MadSci Network: Chemistry |
One of the most accurate methods of the determination of a carbohydrate is through the use of High Performance Liquid Chromatography. High Performance Liquid Chromatography, HPLC, is an efficient way to separate macromolecules, ionic species, natural products, polymers, and other high-molecular weight groups of molecules. HPLC achieves this by pumping a liquid mixture through a column at high pressure. An interactive liquid mobile phase carries the mixture through the column, which contains a stationary phase. Components of the mixture interact differently with both the mobile and stationary phases as they are being pushed through the column at high pressure. Chromatographic separation in HPLC is the result of specific interactions between sample molecules with both the stationary and mobile phases so, components of a mixture elute from the column at different times. Separated fractions are collected by placing an open vessel at the end of the column. Recovery is usually quantitative and separated sample components are readily isolated from the mobile-phase solvent. A typical chromatograph will provide the amount of sample (eluent) vs. the retention time. The area of a peak is proportional to the concentration of the component. Retention time can be used as a qualitative analysis for describing an analyte because analytes elute through at different times. Therefore you are able to detect qualatively the retention time of glucose by the use of a standard. And quantitativly by the measurement of the peak area.
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