Research On The Construction Of Multi-component Mixed Aqueous Two-phase System And Its Application On The Separation And Analysis Of Antibiotic Residues In Environmental Water Samples

Antibiotics have been widely used in medical and health,animal husbandry,aquaculture and agriculture due to their effective antibacterial properties.However,with the irrational and excessive use of antibiotics,large number of drug-resistant bacteria have emerged,causing widespread concern about the abuse of antibiotics.Therefore,the construction of separation and detection methods for antibiotic residues in the environment has become a hot research topic for researchers.Due to the complexity of antibiotic matrix components and the abundance of interfering substances in the environment,antibiotic residues are usually present in trace or ultra-trace amounts.Therefore,it is imperative to construct an easy-to-operate,efficient,rapid and environment friendly method for the separation and detection of antibiotic residues.Based on the unit aqueous two-phase system,the multi-component mixed aqueous two-phase system is constructed by combining the same or different aqueous two-phase system The constructed multi-component mixed aqueous two-phase system overcomes the defects of large viscosity,long phase formation time,large amount of organic solvent and narrow polarity adjustment range of the unit aqueous two-phase system,improves the extraction rate and distribution coefficient of the target extract,reduces the amount of organic solvent and saves the phase formation cost.s in this paper.The liquid-liquid equilibrium behavior and phase separation ability of the multi-component mixed aqueous two-phase system were deeply studied,which provided sufficient theoretical basis for its application.This system was applied to the separation and separation of antibiotic residues in environmental water samples,and the significant influence of various factors in the system on the distribution behavior of target substances was discussed.Through multi-factor experiments,the optimal extraction experimental conditions were determined.Combined with high performance liquid chromatography technology,the antibiotic residues were determined,which provides new technical means and scientific basis for the separation and detection of other trace and ultra-trace pollutant residues in the environment.The specific research contents are as follows:1.Construction of PEG1000/[Bmim]BF₄/small molecular alcohol-small molecular alcohol-salt mixed aqueous two-phase system and study on liquid-liquid equilibrium behavior.?1?Through the fitting of a series of PEG1000/[Bmim]BF₄/small molecular alcohol-small molecular alcohol-salt multi-component mixed aqueous two-phase system of binodal curves data with a plurality of nonlinear equations,the most suitable equation for the system studied in this subject was selected as:₁=exp(a+b₂^0.5+c₂+d₂²).The liquid-liquid equilibrium composition of the system was calculated by using the“lever principle”and successfully fitted by Othmer-Tobias and Bancroft equations.?2?By calculating the effective exclusion volume?EEV?of PEG1000/[Bmim]BF₄/small molecular alcohol-small molecular alcohol-salt mixed aqueous two-phase system under different conditions,and drawing the binodal curves diagram,the influences of the selection of small molecular alcohol in organic components,the ratio of organic components?v/v?,the types of phase forming salts and temperature on the phase separation capability of the system were discussed,and the following conclusions were drawn:?a?in PEG1000/[Bmim]BF₄-small molecular alcohol-salt mixed aqueous two-phase system,the addition of 1-propanol in organic components is most beneficial to the phase separation of the system;?b?in PEG1000-small molecule alcohol-salt mixed aqueous two-phase system,when the ratio of PEG1000 to small molecule alcohol?v/v?is 2:1,the phase separation ability of the system is the strongest.In the mixed aqueous two-phase system of[Bmim]BF4-small molecular alcohol-salt,the phase separation ability of the system is strongest when the ratio of[Bmim]BF4 to small molecular alcohol?v/v?is 1:1;?c?the salting-out capability of cations is Na⁺>K⁺,Na⁺>NH₄⁺,and that of anions is SO₄^2->OH^-,SO₄^2->H₂PO₄^-;?d?for PEG1000-small molecule alcohol-salt mixed aqueous two-phase system,the higher the temperature,the stronger the phase separation ability of the system.For[Bmim]BF₄-small molecule alcohol-salt multi-component mixed aqueous two-phase system,lowering the temperature is beneficial to phase separation.For the double small molecule alcohol-salt multi-component mixed aqueous two-phase system,in the high salt concentration region,the influence of temperature on the phase separation ability of the system constructed by the low-valence salt is higher than that of the high-valence salt.With the increase of temperature,the binodal curves moves to the left,the slope of tie-line simultaneously increases,and the phase separation ability increases.2.PEG1000/[Bmim]BF₄/small molecular alcohol-small molecular alcohol-salt mixed aqueous two-phase system for the separation and detection of antibiotic residues in environmental water samples.?1?PEG1000-1-propanol-?NH₄?₂SO₄ multi-component mixed aqueous two-phase system was constructed to extract and separate levofloxacin hydrochloride residue.The effects of the selection and ratio of organic components?v/v?,types and concentrations of phase salts,extraction temperature,pH value of the system and other factors on the distribution behavior of antibiotics in the system were discussed.The method was combined with high performance liquid chromatography to the extraction and detection of levofloxacin hydrochloride residues in environmental water samples.The minimum detection limit in the water sample labeling experiment is 0.10 ng?ml^-1,the recovery rate of antibiotics is 93.53⁹6.66%,and the relative standard deviation is 1.74².75%.PEG1000-1-propanol-?NH₄?₂SO₄ multi-component mixed aqueous two-phase system was constructed to extract and separate tetracycline hydrochloride residue.The significance of various influencing factors was discussed,and the extraction experimental conditions were optimized through response surface experimental design.Under the determined optimal experimental conditions,the method was combined with high performance liquid chromatography to the extraction and detection of tetracycline hydrochloride residues in actual water samples.The recovery rate of spiked samples is93.96⁹5.84%,the relative standard deviation is 1.26².17%,and the minimum detection limit is 0.20 ng?ml^-1.By measuring the viscosity of organic components,it is fully demonstrated that the multicomponent system overcomes the defect of large viscosity of unit system.Compared with PEG1000-salt unit aqueous two-phase system,the phase formation speed is faster,and the extraction rate and partition coefficient are both improved.?2?Establish[Bmim]BF₄-1-propanol-NaH₂PO₄ multi-component mixed aqueous two-phase system to extract and separate ciprofloxacin residues.The distribution of target substances in the system can be controlled by changing the types and concentrations of phase-forming substances.Thermodynamic data proves that the distribution of CIP in the two phases is mainly the result of hydrophobic interaction.This technique was combined with high performance liquid chromatography to determine antibiotic residues in the lake water.In the standard addition experiment of water sample,the recovery rate of CIP is 91.14⁹7.73%,RSD is 1.03².16%.[Bmim]BF4-1-propanol-?NH₄?₂SO₄ multi-component mixed aqueous two-phase system was applied to extract and separate tetracycline hydrochloride residues.The significance of single factor was discussed,and the optimal extraction conditions were determined by multi-factor experimental design.The multi-system extraction separation technology combined with high performance liquid phase detection technology is used to determine the residual antibiotics in the lake water.In the standard addition experiment of water samples,the TC recovery rate is 91.76⁹8.96%,RSD is 0.76².18%.Compared with the unit aqueous two-phase system,the multi-component mixed organic solvent has less dosage and higher extraction rate,which provides a new direction for the research on the extraction and separation of antibiotic residues in environmental water samples by the aqueous two-phase system.?3?Construct an ethanol-1-propanol-NaH₂PO₄ multi-component mixed aqueous two-phase system to extract and separate norfloxacin residues.The mixed organic phase formed by ethanol and 1-propanol/2-propanol can adjust the polarity of the mixed organic phase by adjusting the ratio of the two alcohols?v/v?,so as to achieve the same polarity as the target extract,thereby improving the extraction rate of the target extract.The multi-component mixed system was combined with high performance liquid chromatography to detect the residual NOR in the lake water,and the standard addition recovery rate is89.88⁹4.87%,and RSD is 1.25¹.96%.Ethanol-2-opropanol-?NH₄?₂SO₄ multi-component mixed aqueous two-phase system was applied to extraction and separate tetracycline hydrochloride residues.The optimal experimental conditions obtained by single factor and response surface design are:the dosage of 2-opropanol is 2.0 mL?the volume ratio of ethanol to 2-opropanol is 1:2?,the concentration of?NH₄?₂SO₄ is 0.250 g?mL^-1,and the pH value is3.5.The multi-component mixed aqueous two-phase system extraction separation technology was combined with a high performance liquid phase detection technology to determine antibiotic residues in actual water samples.In the standard addition experiment of water samples,the TC standard addition recovery rate is 89.96⁹5.43%,RSD is 1.16².10%.Compared with other multi-component mixed aqueous two-phase systems,the small molecule alcohol-small molecule alcohol-salt multi-component mixed aqueous two-phase system constructed in this paper has the advantage of low phase cost.And compared with small molecule alcohol-salt unit aqueous two-phase system,the extraction separation effect is better.