Do you have any suggestions, which SPE cartiges to use for markers of oxidation stress? I do a LC-MS analysis of quite various group of analytes, including bases (e.g. hydroxyguanosine), acids (e.g. ascorbic acid), aminoacids and peptides (e.g. tyrosine, glutathion), and also lipophilic substances (isoprostanes etc.). What first came to my mind was the HLB sorbent. But is there any other/better option? Or should I rather make two extraction protocols, for example one for the lipophilics and bases (maybe MCX?) and second for the acids and aminoacids?
I will be extracting those markers from serum or cell/tissue extracts, so I need to get rid of mainly proteins, phospholipides, anorganic ions.
Vijay Arjun Bagul , thanks for your answer, but I was asking mainly for the type of the sorbent suitable for those various analytes, not the brand.
Dear Jan Dehner,
The common sorbant is used are C18 & C8 then Cyno and other group like Phenyl. Have you tried C18 for the same?
Hello Jan, have you already thought about dispersive SPE? Above all, I'm thinking about phospholipid removal. There are cartridges from Phenomenex (Phree) and also from Waters, Agilent,... We have had good experience in detecting coccidiostats in muscles and eggs. The Phree cartridges are specifically for blood/plasma tests. You may then have to run an SPE (classic) with C18, StrataX, HLB,... to concentrate the sample again - if necessary. Greetings Dirk
You need to fully understand the property of your target analytes and your marrix interference in your sample that you want to keep or get rid of. In your case, your targets are low concentration of the combination of neutral, acid, and basic small molecules. Your matrix interference are high concentration of large molecules of protein/peptides/phospholpids in the serum. You have at least two options to eliminate the majority of proteins by using a) acetonitrile crash (mix 1 part of serum and 3 parts of acetonitrile. Protein molecules will precipitate out of the solution and your analyes will be in a solution of 1:3 water:acetonitrile. The second way is SPE from Phenomenex or Waters or Agilent that will capture protein/phospholipids. Please do the Google search on these products. . Once you get rid of protein/ phospholipids in the samples, you need to capture your analytes and eliminate small polar/ionic compounds to minimi the matrix suppression at the Ion-spray interface of your LCMS instrument. The mixed mode SPE (MCX or MAX) will be a good choice because one cartridge will be needed. Please study further from the Waters application note of this product on the internet. Essentially if you use MCX, it has affinity for neutral/acidic compounds via reversed-phase mode and retain basic analyte via the cation-exchange mode. You diute the sample with warer to lowere the concentration of organic solvent in the sample and lower the pH with acid. At this condition, the acid analyte will not ionize and retain on the stationary phase along with the neutral ones. The basic analytes will ionizeband has positive charge and stay at the stationary phase. The small polar neut molecule such as sugar and small metal ions will pass thru. you can wash the SPE with water and may be 10% methanol/ water. To elute all anayte, you will use methanol having high pH (1% ammonia).
hope this helps.
http://www.the-ltg.org/data/uploads/applicationnotes/development-of-dimple-dast-dpe-protocols-for-basic-analyte-extraction.pdf
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