Question

Briefly explain how you would perform a quantitative analysis of the

components of a mixture of ethanol, n-propanol, and iso-propanol using HPLC.

Begin with the preparation of standard solutions. Explain how you would use the

HPLC chromatogram to both identify the molecules and quantify the amount of each

alcohol in the mixture. You don’t have to provide volume or concentration numbers or

other technical experimental details. Provide a description of the general approach.

Be complete and concise.

Answer 1

A method has been developed on reverse phase high performance liquid chromatography for simultaneous determination of methyl, ethyl and isopropyl alcohols under refractive index detection using pure water as the mobile phase.

A good separation has been achieved between these alcohols. Detector response was linear with a detection limit of 5 mg/100 ml.

Sample Preparation-

Aqueous samples were prepared by diluting the volatiles, ethanol, acetone, isopropanol, methanol and n-propanol, with DI water and stored at 5°C.

Calibration-

Concentrations were 100, 500, 790, 1,580, 3,160 and 4,740 mg/kg for ethanol; 100, 190, 380, 760, 1,580 and 2,980 mg/kg for acetone; 100, 120, 380, 750, 1,570 and 3,020 mg/kg for isopropanol; 100, 130, 380, 760, 1,580 and 2,980 mg/kg for methanol and 110, 140, 400, 790, 1,580 and 3,010 mg/kg for n-propanol. A linear regression of the ratio of the peak area counts of the analyte and ISTD versus the ratio of the concentrations of the analyte and ISTD was used to construct the calibration curves for each analyte.

Linear aqueous calibration curves were generated from six-point calibration curves with calibrators prepared in duplicate in DI water. Concentrations of the analytes were the same as with the matrix prepared calibrators. The average slope (n = 10) of the aqueous calibration curves were compared to those of the matrix prepared calibration curves for determination of differences, if any