Liquid chromatography chromatograph column regeneration method - Database & Sql Blog Articles

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Liquid chromatography has a wide range of applications in the biomedical industry due to its high precision for low molecular weight polymer compounds. Long-term use of polymeric columns used to separate biomolecules can be contaminated, resulting in lower efficiency and affecting the rate and sensitivity of detection. Therefore, it must be cleaned regularly.
The chemical stability of polymeric materials is usually measured in terms of force. In general, it is mainly washed with nitric acid or sodium hydroxide solution. Some reverse-phase polymerization column cleaning methods such as polystyrene-divinylbenzene beads and polymerized monomers, such as CIM RP-SDVB tablets and Swift columns, can withstand a wide pH range (usually pH 11 to 13 or sometimes pH 0 ~ 14), but users should pay attention when washing the column with strong organic solvent, which is determined by the degree of cross-linking. When the column is washed with some organic solvents, it will expand or contract. Polymers having a degree of crosslinking higher than 8 to 10% generally have good mechanical properties and shrink little in an aqueous solution, and are not greatly expanded in an organic solvent.
The method of regenerating polystyrene-divinylbenzene monolithic column is as follows: flushing 10 column volumes at half the working flow rate with 0.1% tribasic 2-propanol; 100% mobile phase B is washed at half the working flow rate 5 column volumes or more; balance at least 10 volumes at 100% mobile phase A at the working flow rate.
A cleaning method comprising a mono- and butyl methacrylate monolithic column, washing the protein residue on the liquid chromatography silica gel-bonded reverse phase column by back-washing 10 column volumes of sodium hydroxide, water, Remove with 20% ethanol solution and working buffer. If there are many hydrophobic proteins, the user should insert a 30% isopropanol or 70% ethanol step after water.
If microbial colonies are to be washed or inactivated, the polystyrene-divinylbenzene can be completely washed with 0.5 to 1.0 M sodium hydroxide. The monolithic column is washed with sodium hydroxide for at least one hour at room temperature.
The use of a column packed with a conventional polymeric matrix to separate some of the less soluble proteins such as membrane proteins, structural proteins, and viral coat proteins requires strong cleaning conditions. For example, a 50% isopropanol solution containing 3 M guanidine hydrochloride can remove these proteins at 60 °C.
Removal of synthetic peptides in solid phase resins can produce positive ions that are reactivated by anisole and thioanisole. The removal of the positive ion reaction produces large aromatic molecules that contaminate the column as the peptide is purified. This contaminant has a very high retention on the C18 column and cannot be washed out with 100% methanol or acetonitrile. To clean such columns, the column must be reversed with 5 volumes of 100% isopropanol, three to five volumes of dichloromethane, then three to five volumes of isopropanol, and finally to the initial solvent system. Elution of aromatic impurities can be detected by 260 nm UV.