NMR metabolite quantification of a synthetic urine sample: an inter-laboratory comparison of processing workflows

Abstract

Introduction Absolute quantification of individual metabolites in complex biological samples is crucial in targeted metabolomic profiling. Objectives An inter-laboratory test was performed to evaluate the impact of the NMR software, peak-area determination method (integration vs. deconvolution) and operator on quantification trueness and precision. Methods A synthetic urine containing 32 compounds was prepared. One site prepared the urine and calibration samples, and performed NMR acquisition. NMR spectra were acquired with two pulse sequences including water suppression used in routine analyses. The pre-processed spectra were sent to the other sites where each operator quantified the metabolites using internal referencing or external calibration, and his/her favourite in-house, open-access or commercial NMR tool. Results For 1D NMR measurements with solvent presaturation during the recovery delay (zgpr), 20 metabolites were successfully quantified by all processing strategies. Some metabolites could not be quantified by some methods. For internal referencing with TSP, only one half of the metabolites were quantified with a trueness below 5%. With peak integration and external calibration, about 90% of the metabolites were quantified with a trueness below 5%. The NMRProcFlow integration module allowed the quantification of several additional metabolites. The number of quantified metabolites and quantification trueness improved for some metabolites with deconvolution tools. Trueness and precision were not significantly different between zgpr- and NOESYpr-based spectra for about 70% of the variables. Conclusion External calibration performed better than TSP internal referencing. Inter-laboratory tests are useful when choosing to better rationalize the choice of quantification tools for NMR-based metabolomic profiling and confirm the value of spectra deconvolution tools.

Optimization of heteronuclear ultrafast 2D NMR for the study of complex mixtures hyperpolarized by dynamic nuclear polarization

“…5), the potential chemical exchange with its creatine form at lower concentration prevents the linear response of creatinine, especially in D 2 O, an effect which has also been described in conventional 1D NMR. 61 Table 1 also shows the LOQ estimated using the linear regression equations, considering that a 2D SNR of 10 is the reasonable limit for a signal to be properly quantified.…”

“…Analytical Methods which has also been described in conventional 1D NMR. 61 Table 1 also shows the LOQ estimated using the linear regression equations, considering that a 2D SNR of 10 is the reasonable limit for a signal to be properly quantied. The slope values are heterogenous among the different tted metabolite 2D peaks (as discussed in Section 3.3.3).…”

NRX-101 (D-Cycloserine + Lurasidone), a Qualified Infectious Disease Product, is Active Against Drug-Resistant Urinary PathogensIn Vitro

“…3 However, we suggest that alanine-free media provides an estimate of MIC of DCS under ideal conditions, but may not reflect the MIC under real world conditions. Both Mueller Hinton broth, derived from beef extract, and artificial urine medium used in our study contain alanine 16 as does human urine. 17,18…”

NRX-101 (D-Cycloserine + Lurasidone) Is Active against Drug-Resistant Urinary Pathogens In Vitro

“…However, we suggest that alanine-free media provides an estimate of the MIC of DCS under ideal conditions, but may not reflect the MIC under real-world conditions. Both Mueller-Hinton broth, derived from beef extract, and the artificial urine medium used in our study contain alanine [19], as does human urine [20].…”