Component analysis method for toxicological safety detection of yang-strengthening heart-tonifying decoction
By performing chromatographic analysis on the detoxification control samples and co-decoction synergistic samples in the preparation process of Fuyang Yixin Decoction, and by adopting a two-way control mode and similarity calculation, the problem of the inability to effectively control the production process of traditional Chinese medicine compound preparations in the existing technology was solved. This enabled the scientific verification of the detoxification process and the stability evaluation of the overall chemical characteristics of the compound, thereby improving the safety and quality consistency of the product.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-04
- Publication Date
- 2026-04-07
- Estimated Expiration
- Not applicable · inactive patent
AI Technical Summary
Existing technologies cannot penetrate into the production process of traditional Chinese medicine compound preparations, especially the key detoxification process, to conduct effective quality control, and cannot fully reflect the stability of the overall chemical characteristics of the compound, resulting in unscientific safety evaluation and difficulty in ensuring product quality consistency.
High performance liquid chromatography was used to analyze the detoxification control samples and co-decoction synergistic samples in the preparation process of Fuyang Yixin Decoction. The toxic components and detoxification products were evaluated through a two-way control mode. The effectiveness of the detoxification process and the consistency of the overall chemical profile of the compound were ensured by combining similarity calculation and evaluation of the proportion of key components.
This enables dynamic monitoring of the production process of traditional Chinese medicine compound preparations, ensuring the scientific nature of the detoxification process and the stability of product quality, improving the controllability of the production process and the safety of the products, and guaranteeing the consistency of the clinical efficacy of the compound preparations.
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Abstract
Description
Technical Field
[0001] This invention relates to the field of traditional Chinese medicine quality control technology, specifically to a component analysis method for toxicological and safety testing of Fuyang Yixin Decoction. Background Technology
[0002] Due to their complex composition and diverse pathways of action, the quality control of traditional Chinese medicine (TCM) compound preparations has always been a key technical challenge in the modernization of TCM. Fuyang Yixin Decoction, a classic formula, contains processed Aconitum carmichaelii (a type of aconite). Aconitum carmichaelii contains toxic components such as aconitine alkaloids, and its toxicity must be addressed through specific processing or decoction techniques to hydrolyze it into low-toxicity or non-toxic derivatives to ensure clinical safety. Therefore, scientific and effective quality control and safety evaluation of such compound preparations containing toxic medicinal materials are particularly important.
[0003] Currently, the mainstream approach to quality control of traditional Chinese medicine compound preparations typically focuses on testing the final product. These methods are mostly based on pharmacopoeias or industry standards, determining the content of one or two easily measurable indicator components in the finished product, or setting residue limits for known toxic components. However, this endpoint-based testing model has inherent limitations. For safety evaluation, simply detecting the residue of toxic components in the final product cannot fundamentally distinguish whether the low residue level is due to the effectiveness of the attenuation process or because the raw materials used themselves have low levels of related components. This results in a lack of direct and effective monitoring of key attenuation process steps during production, making it impossible to confirm the stability and reproducibility of the process.
[0004] Furthermore, traditional methods that focus only on a few indicative or toxic components largely overlook the holistic nature of the synergistic effects of multiple components in traditional Chinese medicine (TCM) compound preparations. The efficacy of TCM is the result of the overall action of its complex chemical basis; the balance and stability of the compatibility among the various medicinal components within a compound preparation are prerequisites for ensuring consistent clinical efficacy. Existing quality control methods are insufficient to fully reflect the stability of the inherent chemical profile of compound preparations and cannot effectively assess the overall quality differences that may arise between batches due to minor fluctuations in raw material origin, harvesting season, or production process parameters.
[0005] Therefore, existing technologies generally lack a process quality control method that can delve into the production process, scientifically verify the effectiveness of key detoxification processes, and take into account the overall chemical stability of the compound. This constitutes a technical problem that urgently needs to be solved in this field. Summary of the Invention
[0006] To address the shortcomings of existing technologies, this invention provides a component analysis method for the toxicological and safety testing of Fuyang Yixin Decoction. This method overcomes the limitations of existing technologies, which can only perform endpoint detection and therefore cannot simultaneously and scientifically confirm the effectiveness of key detoxification processes while also considering the overall chemical profile stability of the compound. To achieve the above objectives, the present invention provides the following technical solution: a component analysis method for toxicological safety testing of Fuyang Yixin Decoction, comprising the following steps: obtaining a toxicity-reducing control sample and a co-decoction synergistic sample derived from the preparation process of Fuyang Yixin Decoction; the toxicity-reducing control sample is the decoction obtained by decocting black aconite slices separately in the raw materials of Fuyang Yixin Decoction; the co-decoction synergistic sample is the extract obtained by decocting all the raw materials of Fuyang Yixin Decoction together.
[0007] The chromatograms of the attenuated control sample and the co-decoction synergistic sample were determined separately using chromatographic methods. The preferred chromatographic method is high-performance liquid chromatography (HPLC) or ultra-high-performance liquid chromatography (UHPLC).
[0008] The chromatograms of the attenuated control samples are evaluated using a creatively designed bidirectional control mode, controlling not only the upper limit of the toxic component but also the lower limit of its attenuated derivative. This evaluation criterion includes the response value of the toxic component in the chromatogram and the response value of the attenuated derivative derived from the toxic component. Specifically, it determines whether the response value of the toxic component is not higher than a preset upper toxicity threshold and whether the response value of the attenuated derivative is not lower than a preset lower toxicity threshold. This method ensures that the detected low toxicity is due to the thorough implementation of the attenuation process and the successful conversion of the toxic component, rather than other factors, thus scientifically confirming the safety of the process. This determination is made using the following formula: ; ; in, This represents the peak area of the toxic component. The upper limit threshold for toxicity; The peak area of the attenuated derivative is [value missing]. The lower limit threshold for toxicity reduction.
[0009] The chromatograms of the co-decocted synergistic samples are evaluated. The evaluation method of this invention embodies the holistic concept of traditional Chinese medicine compound formulas and employs multi-dimensional evaluation standards. First, the chromatograms of the co-decocted synergistic samples are compared with a preset co-decocted synergistic control fingerprint chromatogram to determine if the similarity is not lower than a preset similarity threshold. This step macroscopically ensures the consistency of the chemical profiles of different batches of products. The similarity calculation uses the cosine similarity method, and its formula is:
[0010] in, For similarity, The chromatogram of the co-decoction synergistic sample is shown in the first... Response values at each data point The co-decoction synergistic control fingerprint spectrum is in the first Response values at each data point This represents the total number of data points. Secondly, to further control the balance of the components within the compound formula, the evaluation step also includes: calculating the relative peak area ratio of at least one key component peak to the internal reference peak in the chromatogram of the co-decocted synergistic sample, and determining whether the relative peak area ratio is within a preset range. This step ensures the stability of the material basis for the synergistic effect of the compound formula at a microscopic level.
[0011] Finally, based on the evaluation results of the attenuated control sample and the co-decoction synergistic sample, the production process of Fuyang Yixin Decoction is comprehensively judged. The production process of Fuyang Yixin Decoction is deemed qualified only if both the evaluation results of the attenuated control sample and the co-decoction synergistic sample meet their respective preset standards.
[0012] This invention provides a component analysis method for toxicological safety testing of Fuyang Yixin Decoction. It has the following beneficial effects: 1. This invention achieves a shift from static endpoint detection to dynamic process monitoring by moving the quality control checkpoint upstream. By acquiring and analyzing samples from the two key process nodes of "toxicity reduction control" and "co-decoction synergy," this method can gain real-time insight into the internal state of the production process, thereby enabling early detection and intervention of potential deviations in process parameters. This fundamentally improves the controllability of the production process and effectively avoids potential quality risks for the entire batch of products.
[0013] 2. This invention provides a more scientific and precise evaluation of the attenuation process. Its innovative "two-way" evaluation standard requires not only that the residual amount of toxic components be below the safety upper limit, but also that the amount of attenuated products generated reach a preset lower limit. This design can definitively prove that the reduction in toxicity stems from effective chemical transformation, rather than simply from a low initial content of the raw medicinal materials, thus providing more rigorous and reliable scientific evidence for the product's safety.
[0014] 3. This invention provides a more comprehensive and in-depth control over the overall quality of the compound preparation. By combining the comparison of the overall contour similarity of macroscopic chromatographic fingerprints with the calculation of the relative proportions of key microscopic components, this method not only ensures the overall stability of the chemical composition of different batches of products, but also ensures the compatibility and balance among the core active ingredients within the formula. This is of vital importance for ensuring the stability and consistency of the clinical efficacy of the compound preparation.
[0015] 4. This invention organically integrates safety evaluation and overall control into a unified analytical process. The method smoothly transitions from confirming the detoxification effect of a single medicinal material to evaluating the overall chemical profile of all medicinal materials after co-decoction, forming a logically coherent and data-complementary quality evaluation system. This integrated design overcomes the drawbacks of traditional methods where safety testing and quality evaluation are separated, enabling a more efficient and complete depiction of the product's overall quality.
[0016] 5. This invention significantly improves the objectivity and standardization of quality evaluation. By transforming quality standards into a series of clear, quantifiable mathematical and statistical indicators, such as peak area thresholds, similarity values, and confidence intervals for relative proportions, this method provides a purely data-driven basis for batch product release. This greatly reduces human uncertainty in the evaluation process and lays a solid technical foundation for the standardized and large-scale production of complex traditional Chinese medicine formulas like Fuyang Yixin Tang. Attached Figure Description
[0017] Figure 1 This is a schematic diagram of the overall flow of the analytical method of the present invention; Figure 2 This is a schematic diagram of the internal logic of the analysis result evaluation module. Detailed Implementation
[0018] The technical solutions in 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.
[0019] Please see the appendix Figure 1-2 This invention provides a component analysis method for toxicological safety testing of Fuyang Yixin Decoction, starting with a reference sample preparation step. To ensure the universality and stability of the established standard, multiple batches of Fuyang Yixin Decoction production samples with stable production processes and compliance with internal control standards were selected. Samples from two key stages were obtained from the production process of each batch. A "toxicity reduction control reference sample" was obtained, which was the decoction obtained by decocting only the black aconite root alone for a specified time; simultaneously, a "co-decoction synergistic reference sample" was obtained, which was the final extract obtained by decocting all eight medicinal materials together. These two types of reference samples underwent standardized pretreatment, such as precise dilution and filtration through a microporous membrane, to prepare reference sample solutions of appropriate concentration and clarity for subsequent chromatographic analysis.
[0020] Next, chromatographic analysis was performed. A method-validated high-performance liquid chromatography (HPLC) system was used to analyze multiple batches of prepared reference sample solutions. The chromatographic conditions were established to maximize the separation and visualization of various chemical components in the sample. Preferred chromatographic conditions included: using a C18 reversed-phase column; employing gradient elution as the mobile phase, for example, using acetonitrile as mobile phase B and an aqueous solution containing trace amounts of formic acid or phosphoric acid as mobile phase A; and setting a gradient elution program, flow rate, and column temperature that achieved good separation of the components. A diode array detector was used for multi-wavelength scanning and acquisition to obtain three-dimensional chromatographic information of the sample at different ultraviolet wavelengths, ensuring comprehensive information acquisition.
[0021] Finally, data processing and chromatogram generation steps were performed. Chromatographic data from multiple batches of reference samples obtained under the above chromatographic conditions were imported into the traditional Chinese medicine chromatographic fingerprint similarity evaluation software. Using the software's built-in algorithm, retention time correction and peak matching were performed on each batch of chromatograms to eliminate errors caused by minor instrument fluctuations.
[0022] After processing, "toxicity reduction control fingerprint chromatograms" and "co-decoction synergistic control fingerprint chromatograms" were generated. The control fingerprint chromatograms here are common pattern chromatograms obtained by calculating the median or mean of multiple batches of chromatograms, representing the most stable and core chemical characteristics of a qualified product at a specific process node. Based on this, through comparative analysis with high-purity reference standards, the chromatographic peaks representing toxic components and the chromatographic peaks representing their toxic transformation products were accurately identified in the "toxicity reduction control fingerprint chromatograms." This process lays the foundation for establishing a two-way evaluation standard for upper and lower limits of toxicity. Similarly, in the "co-decoction synergistic control fingerprint chromatograms," common chromatographic peaks of multiple key components from different medicinal materials were identified, providing a basis for subsequent evaluation of overall profile similarity and internal component ratios.
[0023] In the specific analysis process, the first step is to sample the batch of products to be tested. Specifically, from the production line of the batch to be tested, after the decoction process of black aconite slices is completed, a "tolerance control sample" is obtained; after the decoction process of all medicinal materials in the batch is completed, a "co-decoction synergistic sample" is obtained.
[0024] The obtained test samples must undergo the same standardized pretreatment steps as the reference samples to eliminate potential systematic errors introduced during preparation. The obtained "attenuated control sample" and "co-decoction synergistic sample" are precisely measured, diluted and brought to volume using a pre-set solvent, and then filtered through a microporous membrane. This step ultimately yields clarified "attenuated control sample solution" and "co-decoction synergistic sample solution," both of which meet the requirements for direct injection analysis.
[0025] The prepared sample solutions were sequentially injected into the chromatographic analysis system established and validated in Example 1, and run under identical chromatographic conditions. The chromatographic system separated the chemical components in the sample according to the preset gradient elution program, flow rate, column temperature, and other parameters. Simultaneously, the detector monitored the signal response of the eluent in real time. The entire analysis process was controlled by workstation software; the system automatically acquired and recorded chromatographic data, ultimately generating raw chromatograms corresponding to the "analyte attenuated control sample" and the "analyte co-decoction synergistic sample," respectively. These chromatograms graphically and intuitively presented the chemical composition profiles of the sample under these analytical conditions, serving as direct input data for subsequent quality evaluation.
[0026] For the chromatogram of the "to-be-tested attenuated control sample," this invention employs a two-way evaluation standard for analysis. The innovation of this evaluation method lies in its focus not only on the residue of toxic components but also on the degree of their transformation into less toxic components, thus more accurately reflecting the effectiveness of the attenuation process. First, the chromatographic peaks of the toxic components in the chromatogram are integrated, and the peak area must not exceed the upper limit threshold of toxicity preset based on toxicological data. This step is determined using the following formula:
[0027] in, This represents the peak area of the toxic component in the sample to be tested. This is the upper limit threshold for the peak area of the toxic component. Simultaneously, the chromatographic peaks of the attenuated derivatives derived from the toxic component are integrated in the chromatogram; their peak areas must not be lower than the lower limit threshold for attenuation obtained statistically from data of multiple batches of qualified samples. This step is determined using the following formula:
[0028] in, The peak area of the attenuated derivative in the sample to be tested is given. This is the lower limit threshold for the peak area of the attenuated product. Only when the test results of the sample meet both of the above conditions can the attenuation process of the batch of products be deemed qualified.
[0029] Next, for the chromatogram of the "co-decoction synergistic sample to be tested," this invention employs a multi-dimensional evaluation standard, comprehensively evaluating it from both macroscopic profile and microscopic proportion levels. At the macroscopic level, the overall similarity of the chromatogram to be tested is compared with the established "co-decoction synergistic control fingerprint chromatogram" to ensure the overall consistency of the chemical composition of different batches of products. The cosine similarity method is preferably used for similarity calculation, and its formula is:
[0030] in, The calculated similarity value, For the spectrum to be tested in the first Response values at each data point For comparison, the spectrum is in the first Response values at each data point This represents the total number of data points in the map. The calculated... The value must not be lower than the preset similarity threshold.
[0031] At the microscopic level, to further ensure the uniformity of dissolution of the effective components of each medicinal material in the compound formula, this invention also assesses the relative proportions of key components in the chromatogram. A common peak with stable peak shape and good reproducibility is selected as an internal reference peak in the chromatogram, and the peak area ratio of other key component peaks from different core medicinal materials to this internal reference peak is calculated. This ratio must fall within the confidence interval obtained from statistical analysis of data from multiple batches of qualified samples. This step ensures that even if the overall profile is similar, the proportions of the core active pharmaceutical substances remain within a stable and effective range.
[0032] Finally, this invention provides a final comprehensive judgment logic. Whether the production process of a batch of products is qualified depends on the results of all the above evaluation stages. That is, the production process of a batch of products is ultimately judged to be qualified only if the evaluation results of the "toxicity reduction control sample" of the batch of products meet its two-way standard, and the evaluation results of its "co-decoction synergistic sample" meet both the overall contour similarity requirement and the key component relative proportion requirement. This comprehensive judgment method organically combines safety control and overall evaluation, providing comprehensive technical support for the quality control of Fuyang Yixin Decoction.
[0033] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.
Claims
1. A component analysis method for toxicological safety testing of Fuyang Yixin Decoction, characterized in that, Includes the following steps: To obtain samples for toxicity control and co-decoction synergistic effects from the preparation process of Fuyang Yixin Decoction; The chromatograms of the attenuated control sample and the co-decoction synergistic sample were determined by chromatography. The evaluation criteria for the chromatogram of the attenuated control sample include: the response value of the toxic component in the chromatogram, and the response value of the attenuated product derived from the toxic component. Evaluate the chromatograms of the co-decoction synergistic samples; Based on the evaluation results of the detoxification control sample and the evaluation results of the co-decoction synergistic sample, a comprehensive judgment is made on the production process of Fuyang Yixin Decoction.
2. The method according to claim 1, characterized in that, The specific steps for evaluating the chromatogram of the attenuated control sample are as follows: determining whether the response value of the toxic component is not higher than a preset upper limit threshold for toxicity, and determining whether the response value of the attenuated conversion product is not lower than a preset lower limit threshold for toxicity.
3. The method according to claim 2, characterized in that, The determination is made using the following formula: ; ; in, This represents the peak area of the toxic component. The upper limit threshold for toxicity, The peak area of the attenuated derivative is [value missing]. The lower limit threshold for toxicity reduction.
4. The method according to claim 1, characterized in that, The step of evaluating the chromatogram of the co-decoction synergistic sample includes: calculating the similarity between the chromatogram of the co-decoction synergistic sample and a preset co-decoction synergistic control fingerprint chromatogram, and determining whether the similarity is not lower than a preset similarity threshold.
5. The method according to claim 4, characterized in that, The similarity calculation uses the cosine similarity method, and the formula is as follows: ； in, For similarity, Let be the response value of the chromatogram of the co-decoction synergistic sample at the i-th data point. Let be the response value of the co-decoction synergistic control fingerprint spectrum at the i-th data point, and n be the total number of data points.
6. The method according to claim 4, characterized in that, The step of evaluating the chromatogram of the co-decoction synergistic sample further includes: calculating the relative peak area ratio of at least one key component peak to the internal reference peak in the chromatogram of the co-decoction synergistic sample, and determining whether the relative peak area ratio is within a preset range.
7. The method according to claim 1, characterized in that, The detoxification control sample was the decoction obtained by decocting black aconite slices separately in water from the raw materials of Fuyang Yixin Decoction.
8. The method according to claim 1, characterized in that, The co-decoction synergistic sample is the extract obtained after decocting all the raw medicinal materials of Fuyang Yixin Decoction.
9. The method according to claim 1, characterized in that, The comprehensive judgment is as follows: the production process of Fuyang Yixin Decoction is deemed qualified if and only if the evaluation results of the detoxification control sample and the evaluation results of the co-decoction synergistic sample both meet their respective preset standards.
10. The method according to claim 1, characterized in that, The chromatographic method is high performance liquid chromatography or ultra-high performance liquid chromatography.