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.

Production of high loading insulin nanoparticles suitable for oral delivery by spray drying and freeze drying techniques.

Insulin nanoparticles (NPs) with high loading content have found diverse applications in different dosage forms. This work aimed to evaluate the impact of freeze-drying and spray drying process on the structures of insulin-loaded chitosan nanoparticles, with or without mannitol as cryoprotectants. We also assessed the quality of these nanoparticles by redissolving them. Before dehydration, the chitosan/sodium tripolyphosphate/insulin crosslinked nanoparticles were optimized to 318 nm of particle size, 0.18 of PDI, 99.4% of entrapment efficiency, and 25.01% of loading content. After reconstitution, all nanoparticles, except the one produced by the freeze-drying method without using mannitol, maintained their spherical particle structure. The nanoparticles dehydrated by spray drying without mannitol also showed the smallest mean particle size (376 nm) and highest loading content (25.02%) with similar entrapment efficiency (98.7%) and PDI (0.20) compared to mannitol-containing nanoparticles dehydrated by either spray drying or freeze-drying techniques. The nanoparticles dried by spray drying without mannitol also resulted in the fastest release and highest cellular uptake efficacy of insulin. This work shows that spray drying can dehydrate insulin nanoparticles without the need for cryoprotectants, creating a significant advantage in terms of greater loading capacity with lower additive requirements and operating costs as compared to conventional freeze drying approaches.

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.

Optimal ultrasonication process time remains constant for a specific nanoemulsion size reduction system.

This paper theorizes the existence of a constant optimum ultrasound process time for any size-reduction operation, independent of process parameters, and dependent on product parameters. We test the concept using the case of 'ultrasonic preparation of oil-in-water nanoemulsions' as model system. The system parameters during ultrasonication of a hempseed oil nanoemulsion was evaluated by a response surface methodology, comprising lecithin and poloxamer-188 as surfactants. Results revealed that the particle size and emulsion stability was affected significantly (p < 0.05) by all product parameters (content of hempseed oil-oil phase, lecithin and polaxamer-surfactants); but was not significantly (p > 0.05) affected by process parameter ('ultrasonication process time'). Next, other process parameters (emulsion volume and ultrasonic amplitude) were tested using kinetic experiments. Magnitude of particle size reduction decreased with increasing 'ultrasonication process time' according to a first order relationship, until a minimum particle size was reached; beyond which ultrasonication no longer resulted in detectable decrease in particle size. It was found that the optimal ultrasonication process time (defined as time taken to achieve 99% of the 'maximum possible size reduction') was 10 min, and was roughly constant regardless of the process parameters (sample volume and ultrasonic amplitude). Finally, the existence of this constant optimal ultrasonication process time was proven for another emulsion system (olive oil and tween 80). Based on the results of these case studies, it could be theorized that a constant optimum ultrasonication process time exists for the ultrasonication-based size-reduction processes, dependent only on product parameters.

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.

Comparing microfluidics and ultrasonication as formulation methods for developing hempseed oil nanoemulsions for oral delivery applications.

Emerging formulation technologies aimed to produce nanoemulsions with improved characteristics, such as stability are attractive endeavors; however, comparisons between competing technologies are lacking. In this study, two formulation techniques that employed ultrasound and microfluidic approaches, respectively, were examined for relative capacity to produce serviceable oil in water nanoemulsions, based on hempseed oil (HSO). The ultrasound method reached > 99.5% entrapment efficiency with nanoemulsions that had an average droplet size (Z-Ave) < 180 nm and polydispersity index (PDI) of 0.15 ± 0.04. Surfactant concentration (% w/v) was found to be a significant factor (p < 0.05) controlling the Z-Ave, PDI and zeta potential of these nanoparticles. On the other hand, the microfluidic approach produced smaller particles compared to ultrasonication, with good stability observed during storage at room temperature. The Z-Ave of < 62.0 nm was achieved for microfluidic nanoemulsions by adjusting the aqueous : organic flow rate ratio and total flow rate at 4:1 and 12 mL/min, respectively. Further analyses including a morphology examination, a simulated gastrointestinal release behavior study, transepithelial transport evaluations and a toxicity test, using a Caco2-cell model, were performed to assess the functionality of the prepared formulations. The results of this study conclude that both approaches of ultrasound and microfluidics have the capability to prepare an HSO-nanoemulsion formulation, with acceptable characteristics and stability for oral delivery applications.

Antioxidants help favorably regulate the kinetics of lipid peroxidation, polyunsaturated fatty acids degradation and acidic cannabinoids decarboxylation in hempseed oil.

The seed of the hemp plant (Cannabis sativa L.) has been revered as a nutritional resource in Old World Cultures. This has been confirmed by contemporary science wherein hempseed oil (HSO) was found to exhibit a desirable ratio of omega-6 and omega-3 polyunsaturated fatty acids (PUFAs) considered optimal for human nutrition. HSO also contains gamma-linoleic acid (GLA) and non-psychoactive cannabinoids, which further contribute to its' potential bioactive properties. Herein, we present the kinetics of the thermal stability of these nutraceutical compounds in HSO, in the presence of various antioxidants (e.g. butylated hydroxytoluene, alpha-tocopherol, and ascorbyl palmitate). We focussed on oxidative changes in fatty acid profile and acidic cannabinoid stability when HSO was heated at different temperatures (25 °C to 85 °C) for upto 24 h. The fatty acid composition was evaluated using both GC/MS and 1H-NMR, and the cannabinoids profile of HSO was obtained using both HPLC-UV and HPLC/MS methods. The predicted half-life (DT50) for omega-6 and omega-3 PUFAs in HSO at 25 °C was about 3 and 5 days, respectively; while that at 85 °C was about 7 and 5 hours respectively, with respective activation energies (Ea) being 54.78 ± 2.36 and 45.02 ± 2.87 kJ/mol. Analysis of the conjugated diene hydroperoxides (CDH) and p-Anisidine value (p-AV) revealed that the addition of antioxidants significantly (p < 0.05) limited lipid peroxidation of HSO in samples incubated at 25-85 °C for 24 h. Antioxidants reduced the degradation constant (k) of PUFAs in HSO by upto 79%. This corresponded to a significant (p < 0.05) increase in color stability and pigment retention (chlorophyll a, chlorophyll b and carotenoids) of heated HSO. Regarding the decarboxylation kinetics of cannabidiolic acid (CBDA) in HSO, at both 70 °C and 85 °C, CBDA decarboxylation led to predominantly cannabidiol (CBD) production. The half-life of CBDA decarboxylation (originally 4 days) could be increased to about 17 days using tocopherol as an antioxidant. We propose that determining acidic cannabinoids decarboxylation kinetics is a useful marker to measure the shelf-life of HSO. The results from the study will be useful for researchers looking into the thermal treatment of hempseed oil as a functional food product, and those interested in the decarboxylation kinetics of the acidic cannabinoids.

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.

Pea Protein for Hempseed Oil Nanoemulsion Stabilization.

In this paper, we present the possibility of using pea protein isolates as a stabilizer for hempseed oil (HSO)-based water/oil emulsions in conjunction with lecithin as a co-surfactant. A Box-Behnken design was employed to build polynomial models for optimization of the ultrasonication process to prepare the emulsions. The stability of the system was verified by droplet size measurements using dynamic light scattering (DLS) as well as centrifugation and thermal challenge tests. The z-ave droplet diameters of optimized emulsion were 209 and 207 nm after preparation and 1 week storage, respectively. The concentration of free Linoleic acid (C18:2; n-6) was used for calculation of entrapment efficiency in prepared nanoemulsions. At optimum conditions of the process, up to 98.63% ± 1.95 of entrapment was achieved. FTIR analysis and rheological tests were also performed to evaluate the quality of oil and emulsion, and to verify the close-to-water like behavior of the prepared samples compared to the viscous nature of the original oil. Obtained results confirmed the high impact of lecithin and pea protein concentrations on the emulsion droplet size and homogeneity confirmed by microscopic imaging. The presented results are the first steps towards using hempseed oil-based emulsions as a potential food additive carrier, such as flavor. Furthermore, the good stability of the prepared nanoemulsion gives opportunities for potential use in biomedical and cosmetic applications.

Plant Extracts Inhibit the Formation of Hydroperoxides and Help Maintain Vitamin E Levels and Omega-3 Fatty Acids During High Temperature Processing and Storage of Hempseed and Soybean Oils.

Oxidative stability of hempseed and soybean oils, was evaluated after heating at 180 °C, followed by a subsequent 14-day storage at 38 °C. Natural plant extracts (Rosemary, Sage, and Thyme) were added to oils, to evaluate the carry-over stabilizing potential. Heated oils exhibited a relatively faster (P < 0.05) onset of lipid oxidation, as depicted by the analysis of the peroxides and aldehydes formed during the lipid oxidation process, with hempseed oil being more susceptible to lipid oxidation than soybean oil. There were notable losses in ω-3 PUFA and ω-6-GLA during storage of heat treated hempseed oil (P < 0.05). Moreover, peroxide values measured from hempseed oil remained low after high-temperature heating but progressed at a relatively greater rate than that observed in soybean during storage (P < 0.05). The addition of different plant extracts to oils did not prevent oxidation due to heating, but effectively inhibited the generation of hydroperoxides during subsequent storage (P < 0.05). This stabilizing effect was attributed to retention of tocopherols, in particular, γ-tocopherol. PRACTICAL APPLICATION: This research demonstrates the use of plant extracts like rosemary, sage and thyme, for improving the shelf-life and nutritional stability of hempseed and soybean oil. We demonstrate the deterioration of fatty acid profiles and vitamin E levels in the oil on heating and subsequent storage, and show the efficacy of using plant extracts in slowing down these deteriorations. This research will be applicable in food industries using or producing oils for use in food during cooking, and also as dressing on already processed food products.

Effect of solvent type and ratio on betacyanins and antioxidant activity of extracts from Hylocereus polyrhizus flesh and peel by supercritical fluid extraction and solvent extraction.

The main objective of the present study was to investigate the effect of solvent type and ratio as well as the extraction techniques (i.e. supercritical fluid extraction (SFE) and conventional solvent extraction) on betacyanins and antioxidant activity of the peel and fresh extract from the red pitaya (Hylocereus polyrhizus). The peel and flesh extracts obtained by SFE at 25MPa pressure and 10% EtOH/water (v/v) mixture as a co-solvent contained 24.58 and 91.27mg/100ml total betacyanin, respectively; while the most desirable solvent extraction process resulted in a relatively higher total betacyanin in the peel and flesh extracts (28.44 and 120.28mg/100ml, respectively). The major betacyanins identified in the pitaya peel and flesh extracts were betanin, isobetanin, phyllocactin, butyrylbetanin, isophyllocactin and iso-butyrylbetanin. The flesh extract had the stronger antioxidant activity than the peel extract when the higher proportion of ethanol to water (E/W) was applied for the extraction.