Short-chain Peptides Research Articles

BIOLOGICALLY ACTIVE PROPERTIES OF HYDROLYSED AND FERMENTED MILK PROTEINS

A comparative analysis of protein and peptide profile, biological activities of hydrolysed bovine colostrum and whey was performed. It was found that the depth of proteolysis, qualitative and quantitative composition of protein component of samples determined the level of their antiradical, antimutagenic, antimicrobial and antigenic properties. SDS electrophoresis of experimental samples revealed more extensive protein hydrolysis in the course of alcalase treatment than during enzymatic reaction with neutrase. Using fluorimetric method the influence of hydrolysis with endopeptidases and fermentation with Lactobacillus acidophilus on antioxidant properties of milk proteins was established. 1.7–5.5 times increase in antiradical activity of derived samples in comparison with native proteins was recorded. Reduction in mutation rate induced by whey hydrolysate in tested strain Salmonella typhimurium TA 98 ranged from 15.7 % to 49.2 %, whereas antimutagenic effect on strain TA 100 varied from 18.8 % to 52.1 %, exceeding the similar values shown by colostrum hydrolysates. Samples of colostrum and whey hydrolysed with alcalase are enriched with specific short-chain peptides which determine their relatively high antimutagenic and antiradical properties. Immunoprecipitation reaction demonstrated effective splitting of β lactoglobulin by alcalase, resulting in production of hypoallergenic hydrolysates. It was found by impedimetric technique that neutrase-cleaved colostrum accounted for maximum inhibition of Escherichia coli АТСС 8739 (82 %) and Staphylococcus aureus АТСС 6538 (19 %). Samples of enzymatic hydrolysates of colostrum and whey proteins with confirmed antimicrobial, antimutagenic and antioxidant action were obtained. The use of hydrolysed and fermented colostrum with elevated antioxidant potential in special nutrition appears extremely promising.

Presence of Exopeptidase-Resistant and Susceptible Peptides in a Bacterial Protease Digest of Corn Gluten.

Corn gluten hydrolysate (CGH) was prepared by food-grade bacterial proteases, alcalase and neutral protease. Digestion of CGH with carboxypeptidase A and leucine aminopeptidase extensively changed the elution patterns of peptides as observed from reversed phase high performance liquid chromatography-mass spectrometry (LC-MS), whereas digestion with pepsin and trypsin hardly affected the elution patterns. Twenty-five major peptides in CGH were identified. After digestion with exopeptidases, only prolyl dipeptides and pyroglutamyl di- and tripeptides remained, whereas the other 17 peptides completely disappeared. On the other hand, all 25 peptides remained after digestion with pepsin and trypsin. These facts suggest that a majority of short-chain peptides in food protein hydrolysates are degraded by exopeptidases during digestion and absorption processes. Thus, susceptibility to exopeptidases should be considered for prediction of bioactive peptide upon ingestion, which has not been considered in most of previous studies on food-derived bioactive peptides.

SEARCH FOR NEW OLIGOPEPTIDE MOLECULES FOR PHARMACOLOGICAL CORRECTION OF NSAIDS-INDUCED GASTRIC ULCER

This article presents the results of a study aimed at determining the gastroprotective activity of a number of short-chain peptides against the background of experimental gastropathy in rats. Indomethacin ulcer was used as a model of ulceration. Omeprazole, a drug of basic therapy, was used as a comparison drug. According to the results of the study, two peptides — tetrapeptide LA-4 and hexapeptide LF-6 — demonstrated the gastroprotective activity. The effect was manifested in a signifi cant decrease in the average number of erosions on the surface of the gastric mucosa, their depth, the number of stripe-shaped erosions and Paul’s Index. At the same time, the LF-6 peptide had an effect comparable to that of omeprazole, which makes it promising and interesting for further study.

Efficacy of Indolicidin, Cecropin A (1-7)-Melittin (CAMA) and Their Combination Against Biofilm-Forming Multidrug-Resistant Enteroaggregative Escherichia coli.

The present study examined the anti-biofilm efficacy of two short-chain antimicrobial peptides (AMPs), namely, indolicidin and cecropin A (1-7)-melittin (CAMA) against biofilm-forming multidrug-resistant enteroaggregative Escherichia coli (MDR-EAEC) isolates. The typical EAEC isolates re-validated by PCR and confirmed using HEp-2 cell adherence assay was subjected to antibiotic susceptibility testing to confirm its MDR status. The biofilm-forming ability of MDR-EAEC isolates was assessed by Congo red binding, microtitre plate assays and hydrophobicity index; broth microdilution technique was employed to determine minimum inhibitory concentrations (MICs) and minimum biofilm eradication concentrations (MBECs). The obtained MIC and MBEC values for both AMPs were evaluated alone and in combination against MDR-EAEC biofilms using crystal violet (CV) staining and confocal microscopy-based live/dead cell quantification methods. All the three MDR-EAEC strains revealed weak to strong biofilm-forming ability and were found to be electron-donating and weakly electron-accepting (hydrophobicity index). Also, highly significant (P < 0.001) time-dependent hydrodynamic growth of the three MDR-EAEC strains was observed at 48h of incubation in Dulbecco's modified Eagle's medium (DMEM) containing 0.45% D-glucose. AMPs and their combination were able to inhibit the initial biofilm formation at 24h and 48h as evidenced by CV staining and confocal quantification. Further, the application of AMPs (individually and combination) against the preformed MDR-EAEC biofilms resulted in highly significant eradication (P < 0.001) at 24h post treatment. However, significant differences were not observed between AMP treatments (individually or in combination). The AMPs seem to be an effective candidates for further investigations such as safety, stability and appropriate biofilm-forming MDR-EAEC animal models.

Modeling of probiotic gel shampoo recipes with vegetable extracts and marketing analysis of their market promotion

The object of research is the formulation of probiotic gel shampoos enriched with short-chain peptides of whey proteins and probiotic lysate. The intensive growth in demand for new types of cosmetics, including cosmetics with probiotics, causes the desire of manufacturers to constantly improve products to meet consumer requirements. Of the whole complex of factors that form the final quality of natural cosmetics, from the point of view of the consumer, one of the most important is the planning and development of component content of products. Therefore, a promising task is studying the possibilities of creating probiotic gel shampoos based on available natural components, which, thanks to the mathematical modeling of formulations and the use of promising technologies, allows to predict their high popularity and effectiveness.The work substantiates the feasibility of using whey nano-filtrate as an extractant to extract biologically active substances from medicinal raw materials for the production of probiotic gel shampoos. The expediency of using extracts of peppermint grass, nettle leaves and marigold flowers as part of probiotic gel shampoos enriched with short-chain peptides of whey proteins and probiotic lysate has been shown. Due to the optimized ratio of medicinal plant extracts in the composition, the target product has high functional properties. The ratio of extracts of nettle leaves, marigold flowers and peppermint grass is 2:1:1, while the mixture of extracts has a maximum antioxidant activity of 1262.255 act. units. As a result of the research, «Probio-beauty» probiotic gel shampoo formulations are developed for daily use and for strengthening and restoring hair. Quality indicators developed with optimized formulations of «Probio-beauty» probiotic gel shampoos meet the requirements of regulatory documents.Since today there are no cosmetic products with probiotics produced in Ukraine on the market, the implementation of research will allow to occupy the niche of «natural cosmetics». And correctly conducted marketing activities that are offered in this work will allow to gain a foothold in the cosmetics market, make the brand recognizable and expand the product range based on consumer preferences.

Potent activity of bioconjugated peptide and selenium nanoparticles against colorectal adenocarcinoma cells

Nanomaterial based anticancer treatment is promising nowadays because of their small size that can penetrate and interact both inside and outside the cell surface. In this study, a simple protocol was followed for the conjugation of the biologically synthesized selenium nanoparticles (SeNPs) and short chain synthetic peptide. SeNPs was synthesized by using the culture supernatant of Streptomyces griseoruber, actinomycetes isolated from the soil. The short chain peptide Boc-L-F-OMe was synthesized by the conventional solution phase chemistry using a racemization-free fragment condensation strategy. Peptide interaction with different anticancer receptors was preliminarily studied by docking studies. Biosynthesized SeNPs was conjugated with short chain synthetic peptides by means of cysteine conjugation. Characterization of SeNPs with peptide was done by UV–visible spectroscopy and DLS that showed the red shift in the peak and increase in average particle size and zeta potential, respectively. Bioconjugated SeNPs- peptide was tested for its cytotoxicity against the colon cancer cell line HT-29. Bioconjugated SeNPs-peptide showed enhanced cytotoxic activity when compared to the peptide and nanoparticle alone that was tested at 10–50 µg/ml concentration. Further apoptotic studies were done by AO/PI staining and DNA fragmentation assay that confirms the cytotoxicity of the conjugates. Novel peptide-SeNPs conjugates tested in our study has a significant anticancer activity that can be potentially used for targeting the cancer cells.

Synthesis and Characterization of Peptide-Chitosan Conjugates (PepChis) with Lipid Bilayer Affinity and Antibacterial Activity.

Antimicrobial peptides appear among innovative biopolymers with potential therapeutic interest. Nevertheless, issues concerning efficiency, production costs, and toxicity persist. Herein, we show that conjugation of peptides with chitosans can represent an alternative in the search for these needs. To increase solubility, deacetylated and degraded chitosans were prepared. Then, they were functionalized via N-succinimidyl- S-acetylthiopropionate or via glutathione (GSH), an endogenous peptide linker. To the best of our knowledge, it is the first time that GSH is used as a thiolating agent for the conjugation of peptides. Next, thiolated chitosans were conjugated through a disulfide bond with designed short-chain peptides, one of them derived from the antimicrobial peptide Jelleine-I. Conjugates and respective reaction intermediates were characterized by absorciometry, attenuated total reflectance-Fourier transform infrared, and 1H NMR. Zeta potential measurements showed the cationic nature of these biomacromolecules and their preferential partitioning to Gram-positive bacterial-like model membranes. In vitro investigation using representative Gram-positive and -negative bacteria ( Staphylococcus aureus and Escherichia coli, respectively) showed that the conjugation strategies lead to enhanced activity in relation to the unconjugated peptide and to the unconjugated chitosan. The obtained products showed selectivity toward S. aureus at low cytotoxicity as determined in NIH/3T3 cells. Overall, our study demonstrates that an appropriate choice of antimicrobial peptide and chitosan characteristics leads to increased antimicrobial activity of the conjugated product and represents a strategy to modulate the activity and selectivity of antimicrobials resorting to low-cost chemicals. The present proposal starts from less expensive raw materials (chitosan and short-chain peptide), is based on aqueous solvents, and minimizes the use of reactants with a higher environmental impact. The final biopolymer contains the backbone of chitosan, just 3-6% peptide derived from royal jelly and GSH, all of them considered safe for human use or as a physiological molecule.

High resolution liquid chromatography tandem mass spectrometry for the separation and identification of peptides in coffee silverskin protein hydrolysates

An analytical methodology was developed for the first time in this work to investigate the peptide composition of coffee silverskin protein hydrolysates. Coffee silverskin is the only by-product produced in the coffee roasting process and it contains a relatively high amount of proteins (16.2–19.0%). Different extraction procedures were tested to obtain protein extracts from coffee silverskin samples which were subsequently submitted to enzymatic digestion using different enzymes. Protein hydrolysates from Arabica coffee silverskin obtained using three roasting degrees (light, medium and dark) were considered in order to evaluate the influence of this process on peptide composition. Antioxidant and hypocholesterolemic activities were investigated for these hydrolysates. A method based on the use of liquid chromatography coupled to a quadrupole-time-of-flight mass spectrometer was developed enabling the separation and identification of different short chain peptides in the coffee silverskin hydrolysates using de novo sequencing tool. Different peptides, with a number of amino acids ranging from 4 to 12, were identified in the coffee silverskin analyzed. Peptides obtained were different depending on the enzymatic hydrolysis employed. As general trend, the results obtained showed that peptide composition in coffee silverskin protein hydrolysates was not significantly affected by the coffee roasting process.

Scope and Limitations of Boron Fluorescent Complexes from Stable Nitrile Oxides in ABPP Assays.

Anthracenenitrile oxide undergoes 1,3-dipolar cycloaddition reactions with 1-substituted-4-(prop-2-yn-1-yloxy)benzene affording the expected isoxazoles in good yields and as single regioisomers. N–O Bond cleavage and boron complexation afforded the corresponding boron complexes, derivatized with either a triple bond for click-chemistry applications or an oxime group for nitrile oxide 1,3-dipolar cycloaddition. The optical properties of these compounds show potential for employment as fluorescent tags in imaging techniques. The activity-based protein profiling assays showed good reactivity for the synthesized probes even with short peptide chains (epoxomicin). Scope and limitations are discussed in the light of the obtained results.

ReaxFF MD Simulations of Peptide-Grafted Gold Nanoparticles.

Functionalized gold nanoparticles have critical applications in biodetection with surface-enhanced Raman spectrum and drug delivery. In this study, reactive force field molecular dynamics simulations were performed to study gold nanoparticles, which are modified with different short-chain peptides consisting of amino acid residues of cysteine and glycine in different grafting densities in the aqueous environment. Our study showed slight facet-dependent peptide adsorption on a gold nanoparticle with the 3 nm core diameter. Peptide chains prefer to adsorb on the Au(111) facet compared to those on other facets of Au(100) and Au(110). In addition to the stable thiol interaction with gold nanoparticle surfaces, polarizable oxygen and nitrogen atoms show strong interactions with the gold surface and polarize the gold nanoparticle surfaces with an overall positive charge. Charges of gold atoms vary according to their contacts with peptide atoms and lattice positions. However, at the outmost peptide layer, the whole functionalized Au nanoparticles exhibit overall negative electrostatic potential due to the grafted peptides. Moreover, simulations show that thiol groups can be deprotonated and subsequently protons can be transferred to water molecules and carboxyl groups.

Separation and identification of peptides in hydrolysed protein extracts from edible macroalgae by HPLC-ESI-QTOF/MS

Macroalgae contain significant amounts of high-quality proteins which, because of their structural diversity, contain a range of yet undiscovered peptides within their primary structures. In this work, an analytical methodology was developed for the separation and identification of peptides present in protein hydrolysates from three different edible macroalgae used for human consumption (Saccharina latissima (brown macroalga), Codium spp. (green macroalga), and Mastocarpus stellatus (red macroalga)). The extraction of aqueous and alkaline soluble proteins was carried out followed by their precipitation with HCl or acetone. The protein extracts obtained were submitted to enzymatic digestion with alcalase and subsequently analyzed by reversed-phase high-performance liquid chromatography-quadrupole-time-of flight mass spectrometry (RP-HPLC-QTOF/MS) and de novo sequencing tool to separate and identify different short chain peptides. Thirty-seven peptides were identified in the hydrolysed protein extracts from the three macroalgae, five of them being common in brown and red macroalgae. After checking against BIOPEP database, several sequenced peptides were found within longer peptides with potential antibacterial activity. Any of the identified peptides had previously been identified in macroalgae.

Thermally-induced formation of taste-active 2,5-diketopiperazines from short-chain peptide precursors in cocoa.

2,5-diketopiperazines (DKPs) are cyclic dipeptides responsible for the specific bitter taste of cocoa formed during roasting. The 2,5-diketopiperazine and peptide composition of four different cocoa bean samples from different origins were studied using LC-MS techniques. 34 diketopiperazines were identified, of which 10 are newly reported in cocoa. Their formation was followed during two different roasting time-series using a zero-order and an alternative Prout-Tompkins solid-state kinetic models. The activation energies of diketopiperazine formation showed a distribution close to normal with individual values depending on the nature of the substituents. The relative concentrations of the DKPs were correlated with their putative peptide precursors in unroasted cocoa bean samples. The results showed a significant positive correlation, indicating that oligopeptides formed in cocoa bean fermentation are taste-precursors for bitter tasting diketopiperazines. Unexpectedly, for most diketopiperazines, a single major peptide precursor could be suggested.

OPTIMIZATION OF PARAMETERS OF FERMENTOLYSIS OF PROTEINS IN THE COMPOSITION OF SERUM-PROTEIN CONCENTRATE

The food industry is a strategic industry that works quite steadily even during periods of economic crises, providing food security to any state, and is a source of raw material for other industries with a high potential for development, for example, for the production of cosmetics. The modern cosmetics market is represented by various cosmetic products, often expensive, but not always made from natural ingredients. Therefore, the search for the newest ingredients for the production of natural cosmetics on the basis of domestic raw materials is an urgent task of the present. Ingredients from milk serum can be used for the production of natural cosmetics that in large quantities is obtained in milk processing enterprises and often remains unprocessed. Whey protein concentrations can be a source of short-chain peptides and free amino acids for the production of various cosmetic products. The process of fermentolysis of serum proteins in nanofiltration concentrate KSB-65 with the content of dry matter of 20% using neutral peptidase C from the domestic producer at a temperature of 40 ºС with the duration of the process varying from 1 to 5 hours, the content of peptidase – from 0,5 to 2,0 U/g. It is established that the optimal parameters of the fermentolysis of serum proteins in KSB-65 are as follows: temperature 40 º C, neutral peptidase C content – 0.78 U/g, duration of fermentolysis – 3.17 hours. With optimal parameters of the fermentolysis process, the hydrolyzate of the nanofiltration concentrate KSB-65 contains the maximum amount of short chain peptides (57.03 mg/cm3) and a high concentration of free amino acids (54.66 μg/cm3). Recommendations for the further use of serum protein hydrolyzate obtained using the recommended optimal parameters of the enzyme production process from the nanofiltration concentrate KSB-65, in the manufacture of cosmetic products, including with anti-age effect, and hydrolyzates of proteins enriched with probiotic cultures of lactam bifidobacteria or their lysates.

Casein-derived peptides as an alternative ingredient for low-phenylalanine diets.

Introduction: casein-derived peptides can be liberated both in vivo via normal digestion of casein, as well as in vitro via enzymatic hydrolysis. These peptides were suggested to have biological activity. Objectives: the aim of this study was to describe the production and characterization of casein peptides and to explore the potential of these peptides as an option for low-phenylalanine diets. Methods: peptides were produced by tryptic hydrolysis of sodium caseinate and acid precipitation with HCl, followed by precipitation with ethanol or aggregation of CaCl2 or ZnSO4. Results: the amino acid analysis revealed a significant reduction in the amount of phenylalanine from the original protein. Conclusion: casein-derived peptides could be a future alternative of short chain peptides to low-phenylalanine formulations.

Screening of Angiotensin-I Converting Enzyme Inhibitory Peptides Derived from Caulerpa lentillifera.

Peptides with angiotensin-I converting enzyme (ACE) inhibitory activity have received considerable interest due to their potential as antihypertensive agents and consumer concern over the safety of synthetic drugs. The objective of this study was to isolate ACE inhibitory (ACEI) peptides from Caulerpa lentillifera (known commonly as sea grape) protein hydrolysate. In this study, short-chain peptides were obtained after hydrolysis by various enzymes and subsequently by ultrafiltration. Thermolysin hydrolysate showed the highest ACEI activity. Bioassay-guided fractionation was performed using reversed-phase high performance liquid chromatography (RP-HPLC) to uncover the fraction 9 with the highest ACE inhibitory activity from thermolysin hydrolysate. Peptides in this fraction were further identified using liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis coupled with de novo sequencing, which gave two oligopeptides, FDGIP (FP-5) and AIDPVRA (AA-7). The identities and activities of these two peptides were further confirmed using synthetic peptides. Their IC50 values were determined as 58.89 ± 0.68 µM and 65.76 ± 0.92 µM, respectively. Moreover, the inhibition kinetics revealed that both FP-5 and AA-7 are competitive inhibitors. These activities were further explained using molecular docking simulation. The present study is the first report about ACEI peptides derived from Caulerpa lentillifera and it shows the potential for preventing hypertension and for functional food development.

Current Developments in Pt(IV) Prodrugs Conjugated with Bioactive Ligands.

To overcome the side effects of and resistance to cisplatin, a variety of Pt(IV) prodrugs were designed and synthesized via different modifications including combination with lipid chains to increase hydrophobicity, conjugation with short peptide chains or nanoparticles to improve drug delivery, or addition of bioactive ligands to the axial positions of Pt(IV) complexes to exert dual-function effects. This review summarizes the recent progress in the development of Pt(IV) prodrugs conjugated with bioactive-targeting ligands, including histone deacetylase inhibitors, p53 agonists, alkylating agents, and nonsteroidal anti-inflammatory agents. Although Pt(IV) complexes that conjugated with bioactive ligands show satisfactory anticancer effects, none has been approved for clinical use. Therefore, we hope that this review will contribute to further study and development of Pt(IV) complexes conjugated with bioactive and other ligands.

Cytochrome c electrochemistry on peptide self-assembled monolayers

A short chain peptide is covalently attached to an evaporated gold electrode to form a biocompatible self-assembled monolayer (SAM) surface. The peptide is designed to optimize the interfacial interaction of horse heart cytochrome c (cytc) through the electrostatic interaction of the carboxylate group on the terminal amino acid glutamate and the amine on lysine in cytc. Cytc exhibits quasi-reversible voltammetry and is stable on the time scale of the experiments. Marcus theory is used to calculate the electron transfer rates, which are ~5 times faster for cytc/peptide/Au compared to cytc/11-mercaptoundecanoic acid/Au SAMs of similar chain length. The formal potentials also shift for the cytc/peptide system ~20 mV more positive than cytc adsorbed to 11-mercaptoundecanoic acid-SAM/Au electrodes.

QUANTUM-CHEMICAL MODELING OF GLUTATHIONE-PECTIN SYSTEM AND EVALUATION OF THE POSSIBILITY OF THEIR USING IN THE FOODSTUFFS COMPOSITION

OF THE POSSIBILITY OF THEIR USING IN THE FOODSTUFFS COMPOSITIONThe article presents the results of the analytical study of glutathiones properties, as a short chain peptide, and analyzed the possibility of increasing the nutritional and biological value due to its introduction into food products. Glutathione activates assimilation and dismulsion processes in the organism, the results of which are the cells rejuvenation, their protection from negative factors of influence and mutations. Glutathione, like all short-chain peptides, has the ability to break down under the influence of various factors (acidity of the stomach, mechanical effects, and the appearance of moisture in dry foods). The article proposes the solution of the problem of glutathione lack in the organism and the difficulties of the restoration of its level in the organism by means of its stabilization in the pectin matrix. The article describes a method of combining glutathione with functional groups of galacturonic acid residues, using quantum-chemical modeling.

A fluorescence resonance energy transfer biosensor based on carbon dots and gold nanoparticles for the detection of trypsin

Herein we propose a fluorescence turn-on strategy for the sensitive and selective detection of trypsin based on fluorescence resonance energy transfer (FRET) between gold nanoparticles (AuNPs) and amino-functionalized carbon dots (C-dots). In this assay, C-dots were treated as fluorometric reporter, while AuNPs were treated as fluorescence quencher. A specially designed negatively charged short peptide chain with one functioned cysteine group as the substrate for trypsin. The sensor works as follows: the negatively charged part of the peptide can be hydrolyzed by trypsin, resulting in the releasing of shorter and positively charged peptides. These shorter peptides then induces AuNPs aggregation due to the strong electrostatic interaction between positively charged peptides and the negatively charged AuNPs, thus leading to the restoration of the FRET-quenched fluorescence emission of C-dots. Measuring the changes in the fluorescence of C-dots, the concentrations of trypsin can be determined. Under the optimized conditions, we achieved quantitative evaluation of trypsin in a range of 2.5–80 ng mL−1 with the detection limit of 0.84 ng mL−1. Meanwhile, this sensing system also exhibited excellent selectivity and sensitivity for trypsin, and we successfully applied it for the detection of trypsin in human serum samples. Based on the above findings, we conclude that this new FRET based sensor might be significant in disease diagnosis in the future.

Inorganic and Organic Clusters Formed upon Surface-Assisted Laser Desorption/Ionization

Examples of cluster formation from inorganic and organic compounds upon surface-assisted laser desorption/ionization have been considered. It has been shown that the cluster formation is especially efficient upon ionization of silver halides. The addition of silver salts to salts of other metals, in particular, lead and nickel salts and sodium stannate, leads to the formation of mixed clusters. The peak intensities of the ions of such clusters are higher than those of the clusters of initial salts, thereby indicating a higher efficiency of their formation. Nonpolar amino acids and short-chain peptides predominantly form adducts with alkali-metal ions.