*4.3. Statistical Analysis*

Principal component analysis (PCA) was conducted using SampleStat software (Human Metabolome Technologies, Japan). Hierarchical cluster analysis (HCA) was performed and heatmap formation analyzed using PeakStat software (Human Metabolome Technologies). Differences between the relative quantity of metabolites in Wagyu and Holstein samples were evaluated using Welch's t-test. Results are presented as means, and S.D. values of *p* < 0.05 were considered significant.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2218-1989/10/3/95/s1, Table S1: Metabolites of Wagyu and Holstein Beef.

**Author Contributions:** Conceptualization, T.Y. and M.H.; methodology, T.Y.; software, T.Y.; validation, T.Y., M.K., and M.H.; formal analysis, T.Y.; investigation, T.Y., M.K., and M.H.; resources, T.Y., M.K., and M.H.; data curation, T.Y., M.K., and M.H.; writing—original draft preparation, T.Y.; writing—review and editing, T.Y., M.K., and M.H.; visualization, T.Y.; supervision, T.Y.; project administration, T.Y.; funding acquisition, T.Y. All authors have read and agreed to the published version of the manuscript.

**Funding:** This work was supported in part by a Grant-in-Aid for Ito Memorial Foundation and JSPS KAKENHI (18K05958).

**Acknowledgments:** We would like to thank Kazuki Iguchi (Human Metabolome Technologies) for technical support.

**Conflicts of Interest:** The authors declare no conflict of interest.
