The methods presented here can be scaled for longer or shorter analysis times. To facilitate this, the methods adhere to simple linear gradients as much as possible, with only slight deviations in the HILIC and lipid methods. This differs substantially from more highly tailored methods which, in our experience for metabolic profiling of complex biofluids, are less reproducible and reliable. However, when shortening methods, it must be noted that a reduction in the number of features detected and a reduction in the precision of their measurement has been well described. For this reason, shorter methods utilising the same UHPLC technologies may suffer a reduced ability to capture more subtle metabolic effects, although more rapid methods have been shown to be of use where the expected effect sizes are large and the analytes studied are dominant in the profiles. For best results, the throughput provided by a platform should be carefully matched to (not needlessly exceeding) the actual needs of the laboratory. Researchers concerned with data quality should not overlook the addition of LC-MS instruments as a viable, albeit more costly, alternative to reliance on higher throughput methodology alone when looking to increase overall analysis capacity. Finally, the methods presented here can also be adapted for larger scale separations (e.g., using HPLC columns) to allow isolation of metabolites from biological samples in support of MetID efforts. We have achieved this using repeated HPLC-scale separations of urine samples without difficulty using a rescaled version of our RPC assay for small molecules. Beyond global profiling applications, the methods presented here can also be adapted for use as targeted assays. For example, when developing an assay for the direct analysis of bile acids without laborious sample preparation , the lipid RPC method was used as a foundation owing to its ability to cleanly elute very hydrophobic lipid species in blood, bile and other sample types of interest. Doing so precluded the need to separate lipids in the sample preparation step, streamlining the whole analytical procedure. Similarly, when developing a small molecule assay for tryptophan catabolism pathway analysis, the established lower hydrophobicity RPC profiling method (column chemistry and dimensions, column temperature, solvent system and initial conditions) was adapted, substantially accelerating the development of the targeted method.