Identification and Quantification of Deranged Metabolites in Critically Ill Patients Using NMR-Based Metabolomics

Summary

Nuclear magnetic resonance (NMR) spectroscopy is used to identify dysregulation in the metabolites in patients with various diseases. This technique allows the quantification of the deranged metabolites, unraveling the pathophysiological insights. Here, we describe the step-by-step procedure of the NMR-based approach for the metabolic characterization of the patients.

Abstract

Metabolomics is emerging as a significant approach to reflect the individual's response to pathophysiological conditions. Nuclear magnetic resonance (NMR) spectroscopy has evolved as a tool to identify metabolic dysregulations in critically ill patients afflicted with conditions like acute respiratory distress syndrome (ARDS), severe acute pancreatitis (SAP), acute kidney injury (AKI), and sepsis. The spectral data from the serum sample of the study and control group are recorded using an 800 MHz NMR spectrometer and processed using NMR processing and analysis tools. Furthermore, a rigorous statistical analysis, such as univariate and multivariate tests, is performed to pinpoint significant metabolites, which are then accurately identified and quantified using NMR metabolite quantification software. Additionally, pathway analysis highlights the deranged biochemical cycles that result in the severity of illness. Through this comprehensive approach, researchers aim to gain deeper insights into the metabolic alterations associated with these critical illnesses, potentially paving the way for a better understanding of the disease and improved diagnostics and treatment strategies.

Introduction

Despite continuous efforts to provide efficient disease diagnosis worldwide, targeted therapy has still not achieved its true potential. Various approaches, such as transcriptomics, proteomics, etc., have resulted in the identification of several biomarkers, but these did not hold enough clinical utility because of the lack of sensitivity and specificity^1,2. Targeted therapy is a big challenge in some multifactorial diseases, eventually leading to higher mortality. There is a need for a better understanding of the underlying mechanism and pathophysiology of complex diseases existing with ....

Protocol

Ethical approval (IEC code: 2022-71-PhD-126) was obtained from the IEC of Sanjay Gandhi Postgraduate Institute of Medical Sciences (SGPGIMS), Lucknow. Written and informed consents were taken from the patients or their relatives to perform the study and publish the data for research purpose. Moreover, the research was conducted following the institutional guidelines.

1. Study design and ethical clearance

1. Determine the sample size for each group. Carefully review an.......

Discussion

Metabolomics efficiently identifies and quantifies metabolites, targeting the metabolic cycles that become deranged during disease. The quality of the results depends on the meticulous execution of each step in the metabolomics approach. Every stage, from sample selection and collection to pathway identification, is critical in accurately identifying the primary factors contributing to the disease. Before performing metabolomics, a thorough review of the literature is essential, and careful attention must be paid at ever.......

Materials

Name	Company	Catalog Number	Comments
Centirfuge	Sigma aldrich	3-18KS
Chenomx NMR suite	NMR Suite, v9, Chenomx Inc., Edmonton, Canada		NMR metabolite quantification software
Co-axial insert	Sigma aldrich	Z278513
Deuterim oxide	Sigma aldrich	151882
Eppendorf tubes	Tarsons	500020
Metaboanalyst	Wishart Research Group		Metabolomics statistical analysis software
NMR tube	Wilmad	Z412007	5mm diameter
Pipette	Eppendorf research plus	3123000039	0-100 μl
Sample collection vials	Tarsons cryo chill vials	523194
Sodium azide	Sigma aldrich	S2002
Sodium chloride crystal	Sigma aldrich	S9625
Sodium phosphate dibasic	Sigma aldrich	567550
Sodium phosphate monobasic	Sigma aldrich	S0751
Topspin 3.6.4	Bruker		NMR processing and analysis tool
Tsp salt	Sigma aldrich	269913