Highly Selective Determination Of Several Amino Acids In Food Samples Using Pre-column Derivatization Capillary Electrophoresis Coupled With Electrochemiluminescence Detection Method

In this thesis,formaldehyde and sodium borohydride were used as the main derivatization reagents for an N-methylation derivatization reaction of theα-amino group of severalα-amino acid molecules in food samples.The completely derivative products forα-amino group of amino acids can be obtained through the N-methylation reaction including nucleophilic addition,dehydration and chemical reduction reaction steps.And,it was shown that these derivatives would produce more stronger electroluminescence signal for analytic aim when the oxidized tris(2,2-bipyridyl)ruthenium react with them than non-derivedα-amino acids.Based on this principle,a new method for the highly selective determination of several amino acids in food samples by pre-column derivatization capillary electrophoresis coupled with electrochemiluminescence detection(CE-ECL)was successfully established.Moreover,when the inner wall modified capillary coating with chitosan was applied as separation capillary,we experimentally optimized the electrophoresis testing conditions and the amount of separation additive,and achieved the separation and quantitative analysis for some amino acids in the actual food samples such as melon tissue,rice wine and soy sauce.The main research contents are as follows:1.Determination of citrulline content in melon pulp by pre-column derivatization capillary electrophoresis coupled with electrochemiluminescence detection(CE-ECL)In this chapter,formaldehyde and sodium borohydride were used as the main derivatization reagents for an N-methylation derivatization reaction of theα-amino group in citrulline molecule,in order to obtain a single derivative product with the strong electrochemiluminescence signal.Under the optimum experimental conditions,there was a good linear relationship between the electrophoretic peak intensity of the derivative and its original concentration of citrulline in the range of 9.0 to 2.5×10~2μmol/L(r~2=0.9996).The lower detection limit was tested as 3.2μmol/L(S/N=3).The relative standard deviations of peak height and migration time for the same standard sample to perform six parallel testing were 2.6%and 0.92%,respectively.2.Determination of proline,alanine and citrulline in yellow rice wine samples by pre-column derivatization CE-ECL methodIn this chapter,the three kinds ofα-amino acids in the yellow rice wine samples would be subjected to N-methylation derivatization reaction,and some studies on the capillary electrophoresis analysis was carried out by adding separation additives into the running buffer.In addition,the some important analytical conditions such as the applied potential,separation voltage and additive dosage were optimized in order to achieve good separation efficiency and sensitive ECL detection for the threeα-amino acids in yellow rice wine samples.The detection limits of these three amino acids were 3.1μM,6.9μM and 8.9μM(S/N=3),and the relative standard deviations of peak height and migration time for the same L-proline standard sample to perform six parallel testing were 2.7%and 0.73%,respectively.Moreover,the standard addition method was used to quantitatively determination of three amino acids in four kinds of yellow rice wine samples.The recoveries ranged from 87.0%to 110%.3.Simultaneous determination of citrulline and arginine in soy sauce samples by pre-column derivatization CE-ECL methodIn this chapter,citrulline and arginine in soy sauce samples were separated and determined simultaneously by CE-ECL method.Under the optimum experimental conditions,the two electrophoretic peaks in a mixed amino acid standard sample can be completely separated within 10 minutes.The ECL peak values of citrulline and arginine derivatives have a good linear relationship with their original concentration in the range of 20~2.5×10~2μM and 25~5.0×10~2μM(r~2=0.9991-0.9998),and the detection limits were 7.1μM and 8.9μM,respectively.The relative standard deviations of peak height and migration time for the same L-Arginine standard sample to perform six parallel testing were 3.2%and 1.4%respectively.Moreover,citrulline and arginine contents in four kinds of soy sauces were determined by standard addition method.The average recovery was between 87.9%and 109%.