The Metabolomic-Gut-Clinical Axis of Mankai Plant-Derived Dietary Polyphenols

Abstract

Background: Polyphenols are secondary metabolites produced by plants to defend themselves from environmental stressors. We explored the effect of Wolffia globosa 'Mankai', a novel cultivated strain of a polyphenol-rich aquatic plant, on the metabolomic-gut clinical axis in vitro, in-vivo and in a clinical trial.

Methods: We used mass-spectrometry-based metabolomics methods from three laboratories to detect Mankai phenolic metabolites and examined predicted functional pathways in a Mankai artificial-gut bioreactor. Plasma and urine polyphenols were assessed among the 294 DIRECT-PLUS 18-month trial participants, comparing the effect of a polyphenol-rich green-Mediterranean diet (+1240 mg/polyphenols/day, provided by Mankai, green tea and walnuts) to a walnuts-enriched (+440 mg/polyphenols/day) Mediterranean diet and a healthy controlled diet.

Results: Approximately 200 different phenolic compounds were specifically detected in the Mankai plant. The Mankai-supplemented bioreactor artificial gut displayed a significantly higher relative-abundance of 16S-rRNA bacterial gene sequences encoding for enzymes involved in phenolic compound degradation. In humans, several Mankai-related plasma and urine polyphenols were differentially elevated in the green Mediterranean group compared with the other groups (p < 0.05) after six and 18 months of intervention (e.g., urine hydroxy-phenyl-acetic-acid and urolithin-A; plasma Naringenin and 2,5-diOH-benzoic-acid). Specific polyphenols, such as urolithin-A and 4-ethylphenol, were directly involved with clinical weight-related changes.

Conclusions: The Mankai new plant is rich in various unique potent polyphenols, potentially affecting the metabolomic-gut-clinical axis.

Keywords: Mediterranean diet; Wolffia globosa; flavonoids; plant-based nutrition; polyphenols; weight loss.

Figure 1

(A–D) Putative identification of polyphenolic compounds from LC-MS measurements (experiment 3). A. total ion chromatogram (TIC) of Wolffia globosa ‘Mankai’ extract acquired in negative ionization mode (ESI-); inlay: UV-absorption spectrum of compound eluting at 8.8 min. B. extracted ion chromatogram of mass-to-charge ratio (m/z) 447.09. C. background corrected mass spectrum of peak at retention time 8.81 min; two masses are detected m/z 447.0936 assigned as [M-H]⁻ and the dimer of 447.0936, m/z 895.1968 assigned as [2M-H]⁻; elemental composition of the ion m/z 447.0936 was calculated to C₂₁H1₉O₁₁- with a mass error of 2 ppm. The molecular formula corresponds to putative 8-hexosyl-luteolin. D. mass fragmentation spectrum acquired in MS^E mode (MS^E) spectrum of m/z 447.0936 confirms assignment as 8-C-hexosyl-luteolin. Major fragments are shown with structure and elemental composition. EIC: extracted ion chromatogram.

Figure 2

(A,B) Example of a UV spectra of the metabolites (experiment 4). (A). Flavonoid group quercetin and kaempferol derivatives. (B). Cinnamic group caffeoyl and coumaroyl derivatives.

Figure 3

Relative abundance of predicted microbial pathways in Mankai-supplemented artificial gut bioreactors.

Figure 4

(A,B) Differentially detected plasma polyphenols between groups at the end of the intervention. A: Naringenin: out of the three groups, the highest detection was among the green-MED dieters (65.27% of all detection in whole DIRECT PLUS samples), followed by the MED (30.43% detection) and HDG (4.3% detection) groups; p = 0.001. B: 2,5 diOH benzoic acid: green-MED group showed that highest detection (detection of 50.75%), followed by the MED (37.34%) and HDG (11.91%); p = 3.7 × 10⁻⁵. Differences between groups were calculated using the Chi-square test. HDG, Healthy dietary guidelines; MED, Mediterranean.

Figure 5

(A–C) Differential six-month change (relative change, log-transformed) of urine polyphenols, between-group comparisons. Between-group changes are corrected for multiple comparisons. Data presented as means of log change and SEM. A: Urine polyphenol annotated to 6-month hydroxy phenyl acetic acid: p = 1.5 × 10⁻⁴, q = 1.7 × 10⁻³. B: Urine polyphenol annotated to 6-month urolithin A: p = 2.9 × 10⁻⁴, q = 3.6 × 10⁻³. C: Urine polyphenol also annotated to 6-month urolithin A: p = 0.002, q = 0.007.