Detections Of Nitrogen-containing Molecular Adulterants In Milk Based On Fluorescence Spectroscopy

Milk is a nutrient-rich health food,rich in protein,amino acids,vitamins and other nutrients,so it is very popular among consumers.Since protein content in milk is calculated from nitrogen content,it is possible to adulterate milk with non-protein nitrogen molecules to increase milk apparent protein content.Small-molecule compounds such as melamine,urea,and thiourea have the characteristics of high nitrogen content and low cost,and may be illegally added to milk as adulterants,which threatens consumers’health.Therefore,the accurate detection of nitrogen-containing small molecule adulterants in milk can provide the strong guarantee for food safety.Small nitrogen-containing compounds such as melamine,urea and thiourea have weak fluorescence,which are difficult to be directly detected by fluorescence spectroscopy.However,milk has strong fluorescence,which will have a great impact on the detection of adulterants.Therefore,it is particularly difficult to detect these weakly fluorescent substances in milk.Lanthanide complexes,carbon nanomaterials and metal nanoclusters are often used as the fluorescent probes to detect weak fluorescent substances in complex environments because of their good fluorescence properties.Through the fluorescence response between the fluorescent probes and the analytes,the specific determination of the analyte in complex systems can be realized.Based on the fluorescence spectroscopy technology,europium ion complex,blue carbon quantum dots and gold nanocluster fluorescent probes were developed to achieve quantitative detection of melamine,thiourea and urea in milk,respectively.The main works are as follows:1.Detection of melamine based on europium complex fluorescent probe.Through the calculation and analysis of the molecular structure of the molecular characteristics and interaction mechanism of melamine,europium ion and tetracycline,it was found that the melamine and europium ion-tetracycline（Eu TC）can form the ternary complex through hydrogen bond interaction,which can significantly enhance the fluorescence intensity of the Eu TC complex.Therefore,the Eu TC complex fluorescent probe was developed and the method for detecting melamine based on Eu TC complex fluorescent probe was established.The results showed that the fluorescence intensity of the system had a good linear relationship with the concentration of melamine,and the detection limit was 7.85×10^-8 M.The results show that the fluorescent probe with Eu TC complex has a high sensitivity for the detection of melamine,and the results are superior to the reported detection methods.2.Detection of thiourea in milk based on blue carbon quantum dots fluorescence probe.Analysis of the chemical properties of thiourea shows that thiourea has strong coordination with mercury ions(Hg²⁺),and can compete for the Hg²⁺coordinated with the fluorescent material to restore the fluorescence of the material.Therefore,the carbon quantum dots（CDs）with specific fluorescence response to Hg²⁺was developed,which was combined with Hg²⁺as the“turn-off-on”fluorescent probe,and the method for the detection of thiourea in milk based on the switch fluorescent probe was established.The results showed that there was a good linear relationship between fluorescence intensity of the system and the concentration of thiourea in aqueous solution and milk,and the detection limits were 1.96×10^-7 M and7.70×10^-8 M,respectively.The results showed that the method was superior to the existing detection techniques,and realized the rapid,sensitive and specific detection of thiourea in milk.3.Detection of urea in milk based on gold nanocluster fluorescence probe.Analysis of chemical properties of urea shows that ammonia produced by urease hydrolysis of urea can increase the pH of the system.In order to realize the detection of urea,a gold nanocluster fluorescent probe with sensitive response to pH was developed,and the detection method for urea in milk was established through the fluorescence response to pH.The results showed that the fluorescence quenching degree of the reaction system had a good linear relationship with the concentration of urea in milk,and the detection limit was 1.62×10^-5 M.The urea concentration in milk samples obtained by standard addition method was 284.04 mg/L.Compared with other methods,this work has higher detection sensitivity,and achieves the accurate,sensitive and specific detection of urea in milk.In this work,the fluorescent probes were developed by analyzing the structure and properties of analytes.The fluorescence spectroscopy technology and fluorescence probes were applied to detect melamine,thiourea and urea,respectively.It supplements and expands the application of fluorescence technology and provides the new methods for the food safety supervision.