In Shimadzu HPLC-UVD, binary gradient mode with two pumps, I want to use three mobile phase (ACN:MeOH:Water, 40:30:30; v/v/v). How can it be done? If two of them are mixed in one bottle for one pump, is it the same after spraying by pump?
Can anyone have such experience earlier?
Hi Aniruddha,
in the attached article an isocratic solvent mixture is used, i.e. the solvents are mixed before and then pumped with one channel of the pump. So you don't need a gradient. When mixing the solvents you have two possibilities. 1. measure each solvent separately with a measuring cylinder and then mix. 2. mix the solvents together in the appropriate ratio. I would use method 1, because a volume contraction occurs when mixing methanol and water.
Regards
Markus
From what you write, you are doing an isocratic elution (you are not using three mobile phases but one mobile phase composed of three liquids): just mix all the solvents first, then use a single pump.
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