Liposome Model System Research Articles

Benefits of Combined Fluorescence Lifetime Imaging Microscopy and Fluorescence Correlation Spectroscopy for Biomedical Studies Demonstrated by Using a Liposome Model System

Drug delivery systems play a pivotal role in targeted pharmaceutical transport and controlled release at specific sites. Liposomes, commonly used as drug carriers, constitute a fundamental part of these systems. Moreover, the drug–liposome model serves as a robust platform for investigating interaction processes at both cellular and molecular levels. To advance our understanding of drug carrier uptake mechanisms, we employed fluorescence lifetime imaging microscopy (FLIM) and fluorescence correlation spectroscopy (FCS), leveraging the unique benefits of two-photon (2P) excitation. Our approach utilized giant unilamellar vesicles (GUVs) as a simplified model system for cell membranes, labelled with the amphiphilic fluorescent dye 3,3′-dioctadecyloxa-carbocyanine (DiOC18(3)). Additionally, large unilamellar vesicles (LUVs) functioned as a drug carrier system, incorporating the spectrally distinct fluorescent sulforhodamine 101 (SRh101) as a surrogate drug. The investigation emphasized the diverse interactions between GUVs and LUVs based on the charged lipids employed. We examined the exchange kinetics and structural alterations of liposome carriers during the uptake process. Our study underscores the significance of employing 2P excitation in conjunction with FLIM and FCS. This powerful combination offers a valuable methodological approach for studying liposome interactions, positioning them as an exceptionally versatile model system with a distinct technical advantage.

Development of novel broad-spectrum antimicrobial lipopeptides derived from plantaricin NC8 β

Bacterial resistance towards antibiotics is a major global health issue. Very few novel antimicrobial agents and therapies have been made available for clinical use during the past decades, despite an increasing need. Antimicrobial peptides have been intensely studied, many of which have shown great promise in vitro. We have previously demonstrated that the bacteriocin Plantaricin NC8 αβ (PLNC8 αβ) from Lactobacillus plantarum effectively inhibits Staphylococcus spp., and shows little to no cytotoxicity towards human keratinocytes. However, due to its limitations in inhibiting gram-negative species, the aim of the present study was to identify novel antimicrobial peptidomimetic compounds with an enhanced spectrum of activity, derived from the β peptide of PLNC8 αβ. We have rationally designed and synthesized a small library of lipopeptides with significantly improved antimicrobial activity towards both gram-positive and gram-negative bacteria, including the ESKAPE pathogens. The lipopeptides consist of 16 amino acids with a terminal fatty acid chain and assemble into micelles that effectively inhibit and kill bacteria by permeabilizing their cell membranes. They demonstrate low hemolytic activity and liposome model systems further confirm selectivity for bacterial lipid membranes. The combination of lipopeptides with different antibiotics enhanced the effects in a synergistic or additive manner. Our data suggest that the novel lipopeptides are promising as future antimicrobial agents, however additional experiments using relevant animal models are necessary to further validate their in vivo efficacy.

Cholesterol content regulates silica-induced lysosomal membrane permeability.

Inhalation of crystalline silica has been well documented to cause pulmonary inflammation and lung disease such as silicosis. Respirable silica particles deposit in the lungs and are phagocytosed by alveolar macrophages. Subsequently, phagocytosed silica remains undegraded within lysosomes causing lysosomal damage known as phagolysosomal membrane permeability (LMP). LMP can trigger the assembly of the NLRP3 inflammasome resulting in release of inflammatory cytokines that contribute to disease. In order to better understand the mechanisms of LMP this study used murine bone marrow derived macrophages (BMdM) as a cellular model to investigate the mechanism of silica-induced LMP. Reduction of lysosomal cholesterol in bone marrow derived macrophages with 18:1 phosphatidylglycerol (DOPG) liposome treatment increased silica-induced LMP and IL-1β release. Conversely, increasing lysosomal and cellular cholesterol with U18666A reduced IL-1β release. Co-treatment of bone marrow derived macrophages with 18:1 phosphatidylglycerol and U18666A resulted in a significant reduction of the effects of U18666A on lysosomal cholesterol. Phosphatidylcholine 100-nm liposome model systems were used to examine the effects of silica particles on lipid membrane order. Time-resolved fluorescence anisotropy of the membrane probe, Di-4-ANEPPDHQ, was used to determine changes to membrane order. Silica increased lipid order that was attenuated by inclusion of cholesterol in the phosphatidylcholine liposomes. These results demonstrate that increased cholesterol can attenuate silica-induced membrane changes in liposomes and cell models, while decreasing cholesterol exacerbates silica-induced membrane changes. Selective manipulation of lysosomal cholesterol may be a way of attenuating lysosomal disruption and preventing silica-induced chronic inflammatory disease progression.

Role of Stingray (Himantura signifier) Non-Protein Nitrogenous Fraction on the Oxidative Stability of Lipid and Myoglobin.

Non-protein nitrogen (NPN) is abundant in stingray (Himantura signifier) muscle, which also has in vitro antioxidant activity. In this study, NPN from stingray muscle was further investigated for its antioxidant properties in lecithin liposome and oxymyoglobin model systems to validate its protective impact against lipid and myoglobin oxidations during storage for 120 min at various temperatures (4, 25, and 60 °C). NPN solution (10 ppm nitrogen) was added to the lecithin liposome system at different concentrations (0, 0.5, 1, 5, and 10% (v/v)) to investigate its effects on lipid stability by measuring the conjugated diene (CD), peroxide value (PV), and thiobarbituric acid reactive substances (TBARS) contents. In the oxymyoglobin system, NPN solution (10 ppm nitrogen) was also added at different concentrations (0, 0.5, 1, 5, and 10% (v/v)) to the oxymyoglobin solution in order to examine its effect on the stability of myoglobin by determining the contents of oxymyoglobin, metmyoglobin, and protein carbonyl. According to the findings, in all NPN concentrations, the system incubated at 4 °C had the lowest levels of lipid oxidation as measured by CD, PV, and TBARS values, and the lowest levels of myoglobin oxidation. At all incubating temperatures, the oxymyoglobin and lipid oxidation of all model systems tended to rise with the lengthening of the incubation duration. With the addition of 5% NPN, however, the lowest CD, PV, TBARS, oxymyoglobin oxidation, metmyoglobin formation, and protein carbonyl content were all observable, and the remarkable result was discovered during incubation at 4 °C. The results indicate that stingray NPN, especially at 5%, can be used to delay lipid and myoglobin oxidation, particularly at 4 °C. In order to prolong the shelf life of products with dark-fleshed fish and red meat, stingray NPN might be used as an alternative antioxidant to delay the oxidation of lipid and myoglobin during cold chain storage.

Exploring the potential effect of phospholipase A2 antibody to extend beef shelf-life in a beef liposome model system

The objective of this study was to elucidate the effect of phospholipase A2 (PLA2) and a PLA2 antibody (aPLA2) on phospholipid (PL) hydrolysis in beef and to understand how the altered PL composition may affect lipid oxidation and antioxidant capacity of beef in an in vitro system. Various combinations of PLA2 and aPLA2 were introduced to a beef liposome model system and exposed to a retail display. The PL and free fatty acid (FFA) profiles, antioxidant capacity and lipid oxidation were measured for the liposome system. Key PL classes were reduced and the release of polyunsaturated FFAs was increased with the inclusion of PLA2 in the treatments (P < 0.05). There was no inhibition of PL hydrolysis with the addition of aPLA2. PLA2 showed strong antioxidant capacity in the liposome system (P < 0.01), but lipid oxidation still increased in samples treated with PLA2 throughout the retail display (P < 0.01). Finally, aPLA2 treatments demonstrated potential to decrease lipid oxidation (P < 0.01).

Synthesis and Characterization of Ceramide-Containing Liposomes as Membrane Models for Different T Cell Subpopulations

A fine balance of regulatory (Treg) and conventional CD4+ T cells (Tconv) is required to prevent harmful immune responses, while at the same time ensuring the development of protective immunity against pathogens. As for many cellular processes, sphingolipid metabolism also crucially modulates the Treg/Tconv balance. However, our understanding of how sphingolipid metabolism is involved in T cell biology is still evolving and a better characterization of the tools at hand is required to advance the field. Therefore, we established a reductionist liposomal membrane model system to imitate the plasma membrane of mouse Treg and Tconv with regards to their ceramide content. We found that the capacity of membranes to incorporate externally added azide-functionalized ceramide positively correlated with the ceramide content of the liposomes. Moreover, we studied the impact of the different liposomal preparations on primary mouse splenocytes in vitro. The addition of liposomes to resting, but not activated, splenocytes maintained viability with liposomes containing high amounts of C16-ceramide being most efficient. Our data thus suggest that differences in ceramide post-incorporation into Treg and Tconv reflect differences in the ceramide content of cellular membranes.

Design and simulation of the liposomal model by using a coarse-grained molecular dynamics approach towards drug delivery goals

The simulated liposome models provide events in molecular biological science and cellular biology. These models may help to understand the cell membrane mechanisms, biological cell interactions, and drug delivery systems. In addition, the liposomes model may resolve specific issues such as membrane transports, ion channels, drug penetration in the membrane, vesicle formation, membrane fusion, and membrane protein function mechanism. One of the approaches to investigate the lipid membranes and the mechanism of their formation is by molecular dynamics (MD) simulations. In this study, we used the coarse-grained MD simulation approach and designed a liposome model system. To simulate the liposome model, we used phospholipids that are present in the structure of natural cell membranes (1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC) and 1,2-Dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE)). Simulation conditions such as temperature, ions, water, lipid concentration were performed based on experimental conditions. Our results showed a liposome model (ellipse vesicle structure) during the 2100 ns was formed. Moreover, the analysis confirmed that the stretched and ellipse structure is the best structure that could be formed. The eukaryotic and even the bacterial cells have elliptical and flexible structures. Usually, an elliptical structure is more stable than other assembled structures. The results indicated the assembly of the lipids is directed through short-range interactions (electrostatic interactions and, van der Waals interactions). Total energy (Van der Waals and electrostatic interaction energy) confirmed the designed elliptical liposome structure has suitable stability at the end of the simulation process. Our findings confirmed that phospholipids DOPC and DOPE have a good tendency to form bilayer membranes (liposomal structure) based on their geometric shapes and chemical-physical properties. Finally, we expected the simulated liposomal structure as a simple model to be useful in understanding the function and structure of biological cell membranes. Furthermore, it is useful to design optimal, suitable, and biocompatible liposomes as potential drug carriers.

Exploring the Potential Effect of Phospholipase A2 Antibody to Extend Beef Shelf-Life in a Beef Liposome Model System

Objective: The objective of this study was to utilize a beef liposome model system to investigate if phospholipase-A2 antibody (aPLA2) can be used to inhibit phospholipase-A2 (PLA2) activity to potentially improve beef shelf-life.

Electrochemical energy for living systems

Plastoquinone and ubiquinone play essential roles in the electron transport chains of chloroplasts and mitochondria by coupling electron transport to the transfer of protons across membranes. The energy of the resulting proton gradient is used to synthesize adenosine triphosphate (ATP), the energy currency of all life. How did quinones first become involved in this process? We have detected several quinone compounds in carbonaceous meteorites that can transport protons in a liposome model system. It is possible that such compounds were available to transport protons in primitive versions of electron transport in early life.

Enzymatic extraction of antioxidant ingredients from Danish seaweeds and characterization of active principles

The present study investigated the enzymatic extraction of antioxidant ingredients from four Danish seaweeds viz., Fucus vesiculosus, Fucus serratus, Ascophyllum nodosum and Polysiphonia fucoides. The seaweeds were hydrolyzed with five carbohydrases and three proteases. The antioxidant activities and total phenolic content (TPC) of the resulting extracts were studied. Considerable variation was observed in the antioxidant activities and TPC of different enzymatic hydrolysates. Among the 32 enzymatic extracts studied, F. vesiculosus-Alcalase, F. vesiculosus-Viscozyme, F. serratus-Termamyl, and P. fucoides-Termamyl extracts showed high TPC and antioxidant activities. Fractionation and characterization of these extracts revealed that radical scavenging and reducing power was mainly due to the phenolic fractions, whereas protein and polysaccharide fractions mainly contributed to the iron-chelating activity. The antioxidant activity observed in the liposome model system was due to a combined effect of the various fractions and their antioxidant properties. Gallic, gentisic and chlorogenic acids were the main free phenolics in the phenolic fractions. Fucose rich polysaccharides were obtained when F. vesiculosus and F. serratus were hydrolyzed with Viscozyme and Termamyl, respectively. Thus, enzyme assisted extraction is an effective tool for getting antioxidant with specific properties, which can find application in food and cosmetics.

Liposome fusion with orthogonal coiled coil peptides as fusogens: the efficacy of roleplaying peptides.

Biological membrane fusion is a highly specific and coordinated process as a multitude of vesicular fusion events proceed simultaneously in a complex environment with minimal off-target delivery. In this study, we develop a liposomal fusion model system with specific recognition using lipidated derivatives of a set of four de novo designed heterodimeric coiled coil (CC) peptide pairs. Content mixing was only obtained between liposomes functionalized with complementary peptides, demonstrating both fusogenic activity of CC peptides and the specificity of this model system. The diverse peptide fusogens revealed important relationships between the fusogenic efficacy and the peptide characteristics. The fusion efficiency increased from 20% to 70% as affinity between complementary peptides decreased, (from KF ≈ 108 to 104 M−1), and fusion efficiency also increased due to more pronounced asymmetric role-playing of membrane interacting ‘K’ peptides and homodimer-forming ‘E’ peptides. Furthermore, a new and highly fusogenic CC pair (E3/P1K) was discovered, providing an orthogonal peptide triad with the fusogenic CC pairs P2E/P2K and P3E/P3K. This E3/P1k pair was revealed, via molecular dynamics simulations, to have a shifted heptad repeat that can accommodate mismatched asparagine residues. These results will have broad implications not only for the fundamental understanding of CC design and how asparagine residues can be accommodated within the hydrophobic core, but also for drug delivery systems by revealing the necessary interplay of efficient peptide fusogens and enabling the targeted delivery of different carrier vesicles at various peptide-functionalized locations.

Effect of Enzymatic Hydrolysis on the Antioxidant Activity of Red and Green Seaweeds and Characterization of the Active Extracts

AbstractIn the present study, the effect of enzymatic hydrolysis on the antioxidant activity of three red seaweeds (Chondria cornuta, Chondria dasyphylla, and Murrayella periclados) and two green seaweeds (Cladophora sp. and Ulva lactuca) from the Kuwait coast were evaluated. The seaweeds were hydrolyzed with five carbohydrases and three proteases, and the resulting extracts were tested for antioxidant activity. The total phenolic content and yield of the extracts varied greatly depending upon the species and enzyme used for hydrolysis. Of the 40 enzymatic extracts screened for antioxidant activity, the Viscozyme and Alcalase extracts of M. periclados, Neutrase and Ultraflo extracts of Cladophora sp., and Neutrase extracts of C. cornuta had high antioxidant activity compared to other enzymatic extracts in various in vitro assays. Fractionation of the extracts revealed that the radical scavenging and reducing power of the extracts were mainly due to phenolic fractions. In contrast, the iron‐chelating ability was mainly due to protein fractions. The level of prevention of lipid oxidation in the liposome model system varied for different fractions of extracts and did not correlate with total phenolic content and other antioxidant properties. The results of the study show that, by hydrolyzing seaweeds with specific enzymes, customized seaweed extracts with specific bioactivity could be obtained.

Modulation of Coiled-Coil Binding Strength and Fusogenicity through Peptide Stapling.

Peptide stapling is a technique which has been widely employed to constrain the conformation of peptides. One of the effects of such a constraint can be to modulate the interaction of the peptide with a binding partner. Here, a cysteine bis-alkylation stapling technique was applied to generate structurally isomeric peptide variants of a heterodimeric coiled-coil forming peptide. These stapled variants differed in the position and size of the formed macrocycle. C-terminal stapling showed the most significant changes in peptide structure and stability, with calorimetric binding analysis showing a significant reduction of binding entropy for stapled variants. This entropy reduction was dependent on cross-linker size and was accompanied by a change in binding enthalpy, illustrating the effects of preorganization. The stapled peptide, along with its binding partner, were subsequently employed as fusogens in a liposome model system. An increase in both lipid- and content-mixing was observed for one of the stapled peptide variants: this increased fusogenicity was attributed to increased coiled-coil binding but not to membrane affinity, an interaction theorized to be a primary driving force in this fusion system.

Enzyme-assisted extraction of bioactive compounds from brown seaweeds and characterization

Brown algae comprise the largest biomass producers in coastal waters and play important ecological roles. The complex nature of cell wall polysaccharides limits the extraction of bioactive compounds from these seaweeds. The aim of the study was to use enzyme-assisted extraction as a tool to release the bioactive compounds from seven brown seaweeds of Kuwait coast and characterization of the active extracts. The enzymatic extracts obtained by hydrolysing seaweeds with five carbohydrases and three proteases were screened for antioxidant and antimicrobial activity. Yield, total phenolics, and bioactivity were as a function of species difference in cell wall composition and specificity of the enzyme used. Among the six species of brown seaweeds studied, the enzymatic extracts obtained from Sargassum boveanum, Sargassum angustifolium, and Feldmannia irregularis showed high antioxidant activity in different assays. Though antimicrobial activities of the enzymatic extracts were low, Flavourzyme resulted in more number of seaweed extracts with antimicrobial activity against foodborne pathogens. In general, carbohydrases resulted in extracts with high radical scavenging activity whereas proteases resulted in extracts with high iron chelating activity. The extracts with highest antioxidant activity such as S. boveanum-Viscozyme and Alcalase extracts were further fractionated and characterized. The polyphenol and polysaccharide-rich fractions were responsible for the high radical scavenging and reducing power whereas the iron chelating activity and inhibition of lipid oxidation in liposome model system was mainly contributed by polysaccharide and protein-rich fractions. The results of study showed that enzyme-assisted extraction could be useful to make tailor-made seaweed extracts with specific bioactivity.

Comparative Study of Phytochemical Profiles and Antioxidant Properties of Hydrosols from Bulgarian Rosa Alba L. and Rosa Damascena Mill.

This study assessed phytochemical profile and antioxidant capacity of hydrosols, obtained by water-steam distillation of Rosa alba L. and Rosa damascena Mill. plants, grown in experimental field of the Institute of Rose and Essential Oil Plants, Bulgaria. The identification of hydrosols’ compounds was performed using 7890A/5975 GC-MS system, the total phenolic content (TPC) was determined quantitatively using the Folin Ciocalteu reagent, with Gallic acid as the standard and the antioxidant capacity of both hydrosols in concentration of 0.3 % up to 15 % were tested in liposomal suspension, and in systems generating either hydroxyl and superoxide anion radicals. The results showed that hydrosols were rich in geraniol (42.55 % for Rosa alba L., and 27.25 % for Rosa damascena Mill.) and citronellol (28.70 % in both hydrosols). The hydrosolic TPC of Rosa alba L. and of Rosa damascena Mill. were found to be 72 and 32.52 μg/ml of the Gallic acid equivalent, respectively. Both hydrosols demonstrated good capacity to inhibit Fe2+/ascorbic acid - induced lipid peroxidation in egg liposomal model system as the effect of Rosa alba L. hydrosol in all tested concentrations was higher than those of Rosa damascena Mill. However, Rosa damascena Mill. demonstrated better •OH scavenger effect than that of Rosa alba L. These results suggest that the hydrosols from Rosa alba L. and Rosa damascena Mill. have well-expressed antioxidant properties that could be used as antioxidant supplements in conditions with oxidative stress ethology.

Isolation of Fucoxanthin from Brown Algae and Its Antioxidant Activity: In Vitro and 5% Fish Oil‐In‐Water Emulsion

AbstractFucoxanthin, a nonprovitamin A carotenoid, is a yellowish‐brown pigment found abundantly in brown algae. Along with chlorophyll a, it is bound to proteins and acts as a light‐harvesting and a light‐transferring pigment. The occurrence of carotenoids and chlorophylls in photosynthetic tissues complicates the isolation of pure fucoxanthin. The present study deals with a method for the isolation of fucoxanthin by low‐temperature crystallization and testing its antioxidant activity both in in vitro assays and in 5% oil‐in‐water emulsion. The yield of fucoxanthin obtained with this method ranged from 0.1% to 0.5% of dried algae depending on the species. Fucoxanthin showed good α,α‐diphenyl‐β‐picrylhydrazyl radical‐scavenging and iron‐chelating properties. However, it showed low reducing power and was poor in the inhibition of lipid oxidation in a liposome model system. When tested in 5% fish oil‐in‐water emulsion with iron as an oxidation inducer, fucoxanthin showed antioxidant activity by resulting in low levels of volatile oxidation products and low tocopherol loss compared to control and butylated hydroxytoluene.

High Permeation Rates in Liposome Systems Explain Rapid Glyphosate Biodegradation Associated with Strong Isotope Fractionation.

Bacterial uptake of charged organic pollutants such as the widely used herbicide glyphosate is typically attributed to active transporters, whereas passive membrane permeation as an uptake pathway is usually neglected. For 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) liposomes, the pH-dependent apparent membrane permeation coefficients (Papp) of glyphosate, determined by nuclear magnetic resonance (NMR) spectroscopy, varied from Papp (pH 7.0) = 3.7 (±0.3) × 10–7 m·s–1 to Papp (pH 4.1) = 4.2 (±0.1) × 10–6 m·s–1. The magnitude of this surprisingly rapid membrane permeation depended on glyphosate speciation and was, at circumneutral pH, in the range of polar, noncharged molecules. These findings point to passive membrane permeation as a potential uptake pathway during glyphosate biodegradation. To test this hypothesis, a Gram-negative glyphosate degrader, Ochrobactrum sp. FrEM, was isolated from glyphosate-treated soil and glyphosate permeation rates inferred from the liposome model system were compared to bacterial degradation rates. Estimated maximum permeation rates were, indeed, 2 orders of magnitude higher than degradation rates of glyphosate. In addition, biodegradation of millimolar glyphosate concentrations gave rise to pronounced carbon isotope fractionation with an apparent kinetic isotope effect, AKIEcarbon, of 1.014 ± 0.003. This value lies in the range typical of non-masked enzymatic isotope fractionation demonstrating that glyphosate biodegradation was not subject to mass transfer limitations and glyphosate exchange across the cell membrane was rapid relative to enzymatic turnover.

Membrane permeabilization of phosphatidylcholine liposomes induced by cryopreservation and vitrification solutions

Membranes are the primary site of freezing injury during cryopreservation or vitrification of cells. Addition of cryoprotective agents (CPAs) can reduce freezing damage, but can also disturb membrane integrity causing leakage of intracellular constituents. The aim of this study was to investigate lipid-CPA interactions in a liposome model system to obtain insights in mechanisms of cellular protection and toxicity during cryopreservation or vitrification processing. Various CPAs were studied including dimethyl sulfoxide (DMSO), glycerol (GLY), ethylene glycol (EG), dimethyl formamide (DMF), and propylene glycol (PG). Protection against leakage of phosphatidylcholine liposomes encapsulated with carboxyfluorescein (CF) was studied upon CPA addition as well as after freezing-and-thawing. Molecular interactions between CPAs and phospholipid acyl chains and headgroups as well as membrane phase behavior were studied using Fourier transform infrared spectroscopy. A clear difference was observed between the effects of DMSO on PC-liposomes compared to the other CPAs tested, both for measurements on CF-retention and membrane phase behavior. All CPAs were found to inhibit membrane leakiness during freezing. However, exposure to high CPA concentrations already caused leakage before freezing, increasing in the order DMSO, EG, DMF/PG, and GLY. With DMSO, liposomes were able to withstand up to 6M concentrations compared to only 1M for GLY. Cholesterol addition to PC-liposomes increased membrane stability towards leakiness. DMSO was found to dehydrate the phospholipid headgroups while raising the membrane phase transition temperature, whereas the other CPAs caused an increase in the hydration level of the lipid headgroups while decreasing the membrane phase transition temperature.

Analysis of liposome model systems by time‐of‐flight secondary ion mass spectrometry

Time-of-flight secondary ion mass spectrometry (ToF-SIMS) is an important technique for studying chemical composition of micrometer scale objects due to its high spatial resolution imaging capabilities and chemical specificity. In this work we focus on the application of ToF-SIMS to gain insight into the chemistry of micrometer size liposomes as a potential model for neurotransmitter vesicles. Two models of giant liposomes were analyzed: histamine and aqueous two phase system (ATPS)-containing liposomes. Characterization of the internal structure of single fixed liposomes was done both with the Bi3+ and C60+ ion sources. The depth profiling capability of ToF-SIMS was used to investigate the liposome interior.

Synergistic inhibition of lipid oxidation by pea protein hydrolysate coupled with licorice extract in a liposomal model system.

Fourteen pea protein hydrolysates (PPHs) were prepared using different proteases and tested for antioxidant activity in a liposomal model system under oxidative stress (100 μM FeCl3/2 mM ascorbate). Almost all PPHs inhibited lipid oxidation, and those prepared from heated protein with Flavourzyme (Fla-PPH) or Protamex (Pro-PPH) were the most effective. Remarkable synergistic effects were observed on both Fla-PPH and Pro-PPH with licorice extract (LE). Electron microscopy revealed a self-assembled network that appeared to provide crucial protection of liposome against oxidation. The presence of LE enhanced the antioxidant potential by producing a more compact network apparently via PPH-LE complexation. Zeta-potential measurements suggested electrostatic interactions are important driving forces for the accumulation of active peptides at the liposome interface. Peptides rich in leucine, lysine, glutamic acid, glutamine, valine, or proline with a hydrophobic N-terminus, as identified by mass spectrometry, were implicated in the antioxidative protection.