Hi,
I was wondering if anyone had advice on desalinating LC-MS Metabolomics samples. We are studying the composition of uterine fluid which was collected by flushing with 50 mL PBS. I extracted the samples using methanol/ chloroform and dried them down. However, when resuspending the sample in water I read the osmolality of the sample and it was 4700 mOsm which will block the column and cause excess back pressure.
One of the key components we are looking at though are amino acids and all the desalinating columns I have been able to find will remove the amino acids due to their small size.
Does anyone have any advice on how I could desalt the samples without removing the amino acids and metabolites?
Thanks so much
a possible way to desalt the samples is to use a SPE cartridge, that can also be useful to concentrate your analites. If you are using a HPLC-MS with a RP column you can simply dischcarge the initial part of your eluate by using the swithch valve of the MS.
As Ziyaulhaq Auwal Abdulkadir indicated, salt typically is not a problem in reverse phase HPLC, because it flushes out in the solvent front. You may need to dilute your sample and load it slowly, but probably it will be fine. Filtering your samples, or using SPE sample prep, which you should be always doing routinely anyway, should eliminate issues with back-pressure.
After taking some precautions and utilizing an ultra low-volume MS diverter valve in the flow-path, we routinely inject samples into our LC-MS system which may have some residual salts in the solution. *Ideally, samples should always be dissolved in the mobile phase, which for LC-MS analysis, should not contain any non-volatile additives such as PBS salts. In the case where residual salts or other undesirable materials remain in the injection solution, we develop the HPLC method to elute them at or near the column's void volume (Tzero) so we can redirect the column outlet flow directly to waste for a short period of time (often using a timed contact closure to the valve). Once all of the material (e.g. salts) has been safely eluted out of the column to waste, we switch the diverter valve back to the normal flow position to the detectors, restoring normal flow to the LC and MS detectors (in-line). Note: During the time in which the diverter valve is redirecting flow to waste, no liquid flow will be sent to the detectors. Lack of flow to the LC and esp. MS detector should be avoided. To maintain flow to the MS detector we use a secondary pump connected to the diverter valve's ports (an electronically operated, two-position, 6-port valve offers this option). The secondary pump is connected to the same mobile phase composition (same flow rate) as the main analysis uses. It automatically directs the liquid to the inline detectors when the diverter valve toggles to re-direct the main flow to waste. It then switches out of line when normal flow is restored (and the valve switches back). **This functionality is needed to prevent running the MS detector dry, resulting is HIGH noise and baseline disturbances (which may invalidate the analysis data and method). This is especially important if running in negative mode ESI (as an example) as source arcing may occur under conditions of low or no flow, damaging the needle.
For offline strategy as sample prep, use C18 in the spin column, spe, or stage tip format...As online desalting use trap columns and adjust the LC fluidics operating the valves to wash out the salts while retaining the peptides...These are very common approaches for proteomic desalting ...
Have you thought about precipitating the Sodium Chloride with 0.05N Silver Nitrate and filtering it. I have no idea what excess Silver ions will do to the column and MS detector!