Esculin induces endoplasmic reticulum stress and drives apoptosis and ferroptosis in colorectal cancer via PERK regulating eIF2α/CHOP and Nrf2/HO-1 cascades.

ETHNOPHARMACOLOGICAL RELEVANCE: Cortex fraxini (also known as Qinpi), the bark of Fraxinus rhynchophylla Hance and Fraxinus stylosa Lingelsh, constitutes a crucial component in several traditional Chinese formulas (e.g., Baitouweng Tang, Jinxiao Formula, etc.) and has demonstrated efficacy in alleviating intestinal carbuncle and managing diarrhea. Cortex fraxini has demonstrated commendable anticancer activity in the realm of Chinese ethnopharmacology; nevertheless, the underlying mechanisms against colorectal cancer (CRC) remain elusive. AIM OF THE STUDY: Esculin, an essential bioactive compound derived from cortex fraxini, has recently garnered attention for its ability to impede viability and induce apoptosis in cancer cells. This investigation aims to assess the therapeutic potential of esculin in treating CRC and elucidate the underlying mechanisms. MATERIALS AND METHODS: The impact of esculin on CRC cell viability was assessed using CCK-8 assay, Annexin V/PI staining, and Western blotting. Various cell death inhibitors, along with DCFH-DA, ELISA, biochemical analysis, and Western blotting, were employed to delineate the modes through which esculin induces HCT116 cells death. Inhibitors and siRNA knockdown were utilized to analyze the signaling pathways influenced by esculin. Additionally, an azomethane/dextran sulfate sodium (AOM/DSS)-induced in vivo CRC mouse model was employed to validate esculin's potential in inhibiting tumorigenesis and to elucidate its underlying mechanisms. RESULTS: Esculin significantly suppressed the viability of various CRC cell lines, particularly HCT116 cells. Investigation with diverse cell death inhibitors revealed that esculin-induced cell death was associated with both apoptosis and ferroptosis. Furthermore, esculin treatment triggered cellular lipid peroxidation, as evidenced by elevated levels of malondialdehyde (MDA) and decreased levels of glutathione (GSH), indicative of its propensity to induce ferroptosis in HCT116 cells. Enhanced protein levels of protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) and p-eIF2α suggested that esculin induced cellular endoplasmic reticulum (ER) stress, subsequently activating the Nrf2/ARE signaling pathway and initiating the transcriptional expression of heme oxygenase (HO)-1. Esculin-induced excessive expression of HO-1 could potentially lead to iron overload in HCT116 cells. Knockdown of Ho-1 significantly attenuated esculin-induced ferroptosis, underscoring HO-1 as a critical mediator of esculin-induced ferroptosis in HCT116 cells. Furthermore, utilizing an AOM/DSS-induced colorectal cancer mouse model, we validated that esculin potentially inhibits the onset and progression of colon cancer by inducing apoptosis and ferroptosis in vivo. CONCLUSIONS: These findings provide comprehensive insights into the dual induction of apoptosis and ferroptosis in HCT116 cells by esculin. The activation of the PERK signaling pathway, along with modulation of downstream eIF2α/CHOP and Nrf2/HO-1 cascades, underscores the mechanistic basis supporting the clinical application of esculin on CRC treatment.

Esculin targets TLR4 to protect against LPS-induced septic cardiomyopathy.

BACKGROUND: Esculin, a main active ingredient from Cortex fraxini, possesses biological activities such as anti-thrombosis, anti-inflammatory, and anti-oxidation effects. However, the effects of Esculin on septic cardiomyopathy remains unclear. This study aimed to explore the protective properties and mechanisms of Esculin in countering sepsis-induced cardiac trauma and dysfunction. METHODS AND RESULTS: In lipopolysaccharide (LPS)-induced mice model, Esculin could obviously improve heart injury and function. Esculin treatment also significantly reduced the production of inflammatory and apoptotic cells, the release of inflammatory cytokines, and the expression of oxidative stress-associated and apoptosis-associated markers in hearts compared to LPS injection alone. These results were consistent with those of in vitro experiments based on neonatal rat cardiomyocytes. Database analysis and molecular docking suggested that TLR4 was targeted by Esculin, as shown by stable hydrogen bonds formed between Esculin with VAL-308, ASN-307, CYS-280, CYS-304 and ASP-281 of TLR4. Esculin reversed LPS-induced upregulation of TLR4 and phosphorylation of NF-κB p65 in cardiomyocytes. The plasmid overexpressing TLR4 abolished the protective properties of Esculin in vitro. CONCLUSION: We concluded that Esculin could alleviate LPS-induced septic cardiomyopathy via binding to TLR4 to attenuate cardiomyocyte inflammation, oxidative stress and apoptosis.

LC/MS/MS and GC/MS/MS metabolic profiling of Leontodon hispidulus, in vitro and in silico anticancer activity evaluation targeting hexokinase 2 enzyme.

Leontodon hispidulus Boiss is a wild annual plant growing in Egypt. The present study aims for the first time, to evaluate the phytochemical profile of the main secondary metabolites of the optimized ethanolic extract of the plant using Quadrupole Time-of-Flight Liquid chromatography-mass spectrometry and Gas chromatography-mass spectrometry. It also aims to assess the anticancer activity of its different fractions against the prostate carcinoma cell line. Moreover, an in-silico docking study was performed using the Hexokinase-two enzyme. LC-qToF-MS analysis revealed the tentative identification of 36 phenolic compounds including the glycosides of (luteolin, quercetin, kaempferol, apigenin, isorhamnetin, and daidzein), coumarines (esculin, esculetin, and daphnetin), and phenolic acids (chlorogenic, caffeic, quinic, P-coumaric, and rosmarinic). GC-MS/MS analysis revealed the presence of 18 compounds where palmitic acid, myristic acid, alpha-amyrin, and beta-amyrin were the major ones. The cytotoxic activity results revealed that methylene chloride and ethyl acetate fractions showed the highest cytotoxic activity against the PC3 cell line, with IC50 values of 19, and 19.6 µg/ml, respectively. Interestingly, the docking study demonstrated that apigenin-7-O-glucoside, luteolin-7-O-glucoside, kaempferol-3-O-glucuronide, quercetin-4'-O-glucoside, esculin, rosmarinic acid, chlorogenic acid, and α-amyrin exhibited high affinity to the selected target, HEK-2 enzyme.

Recovery and characterization of ß-glucosidase-producing non-Saccharomyces yeasts from the fermentation broth of Vitis labruscana Baily × Vitis vinifera L. for investigation of their fermentation characteristics.

The present study focuses on investigating 60 strains of yeast isolated from the natural fermentation broth of Vitis labruscana Baily × Vitis vinifera L. These strains underwent screening using lysine culture medium and esculin culture medium, resulting in the identification of 27 local non-Saccharomyces yeast strains exhibiting high ß-glucosidase production. Subsequent analysis of their fermentation characteristics led to the selection of four superior strains (Z-6, Z-11, Z-25, and Z-58) with excellent ß-glucosidase production and fermentation performance. Notably, these selected strains displayed a dark coloration on esculin medium and exhibited robust gas production during Duchenne tubules' fermentation test. Furthermore, all four non-Saccharomyces yeast strains demonstrated normal growth under specific conditions including SO2 mass concentration ranging from 0.1 to 0.3 g/L, temperature between 25 and 30 °C, glucose mass concentration ranging from 200 to 400 g/L, and ethanol concentration at approximately 4%. Molecular biology identification confirmed that all selected strains belonged to Pichia kudriavzevii species which holds great potential for wine production.

In vivo comprehensive metabolite profiling of esculetin and esculin derived from chicory in hyperuricemia rats using ultra-high-performance liquid chromatography coupled with quadrupole-orbitrap high-resolution mass spectrometry.

Chicory, renowned for its multifaceted benefits, houses two vital coumarins, esculetin and esculin, both instrumental in reducing uric acid. This study emphasizes the metabolic pathways of esculetin and esculin under both standard and hyperuricemia conditions. Hyperuricemia was induced in Sprague-Dawley rats using oxonic acid potassium salt (300 mg·kg-1 ) and a 10% fructose water regimen over 21 days. Leveraging the ultra-high-performance liquid chromatography-Q Exactive hybrid quadrupole-orbitrap high resolution mass spectrometry, we analyzed the fragmentation behaviors of esculetin and esculin in rat bio-samples. Post oral-intake of esculetin or esculin, a notable dip in serum uric acid levels was observed in hyperuricemia rats. The investigation unveiled 24 esculetin metabolites and 14 for esculin. The metabolic pathways of both compounds were hydrolysis, hydroxylation, hydrogenation, dehydroxylation, glucuronidation, sulfation, and methylation. Interestingly, certain metabolites presented variations between standard and hyperuricemia rats, indicating that elevated levels of uric acid may affect enzyme activity linked to these metabolic reactions. This is the first systematic study on comparison of metabolic profiles of esculetin and esculin in both normal and hyperuricemia states, which was helpful to enrich our understanding of the complicated structure-activity relationships between esculin and esculetin and shed light to their action mechanism.

Pharmacological activities of esculin and esculetin: A review.

Esculin and esculetin are 2 widely studied coumarin components of Cortex Fraxini, which is a well-known herbal medicine with a 2000-year history. In vivo and in vitro studies have demonstrated that both have a variety of pharmacological activities, including antioxidant, anti-tumor, anti-inflammatory, antibacterial, antidiabetic, immunomodulatory, anti-atherosclerotic, and so on. Their underlying mechanisms of action and biological activities include scavenging free radicals, modulating the nuclear factor erythroid 2-related factor 2 pathway, regulating the cell cycle, inhibiting tumor cell proliferation and migration, promoting mitochondrial pathway apoptosis, inhibiting the NF-κB and MAPK signaling pathways, regulating CD4+ T cells differentiation and associated cytokine release, inhibiting vascular smooth muscle cells, etc. This review aims to provide comprehensive information on pharmacological studies of esculin and esculetin, which is of noteworthy importance in exploring the therapeutic potential of both coumarin compounds.

Characterization of the horse chestnut genome reveals the evolution of aescin and aesculin biosynthesis.

Horse chestnut (Aesculus chinensis) is an important medicinal tree that contains various bioactive compounds, such as aescin, barrigenol-type triterpenoid saponins (BAT), and aesculin, a glycosylated coumarin. Herein, we report a 470.02 Mb genome assembly and characterize an Aesculus-specific whole-genome duplication event, which leads to the formation and duplication of two triterpenoid biosynthesis-related gene clusters (BGCs). We also show that AcOCS6, AcCYP716A278, AcCYP716A275, and AcCSL1 genes within these two BGCs along with a seed-specific expressed AcBAHD6 are responsible for the formation of aescin. Furthermore, we identify seven Aesculus-originated coumarin glycoside biosynthetic genes and achieve the de novo synthesis of aesculin in E. coli. Collinearity analysis shows that the collinear BGC segments can be traced back to early-diverging angiosperms, and the essential gene-encoding enzymes necessary for BAT biosynthesis are recruited before the splitting of Aesculus, Acer, and Xanthoceras. These findings provide insight on the evolution of gene clusters associated with medicinal tree metabolites.

Tracing the opposing assimilate and nutrient flows in live conifer needles.

The vasculature along conifer needles is fundamentally different from that in angiosperm leaves as it contains a unique transfusion tissue inside the bundle sheath. In this study, we used specific tracers to identify the pathway of photoassimilates from mesophyll to phloem, and the opposing pathway of nutrients from xylem to mesophyll. For symplasmic transport we applied esculin to the tip of attached pine needles and followed its movement down the phloem. For apoplasmic transport we let detached needles take up a membrane-impermeable contrast agent and used micro-X-ray computed tomography to map critical water exchange interfaces and domain borders. Microscopy and segmentation of the X-ray data enabled us to render and quantify the functional 3D structure of the water-filled apoplasm and the complementary symplasmic domain. The transfusion tracheid system formed a sponge-like apoplasmic domain that was blocked at the bundle sheath. Transfusion parenchyma cell chains bridged this domain as tortuous symplasmic pathways with strong local anisotropy which, as evidenced by the accumulation of esculin, pointed to the phloem flanks as the preferred phloem-loading path. Simple estimates supported a pivotal role of the bundle sheath, showing that a bidirectional movement of nutrient ions and assimilates is feasible and emphasizing the role of the bundle sheath in nutrient and assimilate exchange.

[Rapid determination of aesculin and aesculetin in Fraxini Cortex by high performance liquid chromatography-ultraviolet at equal absorption wavelength].

Fraxini Cortex is a traditional Chinese herbal medicine that has been used for thousands of years to treat dampness-heat diarrhea, dysentery, red or white vaginal discharge, painful swelling or redness of the eyes, and nebula. It contains various chemical components, including coumarins, iridoids, phenolic acids, and flavonoids. Coumarins are important active ingredients in Fraxini Cortex and have antibacterial, anti-inflammatory, antioxidant, antitumor, and antiviral activities. Aesculin and aesculetin are two major coumarin components of Fraxini Cortex that are widely used in its quality evaluation. Previous HPLC methods for determination of aesculin and aesculetin present several limitations, such as long analysis times and high solvent and reference compound consumption. In this study, a rapid, eco-friendly and cost saving HPLC method for the determination of aesculin and aesculetin in Fraxini Cortex was established by using the core-shell column and equal absorption wavelength (EAW). Different factors influencing the extraction process, such as the extraction solvent, temperature, and time, were assessed to obtain the optimal extraction conditions. The results showed that Fraxini Cortex samples could be well extracted by ultrasonic extraction for 5 min with a 25% ethanol aqueous solution. A core-shell column was used, and different mobile phases and flow rates were investigated to obtain the best rapid-HPLC separation conditions. The optimized HPLC conditions were as follows: a Poroshell 120 EC-C18 column (50 mm×4.6 mm, 2.7 µm), acetonitrile-0.1% formic acid aqueous solution (6∶94, v/v) as the eluent, a flow rate of 1.5 mL/min, and a column temperature of 25 ℃. The EAW of aesculin and aesculetin was a key factor in their determination using a single reference compound. EAW selection was performed in two steps. First, the UV spectra of two equimolar concentrations of the reference compounds (aesculin and aesculetin) were compared to determine the EAW of the two analytes. The EAW results were then verified by the HPLC analysis of the reference compound solutions. The final EAW of aesculin and aesculetin was 341 nm. The determination of aesculin and aesculetin using only one reference compound (i. e., aesculin) was achieved by HPLC-UV at this EAW. The newly developed HPLC method revealed a good linear relationship between the two target analytes (r=1.0000). The limits of detection (LODs) and limits of quantification (LOQs) were 1.5 µmol/L and 3.0 µmol/L, respectively, and the average recoveries of aesculin and aesculetin were 99.0% and 97.5%. The stabilities of the sample solutions were examined, and the two analytes demonstrated good stability for 24 h. The contents of the target analytes in 10 batches of Fraxini Cortex were determined using the proposed EAW method and the classic external standard method (ESM), and comparable concentrations were obtained. The contents of aesculin and aesculetin in the 10 batches of Fraxini Cortex were 0.26%-2.80% and 0.11%-1.47%, respectively. A t-test was conducted to compare the results of the proposed EAW technique with those obtained via the method reported in the Chinese Pharmacopoeia, and no significant difference between the two assay methods was noted (P>0.05). Comparison of the newly established EAW method with those reported in the literature revealed that our method required only 10 min to complete and used as little as 0.5 mL of the solvent and only one standard. Therefore, the developed EAW method is a rapid, simple, eco-friendly, and cost-effective analytical method that is suitable for the determination of aesculin and aesculetin in Fraxini Cortex and its related products. The proposed technique is an improved method for determining aesculin and aesculetin and contributes to the enhancement of the quality evaluation of Fraxini Cortex.

Multi-omics reveal key enzymes involved in the formation of phenylpropanoid glucosides in Artemisia annua.

Although mainly known for producing artemisinin, Artemisia annua is enriched in phenylpropanoid glucosides (PGs) with significant bioactivities. However, the biosynthesis of A. annua PGs is insufficiently investigated. Different A. annua ecotypes from distinct growing environments accumulate varying amounts of metabolites, including artemisinin and PGs such as scopolin. UDP-glucose:phenylpropanoid glucosyltransferases (UGTs) transfers glucose from UDP-glucose in PG biosynthesis. Here, we found that the low-artemisinin ecotype GS produces a higher amount of scopolin, compared to the high-artemisinin ecotype HN. By combining transcriptome and proteome analyses, we selected 28 candidate AaUGTs from 177 annotated AaUGTs. Using AlphaFold structural prediction and molecular docking, we determined the binding affinities of 16 AaUGTs. Seven of the AaUGTs enzymatically glycosylated phenylpropanoids. AaUGT25 converted scopoletin to scopolin and esculetin to esculin. The lack of accumulation of esculin in the leaf and the high catalytic efficiency of AaUGT25 on esculetin suggest that esculetin is methylated to scopoletin, the precursor of scopolin. We also discovered that AaOMT1, a previously uncharacterized O-methyltransferase, converts esculetin to scopoletin, suggesting an alternative route for producing scopoletin, which contributes to the high-level accumulation of scopolin in A. annua leaves. AaUGT1 and AaUGT25 responded to induction of stress-related phytohormones, implying the involvement of PGs in stress responses.

The potential of esculin in ameliorating Type-2 diabetes mellitus induced neuropathy in Wistar rats and probing its inhibitory mechanism of insulin aggregation.

Diabetic neuropathy encompasses multiple pathological disturbances, many of which coincide with the pathophysiological mechanisms of neurodegenerative disorders. In the present study, various biophysical techniques like Rayleigh light scattering assay, Thioflavin T assay, far-UV Circular Dichroism spectroscopy, Transmission electron microscopy have unveiled the anti-fibrillatory effect of esculin upon human insulin fibrillation. MTT cytotoxicity assay demonstrated the biocompatibility of esculin and in-vivo studies such as behavioral tests like hot plate test, tail immersion test, acetone drop test, plantar test were performed for validating diabetic neuropathy. Assessment of levels of serum biochemical parameters, oxidative stress parameters, pro-inflammatory cytokines as well as neuron specific markers was done in the current study. Rat brains were subjected to histopathology and their sciatic nerves were subjected to transmission electron microscopy to analyze myelin structure alterations. All these results reveal that esculin ameliorates diabetic neuropathy in experimental diabetic rats. Conclusively, our study demonstrates the anti-amyloidogenic potential of esculin in the form of inhibition of human insulin fibrillation, making it a promising candidate in combating neurodegenerative disorders in the near future and the results of various behavioral, biochemical, and molecular studies reveal that esculin possesses anti-lipidemic, anti-inflammatory, anti-oxidative and neuroprotective properties which help in ameliorating diabetic neuropathy in streptozotocin induced diabetic Wistar rats.

Esculin alleviates LPS-induced acute lung injury via inhibiting neutrophil recruitment and migration.

OBJECTIVES: Acute lung injury (ALI) poses a serious threat to human health globally, particularly with the Coronavirus 2019 (COVID-19) pandemic. Excessive recruitment and infiltration of neutrophils is the major etiopathogenesis of ALI. Esculin, also known as 6,7-dihydroxycoumarin, is a remarkable compound derived from traditional Chinese medicine Cortex fraxini. Accumulated evidence indicates that esculin has potent anti-inflammatory effects, but its pharmaceutical effect against ALI and potential mechanisms are still unclear. METHODS: This study evaluated the protective effect of esculin against ALI by histopathological observation and biochemical analysis of lung tissues and bronchoalveolar lavage fluid (BALF) in lipopolysaccharide (LPS)-challenged ALI mice in vivo. The effects of esculin on N-formyl-met-leu-phe (fMLP)-induced neutrophil migration and chemotaxis were quantitatively assessed using a Transwell assay and an automated cell imaging system equipped with a Zigmond chamber, respectively. The drug affinity responsive target stability (DARTS) assay, in vitro protein binding assay and molecular docking were performed to identify the potential therapeutic target of esculin and the potential binding sites and pattern. RESULTS: Esculin significantly attenuated LPS-induced lung pathological injury, reduced the levels of pro-inflammatory cytokines in both BALF and lung, and suppressed the activation of NF-κB signaling. Esculin also significantly reduced the number of total cells and neutrophils as well as myeloperoxidase (MPO) activity in the BALF. Esculin impaired neutrophil migration and chemotaxis as evidenced by the reduced migration distance and velocity. Furthermore, esculin remarkably inhibited Vav1 phosphorylation, suppressed Rac1 activation and the PAK1/LIMK1/cofilin signaling axis. Mechanistically, esculin could interact with ß2 integrin and then diminish its ligand affinity with intercellular adhesion molecule-1 (ICAM-1). CONCLUSIONS: Esculin inhibits ß2 integrin-dependent neutrophil migration and chemotaxis, blocks the cytoskeletal remodeling process required for neutrophil recruitment, thereby contributing to its protective effect against ALI. This study demonstrates the new therapeutic potential of esculin as a novel lead compound.

Esculin protects human blood cells from bioallethrin-induced toxicity: An ex vivo study.

Bioallethrin, a household insecticide, is a member of the pyrethroid family and is known for its adverse effects on human health. Human exposure to pyrethroids is unavoidable due to their widespread use in controlling several fatal vector-borne diseases, mostly in developing nations. Bioallethrin is known to induce oxidative stress in target cells, including erythrocytes. Here we have studied the protective effect of dietary antioxidant esculin on bioallethrin-induced damage in isolated human erythrocytes. The cells were incubated with 200 µM bioallethrin, without or with different concentrations of esculin (200, 400 and 600 µM), and the results compared to the untreated control samples. Bioallethrin-treated erythrocytes showed a significant increase in oxidative stress markers, like protein and lipid oxidation, accompanied by decrease in free amino groups and ratio of reduced to oxidized glutathione. There was enhanced generation of reactive oxygen and nitrogen species with changes in plasma membrane integrity. Bioallethrin oxidized hemoglobin to methemoglobin, which cannot transport oxygen. It altered the activities of antioxidant enzymes and lowered the electron donating and free radical quenching ability of erythrocytes. The cell morphology and redox system of erythrocyte membrane were also altered by bioallethrin. Treatment with esculin, prior to incubation with bioallethrin, led to significant restoration in all these parameters in an esculin concentration-dependent manner. Thus esculin attenuated the biolletherin-induced oxidative damage to erythrocytes. Esculin can, therefore, be an effective chemoprotectant against xenobiotic-induced toxicity in human erythrocytes.

Molecular interactions of esculin with bovine serum albumin and recognition of binding sites with spectroscopy and molecular docking.

Esculin is structurally a hydroxycoumarin found in various medicinal plants. This study investigates the binding mode of esculin with bovine serum albumin by employing numerous spectroscopic studies and molecular docking approaches. Ultraviolet absorption spectroscopy revealed ground state complex formation between esculin and bovine serum albumin. At the same time, steady-state fluorescence studies showed quenching in the fluorescence emission spectra of BSA in the presence of esculin. To get insight into the location of the binding pocket of esculin on BSA, warfarin and ibuprofen site markers were used. Competitive site marker displacement assay revealed that esculin binds to Sudlow's site I (subdomain IIA) in bovine serum albumin. Thermodynamic parameters suggested that hydrogen bonding and van der Waals interaction stabilizes the esculin-BSA complex. Förster's non-radiation energy transfer analysis described the high propensity of energy transfer between bovine serum albumin and esculin. The molecular docking approach facilitated locating the binding pocket, amino acid residues involved, types of interacting forces, and binding energy (ΔG) between esculin and BSA. Circular dichroism revealed the effect of the binding of esculin on the secondary structure and helped understand the thermal unfolding profile of BSA in the presence of esculin.Communicated by Ramaswamy H. Sarm.

A study on the catalytic activity of polypeptides toward the hydrolysis of glucoside compounds gastrodin, polydatin and esculin.

The self-assembly of a series of catalytically active polypeptides toward hydrolysis of glucoside compounds, namely, gastrodin, polydatin and esculin was investigated. These active peptides are composed of two functional fragments: one is the hydrophobic sequence LHLHLRL, which forms assembling segments in the presence of Zn ions (Zn2+); another functional sequence of active peptides are catalytic sites such as Glu (E), Asp (D) and His (H), where carboxylic acids (-COOH) or imidazole groups act like scissors to cleave glucoside bonds of the compounds (according to the acid-base coupling mechanism). The effects of the amino acid sequence of the peptide, Zn2+ concentration, pH and the size or steric hindrance of glucoside compounds on the hydrolytic activity were studied. It was found that the crystalline structure of assembled peptides was crucial to provide the peptide with catalytic hydrolytic activity. Noncovalent interaction index was used to analyse the noncovalent interaction of PEs with glucoside compounds, including hydrogen bonds, van der Waals, and steric effect in the complexes. The binding energy of complexes, the direction and site of nucleophilic attack during deglycosylation processes were also investigated by molecular docking and the electron density Laplace function. This revealed that the differences in the hydrolytic activity of peptides toward glucoside compounds with different sizes originated from different hydrogen bond interactions between the peptides and substrates. These active peptides may find application in the preparation of drugs by de-glycosylation of natural compounds.

Ideonella alba sp. nov. and Ideonella aquatica sp. nov. isolated from an aquaculture farm.

The three novel bacterial strains designated as 3Y2T, 4Y16 and 4Y11T were isolated from an aquaculture farm and characterized using a polyphasic taxonomic approach. These strains were determined to be catalase- and oxidase-positive and to hydrolyze gelatin and aesculin. The results of 16S rRNA gene-based phylogenetic analysis indicated that the three strains were related to members of the genus Ideonella. The phylogenomic results further indicated that the three strains formed two independent branches distinct from reference type strains within this genus. The digital DNA-DNA hybridization (dDDH), average nucleotide identity (ANI) and average amino acid identity (AAI) values between the three strains and their relatives were far below the thresholds of 70 % dDDH, 95-96 % ANI and 95 % AAI for species definition, respectively, indicating that the three strains represent two novel genospecies. The results of chemotaxonomic characterization indicated that the major cellular fatty acids of the three strains were summed feature 3 (C16 : 1ω6c and/or C16 : 1 ω7c) and C16 : 0; the common main polar lipids were diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol; the respiratory quinone was ubiquinone-8. The genomic DNA G+C contents of the three strains were 70.2, 70.1 and 69.7%, respectively. On the basis of the different genotypes and distinctive phenotypes such as the phosphatidylcholine and glycolipid only in 3Y2T and the utilization of malic acid and trisodium citrate only in 4Y11T, strains 3Y2T and 4Y11T are concluded to represent two novel species of the genus Ideonella, for which the names Ideonella alba sp. nov. (type strain 3Y2T = GDMCC 1.2584T = KCTC 82813T) and Ideonella aquatica sp. nov. (type strain 4Y11T = GDMCC 1.1935T = JCM 34285T) are proposed.

[Effect of Wei's triple nine needling on eye regulation in patients with presbyopia complicated with visual fatigue of liver depression and spleen deficiency].

OBJECTIVE: To compare the clinical efficacy between Wei's triple nine needling combined with esculin and digitalis glycosides eye drops and esculin and digitalis glycosides eye drops alone for presbyopia complicated with visual fatigue of liver depression and spleen deficiency. METHODS: Forty-six cases (92 eyes) with presbyopia complicated with visual fatigue of liver depression and spleen deficiency were randomly divided into an observation group (23 cases) and a control group (23 cases, 2 cases dropped off). The cases in the observation group were treated with Wei's triple nine needling and esculin and digitalis glycosides eye drops. The acupoints included Shangming (Extra), Chengqi (ST 1), Cuanzhu (BL 2) to Jingming (BL 1), Sizhukong (TE 23) to Taiyang (EX-HN 5), etc; the needling was given once every other day, three times a week, and the eye drops were given one drop each time, three times a day. The cases in the control group were only treated with the eye drops. Both groups were treated for 7 days as one course of treatment, and 2 courses of treatment were given. The visual fatigue core symptoms score, adjustment amplitude, adjustment lag and best average corrected visual acuity were observed in the two groups before treatment, 1 week and 2 weeks into treatment, respectively. RESULTS: Compared before treatment, the visual fatigue core symptoms scores in the two groups were decreased after 1-week and 2-week treatment (P<0.05); in the observation group, the adjustment amplitude was increased after 2-week treatment (P<0.05), while in the control group, the adjustment amplitude was increased after 1-week and 2-week treatment (P<0.05); in the observation group, the adjustment lag was decreased after 1-week and 2-week treatment (P<0.05). After 2-week treatment, the visual fatigue core symptoms score in the observation group was lower than that in the control group, and the adjustment amplitude was higher than that in the control group (P<0.05). There were no significant differences in adjustment lag and best average corrected visual acuity between the two groups after 1-week and 2-week treatment (P>0.05). CONCLUSION: Wei's triple nine needling combined with esculin and digitalis glycosides eye drops could improve the visual fatigue and eye regulation ability in patients with presbyopia complicated with visual fatigue of liver depression and spleen deficiency, and the effect is better than esculin and digitalis glycosides eye drops alone.

An improved selective/differential medium for culturing the Bacteroides fragilis group from wastewater.

The aim of this study was to develop an effective selective/differential medium for culturing environmental strains of the Bacteroides fragilis group (BFG). This goal was achieved by modifying standard commercial Bacteroides Bile Esculin Agar (BBE Agar). Bacteroides Bile Esculin Agar was combined with substances that inhibit the growth of non-BFG bacteria, mostly Klebsiella pneumoniae and Fusobacterium mortiferum. The strains isolated from standard and modified BBE Agar were identified as BFG strains by PCR and 16S rRNA gene sequencing. The supplementation of standard BBE Agar with colistin (40 mg L-1), kanamycin (400 mg L-1) and vancomycin (7.5 mg L-1) increases the effectiveness of BFG bacteria isolation from <10% to 35%, and additional Gram staining improves the effectiveness of bacterial isolation five-fold relative to standard BBE Agar. The results of the present study also suggest that the presence of the bfr gene is not a reliable indicator for the identification of BFG strains.

The pharmacological and pharmacokinetic properties of esculin: A comprehensive review.

Cortex fraxini is a widely used traditional Chinese medicine. Esculin is one of the main active ingredients of Cortex fraxini and has attracted more and more attention from scholars. The purpose of the review is to systematically review relevant studies on the pharmacological effects and pharmacokinetic characteristics of esculin to support its further application as therapeutic agents. Pharmacological studies have shown that the anti-inflammatory and anti-oxidative stress effects of esculin are outstanding. This indicates that esculin is promising to be used to treat a variety of diseases closely related to inflammation and oxidative damage. Esculin has anti-diabetic effect, which is closely related to improving pancreas damage, promoting insulin release, and enhancing glucose homeostasis. In addition, esculin has anti-cancer, antibiosis, anti-virus, neuroprotection, anti-thrombosis and treating eye diseases properties. Pharmacokinetic studies show that esculin can be quickly and evenly distributed in the body. However, the first pass effect of esculin is serious. In short, esculin is promising to treat many diseases, but further high quality studies are needed to firmly establish the clinical efficacy of esculin.

Co-treatment with Esculin and erythropoietin protects against renal ischemia-reperfusion injury via P2X7 receptor inhibition and PI3K/Akt activation.

Renal ischemia/reperfusion (RI/R) is a critical clinical outcome with slightly reported improvement in mortality and morbidity. Effective therapies are still crucially required. Accordingly, the therapeutic effects of esculin (ESC, LCESI-MS/MS-isolated compound from Vachellia farnesiana flowers extract, with reported P2X7 receptor inhibitor activity) alone and in combination with erythropoietin (EPO) were investigated against RI/R injury and the possible underlying mechanisms were delineated. ESC and EPO were administered for 7 days and 30 min prior to RI, respectively. Twenty-four hour following reperfusion, blood and kidney samples were collected. Results revealed that pretreatment with either ESC or EPO reduced serum nephrotoxicity indices, renal oxidative stress, inflammatory, and apoptosis markers. They also ameliorated the renal histopathological injury on both endothelial and tubular epithelial levels. Notably, ESC markedly inhibited P2X7 receptors and NLRP3 inflammasome signaling (downregulated NLRP3 and Caspase-1 gene expressions), whereas EPO significantly upregulated PI3K and Akt gene expressions, also p-PI3K and p-Akt levels in renal tissues. ESC, for the first time, demonstrated effective protection against RI/R-injury and its combination with EPO exerted maximal renoprotection when compared to each monotherapy, thereby representing an effective therapeutic approach via inhibiting oxidative stress, inflammation, renal tubular and endothelial injury, apoptosis, and P2X7 receptors expression, and activating PI3K/Akt pathway.