Effect of egg ingestion on trimethylamine-N-oxide production in humans: a randomized, controlled, dose-response study

Abstract

Background: It is important to understand whether eating eggs, which are a major source of dietary choline, results in increased exposure to trimethylamine-N-oxide (TMAO), which is purported to be a risk factor for developing heart disease.

Objective: We determined whether humans eating eggs generate TMAO and, if so, whether there is an associated increase in a marker for inflammation [ie, high-sensitivity C-reactive protein (hsCRP)] or increased oxidation of low-density lipoprotein (LDL).

Design: In a longitudinal, double-blind, randomized dietary intervention, 6 volunteers were fed breakfast doses of 0, 1, 2, 4, or 6 egg yolks. Diets were otherwise controlled on the day before and day of each egg dose with a standardized low-choline menu. Plasma TMAO at timed intervals (immediately before and 1, 2, 4, 8, and 24 h after each dose), 24-h urine TMAO, predose and 24-h postdose serum hsCRP, and plasma oxidized LDL were measured. Volunteers received all 5 doses with each dose separated by >2-wk washout periods.

Results: The consumption of eggs was associated with increased plasma and urine TMAO concentrations (P < 0.01), with ∼14% of the total choline in eggs having been converted to TMAO. There was considerable variation between individuals in the TMAO response. There was no difference in hsCRP or oxidized LDL concentrations after egg doses.

Conclusions: The consumption of ≥2 eggs results in an increased formation of TMAO. Choline is an essential nutrient that is required for normal human liver and muscle functions and important for normal fetal development. Additional study is needed to both confirm the association between TMAO and atherosclerosis and identify factors, microbiota and genetic, that influence the generation of TMAO before policy and medical recommendations are made that suggest reduced dietary choline intake.

Trial registration: ClinicalTrials.gov NCT01906554.

© 2014 American Society for Nutrition.

Figures

FIGURE 1.

Concentration of TMAO in plasma. Six healthy volunteers (subjects) consumed a standardized low-choline diet on the day before each of 5 randomly assigned doses of 0, 1, 2, 4, or 6 egg yolks. Subjects consumed the egg dose for breakfast followed by the same standardized low-choline lunch, dinner, and snacks as consumed the previous day. Each egg dose delivered 1.14 mmol (119 mg) total choline. Therefore, the 0, 1, 2, 4, and 6 egg doses provided 0, 1.1, 2.3, 4.6, and 6.8 mmol (0, 119, 238, 476, and 714 mg) total choline, respectively, in addition to the standardized diet. TMAO was measured in plasma collected from each volunteer immediately before and 1, 2, 6, 8, and 24 h after the egg dose was consumed by using liquid chromatography and electrospray ionization–isotope dilution mass spectrometry. Doses were separated by a 2–4-wk washout period. Doses are represented as follows: 0, diamond with dotted line; 1, square with solid light-gray line; 2, triangle with solid-gray line; 4, circle with solid dark-gray line; and 6, asterisks with black solid line. TMAO, trimethylamine-N-oxide.

FIGURE 2.

Plasma TMAO. Six healthy volunteers (subjects) consumed a standardized low-choline diet on the day before each of 5 randomly assigned doses of 0, 1, 2, 4, or 6 egg yolks. Subjects consumed the egg dose for breakfast, followed by the same standardized low-choline lunch, dinner, and snacks as consumed the previous day. Each egg dose delivered 1.14 mmol (119 mg) total choline. Therefore, 0, 1, 2, 4, and 6 egg doses provided 0, 1.1, 2.3, 4.6, and 6.8 mmol (0, 119, 238, 476, and 714 mg) total choline, respectively, in addition to the standardized diet. TMAO was measured in plasma collected from each volunteer immediately before and 1, 2, 6, 8, and 24 h after the egg dose was consumed by using liquid chromatography and electrospray ionization–isotope dilution mass spectrometry. Doses were separated by a 2–4-wk washout period. A: Area under plasma TMAO curves. Plasma TMAO data were used to calculate the total AUC for each dose for all subjects. Values for the different egg doses are represented by bars patterned as follows: 0 eggs, white; 1 egg, light gray; 2 eggs, diagonal hatch; 4 eggs, dark gray; and 6 eggs, black. B: Mean (±SE) area under plasma TMAO curves after egg doses. Data from A were used to calculate areas under plasma TMAO 24-h dose-response curves for all subjects (n = 6) for each dose. A repeated-measures regression model with the plasma TMAO AUC as the response variable and the egg dose as the predictor was used. Variable estimates (±SEs) resulting from the model were used to make pairwise comparisons for various dietary choline intakes (egg dosages). aP < 0.05, different from 0 eggs; bP < 0.05, different from 1 egg; cP < 0.05, different from 2 eggs. TMAO, trimethylamine-N-oxide.

FIGURE 3.

Amount of TMAO in 24-h urine collections after egg doses. Six healthy volunteers (subjects) consumed a standardized low-choline diet on the day before each of 5 randomly assigned doses of 0, 1, 2, 4, or 6 egg yolks. Subjects were fed the egg dose for breakfast, followed by the same standardized low-choline lunch, dinner, and snacks as consumed the previous day. Each egg dose delivered 1.14 mmol (119 mg) total choline. Therefore, 0, 1, 2, 4, and 6 egg doses provided 0, 1.1, 2.3, 4.6, and 6.8 mmol (0, 119, 238, 476, and 714 mg) total choline, respectively, in addition to the standardized diet. A: Amount of TMAO in 24-h urine collections after egg doses. Urine was collected for 24 h on the day of each egg dose, and the concentration of TMAO was measured by using liquid chromatography and electrospray ionization–isotope dilution mass spectrometry as described in Subjects and Methods. The amount of TMAO was calculated from the concentration and volume of urine collected in 24 h for each dose for each subject. Some samples were not included in the figure because the amount of creatinine excreted per 24 h fell below 75% of the mean amount in all 24-h urine collections for that subject, suggesting that the individual sample was not a complete 24-h collection. TMAO values for the different egg doses are represented by bars patterned as follows: 0 eggs, white; 1 egg, light gray; 2 eggs, diagonal hatch; 4 eggs, dark gray; and 6 eggs, black. B: Mean (±SE) amount of TMAO excreted into urine in 24 h. Data from A were used to calculate the amount of TMAO in 24-h urine collections for all subjects (n = 6) for each dose. A repeated-measures regression model with urine TMAO as the response variable and the egg dose as the predictor was used. Variable estimates (±SE) resulting from the model were used to make pairwise comparisons for the various dietary choline intakes (egg dosages). aP < 0.05, different from 0 eggs; bP < 0.05, different from 1 egg; cP < 0.05, different from 2 eggs. TMAO, trimethylamine-N-oxide.

FIGURE 4.

Mean (±SD) differences (after dose minus before dose) in plasma oxidized LDL concentrations. Six healthy volunteers (subjects) consumed a standardized low-choline diet on the day before each of 5 randomly assigned doses of 0, 1, 2, 4, or 6 egg yolks. Subjects consumed the egg dose for breakfast, followed by the same standardized low-choline lunch, dinner, and snacks as consumed the previous day. Each egg dose delivered 1.14 mmol (119 mg) total choline. Therefore, 0, 1, 2, 4, and 6 egg doses provided 0, 1.1, 2.3, 4.6, and 6.8 mmol (0, 119, 238, 476, and 714 mg) total choline, respectively, in addition to the standardized diet. Oxidized LDL was measured in plasma collected immediately before and 24 h after each egg dose from each subject, and differences were calculated. According to paired t tests, there was no difference between predose and postdose values.

FIGURE 5.

Mean (±SD) differences (after dose minus before dose) in serum high-sensitivity C-reactive protein concentrations. Six healthy volunteers (subjects) consumed a standardized low-choline diet on the day before each of 5 randomly assigned doses of 0, 1, 2, 4, or 6 egg yolks. Subjects consumed the egg dose for breakfast, followed by the same standardized low-choline lunch, dinner, and snacks as consumed the previous day. Each egg dose delivered 1.14 mmol (119 mg) total choline. Therefore, 0, 1, 2, 4, and 6 egg doses provided 0, 1.1, 2.3, 4.6, and 6.8 mmol (0, 119, 238, 476, and 712 mg) total choline, respectively, in addition to the standardized diet. High-sensitivity C-reactive protein was measured in serum collected immediately before and 24 h after each egg dose from each subject, and differences were calculated. According to paired t tests, there was no difference between predose and postdose values.