Animal-derived food allergen: A review on the available crystal structure and new insights into structural epitope.

Immunoglobulin E (IgE)-mediated food allergy is a rapidly growing public health problem. The interaction between allergens and IgE is at the core of the allergic response. One of the best ways to understand this interaction is through structural characterization. This review focuses on animal-derived food allergens, overviews allergen structures determined by X-ray crystallography, presents an update on IgE conformational epitopes, and explores the structural features of these epitopes. The structural determinants of allergenicity and cross-reactivity are also discussed. Animal-derived food allergens are classified into limited protein families according to structural features, with the calcium-binding protein and actin-binding protein families dominating. Progress in epitope characterization has provided useful information on the structural properties of the IgE recognition region. The data reveals that epitopes are located in relatively protruding areas with negative surface electrostatic potential. Ligand binding and disulfide bonds are two intrinsic characteristics that influence protein structure and impact allergenicity. Shared structures, local motifs, and shared epitopes are factors that lead to cross-reactivity. The structural properties of epitope regions and structural determinants of allergenicity and cross-reactivity may provide directions for the prevention, diagnosis, and treatment of food allergies. Experimentally determined structure, especially that of antigen-antibody complexes, remains limited, and the identification of epitopes continues to be a bottleneck in the study of animal-derived food allergens. A combination of traditional immunological techniques and emerging bioinformatics technology will revolutionize how protein interactions are characterized.

Phytochemical compositions of extract from peel of hawthorn fruit, and its antioxidant capacity, cell growth inhibition, and acetylcholinesterase inhibitory activity.

BACKGROUND: Hawthorn fruit (HF) is a well-known traditional medicine in China with the effects of improving digestion and regulating qi-flowing for removing blood stasis. Modern pharmacological experiments showed that HF extract has various pharmaceutical properties and flavonoids are considered as the main bioactive compounds. In this paper, Diaion HP-20 adsorption chromatography was used to enrich flavonoids in PHF, and the phytochemical composition of EPHF was analyzed by high performance liquid chromatography (HPLC) and liquid chromatography tandem mass spectrometry (LC-MS). In addition, EPHF's antioxidant capacity, acetylcholinesterase (AChE) inhibitory activity and cytotoxic activity were evaluated. METHODS: EPHF was obtained by Diaion HP-20 adsorption chromatography. Phytochemical composition of EPHF was analyzed qualitatively and quantitatively using HPLC and LC-MS. Radical scavenging capacity of EPHF was estimated using 2,2-diphenyl-1-picryhydrazyl (DPPH) assay and oxygen radical absorbance capacity (ORAC) assay. The AChE inhibitory activity of EPHF was evaluated by Ellman method. Cytotoxic activity of EPHF was assessed by means of MTT assay. RESULTS: Eight kinds of components were identified, in which ideain with the value of 179.4 mg/g was identified to be present in the highest level in EPHF, followed by (-)-epicatechin, chlorogenic acid, cyanidin 3-arabinoside, hyperoside and isoquercitrin at the concentrations of 40.9, 10.0, 1.4, 0.4 and 0.2 mg/g, respectively. The contents of these compounds in EPHF were much higher than those in PHF and HF. In addition, EPHF exhibited strong antioxidant and AChE inhibitory activity (ORAC value: 11.65 ± 2.37 µM Trolox equivalents (TE)/mg, DPPH IC50 value: 6.72 µg/mL, anti-AChE activity IC50 value: 11.72 µg/mL) compared with PHF and HF. Moreover, EPHF exhibited high levels of cytotoxicity on MCF-7 and SKOV-3 human tumour cell lines in a dose-dependent manner with the IC50 of 2.76 and 80.11 µg/mL, respectively. CONCLUSIONS: Macroporous resin is useful for the extraction and separation of the total flavonoids from PHF. The contents of flavonoids especially anthocyanin in EPHF were increased significantly compared with the PHF, and EPHF exhibited strong antioxidant, AChE inhibitory activity and cytotoxicity on human tumour cells.

Application of NIR and MIR spectroscopy for rapid determination of antioxidant activity of Radix Scutellariae from different geographical regions.

INTRODUCTION: The beneficial health effects of traditional Chinese medicines are often attributed to their potent antioxidant activities, usually established in vitro. However, these wet chemical methods for determining antioxidant activities are time-consuming, laborious, and expensive. OBJECTIVES: This study was conducted to establish a rapid determination of antioxidant activity of Radix Scutellariae using near-infrared (NIR) and mid-infrared (MIR) spectroscopy. MATERIAL AND METHODS: Antioxidant capabilities were evaluated using 2,2-diphenyl-1-picrylhydrazyl hydrazyl (DPPH) and oxygen radical absorbance capacity (ORAC) assays. The total flavonoid contents (TFCs) of Radix Scutellariae were measured by the aluminium chloride colorimetric method. The same sample was then scanned using NIR and MIR spectroscopy. Chemometrics analysis using partial least-squares (PLS) regression was performed to establish the models for predicting the antioxidant activities of Radix Scutellariae. RESULTS: A better predictive performance was achieved using PLS models based on NIR data. The determination coefficient (R(2)) and the residual predictive deviation (RPD) for the validation set were 0.9298 and 2.84 for DPPH, and 0.9436 and 2.66 for TFCs, respectively. MIR-PLS algorithms gave a slightly lower reliability (R(2) = 0.9090 and 0.9374, RPD = 2.01 and 2.42, for DPPH and TFC, respectively). Very comparable results for ORAC were obtained with the two methods. CONCLUSION: The developed spectroscopic method can be successfully applied in high-throughput screening of the antioxidant capability of Radix Scutellariae samples. It can also be a viable and advantageous alternative to laborious chemical procedures.

[Optimization of Aqueous Two-Phase Extraction of Polysaccharide from Schisandra chinensis Using Response Surface Methodology with Box-Behnken Design].

Objective: To optimize the extraction condition of polysaccharide from Schisandra chinensis. Aqueous two-phase extraction（ ATPE） method was used, based on Box-Behnken design with Response surface methodology（ BBD-RSM）. Methods: solvent volume,K2HPO4 and PEG6000 were selected as the investigation factors by the single-factor experiment, and the overall desirability（ OD）of phase volume ratio, partition coefficient and extraction rates were the reponse value. BBD-RSM was used to optimize the extraction process. . Results: The optimal parameters were as follows,the solvent volume was 5 m L,the addition amount of K2HPO4 was 1. 0 g, the addition amount of PEG6000 was 1. 8 g and centrifugation time was 9 min, which indicated that the model had a good predictability. The predicted value was 0. 950,and the deviation between observed and predicted values was 3. 94% Conclusion: The ATPE technology is easy to operate and cost-effective for the extraction of polysaccharides from Schisandra chinensis.

[Study on the mechanism of haitongpi prescription extract ointment in the treatment of knee osteoarthritis based on transcriptome].

OBJECTIVE: To explore the mechanism of Haitongpi Prescription extract in the treatment of knee osteoarthritis based on transcriptome. METHODS: Total of 12 SPF grade rats were divided into control group(group C), model group(group M), and Haitongpi prescription group(group HP). The knee osteoarthritis rat model was established using the Panicker method for group M and group HP, and group HP was intervened by local topical application of Haitongpi Prescription extract for 4 weeks. Total RNA from mouse knee cartilage was extracted and three sets of differential genes were obtained through sequencing.Differential genes were prediction and analysis through GO function and KEGG pathway enrichment analysis. RESULTS: A total of 109 differentially expressed genes were identified in Group C versus Group M, while 118 differentially expressed genes were identified in Group M versus Group HP, resulting in a total of 28 genes. GO functional enrichment analysis showed that the mechanism of HP extract in treating knee osteoarthritis mainly involved immunoglobulin mediated immune response, immunoglobulin complexes, and antigen binding; KEGG pathway enrichment analysis showed correlation with tumor necrosis factor (TNF) signaling pathway, interleukin 17(IL-17) signaling pathway, and estrogen signaling pathway. CONCLUSION: HP extract can exert therapeutic effects on knee osteoarthritis through mechanisms such as immunoglobulin mediated immune response, immunoglobulin complexes, and antigen binding, as well as signaling pathways such as TNF signaling pathway, IL-17 signaling pathway, and estrogen signaling pathway.

Mechanism of icariin for the treatment of osteoarthritis based on network pharmacology and molecular docking method.

BACKGROUND: Icarin's mechanism of action in osteoarthritis (OA) was explored using network pharmacology and the GEO database, and then further validated using molecular docking. METHODS: GEO database using network pharmacology identified differential genes in OA based on Icariin's possible targets predicted by pharmmapper database. Combining the differentially expressed genes in OA with the OA-related targets, the overlapping targets were removed. In order to determine what Icariin's core targets are for treating OA, PPI network analysis was performed using OA-related targets and possible Icariin targets. Furthermore, molecular docking was used to verify the chemical's binding to the targets. Final steps included Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. Cytoscape was used to construct a network of compound-target-pathway-disease. RESULTS: Protein-protein interactions between overlapping targets revealed 151 intersection targets based on a network analysis. The top ten targets with the highest enrichment scores were SRC, MAPK1, HSP90AA1, AKT1, PTPN11, ESR1, EGFR, RhoA, JAK2, and MAPK14. KEGG enrichment analysis showed that the pathways at which Icariin intervention occurs include the OA including FOXO signaling pathway, and estrogen signaling pathway. The GO analysis result showed that various biologic processes such as proteolysis, angiogenesis, innate immune response, and positive regulation of inflammatory response were involved in treatment. Molecular docking analysis confirmed that Icariin could bind well to the targets through intermolecular forces. CONCLUSION: With its multi-targeting and multi-pathway characteristics, Icariin is a promising candidate drug for treating OA.

Immunoglobulin E Epitope Mapping and Structure-Allergenicity Relationship Analysis of Crab Allergen Scy p 9.

Filamin C is an allergen of Scylla paramamosain (Scy p 9), and six IgE linear epitopes of the allergenic predominant region had previously been validated. However, the IgE epitope and structure-allergenicity relationship of Scy p 9 are unclear. In this study, a hydrophobic bond was found to be an important factor of conformation maintaining. The critical amino acids in the six predicted conformational epitopes were mutated, and the IgE-binding capacity and surface hydrophobicity of four mutants (E216A, T270A, Y699A, and V704A) were reduced compared to Scy p 9. Ten linear epitopes were verified with synthetic peptides, among which L-AA187-205 had the strongest IgE-binding capacity. In addition, IgE epitopes were mapped in the protruding surface of the tertiary structure, which were conducive to binding with IgE and exhibited high conservation among filamin genes. Overall, these data provided a basis for IgE epitope mapping and structure-allergenicity relationship of Scy p 9.

Optimisation of infrared-assisted extraction of rutin from crude Flos Sophorae Immaturus using response surface methodology and HPLC analysis.

INTRODUCTION: Rutin, one of main constituents in Flos Sophorae Immaturus, has been proven to possess several pharmacological properties such as anti-oxidant, anti-platelet, anti-inflammatory effects and so on. However, optimisation of the extraction of rutin from Flos Sophorae Immaturus has rarely been reported. Thus, it is important to develop an effective method to obtain maximum yields of rutin from Flos Sophorae Immaturus. OBJECTIVE: To develop an infrared-assisted extraction method for maximum rutin yield from crude Flos Sophorae Immaturus using response surface methodology and HPLC analysis. METHODOLOGY: Through single factor experiments, ranges of the main variables (including methanol concentration, liquid:solid ratio, extraction time and infrared power) affecting the extraction yield of rutin were confirmed. A Box-Behnken design consisting of 24 experimental runs and five replicates at zero point was then applied and a regression model was obtained to predict the optimal extraction yield. RESULTS: The ANOVA analysis indicated that the regression equation fits very well with the actual situation. The optimal conditions were as follows: infrared power 204.90 W, liquid:solid ratio 30.00 mL/g, methanol concentration 70.00% and extraction time 4.80 min. Under optimal conditions the predicted maximum yield (125.70 mg rutin/0.5 g raw material) was consistent with the experimental value (126.32 ± 0.67 mg rutin/0.5 g raw material) (n = 3). CONCLUSION: The application of response surface methodology was reliable and feasible in the optimisation of infrared-assisted extraction of rutin from crude Flos Sophorae Immaturus.

Molecular interaction studies of acetylcholinesterase with potential acetylcholinesterase inhibitors from the root of Rhodiola crenulata using molecular docking and isothermal titration calorimetry methods.

(-)-Epicatechin gallate ((-)-ECG), 1,2,3,4,6-O-pentagalloylglucose (PGG), rhodionin, herbacetin and rhodiosin isolated from the root of Rhodiola crenulata exhibited potent, dose-dependent inhibitory effects on acetylcholinesterase (AChE) with IC50 ranged from 57.50±5.83 to 2.43±0.34µg/mL. With the aim of explaining the differences in activity of these active ingredients and clarifying how they inhibit AChE, the AChE-inhibitor interactions were further explored using molecular docking and isothermal titration calorimetry (ITC) methods in the present study. Molecular docking studies revealed that all compounds except PGG showed binding energy values ranging from -10.30 to -8.00kcal/mol while the binding energy of galantamine, a known AChE inhibitor, was -9.53kcal/mol; they inhibited the AChE by binding into the ligand pocket with the similar binding pattern to that of galantamine by interacting with Glu199 of AChE. Inhibition constant of these active ingredients had a positive correlation with binding energy. The interaction between AChE and PGG was further evaluated with the ITC method and the results indicated that the PGG-AChE interaction was relevant to AChE concentration. The results revealed a possible mechanism for the AChE inhibition activity of these bioactive ingredients, which may provide some help in lead compounds optimization in the future.

Global profiling and rapid matching of natural products using diagnostic product ion network and in silico analogue database: Gastrodia elata as a case study.

Rapid discovery of novel compounds of a traditional herbal medicine is of vital significance for pharmaceutical industry and plant metabolic pathway analysis. However, discovery of unknown or trace natural products is an ongoing challenge. This study presents a universal targeted data-independent acquisition and mining strategy to globally profile and effectively match novel natural product analogues from an herbal extract. The famous medical plant Gastrodia elata was selected as an example. This strategy consists of three steps: (i) acquisition of accurate parent and adduct ions (PAIs) and the product ions data of all eluting compounds by untargeted full-scan MS(E) mode; (ii) rapid compound screening using diagnostic product ions (DPIs) network and in silico analogue database with SUMPRODUCT function to find novel candidates; and (iii) identification and isomerism discrimination of multiple types of compounds using ClogP and ions fragment behavior analyses. Using above data mining methods, a total of 152 compounds were characterized, and 70 were discovered for the first time, including series of phospholipids and novel gastroxyl derivatives. Furthermore, a number of gastronucleosides and phase II metabolites of gastrodin and parishins were discovered, including glutathionylated, cysteinylglycinated and cysteinated compounds, and phosphatidylserine analogues. This study extended the application of classical DPIs filter strategy and developed a structure-based screening approach with the potential for significant increase of efficiency for discovery and identification of trace novel natural products.

Restricted access magnetic core-mesoporous shell microspheres with C8-modified interior pore-walls for the determination of diazepam in rat plasma by LC-MS.

In this study, a novel enrichment technique based on magnetic core-mesoporous shell microspheres with C8-modified interior pore-walls (C8-Fe3O4@mSiO2) was successfully developed for the determination of diazepam in rat plasma by LC-MS. Due to the unique properties of the synthesized C8-Fe3O4@mSiO2 microspheres (C8-modified magnetic mesoporous microsphere), small drug molecules like diazepam can enter the mesopore channels and be efficiently absorbed through hydrophobic interaction by interior C8-groups (Octyl functional groups). Large molecules like proteins are excluded from the mesopore channels as a result of size exclusion effect, leading to direct extraction of drug molecules from protein-rich biosmaples such as plasma without any other pretreatment procedure. Moreover, diazepam adsorbed C8-Fe3O4@mSiO2 microspheres could be simply and rapidly isolated through placing a magnet on the bottom of container, and then diazepam could be easily eluted from C8-Fe3O4@mSiO2 microspheres for further LC-MS analysis. Extraction conditions such as amounts of C8-Fe3O4@mSiO2 microspheres added, adsorption time, elution solvent and elution time were investigated. Method validations including linear range, the limit of detection, precision, and recovery were also studied. The results indicated that the proposed method based on C8-Fe3O4@mSiO2 microspheres was simple and accurate for the analysis of diazepam in the rat plasma. And it will provide new ideas for analyzing plasma concentration and pharmacokinetics of similar drugs.

The successful treatment of flat warts with auricular acupuncture.

BACKGROUND: Flat warts are a common presenting complaint in adolescents and adults and may be a cosmetic problem as well. Patients suffering from flat warts are often unsatisfied with conventional medical care because of adverse effects such as intolerable pain, hyperpigmentation, hypopigmentation, or occasionally allergic contact dermatitis. To offset the possibilities of hyperpigmentation, hypopigmentation, and scar formation, the method of auricular acupuncture was used. METHODS: Single-blind method adopted, 60 subjects with flat warts were all outpatients and randomly allocated to a treatment group or a control group, with 30 patients in each group. Thirty subjects in the treatment group were treated with weekly auricular acupuncture for 10 weeks while the other subjects in the control group were treated with 0.1% of tretinoin ointment topically for 10 weeks. RESULTS: Sixteen subjects in the treatment group (53.33%) recovered fully from flat warts without recurrence during the ensuing six months' follow-up after 10 weeks' surgery compared with only one subject in the control group (3.33%). The therapeutic effect of the treatment group was statistically better than that of the control group by Mann-Whitney U-test with SPSS software (P < 0.01). During the treatment period and the ensuing six months' follow-up, no adverse effects were observed by the investigators or reported by patients. CONCLUSIONS: Our findings suggest that auricular acupuncture may be a viable alternative for the treatment of flat warts. Larger randomized studies are needed to fully evaluate auricular acupuncture against more conventional treatments, and these are planned.

4'-Chlorodiazepam, a translocator protein (18 kDa) antagonist, improves cardiac functional recovery during postischemia reperfusion in rats.

Inhibition of translocator protein (18 kDa) (TSPO) can effectively prevent reperfusion-induced arrhythmias and improve postischemic contractile performance. Mitochondrial permeability transition pore (mPTP) opening, mediated mainly through oxidative stress during ischemia/reperfusion (I/R), is a key event in reperfusion injury. 4'-Chlorodiazepam is a widely used TSPO antagonist. However, whether 4'-chlorodiazepam can improve cardiac functional recovery during postischemia reperfusion by affecting oxidative enzymes, reducing reactive oxygen species (ROS) and thereby inhibiting mPTP opening is still unknown. Cardiac function including heart rate, coronary flow rate, left ventricular developed pressure (LVDP), left ventricular end-diastolic pressure (LVEDP), maximal time derivatives of pressure (+/-dP/dt max) and the severity of ventricular arrhythmias were analyzed in isolated rat hearts during I/R. mPTP opening, ROS and oxidative enzyme activities were measured with fluorometric or spectrophotometric techniques. 4'-Chlorodiazepam did not affect heart rate and coronary flow rate, but abolished the increase in LVEDP, accelerated the recovery of LVDP and +/-dP/dt max, and reduced the severity of ventricular arrhythmias. The mPTP opening probability was reduced by 4'-chlorodiazepam, accompanied by a reduction in ROS level. In addition, the activities of mitochondrial electron transport chain complex I and complex III were increased, while those of xanthine oxidase and NADPH oxidase were reduced. Therefore, 4'-chlorodiazepam may improve cardiac functional recovery during reperfusion, potentially by affecting the activities of oxidative enzymes, reducing ROS and thereby inhibiting mPTP opening. The present study presents evidence that 4'-chlorodiazepam could be a novel adjunct to reperfusion.