How the HPLC colum and ion chromatgraphic colum is re genrate

You regenerate HPLC columns by flushing  it with the following volumes of solvent.

• 20 column volumes of water/acetonitrile (95:5 v/v)
• 20 column volumes of acetonitrile
• 5 column volumes isopropanol
• 20 column volumes hexane
• 5 column volumes isopropanol
• 20 column volumes acetonitrile
• 20 column volumes of water/acetonitrile (95:5 v/v)
• Re-equilibrate with the required mobile phase                                                                                                     Ion chromatographic columns are regenerated by flushing with electrolytes such as acids and bases                                                    

by multiple washing with mobile phase they can be regenerate

HPLC columns is regenerate by washing washing with purified water and preserve with methanol or Acetonytrle 50% depend open the type of mobile phase we are using

To clean and regenerate a reversed phase column flush it with the following volumes of solvent.

•  water/acetonitrile (95:5 v/v)
•  acetonitrile
•  isopropanol
• hexane
•  isopropanol
•  acetonitrile
• water/acetonitrile (95:5 v/v)
• Re-equilibrate with the required mobile phase

BY PASSING 1 M NACL SOLUTION THROUGH CHROMATOGRAPHY COLUMN.

For HPLC Reverse phase;

wash with unbuffered mobile phase

wash with100% Water

wash with Methaol or ACN

wash with THF or IPA

wash with Methylene chloride

wash with N-Heptane

wash with Methylene chloride

wash with Methanol or ACN

Wash with Water 

Return to Solvent

Time:10 times of column volume.

Column volume:   Πr2h

Example: column dimension250*4.6*5

h=250mm=25cm

r=4.6/2 mm=0.23cm

Π=3.14

Volume=3.14*0.23*0.23*25 =4.1527cm3=4.1527 ml

Flow=1.0ml/min

Time: (10*Vol)/flow =10*4.1527/1.0=41.52 minutes 

42 minutes enough for each washing.

regeneration done by organic solvent like acetonitril

For Ion Chromatographic columns, you can regenerate your column by one the following methods:-

1. Hydrophilic contamination of low valence: Concentrated eluent of10 times the concentration of the most concentrated eluent used in the application.

2. High valence hydrophilic  ions contamination & Metal Contamination:1 to3 M HCl.......if Metal Contamination still there try with0.2 Molar Oxalic acid.

3. Nonionic and hydrophobic contamination:80% Acetonitrile solution will be used. 

For HPLC columns; Flush it with water in the reverse direction at a flow of1ml/min. for about30 minutes, then flow Isopropanol, Methylene Chloride, Hexane, Methylene Chloride, then Isopropanol at this exact order each for30 minutes at a flow of1 ml/min.

Then flow the mobile phase you're working with at the normal direction of the column but without the buffer for30 minutes then introduce the buffer.

After this, performance should be restored for the column.