Development of a pre-column derivatization ultra-high-performance liquid chromatography method for polysaccharide and monosaccharide quantification in Lycium barbarum.

Given the limited specificity and accuracy observed in the current official colorimetric quantification of polysaccharide in Lycium barbarum, our study aims to establish a novel, specific, accurate, and economic pre-column derivatization ultra-high-performance liquid chromatography (UHPLC) method for determining the monosaccharide and polysaccharide content in L. barbarum. The optimization of extraction, hydrolysis, and derivatization (using 1-phenyl-3-methyl-5-pyrazolone) processes for polysaccharide from L. barbarum was conducted initially, followed by separation of nine monosaccharides within 20 min using UHPLC with a C18 column. Subsequently, a novel method known as quantitative analysis of multiple components by single marker was developed, utilizing either additive 2-deoxy-D-ribose or any monosaccharide present in the sample as a single reference standard to simultaneously detect the contents of polysaccharide and nine monosaccharides in L. barbarum. To validate the accuracy of the established method, the quantitative results of our approach were compared to both external and internal standard method methods. The minimal relative errors in the quantitative determination of monosaccharides among the three methods confirmed the dependability of the method. By analyzing 20 batches of L. barbarum samples, D-galacturonic acid exhibited the highest content and the polysaccharide levels ranged from 3.02 to 13.04 mg/g. All data implied the specificity and accuracy of the method.

Integrating untargeted and targeted LC-MS-based metabolomics to identify the serum metabolite biomarkers for tuberculosis.

Given the limitations of untargeted metabolomics in precise metabolite quantification, our current research employed a novel approach by integrating untargeted and targeted metabolomics utilizing ultra-high-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UHPLC-QTOF-MS/MS) to analyze the metabolic profile and potential biomarkers for tuberculosis (TB). A cohort of 36 TB patients and 36 healthy controls (HC) was enlisted to obtain serum samples. Multivariate pattern recognition and univariate statistical analysis were employed to screen and elucidate the differential metabolites, whereas dot plots and receiver operating characteristic (ROC) curves were established for the identification of potential biomarkers of TB. The results indicated a distinct differentiation between the two groups, identifying 99 metabolites associated with five primary metabolic pathways in relation to TB. Of these, 19 metabolites exhibited high levels of sensitivity and specificity, as evidenced by the area under curve values approaching 1. Following targeted quantitative analysis, three potential metabolites, namely, L-asparagine, L-glutamic acid, and arachidonic acid, were demonstrated excellent discriminatory ability as evidenced by the results of the ROC curve, dot plots, and random forest model. Particularly noteworthy was the enhanced diagnostic efficacy of the combination of these three metabolites compared to singular biomarkers, suggesting their potential utility as serum biomarkers for TB diagnosis.

Serum metabolomics reveals the metabolic profile and potential biomarkers of ankylosing spondylitis.

Ankylosing spondylitis (AS) is a chronic systemic inflammatory disease that significantly impairs physical function in young individuals. However, the identification of radiographic changes in AS is frequently delayed, and the diagnostic efficacy of biomarkers like HLA-B27 remains moderately effective, with unsatisfactory sensitivity and specificity. In contrast to existing literature, our current experiment utilized a larger sample size and employed both untargeted and targeted UHPLC-QTOF-MS/MS based metabolomics to identify the metabolite profile and potential biomarkers of AS. The results indicated a notable divergence between the two groups, and a total of 170 different metabolites were identified, which were associated with the 6 primary metabolic pathways exhibiting a correlation with AS. Among these, 26 metabolites exhibited high sensitivity and specificity with area under curve (AUC) values greater than 0.8. Subsequent targeted quantitative analysis discovered 3 metabolites, namely 3-amino-2-piperidone, hypoxanthine and octadecylamine, exhibiting excellent distinguishing ability based on the results of the ROC curve and the Random Forest model, thus qualifying as potential biomarkers for AS. Summarily, our untargeted and targeted metabolomics investigation offers novel and precise insights into potential biomarkers for AS, potentially enhancing diagnostic capabilities and furthering the comprehension of the condition's pathophysiology.

A strategy for inhibitors screening of xanthine oxidase based on colorimetric sensor combined with affinity chromatography technology.

The discovery of enzyme inhibitors from natural products is a crucial aspect in the development of therapeutic drugs. However, the complexity of natural products presents a challenge in developing simple and efficient methods for inhibitor screening. Herein, we have developed an integrated analytical model for screening xanthine oxidase (XOD) inhibitors that combines simplicity, accuracy, and efficiency. This model utilizes a colorimetric sensor and affinity chromatography technology with immobilized XOD. The colorimetric sensor procedure can quickly identify whether there are active components in complex samples. Subsequently, the active components in the samples identified by the colorimetric sensor procedure were further captured, separated, and identified through affinity chromatography. The integrated analytical model can significantly enhance the efficiency and accuracy of inhibitor screening. The proposed method was applied to screen for an activity inhibitor of XOD in five natural medicines. As a result, a potential active ingredient for XOD, polydatin, was successfully identified from Polygoni Cuspidati Rhizoma et Radix. This work is anticipated to offer new insights for the screening of enzyme inhibitors from natural medicines.

Establishment and application of a screening method for α-glucosidase inhibitors based on dual sensing and affinity chromatography.

α-Glucosidase plays a direct role in the metabolic pathways of starch and glycogen, any dysfunction in its activity could result in metabolic disease. Concurrently, this enzyme serves as a target for diverse drugs and inhibitors, contributing to the regulation of glucose metabolism in the human body. Here, an integrated analytical method was established to screen inhibitors of α-glucosidase. This step-by-step screening model was accomplished through the biosensing and affinity chromatography techniques. The newly proposed sensing program had a good linear relationship within the enzyme activity range of 0.25 U mL-1 to 1.25 U mL-1, which can quickly identify active ingredients in complex samples. Then the potential active ingredients can be captured, separated, and identified by an affinity chromatography model. The combination of the two parts was achieved by an immobilized enzyme technology and a microdevice for reaction, and the combination not only ensured efficiency and accuracy for inhibitor screening but also eliminated the occurrence of false positive results in the past. The emodin, with a notable inhibitory effect on α-glucosidase, was successfully screened from five traditional Chinese medicines using this method. The molecular docking results also demonstrated that emodin was well embedded into the active pocket of α-glucosidase. In summary, the strategy provided an efficient method for developing new enzyme inhibitors from natural products.

A comparative UHPLC-QTOF-MS/MS-based metabolomics approach reveals the metabolite profiling of wolfberry sourced from different geographical origins.

Wolfberry, known as Goji berry, is the fruit of Lycium barbarum L. (LB). As a famous functional food and TCM, the cost and efficacy of LB are closely linked to its geographical origin. The present study aimed to establish an effective method for distinguishing LB from different geographical origins. By employing UHPLC-QTOF-MS/MS combined with multivariate analysis, the metabolite profiling of LB (199 batches) obtained from Ningxia, Gansu, Qinghai, and Xinjiang, was evaluated. The results demonstrated that the method effectively distinguished LB from the four regions, with a total of 148 different metabolites being detected. Subsequent assessment using heat maps, Venn analysis, receiver operating characteristics curves and dot plots revealed 21 of these metabolites exhibited exceptional sensitivity and specificity, with under-curve values approaching 1, thus indicating their potential as biomarkers for LB. These findings strongly support the suitability of UHPLC-QTOF-MS/MS-based metabolomics as an effective approach to identify the source of LB.

Exploring the therapeutic potential of natural compounds modulating the endocannabinoid system in various diseases and disorders: review.

Cannabinoid receptors, endogenous cannabinoids (endocannabinoids), and the enzymes involved in the biosynthesis and degradation of the endocannabinoids make up the endocannabinoid system (ECS). The components of the ECS are proven to modulate a vast bulk of various physiological and pathological processes due to their abundance throughout the human body. Such discoveries have attracted the researchers' attention and emerged as a potential therapeutical target for the treatment of various diseases. In the present article, we reviewed the discoveries of natural compounds, herbs, herbs formula, and their therapeutic properties in various diseases and disorders by modulating the ECS. We also summarize the molecular mechanisms through which these compounds elicit their properties by interacting with the ECS based on the existing findings. Our study provides the insight into the use of natural compounds that modulate ECS in various diseases and disorders, which in turn may facilitate future studies exploiting natural lead compounds as novel frameworks for designing more effective and safer therapeutics.

Rubus chingii Hu relieved the polycystic ovary syndrome with enhanced insulin sensitivity through inhibiting TXNIP/NLRP3 inflammasome signaling.

BACKGROUND: Polycystic ovary syndrome (PCOS) is one of the most prevalent endocrine disorders in gynecology with severe metabolic abnormalities. Therefore, identifying effective treatments and drugs for PCOS is important. We aimed to investigate effect of the traditional Chinese medicine (TCM) Rubus chingii Hu (R. chingii) on ovarian function and insulin resistance (IR) of PCOS rat models, and to explore the underlying mechanisms. METHODS: A PCOS rat model was established by subcutaneous injection of dehydroepiandrosterone (DHEA) solution for 20 days. PCOS rats were randomly divided into a control group (CON), model group (MOD), metformin group (MET), TCM R. chingii group (RCG), and RCG + Ad-TXNIP groups. After 28 days of treatment, the samples were collected for subsequent experiments. RESULTS: R. chingii treatment alleviated hormone imbalance and IR while improving ovarian pathology in the PCOS model. R. chingi inhibited the activation of the thioredoxin-interacting protein (TXNIP)/NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome in the ovarian tissue of PCOS rats. Furthermore, TXNIP overexpression hindered the protective effect of R. chingii intervention in PCOS rats, as evidenced by the increase of homeostasis model assessment of insulin resistance (HOMA-IR), luteinizing hormone (LH), testosterone (T), C-reactive protein (CRP) levels, and atretic follicles. CONCLUSION: R. chingii intervention improved ovarian polycystic development by suppressing the TXNIP/NLRP3 inflammasome, which may be an effective treatment for PCOS.

Bloodletting Acupuncture at Jing-Well Points on Hand Induced Autophagy to Alleviate Brain Injury in Acute Altitude Hypoxic Rats by Activating PINK1/Parkin Pathway.

OBJECTIVE: To explore the protective effect of bloodletting acupuncture at twelve Jing-well points on hand (BAJP) on acute hypobaric hypoxia (AHH)-induced brain injury in rats and its possible mechanisms. METHODS: Seventy-five Sprague Dawley rats were divided into 5 groups by a random number table (n=15), including control, model, BAJP, BAJP+3-methyladenine (3-MA), and bloodletting acupuncture at non-acupoint (BANA, tail tip blooding) groups. After 7-day pre-treatment, AHH models were established using hypobaric oxygen chambers. The levels of S100B, glial fibrillary acidic protein (GFAP), superoxide dismutase (SOD), and malondialdehyde (MDA) in serum were measured by enzyme-linked immunosorbent assay. Hematoxylin-eosin staining and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method were used to assess hippocampal histopathology and apoptosis. Transmission electron microscopy assay was used to observe mitochondrial damage and autophagosomes in hippocampal tissues. Flow cytometry was used to detect mitochondrial membrane potential (MMP). The mitochondrial respiratory chain complexes I, III and IV activities and ATPase in hippocampal tissue were evaluated, respectively. Western blot analysis was used to detect the protein expressions of Beclin1, autophagy protein 5 (ATG5), microtubule-associated protein 1 light chain 3 beta (LC3B), phosphatase and tensin homolog induced kinase 1 (PINK1), and Parkin in hippocampal tissues. The mRNA expressions of Beclin1, ATG5 and LC3-II were analyzed by quantitative real-time polymerase chain reaction. RESULTS: BAJP treatment reduced hippocampal tissue injury and inhibited hippocampal cell apoptosis in AHH rats. BAJP reduced oxidative stress by decreasing S100B, GFAP and MDA levels and increasing SOD level in the serum of AHH rats (P<0.05 or P<0.01). Then, BAJP increased MMP, the mitochondrial respiratory chain complexes I, III and IV activities, and the mitochondrial ATPase activity in AHH rats (all P<0.01). BAJP improved mitochondrial swelling and increased the autophagosome number in hippocampal tissue of AHH rats. Moreover, BAJP treatment increased the protein and mRNA expressions of Beclin1 and ATG5 and LC3-II/LC3-I ratio in AHH rats (all P<0.01) and activated the PINK1/Parkin pathway (P<0.01). Finally, 3-MA attenuated the therapeutic effect of BAJP on AHH rats (P<0.05 or P<0.01). CONCLUSION: BAJP was an effective treatment for AHH-induced brain injury, and the mechanism might be through reducing hippocampal tissue injury via increasing the PINK1/Parkin pathway and enhancement of mitochondrial autophagy.

L-Arginine Supplementation Improves Vascular Endothelial Dysfunction Induced by High-Fat Diet in Rats Exposed to Hypoxia.

INTRODUCTION: Our previous study showed that high-fat diet inhibited the increase in nitric oxide and endothelial nitric oxide synthase expression in the aortic endothelium of rats exposed to hypoxia, and hypoxia plus a high-fat diet led to earlier and more severe vascular endothelial dysfunction (VED) than hypoxia alone. The purpose of the present study was to investigate the effects of L-arginine on high-fat diet-induced VED of rats in hypoxia. METHODS: Forty male Sprague-Dawley rats were randomly divided into 4 groups and treated with hypoxia (H group), hypoxia plus high-fat diet (H+HFD group), hypoxia plus L-arginine (H+L-Arg group), and hypoxia plus high-fat diet and L-arginine (H+HFD+L-Arg group) for 1 wk. Hypoxia was simulated in a hypobaric chamber with an altitude of 5000 m. Aortic morphology and endothelium-dependent vasorelaxation were used to assess VED. RESULTS: High-fat diet impaired vascular remodeling and reduced endothelium-dependent vasodilator response to acetylcholine in rats exposed to hypoxia, secondary to dysregulation of the nitric oxide pathway. L-arginine supplementation significantly increased plasma nitrates and nitrites and endothelial nitric oxide synthase mRNA levels and improved ultrastructural changes in aortic endothelium and endothelium-dependent vasodilator response. CONCLUSIONS: L-arginine prevents aortic ultrastructural changes and reverses VED induced by high-fat diet in rats exposed to hypoxia, which may have implications for VED induced by high-fat diet in high altitude dwellers.