Residue of Chlormequat and Regulatory Effects on the Specialized Metabolites of Astragali Radix.

Presently, the utilization of chlormequat in Astragalus mongholicus Bunge (Leguminosae) cultivation is prevalent for augmenting rhizome (Astragali Radix) yield. However, indiscriminate and excessive chlormequat employment can detrimentally influence Astragali Radix quality and safety. This research aimed to comprehensively comprehend chlormequat risks and its influence on Astragali Radix metabolites. Diverse chlormequat concentrations were employed in Astragalus mongholicus cultivation, with subsequent analysis of residual chlormequat levels in Astragali Radix across treatment groups. Astragali Radix metabolic profiling was conducted through UPLC-QTOF-MS, and thirteen principal active components were quantified via UFLC-MS/MS. Findings revealed a direct correlation between chlormequat residue levels in Astragali Radix and application concentration, with high-dose residue surpassing 5.0 mg/kg. Metabolomics analysis identified twenty-six distinct saponin and flavonoid metabolites. Notably, the application of chlormequat led to the upregulation of seven saponins (e.g., astragaloside I and II) and downregulation of six flavonoids (e.g., methylnissolin-3-O-glucoside and astraisoflavan-7-O-ß-d-glucoside). Quantitative analysis demonstrated variable contents of active ingredients due to differing chlormequat concentrations, leading to astragaloside I increase (14.59-62.55%) and isoastragaloside II increase (4.8-55.63%), while methylnissolin-3-O-glucoside decreased (22.18-41.69%), as did astraisoflavan-7-O-ß-d-glucoside (21.09-47.78%). In conclusion, chlormequat application influenced multiple active components in Astragali Radix, causing constituent proportion variations. Elevated chlormequat concentrations led to increased active components alongside heightened chlormequat residues in Astragali Radix. Consequently, prudent chlormequat application during Astragali Radix production is imperative to avert potential detriments to its quality and safety.

Advances in DNA methylation and demethylation in medicinal plants: a review.

DNA methylation and demethylation are widely acknowledged epigenetic phenomena which can cause heritable and phenotypic changes in functional genes without changing the DNA sequence. They can thus affect phenotype formation in medicinal plants. However, a comprehensive review of the literature summarizing current research trends in this field is lacking. Thus, this review aims to provide an up-to-date summary of current methods for the detection of 5-mC DNA methylation, identification and analysis of DNA methyltransferases and demethyltransferases, and regulation of DNA methylation in medicinal plants. The data showed that polyploidy and environmental changes can affect DNA methylation levels in medicinal plants. Changes in DNA methylation can thus regulate plant morphogenesis, growth and development, and formation of secondary metabolites. Future research is required to explore the mechanisms by which DNA methylation regulates the accumulation of secondary metabolites in medicinal plants.

Effect of exogenous phytohormones treatment on glycyrrhizic acid accumulation and preliminary exploration of the chemical control network based on glycyrrhizic acid in root of Glycyrrhiza uralensis

ABSTRACT One-year-old Glycyrrhiza uralensis Fisch. ex DC, Fabaceae, was treated with three exogenous phytohormones in June and July, namely gibberellin, auxin (indole-3-acetic acid), methyl jasmonate at different concentrations. Control plants were treated with water. Roots of controls and hormones-treated G. uralensis plants were harvested at different times, and the contents of seven main chemical components were determined. Root glycyrrhizic acid content of plants treated in June increased significantly compared with controls, and the difference was significant. As for plants treated in July, root glycyrrhizic acid content increased in which sprayed with appropriate concentrations of hormones, but the effects of hormones were more evident in plants treated in June coincided with the vigorous growth period than those treated in July. Gibberellin at 40 mg/l and auxin at 40 mg/l applied in the two treatment periods significantly promoted the accumulation of glycyrrhizic acid in G. uralensis root. Treatment with methyl jasmonate at 100 and 25 mg/l in June and July, respectively, also increased glycyrrhizic acid content significantly. The determination of major active compositions indicated that liquiritin, isoliquiritin, isoliquiritin apioside and liquiritin apioside contents were positively related to glycyrrhizic acid content. The study preliminarily found phytohormones and the main chemical components associated with glycyrrhizic acid content, and these discoveries could provide a basis for establishing a chemical control network with glycyrrhizic acid as the core, confirming the secondary product metabolic pathways in the network and completely uncovering synthesis mechanism underlying glycyrrhizic acid-combined functional gene polymorphism.

Sources of varieties and quality of circular Fructus Ligustri Lucidi.

This study aimed to trace sources and quantitatively analyze the specnuezhenide content of circular Fructus Ligustri Lucidi for clinical use. Different specifications of Fructus Ligustri Lucidi were identified using DNA barcoding technology and the specnuezhenide content was analyzed by High Performance Liquid Chromatography (HPLC). The ITS sequence of circular Fructus Ligustri Lucidi was identical to that of standard privet, which was determined through botanical identification. ITS sequence similarity between circular Fructus Ligustri Lucidi and Fructus Ligustri Lucidi which was registered in NCBI ranged from 99.5% to 100%. The sequences of circular and other Fructus Ligustri Lucidi were clustered in a Neighbor-Joining tree with bootstrap value of 95, and these sequences could be distinguished from adulterants. Conforming to pharmacopoeia standard, the average specnuezhenide content of circular Fructus Ligustri Lucidi was higher than that of chicken waist Fructus Ligustri Lucidi. Circular Fructus Ligustri Lucidi derived from Ligustrum lucidum Ait. and the specnuezhenide content was higher in circular Fructus Ligustri Lucidi than that in chicken waist Fructus Ligustri Lucidi.

Spectrum-Effect Relationships Between Chemical Fingerprints and Antibacterial Effects of Lonicerae Japonicae Flos and Lonicerae Flos Base on UPLC and Microcalorimetry.

The traditional Chinese medicines Lonicerae Japonicae Flos (LJF, Jinyinhua in Chinese) and Lonicerae Flos (LF, Shanyinhua in Chinese) refer to the flower buds of five plants belonging to the Caprifoliaceae family. Until 2000, all of these were officially listed as a single item, LJF (Jinyinhua), in the Chinese Pharmacopoeia. However, there have recently been many academic controversies concerning the separation and combination of LJF and LF in administrative regulation. Till now there has been little work completed evaluating the relationships between biological activity and chemical properties among these drugs. Microcalorimetry and UPLC were used along with principal component analysis (PCA), hierarchical cluster analysis (HCA), and canonical correlation analysis (CCA) to investigate the relationships between the chemical ingredients and the antibacterial effects of LJF and LF. Using multivariate statistical analysis, LJF and LF could be initially separated according to their chemical fingerprints, and the antibacterial effects of the two herbal drugs were divided into two clusters. This result supports the disaggregation of LJF and LF by the Pharmacopoeia Committee. However, the sample of Lonicera fulvotomentosa Hsu et S. C. Cheng turned out to be an intermediate species, with similar antibacterial efficacy as LJF. The results of CCA indicated that chlorogenic acid and 3,4-Dicaffeoylquinic acid were the major components generating antibacterial effects. Furthermore, 3,4-Dicaffeoylquinic acid could be used as a new marker ingredient for quality control of LJF and LF.

[Effect of exogenous brassinolide on morphological characters and contents of seven chemical constituents of Glycyrrhiza uralensis].

The transplants of the two-year-old Glycyrrhiza uralensis were subjected to four concentration of brassinolide (BR 0.1, 0.4, 0.7, 1.0 mg•L⁻¹) in July. The morphological characters ( plant height, stem diameter, nodes number, internode length and root length , root thick, root fresh weight and root dry weight ) were measured and seven kinds of chemical constituents (glycyrrhizic acid, liquiritin, isoliquiritin, liquiritigenin, isoliquiritigenin, liquiritin apioside, isoliquiritin apioside) were determined by HPLC with the aim of increasing sinter output and improving quality of G. uralensis. Then the long-term dynamic changes of these morphological characters and chemical compositions' content were analyzed. The results showed that morphological characters of plant height, stem diameter, root length , root thick, root fresh weight and root dry weight increased remarkably with the 0.7 mg•L⁻¹ BR stimulating 2 months later,the increase rates were： 15.09%,6.15%,16.52%,8.46%,21.90%,29.41%, respectively. The content of glycyrrhizic acid, liquiritin, isoliquiritin, liquiritigenin, liquiritin apioside, isoliquiritin apioside were increased 20.16%,45.31%,53.56%,27.66%,23.54%,8.46% with the 0.7 mg•L⁻¹ BR stimulating 2 months later. The best effects were achieved in 2 months after brassinolide stimulating. The conclusions prove that morphological characters and the main chemical constituents accumulation of G. uralensis could be effected by exogenous BR stimulation in certain case.

An ultrahigh-performance liquid chromatography method with electrospray ionization tandem mass spectrometry for simultaneous quantification of five phytohormones in medicinal plant Glycyrrhiza uralensis under abscisic acid stress.

An efficient simplified method was developed to determine multiple classes of phytohormones simultaneously in the medicinal plant Glycyrrhiza uralensis. Ultrahigh-performance liquid chromatography electrospray ionization tandem mass spectrometry (UPLC/ESI-MS/MS) with multiple reaction monitoring (MRM) in negative mode was used for quantification. The five studied phytohormones are gibberellic acid (GA3), abscisic acid (ABA), jasmonic acid (JA), indole-3-acetic acid, and salicylic acid (SA). Only 100 mg of fresh leaves was needed, with one purification step based on C18 solid-phase extraction. Cinnamic acid was chosen as the internal standard instead of isotope-labeled internal standards. Under the optimized conditions, the five phytohormones with internal standard were separated within 4 min, with good linearities and high sensitivity. The validated method was applied to monitor the spatial and temporal changes of the five phytohormones in G. uralensis under ABA stress. The levels of GA3, ABA, JA, and SA in leaves of G. uralensis were increased at different times and with different tendencies in the reported stress mode. These changes in phytohormone levels are discussed in the context of a possible feedback regulation mechanism. Understanding this mechanism will provide a good chance of revealing the mutual interplay between different biosynthetic routes, which could further help elucidate the mechanisms of effective composition accumulation in medicinal plants.

[A new herbs traceability method based on DNA barcoding-origin-morphology analysis--an example from an adulterant of 'Heiguogouqi'].

The fruit of Lycium ruthenicum is a common folk medicine in China. Now it is popular for its antioxidative effect and other medical functions. The adulterants of the herb confuse consumers. In order to identify a new adulterant of L. ruthenicum, a research was performed based on NCBI Nucleotide Database ITS Sequence, combined analysis of the origin and morphology of the adulterant to traceable varieties. Total genomic DNA was isolated from the materials, and nuclear DNA ITS sequences were amplified and sequenced; DNA fragments were collated and matched by using ContingExpress. Similarity identification of BLAST analysis was performed. Besides, the distribution of plant origin and morphology were considered to further identification and verification. Families and genera were identified by molecular identification method. The adulterant was identified as plant belonging to Berberis. Origin analysis narrowed the range of sample identification. Seven different kinds of plants in Berberis were potential sources of the sample. Adulterants variety was traced by morphological analysis. The united molecular identification-origin-morphology research proves to be a preceding way to medical herbs traceability with time-saving and economic advantages and the results showed the new adulterant of L. ruthenicum was B. kaschgarica. The main differences between B. kaschgarica and L. ruthenicum are as follows: in terms of the traits, the surface of B. kaschgarica is smooth and crispy, and that of L. ruthenicum is shrinkage, solid and hard. In microscopic characteristics, epicarp cells of B. aschgarica thickening like a string of beads, stone cells as the rectangle, and the stone cell walls of L. ruthenicum is wavy, obvious grain layer. In molecular sequences, the length of ITS sequence of B. kaschgarica is 606 bp, L. ruthenicum is 654 bp, the similarity of the two sequences is 53.32%.