Rhizobacterial Bacillus enrichment in soil enhances smoke tree resistance to Verticillium wilt.

Verticillium wilt, caused by the soilborne fungus Verticillium dahliae, poses a serious threat to the health of more than 200 plant species worldwide. Although plant rhizosphere-associated microbiota can influence plant resistance to V. dahliae, empirical evidence underlying Verticillium wilt resistance of perennial trees is scarce. In this study, we systemically investigated the effect of the soil microbiota on the resistance of smoke trees (Cotinus coggygria) to Verticillium wilt using field, greenhouse and laboratory experiments. Comparative analysis of the soil microbiota in the two stands of smoke trees suggested that Bacillus represented the most abundant and key microbial genus related to potential disease suppression. Smoke tree seedlings were inoculated with isolated Bacillus strains, which exhibited disease suppressiveness and plant growth-promoting properties. Furthermore, repletion of Bacillus agents to disease conducive soil significantly resulted in reduced incidence of smoke tree wilt and increased resistance of the soil microbiota to V. dahliae. Finally, we explored a more effective combination of Bacillus agents with the fungicide propiconazole to combat Verticillium wilt. The results establish a foundation for the development of an effective control for this disease. Overall, this work provides a direct link between Bacillus enrichment and disease resistance of smoke trees, facilitating the development of green control strategies and measurements of soil-borne diseases.

[Chemical constituents from Alangium chinense subsp. pauciflorum].

Chemical constituents of 70% ethanol extract of Alangium chinense subsp. pauciflorum were investigated. The 70% ethanol extract of A. chinense subsp. pauciflorum was isolated and purified by D-101 macroporous resins, silica gel, Sephadex LH-20 and other methods. As a result, nineteen compounds were isolated and identified as 4-cyclohexene-1α,2α,3α-triol-1-O-ß-D-glucoside(1), 1ß,4α,6α,13-tetrahydroxy-eudesm-11(12)-ene(2), sucrose(3), 1'-O-benzyl-α-L-rhamnopyranosyl-(1″→6')-ß-D-glucopyranoside(4), bis(2-ethylhexyl)benzene-1,2-dicarboxylate(5),(Z)-10-heneicosenoic acid(6), di-O-methylcrenati(7), methyl-α-D-fructofuranoside(8), ß-daucosterol(9), syringic acid(10), vanillicacid(11), octacosanol(12), isoarborinol(13), 2,7-dihydroxy-6-methyl-4-(1-methylethyl)-1-naphthalenecarboxylate(14),vanillin(15), coniferyl aldehyde(16), 9(11)-dehydroergosterolperoxide(17), 5α,8α-epidioxy-(22E,24R)-ergosta-6,22-dien-3ß-ol(18), ß-sitosterol(19), respectively. Compounds 1 and 2 were new compounds, compounds 5-11, 13, 15-18 were isolated from Alangium for the first time.The anti-inflammatory activity of compourd 1 was determinded by the LPS-induced RAW264.7 macrophage inflammation model. The results showed that the new compound 1 has a certain inhibitory effect on LPS-induced NO production of RAW264.7 cells, and the inhibitory rate was 54.57%.

Linker Mediated Electronic-State Manipulation of Conjugated Organic Polymers Enabling Highly Efficient Oxygen Reduction.

Conjugated polymers with tailorable composition and microarchitecture are propitious for modulating catalytic properties and deciphering inherent structure-performance relationships. Herein, we report a facile linker engineering strategy to manipulate the electronic states of metallophthalocyanine conjugated polymers and uncover the vital role of organic linkers in facilitating electrocatalytic oxygen reduction reaction (ORR). Specifically, a set of cobalt phthalocyanine conjugated polymers (CoPc-CPs) wrapped onto carbon nanotubes (denoted CNTs@CoPc-CPs) are judiciously crafted via in situ assembling square-planar cobalt tetraaminophthalocyanine (CoPc(NH2)4) with different linear aromatic dialdehyde-based organic linkers in the presence of CNTs. Intriguingly, upon varying the electronic characteristic of organic linkers from terephthalaldehyde (TA) to 2,5-thiophenedicarboxaldehyde (TDA) and then to thieno/thiophene-2,5-dicarboxaldehyde (bTDA), their corresponding CNTs@CoPc-CPs exhibit gradually improved electrocatalytic ORR performance. More importantly, theoretical calculations reveal that the charge transfer from CoPc units to electron-withdrawing linkers (i.e., TDA and bTDA) drives the delocalization of Co d-orbital electrons, thereby downshifting the Co d-band energy level. Accordingly, the active Co centers with more positive valence state exhibit optimized binding energy toward ORR-relevant intermediates and thus a balanced adsorption/desorption pathway that endows significant enhancement in electrocatalytic ORR. This work demonstrates a molecular-level engineering route for rationally designing efficient polymer catalysts and gaining insightful understanding of electrocatalytic mechanisms.

Molecule and Microstructure Modulations of Cyano-Containing Electrodes for High-Performance Fully Organic Batteries.

Cyano-containing electrodes usually promise high theoretical potentials while suffering from uncontrollable self-dissolution and sluggish reaction kinetics. Herein, to remedy their limitations, an unprecedented core-shell heterostructured electrode of carbon nanotubes encapsulated in poly(1,4-dicyanoperfluorobenzene sulfide) (CNT@PFDCB) is rationally crafted via molecule and microstructure modulations. Specifically, the linkage of sulfide bridges of PFDCB prevents the active cyano groups from dissolving, resulting in a robust structure. The fluorinations modulate the electronic configurations in frontier orbitals, allowing higher electrical conductivity and elevated output voltage. Combined with the core-shell architecture to unlock the sluggish diffusion kinetics for both electrons and guest ions, the CNT@PFDCB exhibits an impressive capacity (203.5 mAh g-1), remarkable rate ability (127.6 mAh g-1 at 3.0 A g-1), and exceptional cycling stability (retaining 81.1 % capacity after 3000 cycles at 1.0 A g-1). Additionally, the Li-storage mechanisms regarding PFDCB are thoroughly revealed by in situ attenuated total reflection infrared spectroscopy, in situ Raman spectroscopy, and theoretical simulations, which involve the coordination interaction between Li ions and cyano groups and the electron delocalization along the conjugated skeleton. More importantly, a practical fully organic cell based on the CNT@PFDCB is well-validated that demonstrates a tremendous potential of cyanopolymer as the cathode to replace its inorganic counterparts.

VdTps2 Modulates Plant Colonization and Symptom Development in Verticillium dahliae.

The trehalose biosynthesis pathway is a potential target for antifungal drugs development. Trehalose phosphate synthase (TPS) and phosphatase are widely conserved components of trehalose biosynthesis in fungi. However, the role of trehalose biosynthesis in the vascular plant-pathogenic fungus Verticillium dahliae remains unclear. Here, we investigated the functions of the TPS complex, including VdTps1, VdTps2, and VdTps3 in V. dahliae. Unlike VdTps2, deletion of VdTps1 or VdTps3 did not alter any phenotypes compared with the wild-type strain. In contrast, the ΔVdTps2 strain showed severely depressed radial growth due to the abnormal swelling of the hyphal tips. Further, deletion of VdTps2 increased microsclerotia formation, melanin biosynthesis, and resistance to cell-wall perturbation and high-temperature stress. Virulence assays and quantification of fungal biomass revealed that deletion of VdTps2 delayed disease symptom development, as evident by the reduced virulence and decreased biomass of the ΔVdTps2 strain in plant stem tissue following inoculation. Additionally, increases in penetration peg formation observed in the ΔVdTps2 strain in the presence of H2O2 suggested that VdTps2 suppresses initial colonization. Our results also revealed the role of VdTps2 as a regulator of autophagy. Together, these results indicate that VdTps2 contributes to plant colonization and disease development. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The Transcription Factor CcRlm1 Regulates Cell Wall Maintenance and Poplar Defense Response by Directly Targeting CcChs6 and CcGna1 in Cytospora chrysosperma.

Maintenance of cell wall integrity is important for fungal cell morphology against external stresses and even virulence. Although the transcription factor Rlm1 is known to play major regulatory roles in the maintenance of cell integrity, the underlying mechanism of how Rlm1 contributes to cell wall integrity and virulence in phytopathogenic fungi remains unclear. Here, we demonstrated that CcRlm1 plays important roles in cell wall maintenance and virulence in the poplar canker fungus Cytospora chrysosperma. Among putative downstream targets, CcChs6 (chitin synthase) and CcGna1 (glucosamine 6-phosphate N-acetyltransferase) were found to be direct targets of CcRlm1 and shown to function in chitin synthesis and virulence. Furthermore, we found stronger induction of poplar defense responses when challenged with these gene deletion mutants. Collectively, these results suggest that CcRlm1 plays a critical role in the regulation of cell wall maintenance, stress response, and virulence by directly regulating CcChs6 and CcGna1 in C. chrysosperma. IMPORTANCE Cytospora chrysosperma causes canker diseases on woody plants, and the molecular basis of its infection is not well understood. This study shows that CcRlm1 is the major regulator of chitin synthesis and virulence of the poplar canker fungus. Our research contributes to further understanding the molecular basis of the interaction between C. chrysosperma and poplar.

Six New Constituents from the Fruit of Hypericum patulum and Their Anti-Inflammatory Activity.

Four new xanthone glucosides, 3-hydroxy-2-methoxyxanthone-4-O-ß-D-glucopyranoside (1), 4,8-dihydroxy-2-methoxyxanthone-3-O-ß-D-glucopyranoside (2), 2-methoxyxanthone-5-O-ß-D-glucopyranoside (3), 4-hydroxy-2-methoxyxanthone-3-O-ß-D-glucopyranoside (4), a new phenolic acid, 4,4'-dihydroxy-3,3'-imino-di-benzoic acid monomethyl ester (5), and a new isoquinoline, methyl 6-hydroxy-1-oxo-1,2,3,4-tetrahydroisoquinoline-4-carboxylate (6) were isolated from the fruit of Hypericum patulum. The structural elucidation of the isolated compounds was primarily based on HR-ESI-MS, UV, IR, 1D and 2D NMR. All compounds were evaluated for their inhibitory effect against LPS-induced NO production in RAW 264.7 cells. Compound 2, 3 exhibited moderate inhibitory activity against NO production.

CRISPR-Cas Detection Coupled with Isothermal Amplification of Bursaphelenchus xylophilus.

The pine wood nematode (PWN), Bursaphelenchus xylophilus, causes significant damage to pine trees and, thus, poses a serious threat to pine forests worldwide, particularly in China, Korea, and Japan. A fast, affordable, and ultrasensitive detection of B. xylophilus is urgently needed for disease diagnosis. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)-based diagnostics have reshaped molecular diagnosis, with high speed, precision, specificity, strength, efficiency, and versatility. Herein, we established two isothermal diagnostics methods based on CRISPR-based platforms (CRISPR/Cas12a and CRISPR/Cas13a) for B. xylophilus-specific detection via fluorescence or lateral-flow strip readout. The guide RNA and CRISPR RNA were designed to target the 5S ribosomal DNA intergenic spacer sequences region of B. xylophilus. Recombinase-aided amplification was used for preamplification whose reaction condition was 37°C for 15 min. The sensitivity of CRISPR/Cas12a could reach 94 copies/µl of plasmid DNA, or 2.37 copies/µl of purified genomic DNA (gDNA) within 45 min at 37°C, while the sensitivity of CRISPR/Cas13a was 1,000 times higher than that of CRISPR/Cas12a of plasmid DNA in 15 min or 100 times higher of purified gDNA at the minimum reaction time of 4 min via fluorescence measurement. The CRISPR/Cas12a assay enabled the detection of 0.01 PWNs per 100 mg of pine wood, 10 times higher than that of the CRISPR/Cas13a assay. This work enriches molecular detection approaches for B. xylophilus and provides huge potential for ultrasensitive and rapid methods to detect B. xylophilus in pine wood, facilitating point-of-sample diagnostic processing for pine wilt disease management.

Metabolites-Based Network Pharmacology to Preliminarily Verify In Vitro Anti-Inflammatory Effect of Ardisiacrispin B.

Ardisiae Crenatae Radix is an ethnic medicinal herb with good anti-inflammatory activity. Ardisiacrispin B is one of the main components in Ardisiae Crenatae Radix extract, with a content of up to 16.27%, and it may be one of the pharmacological components through which Ardisiae Crenatae Radix exerts anti-inflammatory activity. At present, reports on ardisiacrispin B mainly focus on anti-tumor effects, and there have been no reports on anti-inflammatory activities. As a triterpenoid saponin, due to its large molecular weight and complex structure, the composition of substances that function in the body may include other forms after metabolism, in addition to compounds with original structures. Exploring the anti-inflammatory effects on the prototypes and metabolites of the compound may provide a more comprehensive response to the characteristics of ardisiacrispin B's anti-inflammatory action. In this study, ardisiacrispin B was analyzed for metabolites to explore its metabolic processes in vivo. Subsequently, the anti-inflammatory effects of the prototypes and metabolites were further analyzed through network pharmacology, with the expectation of discovering the signaling metabolic pathways through which they may act. Finally, the anti-inflammatory effects of ardisiacrispin B in vitro and the effects on key signaling pathways at the protein level were explored. The results of this study showed that the isolated compounds were confirmed to be ardisiacrispin B. After the metabolite analysis, a total of 26 metabolites were analyzed, and the metabolism process in rats mainly involves oxidation, dehydration, glucuronide conjugation, and others. Speculation as to the anti-inflammatory molecular mechanisms of the prototypes and metabolites of ardisiacrispin B revealed that it may exert its anti-inflammatory effects mainly by affecting the PI3K-AKT pathway. Further anti-inflammatory mechanisms demonstrated that ardisiacrispin B had a good anti-inflammatory effect on LPS-induced RAW264.7 cells and a strong inhibitory effect on NO, TNF-α, and IL-1ß release in cells. Furthermore, it had significant inhibitory effects on the expression of PI3K, P-PI3K, AKT, and P-AKT. This study supplements the gaps in the knowledge on the in vivo metabolic process of ardisiacrispin B and explores its anti-inflammatory mechanism, providing an experimental basis for the development and utilization of pentacyclic triterpenoids.

Polygonum orientale L. Alleviates Myocardial Ischemia-Induced Injury via Activation of MAPK/ERK Signaling Pathway.

Although Polygonum orientale L. (PO) has a beneficial effect on treatment of myocardial ischemia (MI), its mechanism remains unclear. This study aimed to explore the pharmacological mechanism of PO against MI through MAPK signaling pathways. Firstly, the therapeutic effect of PO was evaluated for treatment of MI mice. Using Western blot and immunohistochemistry, the influence of PO on MAPK signaling pathways and cell apoptosis was investigated. Subsequently, one key pathway (ERK) of MAPK signaling pathways was screened out, on which PO posed the most obvious impact. Finally, an inhibitor of ERK1/2 was utilized to further verify the regulatory effect of PO on the MAPK/ERK signaling pathway. It was found that PO could reduce the elevation of the ST segment; injury of heart tissue; the activity of LDH, CK, NOS, cNOS and iNOS and the levels of NO, BNP, TNF-α and IL-6. It is notable that PO could significantly modulate the protein content of p-ERK/ERK in mice suffering from MI but hardly had an effect on p-JNK/JNK and p-p38/p38. Additionally, the expressions of bax, caspase3 and caspase9 were inhibited in heart tissue in the PO-treated group. To evaluate whether ERK1/2 inhibitor (PD98059) could block the effect of PO on treatment of MI, both PO and PD98059 were given to mice with MI. It was discovered that the inhibitor indeed could significantly reverse the regulatory effects of PO on the above indicators, indicating that PO could regulate p-ERK/ERK. This study provides experimental evidence that PO extenuates MI injury, cardiomyocyte apoptosis and inflammation by activating the MAPK/ERK signaling pathway.

[Qualitative and quantitative study of constituents in Lysionoti Herba based on UHPLC-Q-Exactive Orbitrap HRMS and HPLC-UV].

Ultra-high performance liquid chromatography-quadrupole-Exactive Orbitrap high resolution mass spectrometry(UHPLC-Q-Exactive Orbitrap HRMS) was employed to systematically analyze the chemical constituents in Lysionoti Herba, and high perfor-mance liquid chromatography-ultraviolet(HPLC-UV) to determine the content of main compounds. A Synergi~(TM) Hydro-RP 100 Å colu-mn(2 mm×100 mm, 2.5 µm) was used for gradient elution with acetonitrile-0.1% aqueous formic acid as the mobile phase at a flow rate of 0.2 mL·min~(-1) and a column temperature of 40 ℃. MS and MS/MS were conducted with electrospray ionization(ESI) in both positive and negative modes. The chemical components in Lysionoti Herba were identified by comparison with the retention time and mass spectra of reference compounds and the relevant mass spectral data reported in MS databases and relevant literature. Furthermore, the content of five constituents(neochlorogenic acid, chlorogenic acid, forsythoside B, acteoside, and nevadensin) in different Lysiono-ti Herba samples was simultaneously determined by HPLC-UV at the wavelength of 330 nm. A total of 84 compounds were identified in Lysionoti Herba, including 27 flavonoids, 20 phenylethanoid glycosides, 5 amino acids, 18 organic acids, 1 alkaloid, 6 nucleosides, and 7 others. The content of neochlorogenic acid, chlorogenic acid, forsythoside B, acteoside, and nevadensin showed good linear relationship(r>0.999) with the peak area within certain concentration ranges, which were 3.22-102.90, 12.84-410.82, 31.63-1 012.01, 25.00-800.11, and 4.08-130.51 µg·mL~(-1), respectively. The instrument precision, method repeatability, and solution stability all met requirement, and the average recovery rate was 97.31%-100.2%, with RSD ranging from 0.95% to 2.4%. The content of the five components varied among different Lysionoti Herba samples collected from different regions of Guizhou, and the average content of forsythoside B was the highest. The established qualitative method can rapidly and efficiently identify the chemical components of Lysionoti Herba, and the developed HPLC-UV method can simultaneously determine the content of five components in a simple, ra-pid, and accurate manner, providing a scientific basis for the quality evaluation of Lysionoti Herba.

Imbalanced GSH/ROS and sequential cell death.

Reactive oxygen species (ROS) are produced in cells during metabolic processes. Excessive intracellular ROS may react with large biomolecules, such as DNA, RNA, proteins, and small biomolecules, that is, glutathione (GSH) and unsaturated fatty acids. GSH has physiological functions, including free radical scavenging, anti-oxidation, and electrophile elimination. The disruption of ROS/GSH balance results in the deleterious oxidation and chemical modification of biomacromolecules, which eventually leads to cell-cycle arrest and proliferation inhibition, and even induces cell death. Imbalanced ROS/GSH may result from a direct increase of ROS, consumption of GSH, intracellular oxidoreductase interference, or thioredoxin activity reduction. Some chemicals including arsenic trioxide (ATO), pyrogallol (PG), and carbobenzoxy-Leu-Leu-leucinal (MG132) could also disrupt the balance of GSH and ROS. This article reviews the occurrence and consequences of the imbalance between GSH and ROS and introduces factors responsible for the disruption of cellular ROS and GSH balance, resulting in cell death. "GSH" and "ROS" were used as keywords to search the relevant literaturess.

A Cytochrome P450 Monooxygenase in Nondefoliating Strain of Verticillium dahliae Manipulates Virulence via Scavenging Reactive Oxygen Species.

Verticillium dahliae is a broad host-range phytopathogenic fungus that causes destructive vascular wilt on plants worldwide. Cytochrome P450 monooxygenases, also known as CYPs/P450s, are broadly distributed in organisms and are involved in a diverse array of molecular/metabolic processes. In this study, using reverse transcription quantitative PCR analysis, we observed that the expression of a P450 gene (Chr2g00380) in the E-class P450, group IV from V. dahliae isolate JR2 was highly induced during tomato infection. Targeted deletion of Chr2g00380 in JR2 did not affect hyphal growth and morphology; however, the mutants exhibited increased sensitivity to H2O2 and defects in melanized microsclerotia formation compared with the wild type. Loss of Chr2g00380 resulted in reduced virulence on tomato and tobacco plants but did not cause phenotypic changes in infection structure formation or in the penetration of cellophane membranes. These data provide evidence for an involvement of a cytochrome P450 monooxygenase in virulence in V. dahliae.

Comparative pharmacokinetics of 11 components from the active part of Gerberae Piloselloidis Herba after oral administration in control and asthmatic mice.

Gerberae Piloselloidis Herba, a traditional Chinese medicine, is often employed to treat such lung-related diseases as coughs, asthma, and pulmonary carbuncles in southwest China. Our previous study demonstrated that its active fraction, prepared from Gerberae Piloselloidis Herba, exerts an obvious beneficial effect on asthma. However, the pharmacokinetics of its major constituents remain unclear. Therefore, an ultra-performance mass spectrometry-electrospray ionization-tandem mass spectrometry method was successfully established to simultaneously perform the pharmacokinetics of the main 11 components of the active fraction between normal and ovalbumin-induced asthmatic mice. Compared to the normal group, in asthmatic mice the peak concentration of arbutin, marmesin, caffeoylquinic acids, and flavonoid glycosides clearly increased, while for luteolin it significantly declined; the area under the curve for arbutin and luteolin showed an increase, but the values of marmesin, caffeoylquinic acids, and flavonoid glycosides revealed a decline; the peak time for arbutin, caffeoylquinic acids and flavonoid glycosides decreased, while for marmesin and luteolin it significantly augmented; apart from marmesin, the half-life for all compounds shortened significantly. It is indicated that the pathology of asthma could lead to an alteration in the pharmacokinetic profiles of the 11 components in plasma, providing a reference for further exploration of the pharmacodynamic basis of the anti-bronchial effect of Gerberae Piloselloidis Herba.

Protective effects on ovalbumin-induced mouse asthma models and qualitative and quantitative analysis of multiple compounds in Gerberae Piloselloidis Herba.

Gerberae Piloselloidis Herba is widely used to treat cough and asthma in China. However, its effects on allergic asthma as related to its chemical compositions have not been fully elucidated, and there is a scarcity of methods to determine multi-component contents for quality control. In this study, protective effects of Gerberae Piloselloidis Herba on ovalbumin-induced asthma models were investigated, while qualitative and quantitative analyses of multiple constituents in Gerberae Piloselloidis Herba were conducted by using an ultrahigh-performance liquid chromatography-Q Exactive hybrid quadrupole-orbitrap high-resolution accurate mass spectrometry and an ultrahigh-performance liquid chromatography-photodiode array detection. The results showed that Gerberae Piloselloidis Herba could significantly mitigate asthma symptoms, reduce eosinophils counts in the bronchoalveolar lavage fluid, as well as decrease IgE, IL-5, and IL-13 concentration, and inflammatory cellular infiltration in lung tissues. A total of 51 compounds were tentatively identified, in which the content of 10 representative compounds was determined in 24 batches of Gerberae Piloselloidis Herba by using an ultrahigh-performance liquid chromatography method with good linearity, precision, repeatability, accuracy, and stability. This research presents a comprehensive strategy combining biological activity evaluation with chemical profiling, providing a useful and comprehensive reference for further application and quality control of Gerberae Piloselloidis Herba.

Chemical profiling and quantification of multiple components in Jin-Gu-Lian capsule using a multivariate data processing approach based on UHPLC-Orbitrap Exploris 240 MS and UHPLC-MS/MS.

The Jin-Gu-Lian capsule, a Chinese Miao herbal compound, is widely used to treat rheumatoid arthritis. In this study, a rapid, selective, and sensitive UHPLC-Orbitrap Exploris 240 MS method was developed to analyze the chemical composition of Jin-Gu-Lian capsules. A total of 88 compounds were identified, including 23 flavonoids, 23 organic acids, 14 phenylpropanoids, 12 phenols, eight alkaloids, four terpenes, three quinones, and one ketone. Among these, 21 compounds were clearly detected based on a comparison with reference standards and selected as quality control markers. Thereafter, these compounds were simultaneously determined in the Jin-Gu-Lian capsules. The established method was successfully validated and applied for the simultaneous determination of 21 biologically active compounds in Jin-Gu-Lian capsules of 27 sample batches. Quantitative data of the analytes were analyzed using multivariate statistical analysis to determine the quality of the Jin-Gu-Lian capsules. Four compounds (JGLC6 [salidroside], JGLC8 [chlorogenic acid], JGLC12 [liriodendrin], JGLC19 [quercetin]) were identified as chemical markers for quality control of Jin-Gu-Lian capsules. Altogether, the established method was validated as a novel and efficient tool, that can be used for rapid analysis of Jin-Gu-Lian capsules. Accordingly, this study serves as a reference for scientific research on traditional Chinese and ethnic medicine.

[Effects of Gukang Capsules on activity and protein expression of hepatic cytochrome P450 enzymes in rats].

Gukang Capsules are often used in combination with drugs to treat fractures, osteoarthritis, and osteoporosis. Cytochrome P450(CYP450) mainly exists in the liver and participates in the oxidative metabolism of a variety of endogenous and exogenous substances and serves as an important cause of drug-metabolic interactions and adverse reactions. Therefore, it is of great significance to study the effect of Gukang Capsules on the activity and expression of CYP450 for increasing its clinical rational medication and improving the safety of drug combination. In this study, the Cocktail probe method was used to detect the changes in the activities of CYP1A2, CYP3A2, CYP2C11, CYP2C19, CYP2D4, and CYP2E1 in rat liver after treatment with high-, medium-and low-dose Gukang Capsules. The rat liver microsomes were extracted by the calcium chloride method, and protein expression of the above six CYP isoform enzymes was detected by Western blot. The results showed that the low-dose Gukang Capsules could induce CYP3A2 and CYP2D4 in rats, medium-dose Gukang Capsules had no effect on them, and high-dose Gukang Capsules could inhibit them in rats. The high-dose Gukang Capsules did not affect CYP2C11 in rats, but low-and medium-dose Gukang Capsules could induce CYP2C11 in rats. Gukang Capsules could inhibit CYP2C19 in rats and induce CYP1A2 in a dose-independent manner, but did not affect CYP2E1. If Gukang Capsules were co-administered with CYP1A2, CYP2C19, CYP3A2, CYP2C11, and CYP2D4 substrates, the dose should be adjusted to avoid drug interactions.

[Uptake and transport of Laportea bulbifera extract in Caco-2 cell model].

Laportea bulbifera extract is effective in resisting inflammation and shows a good therapeutic effect on rheumatoid arthritis in rats. However, the absorption characteristics of active components in L. bulbifera extract in Caco-2 cells are still unclear, which limits the in-depth development of L. bulbifera resources. The purpose of this study was to investigate the absorption and transport mechanism of the active components of L. bulbifera extract in the Caco-2 cell model and explore the effects of different factors(concentration, time, pH value, temperature, and efflux transporter inhibitor) on its uptake and transport. The results showed that L. bulbifera extract at the concentration of 2.0-8.0 mg·mL~(-1) showed no toxicity to Caco-2 cells. The uptake and transport of L. bulbifera extract in the Caco-2 cell model were concentration-dependent and time-dependent. The main absorption mechanism was passive diffusion, and acidic condition(pH 5.0-6.0) and 37 ℃ were more favorable for drug absorption. P_(app)>1.0×10~(-6 )cm·s~(-1) of each component indicated that L. bulbifera was a moderately absorbed drug. P-gp, MRP2, and BCRP were not involved in its uptake and transport.

[Potential pharmacodynamic substances of Laportea bulbifera in treatment of rheumatoid arthritis based on serum pharmacochemistry and pharmacology].

The present study investigated the pharmacodynamic material basis of Laportea bulbifera in the treatment of rheumatoid arthritis. Firstly, human rheumatoid arthritis fibroblast-like synoviocyte line MH7A was cultured in vitro and treated with tumor necrosis factor alpha(TNF-α, 50 ng·mL~(-1)). The proliferation and the levels of inflammatory cytokines such as prostaglandin E2(PGE2), interleukin-1ß(IL-1ß), and interleukin-6(IL-6) of the MH7A cells exposed to the serum containing L. bulbifera were determined to evaluate the anti-rheumatoid arthritis effects of the serum. Furthermore, the ultra-performance liquid chromatography tandem mass spectrometry fingerprints of the L. bulbifera crude extract, the drug-containing serum, and the drug-free serum were compared to identify the compounds newly generated in the serum after oral administration of the extract. According to the peak areas of common peaks and the results of anti-rheumatoid arthritis effect test, the active components were identified. The serum containing L. bulbifera significantly inhibited the proliferation of the MH7A cells activated by TNF-α and the expression of PGE2, IL-6, and IL-1ß. Thirty newly generated compounds were detected in the drug-containing serum. Among them, neochlorogenic acid, cryptochlorogenic acid, chlorogenic acid, rutin, isoquercitrin, luteoloside, kaempferol-3-O-rutinoside, and quercitrin were also present in the crude extract. Twelve characteristic peaks(3, 7, 8, 14, 18, 19, 21, 23, 24, m6, m7, and m15) were significantly correlated with the pharmaceutical effect. According to the correlations, neochlorogenic acid, cryptochlorogenic acid, and chlorogenic acid had great contributions to the anti-rheumatoid arthritis activity. This study preliminarily clarified the potential pharmacodynamic substances of L. bulbifera in the treatment of rheumatoid arthritis, which laid a theoretical and experimental foundation for further development and application of the medicinal plant.

[Effect of Laportea bulbifera extract on CYP450 enzyme activities in rats by Cocktail probe drug method].

The Cocktail probe drug method was used to evaluate the effect of Laportea bulbifera extract on the different subtypes of CYP450 enzyme activities in rats, and to provide references for its clinical rational drug use. The rats were randomly divided into a high-dose L. bulbifera group(0.45 g·kg~(-1)·d~(-1)) and a low-dose L. bulbifera group(0.09 g·kg~(-1)·d~(-1)). After continuous gavage for 7 and 14 days, the Cocktail probe mixing solution, including caffeine, midazolam, tolbutamide, omeprazole, metoprolol, and chlorzoxazone, was injected into the tail vein, and the blood sample was obtained from the tail vein at different time points. Ultra-high performance liquid chromatography-mass spectrometry(UPLC-MS) was established to determine the concentration of six probe drugs in rat plasma. DAS 2.0 was used to calculate its pharmacokinetic parameters, and the effect of L. bulbifera extract on CYP1 A2, CYP2 C9, CYP2 C19, CYP2 D6, CYP2 E1, and CYP3 A4 in rats was investigated. As compared with the blank control group, under the omeprazole index, the AUC_(0-t) and AUC_(0-∞) of the 7-day administration groups and the 14-day high-dose group were significantly decreased, and the CLz and Vz were significantly increased. Under the midazolam index, the AUC_(0-t) and AUC_(0-∞) of the 7-day low-dose group and the 14-day administration group were significantly decreased, and the CLz was significantly increased. In addition, the AUC_(0-∞) of the 7-day high-dose group was also significantly decreased. Under the index of metoprolol, the AUC_(0-t) and AUC_(0-∞) of each experimental group were decreased significantly, and the CLz and Vz of the 7-day low-dose group and the 14-day low-dose group were increased significantly. Under the caffeine index, the AUC_(0-t) and AUC_(0-∞) of the 7-day administration groups were increased significantly, the CLz was decreased significantly, and the t_(1/2 z) of the 14-day high-dose group was increased significantly. Under the chlorzoxazone index, the AUC_(0-t) and AUC_(0-∞) of the 7-day low-dose group were increased significantly, and the CLz was decreased significantly. Under the tolbutamide index, there was no statistical difference in the pharmacokinetic parameters. In conclusion, L. bulbifera extract induces the activities of CYP2 C19, CYP3 A4, and CYP2 D6, inhibits the activities of CYP1 A2 and CYP2 E1, and does not affect the activity of CYP2 C9.