Chromatography Forum

I am intending to develop a new LC-MS/MS method from scratch for a basic compound. I'm intending to use 0.1% NH4OH as the aqueous mobile phase later down the line. However, firstly I need to tune the compound on the MS in ESI+ mode.

When I have done this previously, I've always used an acid (0.1% Formic acid in water) to carry my compound into the MS via syringe for tuning. These LC-MS methods have used acidic mobile phases going forward.

With my basic compound, it seems counterintuitive to tune with acid. So I think I should dissolve my compound in NH4OH and inject this solution into the MS for tuning.

Is this necessary? Will the compound still ionise if there's no acid? Would different fragments be produced if I tuned in an acidic solution instead?

My lab generally tunes (initially) by direct infusion at low flow using a mixture of mobile phases (including modifiers) that fairly closely mimics the mobile phase conditions at the time of elution. If we don't know the exact elution profile and elution time, we approximate or just use 50:50 mobile phases at the time of tuning.

Tuning at a low flow rate by direct syringe infusion is helpful for the "internal" settings of the mass spectrometer (optimized parent mass, daughter masses, collision energy, cone voltage, etc.) to enable observing a good signal.

Then,

The optimal ionization source settings (capillary voltages, temperatures, gas flows) really need to be at the actual flow conditions being used for the method (flow rate, mobile phase ratios and mobile phase modifiers).

My lab generally tunes (initially) by direct infusion at low flow using a mixture of mobile phases (including modifiers) that fairly closely mimics the mobile phase conditions at the time of elution. If we don't know the exact elution profile and elution time, we approximate or just use 50:50 mobile phases at the time of tuning.

Tuning at a low flow rate by direct syringe infusion is helpful for the "internal" settings of the mass spectrometer (optimized parent mass, daughter masses, collision energy, cone voltage, etc.) to enable observing a good signal.

Then,

The optimal ionization source settings (capillary voltages, temperatures, gas flows) really need to be at the actual flow conditions being used for the method (flow rate, mobile phase ratios and mobile phase modifiers).

This is how we do it also. Try to be as close to the actual mobile phase as possible to account for any adducts that could form or other effects.