Fluorocarbon dot-containing bonded silica gel chromatographic stationary phase as well as preparation method and application thereof

Abstract

The invention discloses a fluorine-containing carbon dot bonded silica gel chromatographic stationary phase as well as a preparation method and application thereof, and belongs to the technical field of chromatographic stationary phases. According to the chromatographic stationary phase, naked silica gel is used as a substrate, and two substances are bonded on the surface, one substance is directly bonded six-carbon perfluoropolyether siloxane, and the other substance is fluorine-containing carbon dots bonded through 3-aminopropyltrimethoxysilane. The method disclosed by the invention has the beneficial effects that (1) the prepared chromatographic stationary phase SiO2-CDs-F3 is relatively high in stability and is applicable to both acidic and alkaline conditions; a hydrophobic effect and a hydrophilic effect exist at the same time, and an RPLC mode is achieved; the compound has relatively strong selectivity on strong polar compounds such as nucleosides, saponins, alkaloids and sulfonamides, and rapid and effective separation can be realized; and (2) the raw materials used in the preparation method of the chromatographic stationary phase SiO2-CDs-F3 provided by the invention are wide in source and low in cost, and the preparation method is suitable for large-scale industrial production.

Description

Technical Field

[0001] The invention relates to a chromatographic stationary phase and a preparation method and application thereof, and in particular to a fluorine-containing carbon point bonded silica gel chromatographic stationary phase and a preparation method and application thereof, belonging to the technical field of chromatographic stationary phases. Background Art

[0002] High performance liquid chromatography (HPLC) has become the most widely used analytical technology due to its advantages of high efficiency, high sensitivity, and high selectivity. It is widely used in the fields of drug analysis, food analysis, environmental monitoring, etc. Nowadays, the components of separated samples are complex, and liquid chromatography with a single retention mechanism cannot meet the separation requirements. For example, the traditional C18 chromatographic column has a poor retention effect on highly polar compounds such as nucleosides, saponins, alkaloids, and sulfonamides.

[0003] Carbon dots (CDs) are a type of zero-dimensional carbon nanomaterials with significant fluorescent properties, usually less than 10 nm in size. Carbon dots have the characteristics of high hydrophilicity, good stability, rich surface functional groups and moderate adsorption capacity. When bonded with silica gel, they can be evenly dispersed on the silica gel surface to avoid agglomeration, and have broad application prospects in the field of chromatographic separation. Summary of the invention

[0004] In order to overcome the deficiencies of the prior art, the object of the present invention is to provide a fluorine-containing carbon point bonded silica gel chromatographic stationary phase having a good retention effect (strong selectivity) for strongly polar compounds such as nucleosides, saponins, alkaloids, sulfonamides, etc., as well as a preparation method and application of the chromatographic stationary phase.

[0005] In order to achieve the above object, the present invention adopts the following technical solution: A fluorine-containing carbon point bonded silica gel chromatographic stationary phase takes bare silica gel as a matrix, and the surface of the bare silica gel is bonded with two substances, one is a directly bonded hexacarbon perfluoropolyether siloxane, and the other is a fluorine-containing carbon point bonded through 3-aminopropyltrimethoxysilane, wherein the bonding amount of the fluorine-containing carbon point is 2%-20%.

[0006] The preparation method of the aforementioned fluorine-containing carbon point bonded silica gel chromatographic stationary phase comprises the following steps: (1) Tetrafluoroterephthalic acid and tetraethylenepentamine are heated to synthesize fluorine-containing carbon dots; (2) Aminosilane reagent and silica gel are heated to synthesize amino silica gel; (3) Fluorine-containing carbon dots are bonded to amino-modified silica gel at room temperature to obtain fluorine-containing carbon dots modified silica gel; (4) Fluorine-containing carbon dot-modified silica gel is heated with a fluorosilane reagent to synthesize a fluorine-containing carbon dot-bonded silica gel chromatographic stationary phase.

[0007] Preferably, in step (1), the method for heating tetrafluoroterephthalic acid and tetraethylenepentamine to synthesize fluorine-containing carbon dots is as follows: (a) adding tetrafluoroterephthalic acid, ultrapure water and tetraethylenepentamine into a reaction container, and ultrasonicating to obtain a mixed solution; (b) pouring the mixed solution into a high pressure reactor and heating it at 180° C. for 24 h to obtain a reaction solution; (c) The reactor was naturally cooled to room temperature, the reaction solution was filtered through a 0.22 μm filter membrane, and the filtrate was dialyzed through a 500 Da dialysis bag to obtain a dialysate; (d) The dialysate is concentrated by rotary evaporation to obtain a fluorine-containing carbon dot solution.

[0008] Preferably, in step (2), the method for heating the aminosilane reagent and silica gel to synthesize the amino silica gel is as follows: (a) dispersing bare silica gel in a nitric acid solution, stirring at room temperature for 2 h, then centrifugally washing and drying to obtain acidified silica gel; (b) Anhydrous toluene, 3-aminopropyltrimethoxysilane and acidified silica gel were uniformly mixed in a ratio of 20 mL:1.5 mL:3 g, and reacted at 110°C for 3 h under the protection of N2, and then stopped heating for 9 h and then reacted at 110°C for 24 h; (c) After the reaction is completed, the product is centrifuged, washed and dried to obtain amino silica gel.

[0009] Preferably, in step (3), the method of bonding the fluorinated carbon dots to the amino-modified silica gel at room temperature to obtain the fluorinated carbon dots modified silica gel is as follows: (a) activating the fluorine-containing carbon dots with an activation solution to obtain an activated fluorine-containing carbon dot solution; (b) dispersing the activated fluorinated carbon dot solution and amino-modified silica gel in a ratio of 10 mL:1 g into 2-morpholineethanesulfonic acid buffer and reacting at room temperature for 12 h; (c) After the reaction is completed, the silica gel modified with fluorinated carbon dots is obtained by centrifugation, washing and drying.

[0010] Preferably, in step (4), the method for synthesizing the fluorinated carbon dot-bonded silica gel chromatographic stationary phase by heating the fluorinated carbon dot-modified silica gel with a fluorosilane reagent is as follows: (a) Dispersing hexafluoropolyether siloxane, triethylamine and silica gel modified with fluorinated carbon dots in anhydrous toluene in a ratio of 0.2576 g:445 μL:1 g, reacting at 105° C. for 12 h under N2 protection, centrifuging, washing and drying after the reaction to obtain product A; (b) dispersing hexafluoropolyether siloxane, triethylamine and product A in anhydrous toluene in a ratio of 0.2675 g:445 μL:total amount, reacting at 105° C. for 12 h under N2 protection, and centrifugally washing and drying after the reaction to obtain product B; (c) Trifluoropropyltrichlorosilane, triethylamine and product B were mixed in a ratio of 449.4 μL:445 μL:total amount, and reacted at 105° C. for 12 h. After the reaction, the mixture was centrifuged, washed and dried to obtain a fluorine-containing carbon point bonded silica gel chromatographic stationary phase.

[0011] The aforementioned fluorine-containing carbon point bonded silica gel chromatographic stationary phase is used for separating highly polar compounds, wherein the highly polar compounds include nucleosides, saponins, alkaloids and sulfonamides.

[0012] The present invention is beneficial in that: (1) The carbon quantum dots derived from tetrafluoroterephthalic acid and tetraethylenepentamine contain polar groups such as carboxyl, amino, double bonds, and fluorine atoms, as well as non-polar groups such as alkyl chains and benzene rings. This makes the chromatographic stationary phase SiO2-CDs-F3 prepared by the present invention have both hydrophobic and hydrophilic effects, and has an RPLC mode, thus enriching the types of liquid chromatography stationary phases. (2) The chromatographic stationary phase SiO2-CDs-F3 prepared by the present invention has strong selectivity for nucleosides, saponins, alkaloids and sulfonamide compounds, and can achieve rapid and effective separation, making up for the deficiency of the traditional C18 chromatographic column in having poor retention effect on highly polar compounds; (3) The chromatographic stationary phase SiO2-CDs-F3 prepared by the present invention has strong stability, is suitable for use under both acidic and alkaline conditions, and has a wide range of applications; (3) The raw materials used in the preparation method of the chromatographic stationary phase SiO2-CDs-F3 provided by the present invention are widely available and inexpensive, and are suitable for large-scale industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 It is a schematic diagram of the preparation process of the fluorine-containing carbon point bonded silica gel chromatographic stationary phase provided by the present invention; Figure 2 are transmission electron micrographs of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3, wherein A is a transmission electron micrograph of SiO2, B is a transmission electron micrograph of SiO2-CDs, C is a transmission electron micrograph of SiO2-F3, and D is a transmission electron micrograph of SiO2-CDs-F3; Figure 3are scanning electron micrographs of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3, wherein E is a scanning electron micrograph of SiO2, F is a scanning electron micrograph of SiO2-CDs, G is a scanning electron micrograph of SiO2-F3, and H is a scanning electron micrograph of SiO2-CDs-F3; Figure 4 It is the infrared spectra of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3; Figure 5 It is the elemental analysis diagram of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3; Figure 6 This is the separation chromatogram of five nucleoside compounds on SiO2-CDs-F3, where 1 is cytidine, 2 is guanosine, 3 is 5'-methyluridine, 4 is thymine, and 5 is β-thymidine; Figure 7 This is the separation chromatogram of 7 sulfonamide compounds on SiO2-CDs-F3, where 1 is sulfonamide, 2 is sulfacetamide, 3 is sulfadiazine, 4 is sulfapyridine, 5 is sulfamerazine, 6 is sulfadimethoxine, and 7 is sulfamethoxazole; Figure 8 This is the separation chromatogram of five saponin compounds on SiO2-CDs-F3, where 1 is ginsenoside Re, 2 is ginsenoside Rh1, 3 is ginsenoside Rb1, 4 is ginsenoside Rb3, and 5 is ginsenoside Rd; Fig. 9 This is the separation chromatogram of 5 alkaloid compounds on SiO2-CDs-F3, where 1 is lycorine hydrochloride, 2 is tetrahydropalmatine, 3 is berberine hydrochloride, 4 is reserpine, and 5 is gefitinib; Fig.10 This is the separation chromatogram of 7 common pesticides on SiO2-CDs-F3, where 1 is imidacloprid, 2 is pyrimethanil, 3 is difenoconazole, 4 is chlorfenapyr, 5 is hexaflumuron, 6 is chlorpyrifos, and 7 is pyridabenzane; Fig.11 This is the gradient elution chromatogram of total ginsenosides on SiO2-CDs-F3. DETAILED DESCRIPTION

[0014] The present invention is described in detail below with reference to the accompanying drawings and embodiments.

[0015] 1. Preparation of Fluorinated Carbon-dot Bonded Silica Gel Chromatographic Stationary Phase The fluorine-containing carbon point bonded silica gel chromatographic stationary phase provided by the present invention has the following structure: bare silica gel (SiO2) is used as a matrix, and two substances are bonded to the surface, one is a directly bonded hexacarbon perfluoropolyether siloxane, and the other is a fluorine-containing carbon point bonded through 3-aminopropyltrimethoxysilane, wherein the bonding amount of the fluorine-containing carbon point is 2%-20%.

[0016] like Figure 1 As shown, the preparation method of the fluorine-containing carbon dot bonded silica gel chromatographic stationary phase provided by the present invention comprises the following four steps: 1. heating tetrafluoroterephthalic acid and tetraethylenepentamine to synthesize fluorine-containing carbon dots; 2. heating aminosilane reagent and silica gel to synthesize amino-modified silica gel; 3. bonding the fluorine-containing carbon dots to amino-modified silica gel at room temperature to obtain fluorine-containing carbon dot-modified silica gel; 4. heating the fluorine-containing carbon dot-modified silica gel with a fluorosilane reagent to synthesize a fluorine-containing carbon dot bonded silica gel chromatographic stationary phase.

[0017] 1. Synthesis of fluorinated carbon dots by heating tetrafluoroterephthalic acid and tetraethylenepentamine Add 1.5 g of tetrafluoroterephthalic acid and 60 mL of ultrapure water to a beaker, then add 20 mL of tetraethylenepentamine, and ultrasonicate for 5 min to obtain a mixed solution. Pour the mixed solution into a polytetrafluoroethylene-lined autoclave and heat at 180 ° C for 24 h. After heating, the autoclave is naturally cooled to room temperature. The cooled reaction solution is filtered with a 0.22 μm filter membrane, and then the filtrate is dialyzed in a 500 Da dialysis bag for 48 h. The dialyzate obtained is concentrated to 10 mL by rotary evaporation to obtain a fluorinated carbon dot (F-CDs) solution.

[0018] After testing, the nitrogen and fluorine doping levels of the F-CDs were 7.25% and 19.63%, respectively.

[0019] 2. Synthesis of aminosilane silica gel by heating aminosilane reagent and silica gel Disperse 3 g of bare silica gel (SiO2) in 30 mL of 15% (v / v) nitric acid solution, stir at room temperature for 2 h, centrifuge and wash after the reaction is completed, first wash with deionized water until the pH of the supernatant is neutral, then wash three times with anhydrous ethanol, and dry at 60°C to obtain acidified silica gel.

[0020] 20 mL of anhydrous toluene, 1.5 mL of 3-aminopropyltrimethoxysilane and 3 g of the acidified silica gel obtained above were mixed evenly, and reacted at 110°C for 3 h under the protection of N2, then stopped heating for 9 h, and then reacted at 110°C for 24 h. After the reaction, centrifuge and wash, wash with toluene (or anhydrous ethanol, 50% (v / v) ethanol, methanol) 3 times, and dry at 60°C to obtain amino silica gel (SiO2-NH2).

[0021] 3. Fluorine-containing carbon dots are bonded to amino-modified silica gel at room temperature to obtain fluorine-containing carbon dots modified silica gel 10 mL of the previously obtained F-CDs solution was activated with 12 mL of activation solution (equal volumes of a 20 mg / mL 1-ethyl-(3-dimethylaminopropyl)carbodiimide solution and a 10 mg / mL N-hydroxysuccinimide solution) at room temperature for 10 min to obtain an activated F-CDs solution.

[0022] 10 mL of the activated F-CDs solution obtained above and 1 g of SiO2-NH2 obtained above were dispersed in 45 mL of 0.1 mM 2-morpholineethanesulfonic acid buffer and reacted at room temperature for 12 h. After the reaction, the mixture was centrifuged and washed 3 times with pure water and 3 times with anhydrous ethanol, and finally dried at 60 ° C to obtain fluorinated carbon dot-modified silica gel (SiO2-CDs).

[0023] After testing, the fluorine-containing carbon point bonding content of the SiO2-CDs was 9.36%.

[0024] 4. Synthesis of fluorinated carbon-dot bonded silica gel chromatographic stationary phase by heating fluorinated carbon-dot modified silica gel with fluorosilane reagent 0.2576g of hexafluoropolyether siloxane, 445μL of triethylamine and 1g of SiO2-CDs obtained above were uniformly dispersed in 25mL of anhydrous toluene, and reacted at 105°C for 12h under N2 protection. After the reaction, centrifugation and washing were performed, first washed with toluene 3 times, then washed with anhydrous ethanol 3 times, and finally dried at 60°C to obtain product A.

[0025] 0.2675g of hexafluoropolyether siloxane, 445μL of triethylamine and all the product A obtained above were uniformly dispersed in 25mL of anhydrous toluene, and reacted at 105°C for 12h under N2 protection. After the reaction, centrifuge and wash, first wash with toluene 3 times, then wash with anhydrous ethanol 3 times, and finally dry at 60°C to obtain product B.

[0026] 449.4 μL trifluoropropyltrichlorosilane, 445 μL triethylamine and the entire amount of product B obtained above were mixed and reacted at 105°C for 12 hours. After the reaction, the mixture was centrifuged and washed with toluene 3 times, then with anhydrous ethanol 3 times, and finally dried at 60°C to obtain a fluorine-containing carbon point bonded silica gel chromatographic stationary phase (SiO2-CDs-F3).

[0027] After testing, the fluorine-containing carbon point bonding content of SiO2-CDs-F3 is 9.36%.

[0028] As a control, amino silica gel (SiO2-NH2) was heated with a fluorosilane reagent to synthesize a silica gel chromatographic stationary phase without carbon dots (SiO2-F3). The preparation method of SiO2-F3 was basically the same as that of SiO2-CDs-F3, with the only difference being that amino silica gel (1 g) was used instead of SiO2-CDs (1 g).

[0029] 2. Analysis of the structure of fluorinated carbon point bonded silica gel chromatographic stationary phase The structures of unreacted bare silica gel (SiO2) and previously obtained SiO2-F3, SiO2-CDs and SiO2-CDs-F3 were analyzed by transmission electron microscopy, scanning electron microscopy, infrared spectrometer and elemental analyzer.

[0030] 1. Transmission electron microscopy analysis Transmission electron micrographs of SiO2, SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are shown in Figure 2 .

[0031] Through transmission electron microscopy, it can be observed that the surface of SiO2 is smooth, the surfaces of SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are all rough, among which the surface of SiO2-CDs-F3 is the roughest.

[0032] This indicates that compounds are bonded to the surfaces of SiO2-CDs, SiO2-F3 and SiO2-CDs-F3, among which SiO2-CDs-F3 has the most compounds bonded to its surface.

[0033] 2. Scanning electron microscopy analysis Scanning electron micrographs of SiO2, SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are shown in Figure 3 .

[0034] Scanning electron microscopy shows that SiO2, SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are all spherical and evenly dispersed. Except for SiO2, which has a smooth surface, the surfaces of SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are all rough to varying degrees, among which the surface of SiO2-CDs-F3 is the roughest.

[0035] This indicates that compounds are bonded to the surfaces of SiO2-CDs, SiO2-F3 and SiO2-CDs-F3, among which SiO2-CDs-F3 has the most compounds bonded to its surface.

[0036] 3. Infrared spectroscopy analysis The infrared spectra of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are shown in Figure 4 .

[0037] It can be observed from the infrared spectra that SiO2, SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 have a wavelength of 3450 cm -1 and 1070cm -1 There are absorption peaks at 2930cm-1, which are attributed to the vibration of ν(OH / NH) and ν(Si-O) respectively; SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 have absorption peaks at 2930cm-1. -1 There are absorption peaks different from SiO2 at 1540 cm-1 and 1560 cm-2, which are attributed to the vibration of ν(CH). -1 、1710cm -1 and 1210cm -1 New absorption peaks appeared at all locations, and these three absorption peaks were attributed to the vibrations of ν (RNC=O), ν (C=O) and ν (CF), respectively.

[0038] This indicates that the surface of SiO2-CDs is successfully bonded with F-CDs, the surface of SiO2-F3 is successfully bonded with hexafluoropolyether siloxane, and the surface of SiO2-CDs-F3 is successfully bonded with F-CDs and hexafluoropolyether siloxane.

[0039] 4. Quantitative analysis of elements The American Thermo Fisher Scientific-ESCALAB 250Xi instrument was used to quantitatively analyze the three elements C, O, and F in bare silica gel (SiO2), SiO2-CDs, SiO2-F3, and SiO2-CDs-F3.

[0040] The quantitative analysis results of C, O and F elements of bare silica gel (SiO2), SiO2-CDs, SiO2-F3 and SiO2-CDs-F3 are shown in Figure 5 .

[0041] From the element quantitative analysis chart, we can see that the content of F in SiO2-CDs-F3 is 40.59%, and the contents of C and O are 7.97% and 40.93% respectively, which also shows that SiO2-CDs-F3 has been successfully prepared.

[0042] 3. Testing the Selectivity of Fluorinated Carbon-dot Bonded Silica Gel Chromatographic Stationary Phase The SiO2-CDs-F3 prepared by the present invention has polar groups such as carboxyl groups, amino groups, double bonds, fluorine atoms, and non-polar groups such as alkyl chains and benzene rings (present in carbon quantum dots), so it has both hydrophobic and hydrophilic effects and has a reverse phase chromatography (RPLC) mode. Under the RPLC mode, the selectivity of SiO2-CDs-F3 for nucleosides, saponins, alkaloids and sulfonamide compounds is tested.

[0043] 1. Selectivity for nucleoside compounds Take cytidine, guanosine, 5'-methyluridine, thymine and β-thymidine as examples.

[0044] Chromatographic conditions: mobile phase: 0.4% (v / v) acetic acid solution; detection wavelength: 260nm; flow rate: 0.2mL / min; injection volume 1μL; column temperature: 30℃.

[0045] Detection process: Cytidine, guanosine, 5'-methyluridine, thymine and β-thymidine were respectively prepared into a solution with a concentration of 1000 ppm using a 0.5% (v / v) methanol solution. 100 μL of cytidine solution, 100 μL of guanosine solution, 100 μL of 5'-methyluridine solution, 150 μL of thymine solution and 200 μL of β-thymidine solution were pipetted with a pipette to prepare a mixed standard. The mixture was injected continuously for 5 times through a high performance liquid chromatography to obtain the separation chromatogram of the compound on SiO2-CDs-F3.

[0046] The separation chromatograms of five nucleoside compounds, cytidine, guanosine, 5'-methyluridine, thymine and β-thymidine, on SiO2-CDs-F3 are shown in Figure 6 .

[0047] Depend on Figure 6 It can be seen that SiO2-CDs-F3 has a good separation effect on five nucleoside compounds, namely cytidine, guanosine, 5'-methyluridine, thymine and β-thymidine. The plate numbers are 8700N / m, 9120N / m, 11240N / m, 15080N / m and 11620N / m in the order of peak elution, and the separation degrees R are all greater than 1.5.

[0048] 2. Selectivity for sulfonamide compounds Take sulfonamide, sulfacetamide, sulfadiazine, sulfapyridine, sulfamethazine, sulfadimethoxine and sulfamethoxazole as examples.

[0049] Chromatographic conditions: mobile phase: methanol and 10 mM acetic acid solution mixed in a volume ratio of 13:87; detection wavelength: 270 nm; flow rate: 0.2 mL / min; injection volume: 1 μL; column temperature: 30°C.

[0050] Detection process: Sulfonamide, sulfacetamide, sulfadiazine, sulfapyridine, sulfamethazine, sulfadimethoxazole and sulfamethoxazole were respectively prepared into a solution with a concentration of 1000 ppm using a 50% (v / v) acetonitrile solution. 25 μL of sulfonamide solution, 50 μL of sulfacetamide solution, 100 μL of sulfadiazine solution, 100 μL of sulfapyridine solution, 100 μL of sulfamethoxazole solution, 150 μL of sulfamethoxazole solution and 300 μL of sulfamethoxazole solution were pipetted into a mixed standard. The mixture was injected continuously for 5 times through a high performance liquid chromatography to obtain the separation chromatogram of the compound on SiO2-CDs-F3.

[0051] The separation chromatograms of sulfonamide, sulfacetamide, sulfadiazine, sulfapyridine, sulfamerazine, sulfadimethoxine and sulfamethoxazole on SiO2-CDs-F3 are shown in Figure 2. Figure 7 .

[0052] Depend on Figure 7 It can be seen that SiO2-CDs-F3 has a good separation effect on seven sulfonamide compounds, including sulfonamide, sulfacetamide, sulfadiazine, sulfapyridine, sulfamerazine, sulfadimethoxine and sulfamethoxazole. The plate numbers are 12750N / m, 12700N / m, 11040N / m, 12200N / m, 11950N / m, 13040N / m and 17030N / m in the order of peak elution, and the separation degrees are all greater than 1.5.

[0053] 3. Selectivity for saponin compounds Take ginsenosides Re, Rh1, Rb1, Rb3 and Rd as examples.

[0054] Chromatographic conditions: mobile phase: 50% (v / v) methanol solution; detection wavelength: 203 nm; flow rate: 0.2 mL / min; injection volume 1 μL; column temperature: 30°C.

[0055] Detection process: Ginsenosides Re, Rh1, Rb1, Rb3 and Rd were respectively prepared into solutions with a concentration of 10000 ppm using a 44% (v / v) methanol solution. 50 μL Rh1 solution, 200 μL Rb1 solution, 200 μL Rb3 solution, 200 μL Rd solution and 70 μL Re solution were pipetted into a mixed standard. The mixture was injected continuously for 5 times through a high performance liquid chromatography to obtain the separation chromatogram of the compound on SiO2-CDs-F3.

[0056] The separation chromatograms of ginsenoside Re, Rh1, Rb1, Rb3 and Rd on SiO2-CDs-F3 are shown in Figure 8 .

[0057] Depend on Figure 8 It can be seen that SiO2-CDs-F3 has a good separation effect on the five saponin compounds, namely ginsenoside Re, Rh1, Rb1, Rb3 and Rd. The plate numbers are 7530N / m, 10460N / m, 11380N / m, 12380N / m and 14630N / m in the order of peak elution, and the separation degrees are all greater than 1.5.

[0058] 4. Selectivity for alkaloid compounds Take lycorine hydrochloride, tetrahydropalmatine, berberine hydrochloride, reserpine and gefitinib as examples.

[0059] Chromatographic conditions: mobile phase: methanol and 10 mM acetic acid solution mixed in a volume ratio of 11:9; detection wavelength: 280 nm; flow rate: 0.2 mL / min; injection volume: 1 μL; column temperature: 30°C.

[0060] Detection process: Lycorine hydrochloride, tetrahydropalmatine, berberine hydrochloride, reserpine and gefitinib were respectively prepared into a solution with a concentration of 1000 ppm using an acetonitrile solution with a concentration of 83% (v / v). 100 μL of tetrahydropalmatine solution, 100 μL of reserpine solution, 100 μL of lycorine hydrochloride solution, 100 μL of berberine hydrochloride solution and 200 μL of gefitinib solution were pipetted into a mixed standard. The mixture was injected continuously for 5 times through a high performance liquid chromatography to obtain the separation chromatogram of the compound on SiO2-CDs-F3.

[0061] The separation chromatograms of five alkaloid compounds, namely, lycorine hydrochloride, tetrahydropalmatine, berberine hydrochloride, reserpine and gefitinib on SiO2-CDs-F3 are shown in Fig. 9 .

[0062] Depend on Fig. 9 It can be seen that SiO2-CDs-F3 has a good separation effect on five alkaloid compounds, namely lycorine hydrochloride, corydaline, berberine hydrochloride, reserpine and gefitinib. The plate numbers are 11790N / m, 15100N / m, 11950N / m, 12450N / m and 4550N / m in the order of peak elution, and the separation degrees are all greater than 1.5.

[0063] 5. Selectivity for common pesticides Take common pesticides such as imidacloprid, pyrimethanil, dimethoate, carbofuran, fluazifop, chlorpyrifos, and pyridaben as examples.

[0064] Chromatographic conditions: mobile phase: 61% (v / v) methanol solution; detection wavelength: 270 nm; flow rate: 0.2 mL / min; injection volume 1 μL; column temperature: 30°C.

[0065] Detection process: imidacloprid, pyrimethanil, dimethoate, chlorfenapyr, fluazifop, chlorpyrifos and pyridaben were respectively prepared into solutions with a concentration of 1000 ppm with methanol, and 10 μL of imidacloprid solution, 10 μL of pyrimethanil solution, 50 μL of fluazifop solution, 70 μL of fluazifop solution, 70 μL of fluazifop solution, 200 μL of chlorpyrifos solution and 200 μL of dimethoate solution were pipetted with a pipette to prepare a mixed standard, which was continuously injected 5 times through a high performance liquid chromatography to obtain the separation chromatogram of the compound on SiO2-CDs-F3.

[0066] The separation chromatograms of 7 common pesticides, including imidacloprid, pyrimethanil, difenoconazole, chlorpyrifos, hexaflumuron, chlorpyrifos and pyridaben on SiO2-CDs-F3 are shown in Figure 2. Fig.10 .

[0067] Depend on Fig.10 It can be seen that SiO2-CDs-F3 has a good separation effect on seven common pesticides, including imidacloprid, pyrimethanil, dimethoate, chlorpyrifos, fluazifop, chlorpyrifos and pyridaben. The plate numbers are 7250 N / m, 12240 N / m, 12510 N / m, 18190 N / m, 18560 N / m, 18750 N / m and 18500 N / m in the order of peak elution, and the separation degrees are all greater than 1.5.

[0068] 4. Practical Application Cases The SiO2-CDs-F3 prepared by the present invention is assembled into a chromatographic column, and the total ginsenosides of ginseng are used as the analyte to separate and analyze R1, Rd, Rh2, and Rg3 in the total ginsenosides of ginseng.

[0069] Sample pretreatment: Weigh 1g of total ginsenosides, dissolve it in 20mL of 70% (v / v) methanol solution, ultrasonicate for 30min, and then filter with a 0.22μm filter membrane. Rotary evaporate the filtrate (to remove excess solvent) to obtain a solid. Then dissolve the obtained solid in 5mL of 70% (v / v) methanol solution, and finally filter with a 0.22μm filter membrane to obtain the filtrate to be tested.

[0070] Use a pipette to draw 0.5 mL of the filtrate to be tested for liquid phase separation. Mobile phase: A: 0.1% (v / v) formic acid solution, B: acetonitrile; injection volume: 1 μL; column temperature: 30°C; flow rate: 0.2 mL / min.

[0071] Mobile phase: 0min: 85%A, 15%B; 0-3min: 15%-20%B; 3-4min: 20%B; 4-10min: 20%-30%B; 10-25min: 30%-32.5%B; 25-27min: 32.5%-60%B; 27-39min: 60-95%B; 39-40min: 95%B; 40-40.5min: 95%-15%B; 40.5-41min: 15%B.

[0072] The separation results of R1, Rd, Rh2 and Rg3 in ginsenosides are shown in Fig.11 The results showed that within 40 minutes of rapid separation, R1, Rd, Rh2, and Rg3 were successfully separated from the total ginsenosides. This proves that the SiO2-CDs-F3 prepared by the present invention has a good application prospect in the separation and analysis of complex samples.

[0073] It should be noted that the above embodiments are merely examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is impossible to list all the embodiments here. Any obvious changes or modifications derived from the technical solution of the present invention are still within the scope of protection of the present invention.

Claims

1. A fluorine-containing carbon point bonded silica gel chromatographic stationary phase, with bare silica gel as the matrix, characterized in that: There are two substances bonded to the surface of bare silica gel, one is directly bonded hexacarbon perfluoropolyether siloxane, and the other is fluorinated carbon points bonded through 3-aminopropyltrimethoxysilane, where the bonding amount of fluorinated carbon points is 2%-20%.

2. The method for preparing the fluorine-containing carbon point bonded silica gel chromatographic stationary phase according to claim 1, characterized in that: The following steps are involved: (1) Tetrafluoroterephthalic acid and tetraethylenepentamine are heated to synthesize fluorine-containing carbon dots; (2) Aminosilane reagent and silica gel are heated to synthesize amino silica gel; (3) Fluorine-containing carbon dots are bonded to amino-modified silica gel at room temperature to obtain fluorine-containing carbon dots modified silica gel; (4) Fluorine-containing carbon dot-modified silica gel is heated with a fluorosilane reagent to synthesize a fluorine-containing carbon dot-bonded silica gel chromatographic stationary phase.

3. The preparation method according to claim 2, characterized in that: In step (1), the method for heating tetrafluoroterephthalic acid and tetraethylenepentamine to synthesize fluorine-containing carbon dots is as follows: (a) adding tetrafluoroterephthalic acid, ultrapure water and tetraethylenepentamine into a reaction container, and ultrasonicating to obtain a mixed solution; (b) pouring the mixed solution into a high pressure reactor and heating it at 180° C. for 24 h to obtain a reaction solution; (c) The reactor was naturally cooled to room temperature, the reaction solution was filtered through a 0.22 μm filter membrane, and the filtrate was dialyzed through a 500 Da dialysis bag to obtain a dialysate; (d) The dialysate is concentrated by rotary evaporation to obtain a fluorine-containing carbon dot solution.

4. The preparation method according to claim 2, characterized in that: In step (2), the method for synthesizing aminosilane silica gel by heating the aminosilane reagent and silica gel is as follows: (a) dispersing bare silica gel in a nitric acid solution, stirring at room temperature for 2 h, then centrifugally washing and drying to obtain acidified silica gel; (b) Anhydrous toluene, 3-aminopropyltrimethoxysilane and acidified silica gel were uniformly mixed in a ratio of 20 mL:1.5 mL:3 g, and reacted at 110°C for 3 h under the protection of N2, and then stopped heating for 9 h and then reacted at 110°C for 24 h; (c) After the reaction is completed, the product is centrifuged, washed and dried to obtain amino silica gel.

5. The preparation method according to claim 2, characterized in that: In step (3), the method of bonding the fluorinated carbon dots to the amino-modified silica gel at room temperature to obtain the fluorinated carbon dots modified silica gel is as follows: (a) activating the fluorine-containing carbon dots with an activation solution to obtain an activated fluorine-containing carbon dot solution; (b) dispersing the activated fluorinated carbon dot solution and amino-modified silica gel in a ratio of 10 mL:1 g into 2-morpholineethanesulfonic acid buffer and reacting at room temperature for 12 h; (c) After the reaction is completed, the silica gel modified with fluorinated carbon dots is obtained by centrifugation, washing and drying.

6. The preparation method according to claim 2, characterized in that: In step (4), the method for synthesizing the fluorinated carbon dot-bonded silica gel chromatographic stationary phase by heating the fluorinated carbon dot-modified silica gel with a fluorosilane reagent is as follows: (a) Dispersing hexafluoropolyether siloxane, triethylamine and silica gel modified with fluorinated carbon dots in anhydrous toluene in a ratio of 0.2576 g:445 μL:1 g, reacting at 105° C. for 12 h under N2 protection, centrifuging, washing and drying after the reaction to obtain product A; (b) dispersing hexafluoropolyether siloxane, triethylamine and product A in anhydrous toluene in a ratio of 0.2675 g:445 μL:total amount, reacting at 105° C. for 12 h under N2 protection, and centrifugally washing and drying after the reaction to obtain product B; (c) Trifluoropropyltrichlorosilane, triethylamine and product B were mixed in a ratio of 449.4 μL:445 μL:total amount, and reacted at 105° C. for 12 h. After the reaction, the mixture was centrifuged, washed and dried to obtain a fluorine-containing carbon point bonded silica gel chromatographic stationary phase.

7. Use of the fluorine-containing carbon point bonded silica gel chromatographic stationary phase according to claim 1 in separating highly polar compounds, wherein the highly polar compounds include nucleosides, saponins, alkaloids and sulfonamides.

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