I am analyzing the biodiesel using reverse-phase HPLC. Methyl Oleate and Methyl Palmitate are overlapping in a single peak. How can I quantify the individual components from the same peak?
Hello Venkatesh,
do you have the possibility of coupling with MS? If so, then you can try to quantify the two compounds using SIM (even if they are not completely chromatographically resolved). Personally, as an analyst I have my problems with this - i.e. I only quantify what shows a good chromatographic resolution, but sometimes you have to use such solutions (but they should not remain the rule, but rather the exception).
Regards
Markus
Which mobile phase are you using? Out of Methyl Oleate and Methyl Palmitate, the one having a longer C-chain length will be more hydrophobic, so if you manipulate the methanol (mobile phase) concentration, the extraction power of mobile phase can be increased or decreased that may better resolve Methyl Oleate and Methyl Palmitate!
Thank you, Markus, for your answer. We do not have MS. We have FLR and UV detector. Is there any other mathematical way to solve it?@
Thank you, Shamsher, for your response. I am using Acetonitrile, acetone and water as the mobile phase. I tried with the methanol, n-Hexane, isopropanol also. But whatever I do changes to the mobile phase, the retention times of MeO and MeP are coming exactly the same.
Hi Venkatesh,
the only reasonable way now is changing the selectivity of your method (changing solvents and/or stationary phase of your column, changing your gradient, temperature etc.).
Good luck.
Markus
Yes sir, I am using XBridge C18 column, 3.5 micrometers
Column dimensions: 4.6 mm * 50mm
Alternative is to us C8 column which shall have a moderate binding capacity towards the solutes required to be separated and the chromatogram may give two distinct peaks.
Try C8 column for better resolution subsequently change the Ph of your mobile phase from acidic to basic use ammonium acetate buffer, ammonia buffer or formic acid buffer. also look in to your gradient prgrame too.
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