Order matters: Methods for extracting cellulose from rice straw by coupling alkaline, ozone and enzymatic treatments.

Rice straw is a widely produced residue that can be converted into value-added products. This work aimed at using greener processes combining mild alkali (A), ozone (O) and enzymatic (engineered xylanase) (E) treatments to extract cellulose and other value-added compounds from rice straw and to evaluate the effects of the order of the treatments. Solid (S) and liquid (L) fractions from the process were collected for physicochemical characterization. AOE treatment showed the best capacity to extract high purity cellulose and other valuable compounds. The lignin content was significantly decreased independently of the order of the treatments and, its content in the extract obtained after the AOE process was lower than the one obtained after the OAE process. Moreover, thermal stability of the samples increased after the enzymatic process, being higher in SAOE. The alkaline treatment increased the hemicellulose and polyphenol content (antioxidant activity) in the liquid fractions (LA and LOA). In contrast, the ozonized liquid fractions had lower polyphenol content. Therefore, alkali was fundamental in the process. In conclusion, the AOE strategy could be a more environmentally friendly method for extracting cellulose and other valuable compounds, which could be used to develop active materials in the future.

Composition, structural properties and immunomodulatory activity of several aqueous Pleurotus ß-glucan-rich extracts.

In this work, aqueous extracts from six different Pleurotus species were obtained and their yield, gross composition, ß-glucan content, monosaccharide profile, thermal stability, molecular weight distribution, and FT-IR were analyzed before and after purification through ethanol precipitation of the carbohydrate-rich fractions. The bioactivity (anti-inflammatory and immunomodulatory activity) of the various fractions obtained was also analyzed in three different cell cultures and compared with a lentinan control. The trend observed after purification of the aqueous fractions was an increase in the concentration of polysaccharides (especially ß-glucans), a decrease in ash, glucosamine and protein content and the elimination of low molecular weight (Mw) compounds, thus leaving in the purified samples high Mw populations with increased thermal stability. Interestingly, all these purified fractions displayed immunomodulatory capacity when tested in THP-1 macrophages and most of them also showed significant activity in HEK-hTLR4 cells, highlighting the bioactivity observed for Pleurotus ostreatus (both the extracts obtained from the whole mushroom and from the stipes). This specific species was richer in heteropolysaccharides, having moderate ß-glucan content and being enriched upon purification in a high Mw fraction with good thermal stability.

Estimation of alginate purity and M/G ratio by methanolysis coupled with anion exchange chromatography.

Alginates are industrially relevant polysaccharides widely used in the food and biomedical industries for their excellent gelling properties. The growing emphasis on the valorization of marine resources has evidenced the need for alternative methods for the determination of both alginate content and the M/G ratio. This study describes the application of acid methanolysis and separation by anion exchange chromatography. Five samples, including alginates extracted from Saccharina latissima, Ascophyllum nodosum, a certified standard, and two poly-uronates (Poly-M and Poly-G), were analysed for their M/G ratio and alginate content at different treatment conditions, and compared with other conventionally used or reference methods (NMR, FTIR, and colorimetric methods). Quantitative estimation of alginate was relatively accurate at optimum conditions (4 h at 100 °C), as compared with the certified standard or with other colorimetric methods. M/G ratios were not significantly different from those determined after the reference method (1H NMR) or compared to FTIR protocols. The results evidence that methanolysis may be applied to simultaneously estimate the purity and M/G ratio of alginate-rich samples in a single analysis.

Impact of molecular weight and deacetylation degree of chitosan on the bioaccessibility of quercetin encapsulated in alginate/chitosan-coated zein nanoparticles.

This work aimed at studying the effect of molecular weight (MW) and deacetylation degree (DD) of chitosan on the quercetin bioaccessibility encapsulated in alginate/chitosan-coated zein nanoparticles (alg/chiZN). The chitosan coating layer produced nanoparticulate systems with good stability parameters, high encapsulation efficiency (EE) and a higher bioaccessibilty of quercetin after in-vitro digestion. By increasing the DD of chitosan, the ζ-potential of the colloidal system significantly increased (≥27.1 mV), while low and very low MW chitosans generated systems with smaller particle sizes (≤ 277.8 nm) and polydispersity index [PDI (0.189)]. The best results, in terms of EE (≥84.44) and bioaccessibility (≥76.70), were obtained when the systems were prepared with low MW chitosan and high DD. Thus, the alg/chiZN nanocapsules may be a promising delivery system for improving the quercetin bioaccessibility or other compounds with a similar chemical nature, especially when higher DD and lower MWs are used.

Structural and functional properties of alginate obtained by means of high hydrostatic pressure-assisted extraction.

The effects of the high hydrostatic pressure (HPP) pre-treatment on the alginate extraction were seen to greatly depend on the recalcitrant nature of two algae species. Alginates were deeply characterized in terms of composition, structure (HPAEC-PAD, FTIR, NMR, SEC-MALS), functional and technological properties. The pre-treatment significantly increased the alginate yield in the less recalcitrant A. nodosum (AHP) also favoring the extraction of sulphated fucoidan/fucan structures and polyphenols. Although the molecular weight was significantly lower in AHP samples, neither the M/G ratio nor the M and G sequences were modified. In contrast, a lower increase in alginate extraction yield was observed for the more recalcitrant S. latissima after the HPP pre-treatment (SHP), but it significantly affected the M/G values of the resulting extract. The gelling properties of the alginate extracts were also explored by external gelation in CaCl2 solutions. The mechanical strength and nanostructure of the hydrogel beads prepared were determined using compression tests, synchrotron small angle X-ray scattering (SAXS), and cryo-scanning electron microscopy (Cryo-SEM). Interestingly, the application of HPP significantly improved the gel strength of SHP, in agreement with the lower M/G values and the stiffer rod-like conformation obtained for these samples.

Compositional differences of ß-glucan-rich extracts from three relevant mushrooms obtained through a sequential extraction protocol.

In this work, a sequential fractionation protocol (cold and hot aqueous and alkaline extractions) and detailed compositional analysis (gross composition, monosaccharide analysis, FTIR, TGA) was applied to three relevant mushrooms in terms of global production (Grifola frondosa, Lentinula edodes and Pleurotus ostreatus) aiming to understand what is preferentially extracted during fractionation and how ß-glucan extraction is affected by mushroom source. Room temperature aqueous extracts showed highest overall yields (56.3-82%) consisting of proteins, sugars and polyphenols. ß-glucan content was highest in P. ostreatus and was concentrated in the more soluble fractions. On the contrary, a recalcitrant ß-glucan in G. frondosa was mainly present in the residue (7.38%). L. edodes showed ß-glucan populations distributed along aqueous and alkaline extracts, higher abundance of non-glucan polysaccharides and higher chitin purity (47.78%) in the residue. This work sets the basis for the rational design of extraction processes aiming to valorise mushroom biomass.

Gaining control of bacterial cellulose colonization by polyhydroxyalkanoate-producing microorganisms to develop bioplasticized ultrathin films.

Synergistic methodological strategies based on the fields of microbial biotechnology and materials science open up an enormous range of possibilities for the sustainable production of advanced materials with predictable properties. This study shows how naturally produced polyhydroxyalkanoate (PHA) particles are introduced into bacterial cellulose (BC) driven by their bacterial producers. Thanks to an extensive knowledge of the internal structure of BC, it was possible to control the colonization process, i.e. loading and localization of PHA. A subsequent acid treatment favored the PHA-BC bonding at the position reached by the bacteria. These biodegradable films showed improved mechanical and barrier properties even with respect to reference plastic films 8 times thicker, reaching a Young's modulus 4.25 times higher and an oxygen permeability 3 times lower than those of polyethylene terephthalate (PET) films. Owing to the versatility of the method, a wide variety of materials can be developed for very diverse fields of application.

Edible oleogels based on high molecular weight oleogelators and its prospects in food applications.

Food industry is actively looking for alternative ingredients to replace saturated and trans fats in foods while preserving their original organoleptic attributes to ensure consumers' acceptance. A plausible approach is the replacement of solid fats with oleogels. Oleogels can be engineered to mimic properties that are commonly played by regular solid fats but using hydrophobic liquid vegetable oil with an optimum fatty acid profile and, they can also act as carriers for lipophilic bioactive substance. Low molecular weight oleogelators (LMOGs) are well studied and reviewed. In contrast, high molecular weight oleogelators (HMOGs) e.g., polysaccharides and proteins, are not fully researched yet. This review focusses on development of HMOG oleogels produced by means of emulsion templated, direct dispersion, foam templated and solvent exchange methods that can influence the stability, physicochemical properties and their potential application in food industry. Multi-component oleogels can solve the inefficiencies in a single component oleogel and, thus, combinations of HMOGs and HMOGs & LMOGs can produce oleogels with desired properties. These new oleogels can find application as fat substitutes in food products, providing better nutritional and sensory acceptance. A comprehensive overview of recent developments in the field of HMOG and multicomponent oleogels with HMOG is deeply reviewed.

Electrosprayed Agar Nanocapsules as Edible Carriers of Bioactive Compounds.

Electrosprayed agar nanocapsules were developed using an acetic acid solution as solvent. The role of solution properties (viscosity, surface tension, and conductivity) in the formation of agar particles was assessed, together with the effect of both agar and acetic acid concentrations on the size and morphology of the resulting particles. Agar solutions with a concentration below 10% w/v were not suitable for electrospraying. Furthermore, the agar-acetic acid ratio was also critical for the formation of agar nanostructures (with an optimum ratio of 1:2). A decrease in particle size was also observed when decreasing agar concentration, with particle diameter values ranging between 50 and 400 nm. Moreover, the suitability of the electrosprayed agar nanocapsules as carriers for a model bioactive compound, chlorophyllin sodium copper salt (CHL), was also evaluated. The release profile of encapsulated CHL, with an estimated encapsulation efficiency of around 40%, was carried out in food simulants with different hydrophilicity (10% v/v and 50% v/v ethanol). While the release of the bioactive was negligible in the hydrophilic food simulant, an initial burst release followed by a slower sustained release was observed when the capsules were immersed in 50% ethanol solution. The results open up a broad range of possibilities that deserve further exploration related to the use of these edible polysaccharide-based nanocapsules.

Alginate industrial waste streams as a promising source of value-added compounds valorization.

The alginate industry processes more than hundred thousand tons per year of algae in Europe, discarding around 80% of the algae biomass as different solid/liquid residual streams. In this work, Saccharina latissima and Ascophyllum nodosum, their generated alginates and all residual fractions generated in the process were characterized in terms of lipid, ash, protein content, and the carbohydrate composition and antioxidant capacities analyzed. The first fraction after acid treatment (ca. 50% of the initial dry biomass) was rich in phlorotannins (15 mg GAE/g) and bioactive fucoidans (15-70%), with a high sulfation degree in A. nodosum. Two fractions generated from the solid residue, one soluble and another insoluble (Ra and Rb, respectively), constituted 9% and 5-8% of the initial biomass and showed great potential as a source of soluble protein (30% for S. latissima), and cellulose (70%) or fucoidan, respectively. Valorization strategies are suggested for these waste streams, evidencing their high potential as bioactive, texturizing or nutritional added-value ingredients for cosmetic, food, feed or pharmaceutical applications.

Grifola frondosa (Maitake) Extract Reduces Fat Accumulation and Improves Health Span in C. elegans through the DAF-16/FOXO and SKN-1/NRF2 Signalling Pathways.

In recent years, food ingredients rich in bioactive compounds have emerged as candidates to prevent excess adiposity and other metabolic complications characteristic of obesity, such as low-grade inflammation and oxidative status. Among them, fungi have gained popularity for their high polysaccharide content and other bioactive components with beneficial activities. Here, we use the C. elegans model to investigate the potential activities of a Grifola frondosa extract (GE), together with the underlying mechanisms of action. Our study revealed that GE represents an important source of polysaccharides and phenolic compounds with in vitro antioxidant activity. Treatment with our GE extract, which was found to be nongenotoxic through a SOS/umu test, significantly reduced the fat content of C. elegans, decreased the production of intracellular ROS and aging-lipofuscin pigment, and increased the lifespan of nematodes. Gene expression and mutant analyses demonstrated that the in vivo anti-obesity and antioxidant activities of GE were mediated through the daf-2/daf-16 and skn-1/nrf-2 signalling pathways, respectively. Taken together, our results suggest that our GE extract could be considered a potential functional ingredient for the prevention of obesity-related disturbances.

Bioactive extracts from persimmon waste: influence of extraction conditions and ripeness.

In this work, a bioactive persimmon extract was produced from discarded fruits. A central composite design was used to evaluate the effect of different extraction parameters and ripeness stages of persimmon fruits on the total phenolic content and antioxidant activity of the resulting extracts. Significantly greater phenolic contents were obtained from immature persimmon (IP) fruits. The optimum IP extract with the conditions set by the experimental design was industrially up-scaled and its composition and functional properties were evaluated and compared with those obtained under lab-scale conditions. Both extracts contained significant protein (>20%) and phenolic contents (∼11-27 mg GA/g dry extract) and displayed significant antiviral activity against murine norovirus and hepatitis A virus. Moreover, the extract showed no toxicity and significantly reduced the fat content and the cellular ageing of Caenorhabditis elegans (C. elegans) without affecting the worm development. These effects were mediated by down-regulation of fat-7, suggesting an anti-lipogenic activity of this extract.

Modelling the Extraction of Pectin towards the Valorisation of Watermelon Rind Waste.

Watermelon is the second largest fruit crop worldwide, with great potential to valorise its rind waste. An experimental design was used to model how extraction parameters (temperature, pH, and time) impact on the efficiency of the process, purity, esterification degree, monosaccharide composition and molar mass of watermelon rind pectin (WRP), with an insight on changes in their structural properties (linearity, branching degree and extraction severity). The models for all responses were accurately fitted (R2 > 90%, lack of fit p ≥ 0.05) and experimentally validated. At optimum yield conditions, WRP yield (13.4%), purity (540 µg/g galacturonic acid) and molar mass (106.1 kDa) were comparable to traditional pectin sources but showed a higher branching degree with longer galactan side chains and a higher protein interaction. Harsher conditions (pH 1) generated purer homogalacturonan fractions with average molar masses (80 kDa) at the expense of yield, while mild extraction conditions (pH ≥ 2) produced highly branched entangled pectin structures. This study underlines novel compositional features in WRP and the possibility of producing novel customized pectin ingredients with a wider potential application scope depending on the targeted structure.

On the Use of Persian Gum for the Development of Antiviral Edible Coatings against Murine Norovirus of Interest in Blueberries.

In the last decades, berries have been identified as important vehicles for the transmission of foodborne viruses and different strategies are being explored to eliminate or reduce viral contamination in these fruits. The aim of this work was to develop novel edible coatings with antiviral properties for inactivating and reducing murine norovirus (MNV). Firstly, the effect of gelatin (G) addition on Persian gum (PG) films was studied in terms of microstructural, mechanical, optical, and water barrier properties. The following PG:G ratios were considered: 100:0, 75:25, 50:50, 25:75, and 0:100. Microstructure analysis revealed the compatibility of both hydrocolloids since no phase separation was observed. The addition of G to PG films provided stiffer and more deformable films than pure PG, with lower water vapor permeability values. Specifically, films prepared with 50:50 PG:G ratio presented better mechanical and barrier performance. Interestingly, pure PG showed antiviral activity on murine norovirus, probably due to the presence of some impurities (mainly tannins). Adding allyl isothiocyanate (AITC) enhanced the PG antiviral activity at refrigerated temperatures in blueberries, not being affected by the AITC concentration. This effect was not observed at ambient temperature, probably due to the volatilization of AITC.

Improved Process to Obtain Nanofibrillated Cellulose (CNF) Reinforced Starch Films with Upgraded Mechanical Properties and Barrier Character.

Nowadays, the interest on nanofibrillated cellulose (CNF) has increased owing to its sustainability and its capacity to improve mechanical and barrier properties of polymeric films. Moreover, this filler shows some drawbacks related with its high capacity to form aggregates, hindering its dispersion in the matrix. In this work, an improved procedure to optimize the dispersability of CNF in a thermoplastic starch was put forward. On the one hand, CNF needs a hydrophilic dispersant to be included in the matrix, and on the other, starch needs a hydrophilic plasticizer to obtain a thermoformable material. Glycerol was used to fulfil both targets at once. CNF was predispersed in the plasticizer before nanofibrillation and later on was included into starch, obtaining thin films. The tensile strength of these CNF-starch composite films was 60% higher than the plain thermoplastic starch at a very low 0.36% w/w percentage of CNF. The films showed a noticeable correlation between water uptake, and temperature and humidity. Regarding permeability, a ca. 55% oxygen and water vapor permeability drop was found by nanofiller loading. The hydrolytic susceptibility of the composite was confirmed, being similar to that of the thermoplastic starch.

Recyclable thermophilic hybrid protein-inorganic nanoflowers for the hydrolysis of milk lactose.

Thermostable ß-galactosidase (TmLac) has been immobilized as hybrid inorganic-protein nanoflowers using salts of Cu2+, Mn2+, Zn2+, Co2+ and Ca2+ as the inorganic component. The incorporation efficiency of enzyme into the nanoflowers was higher than 95% for a protein concentration of 0.05 mg/mL. The structure, activity and recyclability of the nanoflowers with different chemical composition were analyzed. Ca2+, Mn2+ and Co2+ nanoflowers showed a level of lactase activity equivalent to their same content of free enzyme. Cu2+nanoflowers showed only marginal enzyme activity in agreement with the inhibitory effect of this cation on the enzyme. TmLac nanoflowers provide an efficient methodology for enzyme immobilization and recyclability. TmLac-Ca2+ nanoflowers presented the best properties for lactose hydrolysis both in buffered and in milk, and could be reused in five consecutive cycles.

Food Packaging Based on Nanomaterials.

The use of nanomaterials for food applications is a rapidly evolving field and, given the specific properties of nanomaterials and their tremendous potential, an increased number of material innovations that contribute to improved food quality and safety are foreseen [...].

Development of enzymatically-active bacterial cellulose membranes through stable immobilization of an engineered ß-galactosidase.

Enzymatically-active bacterial cellulose (BC) was prepared by non-covalent immobilization of a hybrid enzyme composed by a ß-galactosidase from Thermotoga maritima (TmLac) and a carbohydrate binding module (CBM2) from Pyrococcus furiosus. TmLac-CBM2 protein was bound to BC, with higher affinity at pH 6.5 than at pH 8.5 and with high specificity compared to the non-engineered enzyme. Both hydrated (HBC) and freeze-dried (DBC) bacterial cellulose showed equivalent enzyme binding efficiencies. Initial reaction rate of HBC-bound enzyme was higher than DBC-bound and both of them were lower than the free enzyme. However, enzyme performance was similar in all three cases for the hydrolysis of 5% lactose to a high extent. Reuse of the immobilized enzyme was limited by the stability of the ß-galactosidase module, whereas the CBM2 module provided stable attachment of the hybrid enzyme to the BC support, after long incubation periods (3 h) at 75 °C.

Structural and physicochemical characterization of thermoplastic corn starch films containing microalgae.

This work provides an in-depth analysis on how the addition of different microalgae species (Nannochloropsis, Spirulina and Scenedesmus) affected the structural and physicochemical properties of thermoplastic corn starch biocomposites. Structural characterization was conducted by combined SAXS/WAXS experiments and it was correlated with mechanical and barrier properties of the biocomposites. A water vapour permeability drop of ca. 54% was observed upon addition of the different microalgae species. The oxygen permeability and the mechanical properties of biocomposites containing Spirulina or Scenedesmus were not improved since the presence of microalgae hindered the re-arrangement and packing of the lamellar structure of starch polymeric chains, according to the SAXS results. Nannochloropsis caused a great reduction of the matrix rigidity and, the oxygen permeability was also improved. Therefore, all of these features make the Nannochloropsis biocomposites an alternative to generate biodegradable food packaging materials with the additional advantage that they can be easily scaled-up.

Self-assembled gelatin-ι-carrageenan encapsulation structures for intestinal-targeted release applications.

In this work, natural biopolymeric encapsulation structures were developed through the self-assembly of gelatin and ι-carrageenan in aqueous solutions. The interactions of this binary system and of a ternary system containing a polyphenol-rich extract were deeply explored for the development of intestinal delivery systems. The processing of the structures (extrusion vs. freeze-drying) greatly influenced release properties, explained by the specific interactions between gelatin and polyphenols, thus allowing for tuning the processing conditions depending on the desired target application. Release was further controlled by incorporating a divalent salt, giving raise to extract-loaded ι-carrageenan/gelatin capsules with adequate release profiles for intestinal targeted delivery. These results demonstrate the potential of exploiting biopolymer interactions for designing bioactive delivery systems using environmentally friendly processes which do not involve the use of toxic or harsh solvents or cross-linkers.