Same mass, different retention time. Isomers?

[mcg_2022]

Hi,

I have been working on the analysis of carotenoids for the last 2-3 years for my PhD using UPLC-UV-HRMS. I used an analytical standard of lycopene (extracted from tomato) to detect the presence of this compound in bacteria. During all my tests I consistently found that there are 3 peaks in my chromatogram matching the mass of lycopene. One peak matches the exact retention time of the standard (RT: ~20.3 min). The other two peaks have the mass of lycopene but elute at around 16.3 and 17.6 min respectively.

The person who is helping me with the analysis has told me that I can refer to these other two peaks as lycopene isomers. I was wondering with which certainty I can say that these other peaks are lycopene isomers. I haven't found a lot of information in the literature about other lycopene isomers retention times, there are actually so many isomers that is hard to find possible matches just based on RT. Also this compound has been extensively studied in food matrixes but not in bacteria. It is interesting because I see different proportion of these 3 isomers/peaks in my bacteria depending on the growth conditions.

I just want to be certain while writing my thesis that I am not making a mistake when I refer to these other peaks as "isomers". It is possible that these products are maybe some degradation product of lycopene? I guess in that case the mass should be different? I ran the samples on a UPLC-DAD to check the absorbance spectra of the different peaks. One of the isomers have almost identical peaks than the lycopene from the standard while the other one differers a bit more. Is this any indication of what type of isomer/difference in structure?

Any advice will be greatly appreciated. Thank you

[millerk8561]

Identical mass with differing retention times ... yes my gut feeling is some kind of isomer but to be sure you'd have to compare with analytical standards of any known lycopene isomers. Another thing would be to analyze that same sample with NMR to get an idea of the structures of those compounds.

Additionally, what are you using for sample prep/cleanup? If it's not isomers per se, what possible other products could have made it through your sample prep and cleanup process? These are questions to ask yourself.

[Gaetan Glauser]

Several peaks with same exact masses but different retention times are by definition isomers (unless some peaks would have fragmented in the MS). If you only have a lycopene standard available, then it is not wrong to refer to the other peaks as lycopene isomers. However, I would try to go a bit further and at least see if one of these two isomers could be bêta-carotene, which is commonly found not only in carrot but in most plant leaves for instance.