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.

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.

A Method to Study the Distribution Patterns for Metabolites in Xylem and Phloem of Spatholobi Caulis.

Spatholobi Caulis (SC), the vine stem of Spatholobus suberectus Dunn, is a widely used traditional Chinese medicine (TCM) for the treatment of blood stasis syndrome and related diseases. Xylem and phloem are the main structures of SC and the color of xylem in SC is red brown or brown while the phloem with resin secretions is reddish brown to dark brown. They are alternately arranged in a plurality of concentric or eccentric rings. In order to investigate the distribution patterns of metabolites in xylem and phloem of SC, an analytical method based on UFLC-QTRAP-MS/MS was established for simultaneous determination of 22 constituents including four flavanols, nine isoflavones, two flavonols, two dihydroflavones, one flavanonol, one chalcone, one pterocarpan, one anthocyanidin and one phenolic acid in the samples (xylem and phloem) from Laos. Furthermore, according to the contents of 22 constituents, heat map, principal components analysis (PCA), orthogonal partial least squares discriminant analysis (OPLS-DA) and t-test were used to evaluate the samples and discover the differences between xylem and phloem of SC. The results indicated that the measured ingredients in xylem and phloem were significantly different. To be specific, the contents of flavonoids in xylem were higher than that in phloem, while the content of protocatechuic acid showed a contrary tendency. This study will not only reveal the distribution patterns of metabolites in xylem and phloem of SC but also facilitate further study on their quality formation.

[Stimultaneous determination of multiple bioactive constituents in Panacis Japonici Rhizoma processed by different methods and grey relational analysis].

A method, for determination of saponins, amino acids and nucleosides in Panacis Japonici Rhizoma of ultra fast liquid chromatography with triple quadrupole linear ion trap mass spectrometry (UFLC-QTRAP-MS/MS), was established to investigate the effect of different processing methods on the target components of Panacis Japonici Rhizoma. The chromatographic separation was performed on a XBridgeC18(4.6 mm×100 mm, 3.5 µm) at 30 °C with a gradient elution of 0.1% formic acid solution-0.1% formic acid acetonitrile, and the flow rate was 0.8 mL·min⁻¹, using multiple-reaction monitoring (MRM) mode. The grey relational analysis was adopted for the analysis of different processing samples. The results showed that the thirty-three constituents were in a good linear range and the correlation coefficient was greater than 0.999 0; the precision, repeatability and stability were good; the average recovery rates were between 95.33% and 101.8%, and the relative standard deviations were less than 5%. The result of grey relational analysis showed that the complete rhizomes without peeling, which were adopted for the microwave dried method, had the best quality. The established method was accurate and reliable, which could be used to appraise the quality of Panacis Japonici Rhizoma. Our study may lay the way for the processing method of Panacis Japonici Rhizoma in optimization,normalization and standardization.

[Comparative analysis of multiple index constituents in Ophiopogonis Radix and Liriopes Radix].

An analytical method based on UFLC-QTRAP-MS/MS was established for simultaneous determination of thirty-three components including steroidal saponins, homoisoflavonoids, amino acids and nucleosides in Ophiopogonis Radix. Thirty-three target components of commercial medicinal materials of Maidong were comparative analysis. Synergi™ Hydro-RP 100 column (2.0 mm × 100 mm, 2.5 µm) was used with 0.1% formic acid solution-0.1% formic acid acetonitrile for gradient elution at a flow rate of 0.4 mL·min⁻¹. In addition, multiple reaction monitoring (MRM) mode was employed. The data were comprehensively processed and analyzed with hierarchical clustering analysis(HCA), principal component analysis(PCA) and partial least squares discriminant analysis(PLS-DA) methods. All components showed good linearity(r>0.999 0) within the tested ranges. The average recoveries were between 96.23%-102.0%, and the relative standard deviation(RSD) were less than 5%. The results showed that there were significant differences in components between Ophiopogonis Radix and Liriopes Radix, with seven components obviously different. This method was useful for providing basis for the comprehensive evaluation and intrinsic quality control of Ophiopogonis Radix and Liriopes Radix , and may provide a new method reference for the identification of Ophiopogonis Radix and Liriopes Radix.

Variations in Physiology and Multiple Bioactive Constituents under Salt Stress Provide Insight into the Quality Evaluation of Apocyni Veneti Folium.

As one of the major abiotic stresses, salinity stress may affect the physiology and biochemical components of Apocynum venetum L. To systematically evaluate the quality of Apocyni Veneti Folium (AVF) from the perspective of physiological and the wide variety of bioactive components response to various concentrations of salt stress, this experiment was arranged on the basis of ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) technology and multivariate statistical analysis. Physiological characteristics of photosynthetic pigments, osmotic homeostasis, lipid peroxidation product, and antioxidative enzymes were introduced to investigate the salt tolerance mechanism of AVF under salinity treatments of four concentrations (0, 100, 200, and 300 mM NaCl, respectively). Furthermore, a total of 43 bioactive constituents, including 14 amino acids, nine nucleosides, six organic acids, and 14 flavonoids were quantified in AVF under salt stress. In addition, multivariate statistical analysis, including hierarchical clustering analysis, principal component analysis (PCA), and gray relational analysis (GRA) was employed to systematically cluster, distinguish, and evaluate the samples, respectively. Compared with the control, the results demonstrated that 200 mM and 100 mM salt stress contributed to maintain high quality of photosynthesis, osmotic balance, antioxidant enzyme activity, and the accumulation of metabolites, except for total organic acids, and the quality of AVF obtained by these two groups was better than others; however, under severe stress, the accumulation of the oxidative damage and the reduction of metabolite caused by inefficiently scavenging reactive oxygen species (ROS) lead to lower quality. In summary, the proposed method may provide integrated information for the quality evaluation of AVF and other salt-tolerant Chinese medicines.

Distribution patterns for metabolites in medicinal parts of wild and cultivated licorice.

Licorice is a famous Chinese medicinal material widely applied worldwide in food and drugs. It possesses a substantial share in the international and domestic markets. While the demand is continuously increasing, wild-type of licorice is gradually disappearing. Its cultivated-type is significantly different compared to its wild-type, especially the bioactive compounds in dried roots and rhizomes (the medicinal parts of wild and cultivated licorice) are responsible for their quality difference. In this study, a total of thirty-two constituents, including seventeen secondary compounds and fifteen primary metabolites, were simultaneously analyzed by UFLC coupled with triple quadrupole-linear ion trap mass spectrometry (UFLC-QTRAP-MS/MS) in different medicinal parts of licorice. Our findings indicated that the content of major bioactive compounds in wild licorice were significantly higher than those in its cultivated-type. The metabolites in rhizomes showed totally different outcomes between wild and cultivated licorice. Remarkably high level of some important amino acids related to abiotic stress (drought and salt) were found in wild licorice. Notably, the high contents of phenylalanine and compounds in upstream of flavonoid synthesis in cultivated licorice suggested that the synthesis may differ at the initial stage. The distribution pattern of metabolites in different medicinal parts of wild and cultivated licorice will not only provide a novel clue in agricultural breeding but also facilitate the further study on their quality formation.