New Triterpenoids from Lansium domesticum Corr. cv kokossan and Their Cytotoxic Activity.

Lansium domesticum Corr. is a member of the Meliaceae family that is widely spread in tropical and subtropical region of Asia and America. Traditionally, the fruit of this plant has been consumed because of its sweet taste. However, the fruit peels and the seeds of this plant have been rarely utilized. The previous chemical investigation of this plant showed the presence of secondary metabolites with many biological activities, including cytotoxic triterpenoid. Triterpenoids is a class of secondary metabolites which contain thirty carbon atoms in the main skeleton. The high modification of this type of compound, including the ring opening, highly oxygenated carbons, and the degradation of its carbon chain to give the nor-triterpenoid structure, is responsible for its cytotoxic activity. In this paper, we isolated and elucidated the chemical structure of two new onoceranoid triterpenes, kokosanolides E (1) and F (2), from the fruit peels of L. domesticum Corr., along with a new tetranortriterpenoid, kokosanolide G (3), from the seeds of L. domesticum Corr. The structural determination of compounds 1-3 was undertaken through FTIR spectroscopic analysis, 1D and 2D NMR, mass spectrometry, as well as through a comparison of the chemical shifts of the partial structures of compounds 1-3 with the literature data. The cytotoxic properties of compounds 1-3 were tested against MCF-7 breast cancer cells using the MTT assay. Moderate activity was shown by compounds 1 and 3, with IC50 values of 45.90 and 18.41 µg/mL, respectively, while compound 2 showed no activity (IC50 168.20 µg/mL). For the onoceranoid-type triterpene, the high symmetrical structure of compound 1 is presumably the reason for its better cytotoxic activity compared with that of compound 2. Compound 3 showed moderate activity, mainly because of the presence of the furan ring, which, based on the literature, gives better cytotoxic activity in a tetranortriterpenoid-type structure. The findings of three new triterpenoid compounds from L. domesticum indicate the significant value of this plant as a source of new compounds.

Limonin ameliorates indomethacin-induced intestinal damage and ulcers through Nrf2/ARE pathway.

BACKGROUND: Nonsteroidal anti-inflammatory drugs (NSAIDs) can cause intestinal damage and ulcers and the incidence is increasing. Limonin plays an important role in the regulation of inflammatory diseases, but it has not been reported in the treatment of intestinal injury and ulcers. METHODS: Indomethacin (INDO) induced intestinal injury and ulcer model in rats. The indexes related to intestinal injury were detected. Western blot and molecular docking techniques were used to detect the docking between Limonin and Nrf2. Next, ML385, an inhibitor of Nrf2/ARE signaling pathway, was applied to treat intestinal epithelial IEC-6 cells induced by INDO. And CCK8, Western blot, TUNEL, ELISA, DCFH-DA assay, kits, and immunofluorescence were conducted to detect cell activity, apoptosis, inflammatory response, oxidative stress, and tight junction again. RESULTS: INDO can significantly induce intestinal ulcerative lesions in rats. Limonin could improve intestinal ulcerative lesions induced by INDO in rats. Limonin could reduce INDO-induced inflammatory response and oxidative stress in the small intestine of rats, and improve the intestinal barrier dysfunction induced by INDO. Limonin could dock with Nrf2 structure and activate Nrf2/ARE signaling pathway. ML385 could reverse the protective effect of Limonin against INDO-induced cell damage. CONCLUSION: Limonin ameliorates INDO-induced intestinal damage and ulcers through Nrf2/ARE pathway.

UV light-driven late-stage skeletal reorganization to diverse limonoid frameworks: A proof of concept for photobiosynthesis.

Late-stage skeletal reorganization (LSSR) is a type of fascinating organic transformation processes in natural product total synthesis. However, few facile and effective LSSR methodologies have hitherto been developed. Here, LSSR of limonoid natural products via photochemical cascades is first reported. Starting from xyloelves A and B, nine distinct limonoid products with five unprecedented scaffolds are generated. The photocascade pathways of these natural products and mechanistic rationale via intramolecular triplet energy transfer are revealed by quantum mechanical calculations. Most notably, ultraviolet light-driven transannular and stereoselective C â†’ C 1,4-acyl migration is first found as a photochemical approach, particularly for LSSR of natural products. This approach holds promise for designing LSSR strategies to access bioactive cage-like molecules. Besides that, our findings provide a clear proof of concept for natural product photobiosynthesis. Xyloelf A, substantially ameliorating concanavalin A-induced liver injury in mice, could be used as a unique molecular template for hepatoprotective drug discovery.

Munronin V with 7/7/6 tricarbocyclic framework from Munronia henryi harms inhibits tau pathology by activating autophagy.

Munronin V (1), isolated from Munronia henryi Harms, is the first example, to the best of our knowledge, of an unprecedented 7/7/6 tricarbocyclic framework featuring an unusual A,B-seco-limonoid ring. The structures of munronin V were established from extensive spectroscopic and electronic circular dichroism (ECD) analyses. The novel A,B-seco with two seven-membered lactones was formed as a result of Baeyer-Villiger oxidation. Compound 1 activated autophagy and inhibited Tau pathology as revealed by flow cytometric analyses, confocal imaging analysis and western blotting, and this effect was mediated by transcription factor EB (TFEB). These findings suggested that 1 might have potential as a compound for combating Alzheimer's disease.

Complex scaffold remodeling in plant triterpene biosynthesis.

Triterpenes with complex scaffold modifications are widespread in the plant kingdom. Limonoids are an exemplary family that are responsible for the bitter taste in citrus (e.g., limonin) and the active constituents of neem oil, a widely used bioinsecticide (e.g., azadirachtin). Despite the commercial value of limonoids, a complete biosynthetic route has not been described. We report the discovery of 22 enzymes, including a pair of neofunctionalized sterol isomerases, that catalyze 12 distinct reactions in the total biosynthesis of kihadalactone A and azadirone, products that bear the signature limonoid furan. These results enable access to valuable limonoids and provide a template for discovery and reconstitution of triterpene biosynthetic pathways in plants that require multiple skeletal rearrangements and oxidations.

Hainanxylogranolides A-F: New Limonoids isolated from the seeds of Hainan mangrove plant Xylocarpus granatum.

Six new limonoids, named hainanxylogranolides A-F (1-6), together with nineteen known ones (7-25) were isolated from the seeds of a Hainan mangrove Xylocarpus granatum. The structures of the new compounds were established by extensive NMR spectroscopic data combined with the DFT and TDDFT calculated electronic circular dichroism spectra. Hainanxylogranolide A (1) is the aromatic B-ring limonoid containing a central pyridine ring and a C-17 substituted γ(21)-hydroxybutenolide moiety. Hainanxylogranolide B (2) belongs to the small group of mexicanolides containing a C3-O-C8 bridge, whereas hainanxylogranolides C and D (3 and 4) are mexicanolides comprising a C1-O-C8 bridge. Compounds 9 and 25 posed obvious inhibition effect on the tube formation of HUVECs. There are only about 25% tube-like structures were observed at the concentration of 40.0 µM of compound 25. The antiviral activities of the isolates against herpes simplex virus-1 (HSV-1) and severe fever with thrombocytopenia syndrome virus (SFTSV) were tested in vitro. Compound 23 exhibited moderate anti-SFTSV activity with the IC50 value of 29.58 ± 0.73 µM. This is the first report of anti-angiogenic effect and anti-SFTSV activity of limonoids from the genus Xylocarpus.

A Split Renilla Luciferase Complementation Assay for the Evaluation of Hsp90/Aha1 Complex Disruptors and Their Activity at the Aha1 C-Terminal Domain.

Disruption of interactions between Hsp90 and the cochaperone protein, Aha1, has emerged as a therapeutic strategy to inhibit Aha1-driven cancer metastasis and tau aggregation in models of tauopathy. A combination of split Renilla luciferase assays was developed to screen and quantify the ability of small molecules to disrupt interactions between Hsp90 and both full length Aha1 protein (Aha1-FL) and the Aha1 C-terminal domain (Aha1-CTD). This luminescence-based approach was used to identify withaferin A and gedunin as disruptors of Hsp90/Aha1 interactions and provided insight into the binding regions for gambogic acid and gedunin on the Hsp90 homodimer. All compounds tested that disrupted Hsp90/Aha1-CTD interactions were found to disrupt interactions between Hsp90 and Aha1-FL, suggesting that interactions between Hsp90 and the Aha1-CTD play a key role in the stability of Hsp90/Aha1 complexes.

Limonin, a Component of Immature Citrus Fruits, Activates Anagen Signaling in Dermal Papilla Cells.

Hair loss remains a significant problem that is difficult to treat; therefore, there is a need to identify safe natural materials that can help patients with hair loss. We evaluated the hair anagen activation effects of limonin, which is abundant in immature citrus fruits. Limonin increased the proliferation of rat dermal papilla cells (rDPC) by changing the levels of cyclin D1 and p27, and increasing the number of BrdU-positive cells. Limonin increased autophagy by decreasing phosphorylated mammalian target of rapamycin levels and increasing the phospho-Raptor, ATG7 and LC3B. Limonin also activated the Wnt/ß-catenin pathway by increasing phospho-ß-catenin levels. XAV939, a Wnt/ß-catenin inhibitor, inhibited these limonin-induced changes, including induced autophagy, BrdU-positive cells, and cell proliferation. Limonin increased the phosphorylated AKT levels in both two-dimensional cultured rDPC and three-dimensional spheroids. Treatment with the PI3K inhibitor wortmannin inhibited limonin-induced proliferation, and disrupted other limonin-mediated changes, including decreased p27, increased BrdU-positive cells, induced autophagy, and increased ATG7 and LC3B levels. Wortmannin also inhibited limonin-induced cyclin D1 and LC3 expression in spheroids. Collectively, these results indicate that limonin can enhance anagen signaling by activating autophagy via targeting the Wnt/ß-catenin and/or PI3K/AKT pathways in rDPC, highlighting a candidate nutrient for hair loss treatment.

Phytochemistry and Biological Activities of Guarea Genus (Meliaceae).

Guarea is one of the largest genera of the American Meliaceae family, consisting of over 69 species which are widely distributed in Mexico, Argentina, and Africa and are used in traditional medicine for several diseases. Previous studies reported that the Guarea species produce secondary metabolites such as sesquiterpenoid, diterpenoid, triterpenoid, limonoid, steroid, and aromatic compounds. The preliminary chemical investigation commenced by isolating the limonoid compound, dihydrogedunin, in 1962; then, 240 compounds were obtained from the isolation and hydrodistillation process. Meanwhile, sesquiterpenoid is a significant compound with 52% of Guarea species. The extract and compounds were evaluated for their anti-inflammation, antimalarial, antiparasitic, antiprotozoal, antiviral, antimicrobial, insecticidal, antioxidant, phosphorylation inhibitor, and cytotoxic biological activities. The Guarea genus has also been reported as one of the sources of active compounds for medicinal chemistry. This review summarizes some descriptions regarding the types of Guarea species, especially ethnobotany and ethnopharmacology, such as the compounds isolated from the part of this genus, various isolation methods, and their bioactivities. The information can be used in further investigations to obtain more bioactive compounds and their reaction mechanisms.

A Gedunin-Type Limonoid, 7-Deacetoxy-7-Oxogedunin, from Andiroba (Carapa guianensis Aublet) Reduced Intracellular Triglyceride Content and Enhanced Autophagy in HepG2 Cells.

The seed oil of Carapa guianensis Aublet (Andiroba) has been used in folk medicine for its insect-repelling, anti-inflammatory, and anti-malarial activities. This study aimed to examine the triglyceride (TG) reducing effects of C. guianensis-derived limonoids or other commercially available limonoids in human hepatoblastoma HepG2 cells and evaluate the expression of lipid metabolism or autophagy-related proteins by treatment with 7-deacetoxy-7-oxogedunin (DAOG; 1), a principal limonoid of C. guianensis. The gedunin-type limonoids, such as DAOG (% of control at 20 µM: 70.9 ± 0.9%), gedunin (2, 74.0 ± 1.1%), epoxyazadiradione (4, 73.4 ± 2.0%), 17ß-hydroxyazadiradione (5, 79.9 ± 0.6%), 7-deacetoxy-7α-hydroxygedunin (6, 61.0 ± 1.2%), andirolide H (7, 87.4 ± 2.2%), and 6α-hydroxygedunin (8, 84.5 ± 1.1%), were observed to reduce the TG content at lower concentrations than berberine chloride (BBR, a positive control, 84.1 ± 0.3% at 30 µM) in HepG2 cells pretreated with high glucose and oleic acid. Andirobin-, obacunol-, nimbin-, and salannin-type limonoids showed no effect on the intracellular TG content in HepG2 cells. The TG-reducing effect of DAOG was attenuated by the concomitant use of compound C (dorsomorphin), an AMPK inhibitor. Further investigation on the detailed mechanism of action of DAOG at non-cytotoxic concentrations revealed that the expressions of autophagy-related proteins, LC3 and p62, were upregulated by treatment with DAOG. These findings suggested that gedunin-type limonoids from Andiroba could ameliorate fatty liver, and that the action of DAOG in particular is mediated by autophagy.

Concise Chemoenzymatic Synthesis of Gedunin.

The limonoids have attracted significant attention from the synthetic community owing to their striking structural complexity and medicinal potential. Recent efforts notwithstanding, synthetic access to many intact or ring D-seco limonoids still remains elusive. Here, we report the first de novo synthesis of gedunin, a ring D-seco limonoid with HSP90 inhibitory activity, that proceeds in 13 steps. Two enabling features in our strategy are the application of modern catalytic transformations to set the key quaternary centers in the carbocyclic core and the use of biocatalytic oxidation at C3 to establish a chemical handle to access the A-ring enone motif. The strategy presented herein may provide an entry point to a wider range of oxidized limonoids.

Swietenine and swietenolide from Swietenia macrophylla king improve insulin secretion and attenuate apoptosis in H2 O2 induced INS-1 cells.

Oxidative stress is an important factor that causes pancreatic ß-cell dysfunction leading to the development and aggravation of diabetes. Swietenine (Stn) and swietenolide (Std) were isolated from the fruits of Swietenia macrophylla King and had the potential effects on treatment and prevention of diabetes. The aim of this study is to investigate the effects of Stn and Std on insulin secretion and apoptosis in H2 O2 induced insulinoma cell line (INS-1) cells. In the present study, INS-1 cells were treated with 300 µM H2 O2 for 4 h to establish the oxidative damage model. Cell apoptosis, insulin secretion, reactive oxygen species (ROS), superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione (GSH) levels, and Caspase-3 enzyme activity were measured via corresponding methods. Finally, pancreatic duodenal home box factor-1 (PDX-1), B cell lymphoma-2 (Bcl-2), and Bax protein expression were detected by western blot. Experimental results showed that Stn and Std could significantly improve the INS-1 cell viability, increase the secretion of insulin and reduce the ROS level in H2 O2 induced INS-1 cells. Furthermore, the SOD and GSH levels increased, and the MDA levels decreased compared with the model group after Stn and Std treatment. In addition, after treated with Stn and Std, cell apoptosis was improved, and the activity of Caspase 3 was also significantly inhibited. Meanwhile, Western blot results showed that Stn and Std could up-regulate the expression of PDX-1 protein, and affect the cell apoptosis pathway by up-regulating the expression of Bcl-2 protein and down-regulating the expression of Bax protein. In conclusion, Stn and Std can signifcantly improve the insulin secretion function, protect oxidative stress injury, and reduce apoptosis in H2 O2 induced INS-1 cells, which provides a research basis for Stn and Std to be new drug candidates for the treatment and prevention of diabetes.

TKF, a mexicanolide-type limonoid derivative, suppressed hepatic stellate cells activation and liver fibrosis through inhibition of the YAP/Notch3 pathway.

BACKGROUND: Liver fibrosis is a common scarring response and may ultimately lead to liver cancer, unfortunately, there is currently no effective antifibrotic drug approved for human use. Limonoids exhibit a broad spectrum of biological activities; however, the potential role of limonoids against fibrosis is largely unknown. PURPOSE: This study investigates the antifibrotic activities and potential mechanisms of TKF (3-tigloyl-khasenegasin F), a natural mexicanolide-type limonoid derivative. STUDY DESIGN/METHODS: Two well-established mouse models (CCl4 challenge and bile duct ligation) were used to assess anti-fibrotic effects of TKF in vivo. Human hepatic stellate cell (HSC) line LX-2 and mouse primary hepatic stellate cells (pHSCs) also served as in vitro liver fibrosis models. RESULT: TKF administration significantly attenuated hepatic histopathological injury and collagen accumulation and suppressed fibrogenesis-associated gene expression including Col1a1, Acta2, and Timp1. In LX-2 cells and mouse pHSCs, TKF dose-dependently suppressed HSC activation and the expression levels of fibrogenic markers. Mechanistic studies showed that TKF inhibited Notch3-Hes1 and YAP signalings in vivo and in vitro. Furthermore, YAP inhibition or knockdown downregulated the Notch3 expression; however, Notch3 inhibition or knockdown did not affect the level of YAP in activated HSC. We revealed that TKF inhibited Notch3-Hes1 activation and downregulated hepatic fibrogenic gene expression via inhibiting YAP. CONCLUSION: The therapeutic benefit of TKF against liver fibrosis results from inhibition of YAP and Notch3-Hes1 pathways, indicating that TKF may be a novel therapeutic candidate for liver fibrosis.

Identification of Concomitant Inhibitors against Glutamine Synthetase and Isocitrate Lyase in Mycobacterium tuberculosis from Natural Sources.

Tuberculosis (T.B.) is a disease that occurs due to infection by the bacterium, Mycobacterium tuberculosis (Mtb), which is responsible for millions of deaths every year. Due to the emergence of multidrug and extensive drug-resistant Mtb strains, there is an urgent need to develop more powerful drugs for inclusion in the current tuberculosis treatment regime. In this study, 1778 molecules from four medicinal plants, Azadirachta indica, Camellia sinensis, Adhatoda vasica, and Ginkgo biloba, were selected and docked against two chosen drug targets, namely, Glutamine Synthetase (G.S.) and Isocitrate Lyase (I.C.L.). Molecular Docking was performed using the Glide module of the SchrÓ§dinger suite to identify the best-performing ligands; the complexes formed by the best-performing ligands were further investigated for their binding stability via Molecular Dynamics Simulation of 100 ns. The present study suggests that Azadiradione from Azadirachta indica possesses the potential to inhibit Glutamine Synthetase and Isocitrate Lyase of M. tuberculosis concomitantly. The excellent docking score of the ligand and the stability of receptor-ligand complexes, coupled with the complete pharmacokinetic profile of Azadiradione, support the proposal of the small molecule, Azadiradione as a novel antitubercular agent. Further, wet lab analysis of Azadiradione may lead to the possible discovery of a novel antitubercular drug.

Deacetylated nimbin analog N2 fortifies alloxan-induced pancreatic ß-cell damage in insulin-resistant zebrafish larvae by upregulating phosphoenolpyruvate carboxykinase (PEPCK) and insulin levels.

This study aims to evaluate the protective behaviour of N2, a semi-natural analog of nimbin, for its anti-diabetic efficacy against alloxan-induced oxidative damage and ß-cell dysfunction in in-vivo zebrafish larvae. A 500 µM of alloxan was exposed to zebrafish larvae for 24 h to induce oxidative stress in the pancreatic ß-cells and co-exposed with N2 to study the protection of N2 by inhibiting ROS by DCFH-DA, DHE and NDA staining along with Cellular damage, apoptosis and lipid peroxidation. The zebrafish was further exposed to 500 µM alloxan for 72 h to induce ß-cell destruction along with depleted glucose uptake and co-exposed to N2 to study the protective mechanism. Glucose levels were estimated, and PCR was used to verify the mRNA expression of phosphoenolpyruvate carboxykinase (PEPCK) and insulin. Alloxan induced (24 h) oxidative stress in the pancreatic ß-cells in which N2's co-exposure inhibited ROS by eliminating O-2 radicals and restoring the glutathione levels, thus preventing cellular damage and lipid peroxidation. The zebrafish exposed to 500 µM alloxan for 72 h was observed with ß-cell destruction along with depleted glucose uptake when stained with 2NBDG, wherein N2 was able to protect the pancreatic ß-cells from oxidative damage, promoted high glucose uptake and reduced glucose levels. N2 stimulated insulin production and downregulated PEPCK by inhibiting gluconeogenesis, attenuating post-prandial hyperglycemia. N2 may contribute to anti-oxidant protection against alloxan-induced ß-cell damage and anti-hyperglycemic activity, restoring insulin function and suppressing PEPCK expression.

Terpenoids from the Seeds of Toona sinensis and Their Ability to Attenuate High Glucose-Induced Oxidative Stress and Inflammation in Rat Glomerular Mesangial Cells.

Toona sinensis (A. Juss.) Roem is an edible medicinal plant that belongs to the genus Toona within the Meliaceae family. It has been confirmed to display a wide variety of biological activities. During our continuous search for active constituents from the seeds of T. sinensis, two new acyclic diterpenoids (1-2), together with five known limonoid-type triterpenoids (3-7), five known apotirucallane-type triterpenoids (8-12), and three known cycloartane-type triterpenoids (13-15), were isolated and characterized. Their structures were identified based on extensive spectroscopic experiments, including nuclear magnetic resonance (NMR), high-resolution electrospray ionization mass spectra (HR-ESI-MS), and electronic circular dichroism (ECD), as well as the comparison with those reported in the literature. We compared these findings to those reported in the literature. Compounds 5, 8, and 13-14 were isolated from the genus Toona, and compounds 11 and 15 were obtained from T. sinensis for the first time. The antidiabetic nephropathy effects of isolated compounds against high glucose-induced oxidative stress and inflammation in rat glomerular mesangial cells (GMCs) were assessed in vitro. The results showed that new compounds 1 and 2 could significantly increase the levels of Nrf-2/HO-1 and reduce the levels of NF-κB, TNF-α, and IL-6 at concentrations of 30 µM. These results suggest that compounds 1 and 2 might prevent the occurrence and development of diabetic nephropathy (DN) and facilitate the research and development of new antioxidant and anti-inflammatory drugs suitable for the prevention and treatment of DN.

Identification of Gedunin from a Phytochemical Depository as a Novel Multidrug Resistance-Bypassing Tubulin Inhibitor of Cancer Cells.

The chemotherapy of tumors is frequently limited by the development of resistance and severe side effects. Phytochemicals may offer promising candidates to meet the urgent requirement for new anticancer drugs. We screened 69 phytochemicals, and focused on gedunin to analyze its molecular modes of action. Pearson test-base correlation analyses of the log10IC50 values of 55 tumor cell lines of the National Cancer Institute (NCI), USA, for gedunin with those of 91 standard anticancer agents revealed statistically significant relationships to all 10 tested microtubule inhibitors. Thus, we hypothesized that gedunin may be a novel microtubule inhibitor. Confocal microscopy, cell cycle measurements, and molecular docking in silico substantiated our assumption. Agglomerative cluster analyses and the heat map generation of proteomic data revealed a subset of 40 out of 3171 proteins, the expression of which significantly correlated with sensitivity or resistance for the NCI cell line panel to gedunin. This indicates the complexity of gedunin's activity against cancer cells, underscoring the value of network pharmacological techniques for the investigation of the molecular modes of drug action. Finally, we correlated the transcriptome-wide mRNA expression of known drug resistance mechanism (ABC transporter, oncogenes, tumor suppressors) log10IC50 values for gedunin. We did not find significant correlations, indicating that gedunin's anticancer activity might not be hampered by classical drug resistance mechanisms. In conclusion, gedunin is a novel microtubule-inhibiting drug candidate which is not involved in multidrug resistance mechanisms such as other clinically established mitotic spindle poisons.

Integrated miRNA and transcriptome profiling to explore the molecular mechanism of Spodoptera frugiperda larval midgut in response to azadirachtin exposure.

As a destructive agricultural pest, Spodoptera frugiperda has spread worldwide in the past few years. Azadirachtin, an environmentally friendly and most promising compound, showed adverse effects, including mortality and growth inhibition, against S. frugiperda. While the effects of azadirachtin on the midgut of this pest remain to be determined. In this study, structural damage was observed in the larval midguts of S. frugiperda with azadirachtin exposure. RNA-seq on the larval midguts with different azadirachtin treatments was performed. Compared to the control group, a total of 3344 and 4759 differentially expressed genes (DEGs) were identified in the midguts with 0.1 and 0.5 µg/g azadirachtin exposure, respectively. Among them, the DEGs encoding detoxification enzymes/proteins, immune-related proteins, digestion and absorption-related proteins, and transcript factors were further analyzed. High-throughput sequencing was also used for the identification of differentially expressed microRNAs in different treatments. A total of 153 conserved miRNAs and 147 novel miRNAs were identified, of which 11 and 29 miRNAs were affected by 0.1 and 0.5 µg/g azadirachtin treatments, respectively. The integrated analysis found that 13 and 178 miRNA versus mRNA pairs were acquired in the samples with 0.1 and 0.5 µg/g azadirachtin treatments, respectively. The results of high-throughput sequencing were confirmed by real-time quantitative polymerase chain reaction (RT-qPCR). These results provide useful information for revealing the molecular mechanism of S. frugiperda larval midgut in response to azadirachtin.

[Study on characteristic chromatogram and content determination of Wuzhuyu Decoction reference sample].

In this study, the critical quality attributes of Wuzhuyu Decoction reference sample were explored by using characteristic chromatogram, index component content and dry extract rate as indexes.The dissemination relationship of quantity value between medicinal materials-decoction pieces-reference sample was investigated to preliminarily formulate the quality standard of the reference sample.The characteristic chromatogram of 15 batches of Wuzhuyu Decoction was established by high performance liquid chromatography(HPLC) and the similarity analysis was conducted.Common peaks were demarcated and assigned to medicinal materials.Moreover, quantitative determination of limonin, evodiamine, rutaecarpine and ginsenoside Rb_1 of Wuzhuyu Decoction were performed.The dissemination of quantity value was explored combined with dry extract rate, similarity of characteristic chromatogram and transfer rate of index component content.A total of 18 common peaks were identified in the corresponding materials of Wuzhuyu Decoction reference sample, with the similarity of characteristic chromatogram greater than 0.9, and Fructus Evodiae, Radix Ginseng, Rhizoma Zingiberis Recens and Fructus Jujubae contributed 9, 5, 8 and 2 chromatographic peaks, respectively.The index component content of corresponding materials and the transfer rates of medicinal materials-decoction pieces and decoction pieces-reference sample of different batches of Wuzhuyu Decoction reference sample were as follows: the content of limonin was 0.16%-0.51%, and the transfer rates were 83.66%-115.60% and 38.54%-54.58%, respectively; the content of evodiamine was 0.01%-0.11%, the transfer rated were 80.80%-116.15% and 3.23%-12.93%, respectively; the content of rutaecarpine was 0.01%-0.05%, the transfer rates were 84.33%-134.53% and 5.72%-21.24%, respectively; the content of ginsenoside Rb_1 was 0.06%-0.11%, and the transfer rates were 90.00%-96.92% and 32.45%-67.24%, respectively.The dry extract rate of the whole prescription was 22.58%-29.89%.In this experiment, the dissemination of quantity value of Wuzhuyu Decoction reference sample was analyzed by the combination of characteristic chromatogram, index component content and dry extract rate.A scientific and stable quality evaluation method of the reference sample was preliminarily established, which provided basis for the subsequent development of Wuzhuyu Decoction and the quality control of related preparations.

Azadirachtin inhibits the development and metabolism of the silk glands of Spodoptera frugiperda and affects spinning behavior.

BACKGROUND: Spodoptera frugiperda is a major agricultural pest, and the dispersal of its larvae by spinning silk is one of the causes of crop damage. At present, there are relatively few reports of pest control that affect larvae spinning silk. In this study, the effect of spinning behavior of the S. frugiperda larvae was investigated through a series of experiments. RESULTS: The 3rd instar larvae of S. frugiperda were exposed to azadirachtin, and the pathological changes in the silk glands of S. frugiperda and the differences in their metabolites were analyzed by scanning electron microscopy, histological sectioning, transmission electron microscopy and metabolomics. The results showed that azadirachtin could affect the silk gland of S. frugiperda. After 48 h of treatment with azadirachtin, the silk gland lumen of S. frugiperda appeared vacuolated. KEGG showed that 31 different metabolites were identified, of which 12 were upregulated and 19 were downregulated. These metabolites were enriched in 15 different metabolic pathways, which indicated that the silk gland of S. frugiperda was closely related to the formation of fatty acids and energy metabolism for the silk formation process. CONCLUSIONS: This study provides a preliminary report of the effect of azadirachtin on the spinning behavior of the S. frugiperda larvae. Metabolomic results indicated that histidine, glycine and leucine, which are related to serine protein synthesis, were down-regulated. Azadirachtin can damage the silk glands of S. frugiperda and thus affect spinning behavior. This provides the basis for the control of S. frugiperda by spinning silk. © 2022 Society of Chemical Industry.