Peppermint oil effects on the gut microbiome in children with functional abdominal pain

Abstract

Peppermint oil (PMO) is effective in the treatment of functional abdominal pain disorders, but its mechanism of action is unclear. Evidence suggests PMO has microbicidal activity. We investigated the effect of three different doses of PMO on gut microbiome composition. Thirty children (7-12 years of age) with functional abdominal pain provided a baseline stool sample prior to randomization to 180, 360, or 540 mg of enteric coated PMO (10 participants per dose). They took their respective dose of PMO (180 mg once, 180 mg twice, or 180 mg thrice daily) for 1 week, after which the stool collection was repeated. Baseline and post-PMO stools were analyzed for microbiome composition. There was no difference in alpha diversity of the gut microbiome between the baseline and post-PMO treatment. Principal coordinate analysis revealed no significant difference in overall bacterial composition between baseline and post-PMO samples, as well as between the PMO dose groups. However, the very low abundant Collinsella genus and three operational taxonomic units (one belonging to Collinsella) were significantly different in samples before and after PMO treatment. The Firmicutes/Bacteroidetes ratio was lower in children who received 540 mg of PMO compared to the 180 mg and 360 mg dose groups (p = 0.04). Network analysis revealed separation between pre- and post-PMO fecal samples with the genus Collinsella driving the post-PMO clusters. PMO administration appeared to impact only low abundance bacteria. The 540 mg PMO dose differentially impacted the Firmicutes/Bacteroidetes ratio. A higher dose and/or longer duration of treatment might yield different results.

Conflict of interest statement

The authors declared no competing interests for this work.

© 2021 The Authors. Clinical and Translational Science published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.

Figures

FIGURE 1

Estimators of microbial alpha diversity in baseline and PMO dose groups. Box plots showing Shannon diversity index (a), Simpson diversity index (b), variance in Shannon diversity index (c), and variance in Simpson diversity index (d) between baseline and post‐PMO samples. The p values between baseline and post‐PMO samples were calculated using Wilcoxon rank sum test. The variance of diversity in the baseline samples was significantly greater than that in the post‐PMO samples (1000 bootstraps followed by a Wilcoxon rank sum test p < 0.05) for both Shannon (c) and Simpson (d) indices. PMO, peppermint oil

FIGURE 2

Alpha diversity among PMO dose groups. Box plots of Shannon (a) and Simpson (b) diversity indices plotted among the PMO dose groups. The p values were calculated using pairwise Wilcoxon test. p.adj = adjusted p value after false discovery rate (FDR) correction. PMO, peppermint oil

FIGURE 3

Bray‐Curtis dissimilarity‐based microbial beta diversity and inter‐individual divergence in baseline and PMO dose groups. (a) Principal coordinate analysis (PCoA) plot of the Bray‐Curtis index between the baseline and post‐PMO cohorts. (b) Inter‐individual divergence in the bacterial community composition across samples in the baseline and post‐PMO cohorts. (c) Bray‐Curtis indexbased PCoA plot of the samples grouped by baseline and PMO dose groups. (d) Bray‐Curtis indexbased PCoA plot of the samples grouped by PMO dose groups. The percentage of variance accounted for by the first two principal coordinates (PCo) that explained the largest fractions of variability in our data are shown in the axis labels in the PCoA plots. Each point represents a sample and closeness of the points indicates high similarity in the microbial community. The ellipses were drawn at 95% confidence interval. PMO, peppermint oil

FIGURE 4

Gut microbiome composition of baseline and post‐PMO samples. Stacked bar plots showing the average relative abundance (%) of 16S V4 rRNA gene sequences assigned to each bacterial phylum (a), family (b), and genus (c). Taxa with less than 1% relative abundance were grouped together for visualization. A higher taxonomic level is reported if the genus level classification could not be assigned. Unc = unclassified. Box plots showing differently abundant (FDR‐corrected p < 0.05) family (d) and genus (e) levels taxa in baseline and post‐PMO groups. FDR, false discovery rate; PMO, peppermint oil

FIGURE 5

Genus level bacterial network in pre‐PMO (baseline) and post‐PMO samples. Cytoscape was used to generate the network. The subjects and genera were interpreted as nodes and the quantiles as weights of the connecting edges. The network included only genera that were present in at least five samples, had a Wilcoxon p less than or equal to 0.05, and abundance/edges that remained after removing the lowest 0, 0.1, and 0.2 quantiles. PMO, peppermint oil

FIGURE 6

Gut microbiome composition at baseline and in PMO dose groups. Phylum (a) and genus (b) levels bar graphs by PMO dosing groups. Taxa with less than 3% relative abundance were grouped together as “Others” for visualization. Baseline samples are also included in the plots for comparisons. PMO, peppermint oil