Metabolomics

What is Metabolomics?

Metabolomics is a novel “-omics” strategy, defined as the global or local profiling of metabolites in a given biological system, with broad coverage of different classes of endogenous metabolites.

It is the “systematic study of the unique chemical fingerprints that specific cellular processes leave behind” or “Identification and quantification of the compounds (<1500 Da) in the metabolome”.

A metabolomics analysis provides a deeper understanding of metabolite expression within complex biological systems, and the proper analysis and interpretation of such results may potentially indicate the metabolite function.

At C-CAMP we have a well established Metabolomics Facility to help with identification and quantification  of known compounds/metabolites in biological fluids such as Sera, Saliva, Tissue, Urine as well as from Cell extracts using Targeted Metabolomics approach.

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1. Full Scan (MS)/Product ion scan (MS/MS) analysis of known or purified compounds (Solid/Liquid)

This can be used to identify the precursor ions that yield a product ion for a specific m/z ratio

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2. Analysis of compounds in biological matrix

(i)   Extraction of metabolites (liquid-liquid extraction, Solid phase extraction or both, depending on the nature of the metabolite)

(ii)  Analysis by Selected Reaction Monitoring (SRM) or multiple reaction monitoring (MRM)

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3. Method development for specific metabolites to determine the absolute  quantification

(i)   MS scan, MS/MS scan and picking the most intense ion for the MRM scan

(ii)  Spiking the STD and ISTD and extraction of metabolite from matrix (Sera, Saliva, Water, cell extract and Urine)

(iii) Detection of LOD and LOQ

(iv) Construction of STD curve

(v)  Validation of the method (Inter & Intraday variation, recovery, accuracy and precision)

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4. Derivatization methods

In case metabolites are not ionizing, we can try derivatization methods using different derivatizing  agents. This derivatization will improve the separation efficiency, ionization  efficiency and MS/MS detectability of the analytes. We currently offer the following derivatization methods:

(i)   Derivatization of ketone and aldehydes: Girard P and Girard T reagents

(ii)  Derivatization of alcohol and phenols: Dansylchloride, pyridine sulfonyl chloride

(iii) Derivatization of amines:  ACCQ reagents

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5. Analysis and purification using UHPLC system

Through our UHPLC system equipped with both diode array/fluorescence detectors and fraction collector, UV/ Fluorescence based analysis of metabolites, peptides or bulk purified biomolecules  can also be performed.

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6. Metabolite Profiling/Comparative Metabolomics

Using Q-exactive HRMS system we are setting up comparative metabolite profiling (Hydrophobic-reverse phase and Hydrophilic-HILIC) between control versus sample. The data analysis will be done using SIEVE 2.2 software using publically available data bases like HMDB and KEGG through Chemspider.

TSQ Vantage-Agilent UHPLC (LC-MS)

 Nexera Shimadzu UHPLC

UPLC Q-Exactive -Thermo MS