Effect of Degumming in the Characteristics of Silk Fibroin Nanoparticles

Several studies have stated that the process used for sericin removal, or degumming, from silk cocoons has a strong impact on the silk fibroin integrity and consequently in their mechanical or biochemical properties after processing it into several biomaterials (e.g., fibers, films or scaffolds) but still, there is a lack of information of the impact on the features of silk nanoparticles. In this work, silk cocoons were degummed following four standard methods: autoclaving, short alkaline (Na2CO3) boiling, long alkaline (Na2CO3) boiling, and ultrasounds. The resultant silk fibroin fibers were dissolved in the ionic liquid 1-ethyl-3-methylimidazolium acetate and used for nanoparticle synthesis by rapid desolvation in polar organic solvents. The relative efficiencies of the degumming processes and the integrity of the resulting fibroin fibers obtained were analyzed by mass loss, optical microscopy, thermogravimetric analysis, infrared spectroscopy, and SDS-PAGE. Particle sizes and morphology were analyzed by Dynamic Light Scattering and Field Emission Scanning Electronic Microscopy. The results showed that the different treatments had a remarkable impact on the integrity of the silk fibroin chains, as confirmed by gel electrophoresis, which can be correlated with particle mean size and size distribution changes. The smallest nanoparticles (156 ± 3 nm) and the most negative Z potential (−30.2 ± 1.8 mV) were obtained with the combination of long treatment (2 h) of boiling in alkaline solution (Na2CO3 0.02 eq/L). The study confirms that parameters of the process, such as the composition of the solution and time of the degumming step, must be controlled in order to reach an optimum reproducibility of the nanoparticle production.This work has been partially supported (80%) by the European Commission ERDF/FEDER Operational Programme 'Murcia' CCI N° 2007ES161PO001 (Project No. 14-20/20), and the Spanish MINECO (Ref. CTQ2017-87708-R) and the programme of support to the research of the Seneca Foundation of Science and Technology of Murcia, Spain (Ref. 20977/PI/18). A.A.L.-P.’s research contract was partially supported (80%) by the ERDF/FEDER Operational Programme 'Murcia' CCI N° 2007ES161PO001 (Project No. 14-20/20),. M.G. Montalbán’s research contract is funded by the Spanish MINECO (Juan de la Cierva-Formación contract, Ref. FJCI-2016-28081). S.D.A.-C.’s research contract is funded by the program INIA-CCAA (DOC INIA 2015), announced by the National Institute for Agricultural and Food Research and Technology (INIA) and supported by the Spanish State Research Agency (AEI) under the Spanish Ministry of Economy, Industry and Competitiveness

Utilización de marcadores AFLP y SAMPL en la identificación genética de especies y variedades de cítricos

Many species within the Citrus genus are economically important crops throughout tropical and subtropical regions of the world. Molecular marker technologies have eased and improved the genetic analysis of plants and promise to greatly assist Citrus breeding programs. Several types of molecular markers have been used to characterize the Citrus genome, but this is the first time where Amplified Restriction Fragment Polymorphism (AFLP) and Selective Amplification of Microsatellite Polymorphic Loci (SAMPL) techniques are used to identify varieties within species that belong to Citrus genus such as, C. sinensis, C. clementina, C. paradisi, C. aurantium, and C. limon. Both techniques were used to identify accessions belonging to the CIDA - Murcia, collection (Spain).Key words: AFLP, citrus, molecular markers, SAMPL, variety differentiatioMuchas especies del género Citrus son cultivos de gran importancia económica en regiones tropicales y subtropicales del mundo. La aplicación de marcadores moleculares ha facilitado y potenciado el análisis genético de plantas pudiendo resultar herramientas de gran ayuda en los programas de mejoramiento de cítricos. Se han empleado diversos tipos de marcadores moleculares para la caracterización del genoma de cítricos, pero este es el primer trabajo de empleo de AFLPs (Polimorfismos para la longitud de los fragmentos amplificados) y SAMPLs (Amplificación selectiva de loci polimórficos de microsatélites), en la identificación de variedades pertenecientes a especies del género Citrus, como son C. sinensis, C. clementina, C. paradisi, C. aurantium, y C.limon. Ambas técnicas fueron utilizadas para la identificación de muestras pertenecientes a la colección del CIDA-Murcia (España).Palabras clave: AFLP, citrus, marcadores moleculares, SAMPL, diferenciación varieta

Biopolymeric Nanoparticle Synthesis in Ionic Liquids

Recently, much research has focused on the use of biopolymers, which are regarded as biodegradable, natural, and environmentally friendly materials. In this context, biopolymeric nanoparticles have attracted great attention in the last few years due to their multiple applications especially in the field of biomedicine. Ionic liquids have emerged as promising solvents for use in a wide variety of chemical and biochemical processes for their extraordinary properties, which include negligible vapor pressure, high thermal and chemical stability, lower toxicity than conventional organic solvents, and the possibility of tuning their physical–chemical properties by choosing the appropriate cation and anion. We here review the published works concerning the synthesis of biopolymeric nanoparticles using ionic liquids, such as trimethylsilyl cellulose or silk fibroin. We also mention our recent studies describing how high-power ultrasounds are capable of enhancing the dissolution process of silk proteins in ionic liquids and how silk fibroin nanoparticles can be directly obtained from the silk fibroin/ionic liquid solution by rapid desolvation in polar organic solvents. As an example, their potential biomedical application of curcumin-loaded silk fibroin nanoparticles for cancer therapy is also discussed

Silk fibroin nanoparticles enhance quercetin immunomodulatory properties in DSS-induced mouse colitis

This work has been supported from the European Commission ERDF/FEDER Operational Programme `Murcia' CCI N. 2007ES161PO001 (Project No. 14-20/20), the Junta de Andalucia (CTS164), Instituto de Salud Carlos III (PI19/01058) and the Spanish MINECO (Ref. CTQ201787708-R). P.D.-E. is a postdoctoral from Junta de Andalucia (European Commission FEDER); A.J.R.-M and L.H.-G. are predoctoral fellows from University of Granada ("Programa de Doctorado: Medicina Clinica y Salud Publica"); A.A.L.-P's research contract was supported by the ERDF/FEDER Operational Programme `Murcia' CCI N 2007ES161PO001 (Project No. 14-20/20); A.R.-N. is a postdoctoral fellow of Instituto de Salud Carlos III (Miguel Servet Program); T.V. is a postdoctoral fellow from Instituto de Investigación Biosanitaria de Granada.Inflammatory bowel disease (IBD) is a chronic and idiopathic inflammatory disorder affecting the gastrointes-tinal tract. The pharmacological treatments used currently for its treatment lack efficacy, so new therapeutic strategies should be developed. In this context, flavonoids loaded in biopolymeric nanoparticles can be considered as novel promising candidates. The aim of the present study was to evaluate the intestinal anti-inflammatory effects of quercetin when is administered loaded in silk fibroin nanoparticles (QSFN) in the dextran sulphate sodium experimental model of mouse colitis, which displays some similarities to human IBD. Previously characterized quercetin-loaded silk fibroin nanoparticles (QSFN). QSFN showed a reversible aggre-gation profile induced by the acidification of the solution but did not affect the loaded quercetin. Daily administration of QSFN significantly reduced disease activity index values compared to the control colitic group. This beneficial effect was not only corroborated by the histological examination of the colonic specimens but also the improvement of the colonic expression of the different proinflammatory cytokines (Tnf-alpha, Il-1 beta, Il-6, Mcp-1, Icam-1, Nlrp3 and iNOS). Therefore, these data suggest that QSFN could be a promising alternative to current treatments as a drug delivery system for IBD treatment.European Commission ERDF/FEDER Operational Programme `Murcia' CCI 2007ES161PO001- 14-20/20Junta de Andalucia CTS164Instituto de Salud Carlos III European Commission PI19/01058Spanish MINECO CTQ201787708-RERDF/FEDER Operational Programme 'Murcia' CCI 2007ES161PO001- 14-20/2

Impact of Covalent Functionalization on the Aqueous Processability, Catalytic Activity, and Biocompatibility of Chemically Exfoliated MoS2 Nanosheets

Chemically exfoliated MoS2 (ce-MoS2) has emerged in recent years as an attractive two-dimensional material for use in relevant technological applications, but fully exploiting its potential and versatility will most probably require the deployment of appropriate chemical modification strategies. Here, we demonstrate that extensive covalent functionalization of ce-MoS2 nanosheets with acetic acid groups (∼0.4 groups grafted per MoS2 unit) based on the organoiodide chemistry brings a number of benefits in terms of their processability and functionality. Specifically, the acetic acid-functionalized nanosheets were furnished with long-term (>6 months) colloidal stability in aqueous medium at relatively high concentrations, exhibited a markedly improved temporal retention of catalytic activity toward the reduction of nitroarenes, and could be more effectively coupled with silver nanoparticles to form hybrid nanostructures. Furthermore, in vitro cell proliferation tests carried out with murine fibroblasts suggested that the chemical derivatization had a positive effect on the biocompatibility of ce-MoS2. A hydrothermal annealing procedure was also implemented to promote the structural conversion of the functionalized nanosheets from the 1T phase that was induced during the chemical exfoliation step to the original 2H phase of the starting bulk material, while retaining at the same time the aqueous colloidal stability afforded by the presence of the acetic acid groups. Overall, by highlighting the benefits of this type of chemical derivatization, the present work should contribute to strengthen the position of ce-MoS2 as a two-dimensional material of significant practical utility.Financial support from the Spanish Ministerio de Economía y Competitividad (MINECO) and the European Regional Development Fund (FEDER) through project MAT2015-69844-R is gratefully acknowledged, as well as partial support of the ERDF/FEDER. Operative Program of the Región de Murcia 2007–2013. We also acknowledge partial funding by Plan de Ciencia, Tecnología e Innovación 2013-2017 del Principado de Asturias and FEDER through grant GRUPIN14-056.Peer reviewe

Revealing the Influence of the Degumming Process in the Properties of Silk Fibroin Nanoparticles

Several studies have stated that the process used for sericin removal, or degumming, from silk cocoons has a strong impact in the silk fibroin integrity and consequently in their mechanical or biochemical properties after processing it into several biomaterials (e.g. fibers, films or scaffolds) but still, there is a lack of information of the impact on the features of silk nanoparticles. In this work, silk cocoons were degummed following four standard methods: autoclaving, short alkaline (Na2CO3) boiling, long alkaline (Na2CO3) boiling and ultrasounds. The resultant silk fibroin fibers were dissolved in the ionic liquid 1-ethyl-3-methylimidazolium acetate and used for nanoparticle synthesis by rapid desolvation in polar organic solvents. The relative efficiencies of the degumming processes and the integrity of the resulting fibroin fibers obtained were analyzed by mass loss, optical microscopy, thermogravimetric analysis, infrared spectroscopy and SDS-PAGE. Particle sizes and morphology were analyzed by Dynamic Light Scattering and Field Emission Scanning Electronic Microscopy. The results showed that the different treatments had a remarkable impact on the integrity of the silk fibroin chains, as confirmed by gel electrophoresis, which can be correlated with particle mean size and size distribution changes. The smallest nanoparticles (156 ± 3 nm) and the most negative Z potential (−30.2 ± 1.8 mV) were obtained with the combination of long treatment (2 h) of boiling in alkaline solution (Na2CO3 0.02 eq/L). The study confirms that parameters of the process, such as composition of the solution and time of the degumming step, must be controlled in order to reach an optimum reproducibility of the nanoparticle production.This work has been partially supported (80%) from the European Commission ERDF/FEDER Operational Programme 'Murcia' CCI N° 2007ES161PO001 (Project No. 14-20/20), and the Spanish MINECO (Ref. CTQ2017-87708-R) and the programme of support to the research of the Seneca Foundation of Science and Technology of Murcia, Spain (Ref. 20977/PI/18).A.A. Lozano-Pérez’s research contract was partially supported (80%) by the ERDF/FEDER Operational Programme 'Murcia' CCI N° 2007ES161PO001 (Project No. 14-20/20),. M.G. Montalbán’s research contract is funded by the Spanish MINECO (Juan de la Cierva-Formación contract, Ref. FJCI-2016-28081). S.D. Aznar-Cervantes’s research contract is funded by the program INIA-CCAA (DOC INIA 2015), announced by the National Institute for Agricultural and Food Research and Technology (INIA) and supported by The Spanish State Research Agency (AEI) under the Spanish Ministry of Economy, Industry and Competitiveness

Comparative Study of the Antioxidant and Anti-Inflammatory E ects of Leaf Extracts from Four Di erent Morus alba Genotypes in High Fat Diet-Induced Obesity in Mice

The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.Increased levels of reactive oxygen species (ROS) and a low-grade chronic inflammation in multiple organs have been demonstrated in obesity. Morus alba leaves extracts (MAEs) have been used in traditional medicine as anti-inflammatory agents. In this work, the bioactive compounds of di erent genotypes of M. alba L. (Filipina, Valenciana Temprana, Kokuso, and Italia) were analyzed not only by reverse phase high performance liquid chromatography–electrospray ionization-time of flight-mass spectrometry (RP-HPLC-ESI-TOF-MS) and hydrophilic interaction chromatography–electrospray ionization-time of flight-mass spectrometry (HILIC-ESI-TOF-MS), but also screened for in vitro and in vivo antioxidant activity by means of DPPH radical scavenging assay and Caenorhabditis elegans model. These MAEs were administered daily in a model of diet-induced obesity in mice. Filipina and Italia genotypes significantly reduced weight gain, the glycemic levels in high fat diet, as well as, levels of LDL-cholesterol and triglycerides. Filipina and Italia MAEs also reduced the expression of proinflammatory mediators such as Tnf- , Il-1 , Il-6 and increased the levels of adiponectin and AMPK, which exert anti-inflammatory e ects. Moreover, Italia genotype ameliorated the intestinal barrier function. In conclusion, Filipina and Italia methanolic extracts show the highest antioxidant and anti-inflammatory e ect, due to the presence of compounds such as protocatechuic acid or quercetin-3-glucoside, and they could be developed as a complementary treatment for obesity and metabolic disorders.Junta de Andalucia CTS 164Spanish Ministry of Economy and Competitiveness AGL2015-67995-C3-3-REuropean Commission (FEDER/ERDF)ERDF/FEDER Operational Programme of the Region of Murcia 2007ES161PO001 14-20/20Instituto de Salud Carlos II

Mechanical behaviour and formation process of silkworm silk gut

High performance silk fibers were produced directly from the silk glands of silkworms ("Bombyx mori") following an alternative route to natural spinning. This route is based on a traditional procedure that consists of soaking the silk glands in a vinegar solution and stretching them by hand leading to the so called silkworm guts. Here we present, to the authors’ best knowledge, the first comprehensive study on the formation, properties and microstructure of silkworm gut fibers. Comparison of the tensile properties and microstructural organization of the silkworm guts with those of naturally spun fibers allows gain of a deeper insight into the mechanisms that lead to the formation of the fiber, as well as the relationship between the microstructure and properties of these materials. In this regard, it is proved that an acidic environment and subsequent application of tensile stress in the range of 1000 kPa are sufficient conditions for the formation of a silk fiber