Biotin as a structural component in the detection of small model antigens in BLA-S-ELISA.

An interesting biotin-linked-antigen-Sandwich-ELISA was developed (BLA-S-ELISA), which based on the captured Trinitrophenol-Biotin (TNP-Biotin) molecule between the immobilised monovalent antibody and enzyme-conjugated streptavidin. Monoclonal anti-Trinitrotoluene single chain fragment antibody (anti-TNT-scFv) was cloned and expressed in E. coli cells, and then used as an immobilised component in an assay. Thereafter, the previously synthesised TNP-Biotin was added as antigen followed by the addition of streptavidin-horseradish peroxidase (streptavidin-HRP) conjugate which led finally to the formation of a three-component system (antibody/TNP-Biotin/streptavidin-HRP). The assay was performed with a range of different dilutions of TNP-Biotin to establish its minimal detectable concentration. The detection limit of TNP-Biotin was 4 ngmL-1 (i.e. 200 pg or 0.42 pmol antigen calculated on the basis of 50 µL sample or 8.4 nM expressed in concentration units). According to our best knowledge, this is the very first time for any model antigen to be detected with such a form of biotin-streptavidin sandwich-assay.

Thermal Characterisation and Isoconversional Kinetic Analysis of Osmotically Dried Pork Meat Proteins Longissimus dorsi.

The kinetic properties and thermal characteristics of fresh pork meat proteins (Longissimus dorsi), as well as osmotically dehydrated meat proteins, were investigated using differential scanning calorimetry. Two isoconversional kinetical methods, namely the differential Friedman and integral Ortega methods, were employed to analyze the data. The obtained kinetic triplet, activation energy, pre-exponential factor, and extent of conversion, has been discussed. The resulting activation energy for proteins of fresh meat ranges between 751 kJ·mol-1 for myosin, 152 kJ·mol-1 for collagen and sarcoplasmic proteins, and 331 kJ·mol-1 for actin at a conversion degree of 0.1 to 0.9. For osmotically dried pork meat proteins, the values range from 307 kJ·mol-1 for myosin 272 kJ·mol-1 for collagen and sarcoplasmic proteins, and 334.83 kJ·mol-1 for actin at a conversion degree from 0.1 to 0.9. The proteins of the dry meat obtained by osmotic dehydration in molasses could be described as partly unfolded as they retain the characteristic protein denaturation transition. Concerning the decrease in enthalpies of proteins denaturation, thermodynamic destabilization of dried meat proteins occurred. On the contrary, dried meat proteins were thermally stabilized with respect to increase in the temperatures of denaturation. Knowledge of the nature of meat protein denaturation of each kind of meat product is one of the necessary tools for developing the technology of meat product processing and to achieve desired quality and nutritional value. The kinetic analysis of meat protein denaturation is appropriate because protein denaturation gives rise to changes in meat texture during processing and directly affects the quality of product.

Recovery of Polyphenolic Compounds and Vitamins from the Stinging Nettle Leaves: Thermal and Behavior and Biological Activity of Obtained Extracts.

Stinging nettle (SN) is an extraordinary plant from the Urticaceae botanical family. It is well-known and widely used in food and folk medicine to treat different disorders and diseases. This article aimed to study the chemical composition of SN leaves extracts, i.e., polyphenolic compounds and vitamins B and C, because many studies ascribed high biological potency to these compounds and their significance in the human diet. Besides the chemical profile, the thermal properties of the extracts were studied. The results confirmed presence of many polyphenolic compounds and vitamins B and C. It also showed that the chemical profile closely correlated with the applied extraction technique. The thermal analysis showed that analyzed samples were thermally stable up to about 160 °C. Thermal degradation of samples UAE, MAE, and MAC took place in four steps, and sample SE in three steps. Altogether, results confirmed the presence of health-beneficial compounds in stinging nettle leaves and indicated the possible application of its extract in pharmaceutical and food industries as both a medicinal and food additive.

The effect of various extraction techniques on the quality of sage (Salvia officinalis L.) essential oil, expressed by chemical composition, thermal properties and biological activity.

In this study, influence of the extraction techniques on the quality of the sage essential oil was investigated. Obtained samples were analyzed for chemical composition by GC/MS, thermal properties by thermogravimetric analysis (TGA), and for biological activity: antioxidant (DPPH, CUPRAC, FRAP, ABTS, HRSA and TBARS), microbiological (Staphylococcus aureus, Escherichia coli, Bacillus subtilis, Pseudomonas aeruginosa, Candida albicans, and Aspergillus niger), and cytotoxic (HeLa, LS-174, A549 and MRC-5) activities. Chemical composition showed that viridiflorol was principal compound in all samples followed by camphor, thujones, and verticiol. MWD 400 W was the most potent antioxidant agent, D 200 W and MWD 400 W antimicrobial agents, while hydrodistallates (D 200 W and D 400 W) were the most potent cytotoxic agents. An artificial neural network model was developed for the antioxidant activity anticipation of analyzed samples. These models showed good prediction properties (the r2 value during training cycle for output variables was 0.998).

Design and characterization of whey protein nanocarriers for thyme essential oil encapsulation obtained by freeze-drying.

Innovative coating powders, based on whey protein concentrate (10-15 wt%) as native (WPC) or denatured protein (d-WPC), solely or in combination with alginate (0.75 wt%, AL), containing thyme essential oil, were produced using the freeze-drying technique. The impact of individual components (protein, alginate and oil) as well as the effect of heat-induced protein denaturation, was resolved regarding physicochemical, thermal and morphological properties of powders. High product yield (∼100%), particle size (223-257 nm), low moisture content (0.10-0.13%) and zeta potential (-19 to -25.6 mV) were determined for all samples. Strong antimicrobial activity of thyme oil nanocarriers against foodborne pathogens was demonstrated. Thermogravimetric analysis (TGA) indicated enhanced thermal stability of encapsulated oil. The most specific bands of structural compounds were identified in Raman spectra of the tested formulations, but principal component analysis (PCA) on recorded spectra was necessary to show the differences between carriers of different wall materials.

A novel and effective natural product-based immunodetection tool for TNT-like compounds.

This study aimed to develop a fast and reliable protocol for Trinitrophenol-Tris(hydroxymethyl)aminomethane (TNP-Tris) detection applying a ß-lactamase-fusion protein of choice, the natural product-based immunoreagent tool of competitive sensitivity developed herein for the first time. Since the fusion protein 11B3-scFv-ß-lactamase is constructed from a scFv-antibody (11B3) linked to an enzyme (ß-lactamase), the step related to the use of secondary antibody in the enzyme-linked immunosorbent assay (ELISA) is completely omitted. Indeed, this fusion protein itself serves both as binding mean of the antigen model and detecting agent, due to the presence of the naturally occurring enzyme. In such a way, it actually affords the one-step TNP-Tris detection reaching a promising LOD value of 45 ± 2 fmol or 157 ± 6 pg/mL. Taken all together, the current protocol does represent much cheaper and significantly less-time consuming alternative compared both to the recombinant antibodies and recombinant phages, previously designed means in our labs for the same purpose.

Rosemary Essential Oils as a Promising Source of Bioactive Compounds: Chemical Composition, Thermal Properties, Biological Activity, and Gastronomical Perspectives.

Rosemary (Rosmarinus officinalis L.) is a plant worldwide cultivated mainly for essential oils, extracts, and as a spice. Up-to-date results showed diversity in composition of the essential oils, which may influence their quality, biological activity, and thermal properties. Therefore, the aim of this study was to investigate the chemical composition, antimicrobial activity, and thermal properties of the rosemary essential oils originating from Serbia and Russia. Additionally, oils were added to the sunflower oils in order to investigate possible antioxidant activity during the frying. Investigation of the chemical profile marked α-pinene, eucalyptol, and camphor as the most abundant compounds in both oils. However, overall composition influenced in such manner that Russian oil showed significantly higher antimicrobial activity, while Serbian oil proved to be better antioxidant agent in case of frying of sunflower oil. This would significantly influence possible application of the oils, which could be used as an antioxidant agent for extension of the food shelf life, or antimicrobial agent for protection against different microbial strains.

Functionality and Palatability of Yogurt Produced Using Beetroot Pomace Flour Granulated with Lactic Acid Bacteria.

Following the idea of sustainability in food production, a yogurt premix based on beetroot (Beta vulgaris) pomace flour (BPF) was developed. BPF was granulated with lactose solution containing lactic acid bacteria (LAB) by a fluidized bed. Particle size increased ~30%. A decrease in Carr Index from 21.5 to 14.98 and Hausner ratio from 1.27 to 1.18 confirmed improved flowability of granulated BPF, whereas a decrease in water activity implied better storability. Yogurts were produced weekly from neat starters and granulated BPF (3% w/w) that were stored for up to one month (4 °C). High viability of Streptococcus thermophilus was observed. Less pronounced syneresis, higher inhibition of colon cancer cell viability (13.0-24.5%), and anti-Escherichia activity were ascribed to BPF yogurts or their supernatants (i.e., extracted whey). Acceptable palatability for humans and dogs was demonstrated. A survey revealed positive consumers' attitudes toward the granulated BPF as a premix for yogurts amended to humans and dogs. For the first time, BPF granulated with LAB was used as a premix for a fermented beverage. An initial step in the conceptualization of a novel DIY (do it yourself) formula for obtaining a fresh yogurt fortified with natural dietary fiber and antioxidants has been accomplished.

Low sugar jellies of berry fruits: the impact of low vs. high temperature regime on their chemical composition and antioxidativity.

This study focuses on the impact of low and high temperature regimes on the chemical composition and antioxidativity of low sugar berry fruits jellies. High quality fruits (strawberry, raspberry and blackberry) were collected from Western Serbia region, quite well recognised both nationally and internationally due to an extremely well developed practice in growing berry fruits. The obtained results have clearly indicated the importance of low temperature regime for enriched contents of both total phenolics and anthocyanins followed by an enhanced antioxidativity. Rubus fruticosus L. Cacak Thornless cultivar, the only autochthonous berry fruit variety screened herein, may be firmly recommended as a raw material for industrial production of low sugar blackberry jellies with exceptional characteristics. This innovative procedure of preparing berry fruit jellies have encompassed the application of low temperature regime (55 °C), lower content of sugar (40%), seeds separation from jellies followed by no use of pectin throughout the whole process.

Evaluation of Apple Pomace Flour Obtained Industrially by Dehydration as a Source of Biomolecules with Antioxidant, Antidiabetic and Antiobesity Effects.

Apple pomace flour (APF) obtained at industrial scale level by the application of innovative technological process (dehydration (5 h, T ≤ 55 °C), grinding (300 µm)) was evaluated as a source of bioactive compounds with antioxidative, antiobesity and antidiabetic effects. Proximate composition, individual (HPLC-DAD-MS/MS) and total phenols (TPC) as well as flavonoids content (TFC), antioxidant (AO) activity (DPPH, ABTS, HPMC), water and oil holding capacity (WHC and OHC) of APFs obtained from apple pomace from mixed and individual apple cultivars grown conventionally and organically were compared. The effect of APF supplementation on the glycaemic status and glucose tolerance (oral glucose tolerance test (OGTT)) of C57BL/6J mice exposed to high-fat and sucrose diet was examined. High K content (4.2-6.4 g/kg), dietary fibres (35-45 g/100 g), TPC (4.6-8.1 mg GAE/g), TFC (18.6-34.6 mg QE/g), high water and oil holding capacity (4.7-6.4 and 1.3-1.6 g/g) were observed in the APFs. Content of major phenols (phlorizin, chlorogenic acid, quercetin), TPC and TFC correlated highly with prominent AO activity. APF supplementation lowered the increase of body weight gain and blood glucose, and improved glucose tolerance significantly. Health-promoting biomolecules, AO activity, functional properties and prevention of diet-driven glucose metabolism disorders pave the way to APF exploitation in human nutrition.

Functionality and Storability of Cookies Fortified at the Industrial Scale with up to 75% of Apple Pomace Flour Produced by Dehydration.

Apple pomace flour (APF) with high content of dietary fibers (DF), total polyphenolics (TPCs) and flavonoids (TFCs) was produced at the industrial scale. Bulk and tapped density, swelling, water and oil holding capacity, solubility and hydration density of fine and coarse APF with average particle size 0.16 and 0.50 mm were compared. The effect of wheat flour substitution with 25%, 50% and 75% of fine and coarse APF was studied upon cookies production at the industrial scale and after one year of storage. Coarse APF performed better in respect to sensorial properties, content and retention of dietary compounds and antioxidant (AO) activity. The cookies with optimal share of coarse APF (50%) contained 21 g/100 g of DF and several times higher TPC, TFC as well as AO activity than control cookies, retained well health promoting compounds and maintained an intensely fruity aroma and crispy texture. They were acceptable for consumers according to the hedonic test.