| In this study, we started with the present status of FQs residues in environmental water, and established a SPE-UPLC-MS/MS detection method to investigate the current pollutant situation of the drug in three kinds of water. On this basis, QuEChERS method using improved optimization of the FQs drug residues in aquatic products was carried out to get a better sample preparation in seven kinds of tissues, and detection methodologies were evaluated. The analytical methods that residues of six kinds of FQs in aqueous solution and biological matrices were established. Then the work that the distribution coefficients of tissue-plasma and plasma protein binding rates of norfloxacin and sarafloxacin with the carp were carried out. Finally, with presence of norfloxacin in environmental water which stressed carp in vivo metabolism of the amino acid, the screening of small molecule residue marker was preliminarily developed.A method of ultra-performance liquid chromatography coupled to triple quadrupole tandem mass spectrometry (UPLC-MS/MS) has been developed for the analysis of the six FQs residues in environmental water. The conditions of water sample preparation and analytical instrument were investigated and optimized to get a rapid, sensitive, qualitative and quantitative method to detect the target analytes in surface water and waste water. The method was suitable for robust screening of FQs residues in large basin area. The method validation was investigated and the results showed that the sensitivity was high with the low limit of detection and the limit of quantification being0.4ng·L-1and1.0ng·L-1, respectively. It performed good linearly dependent coefficient and recovery, and the precision was also satisfactory. Finally the method was successfully applied to monitor the six FQs residues in surface water and waste water from June2009to December2011, and the results indicated that enorfloxacin, norfloxacin and ciprofloxacin were found in the water.An improved QuEChERS method was used to pretreat the six FQs residues in seven tissues from aquatic products by comparing the SPE and traditional QuEChERS method, and a new QuEChERS-UPLC-MS/MS method was developed. The choice of extraction agent, style of extraction, methods of purification, and influence of different quality of Na2SO4on recovery and time of evaporation were investigated to get an optimized preparation. Matrix effects study results showed that the liver sample blank matrix interferences was little after treatment by improved QuEChERS method. Sensitivity results showed that in the plasma, liver, kidney, intestine, testis or ovary, gill the detection limits were0.5μg·L-1,0.25μg·kg-1,1.0μg·kg-1,0.25μg·kg-1,0.25μg·kg-1 and0.5μg·kg-1, respectively, and the limits of quantification were2times the detection limit. The recoveries of different spiked levels and the precision were both satisfactory.We carried out the research about the distribution coefficient of tissue-plasma and plasma protein binding rate on the bais of the last results on the methods of FQs residues in water and tissues. The concentration of NOR in plasma of carp showed no significant difference during4h,5h, and6h afer10mg·kg-1oral administration following by0.5mg·L-1drug bath. The distribution coefficient of muscle-plasma, liver-plasma, kindey-plasma, gill-plasma were0.786、1.068、0.699and1.105, separately. On the contrary, the concentration of SAR in plasma of carp kept relative stability during3h,4h and5h afer10mg·kg-1oral administration, following by1.0mg·L-1drug bath. The distribution coefficient of the tissue-plasma were0.646,0.867,0.655and1.093, separately. Plasma protein binding rate of NOR was analysed at18℃，23℃and28℃with equilibrium dialysis method being adopted, and the time that reached the balance were30h,28h and24h, separately. The binding rate was31.7%that calculated by analysis the concentration of NOR in puffer solution. On the other side, the time of balance of SAR between plasma and puffer solution were42h,35h and30h at the same temperature and concentration level as NOR, separately. The binding rate was22.9%that calculated by analysis the SAR concentration in puffer solution. The contact area was discussed to evaluate the influence and the resut indicated that the balance time of the two drugs both showed a downward trend with the increasing area. But the final numerical value of the binding rate would not be changedFinally, the primary screening of the residue markers of small molecule metabolites was carried out. The experiment was investigated under conditions of environmental stress of NOR which was present in water environment at some concentration levels. A rapidly quantitative method of ultra-performance liquid chromatography （UPLC）with ultraviolet detector had been developed for the analysis of17kinds of free amino acids in fish plasma and amino acids in fish eggs. For the content of the free amino acids in fish plasma, the conditions such as wavelength, agent for deproteinisation for plasma, time and temperature of derivatization and pH value of mobile phase were discussed to get an optimized results. The method was validated by analysis of plasma samples fortified with three kinds of concentration levels mixed amino acids standards, respectively. Depending on the biological matrix and analyte, recoveries and relative standard deviations were both suitable for the analysis method. The volume of8mL6.0mol·L Hcl was chosen to hydrolyze the fish eggs sample and the time and temperature of derivatization were chosen the same as plasma samples. The method was also validated by analysis of three kinds of fortified concentration levels of mixed amino acids standards, respectively, and the results showed that the recoveries and relative standard deviations performed well. In the end, the analysis methods were used to study the preliminary metabolic response mechanism of amino acids under environmental stress of NOR and five kinds of markers of small molecule metabolites of amino acids were chosen to show the FQs existence initiatorily. |
PDF Full Text Request