LC-MS/MS assay is a chemical process with a high degree of application in drug development. It is particularly helpful in biomarker, drug and metabolite analysis.
Liquid Chromatography with tandem Mass Spectrometry (LC-MS/MS) is a technique of analytical chemistry. It combines the physical-separating capacities of liquid chromatography (LC) with the sensitive and selective mass analysis capacity of mass spectrometry (MS).
Both the LC-MS assay and the LC-MS/MS test gets applied at different stages of drug development from the first stage of drug discovery to the final stage of clinical trials. These are highly effective methods as the synergy between the two methods enhances their capacities.
Combining chromatography with mass spectrometry dates back to the 1950s. A.T. James and A.J.P. Martin first combined gas chromatography with mass spectrometry in 1952. But the liquid chromatography-mass spectrometry combination started much later in the 1970s.
High Performance Liquid Chromatography (HPLC)
Liquid chromatography (LC) is a process in analytical chemistry used to separate the individual components of a sample mixture. It involves the transfer of the sample mixture through a polar mobile stage and a non-polar stationary stage.
High performance or high pressure liquid chromatography (HPLC) is an advanced variation of LC which has a wide range of applications. A considerable body of published literature is available on HPLC.
The force of gravity makes the sample solvent travel in traditional LC. In HPLC, high pressure induced by a pump makes the solvent travel.
HPLC is a highly popular method in pharmaceutical R&D. The technique is applied at every stage of drug development.
Mass Spectrometry (MS)
Mass spectrometry (MS) is an analytical technique used to measure the mass-to-charge ratio of ions. Biochemical applications of MS form an integral part of pharmaceutical research and development.
MS has emerged as one of the most widely used techniques in drug development covering both qualitative and quantitative aspects.
Tandem mass spectrometry (MS/MS) breaks down precursor ions into fragments known as product ions. Once broken, the product ions expose the chemical properties of the precursor ions.
LC-MS and LC-MS/MS Analyses
A hybrid method of liquid chromatography- mass spectrometry assay (LC-MS assay) and liquid-chromatography-tandem mass spectrometry analysis (LC-MS/MS analysis) is applied in drug R&D as both these tests can directly analyze highly complex protein mixtures after proteolysis.
Over the last couple of decades, LC-MS assay and LC-MS/MS test have emerged as two of the most efficient methods in the pharmaceutical industry. Their application starts right from the drug discovery stage. But their most popular use is during the clinical trial stage.
LC-MS/MS assay plays a crucial role in the metabolite quantitation analysis of biological fluids like blood, plasma, serum, urine and other matrices. This is a vital procedure in the analysis of drugs, metabolites and biomarkers.
How Does LC-MS/MS Assay Work?
Once a solution with analytes to be studied is separated into its components through HPLC, the effluent gets directed to the mass spectrometer. There it gets nebulized, desolvated and ionized to transform into charged particles.
The next stage of LC-MS/MS assay involves the use of several mass analyzers known as quadrupole. A selected precursor ion gets passed through the first quadrupole.
In the second quadrupole the precursor ion gets fragmented into product ions through collision with an inert gas. The third quadrupole targets particular product ions.
Compounds of interest get quantified in the process with a high degree of selectivity and sensitivity based on their mass/charge transitions.