Riboflavin modified carbon cloth enhances anaerobic digestion treating food waste in a pilot-scale system.

Previous laboratory-scale studies have consistently shown that carbon-based conductive materials can notably improve the anaerobic digestion of food waste, typically employing reactors with regular capacity of 1-20 L. Furthermore, incorporating riboflavin-loaded conductive materials can further address the imbalance between fermentation and methanogenesis in anaerobic systems. However, there have been few reports on pilot-scale investigation. In this study, a 10 m2 of riboflavin modified carbon cloth was incorporated into a pilot-scale (2 m3) food waste anaerobic reactor to improve its treatment efficiency. The study found that the addition of riboflavin-loaded carbon cloth can increase the maximum organic loading rate (OLR) by 40% of the pilot-scale reactor, compared to the system using carbon cloth without riboflavin loading, while ensuring efficient operation of the reaction system, effectively alleviating system acidification, sustaining methanogen activity, and increasing daily methane production by 25%. Analysis of the microbial community structure revealed that riboflavin-loaded carbon cloth enriched the methanogenic archaea in the genera of Methanothrix and Methanobacterium, which are capable of extracellular direct interspecies electron transfer (DIET). And metabolic pathway analysis identified the methane production pathway, highly enriched on the reduction of acetic acid and CO2 at riboflavin-loaded carbon cloth sample. The expression levels of genes related to methane production via DIET pathway were also significantly upregulated. These results can provide important guidance for the practical application of food waste anaerobic digestion engineering.

Exploration on pharmacological mechanisms of YZP against neuropathic pain via inhibiting spinal inflammation and the rationality of its compatibility.

ETHNOPHARMACOLOGICAL RELEVANCE: Yuanhu Zhitong Prescription (YZP) is a well-known traditional Chinese medicine (TCM) formula for neuropathic pain (NP) therapy with a satisfying clinical efficacy. However, the underlying pharmacological mechanism and its compatibility principle remain unclear. AIM OF THE STUDY: This study aims to investigate the analgesic and compatibility mechanisms of YZP on neuropathic pain (NP) at the gene and biological process levels. MATERIALS AND METHODS: The chronic constriction injury (CCI) rats were intragastrically administrated with extracts of YZP, YH and BZ separately, and then mechanical hypersensitivity were measured to evaluate the analgesic effects between YH and BZ before and after compatibility. Then, RNA-seq and bioinformatics analyses were performed to elucidate the potential mechanisms underlying YZP's analgesia and compatibility. Finally, the expression levels and significant differences of key genes were analyzed. RESULTS: Behaviorally, both YZP and YH effectively alleviated mechanical allodynia in CCI rats, with YZP being superior to YH. In contrast, we did not observe an analgesic effect of BZ. Genetically, YZP, YH, and BZ reversed the expression levels of 52, 34, and 42 aberrant genes in the spinal cord of CCI rats, respectively. Mechanically, YZP was revealed to alleviate NP mainly by modulating the inflammatory response and neuropeptide signaling pathway, which are the dominant effective processes of YH. Interestingly, the effective targets of YZP were especially enriched in leukocyte activation and cytokine-mediated signaling pathways. Moreover, BZ was found to exert an adjunctive effect in enhancing the analgesic effect of YH by promoting skeletal muscle tissue regeneration and modulating calcium ion transport. CONCLUSIONS: YH, as the monarch drug, plays a dominant role in the analgesic effect of YZP that effectively relieves NP by inhibiting the spinal inflammation and neuropeptide signaling pathway. BZ, as the minister drug, not only synergistically enhances analgesic processes of YH but also helps to alleviate the accompanying symptoms of NP. Consequently, YZP exerted a more potent analgesic effect than YH and BZ alone. In conclusion, our findings offer new insights into understanding the pharmacological mechanism and compatibility principle of YZP, which may support its clinical application in NP therapy.

Design of a reinforcement learning-based intelligent car transfer planning system for parking lots.

In this study, a car transfer planning system for parking lots was designed based on reinforcement learning. The car transfer planning system for parking lots is an intelligent parking management system that is designed by using reinforcement learning techniques. The system features autonomous decision-making, intelligent path planning and efficient resource utilization. And the problem is solved by constructing a Markov decision process and using a dynamic planning-based reinforcement learning algorithm. The system has the advantage of looking to the future and using reinforcement learning to maximize its expected returns. And this is in contrast to manual transfer planning which relies on traditional thinking. In the context of this paper on parking lots, the states of the two locations form a finite set. The system ultimately seeks to find a strategy that is beneficial to the long-term development of the operation. It aims to prioritize strategies that have positive impacts in the future, rather than those that are focused solely on short-term benefits. To evaluate strategies, as its basis the system relies on the expected return of a state from now to the future. This approach allows for a more comprehensive assessment of the potential outcomes and ensures the selection of strategies that align with long-term goals. Experimental results show that the system has high performance and robustness in the area of car transfer planning for parking lots. By using reinforcement learning techniques, parking lot management systems can make autonomous decisions and plan optimal paths to achieve efficient resource utilization and reduce parking time.

Risk and protective factors of suicidal tendencies among freshmen in China revealed by a hierarchical regression model.

This study aimed to identify risk and protective factors for suicidal tendencies among college students by exploring current mental health, personal experiences, family environment, and school adaptation. A total of 11,504 freshmen in China were recruited. Suicidal tendencies were assessed using the Adolescents Suicidal Tendencies Scale (ASTS), while explored risk and protective factors included mental health assessed by the Symptom Checklist-90 (SCL-90), campus adaptation using the College Student School Adaptation Scale, and Personal Situation Survey. Single-factor Logistic regression analysis, correlation analysis, and hierarchical regression analysis were used to analyze the risk and protective factors affecting suicidal tendencies. The results showed that in terms of personal experience, self-injury behavior (OR = 3.522, 95% CI [3.256, 3.811]), sexual assault experience (OR = 2.603, 95% CI [2.374, 2.855]) and lack of friendship relationship (OR = 2.249, 95% CI [2.076, 2.436]) were the most significant risk factors. Regarding family environment, parenting style (OR = 2.455, 95% CI [2.272, 2.652]), parent-child relationship (OR = 2.255, 95% CI [2.092, 2.429]) and violent conflict (OR = 2.164, 95% CI [2.015, 2.324]) were the most prominent risk factors. For protective factors, life satisfaction (OR = 0.330, 95% CI [0.304, 0.359]) and rest quality (OR = 0.415, 95% CI [0.386, 0.447]) were the most significant protective factors. In addition, Symptom Checklist-90 was positively correlated with suicidal tendencies (r = 0.541, 95% CI [0.522, 0.560], p < 0.001), while school adaptation was negatively correlated with suicidal tendencies (r = - 0.590, 95% CI [- 0.579, - 0.601], p < 0.001). After considering demographic variables, psychological symptoms, school adaptation and other risk and protective factors, the hierarchical regression model could explain 48.9% of the variance of suicidal tendencies. The study emphasizes a range of multidimensional risk and protective factors for suicidal tendencies. This enhanced understanding is crucial in aiding the design of future intervention studies targeted at improving the mental health of college students.

Effect of different hydrogen evolution rates at cathode on bioelectrochemical reduction of CO2 to acetate.

Microbial electrosynthesis (MES) offers a promising approach for converting CO2 into valuable chemicals such as acetate. However, the relative low conversion rate severely limits its practical application. This study investigated the impact of different hydrogen evolution rates on the conversion rate of CO2 to acetate in the MES system. Three potentials (-0.8 V, -0.9 V and -1.0 V) corresponding to various hydrogen evolution rates were set and analyzed, revealing an optimal hydrogen evolution rate, yielding a maximum acetate formation rate of 1410.9 mg/L and 73.5 % coulomb efficiency. The electrochemical findings revealed that an optimal hydrogen evolution rate facilitated the formation of an electroactive biofilm. The microbial community of the cathode biofilm highlighted key genera, including Clostridium and Acetobacterium, which played essential roles in electrosynthesis within the MES system. Notably, a low hydrogen evolution rate failed to provide sufficient energy for the electrochemical reduction of CO2 to acetate, while a high rate led to cathode alkalinization, impeding the reaction and causing significant energy wastage. Therefore, maintaining an appropriate hydrogen evolution rate is crucial for the development of mature electroactive biofilms and achieving optimal performance in the MES system.

Riboflavin-loaded carbon cloth aids the anaerobic digestion of cow dung by promoting direct interspecies electron transfer.

Cow dung generates globally due to increased beef and milk consumption, but its treatment efficiency remains low. Previous studies have shown that riboflavin-loaded conductive materials can improve anaerobic digestion through enhance direct interspecies electron transfer (DIET). However, its effect on the practical anaerobic digestion of cow dung remained unclear. In this study, carbon cloth loaded with riboflavin (carbon cloth-riboflavin) was added into an anaerobic digester treating cow dung. The carbon cloth-riboflavin reactor showed a better performance than other two reactors. The metagenomic analysis revealed that Methanothrix on the surface of the carbon cloth predominantly utilized the CO2 reduction for methane production, further enhanced after riboflavin addition, while Methanothrix in bulk sludge were using the acetate decarboxylation pathway. Furthermore, the carbon cloth-riboflavin enriched various major methanogenic pathways and activated a large number of enzymes associated with DIET. Riboflavin's presence altered the microbial communities and the abundance of functional genes relate to DIET, ultimately leading to a better performance of anaerobic digestion for cow dung.

Granular activated carbon enhances volatile fatty acid production in the anaerobic fermentation of garden wastes.

Garden waste, one type of lignocellulosic biomass, holds significant potential for the production of volatile fatty acids (VFAs) through anaerobic fermentation. However, the hydrolysis efficiency of garden waste is limited by the inherent recalcitrance, which further influences VFA production. Granular activated carbon (GAC) could promote hydrolysis and acidogenesis efficiency during anaerobic fermentation. This study developed a strategy to use GAC to enhance the anaerobic fermentation of garden waste without any complex pretreatments and extra enzymes. The results showed that GAC addition could improve VFA production, especially acetate, and reach the maximum total VFA yield of 191.55 mg/g VSadded, which increased by 27.35% compared to the control group. The highest VFA/sCOD value of 70.01% was attained in the GAC-amended group, whereas the control group only reached 49.35%, indicating a better hydrolysis and acidogenesis capacity attributed to the addition of GAC. Microbial community results revealed that GAC addition promoted the enrichment of Caproiciproducens and Clostridium, which are crucial for anaerobic VFA production. In addition, only the GAC-amended group showed the presence of Sphaerochaeta and Oscillibacter genera, which are associated with electron transfer processes. Metagenomics analysis indicated that GAC addition improved the abundance of glycoside hydrolases (GHs) and key functional enzymes related to hydrolysis and acidogenesis. Furthermore, the assessment of major genera influencing functional genes in both groups indicated that Sphaerochaeta, Clostridium, and Caproicibacter were the primary contributors to upregulated genes. These findings underscored the significance of employing GAC to enhance the anaerobic fermentation of garden waste, offering a promising approach for sustainable biomass conversion and VFA production.

Genome-wide identification and comprehensive analysis heat shock transcription factor (Hsf) members in asparagus (Asparagus officinalis) at the seeding stage under abiotic stresses.

Heat shock transcription factors (Hsf) are pivotal as essential transcription factors. They function as direct transcriptional activators of genes regulated by thermal stress and are closely associated with various abiotic stresses. Asparagus (Asparagus officinalis) is a vegetable of considerable economic and nutritional significance, abundant in essential vitamins, minerals, and dietary fiber. Nevertheless, asparagus is sensitive to environmental stresses, and specific abiotic stresses harm its yield and quality. In this context, Hsf members have been discerned through the reference genome, and a comprehensive analysis encompassing physical and chemical attributes, evolutionary aspects, motifs, gene structure, cis-acting elements, collinearity, and expression patterns under abiotic stresses has been conducted. The findings identified 18 members, categorized into five distinct subgroups. Members within each subgroup exhibited analogous motifs, gene structures, and cis-acting elements. Collinearity analysis unveiled a noteworthy pattern, revealing that Hsf members within asparagus shared one, two, and three pairs with counterparts in Arabidopsis, Oryza sativa, and Glycine max, respectively.Furthermore, members displayed tissue-specific expression during the seedling stage, with roots emerging as viable target tissue. Notably, the expression levels of certain members underwent modification under the influence of abiotic stresses. This study establishes a foundational framework for understanding Hsf members and offers valuable insights into the potential application of molecular breeding in the context of asparagus cultivation.

Levo-tetrahydropalmatine ameliorates neuropathic pain by inhibiting the activation of the Clec7a-MAPK/NF-κB-NLRP3 inflammasome axis.

BACKGROUND: Because of the complex pathogenesis of neuropathic pain (NP), the therapeutic efficacy of existing drugs is not satisfactory. Accumulating studies have indicated that neuroinflammation may play a key role in NP onset and progression. Levo-tetrahydropalmatine (l-THP) has been extensively used for relieving chronic pain for decades. However, its potential mechanisms against NP have not yet been fully elucidated. PURPOSE: Exploring and elucidating the therapeutic effect and pharmacological mechanism of l-THP in treating NP. METHODS: RNA-seq and bioinformatics analyses were carried out to identify effective target profiling of I-THP in chronic constrictive injury (CCI) rats. The I-THP related hub targets and signaling pathways were obtained via bioinformatics analysis, then subjected to in-depth analyses through experiments in vivo. A gain-of-function study further confirmed the role of Clec7a in l-THP-mediated pain relief. Finally, the interaction between l-THP and Clec7a was verified through molecular docking and surface plasmon resonance (SPR). RESULTS: l-THP treatment effectively alleviated mechanical and thermal allodynia in NP model rats. Functionally, the I-THP effective targets were mainly enriched in inflammatory response-related pathways. Furthermore, Clec7a-MAPK/NF-κB-NLRP3 inflammasome axis was selected as one of the potential pathways of l-THP against NP. Mechanically, l-THP markedly reduced CCI-induced Clec7a overexpression, significantly inhibited the Clec7a-triggered phosphorylation of MAPK and NF-κB-p65, and decreased the expression of pyroptosis-related protein NLRP3 and Caspase-1-p20. The analgesic effect of l-THP on NP was partly eliminated when transfecting the overexpression vector virus pLVSO5Clec7a. Importantly, molecular docking and SPR data revealed that l-THP directly binds with the Clec7a protein. CONCLUSION: This study is the first to indicate that l-THP may exert an analgesic effect through inhibiting neuroinflammation via the Clec7a-MAPK/NF-κB-NLRP3 inflammasome axis, supporting the clinical utility of l-THP in NP therapy.

ETCM v2.0: An update with comprehensive resource and rich annotations for traditional Chinese medicine.

Existing traditional Chinese medicine (TCM)-related databases are still insufficient in data standardization, integrity and precision, and need to be updated urgently. Herein, an Encyclopedia of Traditional Chinese Medicine version 2.0 (ETCM v2.0, http://www.tcmip.cn/ETCM2/front/#/) was constructed as the latest curated database hosting 48,442 TCM formulas recorded by ancient Chinese medical books, 9872 Chinese patent drugs, 2079 Chinese medicinal materials and 38,298 ingredients. To facilitate the mechanistic research and new drug discovery, we improved the target identification method based on a two-dimensional ligand similarity search module, which provides the confirmed and/or potential targets of each ingredient, as well as their binding activities. Importantly, five TCM formulas/Chinese patent drugs/herbs/ingredients with the highest Jaccard similarity scores to the submitted drugs are offered in ETCM v2.0, which may be of significance to identify prescriptions/herbs/ingredients with similar clinical efficacy, to summarize the rules of prescription use, and to find alternative drugs for endangered Chinese medicinal materials. Moreover, ETCM v2.0 provides an enhanced JavaScript-based network visualization tool for creating, modifying and exploring multi-scale biological networks. ETCM v2.0 may be a major data warehouse for the quality marker identification of TCMs, the TCM-derived drug discovery and repurposing, and the pharmacological mechanism investigation of TCMs against various human diseases.

Identification of the susceptible genes and mechanism underlying the comorbid presence of coronary artery disease and rheumatoid arthritis: a network modularization analysis.

OBJECTIVE: The comorbidities of coronary artery disease (CAD) and rheumatoid arthritis (RA) are mutual risk factors, which lead to higher mortality, but the biological mechanisms connecting the two remain unclear. Here, we aimed to identify the risk genes for the comorbid presence of these two complex diseases using a network modularization approach, to offer insights into clinical therapy and drug development for these diseases. METHOD: The expression profile data of patients CAD with and without RA were obtained from the GEO database (GSE110008). Based on the differentially expressed genes (DEGs), weighted gene co-expression network analysis (WGCNA) was used to construct a gene network, detect co-expression modules, and explore their relation to clinical traits. The Zsummary index, gene significance (GS), and module membership (MM) were utilized to screen the important differentiated modules and hub genes. The GO and KEGG pathway enrichment analysis were applied to analyze potential mechanisms. RESULT: Based on the 278 DEGs obtained, 41 modules were identified, of which 17 and 24 modules were positively and negatively correlated with the comorbid occurrence of CAD and RA (CAD&RA), respectively. Thirteen modules with Zsummary < 2 were found to be the underlying modules, which may be related to CAD&RA. With GS ≥ 0.5 and MM ≥ 0.8, 49 hub genes were identified, such as ADO, ABCA11P, POT1, ZNF141, GPATCH8, ATF6 and MIA3, etc. The area under the curve values of the representative seven hub genes under the three models (LR, KNN, SVM) were greater than 0.88. Enrichment analysis revealed that the biological functions of the targeted modules were mainly involved in cAMP-dependent protein kinase activity, demethylase activity, regulation of calcium ion import, positive regulation of tyrosine, phosphorylation of STAT protein, and tissue migration, etc. CONCLUSION: Thirteen characteristic modules and 49 susceptibility hub genes were identified, and their corresponding molecular functions may reflect the underlying mechanism of CAD&RA, hence providing insights into the development of clinical therapies against these diseases.

Natural Product-Based Screening for Lead Compounds Targeting SARS CoV-2 Mpro.

Drugs that cure COVID-19 have been marketed; however, this disease continues to ravage the world without becoming extinct, and thus, drug discoveries are still relevant. Since Mpro has known advantages as a drug target, such as the conserved nature of the active site and the absence of homologous proteins in the body, it receives the attention of many researchers. Meanwhile, the role of traditional Chinese medicine (TCM) in the control of epidemics in China has also led to a focus on natural products, with the hope of finding some promising lead molecules through screening. In this study, we selected a commercial library of 2526 natural products from plants, animals and microorganisms with known biological activity for drug discovery, which had previously been reported for compound screening of the SARS CoV-2 S protein, but had not been tested on Mpro. This library contains compounds from a variety of Chinese herbs, including Lonicerae Japonicae Flos, Forsythiae Fructus and Scutellariae Radix, which are derived from traditional Chinese medicine prescriptions that have been shown to be effective against COVID-19. We used the conventional FRET method for the initial screening. After two rounds of selection, the remaining 86 compounds were divided into flavonoids, lipids, phenylpropanoids, phenols, quinones, alkaloids, terpenoids and steroids according to the skeleton structures, with inhibition rates greater than 70%. The top compounds in each group were selected to test the effective concentration ranges; the IC50 values were as follows: (-)-gallocatechin gallate (1.522 ± 0.126 µM), ginkgolic acid C15:1 (9.352 ± 0.531 µM), hematoxylin (1.025 ± 0.042 µM), fraxetin (2.486 ± 0.178 µM), wedelolactone (1.003 ± 0.238 µM), hydroxytyrosol acetate (3.850 ± 0.576 µM), vanitiolide (2.837 ± 0.225 µM), ß,ß-dimethylacrylalkannin (2.731 ± 0.308 µM), melanin (7.373 ± 0.368 µM) and cholesteryl sodium sulfate (2.741 ± 0.234µM). In the next step, we employed two biophysical techniques, SPR and nanoDSF, to obtain KD/Kobs values: hematoxylin (0.7 µM), (-)-gallocatechin gallate (126 µM), ginkgolic acid C15:1 (227 µM), wedelolactone (0.9770 µM), ß,ß-dimethylacrylalkannin (1.9004 µM,), cholesteryl sodium sulfate (7.5950 µM) and melanin (11.5667 µM), which allowed better assessments of the binding levels. Here, seven compounds were the winners. Then, molecular docking experiments were specially performed by AutoDock Vina to analyze the mode of interactions within Mpro and ligands. We finally formulated the present in silico study to predict pharmacokinetic parameters as well as drug-like properties, which is presumably the step that tells humans whether the compounds are drug-like or not. Moreover, hematoxylin, melanin, wedelolactone, ß,ß-dimethylacrylalkannin and cholesteryl sodium sulfate are in full compliance with the "Lipinski" principle and possess reasonable ADME/T properties, they have a greater potential of being lead compounds. The proposed five compounds are also the first to be found to have potential inhibitory effects on SARS CoV-2 Mpro. We hope that the results in this manuscript may serve as benchmarks for the above potentials.

Fabrication of a Polysaccharide-Protein/Protein Complex Stabilized Oral Nanoemulsion to Facilitate the Therapeutic Effects of 1,8-Cineole on Atherosclerosis.

Atherosclerosis (AS) is a systemic disease characterized by lipid deposition in the blood vessel wall that urgently requires effective and safe therapeutic drugs for long-term treatment. An essential oil monomer-1,8-cineole (CIN) with ameliorative effects on vascular injuries has considerable potential for preventing the progression of AS because of its antioxidant, anti-inflammation, and cholesterol regulatory effects. However, the high volatility and instability of CIN result in low oral bioavailability and a short half-life, thereby limiting its clinical application. We formulated a nanoemulsion using a polysaccharide-protein/protein complex (dextran-bovine serum albumin/protamine, DEX5k-BSA/PTM) as an emulsifier, with vitamin B12 (VB12) as the ligand to facilitate the transportation across the small intestine. An emulsion preparation method using a microjet followed by ultraviolet irradiation was developed to obtain the CIN-loaded oral nanoemulsion CIN@DEX5k-BSA/PTM/VB12. The nanoemulsion improved the stability of CIN both in vitro and in vivo, prolonged the retention time in the gastrointestinal tract (GIT), and enhanced the permeability across the mucus layer and intestinal epithelial cells to increase oral bioavailability and plaque accumulation of CIN. Validated in an AS mouse model, CIN@DEX5k-BSA/PTM/VB12 achieved prominent therapeutic efficacy combating AS. This study highlights the advantages of DEX5k-BSA/PTM and VB12 in the development of nanoemulsions for CIN and provides a promising oral nanoplatform for the delivery of essential oils.

Identification of the Main Chemical constituents and mechanism of Renshen Guben oral liquid against Renal Fibrosis.

BACKGROUND: Renal fibrosis is the late stage of many chronic kidney diseases (CKD). Clinically, there is almost no effective treatment for renal fibrosis except dialysis. Renshen Guben oral liquid (RSGB) is a Chinese patent medicine approved by National Medical Products Administration (NMPA), which is suitable for clinical patients with chronic nephritis. Currently, the chemical constituents of RSGB remains unclear, and its efficacy and mechanism on renal fibrosis have not been reported. METHODS: In our research, ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) was employed to describe the chemical profile of RSGB, unilateral ureteral obstruction (UUO) model in mice was established to evaluate the beneficial effect of RSGB on renal fibrosis by biochemical indexes, HE and Masson staining. RNA sequencing and "constituents-targets-pathways" multi-dimensional network was established to mine the mechanisms of RSGB. Key targets were verified by quantitative real-time PCR (qRT-PCR) and western bolt (WB). RESULTS: A total of 201 constituents were identified or tentatively characterized, 15 of which were confirmed with standards. The number of triterpenes was the highest with 49, followed by phenols with 46. RSGB ameliorated the blood urea nitrogen (BUN) and serum creatinine (Scr) levels in serum, normalizing pathological structure of kidney tissue. RNA sequencing revealed that RSGB regulates 226 differential genes, which were involved in kidney development. According to the "constituents-targets-pathways" network, 26 key active constituents may mainly regulate the inflammatory immune system through 88 corresponding targets. qRT-PCR and WB results showed that RSGB inhibited the activation of the Tgfß1/Smad2/3 pathway, Wnt4/ß-Catenin pathway and NGFR/NF-κB pathway. CONCLUSIONS: Overall, our study, for the first time, characterized 201 chemical constituents in RSGB, and 26 of them were screened out to alleviates renal fibrosis mainly through Tgfß1/Smad2/3 pathway, Wnt4/ß-catenin pathway and NGFR/NF-κB pathway, which may provide a new research strategy for research on the mechanism of traditional Chinese Medicine.

Bioactive compounds from Huashi Baidu decoction possess both antiviral and anti-inflammatory effects against COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic is an ongoing global health concern, and effective antiviral reagents are urgently needed. Traditional Chinese medicine theory-driven natural drug research and development (TCMT-NDRD) is a feasible method to address this issue as the traditional Chinese medicine formulae have been shown effective in the treatment of COVID-19. Huashi Baidu decoction (Q-14) is a clinically approved formula for COVID-19 therapy with antiviral and anti-inflammatory effects. Here, an integrative pharmacological strategy was applied to identify the antiviral and anti-inflammatory bioactive compounds from Q-14. Overall, a total of 343 chemical compounds were initially characterized, and 60 prototype compounds in Q-14 were subsequently traced in plasma using ultrahigh-performance liquid chromatography with quadrupole time-of-flight mass spectrometry. Among the 60 compounds, six compounds (magnolol, glycyrrhisoflavone, licoisoflavone A, emodin, echinatin, and quercetin) were identified showing a dose-dependent inhibition effect on the SARS-CoV-2 infection, including two inhibitors (echinatin and quercetin) of the main protease (Mpro), as well as two inhibitors (glycyrrhisoflavone and licoisoflavone A) of the RNA-dependent RNA polymerase (RdRp). Meanwhile, three anti-inflammatory components, including licochalcone B, echinatin, and glycyrrhisoflavone, were identified in a SARS-CoV-2-infected inflammatory cell model. In addition, glycyrrhisoflavone and licoisoflavone A also displayed strong inhibitory activities against cAMP-specific 3',5'-cyclic phosphodiesterase 4 (PDE4). Crystal structures of PDE4 in complex with glycyrrhisoflavone or licoisoflavone A were determined at resolutions of 1.54 Å and 1.65 Å, respectively, and both compounds bind in the active site of PDE4 with similar interactions. These findings will greatly stimulate the study of TCMT-NDRD against COVID-19.

Differentiation of Mountain- and Garden-Cultivated Ginseng with Different Growth Years Using HS-SPME-GC-MS Coupled with Chemometrics.

Although there are differences in the appearance of Mountain-Cultivated Ginseng (MCG) and Garden-Cultivated Ginseng (GCG), it is very difficult to distinguish them when the samples are processed to slices or powder. Moreover, there is significant price difference between them, which leads to the widespread adulteration or falsification in the market. Thus, the authentication of MCG and GCG is crucial for the effectiveness, safety, and quality stability of ginseng. In the present study, a headspace solid-phase microextraction gas chromatography mass spectrometry (HS-SPME-GC-MS) coupled with chemometrics approach was developed to characterize the volatile component profiles in MCG and GCG with 5-,10-,15-growth years, and subsequently to discover differentiating chemical markers. As a result, we characterized, for the first time, 46 volatile components from all the samples by using the NIST database and the Wiley library. The base peak intensity chromatograms were subjected to multivariate statistical analysis to comprehensively compare the chemical differences among the above samples. MCG5-,10-,15-years and GCG5-,10-,15-years samples were mainly divided into two groups by unsupervised principal component analysis (PCA), and 5 potential cultivation-dependent markers were discovered based on orthogonal partial least squares-discriminant analysis (OPLS-DA). Moreover, MCG5-,10-,15-years samples were divided into three blocks, and 12 potential growth-year-dependent markers enabled differentiation. Similarly, GCG5-,10-,15-years samples were also separated into three groups, and six potential growth-year-dependent markers were determined. The proposed approach could be applied to directly distinguish MCG and GCG with different growth years and to identify the differentiation chemo-markers, which is an important criterion for evaluating the effectiveness, safety, and quality stability of ginseng.

Mechanism Repositioning Based on Integrative Pharmacology: Anti-Inflammatory Effect of Safflower in Myocardial Ischemia-Reperfusion Injury.

Safflower (Carthamus tinctorius. L) possesses anti-tumor, anti-thrombotic, anti-oxidative, immunoregulatory, and cardio-cerebral protective effects. It is used clinically for the treatment of cardio-cerebrovascular disease in China. This study aimed to investigate the effects and mechanisms of action of safflower extract on myocardial ischemia-reperfusion (MIR) injury in a left anterior descending (LAD)-ligated model based on integrative pharmacology study and ultra-performance liquid chromatography-quadrupole time-of-flight-tandem mass spectrometer (UPLC-QTOF-MS/MS). Safflower (62.5, 125, 250 mg/kg) was administered immediately before reperfusion. Triphenyl tetrazolium chloride (TTC)/Evans blue, echocardiography, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay, lactate dehydrogenase (LDH) ability, and superoxide dismutase (SOD) levels were determined after 24 h of reperfusion. Chemical components were obtained using UPLC-QTOF-MS/MS. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to analyze mRNA and protein levels, respectively. Safflower dose-dependently reduced myocardial infarct size, improved cardiac function, decreased LDH levels, and increased SOD levels in C57/BL6 mice. A total of 11 key components and 31 hub targets were filtered based on the network analysis. Comprehensive analysis indicated that safflower alleviated inflammatory effects by downregulating the expression of NFκB1, IL-6, IL-1ß, IL-18, TNFα, and MCP-1 and upregulating NFκBia, and markedly increased the expression of phosphorylated PI3K, AKT, PKC, and ERK/2, HIF1α, VEGFA, and BCL2, and decreased the level of BAX and phosphorylated p65. Safflower shows a significant cardioprotective effect by activating multiple inflammation-related signaling pathways, including the NFκB, HIF-1α, MAPK, TNF, and PI3K/AKT signaling pathways. These findings provide valuable insights into the clinical applications of safflower.

Systematic identification of chemical components in Fufang Shuanghua oral liquid and screening of potential active components against SARS-CoV-2 protease.

Coronavirus disease (COVID-19) caused by SARS-COV-2 infection has been widely prevalent in many countries and has become a common challenge facing mankind. Traditional Chinese medicine (TCM) has played a prominent role in this pandemic, and especially TCM with the function of "heat-clearing and detoxifying" has shown an excellent role in anti-virus. Fufang Shuanghua oral liquid (FFSH) has been used to treat the corresponding symptoms of influenza such as fever, nasal congestion, runny nose, sore throat, and upper respiratory tract infections in clinic, which are typical symptoms of COVID-19. The content of chlorogenic acid, andrographolide and dehydrated andrographolide as the quality control components of FFSH is not less than 1.0 mg/mL, 60 µg/mL and 60 µg/mL respectively. In this study, UPLC-Q-TOF-MS/MS was employed to describe the chemical profile of FFSH. Virtual screening and fluorescence resonance energy transfer (FRET) were used to screen the effective components of FFSH acting on SARS-CoV-2 main protease (Mpro). As a result, 214 compounds in FFSH were identified or preliminarily characterized by UPLC-Q-TOF-MS/MS, and 61 active ingredients with potential inhibitory effects on Mpro were selected through receptor-based and ligand-based virtual screening. In particular, quercetin, forsythoside A, and linoleic acid showed a good inhibitory effect on Mpro in FRET evaluation with IC50 values of 26.15 µM, 22.26 µM and 47.09 µM respectively, and had a strong binding affinity with the receptor Mpro (6LU7) in molecular docking. CYS145 and HIS41 were the main amino acid residues affected by small molecules in the protein binding domain. In brief, we characterized, for the first time, 214 chemical components in FFSH, and three of them, including quercetin, forsythoside A and linoleic acid, were screened out to exert beneficial anti-COVID-19 effects through CYS145 and HIS41 sites, which may provide a new research strategy for TCM to develop new therapeutic drugs against COVID-19.

Pan-Genome Analysis of TIFY Gene Family and Functional Analysis of CsTIFY Genes in Cucumber.

Cucumbers are frequently affected by gray mold pathogen Botrytis cinerea, a pathogen that causes inhibited growth and reduced yield. Jasmonic acid (JA) plays a primary role in plant responses to biotic stresses, and the jasmonate-ZIM-Domain (JAZ) proteins are key regulators of the JA signaling pathway. In this study, we used the pan-genome of twelve cucumber varieties to identify cucumber TIFY genes. Our findings revealed that two CsTIFY genes were present in all twelve cucumber varieties and showed no differences in protein sequence, gene structure, and motif composition. This suggests their evolutionary conservation across different cucumber varieties and implies that they may play a crucial role in cucumber growth. On the other hand, the other fourteen CsTIFY genes exhibited variations in protein sequence and gene structure or conserved motifs, which could be the result of divergent evolution, as these genes adapt to different cultivation and environmental conditions. Analysis of the expression profiles of the CsTIFY genes showed differential regulation by B. cinerea. Transient transfection plants overexpressing CsJAZ2, CsJAZ6, or CsZML2 were found to be more susceptible to B. cinerea infection compared to control plants. Furthermore, these plants infected by the pathogen showed lower levels of the enzymatic activities of POD, SOD and CAT. Importantly, after B. cinerea infection, the content of JA was upregulated in the plants, and cucumber cotyledons pretreated with exogenous MeJA displayed increased resistance to B. cinerea infection compared to those pretreated with water. Therefore, this study explored key TIFY genes in the regulation of cucumber growth and adaptability to different cultivation environments based on bioinformatics analysis and demonstrated that CsJAZs negatively regulate cucumber disease resistance to gray mold via multiple signaling pathways.