Use only high-purity solvents and buffer salts for preparing mobile phases. Water should be type I reagent grade, with a resistivity of >15 megohms/cm2. Don't forget to thoroughly wash all glassware used to prepare and hold the mobile phase.
Filter all mobile phases through a 0.2µm membrane filter. The figure below illustrates the filtering device recommended by BASi for doing this. It is borosilicate glass throughout, and uses ground glass joints to avoid contamination from stoppers. The filtration step will reduce problems with column degradation significantly.
CLEAN, PARTICLE-FREE MOBILE PHASES ARE CRITICAL TO ANY LC METHOD!
MF-6126 Filtration Apparatus
Mobile phases should be degassed prior to use. Vacuum filtering may be sufficient to degas the mobile phase. But if bubbles are a problem in your system, a more-thorough degassing is required. Allowing the solution to sit under the vacuum generated by an aspirator for 5-15 minutes should be sufficient. The MF-6126 kit also can be used for degassing, by replacing the filtration apparatus on the top of the unit with a stopper. Small amounts of volatile organic modifiers may be lost, but not enough to cause any difficulty.
The easiest and most effective method of degassing is the LC-26 on-line degasser. This highly effective instrument reliably and consistently delivers bubble-free mobile phase to the PM-80 pump. We strongly recommend its use, particularly for gradient applications.
Whichever way you make your mobile phase, be consistent; consistency in mobile phase preparation is critical for reproducible chromatography.