Evaluation of the Feasibility of In Vitro Metabolic Interruption of Trimethylamine with Resveratrol Butyrate Esters and Its Purified Monomers

Abstract

Resveratrol (RSV), obtained from dietary sources, has been shown to reduce trimethylamine oxide (TMAO) levels in humans, and much research indicates that TMAO is recognized as a risk factor for cardiovascular disease. Therefore, this study investigated the effects of RSV and RSV-butyrate esters (RBE) on the proliferation of co-cultured bacteria and HepG2 cell lines, respectively, and also investigated the changes in trimethylamine (TMA) and TMOA content in the medium and flavin-containing monooxygenase-3 (FMO3) gene expression. This study revealed that 50 µg/mL of RBE could increase the population percentage of Bifidobacterium longum at a rate of 53%, while the rate was 48% for Clostridium asparagiforme. In contrast, co-cultivation of the two bacterial strains effectively reduced TMA levels from 561 ppm to 449 ppm. In addition, regarding TMA-induced HepG2 cell lines, treatment with 50 μM each of RBE, 3,4'-di-O-butanoylresveratrol (ED2), and 3-O-butanoylresveratrol (ED4) significantly reduced FMO3 gene expression from 2.13 to 0.40-1.40, which would also contribute to the reduction of TMAO content. This study demonstrated the potential of RBE, ED2, and ED4 for regulating TMA metabolism in microbial co-cultures and cell line cultures, which also suggests that the resveratrol derivative might be a daily dietary supplement that will be beneficial for health promotion in the future.

Keywords: co-cultured; metabolism; probiotic; short-chain fatty acid (SCFA); trimethylamine-N-oxide (TMAO).

Figure A1

The growth curves for C. asparagiforme under (A) PY + X medium, (B) PY + X: MRS medium (1:1; v/v), (C) MRS medium, and (D) PY + X: MRS medium (1:1; v/v) medium against the influence of B. longum.

Figure 1

Effects of resveratrol (RSV) and RSV-butyrate esters (RBE) on bacterial cultures: (A) effects on the growth of C. asparagiforme and B. longum expressed as bacterial population percentage (%); (B) effects on the metabolic choline production of trimethylamine (TMA; ppm) by C. asparagiforme; (C) effects for a single culture of C. asparagiforme or co-culture of C. asparagiforme and B. longum of choline metabolized into TMA (ppm); and (D) effects of RSV and RBE treated with 50 μg/mL each on the metabolic choline yield of TMA in a single culture of C. asparagiforme and a co-culture of C. asparagiforme and B. longum. Different letters indicate that the statistically different from other (p < 0.05).

Figure 2

Effects of different concentrations of samples on HepG2 cell lines treated with (A) resveratrol butyrate esters (RBE); (B) 3,4′-di-O-butanoylresveratrol (ED2, di-butyric acid derivative); (C) 3-O-butanoylresveratrol (ED4); and (D) trimethylamine (TMA). (E) Effects of TMA treatment at different times on the amount of trimethylamine oxide (TMAO) produced by HepG2 cell lines. Different letters indicate that the statistically different from other (p < 0.05).

Figure 3

Effects of the different samples treated on flavin-containing monooxygenase-3 (FMO3) mRNA gene expression in the HepG2 cell line: (A) trimethylamine (TMA) treatment for 0–180 min; (B) resveratrol butyrate esters (RBE), 3,4′-di-O-butanoylresveratrol (ED2, di-butyric acid derivative), 3-O-butanoyl resveratrol (ED4), and TMA treatment for 180 min. Different letters indicate that the statistically different from other (p < 0.05).