MiRNA-470-5p suppresses epithelial-mesenchymal transition of embryonic palatal shelf epithelial cells by targeting Fgfr1 during palatogenesis.

MicroRNAs (miRNAs) have been identified as crucial modulators of gene expression and to play a role in palatogenesis. The aim of this study was to explore the potential role and regulatory mechanisms of miRNAs during palatogenesis. RNA-sequencing was performed to compare the RNA expression profiles of mouse embryonic palatal shelf (MEPS) tissue between an all-trans retinoic acid (ATRA)-induced group and control group, followed by reverse transcription-quantitative polymerase chain reaction for validation, demonstrating upregulated expression of miRNA-470-5p and downregulated expression of Fgfr1 in the ATRA-induced group. The specific binding sites of miRNA-470-5p that potentially govern Fgfr1 expression were predicted by miRanda and TargetScan. The relationship between miRNA-470-5p and Fgfr1 was validated in HEK293T cells by luciferase reporter assays, confirming that miR-470-5p acts directly on the Fgfr1 3'-untranslated region. Fgfr1 mRNA and FGFR1 protein levels were markedly downregulated in MEPS epithelial cells over-expressing miRNA-470-5p. Functional experiments in vitro with CCK-8, cell colony formation, and 5-ethynyl-2'-deoxyuridine (EdU) staining assays revealed that upregulated miRNA-470-5p expression could inhibit the epithelial-mesenchymal transition (EMT) of MEPS epithelial cells by targeting Fgfr1. These findings provide a new molecular mechanism of cleft palate formation, which can inform the development of new treatment and/or prevention targets.

Efficacy and safety of Mudan granules for painful diabetic peripheral neuropathy: A protocol for a double-blind randomized controlled trial.

BACKGROUND: As one of the most challenging complications in the management of diabetes mellitus, painful diabetic peripheral neuropathy (PDPN) is accompanied by various clinical manifestations, including numbness, burning, coldness, and other sensory abnormalities in the extremities. Meanwhile, PDPN seriously affects the life quality of patients and causes great pain. Western medicine mostly provides symptomatic treatments, such as antioxidants, aldose reductase inhibitors, nerve nutrition, microcirculation improvement, and analgesic drugs on the basis of blood sugar control. Although certain efficacy has been achieved, the problem has not been solved at root. Mudan granules have some advantages in the treatment of PDPN, but there is insufficient high-quality clinical studies to verify this. Therefore, the purpose of this study was to evaluate the efficacy and safety of Mudan granules in treating PDPN. METHODS: A randomized, double-blind, placebo, and parallel-controlled trial design was used to study the efficacy and safety of Mudan granules in the treatment of PDPN. In this study, 93 patients with painful diabetic neuropathy were recruited and randomly divided into a treatment group and a placebo group based on 1:1. The treatment group was given Mudan granules and the control group accepted placebo treatment, and the basic treatment was performed according to the recommended guidelines. During the treatment period, the patients' visual analog scores, clinical efficacy, Medical Outcomes Study 36-item Short-Form Health Survey (SF-36) scores, nerve conduction velocity, and drug-induced adverse reactions were observed at baseline after 8 and 10 weeks. DISCUSSION: This study will evaluate the efficacy and safety of Mudan granules in treating PDPN. The experimental results will provide evidence support to treat PDPN with Mudan granules. TRIAL REGISTRATION: DOI 10.17605/OSF.IO/5CE32.

Metabolite Profiling and Transcriptome Analysis Explains Difference in Accumulation of Bioactive Constituents in Licorice (Glycyrrhiza uralensis) Under Salt Stress.

Salinity stress significantly affects the contents of bioactive constituents in licorice Glycyrrhiza uralensis. To elucidate the molecular mechanism underlying the difference in the accumulation of these constituents under sodium chloride (NaCl, salt) stress, licorice seedlings were treated with NaCl and then subjected to an integrated transcriptomic and metabolite profiling analysis. The transcriptomic analysis results identified 3,664 differentially expressed genes (DEGs) including transcription factor family MYB and basic helix-loop-helix (bHLH). Most DEGs were involved in flavonoid and terpenoid biosynthesis pathways. In addition, 121 compounds including a triterpenoid and five classes of flavonoids (isoflavone, flavone, flavanone, isoflavan, and chalcone) were identified, and their relative levels were compared between the stressed and control groups using data from the ultrafast liquid chromatography (UFLC)-triple quadrupole-time of flight-tandem mass spectrometry (TOF-MS/MS) analysis. Putative biosynthesis networks of the flavonoids and triterpenoids were created and combined with structural DEGs such as phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase [4CL], cinnamate 4-hydroxylase [C4H], chalcone synthase [CHS], chalcone-flavanone isomerase [CHI], and flavonoid-3',5' hydroxylase (F3',5'H) for flavonoids, and CYP88D6 and CYP72A154 for glycyrrhizin biosynthesis. Notably, significant upregulation of UDP-glycosyltransferase genes (UGT) in salt-stressed licorice indicated that postmodification of glycosyltransferase may participate in downstream biosynthesis of flavonoid glycosides and triterpenoid saponins. Accordingly, the expression trend of the DEGs is positively correlated with the accumulation of glycosides. Our study findings indicate that key DEGs and crucial UGT genes co-regulate flavonoid and saponin biosynthesis in licorice under salt stress.

Variations in morphology, physiology, and multiple bioactive constituents of Lonicerae Japonicae Flos under salt stress.

Lonicerae Japonicae Flos (LJF) is an important traditional Chinese medicine for the treatment of various ailments and plays a vital role in improving global human health. However, as unable to escape from adversity, the quality of sessile organisms is dramatically affected by salt stress. To systematically explore the quality formation of LJF in morphology, physiology, and bioactive constituents' response to multiple levels of salt stress, UFLC-QTRAP-MS/MS and multivariate statistical analysis were performed. Lonicera japonica Thunb. was planted in pots and placed in the field, then harvested after 35 days under salt stress. Indexes of growth, photosynthetic pigments, osmolytes, lipid peroxidation, and antioxidant enzymes were identified to evaluate the salt tolerance in LJF under different salt stresses (0, 100, 200, and 300 mM NaCl). Then, the total accumulation and dynamic variation of 47 bioactive constituents were quantitated. Finally, Partial least squares discrimination analysis and gray relational analysis were performed to systematically cluster, distinguish, and evaluate the samples, respectively. The results showed that 100 mM NaCl induced growth, photosynthetic, antioxidant activities, osmolytes, lipid peroxidation, and multiple bioactive constituents in LJF, which possessed the best quality. Additionally, a positive correlation was found between the accumulation of phenolic acids with antioxidant enzyme activity under salt stress, further confirming that phenolic acids could reduce oxidative damage. This study provides insight into the quality formation and valuable information to improve the LJF medicinal value under salt stress.

Quality evaluation of Lonicerae Japonicae Flos and Lonicerae Flos based on simultaneous determination of multiple bioactive constituents combined with multivariate statistical analysis.

INTRODUCTION: Lonicerae Japonicae Flos (LJF) and Lonicerae Flos (LF) belong to different genera of Caprifoliaceae. They have been historically utilised as herbal medicine to treat various diseases. However, the comprehensive assessment of them still remains a challenge. OBJECTIVE: To develop a comprehensive method of ultra-fast liquid chromatography-tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) coupled with multivariate statistical analysis for the quality evaluation and reveal differential components of LJF and LF. METHODOLOGY: A validated UFLC-QTRAP-MS/MS method was established for simultaneous determination of 50 constituents, including 12 organic acids, 12 flavonoids, 6 iridoids, 3 saponins, 13 amino acids and 4 nucleosides. The obtained data were employed to multivariate statistical analysis. Principal component anlysis (PCA) and partial least squares determinant analysis (PLS-DA) were performed to classify and reveal differential components of samples; grey relational analysis (GRA) was introduced to assess the samples according to the contents of 50 constituents by calculating the relative correlation degree of each sample. RESULTS: Fifty constituents were simultaneously determined of LJF and LF. Based on obtained data, PCA and PLS-DA were easy to distinguish samples and the classification of the samples was related to 11 chemical constituents. GRA implied the quality of LJF was better, and that the flower buds were superior to the flowers. Moreover, organic acids are the main components of samples. CONCLUSION: This study not only established a method of simultaneous determination of multiple bioactive constituents in LJF and LF, but provided comprehensive information on the quality control of them. The developed method is conducive to distinguish orthologues or paralogues of them, and supply the support for "heterologous effects".

Qualitative and quantitative analysis of the major constituents in Spatholobi Caulis by UFLC-Triple TOF-MS/MS and UFLC-QTRAP-MS/MS.

There have been few comprehensive studies on the holistic chemical composition of Spatholobi Caulis (SC) and consequently, the information is lacking for the in-depth study of the major constituents. SC is a kind of widely used traditional Chinese medicine with its xylem and phloem alternately arranged in 3-10 rings, but the relationship of phloem ring number and the quality remains unclear. In this study, the characterization of the major constituents in SC was analyzed by ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-Triple TOF-MS/MS), and the content of 19 flavonoids in SC with different phloem ring numbers was simultaneously determined by ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap tandem mass spectrometry (UFLC-QTRAP-MS/MS). Correlation analysis was performed to evaluate the quality of SC with different phloem ring numbers according to the content of 19 flavonoids. Results showed that 50 constituents in SC were identified and the fragmentation pathways of different types of compounds were preliminarily deduced by the fragmentation behavior of the 50 constituents. In addition, the content of flavonoids increased with phloem ring number, which demonstrated that the content of flavonoids in SC was positively correlated with the number of phloem rings. Our research will contribute to the variety identification and quality evaluation of SC, and provide a scientific basis for evaluating the quality of medicinal materials based on its appearance and characteristics.

Metabolomics characterizes the metabolic changes of Lonicerae Japonicae Flos under different salt stresses.

Salt stress affects the metabolic homeostasis of medicinal plants. However, medicinal plants are sessile organisms that cannot escape from salt stress. They acclimatize themselves to the stress by reprogramming their metabolic pathways. Lonicerae Japonicae Flos (LJF) with strong antioxidant activity is commonly used in traditional Chinese medicine, tea, and beverage. Nevertheless, the variation of integrated metabolites in LJF under different salt stresses remains unclear. In this study, High Performance Liquid Chromatography tandem triple time-of-flight mass spectrometry (HPLC- triple TOF-MS/MS) coupled with multivariate statistical analysis was applied to comparatively investigate the metabolites changes in LJF under different salt stress (0, 100, 200, 300 mM NaCl). Total 47 differential metabolites were screened from 79 metabolites identified in LJF under different salt stress. Low salt-treated group (100 mM NaCl) appeared to be the best group in terms of relative contents (peak areas) of the wide variety in bioactive components. Additionally, the phenylpropanoid pathway, monoterpenoid biosynthesis, glycolysis, TCA cycle, and alkaloid biosynthesis were disturbed in all salt-stress LJF. The results showed that LJF metabolisms were dramatically induced under salt stress and the quality of LJF was better under low salt stress. The study provides novel insights into the quality assessment of LJF under salt stress and a beneficial framework of knowledge applied to improvement the medicinal value of LJF.

[Analysis and evaluation of dynamic accumulation of multiple bioactive constituents in Spatholobi Caulis].

A method was established for simultaneous determination of 21 active constituents including flavanols, isoflavones, flavonols, dihydroflavones, dihydroflavonols, chalcones, pterocarpan, anthocyanidins and phenolic acids in Spatholobi Caulis by ultra fast liquid chromatography with triple quadrupole linear ion trap mass spectrometry(UFLC-QTRAP-MS/MS). Then, it was employed to analyze and evaluate the dynamic accumulation of multiple bioactive constituents in Spatholobi Caulis. The chromatographic separation was performed on a XBridge®C_(18)(4.6 mm×100 mm, 3.5 µm) at 30 ℃ with a gradient elution of 0.3% formic acid aqueous solution-methanol, and the flow rate was 0.8 mL·min~(-1), using multiple-reaction monitoring(MRM) mode. A comprehensive evaluation of the multiple bioactive constituents was carried out by gray correlation analysis(GRA). The 21 target components showed good linearity(r>0.999 0) in the range of the tested concentrations. The average recovery rates of the 21 components were from 97.46% to 103.6% with relative standard deviations less than 5.0%. There were differences in the contents of 21 components in Spatholobi Caulis at diffe-rent harvest periods. Spatholobi Caulis had high quality from early November to early December, which is consistent with the local tradi-tional harvest period. This study reveals the rule of the dynamic accumulation of 21 components in Spatholobi Caulis and provides basic information for the suitable harvest time. At the same time, it provides a new method reference for the comprehensive evaluation of the internal quality of Spatholobi Caulis.

[Research progress in molecular biology of Lonicerae Japonicae Flos].

Molecular biology is a new subject that clarifies the phenomena and nature of life at the molecular level. Its development provides new biotechnology and methods for the study of traditional pharmacognosy. The formation of molecular biology has brought the development of pharmacognosy into a new era of gene research. Lonicerae Japonicae Flos is a classical Chinese medicine. Many scholars of home and abroad have carried out relevant studies on its molecular biology on the basis of the in-depth study with traditional methods, and have achieved certain results. In order to provide references on the method, technical for promoting the modernization of Lonicerae Japonicae Flos, and the development, protection, and utilization of other traditional Chinese medicine resources. This article summarized the application status of molecular biology methods and techniques on the identification, biosynthesis of active constituents, and molecular mechanism of secondary metabolite under stress conditions of Lonicerae Japonicae Flos in recent years. In hybridization technology of tag(RFLP), molecular markers based on PCR(RAPD, AFLP, SSR and ISSR), based on DNA sequence analysis of SNP and DNA barcode for the variety identification, diagnosis, identification of Lonicerae Japonicae Flos, and so forth in detail. At the same time, it is proposed that multi-omics technology can be used to build systems biology technology and platforms, and establish related models of secondary metabolite biosynthesis, so as to deepen acknowledge the molecular mechanism of the active component biosynthesis of Lonicerae Japonicae Flos and the accumulation of metabolites, life activities of other medicinal plants under adverse environment, then to regulate them.

A comprehensive study of the aerial parts of Lonicera japonica Thunb. based on metabolite profiling coupled with PLS-DA.

INTRODUCTION: Lonicera japonica Thunb. is an economically important species of honeysuckle belonging to the Caprifoliaceae family. All aerial parts of L. japonica (leaf, flower bud, flower, and caulis) are used as herbal remedies in traditional Chinese medicine. The application of plant metabolomics to the study of L. japonica provides the potential for identifying the phytochemical composition and useful chemical markers of the plant. OBJECTIVE: To develop a strategy integrating metabolic profiling and partial least squares discriminant analysis (PLS-DA) to separate the aerial parts of L. japonica based on the occurrence of chemical markers. METHODOLOGY: The two-part strategy consisted of (1) ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-triple TOF-MS/MS), (2) PLS-DA, which was applied to distinguish between the different aerial parts and reveal their differential characteristic metabolites. RESULTS: A total of 71 metabolites were identified from samples, and eight candidate compounds were identified (lonicerin, kaempferol-3-O-rutinoside, loganin, isochlorogenic acid B, isochlorogenic acid C, secologanic acid, luteoloside, astragalin) as optimal chemical markers based on variable importance in projection (VIP) and p-value. The relative contents of eight candidate compounds were compared based on their peak intensities. CONCLUSION: This study established an efficient strategy for exploring metabolite profiling and defining chemical markers among the different aerial parts of L. japonica, and laid the foundation for elucidating the phytochemical differences in efficacy between Lonicerae Japonicae Flos (LJF) and Lonicerae Japonicae Caulis (LJC). Our findings also indicate that the leaves of L. japonica leaf could be used as an alternative medicinal resource for LJF and provide a reference for comprehensive exploitation and utilisation of L. japonica resources.

Distribution Patterns for Bioactive Constituents in Pericarp, Stalk and Seed of Forsythiae Fructus.

Forsythiae Fructus (FF) is a widely used folk medicine in China, Japan, and Korea. The distribution of bioactive constituents throughout the fruit segments has rarely been addressed, although mounting evidence suggests that plant secondary metabolites are synthesized and distributed regularly. The phytochemical profiles of three segments of FF (pericarp, stalk and seed) were firstly revealed by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based quantitative analysis of twenty-one bioactive constituents, including three phenylethanoid glycosides, five lignans, eight flavonoids, and five phenolic acids to explore the spatial distribution of bioactive constituents. Furthermore, the hierarchical clustering analysis (HCA) and one-way analysis of variance (one-way ANOVA) were conducted to visualize and verify the distribution regularity of twenty-one analytes among three segments. The results showed that phytochemical profiles of the three segments were similar, i.e., phenylethanoid glycosides covering the most part were the predominant compounds, followed by lignans, flavonoids and phenolic acids. Nevertheless, the abundance of twenty-one bioactive constituents among three segments was different. Specifically, phenylethanoid glycosides were highly expressed in the seed; lignans were primarily enriched in the stalk; flavonoids were largely concentrated in the pericarp, while the contents of phenolic acids showed no much difference among various segments. The research improves our understanding of distribution patterns for bioactive constituents in FF, and also complements some scientific data for further exploring the quality formation mechanism of FF.

A Comprehensive Review for Phytochemical, Pharmacological, and Biosynthesis Studies on Glycyrrhiza spp.

Licorice is extensively applied in food as well as herbal medicine across the world, possessing a substantial share in the global market. It has made great progress in chemical and pharmacological research in recent years. Currently, Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat., and Glycyrrhiza glabra L. were officially used as Gan-Cao according to the Chinese Pharmacopoeia. Accumulating evidence demonstrated three varieties of licorice have their own special compounds except for two quality markers set by Pharmacopoeia, providing great possibility for better understanding their characteristics, evaluating quality of each species and studying biosynthesis mechanisms of species-specific compounds. As a special "guide drug" in clinic, licorice plays an important role in Chinese herbal formulas. The interaction between licorice with other ingredients and their metabolism in vivo should also be taken into consideration. In addition, draft genome annotation, and success of the final step of glycyrrhizin biosynthesis have paved the way for biosynthesis of other active constituents in licorice, a promising beginning of solving source shortage. Accordingly, we comprehensively explored the nearly 400 chemical compounds found in the three varieties of licorice so far, systematically excavated various pharmacological activities, including metabolism via CYP450 system in vivo, and introduced the complete biosynthesis pathway of glycyrrhizin in licorice. The review will facilitate the further research toward this herbal medicine.

Comparative Proteomic Analysis Reveals the Molecular Mechanisms Underlying the Accumulation Difference of Bioactive Constituents in Glycyrrhiza uralensis Fisch under Salt Stress.

Licorice (Glycyrrhiza uralensis Fisch) possesses a substantial share of the global markets for its unique sweet flavor and diverse pharmacological compounds. Cultivated licorice is widely distributed in northwest regions of China, covered with land with a broad range of salinities. A preliminary study indicated that suitable salt stress significantly increased the content of bioactive constituents in licorice. However, the molecular mechanisms underlying the influence of salinity on the accumulation of these constituents remain unclear, which hinders quality breeding of cultivated licorice. In our study, flavonoid-related structural genes were obtained, and most of them, such as phenylalanine ammonia-lyases, cinnamate 4-hydroxylases, 4-coumarate: CoA ligases, chalcone synthases, chalcone-flavanone isomerase, and flavonol synthase, showed high levels after salt treatment. In the biosynthesis of glycyrrhizin, three key enzymes (bAS, CYP88D6, and CYP72A154) were identified as differentially expressed proteins and remarkably upregulated in the salt-stressed group. Combining these results with the contents of 14 bioactive constituents, we also found that the expression patterns of those structural proteins were logically consistent with changes in bioactive constituent profiles. Thus, we believe that suitable salt stress increased the accumulation of bioactive constituents in licorice by upregulating proteins involved in the related biosynthesis pathways. This work provided valuable proteomic information for unraveling the molecular mechanism of flavonoid and glycyrrhizin metabolism and offered fundamental resources for quality breeding in licorice.

Qualitative and quantitative analysis of saponins in the flower bud of Panax ginseng (Ginseng Flos) by UFLC-Triple TOF-MS/MS and UFLC-QTRAP-MS/MS.

INTRODUCTION: Ginseng Flos (GF), the flower bud of Panax ginseng, is a worthy functional food with medicinal potential. A few studies have focused on the comprehensive and systematic analysis of its major bioactive constituents. OBJECTIVE: The aims are to develop the methods of ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-Triple TOF-MS/MS) and ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap tandem mass spectrometry (UFLC-QTRAP-MS/MS) for the qualitative and quantitative analysis of the saponins in GF. METHODOLOGY: UFLC-Triple TOF-MS/MS and UFLC-QTRAP-MS/MS were established for the qualitative and quantitative analysis of the saponins in GF, separately. RESULTS: Fifty-one saponins were identified in GF using UFLC-Triple TOF-MS/MS method; among them, 21 saponins were characterized by comparing with standards. Furthermore, 12 ginsenosides (ginsenoside Re, Rg1 , Rf, 20(S)-Rg2 , 20(R)-Rg2 , Rb1 , Rc, Ro, Rb2 , F1 , Rd, and F2 ) were synchronously determined by UFLC-QTRAP-MS/MS method after the extraction with 70% methanol. This UFLC-QTRAP-MS/MS method showed good linearity (r >0.9991), the interday and intraday precision, repeatability and stability were all satisfied, the average recoveries of standard addition for the compounds were between 94.01% and 105.16%, and the relative standard deviations were less than 5%. CONCLUSION: The results are available for the comprehensive quality control and assessment of GF and its relative products.

Dynamic Variations in Multiple Bioactive Constituents under Salt Stress Provide Insight into Quality Formation of Licorice.

The demand for licorice and its natural product derivatives in domestic and foreign market is considerably huge. The core production areas of licorice are covered with salinity and drought land in northwestern China. Studies have shown that suitable environmental stress can promote the accumulation of glycyrrhizin and liquiritin to improve its quality as medicinal materials. However, there are few reports on other bioactive constituents of licorice, not to mention their dynamic accumulation under stressed conditions. To explore the quality formation of licorice from the perspective of salt influence, a reliable method based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-MS/MS) was established for simultaneous determination of sixteen bioactive constituents, including triterpenoids, flavonoids, chalcones and their glycosides. Physiological experiments were performed to investigate salt tolerance of licorice under different salinity treatments. The expressions of crucial genes (bAS and CHS), key enzymes of triterpenoid and flavonoid synthesis, were also tested by qRT-PCR. Our study found that 50 mM NaCl treatment (low stress) was the most favorable to promote the accumulation of bioactive constituents in the long term, without harming the plants. Flavonoid accumulation of non-stressed and low-stressed groups became different in the initial synthesis stage, and glycosyltransferases may have great influence on their downstream synthesis. Furthermore, bAS and CHS also showed higher levels in low-stressed licorice at harvest time. This work provides valuable information on dynamic variations in multiple bioactive constituents in licorice treated by salt and insight into its quality formation under stressed conditions.

Comparative Metabolite Profiling of Wild and Cultivated Licorice Based on Ultra-Fast Liquid Chromatography Coupled with Triple Quadrupole-Time of Flight Tandem Mass Spectrometry.

Licorice is one of the ancient and most frequently applied herbs for its diverse phytochemicals. At present, wild resources of licorice have rapidly declined with increasing demand and the proportion of cultivated products in the market is quickly growing. However, the different level in chemical composition between the wild and cultivated licorice may result in the discrepancy in quality and pharmacological activity. Therefore, an ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-Triple TOF-MS/MS) method combined with multivariate statistical analysis technology was employed to explore chemical composition differences. The result showed that total 63 components were identified from licorice samples. The wild and the cultivated licorice are obviously classified into two groups according to principal component analysis (PCA). PCA and partial least squared discrimination analysis (PLS-DA) were also introduced to rapidly find 14 candidate compounds between two ecotypes of licorice. Apart from glycyrrhizin, licorice saponin J2/G2, glyasperin D and dehydroglyasperin D also could be selected as chemical markers based on t-test and variable importance in the projection (VIP) value. Our study successfully established an effective method for exploring metabolite profiling between two ecotypes of licorice and laying the foundation for distinguishing wild and cultivated licorice.

Quality Evaluation of Ophiopogonis Radix from Two Different Producing Areas.

Ophiopogonis Radix, also known as Mai-dong in Chinese, was a commonly used traditional Chinese medicine (TCM) and functional health food. Two products of Ophiopogonis Radix are largely produced in the Sichuan and Zhejiang province, which are called "Chuan maidong (CMD)" and "Zhe maidong (ZMD)" respectively. To distinguish and evaluate the quality of CMD and ZMD, an analytical method based on ultra-fast performance liquid chromatography coupled with triple quadrupole-linear ion trap mass spectrometry (UFLC-QTRAP-MS/MS) was established for simultaneous determination of 32 constituents including 4 steroidal saponins, 3 homisoflavonoids, 15 amino acids, and 10 nucleosides in 27 Mai-dong samples from Sichuan and Zhejiang. Furthermore, principal components analysis (PCA), partial least squares discriminant analysis (PLS-DA), t-test, and grey relational analysis (GRA) were applied to discriminate and evaluate the samples from Sichuan and Zhejiang based on the contents of 32 constituents. The results demonstrated that the bioactive constituents in CMD and ZMD were significantly different, and CMD performed better in the quality assessment than ZMD. This study not only provides a basic information for differentiating CMD and ZMD, but offers a new insight into comprehensive evaluation and quality control of Ophiopogonis Radix from two different producing areas.

Qualitative and Quantitative Analysis of the Saponins in Panacis Japonici Rhizoma Using Ultra-Fast Liquid Chromatography Coupled with Triple Quadrupole-Time of Flight Tandem Mass Spectrometry and Ultra-Fast Liquid Chromatography Coupled with Triple Quadrupole-Linear Ion Trap Tandem Mass Spectrometry.

Panacis Japonici Rhizoma (PJR) contains various kinds of saponins, which possesses extensive pharmacological activities, but studies of comprehensive analysis of its saponins were limited. Thus, ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-Triple TOF-MS/MS) and ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap tandem mass spectrometry (UFLC-QTRAP-MS/MS) methods were established for the qualitative and quantitative analysis of the saponins in PJR, separately. Fifty three saponins in PJR were identified by UFLC-Triple TOF-MS/MS method, 23 saponins of which were unequivocally identified by reference substances. In addition, fragmentation pathways of different types of saponins were preliminarily deduced by fragmentation behavior of 53 saponins. Furthermore, the simultaneous determination of the contents of 13 saponins in PJR samples harvested at different times were analyzed by UFLC-QTRAP-MS/MS method. Furthermore, the quality of the samples was evaluated by grey relational analysis. This study might be beneficial to the quality assessment and control of PJR. Meanwhile, it might provide the basic information for confirming its optimal harvested period.

Comparison of Multiple Bioactive Constituents in the Flower and the Caulis of Lonicera japonica Based on UFLC-QTRAP-MS/MS Combined with Multivariate Statistical Analysis.

Lonicerae japonicae flos (LJF) and Lonicerae japonicae caulis (LJC) are derived from different parts of Lonicera japonica Thunb. (Caprifoliaceae), and have been used as herbal remedies to treat various diseases for thousands of years with confirmed curative effects. However, little attention has been paid to illustrating the differences in efficacy from the perspective of phytochemistry. In the present study, a simultaneous determination of 47 bioactive constituents, including 12 organic acids, 12 flavonoids, six iridoids, 13 amino acids and four nucleosides in 44 batches of LJF and LJC samples from different habitats and commercial herbs was established based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS). Moreover, principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and t-test were then performed to classify and reveal the differential compositions of LJF and LJC according to the content of the tested constituents. The results demonstrated that the types and contents of chemical components (e.g., isochlorogenic acid A, chlorogenic acid, neochlorogenic acid, quinic acid, secologanic acid, luteoloside, loganin, secoxyloganin, morroniside and L-isoleucine) were significantly different, which may lead to the classification and the differences in efficacy of LJF and LJC. Our findings not only provide a basis for the comprehensive evaluation and intrinsic quality control of LJF and LJC, but also pave the way for discovering the material basis contributing to the different properties and efficacies of the two medicinal materials at the phytochemical level.

Quality Evaluation of Wild and Cultivated Schisandrae Chinensis Fructus Based on Simultaneous Determination of Multiple Bioactive Constituents Combined with Multivariate Statistical Analysis.

Schisandrae Chinensis Fructus, also called wuweizi in China, was a widely used folk medicine in China, Korea, and Russia. Due to the limited natural resources and huge demand of wuweizi, people tend to cultivate wuweizi to protect this species. However, the quality of wild and cultivated herbs of the same species may change. Little attention has been paid to comparing wild and cultivated wuweizi based on simultaneous determination of its active components, such as lignans and organic acids. An analytical method based on UFLC-QTRAP-MS/MS was used for the simultaneous determination of 15 components, including 11 lignans (schisandrin, gomisin D, gomisin J, schisandrol B, angeloylgomisin H, schizantherin B, schisanhenol, deoxyschizandrin, γ-schisandrin, schizandrin C, and schisantherin) and 4 organic acids (quinic acid, d(-)-tartaric acid, l-(-)-malic acid, and protocatechuic acid) in wuweizi under different ecological environments. Principal components analysis (PCA), partial least squares discrimination analysis (PLS-DA), independent sample t-test, and gray relational analysis (GRA) have been applied to classify and evaluate samples from different ecological environments according to the content of 15 components. The results showed that the differential compounds (i.e., quinic acid, l-(-)-malic acid, protocatechuic acid, schisandrol B) were significantly related to the classification of wild and cultivated wuweizi. GRA results demonstrated that the quality of cultivated wuweizi was not as good as wild wuweizi. The protocol not just provided a new method for the comprehensive evaluation and quality control of wild and cultivated wuweizi, but paved the way to differentiate them at the chemistry level.