CN108424419B

CN108424419B - Chain double 1,2, 3-triazole rhodamine 6G fluorescent probe and preparation and application thereof

Info

Publication number: CN108424419B
Application number: CN201810279079.4A
Authority: CN
Inventors: 王宇光; 朱冰春; 叶秋娟; 王捍东
Original assignee: Zhejiang University of Technology ZJUT
Current assignee: Zhejiang University of Technology ZJUT
Priority date: 2018-03-31
Filing date: 2018-03-31
Publication date: 2020-10-27
Anticipated expiration: 2038-03-31
Also published as: CN108424419A

Abstract

The invention discloses a chain double 1,2, 3-triazole rhodamine 6G fluorescent probe and preparation and application thereof, and the probe synthesized by the invention is used for detecting Fe³⁺Has high-efficiency and specific selectivity, and can identify Fe through color change and fluorescence change³⁺Fe capable of being applied to environmental water sample³⁺Detection of (3). The probe method for measuring the concentration of the ferric ions has the characteristics of small error, high sensitivity, high accuracy and the like, and particularly shows excellent sensitivity and accuracy in low-concentration ferric ions. The probe designed and synthesized by the invention is characterized by short synthesis route of the receptor, simple and easy operation and Fe³⁺The identification effect is ideal, and the method is qualitatively applied to preliminary Fe in the environmental water sample³⁺Can also be quantitatively used for determining Fe in a water sample³⁺. It can be further applied to the Fe in the environment and biological system in the future³⁺Detection of (3).

Description

Chain double 1,2, 3-triazole rhodamine 6G fluorescent probe and preparation and application thereof

(I) technical field

The invention relates to a chain double 1,2, 3-triazole rhodamine 6G fluorescent probe and preparation and application thereof.

(II) background of the invention

Rhodamine (Rhodamine) is a catechol fluorescent dye, has a xanthene ring in structure, has a rigid plane in molecule, has good stability, has a plurality of modifiable sites, and is less interfered by a sample; the maximum fluorescence emission is located at the 500-700nm position, namely the red visible light region, the molar absorption coefficient is large, and the quantum yield is high. Because of its excellent photophysical properties and photostability, rhodamine is used in laser dyes, fluorescence scales, and stains; the surface modification of the nano polymer, the structure and dynamic research of the particle, the single molecule imaging and the biological imaging are quite widely applied.

Iron, which is an essential trace element in human body content, is present in various proteins and enzymes and is an important component of hemoglobin, heme, many enzymes, immune system compounds, and the like; participate in the transportation and storage of oxygen and directly participate in the release of energy; promoting human development; increasing resistance to disease; regulating tissue respiration, and preventing fatigue. Iron ions are not toxic in nature, but may also lead to iron poisoning if excessive amounts of iron-containing substances are ingested or misused. With the progress of the social industry, the content of iron ions in the environment is increased, and the iron ions enter human bodies through various ways and harm human health. Therefore, designing and synthesizing the probe for detecting the ferric ions has important application value.

Disclosure of the invention

The invention aims to provide a chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown as a formula (IV) and preparation and application thereof, azide alkane and rhodamine 6G are subjected to ring-forming coupling through click reaction, the alkane chain is a linker to connect rhodamine 6G at two ends, the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe is synthesized, and Fe is identified and detected by utilizing the unique 'on-off' mechanism of the probe molecule³⁺. The method has short synthetic route, is simple and easy to operate, and can be used for treating Fe³⁺Has good identification effect, and can effectively detect out the environmental water sampleMiddle Fe³⁺The content of (a).

The technical scheme adopted by the invention is as follows:

the invention provides a chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown as a formula (IV),

in the formula (IV), R is one of the following: CH (CH)₂、CH₂CH₂、CH₂CH₂CH₂CH₂、CH₂CH₂CH₂CH₂CH₂、CH₂CH₂CH₂CH₂CH₂CH₂、CH₂CH₂OCH₂CH₂。

The invention provides a preparation method of the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe, which comprises the following steps: taking a compound shown as a formula (II) and a compound shown as a formula (III) as raw materials, and taking Cu⁺As a catalyst, after the reaction is completed in tetrahydrofuran at 40-60 ℃, separating and purifying reaction liquid to obtain the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown in the formula (IV);

in the formula (III), R is one of the following: CH (CH)₂、CH₂CH₂、CH₂CH₂CH₂CH₂、CH₂CH₂CH₂CH₂CH₂、CH₂CH₂CH₂CH₂CH₂CH₂、CH₂CH₂OCH₂CH₂。

Further, the amount ratio of the compound shown in the formula (II) to the compound feeding material shown in the formula (III) is 2.2:1, the amount of the catalyst is calculated by the amount of Cu, and the amount ratio of Cu to the compound material shown in the formula (III) is 1.2: 1; the volume of the tetrahydrofuran is 44mL/mmol based on the amount of the compound substance shown in the formula (III).

Further, a method for separating and purifying reaction liquid comprises the following steps: after completion of the reaction, the reaction mixture was concentrated to dryness, followed by addition of water to dissolve it, extraction with methylene chloride (3X 50mL), combination of the organic phases, washing with saturated aqueous sodium chloride (2X 100mL), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent from the filtrate under reduced pressure, and thin layer Chromatography (CH)₃OH:CH₂Cl₂1:20, and v/v is a developing solvent), and collecting components with Rf value of 0.4-0.5 to obtain the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown in the formula (IV).

Further, the compound represented by the formula (II) is prepared by the following method: taking a compound shown in a formula (I) and propargylamine as raw materials, completely carrying out reflux reaction in methanol, and carrying out post-treatment on a reaction solution to obtain a compound shown in a formula (II); the ratio of the compound shown in the formula (I) to the propargylamine feeding substance is 1: 5; the volume usage of the methanol is 20mL/mmol based on the amount of the compound shown in the formula (I);

further, the post-treatment method of the reaction liquid comprises the following steps: after completion of the reaction, the reaction mixture was concentrated to dryness, followed by addition of water to dissolve it, extraction with methylene chloride (3X 50mL), combination of the organic phases, washing with saturated aqueous sodium chloride (2X 100mL), drying over anhydrous magnesium sulfate, filtration, evaporation of the solvent from the filtrate under reduced pressure, and thin layer Chromatography (CH)₃OH:CH₂Cl₂1:20, v/v as developing solvent), and collecting components having Rf values of 0.4 to 0.5 to obtain the compound represented by the formula (II).

The invention also provides a method for detecting Fe by using the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe³⁺The fluorescent probe can visually detect Fe in an environmental water sample³⁺。

In one aspect, the probes of the invention can qualitatively detect Fe³⁺The application is as follows: adding a sample to be detected into PBS (phosphate buffer solution) with pH value of 6.5 and 10mM and containing acetonitrile with volume concentration of 50 percent, and adding a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe acetonitrile solution with the concentration of 5 mu mol/mL, wherein the fluorescence probe acetonitrile solution generates color if the color is generatedRaw (color change can be generated within 5-50min at room temperature), and the sample to be detected contains Fe³⁺。

On the other hand, the probe of the invention can quantitatively detect Fe³⁺The application is as follows: adding a sample to be detected into PBS (phosphate buffer solution) containing acetonitrile with the volume concentration of 50% and the pH value of 6.5 and 10mM, adding a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe acetonitrile solution with the volume concentration of 5 mu mol/mL, measuring the fluorescence value at 555nm, and determining the fluorescence value according to Fe³⁺Obtaining Fe in the sample to be measured according to a standard curve³⁺Concentration; the volume ratio of the sample to be detected to the PBS buffer solution is 1:1, and the volume ratio of the sample to be detected to the probe acetonitrile solution is 10: 1; said Fe³⁺The standard curve is Fe under the same conditions³⁺The concentration of the aqueous solution is plotted on the abscissa and the fluorescence value is plotted on the ordinate.

Further, said Fe³⁺The standard curve was prepared as follows: fe with the concentration gradient of 0, 0.01,0.05,0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4.5, 5, 5.5, 6, 6.5, 7, 7.5 mu mol/mL³⁺Adding the aqueous solution into PBS buffer solution with pH 6.5 and 10mM and containing acetonitrile with volume concentration of 50 percent, adding acetonitrile solution of a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe with 5 mu mol/mL, measuring the fluorescence value at 555nm, and using Fe³⁺The concentration is taken as the abscissa and the fluorescence value is taken as the ordinate to obtain Fe³⁺A standard curve; said Fe³⁺The volume ratio of the aqueous solution to the PBS buffer solution is 1:1, and the Fe content is³⁺The volume ratio of the aqueous solution to the probe acetonitrile solution is 10: 1.

Compared with the prior art, the invention has the following beneficial effects: (1) the invention successfully designs and synthesizes a novel chain double 1,2, 3-triazole rhodamine 6G probe shown in a formula (IV); (2) probe pair Fe synthesized by the invention³⁺Has high-efficiency and specific selectivity, and can identify Fe through color change and fluorescence change³⁺. (3) The invention relates to the identification capability of a chain double 1,2, 3-triazole rhodamine 6G derivative probe on metal ions and the identification capability of the probe on Fe by the different chain lengths of the probe³⁺Whether the capability of the compound has influence or not is compared through experiments, the structure-activity relationship of the compound is analyzed, and the result shows that the alkane chain of the five carbon chain can provide the trivalentThe optimal spatial position for iron ions to complex with the probe and the presence of oxygen ions in the alkane chain can affect the effect. (4) The probe synthesized by the invention can be applied to Fe in an environmental water sample³⁺Detection of (3). (5) The probe method for measuring the concentration of the ferric ions has the characteristics of small error, high sensitivity, high accuracy and the like, and particularly shows excellent sensitivity and accuracy in the detection of the ferric ions with low concentration. The probe designed and synthesized by the invention is characterized by short synthetic route, simple and easy operation, and can be used for treating Fe³⁺The identification effect is ideal, and the method is qualitatively applied to preliminary Fe in the environmental water sample³⁺Can also be quantitatively used for determining Fe in a water sample³⁺. It can be further applied to the Fe in the environment and biological system in the future³⁺Detection of (3).

(IV) description of the drawings

FIG. 1 shows the fluorescent probe compounds IV-4 and Fe in example 9 of the present invention³⁺The color of the solution before and after the action is changed (A: under fluorescence; B: visible with naked eyes, wherein the color of the solution of the fluorescent probe compound IV-4 is changed on the left side, and the color of the solution of the fluorescent probe compound IV-4 and the color of the compound IV-4 and Fe are changed on the right side³⁺Change in color of the solution after the action).

FIG. 2 is an ultraviolet absorption spectrum of the fluorescent probe compound IV-4 of example 9 of the present invention with respect to each metal ion, wherein a is an ultraviolet-visible absorption spectrum, the abscissa is the wavelength (nm), and the ordinate is the ultraviolet absorption OD value. b is a column diagram of the ultraviolet absorption value at 555nm of each metal ion.

FIG. 3 is a fluorescence intensity spectrum of each metal ion by the fluorescent probe compound IV-4 in example 9 of the present invention. a is a fluorescence intensity spectrum, the abscissa is the wavelength (nm), and the ordinate is the fluorescence intensity. b is a bar graph of fluorescence intensity of each metal ion.

FIG. 4 shows the fluorescence intensity of the fluorescent probe compound IV-4 of the present invention and IV-4 + Fe³⁺Fluorescence emission patterns (555nm) of the ionic complexes respectively with changes in pH. The abscissa is pH and the ordinate is fluorescence intensity.

FIG. 5 shows fluorescent probe compounds IV 1-6 vs. Fe in example 8 of the present invention³⁺Fluorescence emission map of fluorescence intensity variation for identification ability. The abscissa is the wavelength (nm) and the ordinate is the fluorescence intensity.

FIG. 6 shows the fluorescent probe compound IV-4 + Fe in example 11 of the present invention³⁺Fluorescence emission patterns (555nm) of the ion complexes respectively with the change of the fluorescence intensity with time. The abscissa is time (min) and the ordinate is fluorescence intensity.

FIG. 7 shows the fluorescent probe compounds IV-4 + metal ion complexes (IV-4 + metals) and IV-4 + Fe in example 12 of the present invention³⁺+ metal ion complex (IV-4 + metalions + Fe³⁺) The fluorescence intensity of (5) was compared with a histogram (555 nm). The abscissa is the metal ion and the ordinate is the fluorescence intensity.

FIG. 8 shows the fluorescent probe compound IV-4 and different concentrations of Fe in example 13 of the present invention³⁺(0-30 times) fluorescence intensity Change Pattern (a) the abscissa is wavelength (nm) and the ordinate is fluorescence intensity and Fe³⁺Linear graph (b) of the change in concentration of (a) and fluorescence intensity.

FIG. 9 shows the fluorescent probe compound IV-4 + Fe in example 13 of the present invention³⁺Job's plot of ionic complex with abscissa [ Fe ]³⁺]/[Fe³⁺]+[Ⅳ-4]And the ordinate represents the ultraviolet absorption OD value.

FIG. 10 shows the fluorescent probe compounds IV-4 and Fe in example 15 of the present invention³⁺The fluorescence intensity change pattern of the reversibility test (2). The abscissa is the wavelength (nm) and the ordinate is the fluorescence intensity.

FIG. 11 shows the fluorescent probe compounds IV-4 and Fe in example 15 of the present invention³⁺The recognition mechanism diagram of (1).

FIG. 12 shows that the fluorescent probe compound IV-4 in example 16 of the present invention recognizes Fe in different concentrations in an environmental water sample³⁺The solution color of (1) can be seen in a change diagram, wherein Fe is from left to right³⁺The concentration of the solution is 0, 0.05, 5.0 and 10.0mM in sequence.

(V) detailed description of the preferred embodiments

The ultrapure water is distilled water obtained by once distilling deionized water, and the room temperature is 25-30 ℃. The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:

example 1: synthesis of Compound (II)

6g (compound I) (1.25mmol, 1.0eq, 0.6g) of rhodamine is dissolved in 25mL of methanol reagent (analytical grade), the solution is reddish brown after complete dissolution, propargylamine (6.25mmol, 5.0eq, 0.34g) is added dropwise, after the dropwise addition is finished, the heating reflux reaction is carried out for 24h, and the reaction process is tracked and detected by TLC. After completion of the reaction, the solvent was evaporated by a rotary evaporator, then 30mL of water and methylene chloride were added to conduct extraction (3X 50mL), the organic phases were combined, washed with a saturated aqueous solution of sodium chloride (2X 100mL), dried over anhydrous magnesium sulfate, filtered, the filtrate was evaporated under reduced pressure to dryness, and then thin layer Chromatography (CH)₃OH:CH₂Cl₂1:20, v/v as developing agent), fractions with Rf of 0.4-0.5 were collected and dried to give compound II.

Compound II (pink solid, 65% yield) MS (ESI) with M/z 452.2[ M + H ]]⁺.mp＝259-261℃.¹H NMR(600MHz,CDCl₃)8.02–7.88(m,1H),7.48–7.37(m,2H),7.11–7.02(m,1H),6.36(s,2H),6.28(s,2H),3.93(d,J＝2.5Hz,2H),3.53(dd,J＝45.2,5.3Hz,2H),3.29–3.14(m,4H),1.90(s,6H),1.81–1.73(m,1H),1.32(t,J＝7.2Hz,6H).¹³C NMR(150MHz,CDCl₃)167.63,154.02,151.86,147.50,132.71,130.25,128.81,128.01,123.76,123.08,117.73,105.46,96.60,78.37,77.28,77.03,76.78,70.05,64.99,38.40,28.65,16.66,14.76.IRν_max(cm^-1):3442.53,3278.00,2964.43,2929.15,2870.13,1704.041621.33,1517.19,1446.13,1383.59,1158.70,938.78,811.98,664.70.

Example 2: synthesis of Compound (IV-1)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-1(1.0eq) was added as Cu⁺As catalyst (addition of copper sulfate pentahydrate (1.2)eq) and VC (1.2eq) to Cu⁺(1.2 eq). The reaction is carried out for 8-12h at about 55 ℃, and the reaction process is tracked and detected by TLC. After completion of the reaction, the reaction mixture was concentrated to dryness, then 30mL of water was added, methylene chloride was extracted (3X 50mL), the organic phases were combined, washed with a saturated aqueous solution of sodium chloride (2X 100mL), dried over anhydrous magnesium sulfate, filtered, the filtrate was evaporated to dryness under reduced pressure to remove the solvent, and subjected to thin layer Chromatography (CH)₃OH:CH₂Cl₂1:20, v/v is developing agent), collecting components with Rf of 0.4-0.5, and drying to obtain the target product IV-1.

Compound IV-1 (white solid, yield 35%) MS (ESI) with M/z 1001.5[ M + H ═]⁺.mp＝259-261℃.¹H NMR(600MHz,CDCl₃)8.07–7.94(m,2H),7.55–7.33(m,4H),7.17(s,1H),7.14–6.98(m,2H),6.94–6.86(m,1H),6.32(s,1H),6.28(d,J＝6.0Hz,1H),6.18–6.06(m,2H),5.86(s,1H),5.79(d,J＝11.4Hz,1H),5.32(d,J＝17.7Hz,2H),4.56(s,1H),4.47(s,1H),3.28–3.12(m,6H),2.97(t,J＝16.0Hz,2H),1.88–1.80(m,6H),1.61(s,12H),1.31(t,J＝7.1,4.8Hz,6H),1.23(dd,J＝13.7,6.5Hz,6H).¹³C NMR(150MHz,CDCl₃)168.10,153.82,152.70,151.73,147.28,144.10,132.65,130.62,128.43,128.03,123.80,123.62,122.92,117.60,115.85,105.58,96.60,77.24,77.03,76.82,66.95,65.15,49.14,38.35,35.14,29.70,16.64,16.56,14.75.IRν_max(cm^-1):3440.66,2968.25,1621.43,1517.46,1421.56,1269.83,1216.44,1093.80,877.48,700.58,655.77.

Example 3: synthesis of Compound (IV-2)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-2(1.0eq) was added as Cu⁺Cu was formed as a catalyst (copper sulfate pentahydrate (1.2eq) and VC (1.2eq) were added⁺). Otherwise, the same procedure as in example 2 was repeated to give a target product IV-2.

Compound IV-2 (white solid, yield 40%) MS (ESI) M/z 1015.5[ M + H ═]⁺.¹H NMR(600MHz,CDCl₃)7.94(dd,J＝9.6,3.7Hz,2H),7.52–7.41(m,4H),7.27(d,J＝6.4Hz,1H),7.04(t,J＝5.7Hz,2H),6.70(d,J＝6.2Hz,2H),6.30(d,J＝6.0Hz,4H),6.08(d,J＝5.6Hz,4H),4.42(d,J＝5.9Hz,2H),4.32(d,J＝6.1Hz,2H),3.22–3.12(m,8H),1.83(d,J＝5.7Hz,12H),1.28(dd,J＝13.4,6.9Hz,15H).¹³C NMR(150MHz,CDCl₃)167.63,154.01,151.83,147.49,132.73,130.22,128.79,128.01,123.75,123.07,117.73,105.40,96.58,78.35,77.26,77.05,76.84,70.06,64.98,38.39,29.71,28.64,16.68,14.76.IRν_max(cm^-1):3397.93,2967.70,1689.81,1620.99,1518.51,1384.09,1270.71,1217.46,1141.17,1092.13,1014.56,875.81,738.93,689.08,540.26.

Example 4: synthesis of Compound (IV-3)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-3(1.0eq) was added as Cu⁺Cu was formed as a catalyst (copper sulfate pentahydrate (1.2eq) and VC (1.2eq) were added⁺). Otherwise, the same procedure as in example 2 was repeated to give a target product IV-3.

Compound IV-3 (light yellow solid, 40% yield) MS (ESI) M/z 1043.3[ M + H ═]⁺.

¹H NMR(600MHz,CDCl₃)7.98–7.92(m,2H),7.49–7.42(m,4H),7.09–7.03(m,2H),7.01(d,J＝21.3Hz,2H),6.34(d,J＝21.1Hz,4H),6.06(d,J＝31.8Hz,4H),4.44(s,4H),4.15–4.07(m,4H),3.29(t,J＝6.6Hz,4H),3.20(dd,J＝7.2,4.0Hz,4H),1.83(s,12H),1.51(dt,J＝10.1,6.6Hz,4H),1.34–1.30(m,12H).¹³C NMR(150MHz,CDCl₃)168.07,153.67,151.84,147.32,144.33,132.64,130.76,128.46,128.07,123.85,122.93,122.17,117.56,105.68,96.63,77.25,77.04,76.82,65.18,50.67,49.11,38.36,35.27,27.26,25.83,16.61,14.75.IRν_max(cm^-1):3427.73,2968.69,269.42,2097.17,1620.87,1518.23,1421.34,1384.02,1269.56,1158.01,1091.99,1014.16,920.76,774.84.

Example 5: synthesis of Compound (IV-4)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-4(1.0eq) was added as Cu⁺Cu was formed as a catalyst (copper sulfate pentahydrate (1.2eq) and VC (1.2eq) were added⁺). Otherwise, the same procedure as in example 2 was repeated to give a target product IV-4.

Compound IV-4 (white solid, 45% yield) MS (ESI) M/z 1057.5[ M + H ═]⁺.¹H NMR(600MHz,CDCl₃)7.95(dd,J＝6.2,2.0Hz,2H),7.50–7.40(m,4H),7.10–6.97(m,4H),6.32(s,4H),6.09(s,4H),4.44(s,4H),4.15–3.97(m,4H),3.96–3.80(m,4H),3.75(dd,J＝11.2,5.4Hz,4H),3.31–3.09(m,12H),1.86–1.78(m,16H),1.40–1.25(m,14H).¹³C NMR(150MHz,CDCl₃)168.10,153.70,151.81,147.30,144.22,132.65,130.72,128.45,128.06,123.84,122.92,122.18,117.54,105.65,104.09,96.62,77.24,77.03,76.82,67.11,66.04,65.18,61.68,51.05,49.49,38.36,35.27,32.43,32.02,29.62,28.24,23.68,23.51,16.63,14.75.IRν_max(cm^-1):3428.86,2966.31,2095.48,1681.38,1620.92,1518.05,1420.44,1384.36,1269.70,1201.42,1092.26,877.09,771.54.

Example 6: synthesis of Compound (IV-5)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-5(1.0eq) was added as Cu⁺Cu was formed as a catalyst (copper sulfate pentahydrate (1.2eq) and VC (1.2eq) were added⁺). Otherwise, the same procedure as in example 2 was repeated to give a target product IV-5.

Compound IV-5 (Pink solid, 40% yield) MS (ESI) M/z 1071.5[ M + H ═]⁺.mp＝253-255℃.¹H NMR(600MHz,CDCl₃)8.00–7.92(m,2H),7.51–7.40(m,4H),7.09–6.95(m,4H),6.32(s,4H),6.14–6.04(m,4H),4.44(s,4H),4.08(dd,J＝16.9,9.5Hz,4H),3.25(t,J＝6.8Hz,4H),3.26–3.14(m,8H),1.88–1.79(m,12H),1.77–1.67(m,4H),1.59–1.52(m,4H),1.35–1.28(m,12H).¹³C NMR(150MHz,CDCl₃)168.09,153.71,151.80,147.30,144.16,132.64,130.71,128.45,128.06,123.83,122.91,122.14,117.55,105.62,104.09,96.60,77.26,77.05,76.84,67.10,66.00,65.18,61.62,51.23,49.61,38.35,35.26,32.42,32.03,29.94,29.32,28.65,26.13,26.06,23.51,16.63,14.75.IRν_max(cm^-1):3440.45,2931.24,2094.83,1621.36,1517.74,1420.61,1269.67,1200.83,1092.00,1013.83,876.99,812.60,743.93.

EXAMPLE 7 Synthesis of Compound (IV-6)

Compound II (0.1mmol, 2.2eq, 0.04g) was dissolved in 20mL of anhydrous tetrahydrofuran, and Compound III-6(1.0eq) was added as Cu⁺Cu was formed as a catalyst (copper sulfate pentahydrate (1.2eq) and VC (1.2eq) were added⁺). Otherwise, the same procedure as in example 2 was repeated to give a target product IV-6.

Compound IV-6 (white solid, yield 35%) MS (ESI) M/z 1059.5[ M + H ═]⁺.mp＝263-266℃.¹H NMR(600MHz,CDCl₃)7.93(dd,J＝19.3,7.1Hz,2H),7.41–7.29(m,4H),6.77(d,J＝7.6Hz,2H),5.93(s,7H),4.27–4.17(m,4H),3.99–3.80(m,7H),3.79–3.52(m,8H),3.26–3.04(m,8H),2.05–1.79(m,12H),1.39–1.26(m,12H).¹³C NMR(150MHz,CDCl₃)166.58,152.96,150.77,146.43,131.69,129.17,127.76,126.97,122.71,122.03,116.68,104.34,95.51,77.30,76.20,75.99,75.78,69.02,63.91,37.35,28.68,27.60,15.65,13.72.IRν_max(cm^-1):3397.93,2967.70,1689.81,1620.99,1518.51,1384.09,1270.71,1217.46,1141.17,1092.13,1014.56,875.81,738.93,689.08,540.26.

EXAMPLE 8 Metal ion Selectivity of probes IV-1 to IV-6

(1) Preparation of Probe stock solution

The probe molecule powders (IV-1) to (IV-6) prepared in examples 2 to 7 were accurately weighed and dissolved in chromatographic grade acetonitrile to prepare a probe acetonitrile solution of 5. mu. mol/mL. Storing in dark at low temperature.

(2) Preparation of metal ion mother liquor

Weighing inorganic salt MgSO₄·7H₂O、KCl、CuSO₄·5H₂O、FeCl₃、FeSO₄·7H₂O、MnSO₄·H₂O、Al(NO₃)₃·9H₂O、CaCl₂、NaCl、AgNO₃、Pb(CH₃COO)₂·3H₂O、Co(NO₃)₂·6H₂O、BaCl₂·2H₂O、Zn(CH₃COO)₂·2H₂O、NiSO₄·6H₂O、CrCl₃·6H₂O、LiCl₂·H₂O、HgCl₂、RuCl₃Transferring into a 10mL centrifuge tube, and diluting with ultrapure water to 8mL to obtain metal ion mother liquor with concentration of 1.25mmol/mL, wherein the corresponding ions are Mg²⁺、K⁺、Cu²⁺、Fe³⁺、Fe²⁺、Mn²⁺、Al³⁺、Ca²⁺、Na⁺、Ag⁺、Pb²⁺、Co²⁺、Ba²⁺、Zn²⁺、Ni²⁺、Cr³⁺、Li²⁺、Hg²⁺、Ru²⁺. Storing in dark at low temperature.

(3) Metal ion selectivity of probes IV-1 to IV-6

The probe molecules IV-1 to IV-6 were assayed for Fe in 10mM PBS buffer at pH 6.5 containing acetonitrile 50% by volume at room temperature³⁺Selectivity of (2).

After diluting the metal ion mother liquor of 1.25mmol/mL in step (2) with ultrapure water to 12.5. mu. mol/mL, 100. mu.L of the metal ion mother liquor was taken, and 100. mu.L of 10mM PBS buffer solution at pH 6.5 containing acetonitrile of 50% by volume was added, and then 10. mu.L of the 5. mu. mol/mL probe acetonitrile solutions (IV-1 to IV-6) in step (1) were added, respectively, and the fluorescence spectra thereof were measured, and the results are shown in FIG. 5.

FIG. 5 shows that in the series of probes of compounds IV-1 to IV-6, the compound IV-4 is coupled with Fe³⁺Has the strongest identification ability, and the compounds IV-2 and IV-6 are opposite to Fe³⁺Is the weakest recognition ability, which may beBecause the five-carbon chain alkane chain can provide the best space position for the ferric ion to complex with the probe under the condition of different chain lengths, and the alkane chain can influence the identification effect if the oxygen ion exists.

EXAMPLE 9 Metal ion selectivity of Probe IV-4

The selectivity of the iv-4 probe molecule for metal ions was determined in 10mM PBS buffer at room temperature at pH 6.5 containing 50% acetonitrile by volume.

Adding 12.5 mu mol/mL Fe³⁺After adding 100. mu.L of an aqueous solution containing 50% acetonitrile by volume at pH 6.5 and 100. mu.L of 10mM PBS buffer, and further adding 10. mu.L of 5. mu. mol/mL acetonitrile (IV-4) probe, the change in the corresponding fluorescence property was measured. Under the same conditions, adding Fe³⁺Respectively replaced by Al³⁺、Mg²⁺、K⁺、Cu²⁺、Fe³⁺、Fe²⁺、Mn²⁺、Ca²⁺、Na⁺、Ag⁺、Pb²⁺、Co²⁺、Ba² ⁺、Zn²⁺、Ni²⁺、Cr³⁺、Li²⁺、Hg²⁺、Ru²⁺After the reaction, the color of the solution changes, and the ultraviolet absorption and fluorescence intensity change, so as to obtain corresponding ultraviolet absorption spectrum and fluorescence emission spectrum, which are respectively shown in fig. 1, fig. 2 and fig. 3.

As can be seen from FIG. 1, the compound IV-4 is in Fe³⁺No color before, adding Fe³⁺Later, the solution of the compound IV-4 turns red, the color of the solution under fluorescence becomes yellow, and the experiment proves that the Fe is added³⁺The other metal ions are not colored. From FIGS. 2 and 3, it can be seen that the compound IV-4 is against Fe³⁺The selectivity is good, the ultraviolet absorption spectrum of the compound has almost no absorption peak above 530nm, and the fluorescence emission of the compound IV-4 is weak; the UV-visible spectrum shows a strong absorption peak at 530nm, and the absorption intensity is increased by about 4.5 times. According to the ultraviolet absorption spectrum, when the fluorescence spectrum is measured, 490nm is selected as the excitation wavelength, and the compounds IV-4 and Fe³⁺The strong fluorescence emission wavelength is at 555nm, and the fluorescence intensity is greatly enhanced, which proves that the lactam ring of the compound is opened. It is worth mentioning only thatWhen Al is added in the same times³⁺After that, some ultraviolet absorption enhancement and fluorescence enhancement were caused, but under the same conditions, Al was present³⁺The induced fluorescence enhancement is much lower than that of Fe³⁺The ultraviolet absorption and fluorescence enhancement are caused, which shows that the compound IV-4 is used for Fe³⁺The selectivity of (A) is less interfered by other coexisting ions, and the Fe is shown to be influenced³⁺High selectivity of the process.

Example 10 Effect of pH on the recognition Performance of Probe IV-4

In order to be able to apply probe IV-4 in a more complex system, pH vs. Fe was investigated during the experiment³⁺An impact of performance is identified. At room temperature, probe IV-4 and probe IV-4 + Fe were measured using a PBS solution containing acetonitrile at a volume concentration of 50% at a pH in the range of 3.5 to 12.0³⁺Change in fluorescence intensity of (a).

Adding 12.5 mu mol/mL Fe³⁺mu.L of the aqueous solution was added to 100. mu.L of PBS buffer containing acetonitrile at a volume concentration of 50% at different pH values (3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, 10.0, 10.5, 11.0, 11.5, and 12.0), 10. mu.L of a probe acetonitrile solution (IV-4) at 5. mu. mol/mL was added thereto, and the change in the corresponding fluorescence value was measured at 555 nm. Under the same conditions, Fe is not added³⁺Aqueous solution was used as control. The results are shown in FIG. 4.

FIG. 4 shows that the solution of probe IV-4 is Fe-free in 10mM PBS buffer at pH 6.5 containing acetonitrile 50% by volume³⁺In the presence and presence of Fe³⁺When the fluorescence intensity changes, the change of the fluorescence intensity at the position of 555nm of emission wavelength is recorded by using 490nm as the excitation wavelength under the condition of different pH values. As can be seen from the figure, there is no Fe³⁺When present, no significant change in fluorescence was observed between pH3.5 and 11.5, indicating that probe IV-4 itself is not pH sensitive. When Fe is added³⁺Then, in the same pH range, probe is directed to Fe³⁺Response was different when pH was<The fluorescence intensity was strong at 7.0 and was strongest at pH 5.5. When the solution is alkaline, Fe³⁺The binding ability with probe IV-4 was reduced and no more fluorescence was evident. Overall, probe IV-4 can achieve the effect on Fe over a wide pH range (pH3.5-9)³⁺Efficient identification of (1). In the following performance tests, 10mM PBS buffer solution with a pH of 6.5 and a volume concentration of 50% acetonitrile was selected as a solvent system, taking into account the environment in which other ions acted during the experiment.

EXAMPLE 11 Probe Compound IV-4 vs Fe³⁺Time response of

Probe molecule IV-4 vs Fe was assayed in 10mM PBS buffer pH 6.5 containing 50% acetonitrile by volume at room temperature³⁺The identification performance of (1).

Mixing 12.5 mu mol/mLFe³⁺The length of action time was determined by adding 100. mu.L of an aqueous solution, 100. mu.L of 10mM PBS buffer solution containing 50% acetonitrile by volume at pH 6.5, and 10. mu.L of 5. mu. mol/mL probe acetonitrile solution (IV-4) to the solution, and measuring the change in the corresponding fluorescence value at 555nm every 5min for 120 min. The results are shown in FIG. 6.

As can be seen from FIG. 6, the compounds IV-4 and Fe were present within 5min³⁺The reaction can generate fluorescence, which shows that the probe has good sensitivity, the fluorescence intensity generated along with the increase of time is enhanced, the maximum fluorescence intensity is reached after 45min, and the fluorescence intensity is stable and does not obviously decrease along with the increase of time. This indicates that the probe IV-4 is for Fe³⁺Has fast response and long-time identification stability. Such real-time monitoring is of great significance in practical applications. Therefore, in the subsequent test work, each sample is subjected to spectrometry after being placed for 30-40min by adding metal ions.

EXAMPLE 12 Probe Compounds IV-4 vs Fe³⁺Interference immunity experiment of

Probe molecules IV-4 vs Fe in 10mM PBS buffer B at pH 6.5 with acetonitrile 50% by volume at room temperature³⁺The identification performance of (1).

Separately, 1.25mmol/mL of a metal ion mother liquor (Mg) obtained in step (2) of example 8²⁺、Na⁺、Cu²⁺、K⁺、Al³⁺、Fe²⁺、Ca²⁺、Ag⁺、Ba²⁺、Co²⁺、Pb²⁺、Zn²⁺、Mn²⁺、Ni²⁺、Cr³⁺、Fe³⁺、Li⁺、Ru²⁺、Hg²⁺) After diluting the mixture with ultrapure water to 12.5. mu. mol/mL, 100. mu.L of the mixture was taken, and added with 50% acetonitrile by volume at pH 6.5 and 100. mu.L of 10mM PBS buffer, and then 10. mu.L of 5. mu. mol/mL probe acetonitrile solution (IV-4), and fluorescence was measured at 555nm using no metal ion as a blank, as shown in FIG. 7, which is a gray column.

Adding 12.5 mu mol/m L Fe into each metal ion solution³⁺The fluorescence value of the aqueous solution is measured at 555nm by 100 mu L, and is shown as a black column in figure 7, and the comparison of the fluorescence intensity of two measurements shows that the existence of ions of the aqueous solution is opposite to that of Fe³⁺The influence of (c).

As shown in FIG. 7, the grey bar graph illustrates that ions other than iron do not cause a strong change in fluorescence emission, but the same equivalent of Fe was added to the solution above³⁺After the solution, a sharp increase in fluorescence intensity (black column) was observed. The results show that the compound IV-4 is relative to Fe³⁺Is less interfered by other coexisting ions, and thus, it was confirmed that IV-4 is responsible for Fe³⁺Has good selectivity.

EXAMPLE 13 probes IV-4 vs Fe³⁺Fluorescence titration and determination of the complexation ratio of

1) Probe IV-4 pairs of Fe³⁺Titration and fitting of

To a 10mM PBS buffer solution containing 50% acetonitrile by volume at pH 6.5 at room temperature, 10. mu.L of a 5. mu. mol/mL probe acetonitrile solution (IV-4) was added to carry out Fe³⁺Titration experiment of (1), i.e. titrating 5. mu. mol/mL Fe thereto³⁺In the aqueous solution, the fluorescence emission spectrum is measured once after each 10. mu.L of the measurement solution is added, the titration is finished after the 10. mu.L titration for the 30 th time, and the total 31 times of measurement (namely, Fe is carried out on the probe (IV-4)) are carried out³⁺0-30.0 times the amount of the titration experiment). The results are shown in FIG. 8.

As can be seen from FIG. 8, with Fe³⁺The fluorescence emission intensity at 555nm is gradually enhanced when the concentration is increased, and when the concentration is added to about 15.0 times of Fe³⁺After that, the reaction was substantially saturated and the increase in fluorescence intensity of the solution was insignificant. By fluorescence intensity with Fe³⁺The change rule of the concentration can be calculated to complexBinding constant of substance, assuming Fe³⁺The binding ratio of the compound IV-4 to the compound IV-4 is 1:1, and the fluorescence intensity of the compound IV-4 is related to Fe by Origin software according to the following equation³⁺The concentration variation graph of (2) is fitted by a nonlinear least square method to obtain a smooth curve (b in figure 8), the R value of a linear correlation coefficient is more than 0.99, and the Fe is proved to be true³⁺The binding ratio with the compound IV-4 was 1:1, and the binding constant value was calculated to be 2.25X 10⁵M^-1The binding constant is so large that the complex has better stability.

In the formula (1), Y represents the fluorescence intensity after the addition of ions; y is₀Represents the fluorescence intensity of the organic compound; y is_limA limit value representing a change in fluorescence intensity after addition of an ion; c_MRepresents the concentration of the added metal ions; c_LRepresents the concentration of the organic compound; ks is the binding constant.

2) Probe IV-4 and Fe³⁺Determination of binding ratio

To further illustrate IV-4 and Fe³⁺Is 1:1 bonded according to Fe³⁺The increase in the molar fraction of (A) and the change in the ultraviolet absorption value were plotted in Job's plot (FIG. 9). As can be seen from the figure, IV-4 is associated with Fe³⁺The total concentration of (D) is 100. mu.M, when Fe³⁺At a molar fraction of 0.5, the fluorescence emission intensity reaches a maximum, which indicates 4d vs Fe³⁺Is 1:1, consistent with the above non-linear fit.

Example 14Fe³⁺Determination of regression equation of concentration and probe IV-4 fluorescence intensity variation relation

In order to develop the practical value of the probe, Fe was carried out³⁺The experimental determination of the regression equation of the relationship between the concentration and the fluorescence intensity change of the probe IV-4 comprises the following specific experimental methods: 1.25mmol/mL Fe in example 8³⁺Diluting the ion mother liquor with ultrapure water to make its concentration respectively 0, 0.01,0.05,0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4.5, 55.5, 6, 6.5, 7, 7.5. mu. mol/mL, 100. mu.L of each sample was added to a solution containing 50% acetonitrile by volume at pH 6.5 and 10mM PBS buffer, 10. mu.L of a 5. mu. mol/mL probe acetonitrile solution (IV-4) was further added, and then the fluorescence intensity was measured at 555 nm. According to the measured Fe³⁺The relationship between the ion concentration and the corresponding fluorescence intensity data to obtain the concentration of Fe³⁺The regression equation y with the concentration (. mu. mol/mL) as independent variable x and the fluorescence intensity as dependent variable y is 251.625+2526.47 x.

EXAMPLE 15 probes IV-4 vs Fe³⁺Determination of the reversibility of recognition of

pH 6.5 with acetonitrile 50% by volume, 100. mu.L of 10mM PBS buffer, 12.5. mu. mol/mLFe at room temperature³⁺After adding 10. mu.L of a 5. mu. mol/mL acetonitrile solution of the probe IV-4 to 100. mu.L of the aqueous solution, the change in the reaction fluorescence was observed and the fluorescence spectrum was measured, and the result is shown in curve a in FIG. 10. The excess ETDA complexing agent was added, the change in the fluorescence of the reaction was observed, and the fluorescence spectrum was measured again, the result being shown in curve b in FIG. 10. Finally, 12.5 mu mol/mL of Fe is added into the mixture³⁺The change of the fluorescence of the reaction was observed in 100. mu.L of the aqueous solution, and the fluorescence spectrum was measured, and the result was shown in curve c in FIG. 10.

After the compound IV-4 reacts with the ferric ion solution, the solution changes from colorless to pink, the fluorescence intensity value at this time is measured, after the excessive EDTA is added into the solution, the solution becomes light, and the fluorescence intensity value at this time is measured again. Then adding the same equivalent of Fe³⁺The colour of the latter solution was recovered but was weaker than the first probably due to the presence of excess EDTA in the solution, which as a complex also reacts with Fe³⁺Complexing, thereby affecting Fe³⁺Complexation with Probe IV-4 and re-measurement of fluorescence. This illustrates IV-4 and Fe³⁺There is coordination between them, and Fe³⁺The complex constant between the Fe and the IV-4 is larger, the coordination is stronger, and Fe is added³⁺The lactam ring in IV-4 is opened and the addition of EDTA can convert Fe³⁺Elimination from the probe system, experiments could show that probe IV-4 is for Fe³⁺Is reversible.

According to IV-4 with Fe³⁺The binding ratio between the two is 1:1, and a reversibility experiment judges the type of probesFor Fe³⁺The mechanism of recognition of (2) is shown in FIG. 11.

Example 16 Probe IV-4 d for qualitative determination of Fe in Water samples³⁺Application of

After the probe IV-4 is subjected to a spectrum detection experiment, the probe IV-4 is preliminarily applied to metal ion Fe in an environmental water sample³⁺In the detection, taking Fe with different concentration gradients³⁺Aqueous solution of Fe of different concentrations in water sample³⁺Visual different color changes can be generated after the probe acts on the Fe-Fe color change detection probe, and the qualitative detection of Fe in a water sample is carried out³⁺In (1). The implementation method comprises the following steps:

respectively taking 4 parts of 0.2mL of Fe with different concentration gradients³⁺An aqueous solution (0, 0.05, 5.0, 10.0. mu.M) was sampled, 200. mu.L of 10mM PBS buffer containing 50% acetonitrile by volume at pH 6.5 was added, and 0.02mL of 5. mu. mol/mL probe IV-4 acetonitrile was added, and the color change was shown in FIG. 12.

FIG. 12 shows the concentration gradients of 0, 0.05, 5.0 and 10.0. mu.M Fe from left to right³⁺The color of the solution gradually changes from colorless to red.

Example 17 Probe IV-4 for quantitative determination of Fe in Water samples³⁺Application of concentration

In order to verify that the probe IV-4 detects Fe in a water sample³⁺The application properties of (1) were 7.5, 6.0, 4.0, 2.0, 1.0, 0.5, 0.1, 0.05, 0.01, 0.005 and 0.001. mu. mol/mL of Fe were prepared using ultrapure water³⁺And (5) using an aqueous solution as a sample to be detected. The Fe is measured by adopting a probe method and a phenanthroline method³⁺The concentration of the solution is specifically operated as follows:

(1) determination of Fe by the Probe method of the present invention³⁺The concentration of the solution was measured by adding 100. mu.L of 10mM PBS buffer with pH 6.5 containing acetonitrile 50% by volume to 100. mu.L of Fe prepared as described above at different concentrations³⁺After 10. mu.L of a 5. mu. mol/mL acetonitrile probe solution (IV-4) was added to the aqueous solution, the fluorescence intensity was measured at 555 nm. Fe was obtained by substituting the measured fluorescence value into the regression equation y of 251.625+2526.47x obtained in example 14³⁺Concentrations, see table 1.

(2) Method for measuring Fe by phenanthroline method³⁺Concentration of the solution

a. Preparation of acetic acid-sodium acetate buffer solution (pH 4.5): 164g of sodium acetate was weighed, dissolved in 500mL of ultrapure water, added with 84mL of glacial acetic acid, and diluted to 1000mL with ultrapure water.

b. Preparation of 20g/L ascorbic acid solution: 10.0g of ascorbic acid was dissolved in 200mL of ultrapure water, 0.2g of disodium Ethylenediaminetetraacetate (EDTA) and 8.0mL of formic acid were added, and then diluted to 500mL with ultrapure water, shaken well and stored in a brown bottle.

c. Preparing a 2.0g/L phenanthroline solution: weighing 2.0g of phenanthroline, dissolving in 800mL of ultrapure water, and diluting to 1000mL by using ultrapure water.

d. Preparation of 40.0g/L potassium persulfate solution: 4.0g of potassium persulfate was weighed out, dissolved in ultrapure water and dissolved to 100 mL.

e. Preparation of 0.1mg/mL ferric ammonium sulfate standard solution I: 0.863g of ammonium ferric sulfate is weighed and placed in a 200mL beaker, 100mL of ultrapure water and 10mL of concentrated sulfuric acid are added, and dissolved to 1000 mL.

f. Preparation of 0.01mg/mL ferric ammonium sulfate standard solution II: 1mL of 0.1mg/mL ferric ammonium sulfate standard solution I is diluted by 10 times and is only used for the same day.

g. Drawing a working curve: respectively putting 0mL (blank), 1.00mL, 2.00mL, 4.00mL, 6.00mL, 8.00mL and 10.00mL of the ammonium ferric sulfate standard solution II prepared in the step f into 7 100mL volumetric flasks, adding ultrapure water to about 40mL, adding 0.50mL of sulfuric acid solution (the volume ratio of water to 98% concentrated sulfuric acid with mass concentration is 1: 35), adjusting the pH value to 2, adding 3.0mL of 20g/L of ascorbic acid solution prepared in the step b, 10mL of acetic acid-sodium acetate buffer solution prepared in the step a and 5mL of phenanthroline solution prepared in the step c. Diluting with ultrapure water to scale, and shaking up. The mixture was allowed to stand at room temperature for 15 minutes, and absorbance was measured by spectrophotometry at 510nm with a reagent blank adjusted to zero. Taking the measured absorbance as a vertical coordinate, corresponding Fe³⁺The amount (. mu. mol) is plotted on the abscissa and the working curve y is 0.00064+ 0.37936X.

h. Determination of the ferric ion concentration: taking 8mL of a sample to be detected into a 100mL volumetric flask, adding ultrapure water to about 40mL, adjusting the pH value to 2 (which can be adjusted by ammonium water if necessary) by using a sulfuric acid solution (the volume ratio of water to 98% concentrated sulfuric acid with mass concentration is 1: 35), adding 3.0mL of 20g/L ascorbic acid solution prepared in the step b, 10mL of acetic acid-sodium acetate buffer solution prepared in the step a and 5mL of phenanthroline solution prepared in the step c. Diluting with ultrapure water to scale, and shaking up. The mixture was allowed to stand at room temperature for 15 minutes, and absorbance was measured at 510nm with a spectrophotometer with a reagent blank adjusted to zero, and the results are shown in Table 1.

The concentration C of iron ions in. mu. mol/mL was calculated as follows:

C＝m/(55.8434*V)

m is the amount of iron ions expressed in μ g;

v: sample volume in mL.

TABLE 1 Probe IV-4 Fe in test Water sample³⁺Application of

From the above table 1, the detection method of ferric ions used in the present invention has the characteristics of small error, high sensitivity, high accuracy, etc., and particularly shows superior sensitivity and accuracy when detecting low-concentration ferric ions.

Claims

1. A chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown as a formula (IV),

2. A method for preparing the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe as claimed in claim 1, which comprisesIs characterized in that the method comprises the following steps: taking a compound shown as a formula (II) and a compound shown as a formula (III) as raw materials, and taking Cu⁺As a catalyst, after the reaction is completed in tetrahydrofuran at 40-60 ℃, separating and purifying reaction liquid to obtain the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown in the formula (IV);

3. The method according to claim 2, wherein the amount of the compound of formula (II) to the amount of the compound of formula (III) is 2.2:1, and the amount of the catalyst is 1.2: 1; the volume of the tetrahydrofuran is 44ml/mmol based on the amount of the compound substance shown in the formula (III).

4. The method according to claim 2, wherein the reaction solution is separated and purified by: after the reaction is completed, concentrating the reaction liquid to be dry, adding water to dissolve, extracting by dichloromethane, taking an organic phase, washing by using a saturated sodium chloride aqueous solution, drying by anhydrous magnesium sulfate, filtering, evaporating the solvent from the filtrate under reduced pressure, performing thin layer chromatography, and performing CH chromatography with the volume ratio of 1:20₃OH:CH₂Cl₂Collecting the components with Rf value of 0.4-0.5 as developing agent to obtain the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe shown in formula (IV).

5. The process according to claim 2, wherein the compound of formula (II) is prepared by: taking a compound shown in a formula (I) and propargylamine as raw materials, completely carrying out reflux reaction in methanol, and carrying out post-treatment on a reaction solution to obtain a compound shown in a formula (II); the ratio of the compound shown in the formula (I) to the propargylamine feeding substance is 1: 5; the volume usage of the methanol is 20ml/mmol based on the amount of the compound substance shown in the formula (I);

6. the method according to claim 5, wherein the post-treatment of the reaction solution comprises: after the reaction is completed, concentrating the reaction liquid to be dry, adding water to dissolve, extracting by dichloromethane, taking an organic phase, washing by using a saturated sodium chloride aqueous solution, drying by anhydrous magnesium sulfate, filtering, evaporating the solvent from the filtrate under reduced pressure, performing thin layer chromatography, and performing CH chromatography with the volume ratio of 1:20₃OH:CH₂Cl₂Collecting the components with Rf value of 0.4-0.5 as developing agent to obtain the compound shown in formula (II).

7. The method for detecting Fe by using the chain double 1,2, 3-triazole rhodamine 6G fluorescent probe as claimed in claim 1³⁺The use of (1).

8. The use according to claim 7, characterized in that said use is: adding a sample to be detected into PBS (phosphate buffer solution) with pH value of 6.5 and 10mM and containing acetonitrile with volume concentration of 50 percent, adding a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe acetonitrile solution with the concentration of 5 mu mol/mL, and if the color is generated, the sample to be detected contains Fe³⁺。

9. The use according to claim 7, characterized in that said use is: adding a sample to be detected into PBS (phosphate buffer solution) containing acetonitrile with the volume concentration of 50% and the pH value of 6.5 and 10mM, adding a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe acetonitrile solution with the volume concentration of 5 mu mol/mL, measuring the fluorescence value at 555nm, and determining the fluorescence value according to Fe³⁺Obtaining Fe in the sample to be measured according to a standard curve³⁺Concentration; the volume ratio of the sample to be detected to the PBS buffer solution is 1:1, and the sample to be detected isThe volume ratio of the test sample to the probe acetonitrile solution is 10: 1; said Fe³⁺Standard curve is in Fe³⁺The concentration of the aqueous solution is plotted on the abscissa and the fluorescence value is plotted on the ordinate.

10. Use according to claim 9, characterized in that said Fe³⁺The standard curve was prepared as follows: fe with the concentration of 0, 0.01,0.05,0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4.5, 5, 5.5, 6, 6.5, 7, 7.5 mu mol/mL³⁺Adding the aqueous solution into PBS buffer solution with pH 6.5 and 10mM and containing acetonitrile with volume concentration of 50 percent, adding acetonitrile solution of a chain double 1,2, 3-triazole rhodamine 6G fluorescence probe with 5 mu mol/mL, measuring the fluorescence value at 555nm, and using Fe³⁺The concentration is taken as the abscissa and the fluorescence value is taken as the ordinate to obtain Fe³⁺A standard curve; said Fe³⁺The volume ratio of the aqueous solution to the PBS buffer solution is 1:1, and the Fe content is³⁺The volume ratio of the aqueous solution to the probe acetonitrile solution is 10: 1.