Utilization of Nanotechnology to Improve the Application and Bioavailability of Phytochemicals Derived from Waste Streams.

Phytochemicals are relatively small molecular species found in edible plants that may exhibit a diverse range of techno- and biofunctional attributes. In particular, there has been great interest in the identification, isolation, and utilization of dietary phytochemicals that can be used as natural pigments, antioxidants, or antimicrobials or that may improve human health and wellbeing by preventing chronic diseases, such as cardiovascular diseases, diabetes, obesity, and cancer. Relatively high levels of these phytochemicals are often present in the waste streams produced by the food and agriculture industry, such as the peels, stems, roots, or leaves of plants, that are normally discarded or turned into animal foods. From an economic and environmental perspective, it would be advantageous to convert these waste streams into value-added functional ingredients, which is consistent with the creation of a more circular economy. Bioactive phytochemicals can be isolated from agricultural and food waste streams using green extraction methods and then incorporated into plant-based functional foods or biodegradable active packaging materials. The utilization of phytochemicals in the food industry is often challenging. They may chemically degrade in the presence of light, heat, oxygen, and some pH conditions, thereby altering their biological activity. They may have low solubility in aqueous solutions and gastrointestinal fluids, thereby making them difficult to introduce into foods and leading to a low bioavailability. These challenges can sometimes be overcome using nanoencapsulation, which involves trapping the phytochemicals inside tiny food-grade particles. These nanoparticles may be assembled from edible lipids, proteins, carbohydrates, and/or surfactants and include nanoemulsions, solid lipid nanoparticles, nanoliposomes, and biopolymer nanoparticles. In this manuscript, we review a number of important phytochemicals and nanoencapsulation methods used to improve their efficacy.

Utilization of Nanotechnology to Improve the Handling, Storage and Biocompatibility of Bioactive Lipids in Food Applications.

Bioactive lipids, such as fat-soluble vitamins, omega-3 fatty acids, conjugated linoleic acids, carotenoids and phytosterols play an important role in boosting human health and wellbeing. These lipophilic substances cannot be synthesized within the human body, and so people must include them in their diet. There is increasing interest in incorporating these bioactive lipids into functional foods designed to produce certain health benefits, such as anti-inflammatory, antioxidant, anticancer and cholesterol-lowering properties. However, many of these lipids have poor compatibility with food matrices and low bioavailability because of their extremely low water solubility. Moreover, they may also chemically degrade during food storage or inside the human gut because they are exposed to certain stressors, such as high temperatures, oxygen, light, moisture, pH, and digestive/metabolic enzymes, which again reduces their bioavailability. Nanotechnology is a promising technology that can be used to overcome many of these limitations. The aim of this review is to highlight different kinds of nanoscale delivery systems that have been designed to encapsulate and protect bioactive lipids, thereby facilitating their handling, stability, food matrix compatibility, and bioavailability. These systems include nanoemulsions, solid lipid nanoparticles (SLNs), nanostructured lipid carriers (NLCs), nanoliposomes, nanogels, and nano-particle stabilized Pickering emulsions.

Formation and stabilization of nanoemulsion-based vitamin E delivery systems using natural biopolymers: Whey protein isolate and gum arabic.

Natural biopolymers, whey protein isolate (WPI) and gum arabic (GA), were used to fabricate emulsion-based delivery systems for vitamin E-acetate. Stable delivery systems could be formed when vitamin E-acetate was mixed with sufficient orange oil prior to high pressure homogenization. WPI (d32=0.11 µm, 1% emulsifier) was better than GA (d32=0.38 µm, 10% emulsifier) at producing small droplets at low emulsifier concentrations. However, WPI-stabilized nanoemulsions were unstable to flocculation near the protein isoelectric point (pH 5.0), at high ionic strength (>100mM), and at elevated temperatures (>60 °C), whereas GA-stabilized emulsions were stable. This difference was attributed to differences in emulsifier stabilization mechanisms: WPI by electrostatic repulsion; GA by steric repulsion. These results provide useful information about the emulsifying and stabilizing capacities of natural biopolymers for forming food-grade vitamin-enriched delivery systems.

Nanoemulsion delivery systems for oil-soluble vitamins: Influence of carrier oil type on lipid digestion and vitamin D3 bioaccessibility.

The influence of carrier oil type on the bioaccessibility of vitamin D3 encapsulated within oil-in-water nanoemulsions prepared using a natural surfactant (quillaja saponin) was studied using a simulated gastrointestinal tract (GIT) model: mouth; stomach; small intestine. The rate of free fatty acid release during lipid digestion decreased in the following order: medium chain triglycerides (MCT) > corn oil ≈ fish oil > orange oil > mineral oil. Conversely, the measured bioaccessibility of vitamin D3 decreased in the following order: corn oil ≈ fish oil > orange oil > mineral oil > MCT. These results show that carrier oil type has a considerable impact on lipid digestion and vitamin bioaccessibility, which was attributed to differences in the release of bioactives from lipid droplets, and their solubilization in mixed micelles. Nanoemulsions prepared using long chain triglycerides (corn or fish oil) were most effective at increasing vitamin bioaccessibility.

Antibody affinity maturation in vitro using unconjugated peptide antigen.

Selection of antibody library in vitro is almost always performed on a certain solid-phase with immobilized antigen. However, for the selection of small molecule binders, conjugation of the antigen to a carrier molecule is indispensable, which often leads to the selection of unwanted binders such as conjugate-binders or those with insufficient specificity. Here we describe a rapid and efficient way to improve the affinity of an anti-small molecule antibody without antigen derivatization. The method is based on the open-sandwich (OS) principle, which utilizes the antigen-dependent stabilization of antibody variable domain Fv. We used an anti-osteocalcin C-terminal peptide Fv that showed a good response but with moderate sensitivity in OS ELISA as a model. By selecting PCR-randomized V(H)-displaying phages for superior binders to the immobilized V(L) fragment in the presence of limited amount of antigen peptide, V(H) mutants that show superior detection sensitivity in OS ELISA were obtained, and were characterized to retain improved antigen-binding affinity. Furthermore, saturation mutagenesis of a mutant resulted in further improvement in sensitivity. This 'OS-selection' will be the first to select anti-small molecule antibodies without using conjugated antigens, and especially useful in the affinity maturation of antibodies whose Fv has limited stability in the absence of antigen.

Cloning, expression and characterization of endo-beta-1,4-mannanase from Aspergillus fumigatus in Aspergillus sojae and Pichia pastoris.

To be utilized in biomass conversion, including ethanol production and galactosylated oligosaccharide synthesis, namely prebiotics, the gene of extracellular endo-beta-1,4-mannanase (EC 3.2.1.78) of Aspergillus fumigatus IMI 385708 (formerly known as Thermomyces lanuginosus IMI 158749) was expressed first in Aspergillus sojae and then in Pichia pastoris under the control of the glyceraldehyde triphosphate dehydrogenase (gpdA) and the alcohol oxidase (AOX1) promoters, respectively. The highest production of mannanase (352 U mL(-1)) in A. sojae was observed after 6 days of cultivation. In P. pastoris, the highest mannanase production was observed 10 h after induction with methanol (61 U mL(-1)). The fold increase in mannanase production was estimated as approximately 12-fold and approximately 2-fold in A. sojae and P. pastoris, respectively, when compared with A. fumigatus. Both recombinant enzymes showed molecular mass of about 60 kDa and similar specific activities ( approximately 350 U mg(-1) protein). Temperature optima were at 60 degrees C and 45 degrees C, and maximum activity was at pH 4.5 and 5.2 for A. sojae and P. pastoris, respectively. The enzyme from P. pastoris was more stable retaining most of the activity up to 50 degrees C, whereas the enzyme from A. sojae rapidly lost activity above 40 degrees C.