Determination Of β-lactam Antibiotic Residues In Milk

The unique structure of (3-lactam antibiotics is theβ-lactam ring, it is one of the most widespread antibiotics. Antibioties are widely used for animal clinic therapy, and are frequently added into feeds of the prevention or treatments of animal diseases. However, illegal administration will lead to residues in edible tissues, especially in the milk of galaetophorous animals, and finally do latent harm to human health. Thus, it is significant to develop a series of sensitive, reliable and convenient methods for the determination of penicillin antibiotics. In this paper, fluorescence analysis of common ofβ-lactam class-antibiotic, such as penicillin G, ampicillin, ceftiofur sodium and cefradine have been studied. The main contents are as follows:Part 1:Catalytic kinetic spectrophotometric determination of penicillin G and mechanism analysisThe new catalytic kinetic spectrophotometry with amaranth fading for determining trace amounts of penicillin G was proposed using hydrogen peroxide as oxidizing agent. The low concentration range of penicillin G obeys catalytic kinetic spectrophotometric mechanism analysis, the linear range of determination was 0.20～4.00μg/mL, the regression equation is y=-0.015x+0.311(x:μg/mL),r=0.9998. And the higher concentration range obeys inhibitory kinetics spectrophotometric mechanism analysis, the linear range of determination was 4.00～50.0μg/mL, and y=0.001x +0.280(x:ug/mL), r=0.9997. The wavelength for determination is 520nm. The detection limit is 1.0×10-7g/mL. The maximum relative standard deviation is 1.23%. The recovery is 96.7%～104.1%.Part 2:Kinetic spectrophotometric determination of trace ampicillin in milk with sensitization effect of surfactantA catalytic kinetic spectrophotometric method for the determination ampicillin in milk has been developed. It was based on the catalytic effect of ampicillin on the reaction between xylenol orange and aquae hydrogenii dioxide with Tris-HCl buffer solution (pH=7.0)and Triton100. Experimental conditions, such as reaction time, temperature, and the concentration of reagents were optimized by measuring the absorbance at 580nm.Under the best condition, the detection limit was 2.0×10-8g/mL with linear range of 0.10～ 50.00μg/mL. The linear equation was y=0.240x+2.151, r=0.9972. The recovery was between 98.1%and 103.6%. This method is quick, stable and with little interferences. It has been applied for the detection of ampicillin in milk.Part 3:Fluorescence resonance energy transfer quenching for determina-tion of trace ceftiofur sodium in milkIt was found that the effective energy transfer could occur between acridine orange and neutral red in the surfactant AES solution containing Britton-Robinson solution(pH=7.00), which improved the fluorescence intensity of neutral red. The fluorescence of acridine orange-neutral red systems was quenched with the addition of ceftiofur sodium. Therefore, a novel fluorescence quenching method was developed for indirect determination of ceftiofur sodium by acridine orange-neutral red fluorescence resonance energy transfer. This method has been applied for the determination of ceftiofur sodium in milk, and the analytical results were in agreement with those obtained by the HPLC method. The detection limit was 0.1 X 10-7g/mL with linear range of 0.10μ～60.00μg/mL. The linear equation was y= 399.7x+38.88, r=0.9980. The recovery was between 95.9%～102.7%. This method is quick, stable and with little interferences. It has been applied for the detection of ampicillin in milk.Part 4:Determination of cephalosporins B antibiotics in milk based on the fluorescence quenching of bovine serum albuminA new method for the determination of trace amounts of cephalosporins B antibiotics has been developed. This method is based on the quenching of fluorescence of bovine serum albumin(BSA). The excitation and emission wavelengths were found to be 280 nm and 341 nm, respectively. The detection limit was 0.5×10-7g/mL with linear range of 0.10～30.00μg/mL for the ceftiofur sodium determination, and its linear equation was y= 563.lx+360.5 (x:μg/mL), r=0.9998, its recovery was between 90.2%～98.5%. The detection limit was 1.0×10-7g/mL with linear range of 2.50～80.0μg/mL for the cefradine determination, and its linear equation was y=312x+90.1 (x:μg/mL), r=0.9999, its recovery was between 90.1%～97.4%.This method is quick, stable and with little interferences. It has been applied for the detection of ceftiofur sodium and cefradine in milk.