Mecanismos preventivos de la degradación de la capa híbrida / Preventive mechanisms of the degradation of the hybrid layer

Uno de los mayores retos de la odontología actual es lograr una adhesión duradera y estable entre los materiales de restauración y los tejidos dentales. Los protocolos adhesivos han ido cambiando a lo largo del tiempo para cumplir con dicho objetivo, tratando de mantener la integridad de la capa hibrida, por lo que se revisaron los factores que provocan la degradación de la capa hibrida y los mecanismos propuestos para prevenir esta degradación. Se realizó una investigación y recopilación de información bibliográfica especializadas en el tema, en buscadores científicos como PubMed, Scielo, Redalyc, Medigraphic y Scopus. Para los criterios de inclusión se consideraron años de publicación entre el año 2002 al 2022, enfocados en trabajos de investigación relacionados con la degradación de la interfaz de unión resina-dentina y mecanismos para prevenir esta degradación en la capa híbrida. El mecanismo mas estudiado a corto y largo plazo es la aplicación de clorhexidina, la cual se utiliza después del ácido fosfórico y antes del adhesivo, inhibe la actividad proteolítica de las metaloproteinasas de la matriz (MMPs) y retarda la degradación de las fibras colágenas, consiguiendo de esta manera una mayor vida de las restauraciones adhesivas.

One of the biggest challenges in dentistry is to achieve a long-lasting and stable bond between restorative materials and dental tissues. The adhesive protocols have been changing over time to meet this objective, trying to maintain the integrity of the hybrid layer, so the factors that cause the degradation of the hybrid layer and the mechanisms proposed to prevent this degradation were reviewed. An investigation and compilation of specialized bibliographic information on the subject was carried out, in scientific search engines such as PubMed, Scielo, Redalyc, Medigraphic, Scopus, among others, as well as books. For the inclusion criteria, years of publication between 2002 and 2022 were considered, focused on research works related to the degradation of the resin-dentin bonding interface and mechanisms to prevent this degradation in the hybrid layer. The placement of CHX is used after the application of phosphoric acid and before the adhesive, it inhibits the proteolytic activity of matrix metalloproteinases (MMPs) and the degradation of collagen fibers, thus achieving a longer life of resin dental restorations.

Biomasa residual de industria del Cannabis, una alternativa para la obtención de productos de alto valor agregado

A medida que como sociedad vamos dando más importancia a lograr una economía circular, se hace importante encontrar fuentes renovables aptas para la producción de biocombustibles y bioquímicos. En los últimos años, diversas fuentes de biomasa lignocelulósica han sido estudiadas para estos propósitos. Dentro de estas fuentes de biomasa se encuentra el cáñamo (Cannabis sativa L.), siendo parte de una industria que ha crecido a pasos agigantados en las últimas décadas, en Colombia, desde su legalización. Específicamente, la industria del cannabis medicinal es responsable de generar una enorme cantidad de residuos en forma de los tallos de la planta, considerados un subproducto de bajo valor. En esta revisión se compila la información de diferentes estudios sobre el aprovechamiento de la fracción de polisacáridos de biomasa cáñamo, mediante transformaciones químicas y bioquímicas, para la obtención de productos de valor agregado. Se encontró que la mayoría de estudios están enfocados en la obtención de bioetanol o biogás; se encontraron también reportes de otras moléculas como ácido succínico, ácido láctico, furfural, polihidroxialcanoatos y bisaboleno. La viabilidad a nivel industrial de todos estos procesos permanece siendo una incógnita, pues los pasos de pretratamiento, hidrólisis y de conversión final utilizados suelen ser costosos. Es necesario que los estudios que realicen en el futuro se enfoquen en optimizar las condiciones de estos procesos y hacerlos verdes y así asegurar que puedan ser escalados.

As we as a society, give more importance to achieving a circular economy, it becomes important to find renewable sources suitable for the production of biofuels and biochemicals. In the last years, several different sources of lignocellulosic biomass have been studied for these purposes. One of these biomass sources is hemp (Cannabis sativa L), being part of an industry that has grown through giant steps in the last decades, in Colombia, since its legalization. Specifically, the industry of medicinal hemp is responsible for the generation of huge amounts of residues in the form of the plant stalks, considered a low value subproduct. This review compiles the information of several studies about the exploitation of the polysaccharide portion of hemp biomass through chemical and biochemical transformations, obtaining value-added products. It was found that most of these studies focus on the production of bioetanol or biogas; reports of other molecules such as succinic acid, furfural, polyhydroxyalkanoates and bisabolene were also found. Industrial viability of these processes remains a question, since pretreatment, hydrolysis and final conversion steps are usually expensive. It necessary that future studies focus on optimizing conditions of these processes as well as making them green, ensuring that they can be scaled.

The potential of salivary albumin to degrade composite resin / Albumina salivar: investigando seu potencial de degradar resinas compostas

ABSTRACT Albumin is a salivary enzyme capable of cleaving ester linkages and catalyzing degradation of resin-based dental materials. However, the effect of concentration-dependent esterolytic action on composite resins as yet remains unexplored. Aim: The purpose of this study was to evaluate whether artificial saliva formulations with different concentrations of albumin affected the surface roughness, flexural strength and microhardness of a composite resin. Materials and Method: Specimens (25x2x2mm) of a nanofilled composite (Filtek Z350XT, 3M/ESPE) were prepared and analyzed for average surface roughness (Ra/pm). The specimens were then allocated to 6 groups (n=30), to be treated with different salivary albumin concentrations: 0, 10, 50, 100, 200, 400 pg/mL. The specimens were stored in their respective artificial saliva groups, half of them for 24 h and the remainder for 180 days (artificial saliva renewed weekly), after which they were submitted to a new Ra reading, and tested for three-point flexural strength (FS, MPa). The specimens stored for 180 days were analyzed for Knoop microhardness (KH, Kg/mm2). Data were submitted to two-way ANOVA (Ra and FS) and one-way ANOVA (KH). Results: Although Ra increased (p < 0.001) and FS decreased (p < 0.001) from 24 hours to 180 days of storage, the albumin concentration did not significantly affectRa (p = 0.168), FS (p = 0.477) or KH (p = 0.378). Conclusion: The esterolytic action of albumin did not increase the artificial-saliva-induced hydrolytic degradation of the composite resin.

RESUMO Albumina, uma enzima encontrada na saliva, é capaz de clivar ligagoes éster e catalisar a degradando de materiais dentários á base resina. Apesar da agao esterolítica ser potencialmente concentragao-dependente, a investigando desse efeito sobre resinas compostas ainda permanence inexplorado. Objetivo: O objetivo deste estudo foi avaliar se formulagoes de saliva artificial contendo diferentes concentragoes de albumina afetariam a rugosidade superficial, a resistencia flexural e a microdureza de uma resina composta. Materials e Método: Corpos de pro-va em barra (25x2x2mm) foram confeccionados a partir de uma resina composta nanoparticulada (Filtek Z350XT, 3M/ESPE) e foram submetidos á leitura de rugosidade superficial média inicial (Ra, pm), em rugosímetro. Entao, as amostras foram divididas em 6 grupos (n=30) de acordo com a concentrando de albumina na saliva: 0, 10, 50, 100, 200, 400 pg/mL. As amostras foram armazenadas nas formulagoes de saliva artificial correspondente ao seu grupo, metade por 24 h e as demais por 180 dias (com trocas de saliva semanais). As amostras foram submetidas a no-vas leituras de rugosidade (Rafinal) e avaliadas quanto á resistencia flexural de tres pontos (RF, MPa). As amostras armazenadas por 180 dias foram avaliadas quanto á microdureza Knoop (KH, Kg/mm2). Os dados foram submetidos a análises de variáncia a dois critérios (Ra e RF) e a um critério (KH). Resultados: Apesar de haver aumento na Ra (p < 0,001) e uma diminuigao da RF (p < 0,001) de 24 h para 180 dias, a concentragao de albumina nao afetou significativamente a Ra (p = 0,168), a RF (p = 0,477) ou a KH (p = 0,378). Conclusoes: A agdo esterolítica da albumina nao aumentou a degradagao hidrolítica da resina composta induzida pela saliva artificial.

Multidisciplinary strategies to enhance therapeutic effects of flavonoids from Epimedii Folium:Integration of herbal medicine,enzyme engineering,and nanotechnology / 药物分析学报

Flavonoids such as baohuoside I and icaritin are the major active compounds in Epimedii Folium(EF)and possess excellent therapeutic effects on various diseases.Encouragingly,in 2022,icaritin soft capsules were approved to reach the market for the treatment of hepatocellular carcinoma(HCC)by National Medical Products Administration(NMPA)of China.Moreover,recent studies demonstrate that icaritin can serve as immune-modulating agent to exert anti-tumor effects.Nonetheless,both production effi-ciency and clinical applications of epimedium flavonoids have been restrained because of their low content,poor bioavailability,and unfavorable in vivo delivery efficiency.Recently,various strategies,including enzyme engineering and nanotechnology,have been developed to increase productivity and activity,improve delivery efficiency,and enhance therapeutic effects of epimedium flavonoids.In this review,the structure-activity relationship of epimedium flavonoids is described.Then,enzymatic en-gineering strategies for increasing the productivity of highly active baohuoside I and icaritin are dis-cussed.The nanomedicines for overcoming in vivo delivery barriers and improving therapeutic effects of various diseases are summarized.Finally,the challenges and an outlook on clinical translation of epi-medium flavonoids are proposed.

Two new terpene glycosides from the Alpiniae Oxyphyllae Fructus / 药学学报

Two undescribed terpene glycosides and two compounds were isolated from the n-butanol fraction of Alpiniae Oxyphyllae Fructus by using various chromatographic methods, including MCI Gel, Sephadex LH-20, ODS, silica gel and semi-preparative HPLC. The structures of the isolated compounds were identified by spectroscopy methods (1D, 2D NMR, UV, IR, MS, etc.), and the absolute configuration of the compound 1 was determined by ECD calculation and acid hydrolysis. Compounds 1 and 2 are new compound, and compounds 3 and 4 were isolated from Alpiniae Oxyphyllae Fructus for the first time.

Preparation of Diosgenin from Dioscorea zingiberensis： A Review / 中国实验方剂学杂志

Direct acid hydrolysis of Dioscorea zingiberensis rhizomes for preparation of diosgenin is wildly used in the traditional industry， which uses a large amount of inorganic acid catalysts， with high wastewater discharge and serious environmental pollution. Therefore， exploring clean and efficient preparation methods and processes has become an inevitable choice to realize the sustainable development of industrial production of diosgenin. Herein， the author reviewed and analyzed the research progress and problems of enzymatic hydrolysis， microbial transformation and modified acid hydrolysis in the preparation of diosgenin from D. zingiberensis rhizomes during the last ten years， and their application prospects are analyzed. Enzymatic hydrolysis has mild reaction conditions， but the yield of diosgenin is low， the economic cost is high， and the purification process of active enzyme is complicated. Microorganism shows specific activity to the substrate and high efficiency for diosgenin production， and microbial transformation is clean and environmentally friendly， but microbial transformation is time-consuming and the metabolic intermediates are complicated. For the modified acid hydrolysis， two-phase acid hydrolysis can reduce the amount of acid catalyst， and sulfonic acid-functionalized ionic liquid displays good recyclable performance by replacing the traditional inorganic acid， however， the wastewater discharge should still be considered. Solid acid catalysts are non-corrosive and easy to be recycled， but the need to use ethanol as the reaction solvent has certain safety hazards， and the catalyst preparation process is cumbersome. In conclusion， exploring clean and efficient conversion methods is an important research trend for preparation of diosgenin from D. zingiberensis rhizomes. For the enzymatic hydrolysis， the key glycoside hydrolases in the bioconversion process should be explored in depth， the conversion pathway of enzymatic saponins and enzyme specificity should be fully elucidated， and efforts should be made to improve the efficiency of enzymatic hydrolysis. For the microbial transformation， we should accelerate its industrial application process based on selecting and breeding efficient transformation strains， and optimizing stable transformation systems and processes， and in-depth investigation of the mechanism of microbial transformation， fully elucidating the specific key hydrolases and its catalytic properties， and striving to improve the efficiency of microbial transformation. For the modified acid hydrolysis， novel acid catalytic system with simple structure， stable performance and good biodegradability should be explored and applied， which can effectively solve the problems of environmental pollution and production safety.

Identification the key factor of pulmonary fibrosis following silica nanoparticles exposure based on bioinformatics analysis / 中华劳动卫生职业病杂志

Objective: To investigate the main mechanisms of pulmonary fibrosis following silica nanoparticles (SiNPs) exposure through constructing the macrophage-fibroblast model in vitro, which simulated the process of pulmonary fibrosis. Methods: In January 2021, human mononuclear leukemia cells (THP-1) were treated with 0, 25, 50, 100 μg/ml SiNPs for 24 h. The supernatant of THP-1 cells was collected and applied to human embryonic lung fibroblast cells (MRC-5) which divided into control and low, medium and high dose groups at the logarithmic growth stage for 24 h. MRC-5 cell viability was detected by CCK8. The hydroxyproline (Hyp), interleukin 6 (IL-6), interleukin 1 beta (IL-1β) and tumor necrosis factor-alpha (TNF-α) expression were detected in the supernatants of MRC-5. The changed proteins were detected by liquid-phase mass spectrometry in high dose group. GeneCard database were applied to identity the differential pulmonary fibrosis proteins in high dose group. Gene Ontology (GO) was performed to identity the key biological process in differential pulmonary fibrosis proteins of high dose group. The String database was used to construct the protein-protein interactions (PPI) network of differential pulmonary fibrosis proteins. The APP of CytoHubba was applied to calculate the key protein of differential pulmonary fibrosis proteins in PPI network. Correlation coefficients between key differential pulmonary fibrosis proteins were calculated using Pearson correlation analysis. Western blotting was applied to detect the expression of key proteins of differential pulmonary fibrosis proteins in different groups. Results: CCK8 results showed that MRC-5 cell viability was increasing in low, medium and high dose groups compared with control group (P<0.05). The expression levels of Hyp and IL-1β in different group were increased compared with control group, the expression levels of IL-6 and TNF-α were increased in high dose group compared with control group (P<0.05). GeneCard database identified 26 differential pulmonary fibrosis proteins, which were mainly involved in extracellular matrix hydrolysis, cell inflammatory response, tissue repair, cell proliferation, inflammation response by GO analysis. The APP of CytoHubba was calculated that matrix metalloproteinase 9 (MMP9) and tissue inhibitor metalloproteinase 1 (TIMP1) played an important role in PPI network. The results of correlation analysis showed that MMP9 was correlated with the expression of matrix metalloproteinase 1 (MMP1), matrix metalloproteinase 3 (MMP3), TIMP1 and epidermal growth factor receptor (EGFR) (r=0.97, 0.98, 0.94, 0.93, P<0.05). Western blotting results showed that TIMP1 protein expression was increased in low, medium and high dose groups, while MMP9 protein expression was increased only in high dose group (P<0.05) . Conclusion: Differential expression proteins related with pulmonary fibrosis in MRC-5 cells mainly regulate biological processes of extracellular matrix hydrolysis, tissue repair, and cellular inflammation response following SiNPs exposure. MMP9 and TIMP1 may be the key proteins, which affected the fibrosis process in vitro pulmonary fibrosis model.

Five new terpenoids from Viburnum odoratissimum var. sessiliflorum / 中国天然药物

Five new terpenoids, including two vibsane-type diterpenoids (1, 2) and three iridoid allosides (3-5), together with eight known ones, were isolated from the leaves and twigs of Viburnum odoratissimum var.sessiliflorum. Their planar structures and relative configurations were determined by spectroscopic methods, especially 2D NMR techniques. The sugar moieties of the iridoids were confirmed as β-D-allose by GC analysis after acid hydrolysis and acetylation. The absolute configurations of neovibsanin Q (1) and dehydrovibsanol B (2) were determined by quantum chemical calculation of their theoretical electronic circular dichroism (ECD) spectra and Rh2(OCOCF3)4-induced ECD analysis. The anti-inflammatory activities of compounds 1, 3, 4, and 5 were evaluated using an LPS-induced RAW264.7 cell model. Compounds 3suppressed the release of NO in a dose-dependent manner, with an IC50 value of 55.64 μmol·L-1. The cytotoxicities of compounds 1-5 on HCT-116 cells were assessed and the results showed that compounds 2 and 3 exhibited moderate inhibitory activities with IC50 values of 13.8 and 12.3 μmol·L-1, respectively.

Study on release and antioxidant activity of quercetin after loaded by hot alkali hydrolysis modified UiO-66 material / 药学学报

UiO-66 (University of Oslo 66) is a kind of promising material that can improve the release and bioavailability of poorly water-soluble bioactive compounds of traditional Chinese medicine. However, the loading of quercetin in raw UiO-66 was not ideal. In this study, UiO-66-BH (UiO-66-blend-heating) was obtained by heating UiO-66 and KOH solution following blended them. UiO-66-BH maintained the outline of octahedral structure of UiO-66 but with obvious rough and uneven pores on the surface. UiO-66-BH had good adsorption of quercetin with saturation adsorption was 138.92 mg·g-1, the adsorption process belonged to single molecular layer adsorption and was controlled by chemisorption. UiO-66-BH can control the release of quercetin in simulated gastrointestinal fluid, and the drug concentration was significantly higher than that of free quercetin after long-term release (36% vs 9%). Compared with quercetin, the ABTS (2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) ammonium salt) radical scavenging activity of UiO-66-BH@quercetin drug delivery system decreased, while the DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging activity remained almost unchanged. The drug delivery system showed a strong antioxidant effect similar to quercetin. The findings indicated that UiO-66-BH could control release of quercetin and was expected to be used as a drug carrier material for some insoluble active components of traditional Chinese medicine such as quercetin.

Exploring of Transformation Rule of Saponins in Polygalae Radix Before and After Processing Based on Simulated Processing Technology / 中国实验方剂学杂志

ObjectiveTo investigate the transformation mechanism and content variation of saponins from Polygalae Radix before and after being boiled with licorice juice and water. MethodSimulated licorice juice boiled products and simulated water boiled products of onjisaponin B， onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ were prepared by simulated processing technology， and analyzed by ultra-performance liquid chromatography-quadrupole-electrostatic field orbitrap high resolution mass spectrometry（UPLC-Q-Exactive Orbitrap/MS）. Then the contents of onjisaponin B， onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ and tenuifolin in Polygalae Radix， licorice-boiled Polygalae Radix and water-boiled Polygalae Radix were determined by UPLC-triple quadrupole tandem mass spectrometry（UPLC-QQQ-MS/MS）. ResultDuring the boiling process with licorice juice and water， onjisaponin B could be hydrolyzed to produce 4-methoxycinnamic acid， desacylsenegin Ⅲ， polygalasaponin ⅩⅩⅧ and tenuifolin， onjisaponin Z could be hydrolyzed to produce 3，4，5-trimethoxycinnamic acid， onjisaponin TF， polygalasaponin ⅩⅩⅧ and tenuifolin， onjisaponin F could be hydrolyzed to produce 3，4，5-trimethoxycinnamic acid， onjisaponin G， polygalasaponin ⅩⅩⅧ and tenuifolin， and polygalasaponin ⅩⅩⅧ was hydrolyzed to produce tenuifolin. After being boiled with licorice juice or water， the content of onjisaponin B decreased significantly（P<0.05， P<0.01）， but the contents of onjisaponin Z， onjisaponin F， polygalasaponin ⅩⅩⅧ and tenuifolin increased significantly（P<0.05， P<0.01） in Polygalae Radix. Compared with the water-boiled products， the contents of onjisaponin Z and tenuifolin increased significantly（P<0.05， P<0.01）， and the change of tenuifolin content was the most significant in the licorice-boiled products.However， there was no significant difference in the content of onjisaponin B， onjisaponin F and polygalasaponin ⅩⅩⅧ between the water-boiled products and the licorice-boiled products. ConclusionBeing boiled with licorice juice or water can hydrolyze onjisaponin B， onjisaponin Z， onjisaponin F and polygalasaponin ⅩⅩⅧ， and generate secondary glycosides and aglycones（organic acids） through deglycosylation， which leads to obvious changes in the contents of onjisaponins after Polygalae Radix being processed.It is inferred that licorice juice can promote the hydrolysis of some onjisaponins in Polygalae Radix to onjisaponin Z and tenuifolin.This study provides an experimental basis for revealing processing mechanism of Polygalae Radix.

Reutilization of waste fungal biomass for concomitant production of proteochitinolytic enzymes and their catalytic products by Alcaligenes faecalis SK10

Fungal biomass, being organic waste, could be an excellent source of protein, carbohydrate and minerals. However, it has not been exploited fully until now. Efficient management of this waste can not only address the environmental impact on its disposal but also yield value-added metabolites. In the present study, in order to explore its potential, we subjected dead fungal biomass of Aspergillus niger SKN1 as substrate for both fermentative and enzymatic biodegradation, respectively by potent proteo-chitinolytic bacteria Alcaligenes faecalis SK10 and its enzyme cocktail. The results revealed that reasonable amount of protease and chitinase could be biosynthesized by the fermentative mode of utilization, while a mixture of amino acid, peptides and low-molecular weight amino-sugar (mono and oligomeric form of N-acetylglucosamine) could be generated through enzymatic hydrolysis. The physicochemical condition of both the bioprocess was subsequently optimized through statistical approach. The projected utilization of waste zero-valued fungal biomass offer a sustainable and environmentally sound method for production of microbial metabolites and large scale execution of the same could be proficient and in tune with the principle of circular economy.

Comparative amino acid composition and quality parameters of Moringa oleifera testa and cotyledon

@#Introduction: Moringa oleifera is a drought-resistant plant, widely used in the tropical region. The leaves and stems have been extensively utilised in foods and neutraceuticals preparation, with less attention to the seeds. In this study, amino acid (AA) compositions of M. oleifera testa and cotyledon were examined comparatively. Methods: Samples were separately defatted, hydrolyed, and neutralised. The AA solution was purified by cation-exchange solid-phase extraction, derivatised and analysed by gas chromatography. Results: Glutamic (acidic amino acid) and phenylalanine (essential amino acid, EAA) were the most concentrated in both samples. Total EAA (g/100g crude protein, cp) was higher in cotyledon (51.0) than testa (41.9). Predicted protein efficiency ratios (P-PERs) were higher in testa (0.605-1.530) than cotyledon 0.286-1.460). EAA index ranged between 0.951-1.13 (soybean comparison) and 83.0-96.9 (egg comparison) with corresponding biological value of 78.7-93.9. The following AA had scores >1.0 in comparison to whole hen’s egg, testa: glycine (Gly), glutamic acid (Glu), phenylalanine (Phe), histidine (His), and cysteine (Cys); cotyledon (Gly), proline (Pro), Glu, Phe, His, arginine (Arg) and Cys. In comparison with requirements of pre-school children, six AA (6/9 or 66.7%) had scores >1.0 in each sample. In provisional AA scoring pattern, isoleucine (Ile) (1.25) and Phe + tyrosine (Tyr) (1.68) had scores >1.0 in testa while methionine (Met) + Cys, Phe+Tyr, and tryptophan (Trp) in cotyledon. However, tryptophan and lysine were the limiting AAs in testa and cotyledon, respectively. Conclusion: The study showed that both anatomical parts would complement each other in terms of amino acid supply.

Reuse of Resource Components in Residue of Shaoyao Gancaotang / 中国实验方剂学杂志

Objective:Aiming at the residue of Shaoyao Gancaotang， the extraction， qualitative and quantitative study of the small molecule resource components were carried out to clarify the residual small molecule chemical components in the residue and explore the ways of its resource utilization. Method:The ultra-performance liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry （UPLC-QTOF-MS/MS） was used to qualitatively identify the residual small molecule substances in the dregs of Shaoyao Gancaotang. Agilent C₁₈ reversed-phase chromatographic column （3.0 mm×100 mm， 2.7 µm） was used at the flow rate of 0.4 mL·min^-1， the injection volume was 5 µL， and the mobile phase was gradient eluted with 0.05% formic acid aqueous solution （A）-acetonitrile （B） （0-1 min， 14%-17.5%B； 1-3 min， 17.5%-19%B； 3-4 min， 19%-20%B； 4-5 min， 20%B； 5-6 min， 20%-21%B； 6-9 min， 21%B； 9-22 min， 21%-36%B； 22-23 min， 36%B； 23-32 min， 36%-43%B）， electrospray ionization （ESI） was employed with negative ion mode scanning and scanning range of <italic>m</italic>/<italic>z</italic> 50-1 200. A high performance liquid chromatography （HPLC） was established for the quantitative analysis of its main components with Agilent C₁₈ reversed-phase chromatographic column （4.6 mm×150 mm， 5 µm）， the detection wavelength was set at 235 nm， the flow rate was 0.8 mL·min^-1， and the injection volume was 5 µL. Mobile phase was 0.05% phosphoric acid （A）-acetonitrile （B） for gradient elution （0-1 min， 14%-19%B； 1-4 min， 19%B； 4-18 min， 19%-50%B）. The content changes of main components in the residue of Shaoyao Gancaotang were compared before and after two different techniques of organic solvent extraction and enzymatic extraction. Result:A total of 16 chemical components in the residue of Shaoyao Gancaotang were qualitatively analyzed， and quantitative analysis found that there were many chemical components in the residue， among which the residues of 6 index components such as paeoniflorin and liquiritin reached more than 70% in the original decoction piece. After enzymolysis by cellulase， liquiritin in the residue could be converted into liquiritigenin. The content of crude polysaccharide in enzymatic extract of the residue was 6 times higher than that in the blank group， and the content was up to 12%. Conclusion:There are still many small molecule resource components in the residue of Shaoyao Gancaotang， which has great development potential. Organic solvents can be used to re-extract the target components in the residue， and liquiritin can be converted into liquiritigenin by biological fermentation technology， and the crude polysaccharide from the residue can be extracted by enzymatic method to develop animal feed. This study can provide reference basis and approach for reusing the residues of Shaoyao Gancaotang preparations and dispensing granules， so as to realize the high-value utilization of Shaoyao Gancaotang.

Structure and identification of immune fragments of acid-degraded oligosaccharides from active Astragalus polysaccharides APS-Ⅱ / 药学学报

We previously reported that active Astragalus polysaccharides APS-Ⅱ generate strong immune activity. Here we establish the optimal method for APS-II acid degradation. After preliminary structural studies and separation and preparation of the degradation products, the oligosaccharide active center with the strongest immune activity was identified by in vitro immune cell culture experiments. The optimum acid degradation conditions for APS-II were determined by a single factor experiment and an orthogonal experiment. Astragalus oligosaccharides prepared under the optimal conditions were subjected to structural analysis by hydrophilic interaction chromatography - electrospray ionization source - high resolution time-of-flight mass spectrometry. The products were separated and oligosaccharide fragments with different degrees of polymerization were isolated by preparative purification chromatography. Finally, fragments of the immunologically active centers were identified by in vitro immune cell cultures from multiple perspectives. The results show that the optimal acid hydrolysis conditions for APS-Ⅱ are hydrolysis temperature 80 ℃, trifluoroacetic acid concentration 1.0 mol·L-1, hydrolysis time 1 h. The degradation conditions have good repeatability. The degradation product is a six-carbon aldehyde glycan structure with the main chain 1→4 connected. The immune activity screening experiment for six oligosaccharide fragments showed that larger molecular weight oligosaccharides have stronger immune-promoting effects. It is speculated that the immunologically active center of Astragalus oligosaccharide is located in the sugar chain of DP9-DP19. The animal welfare and the experimental process in this study follow the requirements of the Animal Ethics Committee of Shanxi University. This result suggests a foundation for the structural characterization and structure-activity relationship research of Astragalus oligosaccharides, and may promote the development of Astragalus oligosaccharide drugs.

Establishment of enzymatic hydrolysis method of active Astragalus polysaccharides APS-Ⅱ and study on the immune activity of oligosaccharides after its degradation / 药学学报

italic>Astragalus polysaccharides are the main immunomodulatory substances in Astragali Radix. The structure of polysaccharides is difficult to accurately determine, which limits the in-depth study of the molecular mechanism of Astragalus polysaccharides in vivo. "Polysaccharide receptor theory" believes that there are one or more oligosaccharide fragment "active centers" in immunologically active polysaccharide molecules. Therefore, the degradation of Astragalus polysaccharides into oligosaccharides and the study of the active centers of polysaccharides at the oligosaccharide level provide new ideas in the study of the structure and mechanism of Astragalus polysaccharides. This article adopts endo-α-1,4-glucanase enzymatic hydrolysis, and determines the best degradation conditions through single factor test and orthogonal test to degrade the immunologically active polysaccharide APS-Ⅱ (10 kDa component) into oligomers with different degrees of polymerization. Then through the preparation of polyacrylamide gel chromatography and specific immune and non-specific immune cell tests, the immune activity screening of different oligosaccharide components is carried out. The animal welfare and the experimental process in this study follow the requirements of the Animal Ethics Committee of Shanxi University. The results showed that compared with the immunologically active polysaccharide APS-Ⅱ, different oligosaccharide components have obvious differences in different immunological activities. This paper studies the different immunological activities of Astragalus polysaccharides at the level of oligosaccharides, laying a foundation for further elucidating the structure and function of Astragalus polysaccharides, enriching the theory of polysaccharide receptors, and providing new ideas for the development of Astragalus polysaccharides.

Phenolic glycosides from the stems of Homalium stenophyllum / 中国天然药物

The phytochemical investigation of the stems of Homalium stenophyllum afforded seven new phenolic glycosides (1-5 and 8-9) and two known compounds (6 and 7). Their structures were elucidated by comprehensive analyses of NMR spectroscopic, mass spectrometric data and chemical hydrolysis. Additionally, their anti-inflammatory activities against the NO production in LPS-induced macrophages were evaluated.

Discovery and functional verification of endogenous glucanases for scleroglucan hydrolysis in Sclerotium rolfsii / 生物工程学报

Scleroglucan is a high-molecular water-soluble microbial exopolysaccharide and mainly applied in the fields of petroleum, food, medicine and cosmetics. The high molecular weight of scleroglucan produced by microbial fermentation leads to low solubility, high viscosity and poor dispersibility, thus bringing a series of difficulties to extraction, preservation and application. It is important to explore suitable degradation method to adjust the molecular weight of scleroglucan for expanding its industrial application. Taking Sclerotium rolfsii WSH-G01 as a model strain, in which functional annotations of the glucanase genes were conducted by whole genome sequencing. Based on design of culture system for culture system for differential expression of β-glucanase, endogenous β-glucanase genes in S. rolfsii WSH-G01 were excavated by transcriptomics analysis. Functions of these potential hydrolases were further verified. Finally, 14 potential endogenous hydrolase genes were obtained from S. rolfsii. After heterologous overexpression in Pichia pastoris, 10 soluble enzymes were obtained and 5 of them had the activity of laminarin hydrolysis by SDS-PAGE and enzyme activity analysis. Further investigation of the 5 endogenous hydrolases on scleroglucan degradation showed that enzyme GME9860 has positive hydrolysis effect. The obtained results provide references not only for obtaining low and medium molecular weight of scleroglucan with enzymatic hydrolysis, but also for producing different molecular weight of scleroglucan during S. rolfsii fermentation process with metabolic engineering.

Effect of high hydrostatic pressure on the antioxidant capacity and peptic hydrolysis of whey proteins / Efeito da alta pressão hidrostática na hidrólise péptica das proteínas do soro de leite e na capacidade antioxidante

ABSTRACT: The effect of high hydrostatic pressure (HHP) application on whey protein concentrate was evaluated both before (pre-treatment - PT) and during (hydrolysis assisted - HA) hydrolysis processes. A factorial design 22 with 3 central points was used with pressure (100, 250, 400 MPa) and time (5, 20 and 35 minutes) as independent variables. The hydrolysis was evaluated and monitored by soluble protein, aromatic amino acid contents and RP-HPLC. ABTS and ORAC tests were used to evaluate the in vitro antioxidant capacity. The reduction of soluble protein content was approximately 20% for conventional hydrolysis and for all PT treatments up to 4 h of reaction, while HHP assisted hydrolysis at 100 MPa showed a 35% protein reduction after 35 minutes of reaction. In addition, pressurization favored peptic hydrolysis of β-lactoglobulin by up to 98% and also improved the in vitro antioxidant capacity of the hydrolysates, which increased from 34.25 to 60.89 μmoles TE g-1 of protein in the best treatment. The results suggest that the use of HHP assisted hydrolysis favored the peptic hydrolysis, with a reduction in hydrolysis time and increased antioxidant activity.

RESUMO: Neste estudo, o efeito da aplicação de alta pressão hidrostática (HHP) sobre o concentrado proteico de soro de leite foi avaliado antes (pré-tratamento - PT) e durante os processos de hidrólise (assistida por hidrólise - HA). Utilizou-se o delineamento fatorial 22 com três pontos centrais, onde as variáveis independentes foram pressão (100, 250, 400 MPa) e tempo (5, 20 e 35 minutos). A hidrólise foi avaliada pelo conteúdo de proteínas solúveis e aminoácidos aromáticos, além do perfil peptídico por RP-HPLC. As análises de ABTS e ORAC foram utilizadas para avaliar a capacidade antioxidante in vitro. A redução do teor de proteína solúvel foi de aproximadamente 20% para a hidrólise convencional e para todos os pontos de PT até 4h de reação, enquanto a hidrólise assistida por HHP a 100 MPa mostrou uma redução de 35% de proteína em 35 minutos de reação. Além disso, a pressurização favoreceu a hidrólise péptica da β-lactoglobulina em até 98% e também melhorou a capacidade antioxidante in vitro dos hidrolisados, que aumentaram de 34,25 para 60,89 μmoles de TE g-1 de proteína no melhor tratamento. Os resultados sugerem que o uso da hidrólise assistida por HHP favoreceu a hidrólise péptica, com redução no tempo de hidrólise e aumento da atividade antioxidante.

Biomass Sorghum: Effect of Acid, Basic and Alkaline Peroxide Pretreatments on the Enzymatic Hydrolysis and Ethanol Production

Abstract This study evaluated the effects of three chemical pretreatments of biomass sorghum (BS): dilute alkaline (PTA1 and PTA2), dilute acid (PTB1 and PTB2) and alkaline hydrogen peroxide (PTC1 and PTC2) in the enzymatic hydrolysis and ethanol production. Among the six investigated conditions, the pretreatment with 7.36% H2O2 (PTC2) was the most efficient in the lignin removal and preservation of the polysaccharide fraction. After the enzymatic hydrolysis, increases in the glucose and xylose concentrations were observed in the pretreated BS hydrolysates, mainly in PTB1 and PTC1. All the hydrolysates obtained low concentrations of inhibitors. In the alcoholic fermentations with Pichia stiptis, the greatest ethanol yield was obtained in PTB1 hydrolysate (3.84 g L-1), corresponding to 16.15% of yield. The highest ethanol yield in PTB1 hydrolysate can be justified by the maximum concentration of xylose obtained in this hydrolysate, demonstrating the potential of P. stiptis in the fermentation of pentose to ethanol. The results indicated that biomass sorghum is an alternative lignocellulose source with potential for the production of second generation ethanol, opening up prospects for additional studies.

Hydrothermal Pretreatment as a Strategy for the Improvement of Sugarcane Bagasse Saccharification by Fungal Enzyme Blend

Abstract Obtaining low cost lignocellulolytic enzymes and efficient biomass pretreatment are key to increase the competitiveness of second-generation ethanol in comparison with fossil fuels. The enzymatic cocktail produced by the Chrysoporthe cubensis fungus as well as the mixture prepared with the cocktails of the Chrysoporthe cubensis and Penicillium pinophilum fungi have already proven to be efficient for hydrolyzing biomass pretreated with alkali. In this study, they were evaluated in saccharification of sugarcane bagasse pretreated with dilute acid or hot water at 121°C using an enzyme loading equal to 8 filter paper units per gram of biomass. The most promising results were obtained from the hydrolysis of biomass pretreated with hot water by the C. cubensis-P. pinophilum enzymes blend. In this condition, the glucose and xylose production were 25.2 g.L-1 and 4.6 g.L-1, respectively, that resulted in the conversion of 68% of glucan and 23% of xylan in only 48 hours. This study shows that the hydrothermal pretreatment is a promising alternative to improve the enzymes performance, produced by the fungi C. cubensis and P. pinophilum, in the sugarcane bagasse hydrolysis without the need of chemical compounds, generally used in the acid and alkali pretreatments. Furthermore, the hydrothermal pretreatment for 60 min allowed all cocktails applied to convert the cellulose efficiently with only 24 h of saccharification, which contributes to the energy savings employed in the process.