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1 EXPERIMENT 9 HIGH PERFORMANCE LIQUID CHROMATOGRAPHY SEPARATION OF COMPONENTS OF SOFT DRINKS Spring ' 10 A. PURPOSE The principal purpose of this experiment is to introduce you to procedures and instrumentation used for high performance liquid chromatography (HPLC). B. OVERVIEW In this experiment you will separate mixtures of components frequently found in soft drinks, namely caffeine, benzoate, and aspartame. You will be given seven solutions containing known amounts of the three species and record the chromatograms. C. INSTRUMENTATION 1. Liquid chromatograph A commercially available liquid chromatograph (PM-80 pump and CC-5 Injector, Bioanalytical Systems, West Lafayette, IN) is used in this experiment where separation is performed on a reversed phase column (#MF 8954). Chromatograph and column settings are as follows: Flow rate: 0.5 mL/min Minimum pressure setting: 250 PSI Maximum pressure setting: 4000 PSI Column dimensions: 3(i.d.) × 100 mm Column has silica packing (3 μ m particle size) and is functionalized with ODS (C 18 ). pH range is 2-7. 2. Detector Detection is by absorption spectroscopy. Although the detector (Model UVVIS 200, Linear) is capable of measuring several wavelengths, all the components of interest absorb at 254 nm. Accordingly, only one wavelength, 254 nm, will be used in this study. Detector settings are as follows: Rise time: 0.3 s Wavelength: 254 nm Range: 0.2 AUFS (absorbance units full scale) 3. Data acquisition Data will be recorded on line using a digital voltmeter and a digital computer. The following settings should be used: Time per point: 0.1 s Time per chromatogram: 10 min initially 4. Saving data Data will be saved on floppy disks. Each group should bring at least two floppy disks with full storage capability free of other files
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This note was uploaded on 02/16/2012 for the course CHEM 125 taught by Professor Na during the Fall '11 term at Purdue University-West Lafayette.

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