A rapid detection device for ergosterol in fruiting bodies

Abstract

This invention provides a rapid detection device for ergosterol in fruiting bodies, comprising a PCR tube and an assembly disposed within the PCR tube. The assembly includes a first fixation tube, a second fixation tube, and a chamber located inside the first and second fixation tubes. The chamber is formed by sealing with a hollow fiber membrane and is filled with the analyte. Both the first and second fixation tubes have a larger upper opening and a smaller lower opening, with the tube diameter gradually decreasing from top to bottom. Furthermore, the outer diameter of the upper end of the second fixation tube is smaller than the inner diameter of the lower end of the first fixation tube. The inner diameter of the upper opening of the second fixation tube is larger than the outer diameter of the chamber, and the inner diameter of the lower opening is smaller than the outer diameter of the chamber. This invention has the advantages of requiring less sample and having a simpler procedure.

Description

Technical Field

[0001] This utility model relates to the field of solid phase sample extraction technology, and in particular to a rapid detection device for ergosterol in fruiting bodies. Background Technology

[0002] Ergosterol is primarily found in the fungal kingdom, including yeasts (such as *Saccharomyces cerevisiae*), molds (such as *Aspergillus* and *Penicillium*), and macrofungi (such as mushrooms and *Ganoderma lucidum*). Macrofungi consist of mycelium and fruiting bodies, with a typical fruiting body composed of a cap and stipe; ergosterol is widely present in the fruiting body. It is noteworthy that higher plants and animals do not synthesize ergosterol. The main sterol in animals is cholesterol, while in plants it is mainly sitosterol and stigmasterol. This specific distribution makes ergosterol an extremely important biomarker.

[0003] Ergosterol has a wide range of applications, playing a crucial role in everything from basic biological research to industrial production, pharmaceuticals, and environmental monitoring. 1. As a precursor to vitamin D2 (pharmaceutical and health product industry). 2. As a target for antifungal drugs (pharmaceutical field). 3. As an indicator of fungal biomass (microbiology and environmental science); detecting ergosterol content in a sample allows for rapid and accurate quantification of the total amount of live fungi. 4. Applications in functional foods and traditional Chinese medicine research: The active components of many medicinal fungi (such as Ganoderma lucidum, Poria cocos, and Cordyceps sinensis) are related to their sterols. Ergosterol, as one of the main sterols in these fungi, is often used as an indicator of the quality of medicinal materials.

[0004] Modern analytical chemistry, especially with high-performance liquid chromatography (HPLC) as its core, combined with ultraviolet light, diode arrays, and even advanced mass spectrometry detection technologies, has made the detection of ergosterol more sensitive, rapid, and automated. This will undoubtedly further promote its widespread application in fields such as food safety, environmental assessment, and new drug development.

[0005] Solid-phase extraction (SPE) is a sample pretreatment technique. With the rapid development of modern instruments, the requirements for sample pretreatment have also increased. SPE, with its unique advantages, plays an increasingly important role in sample separation and purification, distinguishing it from traditional liquid-liquid extraction methods. This technique, developed by combining liquid-solid extraction and column-liquid chromatography, can purify samples, transfer target substances from the sample matrix, and enrich and concentrate trace amounts of target substances, thereby improving the sensitivity and accuracy of sample analysis. Currently, the most commonly used form of SPE is the solid-phase extraction column. However, the large column volume and high sample volume required limit the application of this technique. Therefore, miniaturization has become a research hotspot in the field of solid-phase extraction.

[0006] Furthermore, laboratories often separate sample pretreatment—filtration, sample transfer, and solid-phase extraction—into two steps, including setting up filtration devices and adding transfer equipment. This makes sample testing complex and cumbersome, and the use of multiple instruments can easily lead to more contamination and increase the uncertainty of the results. In addition, the prolonged exposure of the filtration process to air can cause evaporation or oxidation of the filtered liquid, as well as residues in the receiving container and pipettes. The more instruments used, the more unnecessary contamination is likely to occur, making it difficult to guarantee the extraction effect and seriously affecting the accuracy of the test results. Utility Model Content

[0007] This invention provides a rapid detection device for ergosterol in fruiting bodies, which solves at least one of the problems mentioned in the background art.

[0008] The technical solution of this utility model is as follows:

[0009] A rapid detection device for ergosterol in fruiting bodies includes a PCR tube and an assembly disposed within the PCR tube. The assembly includes a first fixing tube, a second fixing tube, and a chamber located inside the first fixing tube and the second fixing tube.

[0010] The chamber is made of a hollow fiber membrane and is filled with the test sample.

[0011] Both the first and second fixed pipes have a shape with a larger upper opening and a smaller lower opening, with the pipe diameter gradually decreasing from top to bottom; and the outer diameter of the upper end of the second fixed pipe is smaller than the inner diameter of the lower end of the first fixed pipe; the inner diameter of the upper opening of the second fixed pipe is larger than the outer diameter of the cabin, and the inner diameter of the lower opening is smaller than the outer diameter of the cabin.

[0012] Furthermore, the cabin has a length of 0.8-1.2 cm, an outer diameter of 0.5-1 mm, and a wall thickness of 0.1-0.2 mm.

[0013] Furthermore, the hollow fiber membrane is a hydrophilic or hydrophobic hollow fiber membrane.

[0014] Furthermore, the object to be tested is a powder of medicinal fungi.

[0015] Furthermore, the powder filling amount is 1–5 mg.

[0016] Furthermore, the two ends of the chamber are sealed by instantaneous flame melting to form a completely sealed columnar microextraction chamber.

[0017] Furthermore, the length of the portion of the cabin into which the second fixing tube can be inserted is 1 / 5 to 1 / 10 of the cabin length.

[0018] This invention provides a hollow fiber extraction chamber that is simple in structure, low in cost, and easy to use, enabling efficient and accurate extraction of ergosterol, a biomass marker of fungi.

[0019] Traditional solid-phase extraction processes are cumbersome, typically involving multiple independent steps such as sample weighing, transfer, filtration, and column chromatography, each of which carries the risk of sample loss and contamination. This invention, through structural design, directly encapsulates the sample to be tested (medicinal fungal powder) within a miniature hollow fiber membrane "chamber."

[0020] Extremely simple operation, one-step process: Users simply place the pre-packaged microcapsule into the PCR tube (i.e., the extraction container) and add the extraction solvent. This completely eliminates the need for complex steps such as setting up a filtration device and using pipettes to transfer liquids. It significantly improves work efficiency and lowers the technical barrier for operators.

[0021] A fully enclosed system eliminates contamination and loss: Because the analyte is completely sealed within the chamber, the entire extraction process takes place in a highly closed system. This maximizes sample isolation, effectively preventing cross-contamination and sample adsorption loss caused by residues in the glassware and during transfer steps. This is crucial for trace analysis (such as the detection of low levels of ergosterol), ensuring the accuracy and reliability of the analytical results.

[0022] Traditional solid-phase extraction columns are large in volume and require large sample volumes, which is a significant limitation when processing precious Chinese medicinal materials or trace biological samples. This invention, however, offers significant advantages in miniaturization and high throughput: it is suitable for precious trace samples and aligns with modern analytical trends. Attached Figure Description

[0023] Figure 1 This is a combined sectional view of the device of this utility model;

[0024] Figure 2 This is a schematic diagram of the exploded structure of the device of this utility model.

[0025] Figure 3 This is a schematic diagram of a partial structural assembly of the present invention and its details. Detailed Implementation

[0026] The technical solutions of the present utility model will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present utility model. Obviously, the described embodiments are only some embodiments of the present utility model, and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present utility model without creative effort are within the protection scope of the present utility model.

[0027] In the description of this utility model, it should be noted that the terms "upper," "lower," "inner," "outer," "top / bottom," etc., indicating the orientation or positional relationship are based on the orientation or positional relationship shown in the accompanying drawings, and are only for the convenience of describing this utility model and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this utility model. Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance.

[0028] This invention, based on the principle of solid-phase extraction and leveraging the selective enrichment of specific substances by adsorbent materials after directional treatment, proposes a rapid detection device for ergosterol in fruiting bodies through structural design. Figure 1-3 As shown, the components of this device include centrifuge tubes (in this embodiment, the centrifuge tubes are specifically PCR tubes 1) and an assembly disposed inside the PCR tubes 1. The assembly includes a first fixing tube 2, a second fixing tube 3, and a chamber 4 located inside the first fixing tube 2 and the second fixing tube 3.

[0029] The chamber 4 is a hydrophilic or hydrophobic hollow fiber membrane with a length of about 1 cm, an outer diameter of 0.5–1 mm, and a wall thickness of 0.1–0.2 mm. The hollow fiber membrane is filled with medicinal fungal powder 5, and the filling amount can be selected as 1–5 mg or other required amounts. The two ends of the chamber 4 are sealed by instantaneous flame melting to form a completely sealed columnar microextraction chamber.

[0030] The first fixing tube 2 has a larger upper opening and a smaller lower opening, with its diameter gradually decreasing from top to bottom. Similarly, the second fixing tube 3 also has a larger upper opening and a smaller lower opening, with its diameter gradually decreasing from top to bottom. Furthermore, the outer diameter of the upper end of the second fixing tube 3 is smaller than the inner diameter of the lower end of the first fixing tube 2, allowing the second fixing tube 3 to be inserted into the first fixing tube 2 from top to bottom and preventing it from detaching from the lower end of the first fixing tube 2.

[0031] The inner diameter of the upper opening of the second fixed pipe 3 is larger than the outer diameter of the cabin 4, and the inner diameter of the lower opening is smaller than the outer diameter of the cabin 4, so that a part of the cabin 4 can be inserted into the upper opening of the second fixed pipe 3.

[0032] Preferably, the length of the portion of the chamber 4 that can be inserted into the second fixed tube 3 is 1 / 5 to 1 / 10 of the length of the chamber 4. In this way, the lower end of the chamber 4 can be fixed inside the second fixed tube 3, while most of the upper part can be fully exposed inside the cavity of the first fixed tube 2, making it easier for the solution to enter the chamber 4 for extraction.

[0033] During assembly, after the medicinal powder 5 is filled, both ends of the chamber 4 are sealed by instantaneous flame melting to form a completely sealed columnar microextraction chamber. This columnar microextraction chamber is then inserted into the upper opening of the second fixing tube 3, and the second fixing tube 3 is then inserted from top to bottom into the first fixing tube 2, forming... Figure 3 The assembly process is as follows. Finally, the entire assembly is placed into PCR tube 1 to form... Figure 1 The assembly state.

[0034] During extraction, simply inject the extraction solution into the opening of the first fixed tube 2, soak for a sufficient time, and then remove the solid portion from the PCR tube 1. The remaining extraction solution in the PCR tube 1 is used for the next step of analysis.

[0035] This invention requires very little sample and has a high resource utilization rate: the micro-chamber of this invention is extremely small in size (length 0.8-1.2cm, outer diameter 0.5-1mm), requiring only 1-5mg of medicinal powder to complete the extraction. This extremely low sample consumption makes it possible to perform non-destructive or minimal-destructive testing on precious medicinal materials such as Ganoderma lucidum and Cordyceps sinensis, greatly saving testing costs.

[0036] Perfectly adapted to high-throughput platforms: The core carrier of this microcompartment is a standard PCR tube. This means it can be directly integrated into the standardized workflows of modern molecular biology laboratories. Utilizing multichannel pipettes, PCR plate centrifuges, and automated liquid handling workstations, it can easily achieve high-throughput parallel processing at the 96-well or even 384-well plate scale, making it particularly suitable for large-scale drug material control screening, fungal fermentation process monitoring, or environmental fungal contamination investigations, resulting in an order-of-magnitude improvement in analytical efficiency.

[0037] Hollow fiber membranes possess a uniform microporous structure. The extraction solvent can freely penetrate the membrane wall and enter the chamber to soak the herbal powder, dissolving the target compound, ergosterol. Simultaneously, the dissolved ergosterol molecules can diffuse through the membrane pores into the external solvent. The herbal powder itself is effectively confined within the chamber, resulting in a relatively clean extract and simplifying the purification process.

[0038] Overcoming the challenges of powder handling: Fungal medicinal materials are typically hard in texture, and after being pulverized, they easily generate static electricity and adhere to the container walls, leading to weighing errors. This device pre-positions and standardizes the weighing and packaging steps for microgram-level powders, avoiding the difficulties of directly handling trace amounts of powder in a conventional laboratory environment, fundamentally reducing weighing errors and adsorption losses during operation.

[0039] In summary, this utility model's "rapid detection device for ergosterol in fruiting bodies" transforms a complex and open sample pretreatment process into a simple, closed, and efficient "one-pot" operation through an integrated design. It not only significantly improves work efficiency but also reduces the risk of contamination and conserves valuable samples.

[0040] The above description is only a preferred embodiment of the present utility model, but the design concept of the present utility model is not limited thereto. Any non-substantial modifications made to the present utility model by those skilled in the art within the scope of the technology disclosed in the present utility model using this concept shall be deemed as an infringement of the protection scope of the present utility model.

Claims

1. A rapid detection device for ergosterol in fruiting bodies, characterized in that, The invention includes a PCR tube and an assembly disposed within the PCR tube, the assembly comprising a first fixing tube, a second fixing tube, and a chamber located inside the first fixing tube and the second fixing tube; The chamber is made of a hollow fiber membrane and is filled with the test sample. Both the first and second fixed pipes have a shape with a larger upper opening and a smaller lower opening, with the pipe diameter gradually decreasing from top to bottom; and the outer diameter of the upper end of the second fixed pipe is smaller than the inner diameter of the lower end of the first fixed pipe; the inner diameter of the upper opening of the second fixed pipe is larger than the outer diameter of the cabin, and the inner diameter of the lower opening is smaller than the outer diameter of the cabin.

2. The rapid detection device for ergosterol in fruiting bodies according to claim 1, characterized in that: The cabin has a length of 0.8-1.2 cm, an outer diameter of 0.5-1 mm, and a wall thickness of 0.1-0.2 mm.

3. The rapid detection device for ergosterol in fruiting bodies according to claim 1, characterized in that: The hollow fiber membrane is a hydrophilic or hydrophobic hollow fiber membrane.

4. The rapid detection device for ergosterol in fruiting bodies according to claim 1, characterized in that: The test substance is a powder of medicinal fungi.

5. The rapid detection device for ergosterol in fruiting bodies according to claim 4, characterized in that: Powder filling amount: 1–5 mg.

6. The rapid detection device for ergosterol in fruiting bodies according to claim 1, characterized in that: The two ends of the chamber are sealed by instantaneous flame melting, forming a completely sealed columnar microextraction chamber.

7. The rapid detection device for ergosterol in fruiting bodies according to claim 1, characterized in that: The length of the portion of the cabin into which the second fixed tube can be inserted is 1 / 5 to 1 / 10 of the cabin length.

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