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The analysis of bases is important in many biotechnology research area. The bases are the building block of nucleotide and bases change of structure and function is the main causes of inherited diseases and human cancers. Therefore, they need to be quickly and accurately
analyzed. Separation investigations in this study focus on the base pairs of guanine and cytosine among nucleotide. Expetiments for analysis of guanine and cytosine are performed 3-zone SMB (Simulated Moving Bed). SMB chromatography is a fast and accurate technique for analysis of nucleotide. The SMB unit set up by three C18-HPLC columns was used to separate guanine and cytosine. Aspen simulation was performed to determine the operating conditions such as switching time, eluent flow rate, raffinate and extract flow rates. The outlet streams of raffinate and extract were analyzed in batch HPLC system. The results show that raffinate and extract separation is possible by SMB technology and purity values range over 90%.