Enrichment of bovine milk-derived extracellular vesicles using surface-functionalized cellulose nanofibers

The isolation of extracellular vesicles (EVs) from milk, a complex mixture of colloidal structures having a comparable size to EVs, is challenging. Although ultracentrifugation (UC) has been widely used for EV isolation, this has significant limitations, including a long processing time at high g-force conditions and large sample volume requirements. We introduced a new approach based on nature nanoentities cellulose nanofibers (CNFs) and short time and low g-force centrifugation to isolate EVs from various milk fractions. The flexible and entangled network of CNFs forms nanoporous, which entraps the EVs. Further, positively charged CNFs interact with anionic EVs through an electrostatic attraction, promoting their isolation with efficiency comparable with UC. The functionality and toxicity of isolated milk EVs were tested in Caco2 cells. Overall, the newly developed approach provides straightforward isolation and biocompatibility and preserves the natural properties of the isolated EVs, enabling further applications

Syväeutektisen liuottimen kierrätys nanofibrilloidun selluloosan valmistuksessa

Syväeutektiset liuottimet (deep eutectic solvents, DESs) ovat uudenlaisia haihtumattomia, vihreitä ja monipuolisia nesteitä, jotka koostuvat kahdesta tai useammasta usein edullisesta ja biohajoavasta komponentista. Niiden avulla pyritään löytämään uusia kestävän kehityksen mukaisia ratkaisuja teollisuuden tarpeisiin ja vähentämään haihtuvien orgaanisten liuottimien käyttöä. Selluloosananomateriaalit ovat luonnonmateriaaleja, joissa yhdistyvät selluloosan ja nanokokoisen aineen erinomaiset ominaisuudet. Niitä voidaan käyttää mm. parantamaan komposiittien ja hybridimateriaalien lujuusominaisuuksia, korvaamaan muovin käyttöä ja muodostamaan erilaisia muita rakenteita. DES:sien käyttäminen nanoselluloosan valmistuksessa on osoittautunut mielenkiintoiseksi vaihtoehdoksi perinteisille kemiallisille ja entsymaattisille esikäsittelyille. Selluloosan esikäsittelyä tarvitaan löystyttämään kuitujen lujaa rakennetta ja rikkomaan fibrillien välisiä vetysidoksia, jolloin energiaintensiivisen mekaanisen käsittelyn tarve vähentyy. DES:seihin perustava esikäsittely on yksinkertainen ja edullinen. Esimerkiksi kuitua voidaan käsitellä 100 °C:ssa koliinikloridi-urea moolisuhteessa 1:2) DES:sissä 2 tuntia, jonka jälkeen DES-liuos erotetaan kuidusta suodattamalla ja esikäsitelty kuitu hajotetaan mekaanisesti edelleen selluloosananofibrilleiksi. Valmistetusta nanoselluloosasta voidaan valmistaa esimerkiksi ohuita ja läpinäkyviä filmejä, jotka omaavat erinomaiset mekaaniset ominaisuudet. DES:sin kierrätys ja uudelleenkäyttö ovat vielä vähän tutkittuja mutta tärkeitä aiheita, joihin tämä työ keskittyy. Tutkimuksessa selvitettiin koliinikloridi-urea DES:sin kierrätettävyyttä selluloosananofibrillien valmistusprosessissa. Työssä tutkittiin erityisesti käytetyn DES:sin koostumuksen ja ominaisuuksien sekä käsitellyn kuidun ja selluloosananofibrillien ominaisuuksien muutoksia. DES:sin ominaisuuksien muutoksista helpoimmin havaittava oli sen värin muuttuminen värittömästä kellertäväksi. Tämä johtui todennäköisesti ¹H NMR spektroskopialla havaitusta karbakoliinin muodostumisesta esikäsittelyn yhteydessä. Havaintoa vahvisti myös massaspektrometrin ja nestekromatografin tulokset. Viiden esikäsittelysyklin jälkeen karbakoliinin laskettiin muodostavan 2,77 % mooliosuuden DES:sistä. Koostumuksen muutoksesta huolimatta DES:sin fysikaalis-kemialliset ominaisuudet eivät kuitenkaan merkittävästi muuttuneet ja sen lämpökäyttäytyminen pysyi samankaltaisena tutkittujen viiden kierrätyskerran jälkeen. DES:sin kierrätyksellä ei havaittu olevan vaikutuksia kuidun fibrilloitavuuteen, sillä kuitunäytteiden ominaisuuksissa ei tapahtunut merkittäviä muutoksia kierrätyssyklien lisääntyessä eikä kuidun mekaanisessa hajottamisessa ollut eroja. Selluloosananofibrilleistä valmistetut filmit olivat ulkoisesti vastaavia, eikä niiden mekaanisissa ominaisuuksissa ollut tilastollisesti eroja. Tutkimuksen tuloksena voidaan todeta koliinikloridi-urea DES:sin olevan kierrätettävissä ilman erillisiä puhdistusvaiheita useita kertoja selluloosan esikäsittelyssä, eikä kierrätyksellä havaittu olevan vaikutusta kuitujen ominaisuuksiin tai selluloosananofibrillifilmien laatuun tutkituilla kierrätysasteilla. DES:siin perustuva selluloosan esikäsittely on siis lupaava vihreän kemian periaatteiden mukainen menetelmä selluloosananofibrillien valmistamiseksi.Deep eutectic solvents (DESs) are novel, non-volatile, green and multifunctional compounds composing of two or more inexpensive, natural and biodegradable components. They are used in accordance with sustainable development to find new solutions for industrial needs and to reduce the use of the volatile organic solvents. Cellulose nanomaterials are natural materials combining the excellent properties of both cellulose and nano materials. Among other things, they can be used to improve strength properties of composite and hybrid materials, replace plastics use and form various other structures. Utilization of DESs in fabrication of cellulose nanomaterials has been proved to be an interesting alternative for more traditional chemical and enzymatic pretreatments. The cellulose pretreatments of cellulose are used for loosening the fiber structure and for the degrading of interfibrillar hydrogen bonding network. The weakened structure requires in turn less energy during the mechanical fibrillation of the cellulose fiber. Pretreatment method based on DESs is straightforward and economical. For example, the fibers can be treated two hours in 100 °C choline chloride-urea (molar ratio 1:2) DES, after which the fibers are separated from DES by filtration and further mechanically degraded into individual cellulose nanofibrils. Thin and transparent films possessing excellent tensile properties can be produced from those nanofibrils. Recycling of DESs is yet underresearched but important subject. In this work, the recyclability of choline chloride-urea DES is demonstrated in the production of the cellulose nanofibrils. The effect of the recycling was studied by analyzing the changes in composition and properties of DES and studying the properties of the fibers and cellulose nanofibrils. Of the DES property changes the easiest one to observe was the colour shifting from transparent to yellow. This was most probably resulting from formation of carbacholine during the DES pretreatment and it was seen ¹H NMR samples. The finding was further confirmed with the results from mass spectrometry and liquid chromatography. After five pretreatment cycles carbacholine was calculated to form 2,77 % mole fraction of the whole DES. Despite subtle changes in composition, DES’s physico-chemical properties didn’t significantly change and it’s thermal behaviour remained similar. The DES recycling wasn’t seen to have any effect on the fibrillation of the fiber. The properties of fiber samples remained practically the same as recycling cycles increased and there were no differences in mechanical disintegration. The films fabricated from the cellulose nanofibrils were similar in appearance and their tensile properties didn’t statistically vary. As a result of this research it can be stated that choline chloride-urea DES can be recycled several times without any separate purification steps in the pretreatment of the cellulose. Furthermore the recycling doesn’t have effect on the fiber properties or the quality of the films fabricated from cellulose nanofibrils after studied five recycling cycles. Consequeently, the DES mediated pretreatment of cellulose is a promising method in production of cellulose nanofibrils and is in agreement with the principles of the green chemistry

Dikarboksyyliselluloosan valmistuksen optimointi

Peräkkäinen perjodaatti-kloriittihapetus on osoittautunut toimivaksi esikäsittelymenetelmäksi valmistettaessa selluloosananofibrillejä. Menetelmässä valkaistu selluloosamassa hapetetaan ensin natriumperjodaatilla dialdehydiselluloosaksi ja sen jälkeen natriumkloriitilla dikarboksyyliselluloosaksi. Kloriittihapetusvaihe vaatii menetelmän mukaisesti pitkän 48 tunnin reaktioajan ja huomattavan ylimäärän kloriittia suhteessa selluloosan aldehydimäärään. Tässä työssä tutkitaan, miten natriumkloriitin määrän vähentäminen ja reaktioajan lyhentäminen vaikuttavat karboksyyliryhmien määrään dikarboksyyliselluloosassa. Teoriaosiossa käsitellään selluloosan rakennetta, ominaisuuksia ja siitä muodostuvia suurempia rakenteita, fibrillejä. Lisäksi perehdytään selluloosan kemiallisiin reaktioihin. Työn kokeellisessa osiossa tutkitaan kloriittihapetusta kahdella koesarjalla, joista ensimmäisessä tarkastellaan natriumkloriitin ylimäärän vaikutusta karboksyyliryhmien ja jäännösaldehydin määrään 60 minuutin reaktioajalla. Toisessa koesarjassa varmistetaan ensin propionaldehydin toimivuus reaktion pysäyttäjänä ja tuotetaan näytesarja, jossa käytetään 2,5-kertaista natriumkloriitin ylimäärää ja lyhennetään reaktioaikaa 60 minuutista 15 sekuntiin. Tuloksista nähdään näytteiden karboksyyliryhmien määrän pienenevän selkeästi, kun kloriitin ylimäärää reaktiossa vähennetään. Samalla jäännösaldehydin määrä lisääntyy teorian mukaisesti, molempien summan pysyessä lähes vakiona. Kloriittihapetus osoittautuu huomattavasti oletettua nopeammaksi reaktioksi, sillä jo neljässä minuutissa karboksyyliryhmien määrä saavuttaa yli 70 % tason verrattuna dialdehydiselluloosan kokonaisaldehydimäärään. Huomionarvoinen piirre reaktiossa on karboksyyliryhmäpitoisuuden vakiintuminen lähes samalla tasolle ainakin 60 minuuttiin asti. Dikarboksyyliselluloosan karboksyyliryhmien määrään on mahdollisesti helpointa vaikuttaa natriumkloriitin ylimäärän avulla, käyttäen samalla kohtuullisen pitkiä reaktioaikoja

Direct sulfation of cellulose fibers using a reactive deep eutectic solvent to produce highly charged cellulose nanofibers

Abstract Wood cellulose pulp was sulfated using a reactive deep eutectic solvent (DES). DES was prepared by heating sulfamic acid and urea together at 80 °C at a molar ratio of 1:4, 1:3, or 1:2. Sulfation of cellulose was performed by mixing dry cellulose fibers with DES at 80 °C, followed by heating at 150 °C for half an hour. Anionic charge as high as 3 mmol/g (degree of substitution of 0.68) was obtained with this simple chemical modification of cellulose at an elevated temperature using DES both as reaction media and reagent without any external solvent. The decrease in the urea content of DES improved the sulfation efficiency. In addition, the presence of urea led to the carbamation of cellulose to some extent. Cellulose sulfate (charge of 2.40 mmol/g) became a gel-like material in water, and after passing once through a microfluidizator, a highly transparent nanocellulose gel (transmittance of 0.1% solution at a visible light range was over 95%) was obtained. Sulfated cellulose nanofibers (SCNFs) exhibited a width of around 4 nm with a minor presence of elemental fibril aggregates (containing five or less elemental fibrils). SCNFs with high aspect ratio can have a potential end-use as a rheology modifier because of their high viscosity even at low concentrations or act as reinforcing additives

Effect of plasticizers on the mechanical and thermomechanical properties of cellulose-based biocomposite films

Abstract Biocomposites based on natural cellulose fibers (CF) and hydroxyethyl cellulose (HEC), were produced in the form of green packaging films. The effect of the different single-component plasticizers (glycerol, propylene carbonate and ethylene carbonate) on the mechanical and dynamic thermomechanical properties of the films were studied. Moreover, the softening effect of the two-component plasticizer based on deep eutectic solvents (DESs) was addressed. Of the single-component plasticizers, glycerol was found to be the most efficient by increasing the elongation at break of the composite by 53%. A similar, or even better, increase in elongation at break (up to 81%) was obtained with DESs based on choline chloride and glycerol, glucose or urea. Based on the dynamic mechanical analysis at varying humidity, the performance of plasticizers was strongly attributed to the humidity. The DES based on tetrabutylammonium bromide and propylene carbonate was most efficient at providing thermoformability to the composite by lowering the thermal softening temperature. Based on the obtained results, DESs are a highly promising plasticizers for the cellulose-based biocomposites with similar or even better plasticizing effect compared to conventional plasticizer. In addition, DESs can be used to improve the thermoformability of biocomposites, by lowering the thermal softening temperature

Solid air-low temperature manufacturing of ultra-low permittivity composite materials for future telecommunication systems

Abstract The frequency spectrum to be used by future wireless telecommunication systems such as 5G and beyond requires novel materials which are environment-friendly, are low cost and, most importantly, have low dielectric loss and permittivity when approaching higher frequencies. In this work, the development of all-inorganic composites with a relative permittivity of ~1.2 and loss tangents in the range of 10−3 is presented. The composites were fabricated at the exceptionally low temperature of 120°C and were based on lithium molybdate (Li2MoO4) ceramic as a water-soluble binder reinforced by quartz fibers. The relative permittivity was further decreased by the addition of hollow micron-sized glass spheres having very low dielectric loss. A simple manufacturing method through filtration, stencil printing and drying is presented. The microstructure of the composites was investigated with FESEM microscopy and the dielectric properties by SPDR. Printing tests were carried out in order to evaluate the possibility of using the proposed composites in, for example, printed antenna applications

Wood-based composite materials for ultralight lens antennas in 6G systems

Abstract Extremely high frequencies used in future wireless communication systems such as 6G require low loss materials to avoid wasting power and maintain acceptable efficiency. Furthermore, especially in the internet of things (IoT) applications, low weight and the possibility to focus the radiation pattern in the desired direction would also improve the communication between units and reduce the required signal power. Radio frequency (RF) lenses for signal focusing can be made from low loss and low relative permittivity materials. In this work, the development of sustainable composites with a relative permittivity of ∼1.15 and loss tangents in the range of 10⁻³ is presented. The composites were fabricated at the exceptionally low temperature of 95 °C and were based on hollow micron-sized glass spheres and three different types of cellulose nanofibers as the water-soluble binder. A simple manufacturing method through casting and drying is presented. The surface properties of the composites were investigated with surface profile analysis and the dielectric properties by SPDR and terahertz spectroscopy. The weight of the fabricated lens was 0.6 g and the transmittance was 99.85%. The lens improved the antenna gain by 14–18 dBi depending on the surface smoothening. Also, coating the lens with a moisture protecting agent did not alter the lens performance

Enrichment of bovine milk-derived extracellular vesicles using surface-functionalized cellulose nanofibers

Abstract The isolation of extracellular vesicles (EVs) from milk, a complex mixture of colloidal structures having a comparable size to EVs, is challenging. Although ultracentrifugation (UC) has been widely used for EV isolation, this has significant limitations, including a long processing time at high g-force conditions and large sample volume requirements. We introduced a new approach based on nature nanoentities cellulose nanofibers (CNFs) and short time and low g-force centrifugation to isolate EVs from various milk fractions. The flexible and entangled network of CNFs forms nanoporous, which entraps the EVs. Further, positively charged CNFs interact with anionic EVs through an electrostatic attraction, promoting their isolation with efficiency comparable with UC. The functionality and toxicity of isolated milk EVs were tested in Caco2 cells. Overall, the newly developed approach provides straightforward isolation and biocompatibility and preserves the natural properties of the isolated EVs, enabling further applications