Multi­omics­based analysis of the regulatory mechanism of gypenosides on bile acids in hypercholesterolemic mice.

Gynostemma pentaphyllum is a traditional medicine used by ethnic minorities in southwest China and gypenosides are currently recognized as essential components of the pharmacological substances of Gynostemma pentaphyllum, which are effective in regulating metabolic syndrome, especially in improving hepatic metabolic disorders. The present study randomly divided C57BL/6J male mice into the normal diet control group (ND), high-fat diet modeling group (HFD) and gypenosides group (GP). Liquid chromatography-mass spectrometry (UPLC-MS) was applied to quantify bile acids in the liver, bile and serum of mice in ND, HFD and GP groups. Liver proteins were extracted for trypsin hydrolysis and analyzed quantitatively using UPLC-MS + MS/MS (timsTOF Pro 2). Total mouse liver RNA was extracted from ND, HFD and GP groups respectively, cDNA sequencing libraries constructed and sequenced using BGISEQ-500 sequencing platform. The expression of key genes Fxr, Shp, Cyp7a1, Cyp8b1, and Abab11 was detected by RT-qPCR. The results showed that gypenosides accelerated free bile acid synthesis by promoting the expression of bile acid synthase CYP7A1 and CYP8B1 genes and proteins and accelerating the secretion of conjugated bile acids from the liver to the bile ducts. GP inhibited the bile acid transporters solute carrier organic anion transporter family member (SLCO) 1A1 and SLCO1A4, reducing the reabsorption of free bile acids and accelerating the excretion of free bile acids from the blood to the kidneys. It also promoted the metabolic enzyme CYP3A11, which accelerated the metabolism and clearance of bile acids, thus maintaining the balance of the bile acid internal environment.

Widely Targeted Metabolomics Analysis of Dendrobium officinale at Different Altitudes.

In order to investigate the differences in the chemical composition of Dendrobium officinale at different altitudes, we collected Dendrobium officinale from different altitudes in Guizhou Province, China, and firstly determined its polysaccharide content by using a sulfuric acid-phenol color development method with reference to the Chinese Pharmacopoeia, and further determined its metabolites by using widely targeted metabolomics, and explored the differences in the chemical composition of Dendrobium officinale at different altitudes using multivariate statistical analysis. We found that the polysaccharide content was higher in plants growing at 1122 m, a total of 902 secondary metabolites were detected by wildly targeted metabolomics, and amino acids and their derivatives were more highly expressed at 1122 m, while other metabolites were more highly expressed at 835 m. Furthermore, we found that the phenolic acid compound nerugein was only present in plants at 835 m, and two lipid compounds, namely Lyso PE 20 : 4 and its isomer, were only present at 1122 m. Taken together, these results may provide a basis for the selection and clinical application of D. officinale cultivated at different altitudes.

An UPLC-Q-TOF/MS-Based Analysis of the Differential Composition of Dendrobium officinale in Different Regions.

Dendrobium officinale (D. officinale) is a valuable traditional Chinese herbal medicine with high commercial value. In Chinese Pharmacopoeia (Ch.P., 2020 edition), the quality of D. officinale is mainly evaluated by its polysaccharide content. However, varying growth and production conditions, such as cultivation environment, origin, harvesting process, or processing methods, resulting in highly variable yields, quality, and composition. The aim of this study was to investigate whether the content of secondary metabolites in D. officinale from different origins is consistent with the polysaccharide content. The results showed that the polysaccharide content and pass rate were ranked as GX > AH > GZ > YN. Based on the nontargeted metabolomics approach, we searched for differential components in 22 different regions of D. officinale, including amides, bibenzyls, disaccharide, flavonoids, organic nitrogenous compounds, and phenolic glycosides. The overall expression was opposite to the polysaccharide, and the most expressed was YN, followed by GZ, AH, and GX. These results indicated that the current quality standard for evaluating the quality of D. officinale by polysaccharide content alone is imperfect, and small molecule compounds need to be included as quality markers.

Determination of Scopolamine Distribution in Plasma and Brain by LC-MS/MS in Rats.

Scopolamine, as a tropane alkaloid found in plants such as belladonna and datura, is used clinically as a transdermal patch and is highly neurotoxic. This study aimed to develop a simple, sensitive, and selective LC-MS/MS method for the determination of the content and distribution of scopolamine in rat plasma and brain after drug administration. In our study, sample pretreatment consisted of protein precipitation with acetonitrile followed by nitrogen blow concentration. Gradient elution of scopolamine and internal standard was performed on a ZORBAX Eclipse Plus C18 (2.1∗100 mm, 3.5 µm) column with water containing 0.1% formic acid (v/v) and acetonitrile as a mobile phase. Those samples were quantified in ESI positive ion mode using an API 4000 triple quadrupole mass spectrometer. The results showed that scopolamine was linear in the calibration range of 2-2500 ng/mL, and the selectivity, accuracy, precision, matrix effect, stability, and recovery of the method were within acceptable limits. The method has been validated and has been successfully used for toxicokinetic studies of scopolamine. After intraperitoneal injection, the time to peak toxic concentrations of scopolamine in rats was 0.5 h. The concentrations of scopolamine in the hippocampus and cortex were much higher than those in the striatum, indicating that the likely targets of its neurotoxic damage were the hippocampus and cortex. Overall, this study provides the basis for the neurotoxicity of scopolamine and provides a reference for its toxicokinetic studies.

Impact of Clarified Apple Juices with Different Processing Methods on Gut Microbiota and Metabolomics of Rats.

The consumption of processed foods has increased compared to that of fresh foods in recent years, especially due to the coronavirus disease 2019 pandemic. Here, we evaluated the health effects of clarified apple juices (CAJs, devoid of pectin and additives) processed to different degrees, including not-from-concentrate (NFC) and from-concentrate (FC) CAJs. A 56-day experiment including a juice-switch after 28 days was designed. An integrated analysis of 16S rRNA sequencing and untargeted metabolomics of cecal content were performed. In addition, differences in the CAJs tested with respect to nutritional indices and composition of small-molecule compounds were analyzed. The NFC CAJ, which showed a higher phenolic content resulting from the lower processing degree, could improve microbiota diversity and influence its structure. It also reduced bile acid and bilirubin contents, as well as inhibited the microbial metabolism of tryptophan in the gut. However, we found that these effects diminished with time by performing experiment extension and undertaking juice-switching. Our study provides evidence regarding the health effects of processed foods that can potentially be applied to public health policy decision making. We believe that NFC juices with a lower processing degree could potentially be healthier than FC juice.

Opposite trends of glycosides and alkaloids in Dendrobium nobile of different age based on UPLC-Q/TOF-MS combined with multivariate statistical analyses.

INTRODUCTION: Alkaloids and glycosides are the active ingredients of the herb Dendrobium nobile, which is used in traditional Chinese medicine. The pharmacological effects of alkaloids include neuroprotective effects and regulatory effects on glucose and lipid metabolism, while glycosides improve the immune system. The pharmacological activities of the above chemical components are significantly different. In practice, the stems of 3-year-old D. nobile are usually used as the main source of Dendrobii Caulis. However, it has not been reported whether this harvesting time is appropriate. OBJECTIVE: The aim of this study was to compare the chemical characteristics of D. nobile in different growth years (1-3 years). METHODS: In this study, ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC-Q/TOF-MS) was employed to analyze the constituents of D. nobile. The relative abundance of each constituent was analyzed with multivariate statistical analyses to screen the characteristic constituents that contributed to the characterization and classification of D. nobile. Dendrobine, a component of D. nobile that is used for quality control according to the Chinese Pharmacopoeia, was assayed by gas chromatography. RESULTS: As a result, 34 characteristic constituents (VIP > 2) were identified or tentatively identified as alkaloids and glycosides based on MS/MS data. Moreover, the content of alkaloids decreased over time, whereas the content of glycosides showed the opposite trend. The absolute quantification of dendrobine was consistent with the metabolomics results. CONCLUSION: Our findings provide valuable information to optimize the harvest period and a reference for the clinical application of D. nobile.

CYP3A1 metabolism-based neurotoxicity of strychnine in rat.

Strychnine is one of the main bioactive and toxic constituents of Semen Strychni. In the present study, the neurotoxic effects of strychnine, and the role of individual differences in metabolism on susceptibility to neurotoxicity of strychnine were investigated. The acute toxicity was observed by a single dose of strychnine (2.92 mg/kg, i.g.) in rats, the epileptic stages of rats were scored according to Racine's scale. The neurotoxicity of strychnine was evaluated by the levels of ROS, MDA, SOD and GSH in hippocampus, striatum, and cortex tissues measurements and histopathological analysis. The concentrations of strychnine in the plasma, hippocampus, striatum, and cortex tissues were determined using high performance liquid chromatography tandem mass spectrometry (LC-MS/MS). The expressions of the cytochrome P450, which is the most critical protein family involved in drugs metabolism, were detected by proteomics. The mechanism of susceptibility to neurotoxicity of strychnine was elucidated by correlation analysis among above indicators. The results indicated that striatum and cortex were the main toxic targets of strychnine, and the CYP3A1 might be a susceptible biomarker to neurotoxicity of strychnine. These results provide valuable insights into the neurotoxic susceptibility of strychnine that will aid in the rational clinical use of strychnine (possibly including Semen Strychni).

Application of single-cell RNA sequencing technology in liver diseases: a narrative review.

OBJECTIVE: This review aimed to summarize the application of single-cell transcriptome sequencing technology in liver diseases. BACKGROUND: The increasing application of single-cell ribonucleic acid (RNA) sequencing (scRNA-seq) in life science and biomedical research has greatly improved our understanding of cellular heterogeneity in immunology, oncology, and developmental biology. scRNA-seq has proven to be a powerful tool for identifying and classifying cell subsets, characterizing rare or small cell subsets and tracking cell differentiation along the dynamic cell stages. Globally, liver disease has high rates of morbidity and mortality, and its exact pathological mechanism remains unclear, current treatment options are limited to clearance of the underlying cause or liver transplantation, which cannot overwhelm and cure liver diseases. scRNA-seq provides many novel insights for healthy and diseased livers. METHODS: In this review, we searched for related articles in the PubMed database and summarized the advances of scRNA-seq in revealing the molecular mechanisms of liver development, regeneration, and disease. We also discussed the challenges and future application potential of scRNA-seq, which is expected to enhance the ability to explore the field of liver research and accelerate the clinical application of liver precision medicine. CONCLUSIONS: With the continuous improvement of scRNA-seq technology, scRNA-seq is expected to unlock new avenues for liver biology exploration, liver disease diagnosis, and personalized treatment, which will pave the way for breakthrough innovation in personalized medicine.

Identification of Pyrrolizidine Alkaloids in Senecio Plants by Liquid Chromatography-Mass Spectrometry.

Pyrrolizidine alkaloids (PAs) are considered as the major constituents that cause hepatoxicity in Senecio plants. PAs can be found in about 3%-5% of the world's flowering plants. Nowadays, the identification method of PAs by separation and preparation was too slow and lacked effective power. A rapid method to identify PAs in plants must be developed. Based on the fragmentation regularity, the hepatoxic PAs and nonhepatoxic PAs were characterized by liquid chromatography-mass spectrometry (LC-MS). The detailed structures of PAs in five Senecio plants were identified based on tandem mass spectrometry (MS/MS) spectrum and chemical research information. In the present study, some new fragmentation regularities of PAs have been found, such as product ions at m/z 122, m/z 140 and m/z 124, m/z 142, which have been discovered as the characteristic fragments of lactone and mono-esterase type of saturated PAs, respectively. Moreover, two product ions at m/z 120 and m/z 138 have been reported as the characteristic fragments of unsaturated PAs. Some of them were found in Senecio species for the first time, and some of them may be new nature product or even new compound. Finally, we classified these plants into five categories based on PAs which were identified in the present study; the result corresponded with the classification by morphology. In addition, we have found some constituents that have odd molecular weight number only in Senecio species but not in Ligularia species; the detailed structures of these non-PAs constituents need penetrating study. LC-MS was rapid and sensitive method for detecting and identifying PAs in plants. Pyrrolizidine alkaloids were the toxiferous constituent of Senecio plants. In this study, we found that PAs can be used as the characteristic constituent of Senecio species.

Identification and Simultaneous Determination of the Main Toxical Pyrrolizidine Alkaloids in a Compound Prescription of Traditional Chinese Medicine: Qianbai Biyan Tablet.

Qianbai biyan tablet (QT) is a compound prescription of traditional Chinese medicine which is used to treat nasal congestion, rhinitis, and nasosinusitis, with Senecio scandens as its main plant material. Several pyrrolizidine alkaloids (PAs) were reported in Senecio scandens and others of Senecio species. Although Senecio scandens is assigned as the legal plant material of QT, whether replaced use of it by other Senecio plants can bring toxicity is unknown because of the lack of quantitative data about toxic PAs between different Senecio species. In the present study, adonifoline, senkirkine, and another PA presumed as emiline have been identified in QT; however, there was no senecionine detected in all tablets. PA contents in QTs varied in different companies and different batches. Adonifoline existed only in Senecio scandens, and senecionine was detected in all eight Senecio plants investigated in the present study. Data showed that replaced use of Senecio scandens with a low level of senecionine by other Senecio plants such as Senecio vulgaris containing a high level of senecionine is advertised to be forbidden. Data of the present study may be used as a reference to make new drug quality regularity and recommendation guideline for the safety of QT.