Chromatography Forum

I am developing an LC-MS/MS amino acid method using an Agilent HILIC-Z column. The standard procedure (e.g. AOAC methods) for sample prep, after acid hydrolysis, is to rotovap down and reconstitute, to avoid injecting HCl into the stainless steel LC system. This would create a significant manpower / time burden due to the difficulty of rotovapping aqueous solutions.

I am wondering if it is possible to simply neutralize the HCl with ammonium hydroxide and rely on the MS diverter valve to send the chloride salts to waste. The initial pre-equilibration period at the beginning of each LC run is enough for about 10 column volumes to go to waste (after the injection but before the diverter valve switches). However, I am concerned that with the HILIC phase these ions may still be making their way through the column, and will then end up in the MS. I also worry about the potential impact on retention times, peak shapes, etc. I would verify that salt is soluble at this level, but it also seems like a potential concern. I am still determining exactly what dilution level will best bring my amino acids in range, but I would expect (order of magnitude estimate) between ~2-20 mM ammonium chloride.

Is it a bad idea to treat these samples the ways I'm envisioning? Is there a way to handle a high sample load that doesn't take, like, an hour per sample in the rotovap?

Sounds like you're concerned about the salt load and I am assuming that it would be high enough to be a concern for the detection. Depending on the column and the method, I would be less concerned about the salts sticking around on the column, but without knowing too much about the method plan I can't make a strong case.

For the sample prep, you might want to look into a green extraction technique like dispersive liquid-liquid extraction. That might help avoid long roto-vap times and desalt the sample prior to analysis.

Are you using the mobile phase conditions in the Agilent HILIC-Z app note for amino acids? That uses a 20 mM ammonium formate mobile phase modifier. Since you're doing this by MS, I'm assuming you're not injecting a ridiculous volume. With those conditions, even with HILIC, I wouldn't be too concerned, and especially if you're using a diverter. I'd definitely give it a shot.