Aesculus hippocastanum L. as a Stabilizer in Hemp Seed Oil Nanoemulsions for Potential Biomedical and Food Applications.

Nanoemulsion systems receive a significant amount of interest nowadays due to their promising potential in biomedicine and food technology. Using a two-step process, we produced a series of nanoemulsion systems with different concentrations of hemp seed oil (HSO) stabilized with Aesculus hippocastanum L. extract (AHE). Water and commercially-available low-concentrated hyaluronic acid (HA) were used as the liquid phase. Stability tests, including an emulsifying index (EI), and droplet size distribution tests performed by dynamic light scattering (DLS) proved the beneficial impact of AHE on the emulsion's stability. After 7 days of storage, the EI for the water-based system was found to be around 100%, unlike the HA systems. The highest stability was achieved by an emulsion containing 5% HSO and 2 g/L AHE in water, as well as the HA solution. In order to obtain the detailed characteristics of the emulsions, UV-Vis and FTIR spectra were recorded, and the viscosity of the samples was determined. Finally, a visible microscopic analysis was used for the homogeneity evaluation of the samples, and was compared with the DLS results of the water system emulsion, which showed a desirable stability. The presented results demonstrate the possible use of oil emulsions based on a plant extract rich in saponins, such as AHE. Furthermore, it was found that the anti-inflammatory properties of AHE provide opportunities for the development of new emulsion formulations with health benefits.

A Rapid Gas-Chromatography/Mass-Spectrometry Technique for Determining Odour Activity Values of Volatile Compounds in Plant Proteins: Soy, and Allergen-Free Pea and Brown Rice Protein.

Plant-based protein sources have a characteristic aroma that limits their usage in various meat-alternative formulations. Despite being the most popular plant-based protein, the allergenicity of soy protein severely restricts the potential adoption of soy protein as an animal substitute. Thereby, allergen-free plant-protein sources need to be characterized. Herein, we demonstrate a rapid solid-phase-microextraction gas-chromatography/mass-spectrometry (SPME-GC/MS) technique for comparing the volatile aroma profile concentration of two different allergen-free plant-protein sources (brown rice and pea) and comparing them with soy protein. The extraction procedure consisted of making a 1:7 w/v aqueous plant protein slurry, and then absorbing the volatile compounds on an SPME fibre under agitation for 10 min at 40 °C, which was subsequently injected onto a GC column coupled to an MS system. Observed volatile concentrations were used in conjunction with odour threshold values to generate a Total Volatile Aroma Score for each protein sample. A total of 76 volatile compounds were identified. Aldehydes and furans were determined to be the most dominant volatiles present in the plant proteins. Both brown rice protein and pea protein contained 64% aldehydes and 18% furans, with minor contents of alcohols, ketones and other compounds. On the other hand, soy protein consisted of fewer aldehydes (46%), but a more significant proportion of furans (42%). However, in terms of total concentration, brown rice protein contained the highest intensity and number of volatile compounds. Based on the calculated odour activity values of the detected compounds, our study concludes that pea proteins could be used as a suitable alternative to soy proteins in applications for allergen-free vegan protein products without interfering with the taste or flavour of the product.

Whey Proteins as a Potential Co-Surfactant with Aesculus hippocastanum L. as a Stabilizer in Nanoemulsions Derived from Hempseed Oil.

The use of natural surfactants including plant extracts, plant hydrocolloids and proteins in nanoemulsion systems has received commercial interest due to demonstrated safety of use and potential health benefits of plant products. In this study, a whey protein isolate (WPI) from a byproduct of cheese production was used to stabilize a nanoemulsion formulation that contained hempseed oil and the Aesculus hippocastanum L. extract (AHE). A Box-Behnken experimental design was used to set the formulation criteria and the optimal nanoemulsion conditions, used subsequently in follow-up experiments that measured specifically emulsion droplet size distribution, stability tests and visual quality. Regression analysis showed that the concentration of HSO and the interaction between HSO and the WPI were the most significant factors affecting the emulsion polydispersity index and droplet size (nm) (p < 0.05). Rheological tests, Fourier transform infrared spectroscopy (FTIR) analysis and L*a*b* color parameters were also taken to characterize the physicochemical properties of the emulsions. Emulsion systems with a higher concentration of the AHE had a potential metabolic activity up to 84% in a microbiological assay. It can be concluded from our results that the nanoemulsion system described herein is a safe and stable formulation with potential biological activity and health benefits that complement its use in the food industry.

Multidisciplinary Studies of Folk Medicine "Five Thieves' Oil" (Olejek Pieciu Zlodziei) Components.

To meet the growing interest in natural antibacterial agents, we evaluated the physicochemical and biological properties of the folk medicine known as "five thieves' oil" (Polish name: olejek pieciu zlodziei). Five thieves' oil consists of a mixture of five oils: rosemary, lemon, clove, eucalyptus, and cinnamon. In this study, we performed gas chromatography, FTIR, and UV-vis spectroscopic analysis, as well as L-a-b color tests, contact angle determination, and surface tension determination. To verify its antibacterial activity, the metabolic activity and changes in cell membrane permeability of bacteria of the genus Pseudomonas were studied. As a result, it was found that among the constituent oils, the oils of clove and cinnamon were the least volatile and, at the same time, had the strongest antibacterial activity. However, a mix of all the oils also showed comparable activity, which was even more pronounced for the oils after 4 weeks of aging. This effect can be linked to the high content of terpene derivatives such as eugenol and cinnamaldehyde, which can cause changes in bacterial membrane permeability, affecting cell activity and survival. This study is the first to characterize the constituents of the popular folk medicine five thieves' oil, confirming and explaining its strong antibacterial activity, thus constituting a significant contribution to contemporary health education.

Fluorescent Submicron-Sized Poly(heptafluoro-n-butyl methacrylate) Particles with Long-Term Stability.

Fluorescent submicron particles of fluorinated methacrylate (HFMBA) with long-term stability have been synthesized and characterized with regard to their potential applications. Rhodamine B (RBITC) isothiocyanate was used as the fluorescent component. The core-shell structure of the particles effectively protected the dye against bleaching. HFBMA nanoparticle (NP) stability was confirmed after seven years of storage. Only slight differences were found in the polydispersity index (pdi) from 0.002 to 0.010. Particle size measurements were carried out using dynamic light scattering (DLS), nanoparticle tracking (NTA), and fluorescence correlation spectroscopy (FCS). The hydrodynamic diameter evaluated by different methods were in good agreement, respectively: 184-550 nm, 218-579 nm, and 236-508 nm. Particle and core morphology was estimated by using scanning and transmission electron microscopy (SEM and TEM). The ability to recognize particles in 3D as a reference sample in biological media has been confirmed by epifluorescence optical microscopy, confocal laser scanning microscopy, and super-resolution confocal microscopy (STED).

Plant Extracts Containing Saponins Affects the Stability and Biological Activity of Hempseed Oil Emulsion System.

In this study, two saponins-rich plant extracts, viz. Saponaria officinalis and Quillaja saponaria, were used as surfactants in an oil-in-water (O/W) emulsion based on hempseed oil (HSO). This study focused on a low oil phase content of 2% v/v HSO to investigate stable emulsion systems under minimum oil phase conditions. Emulsion stability was characterized by the emulsification index (EI), centrifugation tests, droplet size distribution as well as microscopic imaging. The smallest droplets recorded by dynamic light scattering (droplets size v. number), one day after the preparation of the emulsion, were around 50-120 nm depending the on use of Saponaria and Quillaja as a surfactant and corresponding to critical micelle concentration (CMC) in the range 0-2 g/L. The surface and interfacial tension of the emulsion components were studied as well. The effect of emulsions on environmental bacteria strains was also investigated. It was observed that emulsions with Saponaria officinalis extract exhibited slight toxic activity (the cell metabolic activity reduced to 80%), in contrast to Quillaja emulsion, which induced Pseudomonas fluorescens ATCC 17400 growth. The highest-stability samples were those with doubled CMC concentration. The presented results demonstrate a possible use of oil emulsions based on plant extract rich in saponins for the food industry, biomedical and cosmetics applications, and nanoemulsion preparations.

Protein Fractions from Flaxseed: The Effect of Subsequent Extractions on Composition and Antioxidant Capacity.

Flaxseed proteins exhibit functionalities interesting for the food industry, including antioxidant capacity. Antioxidant activity depends on the protein composition and the presence of phenolic compounds extracted with them from the matrix. The research focused on the effect of subsequent protein extractions (water, salt and alkaline) of flaxseed meals (of three cultivars) on the protein fraction composition and its relations to antioxidant capacity. The protein and phenolic profiles and antioxidant functionalities (in antiradical ORAC and emulsion assays) were analysed. Spectroscopic characteristics of the fractions (fluorometric and FT-IR analysis) were also included. Our study has shown the effect of fractionation on the share of proteins at MW from 56-38 kDa (globulin-like) and <15 kDa (albumin-like) in the protein profiles. The highest globulin share was in the alkaline-extracted fractions (AEF) and albumin in the salt-extracted (SEF) ones. SDG (secoisolariciresinol diglucosides) and phenolic acids (p-coumaric and ferulic) were extracted with flaxseed proteins. Their contents were fraction-dependent and the highest in AEF. The concentration of phenolics in AEF corresponded with the highest antiradical capacity (ORAC) compared with the other fractions. However, the SEF showed a higher ability to inhibit oxidation in emulsions than AEF, which could be associated with the higher content of the low MW proteins.

Extracts from Frangula alnus Mill. and Their Effects on Environmental and Probiotic Bacteria.

The bark of Frangula alnus Mill (FAM), the so-called alder buckthorn, has been widely investigated for its medicinal properties, especially its laxative effects and the bioactive properties of the plant material extract. Still, there is no wider study devoted to its antibacterial properties. This is important in the context of its impact on probiotic gut bacteria. The aim of the research was to recognize the effect of FAM extract on bacterial cells, and to determine how the bioactive properties and composition of the extract are influenced by the type of solvent used for the extraction. To find the most suitable conditions for the FAM extraction, we used four solvent solutions with different polarities, including water, methanol, ethanol, and isopropanol. We assessed the quality and composition of the extracts with spectral analysis, using spectrophotometric (FTIR, UV-Vis) and chromatographic methods (GC-MS). Finally, we analyzed the extractant impact of the extracts on the selected bacterial cells. The results showed that the chemical diversity of the extracts increased with the increase in solvent polarity, in which the abundance of frangulin, the main bioactive compound in buckthorn bark, was confirmed. Pseudomonas fluorescens ATCC 17400 was particularly sensitive to the action of extracts, whereas other strains of the Pseudomonas genus showed practically no adverse effects. Ethanolic extracts had the strongest effect on most of the selected bacteria strains. We found that the probiotic Lactobacillus strain, which represents intestinal microflora, has no direct effect on probiotic microorganisms. The research shown FAM extracts can be safe for probiotic bacteria present in human gut microflora. Moreover, the study indicated that contact with the extracts may reduce the total permeability of the bacterial membranes. This opens up the possibility of using FAM extracts as a factor regulating transport into cells, which may be used to support the action of other bioactive substances.

Studies of Peculiar Mg-Containing and Oscillating Bioapatites in Sheep and Horse Teeth.

New types of biological apatites have been discovered in molar sheep and horse teeth and are divided in two types. In the first and more general type, the release of Mg ions is parallel to the changes in composition of apatite leading to a final stoichiometric ratio of Ca to P ions, going from dentin depth towards the boundary of enamel with air. Inside dentin, another apatite sub-types were discovered with alternating layers of Mg-rich and C-rich apatites. The approximate formal stoichiometric relationships for these peculiar types of bioapatites are suggested. We identified two kinds of ion-exchanges responsible for formation of peculiar apatites. Various combinations of main and minor elements lead to new versions of biological apatites.

Disruptive effect of tocopherol oxalate on DPPC liposome structure: DSC, SAXS, and fluorescence anisotropy studies.

α-Tocopherol oxalate (TO), a tocopherol ester derivative, was investigated for its effect on the structural changes of fully hydrated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) liposomes, as a function of concentration and temperature, by applying differential scanning calorimetry (DSC), small angle X-ray scattering (SAXS), and DPH fluorescence anisotropy methods. The DSC and DPH anisotropy data indicated that TO embedded into DPPC membrane lowered the enthalpy (ΔHm) and temperature (Tm) of the main phase transition as well its cooperativity. Fluidization of the membrane at a lowered temperature was accompanied by formation of mixed structures of tocopherol-enriched domains. SAXS studies showed the formation of various ordered structures in DPPC gel-phase during incorporation of TO into the bilayer, as evidenced by the existence of lamellar phases with repeat distances (d) of 6.13 and 6.87 nm, assigned to TO-enriched domains and a lamellar, liquid-ordered DPPC phase with d = 8.45 nm at increasing TO concentrations with lowering and broadening of the Bragg peaks, and diffuse scattering, characteristic of a fluid Lα phase, were observed. In DPPC fluid-phase, the increasing presence of TO at low concentrations resulted in the appearance of a liquid-ordered phase with repeat d = 6.9 nm coexistent with a lamellar structure with d = 9.2 nm, assigned to liquid-disordered structures. An increasing repeat distance observed with raising the TO amount in the DPPC bilayer evolved from an increasing interlamellar water layer of increasing thickness. Presence of TO facilitated penetration of water molecules into the acyl chain region which decreased van der Waals interactions in the bilayer. The DSC, SAXS, and fluorescence anisotropy data established that TO exhibited pronounced disruptive activity in DPPC membranes compared to α-tocopherol. The driving force of the observed action was attributed to electrostatic and dipole interactions of the acidic moiety with the polar head group of phospholipids in the interface region of the bilayer.

Interactions between colloidal silver and photosynthetic pigments located in cyanobacteria fragments and in solution.

Changes in the yield of the fluorescence emitted by pigments of photosynthetic organisms could be used for the establishment of the presence of some toxic substances. The presence of colloidal metals can be indicated by enhancement of pigments' emission as a result of plasmons generation. The spectra of the pigments of cyanobacterium Synechocystis located in the bacterium fragments and in solutions with and without colloidal silver additions have been measured. The quantum yield of the pigments' fluorescence in solution has been observed to increase at some wavelength of excitation, while the fluorescence of the pigments in the bacteria fragments has been only quenched as a consequence of interactions with colloidal silver particles. Close contact between pigment molecules located in bacteria fragments and silver particles is probably not possible. We plan in future to investigate the influence of other, more typical metal pollutants of water, using similar spectral methods and several other photosynthetic bacteria pigments, in solution, in cell fragments and in the whole bacteria organisms.

Spectral properties of fluorescein molecules in water with the addition of a colloidal suspension of silver.

The absorption, fluorescence excitation, and fluorescence emission spectra of water solutions of fluorescein dye with the addition of various amounts of a colloidal silver suspension have been measured in order to check if in such systems it is possible to distinguish the change in photonic mode density due to the metal presence from the other effects such as the influences of the microemulsion system on the spectral properties of the dye. It has been found that the presence of the silver colloid changes the concentrations of the various ionic forms of fluorescein, characterized by different yields of fluorescence. This effect is partially responsible for the change in the yield of the fluorescence emission observed at certain concentrations of the dye and the colloids. But even at the same concentration of various ionic forms of fluorescein (at the same pH of the dye solution and the dye-colloid mixture), at certain concentrations of fluorescein and the colloid, the yield of the dye fluorescence increases, which must be due to the interaction between the dye and the silver colloid. Because of the superposition of several processes influencing the dye yield of fluorescence, it is necessary to carefully establish the properties of the dye in a given environment, before considering its practical application as a marker of the metal presence. It is not excluded that similar complex effects could also occur in biological samples containing natural pigments and colloids of metals. Investigations of other dyes with other forms of metallic samples are in progress.