Top rated Liquid Chromatography (HPLC) is an deductive tool that separates, identifies plus quantifies components in a sample. It 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 compounds occurs. A detector captures the separated compounds and signals are usually sent to the integrator to generate a graphic visual.
HPLC consists of the constituents below:
᾿ Mobile Phase – this is the solvent or usually a mixture of solvents used to transport the samples through the whole system. The solvents have to be miscible in the mixture; otherwise the immiscible solvents will cause pressure build-up in the HPLC system. The particular ratios of each solvent component within the mobile phase affect the separation of compounds as well as analysis length.
᾿ Pump or solvent delivery device – this component is to provide the mobile phase and examples throughout the system at a constant circulation rate or pressure. Usually, to get analytical purposes, HPLC pump is set to operate at constant flow rate.
᾿ Injector Port or car sampler – analytical samples are usually introduced through this component.
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Samples introduced through injector port need to be manually injected using an appropriate HPLC syringes. Auto sampler enables an analyst to load all the samples in 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 center of separation. It is made of tightly packed material in a stainless steel line. Due to the compactness of the packed materials, high pressure are required to pump or deliver solvents throughout the system, hence HPLC sometimes are term as High Pressure Liquid Chromatography. As the samples flow through the column, the compounds within the sample will interact simultaneously using the stationary and mobile phase in a different manner to yield different elution time of each compound. The purpose of each analysis is to separate the peak of interest from other present compounds.
᾿ Detector – this device 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 form of computer systems instead of the conventional ones which use paper charts.