Separation of Adenosine Nucleotides by Reversed-Phase Mixed-Mode HPLC Using LC/MS-Compatible Conditions

Description

Adenosine mono-, di-, and triphosphate (AMP, ADP, and ATP) are hydrophilic nucleotides that play crucial roles as the building blocks of DNA and RNA. Each molecule consists of one or more phosphate groups, an adenine base, and a ribose sugar. Due to their high polarity, these nucleotides exhibit poor retention on traditional reversed-phase columns.

Using the Zodiac HST B2 reversed-phase anion-exchange column, effective retention and separation of the three nucleotides were achieved. Retention times are primarily influenced by buffer concentration and pH. Notably, ADP and ATP require higher buffer concentrations to facilitate proper elution.

This method is compatible with LC/MS and suitable for the analysis of nucleotides in complex biological matrices such as plasma, blood, urine, and other biofluids. It is also adaptable to other detection techniques. The approach is robust, reliable, and demonstrates good tolerance to matrix interference. However, additional sample preparation may be necessary depending on the sample type.

Condition

1. Sodium
2. Adenosine monophosphate
3. Adenosine diphosphate
4. Adenosine triphosphate