Development and validation of a high performance liquid chromatography method for oligodeoxynucleotides determination in a novel liquid crystal-based formulation

ULLIO GAMBOA G; JUAN M. LLABOT; SANCHEZ VALLECILLO, MARIA FERNANDA; MALETTO, B.; PALMA S. D; ALLEMANDI D,

Analytical Chemistry Research

ELSEVIER

Año: 2015 vol. 4

2214-1812

The therapeutic benefit of phosphorothioate oligodeoxynucleotides (PS-ODN) containing immune stimulatorysequences has been demonstrated in animal models of cancer and infection. Several tools areavailable for the determination of these oligonucleotides in biological samples and pharmaceuticalpreparations, including UV spectroscopy, dye binding, isotopic tracing, capillary gel electrophoresis(CGE), hybridization-based enzyme-linked immunosorbent assay (ELISA), and chromatography techniques.However, due to inter-assay variability and accuracy problems associated with the afore mentionedmethods, we have developed and validated an isocratic high performance liquid chromatographic(HPLC) for analytical determination of PS-ODN containing unmethylated CpG motifs (CpG-ODN).Validation under Food and Drug Administration (FDA) guidelines of the analytical parameters include:linearity (r2 0.9996), LOD (0.86 lg/ml) and LOQ (6.25 lg/ml), intra (0.19?3.37%) and inter-day precision(0.63?3.75%) expressed as relative standard deviation (RSD), and robustness parameters (less than2.80%). Using this method, recoveries ranging from 89.9% to 99.9% were obtained. Thus, this method providesa simple, sensitive, precise and reproducible examination which can be readily adapted for theassessment of CpG-ODN in different pharmaceutical preparations.2 0.9996), LOD (0.86 lg/ml) and LOQ (6.25 lg/ml), intra (0.19?3.37%) and inter-day precision(0.63?3.75%) expressed as relative standard deviation (RSD), and robustness parameters (less than2.80%). Using this method, recoveries ranging from 89.9% to 99.9% were obtained. Thus, this method providesa simple, sensitive, precise and reproducible examination which can be readily adapted for theassessment of CpG-ODN in different pharmaceutical preparations.