Construction Of "Carbon Dots"-Based Fluorescence Detection System For Malachite Green In Aquatic Products

Malachite green(MG)is a triphenylmethane compound with a green metallic luster and has certain tritoxicity and genotoxic potential,therefore,eating MG-containing aquatic products poses a threat to human health.In 2002,China included MG in the "List of Veterinary Drugs and Their Compounds Banned in Food Animals",but due to its high efficacy and low cost,illegal use occurs from time to time.Therefore,the development of a fast,simple and accurate MG test method is of great significance to protect the health of consumers.In recent years,carbon quantum dots(CDs)probes have provided new ideas for efficient detection of MG because of their simple preparation,low cost,good selectivity and high sensitivity.At present,most CDs probes used for MG detection are short-wavelength blue fluorescence emission,which are easily interfered by background fluorescence and environmental factors,which limits their detection sensitivity.In this paper,the development of long-wavelength orange-red fluorescent CDs to overcome this defect,and the preparation of fluorescent microspheres with signal amplification effect and the construction of a dual-signal ratiometric detection platform were further explored to improve the sensitivity of CDs-based probes,and the main research contents were as follows:(1)Using rhodamine B as raw material,long-wavelength orange-red CDs(O-CDs)were prepared by hydrothermal synthesis(160℃,2 h),with an optimal excitation/emission wavelength of 555/575 nm and a quantum yield of 37.1%.Its physicochemical properties were characterized by transmission electron microscopy,elemental analysis,Fourier infrared spectroscopy,UV-vis absorption spectroscopy,and fluorescence spectroscopy.Based on the quenching effect of MG on O-CDs probes,a fluorescence detection method based on O-CDs was constructed.Under the optimal operating parameters(O-CDs concentration was 0.1 mg/mL,detection system pH was 5.0,reaction time was 1.0 min),the probe showed good selectivity and anti-interference for MG detection,with linear ranges of 0.01-0.1 μM and 0.1-10.0 μM,and detection limits of 80.0 pM and 0.93 nM,respectively.The sensing mechanism between MG and O-CDs was explored by UV-vis absorption spectroscopy,fluorescence spectroscopy,fluorescence lifetime and zeta potential analysis,and it was found that the fluorescence quenching mechanism of MG to O-CDs stemmed from the synergistic effect of internal filtering effect and fluorescence resonance energy transfer.(2)Using O-CDs as the fluorescence sensing unit and melamine formaldehyde(MF)microspheres as the carrier,bright orange fluorescent microspheres O-CDs-M were prepared by acid catalysis and heating reaction,with an optimal excitation/emission wavelength of 564/581 nm and a quantum yield of 41.5%.Its physicochemical properties were characterized by scanning electron microscopy,energy dispersive spectroscopy,Fourier infrared spectroscopy,UV-vis absorption spectroscopy,and fluorescence spectroscopy.Under the optimal operating parameters(O-CDs-M concentration was 0.07 mg/mL,detection system pH was 5.8,reaction time was 1.0 min),the probe showed good selectivity and anti-interference for MG detection,with linear ranges of 0.01-0.1 μM and 0.1-10.0 μM,and detection limits of 56.7 pM and 0.57 nM,respectively.The sensing mechanism between MG and O-CDs-M was explored by UV-vis absorption spectroscopy,fluorescence spectroscopy,fluorescence lifetime and zeta potential analysis,and it was found that the fluorescence quenching mechanism of MG to O-CDs-M was attributed to the internal filter effect.(3)Blue carbon dots(B-CDs)were prepared by self-release heat using glucose,ethylenediamine,70%H2SO4 solution and concentrated HCl as raw materials.Using the O-CDs prepared in the first part as the working probe and B-CDs as the reference probe,a two-signal ratiometric probe O,B-CDs was constructed for MG detection.Under optimal operating parameters(B-CDs and O-CDs concentration were 0.1 mg/mL,incubation time of the ratiometric probe was 1.0 min,optimal excitation wavelength was 400 nm,detection system pH was 5.8,reaction time was 1.0 min),the ratiometric probe showed good selectivity and immunity for MG detection,with linear ranges of 0.01-0.1 μM and 0.1-15.0μM,and detection limits of 55.0 pM and 0.25 nM,respectively.The sensing mechanism between MG and O,B-CDs was explored by UV-vis absorption spectroscopy,fluorescence spectroscopy,fluorescence lifetime and zeta potential analysis,and it was found that the fluorescence quenching mechanism of MG to O,B-CDs was attributed to the internal filter effect.At the same time,based on the phenomenon that MG caused the color change of the sensing system,the system was prepared into a portable test strip using glass fiber filter paper as the carrier.With the assistance of smart phone color recognizer,the luminous intensity and color change of the sensing system are quantitatively output in the form of fluorescence signals,and a new and efficient CDs-based ratiometric visual detection platform was constructed,which realized the visual detection of the naked-eye.Finally,this work explores the feasibility of this visual paper-based sensor for actual sample analysis for the on-site visual quantitative detection of MG in aquatic products(including silver carp,blackfish,blunt-snout bream,common carp and grass carp),with a recovery rate of 91.7-107.7%and a relative standard deviation of 1.74-6.34%.The detection results of this sensor are consistent with high performance liquid chromatography-ultraviolet method(national standard method),indicating that the detection method has high accuracy and reliability.