Dammarane-type triterpenoids from Gynostemma longipes and their protective activities on hypoxia-induced injury in PC12 cells.

Sixteen new dammarane-type triterpenoid saponins (1-16) featuring diverse structural variations in the side chain at C-17, along with twenty-one known analogues (17-37), have been isolated from the rhizomes of Gynostemma longipes C. Y. Wu, a plant renowned for its medicinal and edible properties. The structural elucidation of these compounds was accomplished through comprehensive analyses of 1D and 2D NMR and HRMS spectroscopic data, supplemented by comparison with previously reported data. Subsequent assays on the isolates for their protective effects against hypoxia-induced damage in pheochromocytoma cells (PC12 cells) revealed that nine saponins exhibited significant anti-hypoxic activities. Further investigation into the anti-hypoxia mechanisms of the representative saponins demonstrated that compounds 22 and 36 markedly reduced the levels of hypoxia-induced apoptosis. Additionally, these compounds were found to decrease the release of lactate dehydrogenase (LDH) and malondialdehyde (MDA), while increasing the activity of superoxide dismutase (SOD), thereby indicating that the saponins could mitigate hypoxia-induced injuries by ameliorating apoptosis and oxidative stress. These findings offer substantial evidence for the future utilization and development of G. longipes, identifying dammarane-type triterpenoid saponins as its active anti-hypoxic constituents.

Exploring the chemical compositions of Fufang Yinhua Jiedu granules based on ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry combined with multistage intelligent data annotation strategy.

Fufang Yinhua Jiedu granules (FYJG) is a Traditional Chinese Medicine (TCM) compound formulae preparation comprising ten herbal drugs, which has been widely used for the treatment of influenza with wind-heat type and upper respiratory tract infections. However, the phytochemical constituents of FYJG have rarely been reported, and its constituent composition still needs to be elucidated. The complexity of the natural ingredients of TCMs and the diversity of preparations are the major obstacles to fully characterizing their constituents. In this study, an innovative and intelligent analysis strategy was built to comprehensively characterize the constituents of FYJG and assign source attribution to all components. Firstly, a simple and highly efficient ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC-QTOF MSE) method was established to analyze the FYJG and ten single herbs. High-accuracy MS/MS data were acquired under two collision energies using high-definition MSE in the negative and positive modes. Secondly, a multistage intelligent data annotation strategy was developed and used to rapidly screen out and identify the compounds of FYJG, which was integrated with various online software and data processing platforms. The in-house chemical library of 2949 compounds was created and operated in the UNIFI software to enable automatic peak annotation of the MSE data. Then, the acquired MS data were processed by MS-DIAL, and a feature-based molecular networking (FBMN) was constructed on the Global Natural Product Social Molecular Networking (GNPS) to infer potential compositions of FYJG by rapidly classifying and visualizing. It was simultaneously using the MZmine software to recognize the source attribution of ingredients. On this basis, the unique chemical categories and characteristics of herbaceous plant species are utilized further to verify the accuracy of the source attribution of multi-components. This comprehensive analysis successfully identified or tentatively characterized 279 compounds in FYJG, including flavonoids, phenolic acids, coumarins, saponins, alkaloids, lignans, and phenylethanoids. Notably, twelve indole alkaloids and four organic acids from Isatidis Folium were characterized in this formula for the first time. This study demonstrates a potential superiority to identify compounds in complex TCM formulas using high-definition MSE and computer software-assisted structural analysis tools, which can obtain high-quality MS/MS spectra, effectively distinguish isomers, and improve the coverage of trace components. This study elucidates the various components and sources of FYJG and provides a theoretical basis for its further clinical development and application.

Cannabidiol ameliorates PTSD-like symptoms by inhibiting neuroinflammation through its action on CB2 receptors in the brain of male mice.

Post-traumatic stress disorder (PTSD) is a debilitating mental health disease related to traumatic experience, and its treatment outcomes are unsatisfactory. Accumulating research has indicated that cannabidiol (CBD) exhibits anti-PTSD effects, however, the underlying mechanism of CBD remains inadequately investigated. Although many studies pertaining to PTSD have primarily focused on aberrations in neuronal functioning, the present study aimed to elucidate the involvement and functionality of microglia/macrophages in PTSD while also investigated the modulatory effects of CBD on neuroinflammation associated with this condition. We constructed a modified single-prolonged stress (SPS) mice PTSD model and verified the PTSD-related behaviors by various behavioral tests (contextual freezing test, elevated plus maze test, tail suspension test and novel object recognition test). We observed a significant upregulation of Iba-1 and alteration of microglial/macrophage morphology within the prefrontal cortex and hippocampus, but not the amygdala, two weeks after the PTSD-related stress, suggesting a persistent neuroinflammatory phenotype in the PTSD-modeled group. CBD (10 mg/kg, i.p.) inhibited all PTSD-related behaviors and reversed the alterations in both microglial/macrophage quantity and morphology when administered prior to behavioral assessments. We further found increased pro-inflammatory factors, decreased PSD95 expression, and impaired synaptic density in the hippocampus of the modeled group, all of which were also restored by CBD treatment. CBD dramatically increased the level of anandamide, one of the endocannabinoids, and cannabinoid type 2 receptors (CB2Rs) transcripts in the hippocampus compared with PTSD-modeled group. Importantly, we discovered the expression of CB2Rs mRNA in Arg-1-positive cells in vivo and found that the behavioral effects of CBD were diminished by CB2Rs antagonist AM630 (1 mg/kg, i.p.) and both the behavioral and molecular effects of CBD were abolished in CB2Rs knockout mice. These findings suggest that CBD would alleviate PTSD-like behaviors in mice by suppressing PTSD-related neuroinflammation and upregulation and activation of CB2Rs may serve as one of the underlying mechanisms for this therapeutic effect. The present study offers innovative experimental evidence supporting the utilization of CBD in PTSD treatment from the perspective of its regulation of neuroinflammation, and paves the way for leveraging the endocannabinoid system to regulate neuroinflammation as a potential therapeutic approach for psychiatric disorders.

A universal method for profiling and characterization of oligosaccharides in traditional Chinese medicines.

Oligosaccharides constitute fundamental components in numerous traditional Chinese medicines (TCMs). Conventional chromatographic methods for natural product analysis are not suitable for oligosaccharides due to their large polarity and structural similarity. Herein, an ultra-high performance liquid chromatography with charged aerosol detector (UHPLC-CAD) method was developed for the profiling of oligosaccharides using 9 neutral (DP3-DP11) reference oligosaccharides. Various factors, including columns, mobile phase, elution conditions, flow rate, and column temperature were systematically examined. Optimal separation was achieved using an Amide column with gradient elution within 18 min, at 0.5 mL/min flow rate and 30°C column temperature. Moreover, an ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) method was also optimized to provide structural information. The developed method was applied to detect oligosaccharides in several TCMs, including Morindae Officinalis Radix (MOR), Ziziphi Spinosae Semen (ZSS), Menthae Haplocalycis Herba (MHH) and Chrysanthemi Indici Flos (CIF), revealing 9 and 16 oligosaccharides being uncovered from MHH and CIF respectively for the first time. This study presents a versatile UHPLC-CAD and UHPLC-Q-TOF/MS method with the potential for advancing oligosaccharides discovery and contributing to the quality analysis of TCMs.

[Qualitative and semiquantitative analyses of the chemical components of the seed coat and kernel of Ziziphi Spinosae Semen].

Ziziphi Spinosae Semen refers to the dried seed of Ziziphus jujuba Mill. var. spinosa (Bunge) Hu ex H. F. Chou. The seed is composed of a reddish brown coat and a yellow kernel. A comparative study was conducted to investigate differences in the chemical composition and their relative contents between the seed coat and kernel of Ziziphi Spinosae Semen. First, the chemical compounds found in the seed coat and kernel were characterized and identified using ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). The analytical results tentatively identified 57 chemical compounds based on reference-compound comparison, literature retrieval, and chemical-database (e. g., MassBank) searches; these compounds included 14 triterpenes, 23 flavonoids, 7 alkaloids, 6 carboxylic acids, and 7 other types of compounds. The mass error of the identified compounds was within the mass deviation range of 5×10-6 (5 ppm). Next, two methods of multivariate statistical analysis, namely, principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA), were used to compare the differential compounds between the two seed parts. A total of 17 differential compounds were screened out via OPLS-DA based on a variable importance in projection (VIP) value of >5. The results revealed that betulinic acid, betulonic acid, alphitolic acid, and jujuboside Ⅰ mainly existed in the seed coat whereas the 13 other compounds, such as spinosin, jujuboside A, and 6‴-feruloylspinosin, mainly existed in the seed kernel. Therefore, these 17 differential compounds can be used to distinguish between the two seed parts. Finally, a semiquantitative method was established using UPLC and a charged aerosol detector (CAD) with inverse gradient compensation in the mobile phase. Six representative compounds with different types were selected to examine the CAD response consistency: magnoflorine (alkaloid), spinosin (flavone), 6‴-feruloylspinosin (flavone), jujuboside A (triterpenoid saponin), jujuboside B (triterpenoid saponin), and betulinic acid (triterpenoid acid). The results showed that the relative standard deviation (RSD) of the average response factors at different levels of these six compounds was 7.04% and that their response intensities were similar. Moreover, each compound in the fingerprint demonstrated good response consistency, and the peak areas obtained directly reflected the contents of each compound. Based on the semiquantitative fingerprints obtained, betulinic acid and oleic acid were considered the main components of the seed coat. The betulinic acid content in the seed coat was approximately 7 times higher than that in the seed kernel. Spinosin, jujuboside A, linoleic acid, betulinic acid, and oleic acid were the main components of the seed kernel. The spinosin content in the seed kernel was 18 times higher than that in the seed coat. In addition, the jujuboside A content in the seed kernel was 24 times higher than that in the seed coat. The proposed method can accurately determine the main components and compare the relative contents of these components in different seed parts. In summary, this study identified the differences in chemical components between the seed coat and kernel of Ziziphi Spinosae Semen and clarified the main components and their relative contents in these parts. The findings can not only provide a basis for the identification of chemical compounds and quality research on different parts of Ziziphi Spinosae Semen but also promote the development and utilization of this traditional Chinese medicine.

A quantitative chemomics strategy for the comprehensive comparison of Murraya paniculata and M. exotica using liquid chromatography coupled with mass spectrometry.

Murrayae Folium et Cacumen (MFC) is a traditional Chinese medicine (TCM) derived from two plant species, Murraya exotica L. and Murraya paniculata (L.) Jack, as recorded in the Chinese Pharmacopoeia. However, there is no research available on the comprehensive analysis and comparison of the chemical constituents of these two species. In the present study, an integrated LC-MS-based quantitative metabolome strategy was proposed to conduct a comprehensive and in-depth qualitative and quantitative analysis and comparison of the chemome of M. exotica and M. paniculata. Firstly, the universal chemical information of two plants was obtained by quadrupole-time-of-flight mass spectrometry (Q-TOF-MS) combined with hybrid triple quadrupole-linear ion trap mass spectrometry (Qtrap-MS). Subsequently, a UNIFI in house database, the proposed fragmentation patterns, and a quantitative structure chromatographic retention relationship (QSRR) model were integrated for the rapid, comprehensive, and accurate structural elucidation of the chemical constituents of these two species. Thirdly, a large-scale quantitation method was established using scheduled multiple reaction monitoring mode (sMRM) and 76 primary components were selected as quantitative markers for the method validation. The obtained dataset was then subjected for multivariate statistical analysis to comprehensive comparison of these two plants. As a result, a total of 209 and 212 compounds were identified from M. exotica and M. paniculata, respectively. Among them, 103 common constituents were disclosed in both plants. The multivariate statistical analysis and absolute quantitative analysis revealed noticeable differences in the contents of specific chemical constituents between these two plants. The higher quantity constituents in M. exotica are 7-methoxycoumarins, while polymethoxylated flavonoids are the major constituents in M. paniculata. The common compounds accounted for approximately 80 % of the quantitative components in both plants, which provides a theoretical basis for their common use as the official source of MFC. In sum, the established quantitative chemomics strategy supplies an effective means for comprehensive chemical comparison of multi-source TCMs.

Isolation and structural elucidation of prebiotic oligosaccharides from Ziziphi Spinosae Semen.

Six oligosaccharides were discovered and isolated for the first time from Ziziphi Spinosae Semen. On the basis of spectroscopic analysis, their structures were determined to be verbascose (1), verbascotetraose (2), stachyose (3), manninotriose (4), raffinose (5), and melibiose (6). The prebiotic effect of the oligosaccharide fraction was assayed by eight gut bacterial growth in vitro, revealing a significant increase in cell density, up to 4-fold, for Lactobacillus acidophilus, Lactobacillus gasseri, and Lactobacillus johnsonii. The impact of six oligosaccharides with different degrees of polymerization (DPs) and structures on the growth of Lactobacillus acidophilus was evaluated. As a result, stachyose and raffinose demonstrated superior support for bacterial growth compared to the other oligosaccharides. This study explored the structure-activity relationship of raffinose family oligosaccharides (RFOs) and showed that the more the monosaccharide type, the more supportive the gut bacteria growth when oligosaccharides have the same molecular weight.

Anti-inflammatory quinoline-4(1H)-one derivatives from the aerial parts of Waltheria indica linn.

Eight previously undescribed quinoline-4(1H)-one derivatives (1-8) and five known analogues (9-13) were isolated from the 95% aqueous extract of the aerial parts of Waltheria indica Linn. Their chemical structures were determined by analyzing 1D NMR, 2D NMR and HRESIMS data comprehensively. Compounds 1-8 possess diverse side chains at C-5 of quinoline-4(1H)-one or tetrahydroquinolin-4(1H)-one skeleton. The absolute configurations were assigned via comparison of the experimental and calculated ECD spectra, and analysis of the ECD data of the in situ formed [Rh2(OCOCF3)4] complex. Additionally, all 13 isolated compounds were evaluated for their anti-inflammatory activities by measuring the inhibitory effects of nitric oxide (NO) production in lipopolysaccharide-induced BV-2 cells. Compounds 2, 5 and 11 showed moderate inhibition toward NO production with IC50 values of 40.41 ± 1.01, 60.09 ± 1.23 and 55.38 ± 0.52 µM, respectively.

Dammarane-type saponins from Gynostemma pentaphyllum and their anti-aging activities via up-regulating mitochondria related proteins.

The importance of mitochondria in regulation of aging has been extensively recognized and confirmed. Gynostemma pentaphyllum (Thunb.) Makino, a homology of medicine and food, has been widely utilized as dietary supplement. In this study, the transcriptome of normal cells (wild type mouse embryo fibroblasts) regulated by the 30% aqueous EtOH extract of G. pentaphyllum was firstly evaluated by RNA sequencing and the results revealed that the G. pentaphyllum could up-regulate the genes involved in oxidative phosphorylation (OXPHOS) and sirtuin (SIRT) signaling pathways, indicating its effect in promoting cell viability might be attributed to the role of improving mitochondrial functions. To further discover the bioactive compounds, sixteen undescribed dammarane-type saponins along with twenty-eight known analogues were isolated from the active extract of G. pentaphyllum. Their structures were elucidated by means of comprehensive analysis of NMR and HRMS spectroscopic data. All isolates were evaluated for the regulatory effects on SIRT3 and translocase of the outer membrane 20 (TOM20), and thirteen of them exhibited satisfactory agonist activities on both SIRT3 and TOM20 at 5 µM. Furthermore, the preliminary structure-activity relationships analysis demonstrated the additional hydroxymethyl and carbonyl groups or less sugar residues in saponins could contribute positively to the up-regulatory effect on SIRT3 and TOM20. These findings encouraged the potential roles of G. pentaphyllum and its bioactive saponins in the development of natural drugs for the treatment of aging-related diseases.

An effective and high-throughput sample preparation method involving demalonylation followed by an ultrahigh-performance liquid chromatography-charged aerosol detector for analyzing gypenoside XLIX and gypenoside A in Gynostemma longipes.

Gypenosides (Gps) are the major bioactive components in Gynostemma species. They include neutral Gps and acidic malonylgypenosides (MGps). MGps are abundant in Gynostemma species and can be transformed into corresponding Gps via extraction, concentration, and drying. If only the Gps were quantified and MGps were ignored, the quality of Gynostemma species would be underestimated. This study aimed to develop a sample preparation method involving demalonylation and ultrahigh-performance liquid chromatography-charged aerosol detector (UHPLC-CAD) analysis to determine the contents of gypenoside XLIX (Gp XLIX) and gypenoside A (Gp A). First, the optimized ultrasonic extraction method was established to extract G. longipes powder ultrasonically. Then, the extracted solution was put into a closed container (centrifuge tube) and heated in a water bath at 95 °C. Then, MGps were converted into corresponding Gps. The proposed preparation method was compared with the other three methods, including water bath reflux heating, alkali hydrolysis, and extraction of heated powder, and was shown to exhibit higher conversion and better convenience. Subsequently, an UHPLC-CAD method was established and validated. Gp XLIX and Gp A showed excellent linear correlations between 15.55 and 248.8 µg/mL and 24.10-385.5 µg/mL, respectively (R2 > 0.999). The limit of detection was 1.40 ng (Gp XLIX) and 2.41 ng (Gp A), and the limit of quantification was 7.77 ng and 14.46 ng, respectively. The relative standard deviation for precision, stability, and repeatability was 0.63-3.15%. The average recovery of Gp XLIX and Gp A was 98.97% and 98.23%, respectively. The established method was applied for determining Gp XLIX and Gp A contents in wild or cultivated G. longipes samples collected from the Qinba Mountains area. The contents of Gp XLIX and Gp A were 5.16-23.02 mg/g and 15.78-54.55 mg/g, respectively. Conclusively, the proposed sample preparation and analysis method could be used for the quality control and evaluation of G. longipes.

Qualitative and quantitative studies on two commercial specifications of Polygonatum odoratum.

The rhizoma of Polygonatum odoratum (PO) is used to treat yin injuries of the lung and stomach in traditional Chinese medicine. The chemical constituents of this herb are steroidal saponins, homoisoflavanones, and alkaloids. Xiangyuzhu (XPO) and Guanyuzhu (GPO) are available in the market as two specifications of the commodity. Nonetheless, systematic research on the identification and comparison of chemical constituents of these two commercial specifications is yet lacking. Herein, an integrated method combing ultra-high-performance liquid chromatography-quadruple time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) with ultra-high-performance liquid chromatography-charged aerosol detection (UHPLC-CAD) was employed for the comprehensively qualitative and quantitative analyses of PO. A total of 62 compounds were identified by UHPLC-Q-TOF/MS, among which 13 potential chemical markers were screened out to distinguish two commercial specifications. Subsequently, the absolute determination method for polygodoraside G, polygonatumoside F, and timosaponin H1 was established and validated by UHPLC-CAD. The contents of the three compounds were 13.33-236.24 µg/g, 50.55-545.04 µg/g, and 13.34-407.83 µg/g, respectively. Furthermore, the ratio of timosaponin H1/polygodoraside G could be applied to differentiate the two specifications. Samples with a ratio <2 are considered XPO and >5 are considered GPO. Therefore, the above results provide a valuable means for the quality control of PO.

[Determination of gypenoside XLVI and LVI in Gynostemma pentaphyllum from Fujian by ultra-high performance liquid chromatography-charged aerosol detector].

Gynostemma pentaphyllum (Thunb.) Makino contains dammarane-type triterpenoid saponins, similar to ginseng, with a host of pharmacological activities. However, its planting resources and chemical composition are quite complex. The chemical constituents of Gynostemma pentaphyllum vary drastically among different origins and varieties. Thus, the corresponding quality control methods also need to be different. Currently, limited information is available about the quality control of Gynostemma pentaphyllum from Fujian. A new method based on ultra-high performance liquid chromatography-charged aerosol detection (UHPLC-CAD) was established for the determination of gypenoside XLVI and LVI in Gynostemma pentaphyllum. The major components of Gynostemma pentaphyllum were characterized using UHPLC-quadrupole time-of-flight-mass spectrometry (UHPLC-Q-TOF/MS) combined with UHPLC-CAD. The results revealed gypenoside XLVI, LVI, and their corresponding malonyl-containing acidic saponins as the main components. However, malonylgypenoside XLVI and LVI can easily remove their malonyl group and convert to gypenoside XLVI and LVI during the application of Gynostemma pentaphyllum. In this study, the samples were pretreated using alkali hydrolysis to transform the acid saponins completely, and the final contents of gypenoside XLVI and LVI were determined via UHPLC-CAD. The optimal alkaline hydrolysis, extraction, and liquid chromatography conditions were established. First, the alkaline hydrolysis conditions were optimized. The effects of the volume of ammonia and reaction time on the contents of gypenoside XLVI, LVI, malonylgypenoside XLVI, and LVI were examined. Malonylgypenoside XLVI and LVI could be transformed completely to gypenoside XLVI and LVI by standing for 24 h in an ethanol-water-ammonia (50∶46∶4, v/v/v) mixture. Furthermore, the extraction conditions were optimized. Next, effects of the different solvents, extraction time, and solid-liquid ratio on the extraction rates of gypenoside XLVI and LVI were investigated. The extraction method for Gynostemma pentaphyllum powder using the ethanol-water-ammonia (50∶46∶4, v/v/v) and a solid-liquid ratio of 1∶150 (g∶mL) for 30 min was established. Finally, a prepared test solution was separated on a Waters ACQUITY UPLC BEH C18 chromatographic column (100 mm×2.1 mm, 1.7 µm). Acetonitrile and 0.1% (v/v) formic acid aqueous solution were used as the mobile phases for gradient elution. The flow rate was set to 0.5 mL/min and column temperature was maintained at 40 ℃. The separation was detected using a charged aerosol detector. Results indicated that the logarithm of the mass concentrations of gypenoside XLVI and LVI had a linear relationship with the logarithm of the peak area in the range of 9.94-318.00 µg/mL and 12.78-409.00 µg/mL, respectively. The correlation coefficients (r) were 0.9993 and 0.9995, respectively. The limit of detection (LOD) and the limit of quantification (LOQ) of gypenoside XLVI were 1.58 µg/mL and 6.36 µg/mL, respectively. The LOD and LOQ of gypenoside LVI were 2.05 µg/mL and 8.18 µg/mL, respectively. The relative standard deviations (RSDs) of precision, repeatability, and 24 h stability were less than 2.0% (n=6). The spiked recoveries of gypenoside XLVI were 100.2%-107.2% and the RSD value was 2.4%. The spiked recoveries of gypenoside LVI were 97.9%-104.2% and the RSD value was 2.6%. The results of 16 batches of Gynostemma pentaphyllum samples indicated that the gypenoside XLVI content was 0.57%-2.57%, and gypenoside LVI content was 0.66%-2.99%. Hence, this method has high sensitivity and good reproducibility. Therefore, it can be used for quality research and quality control of Gynostemma pentaphyllum from Fujian.

New prenylated flavonoid glycosides derived from Epimedium wushanense by ß-glucosidase hydrolysis and their testosterone production-promoting effects.

Six new prenylated flavonoid glycosides, including four new furan-flavonoid glycosides wushepimedoside A-D (1-4) and two new prenyl flavonoid derivatives wushepimedoside E-F (5-6), and one know analog epimedkoreside B (7) were isolated from biotransformation products of the aerial parts of Epimedium wushanense. Their structures were elucidated according to comprehensive analysis of HR-MS and NMR spectroscopic data, and the absolute configurations were assigned using experimental and calculated electronic circular dichroism (ECD) data. The regulatory activity of compounds 1-7 on the production of testosterone in primary rat Leydig cells were investigated, and 4 and 5 exhibited testosterone production-promoting activities. Molecular docking analysis suggested that bioactive compounds 4 and 5 showed the stable binding with 3ß-HSD and 4 also had good affinity with Cyp17A1, which suggested that these compounds may regulate testosterone production through stimulating the expression of the above two key proteins.

Quality analysis of hawthorn leaves (the leaves of Crataegus pinnatifida Bge. var major N.E.Br) in different harvest time.

INTRODUCTION: Harvest time plays an important role on the quality of medicinal plants. The leaves of Crataegus pinnatifida Bge. var major N.E.Br (hawthorn leaves) could be harvested in summer and autumn according to the Pharmacopoeia of the People's Republic of China (Pharmacopoeia). However, little is known about the difference of the chemical constituents in hawthorn leaves with the harvest seasonal variations. OBJECTIVE: The chemical constituents of hawthorn leaves in different months were comprehensively analysed to determine the best harvest time. METHODS: Initially, the chemical information of the hawthorn leaves were obtained by ultra-high-performance liquid chromatography and quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS). Subsequently, principal component analysis (PCA) was applied to compare the chemical compositions of hawthorn leaves harvested in different months. Then, an absolute quantitation method was established using high-performance liquid chromatography-charged aerosol detector (HPLC-CAD) to determine the contents of five compounds and clarify the changes of these components with the harvest seasonal variations. Meanwhile, a semi-quantitative method by integrating HPLC-CAD with inverse gradient compensation was also established and verified. RESULTS: Fifty-eight compounds were identified through UHPLC-Q-TOF-MS. PCA revealed that the harvest season of hawthorn leaves had a significant effect on the chemical compositions. The contents of five components were relatively high in autumn. Other four main components without reference standards were further analysed through the semi-quantitative method, which also showed a high content in autumn. CONCLUSIONS: This work emphasised the effect of harvest time on the chemical constituents of hawthorn leaves and autumn is recommended to ensure the quality.

Comparison of Murraya exotica and Murraya paniculata by fingerprint analysis coupled with chemometrics and network pharmacology methods.

There are two source plants for the traditional Chinese medicine Murrayae Folium et Cacumen (MFC) in Chinese Pharmacopoeia, i.e. Murraya exotica L. and M. paniculata (L.) Jack. Herein, a chemical comparison of M. exotica and M. paniculata by high performance liquid chromatography (HPLC) fingerprint analysis coupled with chemometrics and network pharmacology was performed. The main peaks in the fingerprints were identified by liquid chromatography coupled with ion trap/time-of-flight mass spectrometry (LC-IT-TOF-MS) and authenticated by references. The chemometrics results showed that the HPLC fingerprints of these two species were clearly divided into two categories using hierarchical cluster analysis (HCA) and principal component analysis (PCA), and a total of 13 significantly differentiated markers were screened out by orthogonal partial least squares-discriminant analysis (OPLS-DA). However, the following network pharmacology analysis showed that these discriminated markers were found to act via many common targets and metabolic pathways, indicating the possibly similar pharmacological effects and mechanisms for M. exotica and M. paniculata. The above results provide valuable evidence for the equivalent use of these two plants in clinical settings. Moreover, the chromatographic fingerprint analysis coupled with chemometrics and network pharmacology supplies an efficient approach for the comparative analysis of multi-source TCMs like MFC.

Coumarin derivatives from the leaves and twigs of Murraya exotica L. and their anti-inflammatory activities.

Fifteen previously undescribed coumarin derivatives, murratins A-M, including two pairs of coumarin enantiomers with a cyclopropane unit, two benzocoumarins, a dimeric coumarin, and eight C-8-substituted coumarins, together with six known analogues were isolated from the extract of the leaves and twigs of Murraya exotica L., a medicinal plant named "Jiulixiang" in Chinese. Their structures were elucidated on the basis of comprehensive analysis of 1D and 2D NMR and HRMS spectroscopic data, and the absolute configurations were assigned via comparison of the specific rotations, the ECD exciton coupling method, comparison of experimental and calculated ECD data, and the ECD data of in situ formed transition metal complexes. All the isolates were evaluated for their inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW 264.7 cells, and five coumarin derivatives showed moderate inhibitory activities. The possible mechanism for NO inhibition of undescribed bioactive compounds was deduced to interact with iNOS protein via molecular docking. The above results shed some light on a better understanding of the traditional anti-inflammation effect of M. exotica and coumarins are disclosed to be its potential anti-inflammatory constituents.

Coumarin and spirocyclopentenone derivatives from the leaves and stems of Murraya paniculata (L.) Jack.

Seven previously undescribed compounds, including five coumarins, (+/-)-murpanitin A and murpanitins B-D, and a pair of spirocyclopentenone enantiomers, (+/-)-murrayaspiroketone, along with 14 known coumarin derivatives were isolated from the leaves and stems of Murraya paniculata (L.) Jack. Their structures were elucidated on the basis of comprehensive analysis of 1D and 2D NMR and HRMS spectroscopic data, and the absolute configurations were assigned via calculated and experimental ECD data. Three compounds showed moderate inhibitory effects on nitric oxide production stimulated by lipopolysaccharide in BV-2 microglial cells with IC50 values of 53.2 ± 8.9, 57.7 ± 5.8, and 53.2 ± 4.4 µM, respectively.

Panitins A-G: Coumarin derivatives from Murraya paniculata from Guangxi Province, China show variable NO inhibitory activity.

Seven previously undescribed coumarin derivatives, panitins A‒G, an ethoxylated artifact, and 34 known analogues, were isolated from the roots of Murraya paniculata. Their structures were elucidated on the basis of comprehensive analysis of 1D and 2D NMR and HRMS spectroscopic data, and comparison with the data reported in literature. The absolute configurations of undescribed compounds were assigned via comparison of the specific rotation, Mosher's method, exciton chiral method, and comparison of calculated and experimental ECD data. Panitin D, trans-dehydroosthol, and exotimarin I showed potent inhibition against LPS-induced NO production in BV-2 microglial cells with IC50 values of 19.6 ±â€¯2.3, 12.4 ±â€¯0.9, and 26.9 ±â€¯0.8 µM, respectively.