I am doing lipid research from extracellular vesicles, and in near future I am going to use Mass-Spec. I am still planning my experiment. I read some literatures but I found it is little confused for me to figure out which solvent should I use to dissolve my lipid samples. Generally, I after I extracted lipids from my samples using mixture of chloroform and methanol, I will need to evaporate the organic solvent through speedvac/nitrogen evaporator to prevent the lipid from being oxidized. And if my column is Thermo Accucore C30 column, my mobile phase is A:(60% acetonitrile: 40% H2O with 10 mM ammonium formate and 0.1% formic acid) and B: (90% isopropanol: 10% acetonitrile with 10 mM ammonium formate and 0.1% formic acid). Is there any suggestions for me to try about the solvent to dissolve the lipid in before inject into Mass-Spec? Thank you ahead!
ethanol and dimethylsulfoxide (DMSO). ...
Some lipids may require heat and sonication to dissolve in ethanol/water.
Acetonitrile/water (50/50) and MeOH/water (50:50) with 1% formate or acetic acid are typical solvent systems for positive ion ESI MS. 10 mM salts (e.g., NH4 formate) shouldn't create too much problem, but you will see some NH4+ adducts along. Isopropanol/water may also work, but you will likely need to increase the nozzle temperatures to get complete solvent evaporation. You should be able to use your existing mobile phases. ACN/H2O and MeOH/H2O also work in negative ion mode. I've had good luck with 1% phenol as the proton sink in negative ion mode but this is far from traditional, most people still use acetate.
Oxidation and temperature degradation are always issues for the polyunsaturated fatty acids, so its best to run things as cold as possible and maybe with an N2 sweep to displace the air in the ionization zone.
Dear Yiwen,
Degradation of your sample by lipid oxidation is something to (always!) bear in mind when analysing lipids as this (unwanted) reaction is likely to occur during lipid extraction, storage, and analysis. The best option is to minimise it... I normally add BHT (50ug/mL) to MeOH used in the extraction.
Regarding the solvent to dissolve your extract before LC-MS injection you should remember that, as you say, you used MeOH and CHCl3 for the extraction so the lipids in your extract are soluble in those solvents. So, I would just dissolve it in a MeOH (which has similar polarity to AcN). Lipids by its characteristics tend to be apolar and thus not soluble in water
Hope this helps.
Ana
Try to dissolve your sample with mobile phase A for best chromatography. If it is not fully soluble (%60 ACN) then modify mobile phase A composition with the addition of MeOH a bit to increase solubility. Perform this advice only for solubilization reagent composition, not for chromatographic buffer A.
You can dissolve your analyte in (3:1) Chloroform/Methanol or (1:1)Methyl-tert butyl ether(MTBE)/water, these are mostly used solvents for lipid extraction for Mass spec analysis, you can keep the sample with solvent in a water sonication bath for couple of min & incubate @30°C for better result.
Hi
The calculation and processing protocols are well explained in the following paper:
https://doi.org/10.1007/s12161-021-02203-0
I hope this helps!
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