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Investigation of Glucose and Mannose Separation by HPLC Method

Glucose (Glc) and mannose (Man) are monosaccharides. Due to the six atoms of carbon and one aldehyde group both belong to the group of aldohexoses. Possible configurations of these monosaccharides are the D- or L-form, depending on configuration of fifth carbon atom in chain. D-forms mostly occur in nature. The aim of this work was to separate the mixture of glucose and mannose using cation exchanger in Na form. The data obtained from tests are used for simulation of SMB mannose isolation and separation efficiency guess estimation. D-mannose and D-glucose were used for chromatographic separation on SMB unit and HPLC analysis Solution of 5 % glucose and 5 % of mannose has been prepared for this work. Separation was firstly realized on two columns which were subsequently temperate on 35.5 °C, 44.8 °C, 54.5 °C and 62.5 °C. The feed mixture has been injected for 3 min at flow rate 1.5 l h-1. The fraction collecting started, when the signal generated by refractometer at the column output differs from its zero level base line. Fractions have been collected in time intervals of 2 to 3 min. The aim of this measurement was the searching of optimal temperature. In this case separation of binary mixture glucose and mannose is programmed with highest separation efficiency. Comparing the obtained data from both views was found, that the separation efficiency in given range is the highest at temperature 62.5 °C. At this temperature the continuous chromatographic separation was performed using chromatographic separator KCHS-SMB-8-N. The achieved concentration profile of separated compounds shows, that the mixture did not ideally separated and the resulting course of compounds cannot verify the steady state. In this work the separation was performed again with wider set of operational parameters. The possibility of continuous separation and the separation efficiency of column were verified firstly with mixture glucose and fructose using chromatographic separator on two serially connected columns at four different temperatures 35.5 °C, 44.8 °C, 54.5 °C and 62.5 °C . The maximum achieved purity and HETP were chosen as the criteria for evaluating the separation efficiency at given temperatures.