A practical guide to structural analysis of carbohydrates

Removal of RNA using Ribonuclease A

RNA is a common contaminant in LPS preparations due to cell rupture and leakage of RNA. Ribose appearing in a hydrolysate (sugar analysis) is most often a sign of contaminating RNA. RNA may be removed by treatment with RNAse or alkali, both followed by purification by gel filtration or dialysis.


  • Ribonuclease A
  • 0.1 M CaCl2 or MgCl2
  • 0.1 M Sodium acetate buffer


  1. Dissolve the LPS in 0.1 M acetate buffer (pH 5) to 20 mg/mL (an opalescent solution may be obtained) or take solution direct from dialysis.
  2. Add 1 mL of 0.1 M CaCl2 solution per 100 mL.
  3. Add 0.4 mg RNAse per mL LPS solution.
  4. Leave at 37 °C for 3-4 h or in a dialysis bag at 22 °C over night, in either case with 1 drop of CHCl3 .
  5. Desalt by gel filtration or dialysis over night.
  6. Freeze-dry.


  1. Chloroform is added as an antibacterial agent.
  2. If phenol extraction has preceded, it is possible to concentrate the solution and run directly.
  3. The enzyme is normally such a small amount that it can be neglected. If desired it may be removed by conventional methods, e.g. phenol extraction.