| ?-Dicarbonyl compounds (?-DCs) such as 2,3-butanedione (also called diacetyl), 2,3-pentanedione, glyoxal, methylglyoxal, phenyl glyoxal, and glucosone et al. exist in many foods as important flavor compoundss; they also present in human physiological system as intermediate products of sugar metabolism. Numerous studies have shown that accumulation of the ?-DCs in vivo relates to a variety of human diseases, such as kidney disease, Alzheimer's psychosis. There are many studies about the qualitative and quantitative analysis of this kind of compounds. Although these methods can meet the analysis need in many fields, new derivatization reagents are still needed to improve the detection speed and sensitivity. HPLC was widely used in ?-DCs determination for its quickness, high sensitivity and relatively lower cost. In this study, we established a precise and accurate precolumn derivatization-HPLC method using 3,3'-diaminobenzidine(DAB)and its single diacetyl derivative DAB-dia as derivatizing reagents to determine the content of 2,3-butanedione in alcohol samples.When DAB (HCl salt) was used as a derivatizing reagent, it reacted in a large excess with diacetyl (ratio of DAB to diacetyl?10:1) in water at room temperature for 5min. For DAB-dia as a derivatizing reagent, the reaction was carrired out in water at room temperature for 10min, but the ratio of DAB-dia to diacetyl was more than 10:1. The HPLC conditions were as follows:ODS column with methanol-water gradient as mobile phase.At the begining 10 minutes, methanol was increased from 60% to 100%, and from 10 to 15min, the proportion of methanol from 100% back down to 60%. At the last two minutes, the column was equilibrated with 60% methanol. Detector wavelength:254 nm. Results showed that the correlation coefficient (R2) for the DAB method was 0.9992 in diacetyl concentration range of 0.18?M-118.82?M. The limit of diacetyl detection was 0.09?M and the limit of its quantification was 0.18?M. The RSD(%)of multi-determination was 0.06-1.35. Finally, the content of diacetyl in two alcohols was successful measured with the recoveries in the range of 85.2%-103.6%. For the DAB-dia method, R2 was 0.9991 in diacetyl concentration range of 0.1-100?M. The limits of detection and quantification were 0.02 ?M and 0.1 ?M, respectively. The RSD (%) of multi-determination was 1.3-3.7.We successfully determined five kinds of beers with recoveries in the range of 81.7%-88.5%.In summary, the precolumn derivatization-HPLC method set up in this thesis using 3,3'-diaminobenzidine (DAB) and DAB-dia as derivatizing reagents to determine 2,3-butanedione in alcohol samples is simple, rapid, and sensitive. Therefore, it could also be developed to analyze other ?-dicarbonyl compounds in foods, beverages and some biological samples as well. |
PDF Full Text Request