I would like to develop a method for analysis of many organic acids in the same time. how to separate the chromatographic pics of each one?
High performance liquid chromatography (HPLC) is used for analyzing organic acids because of the simplicity, speed and stability using the C18 column, as a common HPLC column, and phosphate solution as a mobile phase.
I understand that your aim is to develop an analytical method for the analysis of a mixture of organic acids.If this is correct, my answer would be the following.
Normally an acid is converted into its methyl/ethyl ester and then subjected to gas-chromatographic analysis (GC). This is done (a) to convert the acid into a relatively more volatile derivative and (b) to avoid a typical broad peak and dragging in the chromatograph observed in case of organic acids injected directly.
Your acid mixture needs to be converted quantitatively into the corresponding mixture of methyl /ethyl esters. This mixture can then be subjected to GC analysis using a column that separates the components based on the esters' individual boiling points. You may try a neutral column for this purpose.
With best wishes,
A methanol-water gradient should work, but both solvents must have 0.01% TFA in them. If TFA if difficult to get, you may also add 0.01% of acetic acid or formic acid in both solvents.
If your aim is to see the separated peaks and calculate yields based on the concentration, then you can use the Biorad Aminex 87H column for this purpose, with RI detector. It is packed with sulphonated divinyl benzene-styrene copolymer and helps in analyzing sugars and organic acids.
In case you wish to collect the separated acids, you will have to get details on preparative HPLC.
Choose mobile phase A as 0.01%Formic acid or TFA or ammonium acetate or ammonium formate or ammonium bicarbonate etc and then mobile phase B as either acetonitrile or methanol and use column which u want...
With different gradient and flow rate
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