One of the most
popular compound separation techniques used across different bioanalytical labs
is that of liquid chromatography. In this technique, you have an interaction
between individual sample molecules and the stationary and mobile phases of the
Liquid Chromatography chamber. The liquid chromatography process is either
performed on a sheet or within a chamber with solid support and a supporting
liquid mobile phase.
The liquid mobile
phase starts flowing down the solid stationary phase. This allows the mobile
phase to carry all the sample components together. In the liquid chromatography
unit, the interaction between your sample molecules and the chromatographic
medium depends on factors such as charge, affinity binding, size, and
hydrophobicity.
An advanced version of
the liquid chromatography technique utilizing high pressure for forcing the
sample through the entire column length is called high-performance liquid
chromatography. This is currently the most extensively employed quantitative
analysis procedure for pharmaceutical industries.
Mechanism of HPLC unit
–
HPLC or theHigh-Performance Liquid Chromatography is an advanced version of the Liquid
Chromatographic technique. This process is amenable to a wide range of
applications, such as food and pharmaceutical analysis. The main advantage of
employing the High-Performance Liquid Chromatography technique is that it is
easy to analyze the non-volatile compounds which are otherwise difficult to
handle with the gas chromatographic unit.
There are three types
of pumping systems involved here in HPLC –
Syringe Pumps –
Syringe pumps help in
providing a continuous pulseless flow rate. These pumps are relatively
reliable, precise, accurate, and exhibit the potential to hold a large
capacity.
Direct gas-pressure
system –
This is both an
inexpensive and reliable HPLC pumping system. However, the process involved in
assisting solvent change is a bit difficult.
Reciprocating pumps
–
Reciprocating pumps
are an economical solution specifically designed for providing a constant flow
and high pressure. However, sometimes, it may result in pulsing.
Mass Spectroscopy
Analysis –
Mass Spectrometry is a
process wherein the atoms or your molecules undergo the process of ionization
for facilitating their separation and detection. The separation and detection
of these atoms and molecules are accomplished in accordance with their charges
and masses.
What is LC MS?
When the Liquid
Chromatography's separation potential is combined with the detection ability of
your Mass Spectrometry, you get an LC MS unit. Combining the analytical methods
helps improve the accuracy of the quantification, reducing experimental errors
of the entire process.
Working Mechanism of
LC MS –
LC MS technique
separates the sample mixture in accordance with both their physical and
chemical properties. This is followed by individual component identification
within the dimensions of each peak and detecting them with the help of a mass
spectrum. The flow rates involved in your LC MS detection is always less than
what you have in your HPLC. The reason for low lc ms method detection is ensuring acomplete ionization processc ms method detection is ensuring a
complete ionization process, thereby further maintaining the detection
sensitivity of your MS detector beyond 200 microlitres per min. Thus, the LC MS
columns are much smaller compared to your HPLC columns, thereby facilitating
the accommodation of the lower flow rates of your sample and the sample
volumes.
