Determination Of Monoamine Neurotransmitters In Biological Fluid With Functionalized Magnetic Particles Extraction Coupled With High Performance Liquid Chromatography Analysis

Monoamine neurotransmitters, norepinephrine (NE), epinephrine (E), dopamine (DA) and5-hydroxytryptamine (5-HT), play significant functional roles in the central nervous systems. The concentration of monoamine neurotransmitters in body fluids was related to the human health closely. Analysis of monoamine neurotransmitters in biological samples is still a challenge because of the complexity in composition and low concentration of monoamine neurotransmitters in body fluids. Therefore, it is significant to develop a high efficient separation and enrichment method of monoamine neurotransmitters. Compared with conventional sample pre-treatment methods, functional magnetic particles for solid phase extraction (SPE) have received a great deal of attention from medicine, biology, environmental protection and many other fields due to the safety, rapidness and simplicity in separating and enriching target analytes from the complex matrixes. However, the researches on separation and enrichment for monoamine neurotransmitters with functional magnetic particles are not found.In this study, three types of functionalized magnetic nanoparticles were successfully synthesized and characterized by a series of methods. The as-prepared nanoparticles were used for the separation and enrichment of monoamine neurotransmitters in urine, respectively, and showed satisfactory results.The details are summarized as follows:1. The concepts, the analysis methods and the sample pre-treatment methods of monoamine neurotransmitters are summarized, especially for the application of solid-phase extraction and the synthesis methods of functional magnetic particles. 2. A novel phenlyboronic acid functionalized magnetic nanoparticle (Fe3O4@APBA NP) was prepared using chemical multi-step covalent modification. The prepared Fe3O4@APBA NPs were characterized by transmission electron microscopy (TEM), Fourier-transformed infrared spectroscopy (FT-IR), X-ray diffraction (XRD) and vibrating sample magnetometer (VSM). These particles were used for the separation and enrichment of four monoamine neurotransmitters in urine, and exhibited high selective to catecholamines (NE, E, and DA) to demonstrate the highly selective separation ability of boronate to cis-diol containing analytes.10mg of Fe3O4@APBA particles were required for5μg/mL catecholamines (CAs) at pH8.0when the extraction time was5min, and the analytes were eluted with0.02mol/L of HC1-CH3OH solution easily. Combined with high performance liquid chromatography-electrochemical detector analysis (HPLC-ECD), the limits of detection of NE, E and DA were reached to7.8,2.0,7.9ng/mL respectively, and the relative standard deviations were less than5.3%while the linear ranges were all0.01-2.0μg/mL. The propesed method was used to determine the concentrations of NE, E, and DA in human urine. The results demonstrate that the proposed method is simple, rapid and sensitive for analyzing of CAs in urine.3. A novel titania functionalized magnetic mesoporous nanoparticle (Fe3O4@nSiO2@mTiO2NPs) were successfully synthesized through sol-gel method. The prepared magnetic nanoparticles were characterized by TEM, FT-IR, XRD, VSM and BET. These particles were used for the separation and enrichment of four monoamine neurotransmitters in urine, and showed high selective adsorption capacity to the three catecholamines, NE, E, and DA. The separation and enrichment conditions, including sample pH, the extraction time, the kinds of buffer solutions, the amount of magnetic nanoparticles and the elution conditions were investigated. Combined with HPLC-ECD analysis, the limits of detection of NE, E and DA were reached to11.8,0.66, and6.0ng/mL, respectively, while the linear ranges were all0.01-1.5μg/mL. The enrichment factors for NE, E, and DA were8,8, and10, respectively. The propesed method was used to determine the concentrations of NE, E, and DA in human urine. The results demonstrate that the proposed method is simple, rapid and sensitive for analyzing of CAs in urine.4. A novel carboxymethyl cellulose functionalized magnetic nanoparticles (Fe3O4@CMC NPs) were successfully prepared by one-step chemical coprecipitation and characterized by TEM, FT-IR, XRD, and VSM. These particles were used for the separation and enrichment of four monoamine neurotransmitters (NE, E, DA,5-HT) in urine. The separation conditions, including sample pH, the extraction time, the amount of magnetic particles and the elution conditions were investigated. Combined with HPLC-ECD analysis, the limits of detection of the two monoamine neurotransmitters (E, DA) were reached to7.5,9.6ng/mL, respectively, while the linear ranges were all0.01-1.5μg/mL. The propesed method was used to determine the concentrations of E and DA in human urine. The results demonstrate that the proposed method is simple, rapid and sensitive for analyzing of E and DA in urine.