A 3D printed Lay-Fomm60 coated stirring rod and its preparation method and application

Abstract

The present invention discloses a 3D-printed Lay-Fomm60-coated stir bar and its preparation method and application. The stir bar includes a coated cylinder and a glass capillary; the coated cylinder is a hollow cylinder made of Lay-Fomm60 material by 3D printing; the coated cylinder is subjected to surface activation treatment; the glass capillary is embedded with iron wire and is externally sheathed with the coated cylinder. The present invention also discloses the application of the stir bar in enriching and detecting estrogens in samples. The stir bar is used for stir bar sorptive extraction and is combined with a high performance liquid chromatography-ultraviolet detector to detect five estrogens in environmental water samples. The method for printing the stir bar is simple, convenient and low-cost, and has good analytical and detection capabilities in combination with the detection method.

Description

Technical Field

[0001] The present invention belongs to the technical field of separation and analytical detection, and specifically relates to a 3D printed stir bar and a preparation method thereof, as well as a method for performing stir bar sorptive extraction and coupling with a high performance liquid chromatography-ultraviolet detector to detect five estrogens in environmental water samples. Background Art

[0002] Estrogens are important chemicals that affect the endocrine system, so their content in environmental samples has attracted much attention. Since the early 1980s, studies on estrogens in surface water have been reported. Endogenous estrogens and synthetic estrogens are the main sources of environmental estrogens. They can enter the water environment through sewage, agricultural, pharmaceutical and aquaculture waste treatment systems. A large number of studies have shown that estrogens may have health effects on aquatic wildlife and humans, leading to problems in growth, development and reproduction, such as feminization and hermaphroditism. Once estrogens enter the human body excessively through the food chain, they will interfere with the endocrine system and increase the risk of cancer, such as prostate cancer and breast cancer. Therefore, the development of analytical techniques for effectively enriching and analyzing low-concentration estrogens is of great significance for scientific and health issues.

[0003] Generally speaking, a complete sample analysis process consists of four parts: sample collection, sample pretreatment, analytical testing and data processing. Sample pretreatment is an important part of the analysis process, and its main purpose is to remove matrix interference in complex samples and enrich trace / ultratrace target analytes. The selection and operation of sample pretreatment directly affect the precision and accuracy of analytical data, and even directly determine the success or failure of the entire analytical method. Traditional sample pretreatment methods (such as Soxhlet extraction, liquid-liquid extraction, etc.) have disadvantages such as complex operation, long time consumption, low extraction efficiency, and large consumption of organic solvents. To solve these problems, new miniaturized, green, simple, rapid and effective sample pretreatment technologies such as solid phase microextraction (SPME), magnetic solid phase extraction (MSPE), stir bar sorptive extraction (SBSE), etc. have emerged. Among them, SBSE has developed vigorously due to its advantages such as good reproducibility, high recovery rate, simple operation and less consumption of organic solvents.

[0004] Stir bar sorptive extraction is a novel sample pretreatment technique that realizes the extraction and enrichment of analytes based on the different equilibrium distribution ratios of the analytes between the sample and the extraction coating. The typical structure of a coated stir bar: there is an iron core that provides the magnetic stirring power inside the glass shell, and various functional materials can be coated on this substrate using appropriate coating preparation techniques to obtain a coated stir bar. Since the stir bar coated with materials can automatically stir, thus accelerating the attainment of extraction equilibrium, it avoids the competitive adsorption caused by using a magnetic stir bar; compared with SPME, the coating on the surface of the stir bar is large in amount and high in adsorption capacity, which is conducive to improving its extraction efficiency for target analytes; the magnetism of the coated stir bar also enables rapid phase separation using a magnet.

[0005] In SBSE, since the mechanical properties of the coating are required to be relatively high during the extraction and desorption processes, it is necessary to select a suitable coating preparation method according to the physical and chemical properties of the functional materials and load them on the surface of the glass stir bar. The methods mainly used for coating preparation include: sol-gel method, physical adhesion method, monolithic column method, in-situ growth method, molecular imprinting technique. In addition, there are other novel coating preparation methods (such as electrodeposition method, self-assembly method, solvent exchange method, physical magnetic adsorption method, electrospinning method, etc.) introduced into the preparation of stir bars. However, no one has used 3D printing technology for the preparation of stir bar coatings, nor have there been any related reports.

[0006] 3D printing belongs to a kind of rapid prototyping technology. It is an additive manufacturing technology based on digital model files, using powdery metals or plastics and other bondable materials to construct objects by means of layer-by-layer stacking and accumulation. This technology was developed by Charles Hull in 1986 during the production of stereolithography (SLA). Compared with traditional manufacturing methods, 3D printing has various advantages: fast, low cost, and less waste generated during the manufacturing process. Nowadays, 3D printing is an important tool for manufacturing and prototyping. Currently, 3D printing technologies include fused deposition modelling (FDM), stereolithography (SLA), selective laser sintering (SLS), inkjet and multi-jet printing, and laminated object manufacturing (LOM). 3D printing has become a popular manufacturing tool in various disciplines, and the application fields include analytical chemistry, environmental science, biology, medical devices, and other manufacturing fields. Summary of the Invention

[0007] Aiming at the deficiencies of existing stir bars and their preparation methods, one of the objectives of the present invention is to introduce 3D printing technology into the preparation of stir bar coatings, establish a new stir bar coating preparation technology, and thus overcome the defects of stir bars prepared by some conventional methods, and solve the problem of cumbersome preparation operations of coated stir bars in SBSE. The second objective of the present invention is to expand the application scope of 3D printing technology in the field of analytical chemistry, realize the coupling of 3D printed stir bar sorptive extraction and high performance liquid chromatography-ultraviolet detector, and detect estrogens in actual samples.

[0008] The objectives of the present invention are achieved through the following technical solutions:

[0009] In the first aspect, the present invention provides a 3D printed Lay-Fomm60 coated stir bar, which is characterized in that it includes a coated cylinder and a glass capillary;

[0010] The coated cylinder is a hollow cylinder made of Lay-Fomm60 material by 3D printing; the coated cylinder has been subjected to surface activation treatment;

[0011] The glass capillary is embedded with iron wire and is sheathed with the coated cylinder.

[0012] Further, the Lay-Fomm60 material is a composite of polyurethane and polyvinyl alcohol.

[0013] Further, the inner diameter of the coated cylinder is 2.0 mm, the outer diameter is 5.0 mm, and the cylinder height is 2.00 cm.

[0014] In the second aspect, the present invention provides a preparation method for the stir bar described in the first aspect, and the steps are as follows:

[0015] (1) Coated cylinder design: Design the 3D printed stir bar coated cylinder as a hollow cylinder structure;

[0016] (2) Printing the stir bar coated cylinder by fused deposition modeling 3D printing technology: Using a 3D printer to process the digital model through an open source 3D slicing software for the designed coated cylinder, and performing 3D printing on the Lay-Fomm60 material to obtain the coated cylinder;

[0017] (3) Surface activation of the coated cylinder in step (2): Soak the stir bar coating in deionized water, then soak it in methanol, and then place it at room temperature to dry;

[0018] (4) Stir bar assembly: Insert the glass capillary embedded with iron wire into the stir bar coating to obtain it.

[0019] Further, the design software in step (1) is Rhinoceros software.

[0020] Further, in the step (2), the open-source 3D slicing software is Cura.

[0021] In a third aspect, the present invention provides the use of the stirring rod described in the first aspect in enriching and detecting estrogens in a sample.

[0022] In a third aspect, the present invention provides a method for detecting estrogens using the stirring rod described in the first aspect, comprising the following steps:

[0023] (1) Add a sample solution containing the target analyte to an extraction container, place the stirring rod, and perform extraction under stirring;

[0024] (2) After the extraction is completed, take out the stirring rod, wipe the residual solution on the surface of the stirring rod with filter paper, and then place the stirring rod in a desorption tube containing a desorbent for desorption;

[0025] (3) Take the desorbed solution and perform detection using a high performance liquid chromatography-ultraviolet detector.

[0026] Further, in the step (2), the desorbent is methanol, the desorption volume is 2 mL, and the desorption time is 8 min.

[0027] Further, in the step (3), the parameters of the high performance liquid chromatography-ultraviolet detector are as follows: the mobile phase composition is acetonitrile and water, an EC-C18 column is used, the flow rate is 0.70 mL / min, the wavelength of the ultraviolet detector is 200 nm, and the injection volume is 20 μL.

[0028] Further, in the mobile phase, the volume ratio of acetonitrile to water is 52 / 48; the size of the EC-C18 column is 150 mm × 4.6 mm, and the particle size is 4 μm.

[0029] The principle of the present invention is as follows: The 3D printed Lay-Fomm60 material is a composite material of polyurethane and polyvinyl alcohol. After being washed and activated with water for a long time, the polyvinyl alcohol in the material can be dissolved, and pore structures are generated on the surface. Through hydrophobic interaction and size matching effect, the enrichment and detection of the target analyte (estrogen) are realized.

[0030] The beneficial effects of the present invention are as follows: Compared with the conventional stirring rod coating preparation technology, the 3D printing stirring rod coating preparation method can avoid the phenomena of material embedding and shedding, and is faster and simpler. And the present invention can realize the adsorption extraction of the stirring rod and the combination with a detection instrument for the analysis of actual samples. The analytical detection ability of this method is compared with the related research based on solid phase adsorption materials in recent years: Compared with the methods with the same ultraviolet detector, the detection limit of this method is lower or equivalent; for the methods with a mass spectrometry detector, the detection limit of this method is higher, but the service life of this method is longer. Description of the Drawings

[0031] Figure 1 This is the schematic diagram of the technical solution of the present invention.

[0032] Figure 2 This is the scanning electron microscope (SEM) image (2 mm) of the cross-section of the 3D printed stirring rod prepared in Example 1 after activation.

[0033] Figure 3 This is the scanning electron microscope (SEM) image (500 μm) of the 3D printed stirring rod prepared in Example 1 before (A) and after (B) activation.

[0034] Figure 4 This is the scanning electron microscope (SEM) image (100 μm) of the 3D printed stirring rod prepared in Example 1 before (A) and after (B) activation.

[0035] Figure 5 This is the scanning electron microscope (SEM) image (50 μm) of the 3D printed stirring rod prepared in Example 1 before (A) and after (B) activation.

[0036] Figure 6 This is the scanning electron microscope (SEM) image (5 μm) of the 3D printed stirring rod prepared in Example 1 before (A) and after (B) activation.

[0037] Figure 7 This is the infrared spectrum (IR) image of the 3D printed stirring rod prepared in Example 1 before and after activation.

[0038] Figure 8 This is the water contact angle characterization image of the 3D printing material used in Example 1 before and after activation.

[0039] Figure 9 This is the design and physical image of the 3D printed stirring rod in Example 1.

[0040] Figure 10 This is the adsorption efficiency of the 3D printed Lay-Fomm60 coated stirring rod for 4 kinds of chlorophenols in Example 1.

[0041] Figure 11 This is the adsorption efficiency of the 3D printed Lay-Fomm60 coated stirring rod for 5 kinds of estrogens in Example 1.

[0042] Figure 12 This is the optimization diagram of the pH value of the adsorption extraction solution of the 3D printed stirring rod prepared in Example 1.

[0043] Figure 13 This is the optimization diagram of the ionic strength of the adsorption extraction solution of the 3D printed stirring rod prepared in Example 1.

[0044] Figure 14 This is the optimization diagram of the extraction time of the adsorption extraction of the 3D printed stirring rod prepared in Example 1.

[0045] Figure 15 Optimization diagram of the types of desorbing agents for the 3D printed stir bar prepared in Example 1 for adsorption extraction.

[0046] Figure 16 Optimization diagram of the desorbing volume for the 3D printed stir bar prepared in Example 1 for adsorption extraction.

[0047] Figure 17 Optimization diagram of the desorbing time for the 3D printed stir bar prepared in Example 1 for adsorption extraction.

[0048] Figure 18 Service life diagram of the 3D printed stir bar prepared in Example 1. Detailed implementation manners

[0049] The features and advantages of the present invention can be further understood through the following detailed description in conjunction with the accompanying drawings. The provided embodiments are only illustrative of the methods of the present invention and do not limit the remaining content disclosed by the present invention in any way.

[0050]

Example 1

[0051] Preparation of 3D printed Lay-Fomm60 coated stir bar

[0052] (1) Styling design of the stir bar coating: The 3D printed stir bar coating designed in Rhinoceros software is a hollow cylinder shape. The inner diameter of the hollow cylinder is 2.0 mm, the outer diameter is 5.0 mm, and the height of the cylinder is 2.00 cm.

[0053] (2) Printing the stir bar coating by FDM 3D printing technology: According to the stir bar coating shape designed in Rhinoceros software, using an N2 3D printer (China), processing the digital model through the open-source 3D slicing software cura, and performing 3D printing on the Lay-Fomm60 material to obtain a hollow cylinder stir bar coating; the printing temperature is 210 °C, the bottom plate temperature is 60 °C, and the printing speed is 10 mm / s.

[0054] (3) Surface activation of the stir bar coating in step (2): Soak the stir bar coating in deionized water for 3 days, then soak it in methanol for 2 days. Place the activated stir bar to dry at room temperature. After the activation step is completed, wash away part of the polyvinyl alcohol in the Lay-Fomm60 material, thereby generating pore structures on the coating surface, which helps with adsorption extraction.

[0055] (4) Processing the stir bar coating in step (3) to obtain a complete stir bar: Insert a glass capillary (3.00 cm) sealed with a 2.50 cm iron wire into the stir bar coating, and the stir bar is prepared.

[0056] Scanning electron microscope images of the 3D-printed Lay-Fomm60 stir bar coating before and after activation are shown as Figure 3-6 follows. It can be seen that after activation, part of the polyvinyl alcohol was washed away, and pore structures were generated on the coating surface.

[0057] The Lay-Fomm60 stir bar coating before and after activation was characterized by Fourier transform infrared spectroscopy ATR respectively ( Figure 7 ). Compared with the Lay-Fomm60 stir bar coating before activation, the activated Lay-Fomm60 stir bar coating had a strong absorption at around 3300 cm -1 , which was the stretching vibration of -OH, indicating that part of the polyvinyl alcohol in the Lay-Fomm60 material was successfully washed away.

[0058] The water contact angles of the Lay-Fomm60 stir bar coating before and after activation were characterized respectively ( Figure 8 ). Compared with the Lay-Fomm60 stir bar coating before activation (58°), the water contact angle of the activated Lay-Fomm60 stir bar coating was 92°. After water washing and activation, the material was more hydrophobic, and the hydrophilic polyvinyl alcohol on the surface was partially washed away, exposing more hydrophobic polyurethane.

[0059]

Example 2

[0060] The adsorption effects of the 3D-printed Lay-Fomm60 coated stir bar on different pollutants were investigated

[0061] The extraction effects of the 3D-printed Lay-Fomm60 coated stir bar on the following target analytes were investigated: 4 kinds of chlorophenols (hydroquinone, o-nitrophenol, 2,4-dichlorophenol, 4-chloro-3-methylphenol), 3 kinds of benzotriazole ultraviolet absorbers (UV-329, UV-234, UV-320), and five estrogens (estradiol, hexestrol, mestranol, estrone, ethinyl estradiol). Among them, the extraction effect on benzotriazole ultraviolet absorbers was not good; the extraction efficiency of some substances in chlorophenols could reach 50%; the extraction efficiency of the five estrogens was the highest. Therefore, the extraction of chlorophenols and estrogens by the 3D-printed Lay-Fomm60 coated stir bar was further explored. The adsorption results (the ratio of the peak area of the residual liquid to the peak area of the corresponding standard solution with the same concentration) are shown as Figure 10 、 11As shown, the adsorption efficiency of the 3D-printed Lay-Fomm60-coated stir bar for 4-chloro-3-methylphenol, 2,4-dichlorophenol, and 2,4,6-trichloromethylphenol is 50-80%. However, the adsorption efficiency for o-nitrophenol is only 20%. The adsorption efficiency of the 3D-printed Lay-Fomm60-coated stir bar for 5 estrogens is above 80%. Therefore, the Lay-Fomm60-coated stir bar was selected for subsequent studies, with estrogens as the target analytes, to further investigate its extraction performance for estrogens.

[0062] The 3D-printed Lay-Fomm60-coated stir bar hardly adsorbs benzotriazole UV adsorbents, can adsorb chlorophenols (adsorption percentage is 20-80%), and has the best adsorption effect on estrogens (adsorption percentage is greater than 80%). The adsorption efficiency of the 3D-printed Lay-Fomm60-coated stir bar for 4-chloro-3-methylphenol, 2,4-dichlorophenol, and 2,4,6-trichloromethylphenol (lg P is 2.89-3.67) is 50-80%. However, the adsorption efficiency for o-nitrophenol (lg P is 1.67) is only 20%. It is speculated that hydrophobic interaction plays an important role in adsorption, and the 3D-printed Lay-Fomm60 coating is more suitable for adsorbing target analytes with lg P between 3 and 5.

[0063]

Example 3

[0064] Method for Adsorptive Extraction with a Lay-Fomm60-Coated Stir Bar Based on FDM 3D Printing and Detection of Five Estrogens by HPLC-VWD

[0065] The 3D-printed Lay-Fomm60-coated stir bar prepared in Example 1 was used for stir bar sorptive extraction coupled with HPLC-VWD to establish a method for detecting five estrogens by 3D-printed SBSE (Lay-Fomm60)-HPLC-VWD:

[0066] Including the following steps:

[0067] (1) Add 20 mL of the sample solution containing the target analyte to a 60 mL glass extraction bottle, place the 3D-printed stir bar, and extract for 90 min at a stirring rate of 350 rpm.

[0068] (2) After the extraction is completed, take out the stir bar, wipe the residual solution on the surface of the stir bar with filter paper, and then place the stir bar in a desorption tube containing 2 mL of desorbent and ultrasonically desorb for 8 min.

[0069] (3) Take 200 μL of the desorbing solution for subsequent detection using an Agilent 1260 high performance liquid chromatography - ultraviolet detector. The mobile phase composition is acetonitrile and water (52 / 48, v / v). Use an EC - C18 column (150 mm × 4.6 mm, 4 μm particle size), the flow rate is 0.70 mL / min, the wavelength of the ultraviolet detector is 200 nm, and the injection volume is 20 μL;

[0070] (4) Put the desorbed stir bar into 4 mL of methanol and ultrasonicate for 5 min for regeneration.

[0071]

Example 4

[0072] Optimize the extraction and desorption conditions of the Lay - Fomm60 coated stir bar for adsorption extraction based on FDM 3D printing

[0073] (1) First, optimize the extraction conditions of the Lay - Fomm60 coated stir bar for adsorption extraction based on FDM 3D printing. As Figures 12-17 shown, the single - variable method is used to optimize the extraction conditions one by one. The finally determined optimal conditions are that the sample solution is not adjusted for pH, 15% NaCl, the extraction time is 90 min, the desorbent is methanol, the desorption volume is 2 mL, and the desorption time is 8 min.

[0074] (2) Investigate the preparation reproducibility and service life of the Lay - Fomm60 coated stir bar for adsorption extraction based on FDM 3D printing. As Figure 18 shown, under the optimal conditions, the stir bar can be reused more than 55 times without a significant decrease in extraction recovery. The preparation reproducibility of the Lay - Fomm60 coated stir bar based on FDM 3D printing was investigated. Under the optimal experimental conditions, the extraction recoveries of six 3D - printed Lay - Fomm60 coated stir bars for five estrogens were investigated, and their relative standard deviations were calculated, as shown in Table 1. The RSDs of the six stir bars for the five target analytes are all less than 4.4%. In addition, the 3D - printed Lay - Fomm60 coated stir bar can be regenerated by ultrasonication in 4 mL of methanol for 5 min.

[0075] Table 1 Reproducibility of 3D - printed Lay - Fomm60 coated stir bar (c = 50 μg / L)

[0076]

[0077]

[0078] Note: E2 is estradiol, EE is ethinylestradiol, E1 is estrone, HES is hexestrol, MES is mestranol.

[0079] (3) Under their respective optimal conditions, the extraction of five estrogens by the 3D-printed Lay-Fomm60-coated stir bar and the commercial PDMS-coated stir bar was compared. Compared with the extraction efficiency (13 - 69%) of the commercial PDMS-coated stir bar, the 3D-printed Lay-Fomm60-coated stir bar showed higher extraction efficiency (52 - 76%) for the target estrogens.

[0080] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent replacement methods and are all included in the protection scope of the present invention.

Claims

1. A 3D printed Lay-Fomm60 coated stirring rod, characterized in that: It includes a coated cylinder and a glass capillary tube; The coated cylinder is a hollow cylinder made of Lay-Fomm60 material by 3D printing; the coated cylinder has undergone surface activation treatment; the Lay-Fomm60 material is a composite of polyurethane and polyvinyl alcohol; The glass capillary tube is embedded with iron wire and is sleeved with the coated cylinder; The method of the surface activation treatment is: soaking the coated stirring rod in deionized water, then soaking it in methanol, and then drying it at room temperature.

2. The preparation method of the stirring rod according to claim 1, characterized in that The steps are as follows: (1) Coated cylinder design: Design the 3D printed stirring rod coated cylinder to be a hollow cylinder structure; (2) Printing the stirring rod coated cylinder by fused deposition modeling 3D printing technology: For the designed coated cylinder, use a 3D printer to process the digital model through an open-source 3D slicing software, and perform 3D printing on the Lay-Fomm60 material to obtain the coated cylinder; (3) Performing surface activation on the coated cylinder in step (2); (4) Stirring rod assembly: Insert the glass capillary tube embedded with iron wire into the stirring rod coating to obtain it.

3. The preparation method according to claim 2, characterized in that: In step (1), the design software is Rhinoceros software.

4. The preparation method according to claim 2, characterized in that: In step (2), the open-source 3D slicing software is Cura.

5. Application of the Lay-Fomm60 coated stirring rod as described in claim 1 in enriching and detecting estrogens in samples.

6. The method for detecting estrogen by the Lay-Fomm60 coating stirrer according to claim 1, characterized in that It includes the following steps: (1) Add the sample solution containing the target analyte to an extraction container, put in the stirring rod, and perform extraction under stirring; (2) After the extraction is completed, take out the stirring rod, wipe the residual solution on the surface of the stirring rod with filter paper, and then place the stirring rod in a desorption tube containing a desorbent for desorption; (3) Take the desorbed solution and perform detection with a high performance liquid chromatography-ultraviolet detector.

7. The method according to claim 6, wherein: In step (2), the desorbent is methanol, the desorption volume is 2 mL, and the desorption time is 8 min.

8. The method according to claim 6, wherein: In step (3), the parameters of the high performance liquid chromatography-ultraviolet detector are: the mobile phase composition is acetonitrile and water, use an EC-C18 column, the flow rate is 0.70 mL / min, the wavelength of the ultraviolet detector is 200 nm, and the injection volume is 20 μL.

9. The method according to claim 8, characterized in that: In the mobile phase, the volume ratio of acetonitrile to water is 52 / 48; the size of the EC-C18 column is 150 mm×4.6 mm, and the particle size is 4 μm.

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