Determination of Reference Interval of Serum Trimethylamine-N-Oxide In Humans By LC-MS/MS Method: Retrospective Study

Aim: The aim of our study is to determine the reference range of serum TMAO levels in individuals aged 18-65 using the LC-MS/MS method. Methods: TMAO levels in serum samples from 185 healthy individuals aged 18-65 were determined using the LC-MS/MS method. Statistical analysis classified the data by gender and age. The lower and upper limits with a 90% confidence interval were estimated using the robust method. Outliers were identified and excluded from the log-transformed data using the Tukey criterion. Skewness and kurtosis estimates were based on log-transformed data. Results: TMAO levels were similar between men and women, with median values of 180 ng/mL and 200 ng/mL, respectively (Z = –0.476, p=0.633). Additionally, no significant difference was observed in TMAO levels between women under 30 years of age and those aged 30 and above (median: 178 ng/mL vs. 186 ng/mL; Z = –0.739, p = 0.460). However, men aged 30 and above had significantly higher TMAO levels compared to men under 30 (median: 215 ng/mL vs. 191 ng/mL; Z = –2.225, p = 0.026). The reference interval for serum TMAO was determined as follows: 27–974 ng/ mL in men under 30 years, 52–942 ng/mL in men aged 30 and above, 45–655 ng/mL in women under 30 years, and 33–539 ng/mL in women aged 30 and above. Conclusions: This study determined age-and gender-specific reference intervals for serum TMAO levels using the LC-MS/MS method. To our knowledge, this is the first study to establish such reference intervals using this analytical technique. The findings are expected to provide valuable data for future clinical and epidemiological studies involving TMAO as a biomarker.