Determination Of Amino Acids By Fluorescence Method

Amino acids are the basic element of protein which is essenitial to huaman life. They are the main material in the synthesis of protein, and also the nutrition of feed and food. They can also be used to prevent and cure diseases. Besides, they can be used as an effective component of cosmetics and synthetic surface active agents, chemical raw material medicine andother industrial products. So, it is an important technology for the qualitative and quantitative detection of amino acids both in agricultural, ndustrial production and life science. At present, there exist limitted methods for the detection of amino acids, such as chemical method(including formaldehyde titration method and Kjeldahl method), spectrophotometry, thin layer chromatography, paper chromatog raphy, liquid chromatography(LC), gas chromatography(GC), capillary electrophoresis, electrochemical method and amino acidanalyzer and so on. But methods metioned above have the disadvantages of long analysis time, complicated operation and so on. In contrast, fluorescent spectro metryhas the advantage of rapid detection, low detection limit, hig h sensitivity, wide response range, less sample etc. So it is of great significance to establish new fluorescence spectrometry for the determination of amino acids.The thesis mainly includes the following three parts:1.In this paper, we the classification and function of amino acids, and the methods for determination of amino acids and its applications are reviewed in details.Moreover,we also introduce the basic principles of fluorescence spectrometry.2. Several kinds of new composite fluorescence probes for the determination of amino acides have been explored:①In the Kolthoff buffer solution(p H=10.7), fluorescence intensity of Acridine red, which is quenched by Pd2+ can be enhanced when adding a certain amount of laevorotatory-isoleucine(L-isoleucine), Thereby, we establish a fluorescence spectrometry method for the detection of L-isoleucine. When the excitation slit width is 5nm and emission slit width is 10 nm, we get the following results: the linear regression equation: F=-33.884+468.07C(mg/L); correlation coefficient: R=0.9984, linear range: 0.125~1.375mg/L; the detection limit: 0.00027 mg/L. Besides, we calculate the association constants of Pd2+- Acridine red(k1=1.62×102) and Pd2+-isoleucine(k2=7.04×102) by the method of spectrophotometric.②With a certain concentration of B-R buffer solution(p H=9.5), and surfactant of Tween-80, fluorescence intensity of calcein, which is quenched by Pd2+ can be enhanced after adding laevorotatory-arginine. Thereby, we establish a fluorescence spectrometry method to detect the content of laevorotatory-arginine, of which the added amount is proportional to the enhancement rate. It is established that the linear range is different when the slit width changes, so is the detection limit. Thus we can use different slit width to meet various requirements according to the samples we treat. By testing actual samples and the reliability of our experimental method, we discover that our method is reliable for determining the content of laevorotatory-arginine in Radix according to the results.③With a certain concentration of Michaelis buffer solution(pH=6.1), and the presence of CTMAB, fluorescence intensity of fluorescein, which is quenched by Pd2+, can be enhanced after adding L-leucine, and the added amount is proportional to the enhancement rate. So, we establish a new method for the determination of L-leucine by using a composite fluorescent probe.3. In the paper, we have studied for the interaction theory between amino acids and fluorescent probes, and theabsorption spectra and fluorescence spectra of the fluorescent probes, the complex. In addition, the mechanisms of them are analyzed preliminarily.