The gut microbiome radically influences immune development, homeostasis, and stress tolerance, making it a critical contributor to human health. Antibiotics, probiotics, and dietary changes can dramatically affect the microbiome composition. Such changes can result in negative or positive health outcomes, partially through inducing changes in the metabolism of the host organism. These complex interactions are only partially understood so far. The overall objective of this study is to detect metabolic changes caused by antibiotics, high protein diets, and the New Generation Probiotics (Wang et al., patent pending) using Nuclear Magnetic Resonance (NMR) metabolomics. We used mice as a model to observe metabolomic differences between groups within serum, liver, and brain tissue. To detect metabolic differences, we optimized a nontargeted metabolomics NMR method. In order to increase the NMR sensitivity sample preparation methods and data acquisition parameters were carefully optimized. 1D 1H and 2D 1H-13C HSQC spectra were recorded for all samples. Univariate analysis was performed to identify signals contributing to group differences. Results show strong metabolic differences between diets and after antibiotic or probiotics administration both in the tissue samples and the serum. As we discover more about the phenomena occurring within the gut after antibiotic administration and factors contributing to a loss of homeostasis, we can more accurately define the mechanisms of interaction between the host, commensal microbes and pathogens to make better decisions for our health when it comes to standards of practice, and choice of diet.