Reduning injection (RDN) is a traditional chinese medicine injection widely used in clinical practice. In this study, qualitative and quantitative analysis of RDN were conducted by UPLC-Orbitrap MS/MS simultaneously. Totally 118 compounds were identified and 34 compounds were quantified in RDN. The method with completed method validation proved the high sensitivity and efficiency of the method and it was applied to quantify compounds in RDN. Multivariate statistical analysis method selected 11 key variables that affect the content consistency of RDN. 20 batches with high biological potency were screened by cox-2 enzyme activity assay. Spectrum-effect relationship analysis and multivariate statistical analysis showed that 7 batches were high-quality selected after comprehensive quality evaluation and 9 compounds were key indicators for screening it. This strategy including fingerprint, qualitative analysis and multiple-component quantification could be well applied to modern quality evaluation of RDN, which could be valuable for the further quality control of more other traditional Chinese medicines.

BACKGROUND: YiXinShu capsule (YXSC), originally from the classical TCM formula named \"Sheng-Mai-San\", has been extensively utilized in clinic for the treatment of cardiovascular diseases. However, there were few reports about the quality assessment of YXSCs both internationally and domestically.
OBJECTIVE: The objective was to develop a multi-strategy platform incorporating systematic quantitative fingerprint analysis and antioxidant activity determination, with chemometric analysis and bivariate correlation analysis as the auxiliary approaches, to assess and monitor the quality of YXSCs.
METHODS: Firstly, according to the Chinese Pharmacopoeia (2020 edition), 12 key indicator components from seven herb medicines were quantified by HPLC method. Then, three-dimensional fingerprints comprising five-wavelength fusion fingerprint (FWF-FP), electrochemical fingerprint (EC-FP) and Differential Scanning Calorimetry fingerprint (DSC-FP) were established to assess and monitor YXSCs using systematically quantified fingerprint method (SQFM) and principal component analysis (PCA). Moreover, by integrating the analysis of the three-dimensional fingerprints, the quality of YXSCs from different batches was effectively screened. Finally, the antioxidant activity of this TCM was assessed through DPPH and ABTS methods, and the L-ascorbic acid equivalent antioxidant capacity (AEAC) values were compared to evaluate the antioxidant activities of the two methods. A Partial Least Squares (PLS) model was used to develop the spectrum-activity relationship between FWF-FP and AEAC, and a bivariate correlation analysis (BCA) was used to assess the correlation between FWF-FP and EC-FP.
RESULTS: The key indexes including tanshinone I, tol, toe, Atp, first exothermic peak, and second exothermic peak can differentiate between various batches of YXSCs based on their three-dimensional fingerprint profiles. The integration evaluation results from 42 batches of YXSCs were categorized into 2-5 grades, indicating good quality consistency across different batches. In vitro studies have indicated a significant antioxidant activity capacity of YXSCs. The PLS model revealed that 37 out of the 41 fingerprint peaks exhibited antioxidant activity. The overall trend of BCA was consistent with PLS model results.
CONCLUSIONS: This research presents a scientific and holistic strategy for the quality consistency evaluation of YXSCs, thereby offering an effective approach for the thorough evaluation of TCMs.

This paper aims to study the spectrum-effect relationship between the fingerprints before and after salt processing of Dipsacus asper and the efficacy of warming and tonifying kidney Yang and find the main active components against kidney Yang deficiency before and after salt processing of D. asper, so as to provide the basis for clarifying the effect of salt processing on kidney Yang deficiency. The HPLC fingerprint before and after salt processing of D. asper was established by the HPLC-DAD. 15 common peaks were obtained, and 11 components were identified. The content changes of various components in rat serum were detected, and the difference in efficacy before and after salt processing was compared. The results of pharmacological experiments showed that salt processing of D. asper could enhance the kidney index. At the same dose, there was a significant difference between the raw D. asper and D. asper after salt processing groups. Compared with the model group, the contents of ACTH, cAMP, CORT, E_2, GH, Na~+-K~+-ATPase, T, and T4 in the serum of rats in the administration group increased to a certain extent, and the contents of cGMP and TNF-α decreased to a certain extent. Among them, there were significant differences in the above indexes in the serum of rats in the high-dose group of raw D. asper, middle-dose group of D. asper after salt processing, high-dose group of D. asper after salt processing, and the positive drug group. The overall results showed that D. asper after salt processing was more effective than raw D. asper in preventing kidney yang deficiency. The efficacy of D. asper was evaluated by grey correlation analysis, entropy method, and Pearson correlation analysis, and the components of D. asper after salt processing against kidney yang deficiency were screened out. According to the results of correlation degree ranking, the components with increased ranking before and after salt processing of D. asper were loganin, chlorogenic acid, dipsacoside A, asperosaponin Ⅵ, caffeic acid, and isochlorogenic acid B. It was preliminarily speculated that these compounds may be the potential pharmacodynamic components for the treatment of kidney yang deficiency before and after salt processing of D. asper. The changing components before and after the salt processing of D. asper were determined, which proved that D. asper after salt processing was superior to D. asper in the treatment of kidney yang deficiency. The spectrum-effect relationship between the efficacy of D. asper before and after salt processing and the treatment of kidney yang deficiency was established, which laid a foundation for the subsequent study on the pharmacodynamic components and molecular mechanism of salt processing of D. asper.

OBJECTIVE: Farfarae Flos has the effect of cough suppression and phlegm elimination, with cough suppression as the main function. Studies have revealed that certain components of Farfarae Flos may be related to its cough suppressant effect, and some components have been confirmed to have cough suppressant activity. However, the antitussive material basis of Farfarae Flos has not been systematically elucidated. This study aims to elucidate the group of active ingredients in Farfarae Flos with cough suppressant activity by correlating the high performance liquid chromatography (HPLC) fingerprint of Farfarae Flos extract with its cough suppressant activity.
METHODS: HPLC was used to establish the fingerprint profiles of 10 batches of Farfarae Flos extract and obtain their chemical composition data. Guinea pigs were selected as experimental animals and the citric acid-induced cough model was used to evaluate the antitussive efficacy data of 10 batches of Farfarae Flos extract. SPF-grade healthy male Hartley guinea pigs were randomly divided into the S1 to S10 groups, a positive control group, and a blank control group (12 groups in total), with 10 guinea pigs in each group. The S1 to S10 groups were respectively administered Farfarae Flos extract S1 to S10 (4 g/kg), the positive control group was administered pentoverine citrate (10 mg/kg), and the blank control group was administered purified water. Each group received continuous oral administration for 5 days. The guinea pigs were placed in 5 L closed wide-mouth bottles, and 17.5% citric acid was sprayed into the bottle with an ultrasonic atomizer at the maximum spray intensity for 0.5 minutes. The cough latency period and cough frequency in 5 minutes were recorded for each guinea pig. Grey relational analysis (GRA) and partial least squares regression (PLSR) were used to conduct spectral-effect correlation analysis of the chemical composition data of Farfarae Flos extract and the antitussive efficacy data, and predict the group of active ingredients in Farfarae Flos with antitussive activity. The bioequivalence verification was conducted to verify the predicted group of active ingredients in Farfarae Flos with antitussive activity: SPF-grade healthy male Hartley guinea pigs were randomly divided into a S9 group, an active ingredient group, a positive control group, and a blank control group (4 groups in total), with 10 guinea pigs in each group. The S9 group was administered Farfarae Flos extract S9 (4 g/kg), the active ingredient group was administered the predicted combination of antitussive active ingredients (dose equivalent to 4 g/kg of Farfarae Flos extract S9), the positive control group was administered pentoverine citrate (10 mg/kg), and the blank control group was administered purified water. Each group received continuous oral administration for 5 days, and animal modeling and observation of efficacy indicators were the same as above.
RESULTS: The HPLC fingerprint of 10 batches of Farfarae Flos extract was established, and the peak area data of 14 main common peaks were obtained. The antitussive effect data of 10 batches of Farfarae Flos extract were obtained. Compared with the blank control group, the cough latence in the positive control group and S1, S2, S3, S4, S6, S7, S8, S9, S10 groups was prolonged (all P<0.01), while the cough frequency in 5 minutes in the positive control group and S1, S2, S4, S6, S8, S9, S10 groups was decreased (all P<0.05). The analysis of spectrum-effect relationship revealed that isochlorogenic acid C, isochlorogenic acid A, chlorogenic acid, isochlorogenic acid B, isoquercitrin, and rutin had high contribution to the antitussive effect of Farfarae Flos, and the 6 components were predicted to be the antitussive component group of Farfarae Flos. The verification of bioequivalence showed that there were no statistically significant differences in the antitussive effect between the S9 group and the antitussive component composition group(all P>0.05), which confirmed that isochlorogenic acid C, isochlorogenic acid A, chlorogenic acid, isochlorogenic acid B, isoquercetin, and rutin were the antitussive component group of Farfarae Flos.
CONCLUSIONS: The analysis of spectrum-effect relationship combined with the verification of bioequivalence could be used to study the antitussive material basis of Farfarae Flos. The antitussive effect of Farfarae Flos is the result of the joint action of many components.
目的: 款冬花具有止咳化痰的作用，以止咳为主。研究显示款冬花的某些成分可能与止咳作用相关，另有某些成分被证实具有止咳活性，但款冬花止咳药效的物质基础尚未阐明。本研究通过对款冬花提取物的高效液相色谱指纹图谱与止咳作用进行谱效相关分析，旨在阐明款冬花的止咳活性成分群。方法: 采用高效液相色谱法建立10批款冬花提取物的指纹图谱并获得其化学成分数据；选用豚鼠作为实验动物，采用枸橼酸引咳法获得10批款冬花提取物的止咳药效数据。将SPF级健康雄性Hartley豚鼠随机分为S1~S10组、阳性对照组和空白对照组(共12组)，每组10只，S1~S10组分别给予款冬花提取物S1~S10(4 g/kg)，阳性对照组给予枸橼酸喷托维林(10 mg/kg)，空白对照组给予纯化水，各组连续给药5 d，将豚鼠置于5 L容积的密闭广口瓶内，用超声雾化器以最大喷雾力度喷入17.5%的枸橼酸并持续0.5 min，记录每只豚鼠的咳嗽潜伏期和5 min内咳嗽次数。采用灰色关联度分析和偏最小二乘回归对款冬花提取物的化学成分数据和止咳药效数据进行谱效相关分析，预测款冬花的止咳活性成分群。对预测的款冬花止咳活性成分群进行等效性验证：将SPF级健康雄性Hartley豚鼠随机分为S9组、活性成分群组、阳性对照组和空白对照组(共4组)，每组10只，S9组给予款冬花提取物S9(4 g/kg)，活性成分群组给予预测的止咳活性成分组合物(剂量相当于4 g/kg的款冬花提取物S9)，阳性对照组给予枸橼酸喷托维林(10 mg/kg)，空白对照组给予纯化水，各组连续给药5 d，动物造模和药效指标观察同前。结果: 共建立10批款冬花提取物的高效液相色谱指纹图谱并获得14个主要共有峰的峰面积数据；获得10批款冬花提取物的止咳药效数据。与空白对照组比较，阳性对照组及S1、S2、S3、S4、S6、S7、S8、S9、S10组的咳嗽潜伏期加长(均P<0.01)，阳性对照组及S1、S2、S4、S6、S8、S9、S10组的5 min内咳嗽次数减少(均P<0.05)。谱效相关分析显示：异绿原酸C、异绿原酸A、绿原酸、异绿原酸B、异槲皮苷、芦丁6种成分对款冬花止咳作用的贡献度较高，初步判断为款冬花的止咳活性成分群。等效性验证显示：S9组、活性成分群组的2个药效指标的差异均无统计学意义(均P>0.05)，证实异绿原酸C、异绿原酸A、绿原酸、异绿原酸B、异槲皮苷、芦丁为款冬花的止咳活性成分群。结论: 谱效相关分析结合等效性验证可用于款冬花止咳药效物质基础研究。款冬花发挥止咳作用是多成分共同作用的结果。.

Gastrodiae Rhizoma was proven to have anti-inflammatory activity based on its main component of 4-hydroxybenzyl alcohol (4-HBA) and gastrodin (GAS). However, the anti-inflammatory activity of other phenols has been less reported. In this study, the n-BuOH extract was selected as the active anti-inflammatory part of Gastrodiae Rhizoma based on the LPS-induced inflammatory BV-2 cells. The spectral-effect relationship analysis of the n-BuOH extract showed the main effective components were GAS, 4-HBA, parishin A (PA), parishin B (PB), and parishin C (PC). Among them, PB could reduce LPS-induced expression of nitric oxide (NO), intracellular ROS, TNF-α, IL-6, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). Molecular docking predicted that PB had a good binding capacity to AMPKα and SIRT1 proteins of -12.1 kJ/mol and -7.6 kJ/mol, respectively. The Western Blot results further demonstrated that PB could inhibit NF-κB pathway by activating AMPK/SIRT1 pathway, thus exerting anti-LPS-induced neuroinflammatory effects. This study provides a referable idea for solving the problem of unclear action of TCM with complex compositions and is of great significance for the development of innovative medicines of traditional Chinese medicine.

OBJECTIVE: To explore the therapeutic effect of transdermal patches containing Cassia seed extract applied at the navel on slow transit constipation (STC) in rats and explore the spectrum-effect relationship of the patches.
METHODS: In a STC rat model established by gavage of compound diphenoxylate suspension for 14 days, the transdermal patches containing low, medium and high doses of Cassia seed extract (41.75, 125.25, and 375.75 mg/kg, respectively) were applied at the Shenque acupoint on the abdomen for 14 days after modeling, with constipation patches (13.33 mg/kg) as the positive control. After the treatment, fecal water content and intestinal propulsion rate of the rats were calculated, the pathological changes in the colon were observed with HE staining. Serum NO and NOS levels and the total protein content and NO, NOS and AChE expressions in the colon tissue were determined. HPLC fingerprints of the transdermal patches were established, and the spectrum-effect relationship between the common peaks of the patches and its therapeutic effect were analyzed.
RESULTS: Treatment with the transdermal patches containing Cassia seed extract significantly increased fecal water content and intestinal propulsion rate of the rat models, where no pathological changes in the colon tissue were detected. The treatment also suppressed the elevations of serum and colonic NO and NOS levels and reduction of AChE in STC rats. Twenty-eight common peaks were confirmed in the HPLC fingerprints of 6 batches of Cassia seed extract-containing patches. Analysis of the spectrum-effect relationship showed that autrantio-obtusin had the greatest contribution to the therapeutic effect of the patches in STC rats.
CONCLUSIONS: The Cassia seed extract-containing patches alleviates STC in rats via synergistic actions of multiple active ingredients in the extract, where autrantio-obtusin, rhein, chrysoobtusin, obtusin, obtusifolin, emodin, chrysophanol, and physcion are identified as the main active ingredients.

BACKGROUND: Cistanche tubulosa (CT) is the dried fleshy stem with scaly leaves of Cistanche tubiflora (Schenk) Wight, which has the effects of tonifying the kidney-yang, benefiting the vital essence and blood, and moisturizing the intestines and laxatives. There are differences in the activity of CT before and after processing, but the mechanism of processing is not clear.
OBJECTIVE: The study aimed to compare the strength of action of CT before and after yellow-wine processing in the treatment of constipation and kidney yang deficiency and to identify the active ingredients responsible for the differences in activity before and after yellow-wine processing.
METHODS: This study established the fingerprints of CT and PCT using HPLC to identify their shared components. Then efficacy of KYDS and FC were carried out to compare the differences between CT and PCT in terms of efficacy. Next, this study established the spectrum-effect relationship between the shared chemical components and the medical effects of CT and PCT using the gray correlation analysis and entropy methods. Ultimately, the activity of the analyzed chemical components was verified using the zebrafish model.
RESULTS: CT was more effective than PCT in promoting intestinal peristalsis, regulating gastrointestinal hormone levels, and thus treating FC. PCT was more effective than CT in improving the level of hormone indexes of the hypothalamus-pituitary-target gland axis, replenishing blood, and enhancing immunity. Through the analysis of the spectrum-effect relationship, it was finally found that 5, 6, 12 (tubuloside A), and 13 (isoacteoside) might be more closely related to the activity of tonifying kidney yang, and peaks 9, 10, and 11 (acteoside) are more closely associated with the treatment of constipation, and peaks 3 (salidroside), 4, 1, 2 (geniposidic acid), and 8 (echinacoside) were associated with both kidney yang tonic and treatment of constipation. At the same time, an activity verification experiment showed that echinacoside, geniposidic acid, and salidroside were effective in the treatment of FC and KYDS, while acteoside was very effective in the treatment of FC, and tubuloside A was significant in supplementing the blood, which validated the spectrum-effect relationship analysis.
CONCLUSIONS: This study proved that the raw CT had a better laxative effect, while the yellow-wine processed CT had a better kidney-yang tonic effect; moreover, spectrum-effect relationships were established to analyze the chemical components leading to changes in the activity of CT before and after yellow-wine processing.

Ardisia crenata Sims, an important ethnic medicine, is recorded in the Chinese Pharmacopoeia for treating laryngeal diseases and upper respiratory tract infections. This study aimed to evaluate the antimicrobial effect of extracts and potential antimicrobial compounds of A. crenata Sims. It was found that the roots of A. crenata Sims have a potential inhibitory effect on Candida albicans and Aspergillus flavus, with MICs of 1.56 mg/mL and 0.39 mg/mL, and the leaves of A. crenata Sims have a potential inhibitory effect on Pseudomonas aeruginosa and Staphylococcus aureus, with MICs of 3.12 mg/mL and 6.77 mg/mL, respectively. Meanwhile, five compounds including one catechin and four bergenins were obtained from roots. These components were identified on the fingerprint spectrum, representing chromatographic peaks 16, 21, 22, 23, and 25, respectively. Among these, 11-β-d-glucopyranosyl-bergenin and (-)-gallocatechin showed potential inhibition for Staphylococcus aureus and Pseudomonas aeruginosa with MIC of 0.26 and 0.33 mg/mL, respectively. The roots, stems, and leaves of A. crenata Sims are very similar in chemical composition, with large differences in content. Principal component analysis (PCA) and Hierarchical cluster analysis (HCA) showed that 16 batches of A. crenata Sims could be divided into four main production areas: Guizhou, Jiangsu, Guangxi, and Jiangxi. Furthermore, molecular docking results showed that 11-β-d-glucopyranosyl-bergenin had a better affinity for Casein lytic proteinase P (ClpP), and (-)-gallocatechin possessed a strong affinity for LasA hydrolysis protease and LasB elastase. These findings suggest catechin and bergenins from A. crenata Sims can be used as antimicrobial activity molecules.

Hawthorn has the efficacy of eliminating turbidity and lowering the blood lipid level, and it is used for treating hyperlipidemia in clinic. However, the bioactive components of hawthorn are still unclear. In this study, the spectrum-effect relationship was employed to screen the bioactive components of hawthorn in the treatment of hyperlipidemia, and then the bioactive components screened out were verified in vivo. Furthermore, the quality control method for hawthorn was developed based on liquid chromatography-mass spectrometry(LC-MS). The hyperlipidemia model of rats was built, and different polar fractions of hawthorn extracts and their combinations were administrated by gavage. The effects of different hawthorn extract fractions on the total cholesterol(TC), triglycerides(TG), and low-density lipoprotein-cholesterol(LDL-C) in the serum of model rats were studied. The orthogonal projections to latent structures(OPLS) algorithm was used to establish the spectrum-effect relationship model between the 24 chemical components of hawthorn and the pharmacodynamic indexes, and the bioactive components were screened out and verified in vivo. Finally, 10 chemical components of hawthorn, including citric acid and quinic acid, were selected to establish the method for evaluating hawthorn quality based on LC-MS. The results showed that different polar fractions of hawthorn extracts and their combinations regulated the TG, TC, and LDL-C levels in the serum of the model rats. The bioactive components of hawthorn screened by the OPLS model were vitexin-4″-O-glucoside, vitexin-2″-O-rhamnoside, rutin, citric acid, malic acid, and quinic acid. The 10 chemical components of hawthorn, i.e., citric acid, quinic acid, rutin, gallic acid, vitexin-4″-O-glucoside, vitexin-2″-O-rhamnoside, malic acid, vanillic acid, neochlorogenic acid, and fumaric acid were determined, with the average content of 38, 11, 0.018, 0.009 5, 0.037, 0.017, 8.1, 0.009 5, 0.073, and 0.98 mg·g~(-1), respectively. This study provided a scientific basis for elucidating the material basis of hawthorn in treating hyperlipidemia and developed a content determination method for evaluating the quality of hawthorn.

To enhance the quality evaluation and control of traditional Chinese medicine (TCM) and ensure the safety and efficacy of clinical medication, it is imperative to establish a comprehensive quality assessment method aligned with TCM efficacy. This study uses a representative Chinese medicine with multi-origin and multi-efficacy, Paris polyphylla var. yunnanensis (PY), as an illustrative example. Surprisingly, despite the high fingerprint similarity among the 12 batches of PY samples collected from various regions in Yunnan, a notable variation in the composition and content of components was observed. The chromatographic analysis identified seven common peaks, namely, polyphyllin I, polyphyllin II, polyphyllin V, polyphyllin VI, polyphyllin VII, polyphyllin H, and polyphyllin D. In the bioactivity evaluation, an in vitro antiplatelet aggregation model induced by adenosine diphosphate was established, showcasing excellent stability. The maximum antiplatelet aggregation inhibition rate for all PY samples consistently remained stable at 73.1%-99.1%. However, the 50% inhibitory concentration (IC50 ) values exhibited a range from 1.615 to 18.200 mg/mL. This approach not only meets high-throughput screening requirements but also demonstrates remarkable discrimination. The results of chemical and bioactivity evaluations were analyzed using hierarchical cluster analysis and canonical correlation analysis. Polyphyllin I, polyphyllin II, polyphyllin VII, polyphyllin H, and polyphyllin D were identified as the Q-markers for antiplatelet aggregation in PY samples. Validation of the bioactivity for these monomer components aligned with the previously mentioned findings. Notably, this study established a spectrum-effect model for PY samples, enhancing the scientific robustness of the quality evaluation method. Furthermore, these findings offer valuable research insights for improving the quality assessment of other TCMs.