| 1. |
- Boge, Lukas, 1987, et al.
(författare)
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Cubosomes for topical delivery of the antimicrobial peptide LL-37
- 2019
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Ingår i: European Journal of Pharmaceutics and Biopharmaceutics. - : Elsevier BV. - 1873-3441 .- 0939-6411. ; 134, s. 60-67
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Tidskriftsartikel (refereegranskat)abstract
- In this study, the use of cubosomes for topical delivery of the antimicrobial peptide (AMP) LL-37 was investigated. Topical delivery of AMPs is of great interest for treatment of skin infections caused by bacteria, such as Staphylococcus aureus. AMP containing cubosomes were produced by three different preparation protocols and compared: (i) pre-loading, where LL-37 was incorporated into a liquid crystalline gel, which thereafter was dispersed into nanoparticles, (ii) post-loading, where LL-37 was let to adsorb onto pre-formed cubosomes, and (iii) hydrotrope-loading, where LL-37 was incorporated during the spontaneously formed cubosomes in an ethanol/glycerol monooleate mixture. Particle size and size distribution were analyzed using dynamic light scattering (DLS), liquid crystalline structure by small angle x-ray scattering (SAXS) and release of LL-37 by a fluorescamine assay. Proteolytic protection of LL-37 as well as bactericidal effect after enzyme exposure was investigated. The skin irritation potential of cubosomes was examined by an in vitro epidermis model. Finally, the bacterial killing property of the cubosomes was examined by an ex vivo pig skin wound infection model with Staphylococcus aureus. Data showed that a high loading of LL-37 induced formation of vesicles in case of cubosomes prepared by sonication (pre-loading). No release of LL-37 was observed from the cubosomes, indicating strong association of the peptide to the particles. Proteolysis studies showed that LL-37 was fully protected against enzymatic attacks while associated with the cubosomes, also denoting strong association of the peptide to the particles. As a consequence, bactericidal effect after enzyme exposure remained, compared to pure LL-37 which was subjected to proteolysis. No skin irritation potential of the cubosomes was found, thus enabling for topical administration. The ex vivo wound infection model showed that LL-37 in pre-loaded cubosomes killed bacteria most efficient.
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| 2. |
- Boge, Lukas, 1987, et al.
(författare)
-
Cubosomes post-loaded with antimicrobial peptides: Characterization, bactericidal effect and proteolytic stability
- 2017
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Ingår i: International Journal of Pharmaceutics. - : Elsevier BV. - 0378-5173 .- 1873-3476. ; 526:1-2, s. 400-412
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Tidskriftsartikel (refereegranskat)abstract
- Novel antibiotics, such as antimicrobial peptides (AMPs), have recently attended more and more attraction. In this work, dispersed cubic liquid crystalline gel (cubosomes) was used as drug delivery vehicles for three AMPs (AP114, DPK-060 and LL-37). Association of peptides onto cubosomes was studied at two cubosome/peptide ratios using high performance liquid chromatography, ?-potential and circular dichroism measurements. AMPs impact on the cubosome structure was investigated using small angle x-ray scattering and cryogenic transmission electron microscopy. The antimicrobial effect of the AMP loaded cubosomes was studied in vitro by minimum inhibitory concentration and time-kill assays. Proteolytic protection was investigated by incubating the formulations with two elastases and the antimicrobial effect after proteolysis was studied using radial diffusion assay. Different association efficacy onto the cubosomes was observed among the AMPs, with LL-37 showing greatest association (>60%). AP114 loaded cubosomes displayed a preserved antimicrobial effect, whereas for LL-37 the broad spectrum bacterial killing was reduced to only comprise Gram-negative bacteria. Interestingly, DPK-060 loaded cubosomes showed a slight enhanced effect against S. aureus and E. coli strains. Moreover, the cubosomes were found to protect LL-37 from proteolytic degradation, resulting in a significantly better bactericidal effect after being subjected to elastase, compared to unformulated peptide.
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| 3. |
- Boge, Lukas, et al.
(författare)
-
Cubosomes for topical delivery of the antimicrobial peptide LL-37
- 2019
-
Ingår i: European journal of pharmaceutics and biopharmaceutics. - : Elsevier BV. - 0939-6411 .- 1873-3441. ; 134, s. 60-67
-
Tidskriftsartikel (refereegranskat)abstract
- In this study, the use of cubosomes for topical delivery of the antimicrobial peptide (AMP) LL-37 was investigated. Topical delivery of AMPs is of great interest for treatment of skin infections caused by bacteria, such as Staphylococcus aureus. AMP containing cubosomes were produced by three different preparation protocols and compared: (i) pre-loading, where LL-37 was incorporated into a liquid crystalline gel, which thereafter was dispersed into nanoparticles, (ii) post-loading, where LL-37 was let to adsorb onto pre-formed cubosomes, and (iii) hydrotrope-loading, where LL-37 was incorporated during the spontaneously formed cubosomes in an ethanol/glycerol monooleate mixture. Particle size and size distribution were analyzed using dynamic light scattering (DLS), liquid crystalline structure by small angle x-ray scattering (SAXS) and release of LL-37 by a fluorescamine assay. Proteolytic protection of LL-37 as well as bactericidal effect after enzyme exposure was investigated. The skin irritation potential of cubosomes was examined by an in vitro epidermis model. Finally, the bacterial killing property of the cubosomes was examined by an ex vivo pig skin wound infection model with Staphylococcus aureus. Data showed that a high loading of LL-37 induced formation of vesicles in case of cubosomes prepared by sonication (pre-loading). No release of LL-37 was observed from the cubosomes, indicating strong association of the peptide to the particles. Proteolysis studies showed that LL-37 was fully protected against enzymatic attacks while associated with the cubosomes, also denoting strong association of the peptide to the particles. As a consequence, bactericidal effect after enzyme exposure remained, compared to pure LL-37 which was subjected to proteolysis. No skin irritation potential of the cubosomes was found, thus enabling for topical administration. The ex vivo wound infection model showed that LL-37 in pre-loaded cubosomes killed bacteria most efficient.
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| 4. |
- Boge, Lukas, et al.
(författare)
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Freeze-dried and re-hydrated liquid crystalline nanoparticles stabilized with disaccharides for drug-delivery of the plectasin derivative AP114 antimicrobial peptide
- 2018
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Ingår i: Journal of Colloid and Interface Science. - : Elsevier BV. - 0021-9797 .- 1095-7103. ; 522, s. 126-135
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Tidskriftsartikel (refereegranskat)abstract
- Liquid crystalline nanoparticles (LCNPs), e.g. cubosomes and hexosomes, are receiving more and more attraction as drug delivery vehicles. Dry powder formulation that forms LCNPs upon hydration can be advantageous to make new routes of administration accessible. In this work, we investigate use of three disaccharides (lactose, trehalose and sucrose) as protective matrices for glycerol monooleate based LCNP forming powders produced by freeze-drying. Phase behavior, particle size and size distributions at the different preparation steps were monitored by small angle x-ray scattering (SAXS) and dynamic light scattering (DLS). Particle appearance was imaged by cryogenic transmission electron microscopy (cryo-TEM). Moreover, the therapeutic relevant antimicrobial peptide AP114 (plectasin derivative) was incorporated in the formulations. Peptide encapsulation and release as well as in vitro antibacterial effect were investigated. Results showed that all freeze-dried powders did form particles with liquid crystalline structure upon hydration. However, a phase transition from the bicontinuous cubic Pn3m to the reversed hexagonal was observed, as a consequence of sugar addition and the freeze-drying procedure. Data indicates that trehalose is the preferred choice of lyo-protectant in order to maintain a mono-modal particle size distribution. In addition, antimicrobial activity of AP114-containing formulations was found to be highest for the formulation containing trehalose. The release kinetics of AP114 from the nanoparticles was strongly affected by the dimensions of the hexagonal phase. Larger dimension of the hexagonal phase, significantly improved the release of AP114 and antimicrobial activity of the formulation.
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| 5. |
- Boge, Lukas, et al.
(författare)
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Lipid-based liquid crystals as carriers for antimicrobial peptides : Phase behavior and antimicrobial effect
- 2016
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Ingår i: Langmuir. - : American Chemical Society (ACS). - 0743-7463 .- 1520-5827. ; 32:17, s. 4217-4228
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Tidskriftsartikel (refereegranskat)abstract
- The number of antibiotic-resistant bacteria is increasing worldwide, and the demand for novel antimicrobials is constantly growing. Antimicrobial peptides (AMPs) could be an important part of future treatment strategies of various bacterial infection diseases. However, AMPs have relatively low stability, because of proteolytic and chemical degradation. As a consequence, carrier systems protecting the AMPs are greatly needed, to achieve efficient treatments. In addition, the carrier system also must administrate the peptide in a controlled manner to match the therapeutic dose window. In this work, lyotropic liquid crystalline (LC) structures consisting of cubic glycerol monooleate/water and hexagonal glycerol monooleate/oleic acid/water have been examined as carriers for AMPs. These LC structures have the capability of solubilizing both hydrophilic and hydrophobic substances, as well as being biocompatible and biodegradable. Both bulk gels and discrete dispersed structures (i.e., cubosomes and hexosomes) have been studied. Three AMPs have been investigated with respect to phase stability of the LC structures and antimicrobial effect: AP114, DPK-060, and LL-37. Characterization of the LC structures was performed using small-angle X-ray scattering (SAXS), dynamic light scattering, ζ-potential, and cryogenic transmission electron microscopy (Cryo-TEM) and peptide loading efficacy by ultra performance liquid chromatography. The antimicrobial effect of the LCNPs was investigated in vitro using minimum inhibitory concentration (MIC) and time-kill assay. The most hydrophobic peptide (AP114) was shown to induce an increase in negative curvature of the cubic LC system. The most polar peptide (DPK-060) induced a decrease in negative curvature while LL-37 did not change the LC phase at all. The hexagonal LC phase was not affected by any of the AMPs. Moreover, cubosomes loaded with peptides AP114 and DPK-060 showed preserved antimicrobial activity, whereas particles loaded with peptide LL-37 displayed a loss in its broad-spectrum bactericidal properties. AMP-loaded hexosomes showed a reduction in antimicrobial activity.
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| 6. |
- Boge, Lukas
(författare)
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Lipid-based liquid crystals as drug delivery vehicles for antimicrobial peptides
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The development of antimicrobial resistance is a great challenge within health sectors worldwide. Thus, demand for new, efficient treatments is urgent in order to treat various bacterial infections. Antimicrobial peptides (AMPs) are a group of antibiotics that have gained more and more attraction in the past decade. AMPs suffer from relatively low stability due to proteolytic and chemical degradation. As a consequence, carrier systems to protect the AMPs are highly needed to achieve efficient treatments in the clinic. In this thesis, lipid-based liquid crystalline (LC) structures have been examined as carriers for AMPs. LC structures of polar lipids have potential to be used as carriers and delivery systems in various pharmaceutical applications. This is due to their ability to solubilize and encapsulate hydrophilic, hydrophobic and amphiphilic substances. An important feature of these LC systems is that they can coexist with an excess of water, which enables fragmentation of the highly viscous gels into LC nanoparticles (LCNPs), i.e. cubosomes and hexosomes, in the presence of a suitable stabilizer. Peptides and proteins can be incorporated into the lipid self-assembled structures, thereby protecting them from chemical and proteolytic degradation. Cubosomes and hexosomes were investigated as drug delivery vehicles for the three AMPs: i) AP114, an improved plectasin derivative originating from the fungus Pseudoplectania nigrella, ii) DPK-060, derived from the endogenous human protein kininogen and iii) LL-37, a human AMP found in the cathelicidin family. Phase behavior, different preparation methodologies of the LCNPs, antimicrobial effect and proteolytic protection of the AMPs were studied. Moreover, the interaction between AMP-loaded particles with bacteria and bacterial mimicking membranes was investigated. Formulations aimed for pulmonary and topical administration were also evaluated. Results showed that cubic LC phases were most sensitive to the incorporation of AMPs. Depending on the nature of the AMP, different changes in the curvature of the systems were observed. Cubosomes loaded with AMPs exhibited good antimicrobial activity and were found to protect the proteolytic sensitive LL-37 from enzymatic degradation. Data strongly suggested that the release of AMP from the particles cannot solely be explained by the antimicrobial effect. Cubosomes loaded with LL-37 are thought to adsorb onto bacterial membranes, resulting in cell death.
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| 7. |
- Boge, Lukas, 1987
(författare)
-
Lipid-based liquid crystals as drug delivery vehicles for antimicrobial peptides
- 2017
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The development of antimicrobial resistance is a great challenge within the health sector around the world. The demand for new efficient treatments is alarming in order to treat various bacterial infections in the near future. Antimicrobial peptides (AMPs) are a group of novel antibiotics that have gain more and more attraction the past decade. However, AMPs suffers from relatively low stability due to proteolytic and chemical degradation. As a consequence, carrier systems protecting the AMPs are highly needed for achieving efficient treatments. In this thesis, lyotropic liquid crystalline (LC) structures consisting of cubic glycerol monooleate/water and hexagonal glycerol monooleate/oleic acid/water have been examined as carriers for three AMPs (AP114, DPK-060 and LL-37). Both bulk gels and discrete dispersed structures, i.e. cubosomes and hexosomes have been studied. Moreover, two different peptide loading approaches for the cubosomes were tested and compared; pre- and post-loading. Characterization of the LC structures was performed using small-angle x-ray scattering (SAXS), dynamic light scattering, ζ-potential, and cryogenic transmission electron microscopy (Cryo-TEM) and peptide loading efficacy by liquid chromatography. The antimicrobial effect of the AMP loaded LC nanoparticles (LCNPs) was studied in vitro using minimum inhibitory concentration (MIC) and time-kill assays. Proteolytic protection was investigated by incubating the formulations with two elastases and the antimicrobial effect after proteolysis was studied using radial diffusion assay (RDA). Results showed that the most hydrophobic peptide (AP114) was prone to induce an increase in negative curvature of the bulk cubic LC gel, hence pushing the system towards a hexagonal structure. The most polar peptide (DPK-060) induced a decrease in negative curvature while LL-37 did not change the LC phase at all. The hexagonal LC phase was not affected by any of the AMPs. The cubic pre- and post-loaded LCNPs displayed promising antimicrobial activity, and sometimes could a synergetic effect be observed, resulting in a slightly better activity than the unformulated AMP. The hexagonal LCNPs were found to be very efficient in encapsulating the AMPs, but did not display any antimicrobial effect, indicating insufficient delivery of peptide to the bacteria. Moreover, cubosomes post-loaded with LL-37 was found to protect the peptide from proteolytic degradation, resulting in a significant better bactericidal effect after proteolysis.
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| 8. |
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| 9. |
- Boge, Lukas, et al.
(författare)
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Peptide-Loaded Cubosomes Functioning as an Antimicrobial Unit against Escherichia coli
- 2019
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Ingår i: ACS Applied Materials and Interfaces. - : American Chemical Society. - 1944-8244 .- 1944-8252. ; 11:24, s. 21314-21322
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Tidskriftsartikel (refereegranskat)abstract
- Dispersions of cubic liquid crystalline phases, also known as cubosomes, have shown great promise as delivery vehicles for a wide range of medicines. Due to their ordered structure, comprising alternating hydrophilic and hydrophobic domains, cubosomes possess unique delivery properties and compatibility with both water-soluble and -insoluble drugs. However, the drug delivery mechanism and cubosome interaction with human cells and bacteria are still poorly understood. Herein, we reveal how cubosomes loaded with the human cathelicidin antimicrobial peptide LL-37, a system with high bacteria-killing effect, interact with the bacterial membrane and provide new insights into the eradication mechanism. Combining the advanced experimental techniques neutron reflectivity and quartz crystal microbalance with dissipation monitoring, a mechanistic drug delivery model for LL-37-loaded cubosomes on bacterial mimicking bilayers was constructed. Moreover, the cubosome interaction with Escherichia coli was directly visualized using super-resolution laser scanning microscopy and cryogenic electron tomography. We could conclude that cubosomes loaded with LL-37 adsorbed and distorted bacterial membranes, providing evidence that the peptide-loaded cubosomes function as an antimicrobial unit.
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| 10. |
- Heuser, Sebastian, et al.
(författare)
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Angular dependence of photoemission time delay in helium
- 2016
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Ingår i: Physical Review A. - 2469-9926. ; 94:6
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Tidskriftsartikel (refereegranskat)abstract
- Time delays of electrons emitted from an isotropic initial state with the absorption of a single photon and leaving behind an isotropic ion are angle independent. Using an interferometric method involving XUV attosecond pulse trains and an IR-probe field in combination with a detection scheme, which allows for full three-dimensional momentum resolution, we show that measured time delays between electrons liberated from the 1s(2) spherically symmetric ground state of helium depend on the emission direction of the electrons relative to the common linear polarization axis of the ionizing XUV light and the IR-probing field. Such time delay anisotropy, for which we measure values as large as 60 as, is caused by the interplay between final quantum states with different symmetry and arises naturally whenever the photoionization process involves the exchange of more than one photon. With the support of accurate theoretical models, the angular dependence of the time delay is attributed to small phase differences that are induced in the laser-driven continuum transitions to the final states. Since most measurement techniques tracing attosecond electron dynamics involve the exchange of at least two photons, this is a general and significant effect that must be taken into account in all measurements of time delays involving photoionization processes.
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| 11. |
- Håkansson, Joakim, et al.
(författare)
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Characterization of the in vitro, ex vivo, and in vivo Efficacy of the Antimicrobial Peptide DPK-060 Used for Topical Treatment
- 2019
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Ingår i: Frontiers in Cellular and Infection Microbiology. - : Frontiers Media SA. - 2235-2988. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Antimicrobial peptides, also known as host defense peptides, have recently emerged as a promising new category of therapeutic agents for the treatment of infectious diseases. This study evaluated the preclinical in vitro, ex vivo, and in vivo antimicrobial activity, as well as the potential to cause skin irritation, of human kininogen-derived antimicrobial peptide DPK-060 in different formulations designed for topical delivery. We found that DPK-060 formulated in acetate buffer or poloxamer gel caused a marked reduction of bacterial counts of Staphylococcus aureus in vitro (minimum microbicidal concentration <5 μg/ml). We also found that DPK-060 in poloxamer gel significantly suppressed microbial survival in an ex vivo wound infection model using pig skin and in an in vivo mouse model of surgical site infection (≥99 or ≥94% reduction in bacterial counts was achieved with 1% DPK-060 at 4 h post-treatment, respectively). Encapsulation of DPK-060 in different types of lipid nanocapsules or cubosomes did not improve the bactericidal potential of the peptide under the applied test conditions. No reduction in cell viability was observed in response to administration of DPK-060 in any of the formulations tested. In conclusion, the present study confirms that DPK-060 has the potential to be an effective and safe drug candidate for the topical treatment of microbial infections; however, adsorption of the peptide to nanocarriers failed to show any additional benefits.
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| 12. |
- Matougui, Nada, et al.
(författare)
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Lipid-based nanoformulations for peptide delivery
- 2016
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Ingår i: International Journal of Pharmaceutics. - : Elsevier. - 0378-5173 .- 1873-3476. ; 502:1-2, s. 80-97
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Forskningsöversikt (refereegranskat)abstract
- Nanoformulations have attracted a lot of attention because of their size-dependent properties. Among the array of nanoformulations, lipid nanoformulations (LNFs) have evoked increasing interest because of the advantages of their high degree of biocompatibility and versatility. The performance of lipid nanoformulations is greatly influenced by their composition and structure. Therapeutic peptides represent a growing share of the pharmaceutical market. However, the main challenge for their development into commercial products is their inherent physicochemical and biological instability. Important peptides such as insulin, calcitonin and cyclosporin A have been incorporated into LNFs. The association or encapsulation of peptides within lipid-based carriers has shown to protect the labile molecules against enzymatic degradation. This review describes strategies used for the formulation of peptides and some methods used for the assessment of association efficiency. The advantages and drawbacks of such carriers are also described.
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| 13. |
- Mira, Isabel C., et al.
(författare)
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Foam forming revisited Part I. Foaming behaviour of fibre-surfactant systems
- 2014
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Ingår i: Nordic Pulp & Paper Research Journal. - : SPCI. - 0283-2631 .- 2000-0669. ; 29:4, s. 679-688
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Tidskriftsartikel (refereegranskat)abstract
- The foaming properties of typical chemithermo-mechanical (CTMP) and kraft pulp paper making formulations in the presence of a series of surfactants were investigated using a lab-scale foaming set up. Foamability, foam stability, and bubble size distribution of the generated foam-fibre systems were measured. The foaming behaviour of the fibre/surfactant systems was found to be dependent on the surfactant concentration. Foams fulfilling the target requirements of air content (ca. 65% v/v air) and average bubble size (25 to 75 μm in radius) were obtained with all the seven surfactants tested. Three of the surfactants were found to allow for a rapid foaming in the system, namely sodium dodecyl sulphate (SDS), a commercial mixture of alkyl and ethoxylated alkyl sulphates, (MixSAES) and a commercial mixture of short chain alkyl glucosides (C8/C10Gluc). The rapid foaming is believed to be an intrinsic property of mixtures of surfactants with the right molecular structures and in the right proportion with respect to each other. On the other hand, the minimum surfactant concentrations required to reach the target foam volumes were lowest for surfactants with an anionic character. Further, the type of pulp fibre and the presence of GCC in the surfactant/pulp formulation were found to have very little effect on the foaming performance of the suspensions.
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| 14. |
- Simoni, Edi, et al.
(författare)
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Biocompatibility of glycerol monooleate nanoparticles as tested on inner ear cells
- 2019
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Ingår i: International Journal of Pharmaceutics. - : Elsevier B.V.. - 0378-5173 .- 1873-3476. ; 572
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Tidskriftsartikel (refereegranskat)abstract
- Sensorineural hearing loss due to aging, noise exposure, trauma or drug ototoxicity is irreversible because cochlear hair cells and neurons cannot regenerate. Recently, therapeutic strategies involving nanoparticles have been developed as innovative drug delivery systems. Thermodynamically stable liquid crystalline nanoparticles based on the polar lipid glycerol monooleate (GMO NP, cubosomes), nontoxic and able to encapsulate both hydrophilic and hydrophobic compounds, were produced and tested for biocompatibility in an immortalized Organ of Corti derived cell line (OC-k3), through cell viability and cytomorphological assays, and Western blot expression profiles of apoptotic markers. Overall, the GMO NP were biocompatible in OC-k3 at the doses and time tested, supporting previous data obtained in a neuronal cell line (PC12). The results encourage further tests on GMO NP-mediated drug release with improved target specificity and could be useful to develop innovative therapies against sensorineural hearing loss.
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| 15. |
- Valente, Filippo, et al.
(författare)
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Evaluation of toxicity of glycerol monooleate nanoparticles on PC12 cell line.
- 2018
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Ingår i: International Journal of Pharmaceutics. - : Elsevier BV. - 0378-5173 .- 1873-3476. ; 539:1-2, s. 23-30
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Tidskriftsartikel (refereegranskat)abstract
- An innovative approach to improve drug delivery is the use of glycerol monooleate nanoparticles. Numerous studies describe their high versatility, low toxicity and ability to carry relatively high loads of conjugated compounds including scarcely soluble ones, providing sustained drug release and increasing drug diffusion and half-life. Despite a growing interest in their potential use for therapeutic applications, there are surprisingly few literature data concerning the toxic effects of these nanoparticles at high concentrations in vitro and in vivo, and their effects on cell metabolism. We produced and characterized from a physical-chemical point of view glycerol monooleate nanoparticles and tested them on the PC12 cell line, a rat model of neuronal differentiation. The toxicity of these nanoparticles was evaluated by molecular methods on cell viability, cell cycle, nanoparticle uptake and induction of apoptosis. The results showed that glycerol monooleate nanoparticles up to 100 μg/mL had no toxic effects on PC12 cells, did not induce significant changes in the cell cycle nor cause apoptosis. The nanoparticles entered PC12 cells 8 h after treatment, successfully delivering the conjugate compound inside cells. Overall, glycerol monooleate nanoparticles did not exhibit significant toxicity on PC12 cell line in concentrations up to 100 µg/mL, supporting their therapeutic use as drug delivery systems.
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