Full set of Notes with Fill-Ins - San Jose State University

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Chem 155 Unit 1 Page 54 of 3161.1.27 Internal Standards Example:In HPLC the sample is injected into a flowing stream, and the signalis a peak in a plot of absorbance versus time (a chromatogram).Often the volume of the injection will vary from injection to injection,so the peaks will vary in size!Peak 1 is theinternal standard.100 ppm NaNO 3 ,480 All mAu . sabsorbance10.5Peaks 2, 3 and 4 are otheringredients in the sample.1Peak 5 is theanalyte, a caffeinestandard, 25 ppm,530 mAu . sabsorbance100 ppm NaNO 3 ,520 mAu . sabsorbance00 400 800 1200 1600 2000 2400 2800 3200 3600 4000time / s10.500 400 800 1200 1600 2000 2400 2800 3200 3600 4000time / s10.523Chromatogram1 is a standard.Did the caffeineconcentrationincrease fromchromatogram1 to 2?Did the caffeineconcentrationincrease inchromatogram1 to 3?Unknowncaffeine conc.630 mAu . s00 400 800 1200 1600 2000 2400 2800 3200 3600 4000time / sPage 54 of 316
Chem 155 Unit 1 Page 55 of 3161.1.28 Internal Standards CalculationHow about calculating the actual concentration of the sample in 3above?StandardS'A − STDCA − STDS'IS − STDCIS− STD530 mAu . s25 ppm480 mAu . s100 ppmSCA−STDIS−STDk ==S'IS−STDCA−STDS'ANmm' 530⋅100ANIS480⋅25= 4.417 unitless630 mAu . sSampleS'ISCISCANS'S'ISCk520 mAu . s100 ppmAN IS=630⋅100520⋅4.417= 27.4 ppmPage 55 of 316
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Full set of Notes with Fill-Ins - San Jose State University

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