Food Neotame And Vitamin B12 Column Switching Hplc Determination Study

A online column-switching HPLC system was set up with two HPLC instruments and a VICI six-port valve.It could be applied to determine low content analytes in many food samples with complex matrix.In this paper,new and simple methods to qualitify neotame and vitamin B12 in foods and health foods by column-switching HPLC with PDA has been developed.It was composed of two parts.Part one:Determination of neotame in beverages preserved fruits and cakes by column-switching HPLCNeotame was extracted by sonication with water followed by centrifuge(at 5000rpm for 5min) in the presence of precipitant.The sediment was washed by 20% methanol aqueous solution for three times.The extract was filtered through a filter paper and was then applied to Waters HLB SPE column prior to HPLC analysis. Neotame was determined by PDA at 210nm after its separation on two reversedphase C18 columns in a column-switching HPLC system with a isocratic phase made of water/acetonitrile and phosphate acid(0.5%).The calibration graphs plotted with six concentrations was linear with regression coefficient R=0.9999.The repeatability of the method was evaluated of five shares of one product and the relative standard deviation was 4.7%.The detect limit of the method was 40μg/kg.The recoveries evaluated at spiking three different concentrations on fortified products were above 80%.The method was successfully applied to beverages and preserved fruits.Part two:Determination of Vitamin B12 in health food and infant formula foodVitamin B12 in infant formula food was extracted with 100mM sodium acetate buffer pH4-5(at 105℃for at least 60 min) in the presence of sodium cyanide, followed by filtration step in Whatman 2V filter papers and purification step on a C8 SPE column.The eluent was evaporated to dryness by nitrogen and metered volume to 0.5 milliliter prior to HPLC analysis.An enzymatic hydrolysis(taka-diastase at room temperature for at least 1 min ) prior to the extraction step efficiently released the bound vitamin B12.Vitamin B12 in health food was extracted with methanol and ethanol.The extract was evaporated to dryness in water-bath followed by metered volume to 5 milliliter prior to HPLC analysis.Vitamin B12 was determined by PDA at 361nm after its separation on a size exclusion column and a reversed-phase C18 column in a column-switching HPLC system with a gradient mobile phase made of water and acetonitrile.The calibration graphs plotted with six concentrations was linear with regression coefficient R=0.9996.The repeatability of the method was evaluated of six shares of one product and the relative standard deviation was 4.7%and 8.4%.The recoveries evaluated at spiking three different concentrations on fortified products were above 80%.The method was successfully applied to several health food and infant formula food and was simple reliable as well as short time consumed in comparison with MBA.