High end Liquid Chromatography (HPLC) is an deductive tool that separates, identifies plus quantifies components in a sample. This is a commonly used system in analytical chemistry and biochemistry fields. Basically, the system carries the sample using a solvent or mixture of solvents to the stationary phase, where separation of substances occurs. A detector captures the particular separated compounds and signals are sent to the integrator to generate the graphic visual.
HPLC consists of the components below:
᾿ Mobile Phase – this is the solvent or usually a combination of solvents used to transport the examples through the whole system. The solvents have to be miscible in the mixture; else the immiscible solvents will cause stress build-up in the HPLC system. The ratios of each solvent component within the mobile phase affect the separation associated with compounds as well as analysis length.
᾿ Pump or solvent delivery device – this component is to deliver the mobile phase and samples throughout the system at a constant movement rate or pressure. Usually, for analytical purposes, HPLC pump is placed to operate at constant flow price.
᾿ Injector Port or auto sampler – analytical samples are usually introduced through this component. Samples introduced through injector port need to be manually injected using an appropriate HPLC syringes. Auto sampler enables a good analyst to load all the samples to the HPLC system and the system will automatically select the correct sample to inject at preset conditions.
᾿ Stationary phase – also known as column. This part of the system is actually the very center of separation. It is made of firmly packed material in a stainless steel column.
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Due to the compactness of the packed material, high pressure are required to pump or provide solvents throughout the system, hence HPLC sometimes are term as High Pressure Liquid Chromatography. As the samples stream through the column, the compounds in the sample will interact simultaneously using the stationary and mobile phase within a different manner to yield different elution time of each compound. The objective of each analysis is to separate the peak of interest from other present substances.
᾿ Detector – this unit detects the separated compounds in the sample. There are various detectors using different mode of detection such as ultra-violet, fluorescence, mass spectroscopy and refractive index.
᾿ Integrator – integrator turns the signals conveyed from your detector into visual output called chromatograms. Nowadays integrators come in the shape of computer systems instead of the conventional types which use paper charts.