A method for detecting polysaccharide based on raman spectrum of metal organic framework blending composite film

Abstract

This invention discloses a polysaccharide detection method based on Raman spectroscopy of a metal-organic framework (MOF) blended composite thin film, comprising the following steps: S1, Preliminary reaction: The product to be detected is placed in a glass container, and the opening of the glass container is sealed to isolate the inner cavity of the glass container from the external environment; S2, Reaction formation: The glass container in S1 is opened, and a mercapto Raman dye and a halogen high-valence oxidant are added to the interior, allowing the product to further react with the mercapto Raman dye and the halogen high-valence oxidant to generate a polysaccharide oxidation product. This invention relates to the field of polysaccharide detection technology. This polysaccharide detection method based on Raman spectroscopy of a MOF blended composite thin film has the advantages of not requiring enzyme reactions and complex chemical synthesis and labeling. By introducing a metal-organic framework blended with a polymer, the surface aggregation of gold nanoparticles is prevented and the surface-enhanced Raman scattering intensity is improved, thereby enhancing the stability and sensitivity of the detection.

Description

Technical Field

[0001] This invention relates to the field of polysaccharide detection technology, specifically to a polysaccharide detection method based on Raman spectroscopy of metal-organic framework blended composite thin films. Background Technology

[0002] Polysaccharides lack distinct ultraviolet or fluorescence spectra, exhibiting weak characteristic spectral signals, making direct detection difficult. Currently, laboratories primarily rely on large instruments (such as chromatographic instruments) and the detection of hydrolyzed monosaccharides. These methods are time-consuming, fail to directly reflect the actual polysaccharide composition, and make on-site monitoring of polysaccharide components challenging. Polysaccharides are major components of plants, and many possess significant pharmacological activities. Therefore, polysaccharides are used as quality indicators for many traditional Chinese medicines, and polysaccharide content monitoring is essential during the pharmaceutical process. For example, both Codonopsis pilosula and Polygonatum sibiricum contain high levels of polysaccharides. Codonopsis pilosula and Polygonatum sibiricum polysaccharides possess antioxidant, cardioprotective, neuroprotective, and immunomodulatory effects, making them the active pharmaceutical ingredients of these herbs. Currently, polysaccharide determination typically employs ultraviolet spectrophotometry based on the phenol-sulfuric acid method or the anthrone-sulfuric acid method. This method is cumbersome and time-consuming, failing to meet the rapid analytical needs of pharmaceutical process monitoring. Furthermore, this method requires the use of large quantities of highly corrosive concentrated sulfuric acid, posing potential safety hazards.

[0003] The FDA's 2004 guidance on Process Analytical Technology (PAT) promoted the application of PAT technology in the manufacturing process of traditional Chinese medicine. PAT technologies based on spectroscopic analysis methods, such as near-infrared spectroscopy and Raman spectroscopy, can provide real-time, economical, and non-destructive determination of key quality parameters in the manufacturing process. However, water has a strong signal in near-infrared spectroscopy, so when using near-infrared spectroscopy to determine the chemical components in aqueous solutions, the near-infrared absorption signal of water can mask the signals of other components, which is not conducive to the determination of these components. In contrast, water has a weaker signal in Raman spectroscopy, making it suitable for determining the content of chemical components in aqueous solutions. Currently, Raman spectroscopy methods for determining monosaccharides and disaccharides in products such as honey and beverages have been reported in the literature, but there are no reported Raman spectroscopy methods for determining polysaccharides.

[0004] Currently, the detection of polysaccharides in plant extracts mainly involves treating the extracts through biological or chemical reactions to determine the polysaccharide content. These methods primarily include HPLC, ELISA, dinitrosalicylic acid (DNAS) method, and phenol-sulfuric acid method. Existing technologies such as HPLC, ELISA, DNAS, and phenol-sulfuric acid are well-developed. HPLC detection devices offer high precision, but this method is time-consuming and requires complex pretreatment. ELISA detection methods have low detection limits and high sensitivity, but the detection steps are cumbersome, requiring multiple washing steps. The DNAS and phenol-sulfuric acid methods are very complex, and traditional methods cannot reflect the components in the fermentation broth in real time, resulting in a general lag in the detection results. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a polysaccharide detection method based on Raman spectroscopy of metal-organic framework blended composite thin films, which solves the problem that traditional methods cannot reflect the components in fermentation broth in real time and that the detection results generally have a lag.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution: a method for detecting polysaccharides based on Raman spectroscopy of metal-organic framework blended composite thin films, comprising the following steps: S1, preparatory reaction: take the product to be detected and store it in a glass container, seal the opening of the glass container, and isolate the inner cavity of the glass container from the external environment.

[0007] S2, reaction formation: Open the glass container in S1, add mercapto Raman dye and halogen high-valence oxidant into it, so that the product reacts further with mercapto Raman dye and halogen high-valence oxidant to form polysaccharide oxidation product;

[0008] S3, Preliminary detection: The reaction product from S2 is drawn into the dropper and added to the surface of gold nanoparticles on the metal-organic framework hybrid composite film;

[0009] S4. On-site testing: Set the parameters of the Raman spectroscopy equipment and use surface-enhanced Raman scattering technology on the metal-organic framework blended composite film in S3 to obtain the polysaccharide content in the functional plant extract.

[0010] S5. Record Information: Place the sample remaining after testing into a glass container, attach a label with the recorded information to the surface of the glass container, and then seal the opening of the glass container to isolate the inner cavity of the glass container from the external environment.

[0011] Furthermore, in S1, the prepared product is 25 mg.

[0012] Furthermore, in both S1 and S5, the glassware is stored in a dark place.

[0013] Furthermore, in step S2, after adding mercapto Raman dye and halogen high-valence oxidant to the inside of the glassware, it is gently shaken.

[0014] Furthermore, in both S1 and S5, the glassware used is colorless and sterile glassware.

[0015] Furthermore, in S5, the information recorded on the label includes sample type, sample name, preparation time, storage time, detection time, and polysaccharide detection results.

[0016] Furthermore, in S5, the distance between the multiple glass containers is 5-10 cm.

[0017] Furthermore, in both S1 and S5, sterile silicone sealing blocks are used to seal the glassware.

[0018] Furthermore, in S3, the dropper is a disposable plastic dropper.

[0019] Compared with existing technologies, the advantages of this invention are: the method is simple, convenient, fast, and sensitive, and does not require enzyme reactions or complex chemical synthesis and labeling. By introducing metal-organic frameworks and polymer blends, and surface-modifying gold nanoparticles, the surface aggregation of gold nanoparticles is prevented and the surface-enhanced Raman scattering intensity is improved, thereby enhancing the stability and sensitivity of detection. The operation is simple and suitable for large-scale production applications. Raman spectroscopy can provide rapid, simple, repeatable, and, more importantly, non-destructive qualitative and quantitative analysis. Samples can be directly measured through fiber optic probes or through glass, quartz, and optical fibers. Different types of polysaccharides can be distinguished and identified, thereby enabling quality control of polysaccharides. This invention requires short measurement and analysis time, is fast and simple, requires very small sample amounts, and has low detection costs. Attached Figure Description

[0020] Figure 1 This is a flowchart of a polysaccharide detection method based on Raman spectroscopy of metal-organic framework blended composite thin films. Detailed Implementation

[0021] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0022] Please see Figure 1 The present invention provides a technical solution: a method for detecting polysaccharides based on Raman spectroscopy of metal-organic framework blended composite thin films, comprising the following steps: S1, preparatory reaction: take the product to be detected and store it in a glass container, seal the opening of the glass container, and store it in isolation from the external environment.

[0023] S2, reaction formation: Open the glass container in S1, add mercapto Raman dye and halogen high-valence oxidant into it, so that the product reacts further with mercapto Raman dye and halogen high-valence oxidant to form polysaccharide oxidation product;

[0024] S3, Preliminary detection: The reaction product from S2 is drawn into the dropper and added to the surface of gold nanoparticles on the metal-organic framework hybrid composite film;

[0025] S4. On-site testing: Set the parameters of the Raman spectroscopy equipment and use surface-enhanced Raman scattering technology on the metal-organic framework blended composite film in S3 to obtain the polysaccharide content in the functional plant extract.

[0026] S5. Record Information: Place the sample remaining after testing into a glass container, attach a label with the recorded information to the surface of the glass container, and then seal the opening of the glass container to isolate the inner cavity of the glass container from the external environment.

[0027] Polysaccharides lack obvious ultraviolet or fluorescence spectra, and their characteristic spectral signals are weak, making direct detection difficult. Currently, laboratories mainly rely on large instruments (such as chromatography instruments) and the detection of hydrolyzed monosaccharides. However, the detection cycle is long, and it is difficult to directly reflect the actual polysaccharide composition. It is also difficult to monitor polysaccharide components on-site.

[0028] Polysaccharides are the main components of plants, and many polysaccharides possess important pharmacological activities. Therefore, polysaccharides are used as quality indicators for many traditional Chinese medicines, and polysaccharide content monitoring is required during the manufacturing process of traditional Chinese medicines. For example, both Codonopsis pilosula and Polygonatum sibiricum contain high levels of polysaccharides. Codonopsis pilosula polysaccharides and Polygonatum sibiricum polysaccharides have antioxidant, cardioprotective, neuroprotective, and immunomodulatory effects, and are the active medicinal components of Codonopsis pilosula and Polygonatum sibiricum.

[0029] Traditional methods for determining polysaccharides typically employ ultraviolet spectrophotometry based on the phenol-sulfuric acid method or the anthrone-sulfuric acid method. These methods are cumbersome and time-consuming, failing to meet the rapid analytical needs of pharmaceutical process monitoring. Furthermore, these methods require the use of large quantities of highly corrosive concentrated sulfuric acid, posing potential safety hazards.

[0030] Polysaccharides are formed by the condensation and dehydration of multiple monosaccharide molecules, and are a class of sugar substances with complex and large molecular structures.

[0031] All carbohydrates and their derivatives that meet the definition of macromolecular compounds are called polysaccharides. Polysaccharides exist in plants and animals. In higher plants and algae, they are components of the cell wall or cell interior; in bacteria and fungi, they may be both cellular components and metabolic products.

[0032] Polysaccharides have a large molecular weight, and their properties differ significantly from monosaccharides and oligosaccharides. Studies have found that polysaccharides are one of the bioactive components of many traditional Chinese medicines, including Poria cocos, Astragalus membranaceus, Polyporus umbellatus, Lycium barbarum, Bupleurum chinense, Ginseng, Lentinus edodes, Trametes versicolor, Tremella fuciformis, Ganoderma lucidum, and Cordyceps sinensis. These polysaccharides possess immunomodulatory, anti-radiation, anticoagulant, hypoglycemic, and hypolipidemic effects, and their physiological activities are closely related to their molecular weight, molecular weight distribution, and chemical structure.

[0033] Traditional methods for determining the molecular weight of polysaccharides include gel permeation chromatography, infrared spectroscopy and ultraviolet spectroscopy, nuclear magnetic resonance spectroscopy, mass spectrometry and gas chromatography-mass spectrometry. In the structural analysis of polysaccharides, X-ray diffraction, capillary electrophoresis, nuclear magnetic resonance, optical rotation and circular dichroism spectroscopy, fast atom bombardment mass spectrometry, gas chromatography-mass spectrometry, and atomic force microscopy are commonly used. Among these, optical rotation is the most common instrument, but it can only be used for the simplest analyses.

[0034] This method is simple, convenient, rapid, and sensitive, and does not require enzyme reactions or complex chemical synthesis and labeling. By introducing metal-organic frameworks and polymer blends, and surface-modifying gold nanoparticles, the surface aggregation of gold nanoparticles is prevented and the surface-enhanced Raman scattering intensity is improved, thereby enhancing the stability and sensitivity of the detection.

[0035] Furthermore, this Raman spectroscopy method provides rapid, simple, repeatable, and, more importantly, non-destructive qualitative and quantitative analysis. Samples can be measured directly through fiber optic probes or through glass, quartz, and optical fibers. It can distinguish and identify different types of polysaccharides, thereby enabling quality control of polysaccharides. This invention requires short measurement and analysis time, is rapid and simple, requires very small sample amounts, and has low detection costs.

[0036] In S1, the prepared product is 25 mg.

[0037] In both S1 and S5, the glassware is stored in a dark place to prevent it from decomposing when exposed to light and to prevent the temperature from rising due to prolonged exposure to light, which would promote bacterial growth.

[0038] In step S2, after adding mercapto Raman dye and halogen high-valence oxidant to the inside of the glass container, it is gently shaken to accelerate the reaction efficiency.

[0039] In both S1 and S5, the glassware used is colorless and sterile, which makes it easy for staff to observe the reaction.

[0040] In step S5, the information recorded on the label includes sample type, sample name, preparation time, storage time, detection time, and polysaccharide detection results, which is beneficial for data organization in the later stages.

[0041] In step S5, the distance between multiple glass containers is 5-10 cm to prevent them from colliding with each other.

[0042] In both S1 and S5, sterile silicone sealing blocks are used to seal the glassware to prevent oxidation and deterioration.

[0043] In step S3, the dropper is a disposable plastic dropper to avoid mixing of samples later.

[0044] During operation, the product to be tested is placed in a glass container, and the opening of the glass container is sealed to isolate the inner cavity from the external environment. The glass container is then opened, and a mercapto Raman dye and a halogen high-valence oxidant are added inside. The product reacts further with the mercapto Raman dye and the halogen high-valence oxidant to generate a polysaccharide oxidation product. The reaction product is then aspirated with a dropper and added to the surface of gold nanoparticles on a metal-organic framework (MOF) composite film. Raman spectroscopy parameters are set, and surface-enhanced Raman scattering (SERS) is applied to the MOF composite film in S3 to obtain the polysaccharide content in the functional plant extract. The sample remaining after testing is placed in a glass container, and a label for recording information is attached to the surface of the glass container. The opening of the glass container is then sealed to isolate the inner cavity from the external environment. This method is simple, convenient, rapid, and sensitive, and does not require enzyme reactions or complex chemical synthesis and labeling. It prevents surface aggregation of gold nanoparticles and improves the surface-enhanced Raman scattering intensity, thereby enhancing the stability and sensitivity of the detection.

[0045] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such process, method, article, or apparatus.

[0046] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for detecting polysaccharides based on Raman spectroscopy of metal-organic framework blended composite thin films, comprising the following steps, characterized in that: S1. Preliminary reaction: Take the product to be tested and store it in a glass container. Seal the opening of the glass container to isolate the inner cavity of the glass container from the external environment. S2, reaction formation: Open the glass container in S1, add mercapto Raman dye and halogen high-valence oxidant into it, so that the product reacts further with mercapto Raman dye and halogen high-valence oxidant to form polysaccharide oxidation product; S3, Preliminary detection: The reaction product from S2 is drawn into the dropper and added to the surface of gold nanoparticles on the metal-organic framework hybrid composite film; S4. On-site testing: Set the parameters of the Raman spectroscopy equipment and use surface-enhanced Raman scattering technology on the metal-organic framework blended composite film in S3 to obtain the polysaccharide content in the functional plant extract. S5. Record Information: Place the sample remaining after testing into a glass container, attach a label with the recorded information to the surface of the glass container, and then seal the opening of the glass container to isolate the inner cavity of the glass container from the external environment.

2. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In S1, the prepared product is 25 mg.

3. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In both S1 and S5, the glassware is stored in a dark place.

4. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In step S2, a mercapto Raman dye and a halogen high-valence oxidant are added to the inside of the glassware and then gently shaken.

5. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In both S1 and S5, the glassware used is colorless and sterile glassware.

6. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In step S5, the information recorded on the label includes sample type, sample name, preparation time, storage time, detection time, and polysaccharide detection results.

7. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In step S5, the distance between multiple glass containers is 5-10 cm.

8. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In both S1 and S5, sterile silicone sealing blocks are used to seal the glassware.

9. The method for polysaccharide detection based on Raman spectroscopy of metal-organic framework blended composite thin films according to claim 1, characterized in that: In S3, the dropper is a disposable plastic dropper.

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