Question: I am analyzing phosphorylated sugars and obtain poor HPLC peak shapes with very broad tailing. What are some possible solutions?
Answer: We have a number of strategies for fixing problems with peak shape for phosphorylated compounds:
• Ensure the column is not overloaded. We recommend an injection volume of 1 µL or less with ANP methods.
• Use Teflon bottles instead of borosilicate glass for your mobile phase reservoirs. Sodium from the glass can cause peak shape problems for anionic compounds.
• Use 5–10 microM EDTA in the mobile phase and sample diluent. This will help chelate metals that cause problems for phosphates.
• Try adding 0.01% ammonia to the sample diluent.