–I) are depicted separately. Samples from the same participant are connected by solid lines. P-values are from permutational multivariate analysis of variance.

Figure 3

Treatment effect on OTU composition. Change (%) in geometric means of relative abundance and corresponding P-values derived from linear mixed models of treatment effect i.e. alkaline versus neutral water. Prevalence indicates number of participants in which a given OTU is present in at least one sample. Abundance indicates mean relative abundance of a given OTU in baseline samples. Taxonomy of OTUs [Greengenes ID] is given at the lowest classified rank. Only OTUs with P ≤ 0.05 are annotated.

Figure 4

Intervention effect on OTU composition. Change in relative abundance (% increase/decrease in geometric mean) of core OTUs during the neutral water (A) and alkaline (B) water interventions. Red dotted line indicates the cut-off corresponding to a Q-value of 0.05, and the blue dotted line indicates a P-values of 0.05. Prevalence indicates number of participants in which a given OTU is present in at least one sample. Abundance indicates mean relative abundance (‰) of a given OTU in baseline samples. Taxonomy of OTUs [Greengenes ID] is given at the lowest classified rank. o, order. f, family. g, genus.

Figure 5

Treatment effect on glucose and insulin during an oral glucose tolerance test. Curves are mean (±SEM) plasma glucose (A) and serum insulin (B) sampled at 5 time-points during an oral glucose tolerance test. P-value for difference in plasma glucose at 30 min from linear mixed model adjusted for age and body-mass index.