| Fluoroquinolones have been widely used to prevent and remedy animal diseases in animal husbandry because of its wide-spectrum and strong antibiotic activity. But unreasonable or abuse adding medicated feed additives in feeds will lead to the drugs remaining in animal derived food, which will harm the human health through food chain. Thus, it's necessary to establish a method of high selectivity, many types of multi-component, efficiency and rapid for fluoroquinolones detection.With norfloxacin, cipfloxacin, enrofloxacin and sarafloxacin as detecting objects, the pretreatment method and detection technology of suckling piglets and meat chicken feeds were studied. Then established the assay method with four fluoroquinolones by high performance capillary electrophoresis; the high selectivity molecularly imprinted solid phase extraction column was synthesized with norfloxacin as the template. Finally, detection method of norfloxacin's MISPE-HPCE and four kinds of fluoroquinolones'LLE-HPCE in feed was established.1 Establishment of HPCE method for determining four fluoroquinolonesThe seperation method of four kinds of fluoroquinolones was established by high performance capillary electrophoresis through investigating the effects of the pH, composition and concentration of running buffer, separation voltage and temperature on seperating FQs. The optimum conditions obtained from the the orthogonal test were as follows: running buffer of 80 mmol/L citric acid-80 mmol/L sodium citrate (pH of 6.5), separation voltage of 22 kV, temperature of 22℃and detection wavelength of 274 nm. The four drugs seperated completely in 10 min. And correlation coefficient from 0.9991 to 0.9995 in the range of 1.56~25μg/mL, RSD of migration time from 1.36% to 1.69%, RSD of peak area from 3.69% to 5.89%, and the recovery rate from 84.42 % to 107.5%.2 Preparation of molecularly imprinted solid phase extraction of norfloxacinA molecularly imprinted polymer (MIP) properties of norfloxacin (NOR) was prepared by thermal polymerization in which norfloxacin, methacrylic acid (MAA), ethylene glycol dimethacrylate (EDMA) and azobisisobutyronitrile (AIBN) were as the template molecule, functional monomer, crosslinker, and ignition primer respectively. The optimum preparation conditions were as follows: solvent of 80% acetonitrile(v/v), n (NOR)∶n (MAA)∶n (EDMA) = 1∶6∶30, which were obtained by investigating the effects of kinds of solvent, dosage of crosslinker and functional monomer on the binding properties of MIP. The static equilibrium adsorption was used to study the binding capacity and selectivity properties of MIP. The static adsorbance of the MIP and NMIP for norfloxacin was 4.57-154.83μmol/g and 1.59-80.91μmol/g respectively, which revealed that the adsorption capacity of MIP was better. The static adsorption distribution coefficient of MIP to norfloxacin, cipfloxacin, enrofloxacin and sarafloxacin were 59.98, 32.87, 25.33 and 22.11 mL/g respectively. Meanwhile, the binding capacity of MIP to norfloxacin and sulfamonomethoxine were 56.24 and 3.37μmol/g respectively. The results of substrate selecting experiments showed that the MIP had specific combination characteristic for norfloxacin, and the molecularly imprinted solid phase extraction of norfloxacin could be prepared with composite MIP.3 Optimization of the pretreatment of four fluoroquinolonesThe effects of extraction solvents on the feeds extraction were studied through optimizing the pretreatment, and the samples were extracted with ultrasonic for 20 min after being severely mixed on vortex mixer for 3 min, using 50% acetonitrile (v/v) as the extraction solvent. The average recovery rates of four FQs were from 85.65% to 105.6%.4 Establishment of MISPE-HPCE method for norfloxacinWith 50% acetonitrile as extracting solvent and MISPE of norfloxacin as study object, the different parameters were optimized, such as components of activators, eluate and volume of eluent, and evaluation for the recovery rates of NOR. The established optimum pretreatment method were as follows: activators of methanol and water, eluent of methanol-acetic acid (v: v=8:2), with which the recovery rates of NOR were 80.5-86.67%, RSD value were 1.36-4.9%. The recovery rate and the purity of NOR extracted with MISPE was significant higher than that with SPE. The MISPE-HPCE detection method of norfloxacin was established by combining the optimum pretreatment method and the detection method.Generally, it is shown that the MISPE-HPCE of NOR is with high selectivity, precision and accuracy, and the four FQs are separated by simple, rapid and easy HPCE in 10 min, both of which are important for monitoring FQs in feeds and veterinary drug residues in animal derived food and will provide the available data for establishing standard monitoring system for the veterinary drug residue. |
