研究目的
To develop a selective and sensitive near-infrared fluorescent probe for the quantitative detection of tris(2-carboxyethyl)phosphine (TCEP) and its application in determining E. coli concentrations in complex biological systems.
研究成果
HC-N3 is a highly sensitive and selective near-infrared fluorescent probe for TCEP detection with a LOD of 92 nM, the lowest reported. It shows rapid response at physiological pH and good selectivity against common interferents. Successfully applied for E. coli quantification in complex biosystems, demonstrating potential for biological and environmental applications.
研究不足
The probe may have limitations in environments with high concentrations of other reducing agents not tested, and the reaction time of 60 minutes might be optimized for faster detection. Specificity to TCEP over H2S is claimed but requires validation in more complex matrices.
1：Experimental Design and Method Selection:
Designed and synthesized an azido near-infrared hemicyanine dye (HC-N3) for TCEP detection based on the Staudinger reaction, where TCEP reduces the azido group to an amino group, leading to fluorescence enhancement. Used UV-Vis absorption and fluorescence spectroscopy for characterization and detection.
2：Sample Selection and Data Sources:
Prepared solutions of HC-N3 and TCEP in phosphate-buffered saline (PBS) at pH 7.4. Tested selectivity with various species including H2S (from NaHS), DTT, GSH, TCEPO, and 20 natural amino acids. Applied to E. coli samples for quantification.
3：Tested selectivity with various species including H2S (from NaHS), DTT, GSH, TCEPO, and 20 natural amino acids. Applied to E. coli samples for quantification.
List of Experimental Equipment and Materials:
3. List of Experimental Equipment and Materials: Spectrophotometer for UV-Vis and fluorescence measurements, electrospray ionization mass spectrometer (ESI-MS) for product verification, buffers (PBS, Britton-Robinson buffer), chemicals including HC-N3, TCEP, and other analytes.
4：Experimental Procedures and Operational Workflow:
Synthesized HC-N3 from HCA via a one-step reaction, confirmed with NMR and HR-MS. Measured absorption and fluorescence spectra before and after adding TCEP. Studied effects of reaction time and pH. Performed sensitivity tests with varying TCEP concentrations and selectivity tests with potential interferents. Applied probe to E. coli detection by measuring fluorescence decrease with increasing bacterial concentration.
5：Data Analysis Methods:
Analyzed fluorescence intensity at 700 nm, calculated limit of detection (LOD) using 3σ/slope method, and established calibration curves for TCEP and E. coli concentrations. Used linear regression for dynamic range analysis.
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