Characterization of carotenoids extracted from Haloferax larsenii NCIM 5678 isolated from Pachpadra salt lake, Rajasthan.

Carotenoids are a diverse group of pigments known for their broad range of biological functions and applications. This study delves into multifaceted potential of carotenoids extracted from Haloferax larsenii NCIM 5678 previously isolated from Pachpadra Salt Lake in Rajasthan, India. H. larsenii NCIM 5678 was able to grow up to OD600 1.77 ± 0.03 with carotenoid concentration, 3.3 ± 0.03 µg/ml. The spectrophotometric analysis of carotenoid extract indicated the presence of three-fingered peak (460, 490 and 520 nm) which is a characteristic feature of bacterioruberin and its derivatives. The bacterioruberin was purified using silica gel column chromatography and thin layer chromatography. The carotenoid extract showed 12.3 ± 0.09 mm zone of growth inhibition with a minimum inhibitory concentration 546 ng/ml against indicator strain, H. larsenii HA4. The percentage antioxidant activity of carotenoid was found to be 84% which was higher as compared to commercially available ascorbic acid (56.74%). Thus, carotenoid extract from H. larsenii NCIM 5678 possesses unique attributes with compelling evidence of antimicrobial and antioxidant potential for the development of novel pharmaceuticals and nutraceuticals.

Probiotics for Aquaculture: Hope, Truth, and Reality.

The use of microorganisms as beneficial crops for human and animal health has been studied for decades, and these microorganisms have been in practical use for quite some time. Nowadays, in addition to well-known examples of beneficial properties of lactic acid bacteria, bifidobacteria, selected Bacillus spp., and yeasts, there are several other bacteria considered next-generation probiotics that have been proposed to improve host health. Aquaculture is a rapidly growing area that provides sustainable proteins for consumption by humans and other animals. Thus, there is a need to develop new technologies for the production practices associated with cleaner and environment-friendly approaches. It is a well-known fact that proper selection of the optimal probiotics for use in aquaculture is an essential step to ensure effectiveness and safety. In this critical review, we discuss the evaluation of host-specific probiotics in aquaculture, challenges in using probiotics in aquaculture, methods to improve the survival of probiotics under different environmental conditions, technological approach to improving storage, and delivery along with possible negative consequences of using probiotics in aquaculture. A critical analysis of the identified challenges for the use of beneficial microbes in aquaculture will help in sustainable aquafarming, leading to improved agricultural practices with a clear aim to increase protein production.

Mechanism of Cell-Killing Activity of Plantaricin LD1 Against Escherichia coli ATCC 25922.

Plantaricin LD1 was purified from a potential probiotic strain, Lactobacillus plantarum LD1 previously isolated from indigenous food, Dosa. In this study, we have performed a detailed mechanism of action of plantaricin LD1 against Escherichia coli ATCC 25922 considering Micrococcus luteus MTCC 106 as control. The plantaricin LD1 showed a minimum inhibitory concentration (MIC) of 34.57 µg/mL and a minimum bactericidal concentration (MBC) of 138.3 µg/mL against M. luteus MTCC 106, whereas MIC 69.15 µg/mL and MBC 276.6 µg/mL were found against E. coli ATCC 25922. The efflux of potassium ions, dissipation of membrane potential (∆ψ), and transmembrane pH gradient (∆pH) of plantaricin LD1-treated cells suggested the membrane-acting nature of plantaricin LD1. Plantaricin LD1 also caused degradation of the genomic DNA of the target strains tested. The cell killing was confirmed by staining with propidium iodide and visualized under light and electron microscopes. The bacteriocin-treated cells were found to be ruptured, swollen, and elongated. Thus, the findings indicate plantaricin LD1 kills E. coli ATCC 25922 by interacting with the cell membrane resulting in the efflux of intracellular contents and also causing degradation of nucleic acids leading to cell death.

Production of fungal phytases in solid state fermentation and potential biotechnological applications.

Phytases are important enzymes used for eliminating the anti-nutritional properties of phytic acid in food and feed ingredients. Phytic acid is major form of organic phosphorus stored during seed setting. Monogastric animals cannot utilize this phytate-phosphorus due to lack of necessary enzymes. Therefore, phytic acid excretion is responsible for mineral deficiency and phosphorus pollution. Phytases have been reported from diverse microorganisms, however, fungal phytases are preferred due to their unique properties. Aspergillus species are the predominant producers of phytases and have been explored widely as compared to other fungi. Solid-state fermentation has been studied as an economical process for the production of phytases to utilize various agro-industrial residues. Mixed substrate fermentation has also been reported for the production of phytases. Physical and chemical parameters including pH, temperature, and concentrations of media components have significantly affected the production of phytases in solid state fermentation. Fungi produced high levels of phytases in solid state fermentation utilizing economical substrates. Optimization of culture conditions using different approaches has significantly improved the production of phytases. Fungal phytases are histidine acid phosphatases exhibiting broad substrate specificity, are relatively thermostable and protease-resistant. These phytases have been found effective in dephytinization of food and feed samples with concomitant liberation of minerals, sugars and soluble proteins. Additionally, they have improved the growth of plants by increasing the availability of phosphorus and other minerals. Furthermore, phytases from fungi have played an important roles in bread making, semi-synthesis of peroxidase, biofuel production, production of myo-inositol phosphates and management of environmental pollution. This review article describes the production of fungal phytases in solid state fermentation and their biotechnological applications.

Enterocin LD3 from Enterococcus hirae LD3 Inhibits the Growth of Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311 in Fruit Juice.

In order to prevent the growth of pathogens in food, bacteriocins produced by various probiotic lactic acid bacteria have been recognized as potential substitutes of chemical preservatives. In this study, enterocin LD3 was purified from the cell-free supernatant of a food isolate, Enterococcus hirae LD3 using multistep chromatography. In the fruit juice, lethal concentration (LC50) of enterocin LD3 was found to be 260 µg/mL against Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311. The cells treated with enterocin LD3 were red colour indicating dead cells after propidium iodide staining, while untreated cells were found blue after staining with 4', 6-diamidino-2-phenylindole. The mechanism of cell killing was analyzed using infrared spectrum of cells treated with enterocin LD3 which was found altered in the range of 1,094.30 and 1,451.82 cm-1 corresponding to nucleic acids and phospholipids, respectively. The morphology of target cells were severely ruptured and lysed as observed under electron microscopy. Thus, the present study suggested that enterocin LD3 showed bactericidal activity against Salm. enterica subsp. enterica serovar Typhimurium ATCC 13311 and may be applied as a bio-preservative for the safety of fruit juices.

Bacteriocin-Producing Probiotic Lactic Acid Bacteria in Controlling Dysbiosis of the Gut Microbiota.

Several strains of lactic acid bacteria are potent probiotics and can cure a variety of diseases using different modes of actions. These bacteria produce antimicrobial peptides, bacteriocins, which inhibit or kill generally closely related bacterial strains and other pathogenic bacteria such as Listeria, Clostridium, and Salmonella. Bacteriocins are cationic peptides that kill the target cells by pore formation and the dissipation of cytosolic contents, leading to cell death. Bacteriocins are also known to modulate native microbiota and host immunity, affecting several health-promoting functions of the host. In this review, we have discussed the ability of bacteriocin-producing probiotic lactic acid bacteria in the modulation of gut microbiota correcting dysbiosis and treatment/maintenance of a few important human disorders such as chronic infections, inflammatory bowel diseases, obesity, and cancer.

Probiotics, prebiotics and synbiotics: Safe options for next-generation therapeutics.

Probiotics have been considered as an economical and safe alternative for the treatment of a large number of chronic diseases and improvement of human health. They are known to modulate the host immunity and protect from several infectious and non-infectious diseases. The colonization, killing of pathogens and induction of host cells are few of the important probiotic attributes which affect several functions of the host. In addition, prebiotics and non-digestible food substances selectively promote the growth of probiotics and human health through nutrient enrichment, and modulation of gut microbiota and immune system. This review highlights the role of probiotics and prebiotics alone and in combination (synbiotics) in the modulation of immune system, treatment of infections, management of inflammatory bowel disease and cancer therapy. KEY POINTS: • Probiotics and their derivatives against several human diseases. • Prebiotics feed probiotics and induce several functions in the host. • Discovery of novel and biosafe products needs attention for human health.

Polydiacetylene vesicles acting as colorimetric sensor for the detection of plantaricin LD1.

The interaction of antimicrobial peptides with membrane lipids plays a major role in numerous physiological processes. In this study, polydiacetylene (PDA) vesicles were synthesized using 10, 12-tricosadiynoic acid (TRCDA) and 1, 2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC). These vesicles were applied as artificial membrane biosensor for the detection of plantaricin LD1 purified from Lactobacillus plantarum LD1. Plantaricin LD1 (200 µg/mL) was able to interact with PDA vesicles by changing the color from blue to red with colorimetric response 30.26 ± 0.59. Nisin (200 µg/mL), used as control, also changed the color of the vesicles with CR% 50.56 ± 0.98 validating the assay. The vesicles treated with nisin and plantaricin LD1 showed increased infrared absorbance at 1411.46 and 1000-1150 cm-1 indicated the interaction of bacteriocins with phospholipids and fatty acids, respectively suggesting membrane-acting nature of these bacteriocins. Further, microscopic observation of bacteriocin-treated vesicles showed several damages indicating the interaction of bacteriocins. These findings suggest that the PDA vesicles may be used as bio-mimetic sensor for the detection of bacteriocins produced by several probiotics in food and therapeutic applications.

Halocins, natural antimicrobials of Archaea: Exotic or special or both?

Haloarchaea are adapted to survive under extreme saline conditions by accumulating osmolytes and salts to counteract the high osmotic pressure in their habitats. As a consequence, their proteins have evolved to remain active, or even most active, at very high ionic strength. Halocins are proteinaceous antimicrobial substances that are ribosomally-synthesized by haloarchaea and they provide the producers an advantage in the competition for nutrients and ecological niches. These antimicrobials are stable at high temperature, elevated salt concentrations, and alkaline pH conditions. These properties have endowed them with great potential in diverse biotechnological applications, which involve extreme processing conditions (such as high salt concentrations, high pressure, or high temperatures). They kill target cells by inhibition of Na+/H+ antiporter in the membrane or modification/disruption of the cell membrane leading to cell lysis. In general, the taxonomy of haloarchaea and their typical phenotypic and genotypic characteristics are well studied; however, information regarding their halocins, especially aspects related to genetics, biosynthetic pathways, mechanism of action, and structure-function relationship is very limited. A few studies have demonstrated the potential applications of halocins in the preservation of salted food products and brine-cured hides in leather industries, protecting the myocardium from ischemia and reperfusion injury, as well as from life-threatening diseases such as cardiac arrest and cancers. In recent years, genome mining has been an essential tool to decipher the genetic basis of halocin biosynthesis. Nevertheless, this is likely the tip of the iceberg as genome analyses have revealed many putative halocins in databases waiting for further investigation. Identification and characterization of this source of halocins may lead to antimicrobials for future therapeutics and/or food preservation. Hence, the present review analyzes different aspects of halocins such as biosynthesis, mechanism of action against target cells, and potential biotechnological applications.

Purification and Characterization of a New Halocin HA4 from Haloferax larsenii HA4 Isolated from a Salt Lake.

Halocins are antimicrobial peptides secreted by different members of haloarchaea. Halocin HA4 was purified from Haloferax larsenii HA4 using combination of ultrafiltration and chromatographic techniques. It was found to be ~ 14 kDa with unique N-terminal sequence, H2N-AEEEIFXPDX, which did not show homology with the known sequence suggesting a new/novel compound. It was found to be heat resistant up to 100 °C, stable at pH 2.0-10.0, and retained complete activity in the presence of different organic compounds such as methanol, ethanol, acetone, isopropanol, ethyl acetate, Tween 80, acetonitrile, SDS, Triton X-100, and urea. However, complete activity was reduced after the treatment with trypsin, papain, and proteinase K suggesting proteinaceous nature of the compound. The cytocidal nature of halocin HA4 was evidenced with complete loss of viable count of indicator strain, H. larsenii HA10. The change in FTIR spectrum of halocin-treated cells suggested halocin HA4 interacts with cell membrane and nucleic acids of the target cells. Thus, we report a new halocin inhibitory to related strains and may be applied in the preservation of salted foods and leather hides in the respective industries.

Can Probiotics Emerge as Effective Therapeutic Agents in Apical Periodontitis? A Review.

Apical periodontitis (AP) is a biofilm-associated disease initiated by the invasion of dental pulp by microorganisms from the oral cavity. Eradication of intracanal microbial infection is an important goal of endodontic treatment, and this is typically accomplished by mechanical instrumentation and application of sodium hypochlorite and chlorhexidine. However, these agents are tissue-irritating at higher concentrations and cytotoxic. Certain probiotics have been found effective in controlling marginal periodontitis, as evidenced by reduction of pathogenic bacterial loads, gains in clinical attachment levels, and reduced bleeding on probing. In vitro studies have shown inhibitory activity of some probiotics against endodontic pathogens. Similarly, in vivo studies in rats have demonstrated a positive immuno-modulatory role of probiotics in AP, as manifested by decreased levels of proinflammatory markers and increased levels of anti-inflammatory markers. A role for probiotics in effecting a reduction of bone resorption has also been reported. This review provides an outline of current research into the probiotic management of AP, with a focus on understanding the mechanisms of their direct antagonistic activity against target pathogens and of their beneficial modulation of the immune system.

Synergistically-acting Enterocin LD3 and Plantaricin LD4 Against Gram-Positive and Gram-Negative Pathogenic Bacteria.

The efficacy of antimicrobials is an important aspect during their applications in food and therapeutics. In this study, combination of two bacteriocins, enterocin LD3 and plantaricin LD4, was studied against two pathogenic bacteria, Staphylococcus aureus subsp. aureus ATCC25923 and Salmonella enterica subsp. enterica serovar Typhimurium ATCC13311 for increasing their potency and bactericidal activity. The minimal inhibitory concentrations (MICs) of enterocin LD3 and plantaricin LD4 against Staph. aureus subsp. aureus ATCC25923 were 180 and 220 µg/mL, whereas in combination, reduced to 115 µg/mL, respectively. The MICs of enterocin LD3 and plantaricin LD4 against Salm. enterica subsp. enterica serovar Typhimurium ATCC13311 were 240 and 320 µg/mL, respectively, whereas in combination, these were found to be 130 µg/mL, respectively. The fractional inhibitory concentration (FIC) indices calculated as 0.50 against Staph. aureus subsp. aureus ATCC25923 and 0.43 against Salm. enterica subsp. enterica serovar Typhimurium ATCC13311 were found to be ≤ 0.5 indicating the synergy. The isobologram showed MIC of combined bacteriocins falls below the plotted straight line further signifies synergy. The growth response of Staph. aureus subsp. aureus ATCC25923 and Salm. enterica subsp. enterica serovar Typhimurium ATCC13311 was significantly reduced in the presence of combined bacteriocins in comparison with their individual effects. The number of dead cells was higher as a result of combined effect as compared with their independent effect evidenced by fluorescent microscopy. Transmission electron microscopy (TEM) revealed the higher disruption of cell membrane in the combined bacteriocin-treated cells as compared with alone effects. The FTIR spectra of enterocin LD3-treated cells showed alteration at ~ 1,451.82 and ~ 1,094.30/cm corresponding to nucleic acids and phospholipids suggesting its interaction with cell membrane and nucleic acids. In contrast, plantaricin LD4-treated cells did not show such alterations suggesting plantaricin LD4 may kill target cells using other mechanism. Our data suggest that different mode of action of both bacteriocins results in division of labour and may be responsible for their synergistic activity against target cells. Similarly, the synergistic effect of bacteriocins was also observed against other pathogenic bacteria such as Proteus mirabilis ATCC43071, Pseudomonas aeruginosa ATCC27853 and Escherichia coli ATCC25922. These bacteriocins, therefore, act synergistically against target pathogens and may be applied in appropriate combinations for food safety and medical applications.

Probiotics at War Against Viruses: What Is Missing From the Picture?

Our world is now facing a multitude of novel infectious diseases. Bacterial infections are treated with antibiotics, albeit with increasing difficulty as many of the more common causes of infection have now developed broad spectrum antimicrobial resistance. However, there is now an even greater challenge from both old and new viruses capable of causing respiratory, enteric, and urogenital infections. Reports of viruses resistant to frontline therapeutic drugs are steadily increasing and there is an urgent need to develop novel antiviral agents. Although this all makes sense, it seems rather strange that relatively little attention has been given to the antiviral capabilities of probiotics. Over the years, beneficial strains of lactic acid bacteria (LAB) have been successfully used to treat gastrointestinal, oral, and vaginal infections, and some can also effect a reduction in serum cholesterol levels. Some probiotics prevent gastrointestinal dysbiosis and, by doing so, reduce the risk of developing secondary infections. Other probiotics exhibit anti-tumor and immunomodulating properties, and in some studies, antiviral activities have been reported for probiotic bacteria and/or their metabolites. Unfortunately, the mechanistic basis of the observed beneficial effects of probiotics in countering viral infections is sometimes unclear. Interestingly, in COVID-19 patients, a clear decrease has been observed in cell numbers of Lactobacillus and Bifidobacterium spp., both of which are common sources of intestinal probiotics. The present review, specifically motivated by the need to implement effective new counters to SARS-CoV-2, focusses attention on viruses capable of co-infecting humans and other animals and specifically explores the potential of probiotic bacteria and their metabolites to intervene with the process of virus infection. The goal is to help to provide a more informed background for the planning of future probiotic-based antiviral research.

Anti-staphylococcal activity of bacteriocins of food isolates Enterococcus hirae LD3 and Lactobacillus plantarum LD4 in pasteurized milk.

Bacteriocins of Enterococcus hirae LD3 and Lactobacillus plantarum LD4 have been applied in milk for growth inhibition of Staphylococcus aureus. The enumeration of S. aureus cells in nutrient broth and milk was found log10 9.7 and 10.2 CFU/mL, respectively, whereas it was reduced with increasing concentration of bacteriocins suggesting loss of cell viability. The lethal concentration (LC50) of enterocin LD3 and plantaricin LD4 against S. aureus was 160 and 220 µg/mL, respectively. Bacteriocin-treated cells were stained red with propidium iodide (PI) indicating dead cells further confirms bactericidal nature. The enterocin LD3-treated cells showed higher infrared absorbance at 1451.82 cm- 1 corresponding to phospholipids suggesting membrane-acting nature of the bacteriocin. However, plantaricin LD4-treated cells did not show such alterations suggesting different mode of action. Both bacteriocins caused disruption and shrinkage of target cells, and leakage of intracellular contents as observed in transmission electron microscope (TEM). The present study suggests killing of S. aureus in milk, therefore, enterocin LD3 and plantaricin LD4 may be applied in biopreservation of milk and related food products.

Enterocin LD3 from Enterococcus hirae LD3 causing efflux of intracellular ions and UV-absorbing materials in Gram-negative bacteria.

AIMS: To understand the bactericidal action of enterocin LD3 against Gram-negative bacteria. METHODS AND RESULTS: Minimum inhibitory concentration (MIC) of enterocin LD3 against Micrococcus luteus MTCC 106 and Escherichia coli NCDC 135 was 80 and 112 µg ml-1 , and minimum bactericidal concentration (MBC) was 128 and 180 µg ml-1 , respectively. The efflux of potassium ion (K+ ) was 14 and 13 ppm and electrical conductivity 10·5 and 8·3 mS cm-1 in cell-free supernatant of MIC-treated cells of M. luteus and E. coli respectively. The increased absorbance (OD260/280 ) 0·422/0·260 and 0·110/0·075 in the bacteriocin-treated cells of M. luteus MTCC 106 and E. coli, NCDC 135, respectively, suggested the release of nucleic acids and proteins. The higher infrared absorbance at 1451·82 and ~1094·30 cm-1 further suggested its interaction with cell membrane and nucleic acids of the target bacteria. The interaction of bacteriocin with nucleic acids was also confirmed using gel retardation assay. Transmission electron microscopy of the bacteriocin-treated cells revealed disruption of cell membrane and leakage of cytoplasmic contents. CONCLUSIONS: Enterocin LD3 demonstrates bactericidal activity against Gram-negative bacteria interacting with cell membrane and nucleic acids. SIGNIFICANCE AND IMPACT OF THE STUDY: The study discloses the possible mechanism of action of enterocin LD3 against Gram-negative bacteria which is a rare phenomenon.

Comparative Analysis of Inhibition-Based and Indicator-Independent Colorimetric Assay for Screening of Bacteriocin-Producing Lactic Acid Bacteria.

Screening of bacteriocin-producing lactic acid bacteria (LAB) is an important aspect for the search of new/novel probiotic strains. Here, a vesicle-based colorimetric assay was compared with conventional inhibition-based antimicrobial assays using 54 isolates of LAB. All isolates demonstrated zone of growth inhibition ranging from 2.5 to 7.5 mm against indicator strain, Micrococcus luteus MTCC106 using point inoculation method. Cell-free supernatant of the isolates showed zone of growth inhibition varying from 14.5 to 25 mm using agar well diffusion assay. These isolates inhibited the growth of indicator strain by 89.56-98.65%. The antimicrobial activity present in cell-free supernatant of different isolates was found to be in the range of 10-160 AU ml- 1. The treatment of polydiacetylene (PDA) vesicles with cell-free supernatant of selected isolates led to blue-red color transition, and presence of protein band on tricine SDS-PAGE confirmed the presence of membrane-acting peptides, bacteriocins. The colorimetric responses (CR%) varied between 0 and 59%, and the assay was found to be more sensitive, faster, and reliable as compared to the other conventional indicator-based methods used. Therefore, the colorimetric assay may be specifically applied for screening of bacteriocin-producing LAB.

Membrane-acting bacteriocin purified from a soil isolate Pediococcus pentosaceus LB44 shows broad host-range.

Bacteriocin LB44 was purified from cell-free supernatant (CFS) of Pediococcus pentosaceus LB44 using activity-guided chromatography techniques. It was stable up to 121 °C, pH 2.0-6.0, sensitive to proteinase K, papain and trypsin, and retained complete activity in the presence of organic solvents tested. The molecular weight of bacteriocin was ∼6 kDa and initial ten amino acid residues (GECGMCXECG) suggested a new compound. The loss in viable cell count and K+ ion efflux of target cells of Micrococcus luteus suggested bactericidal activity. The cell membrane of bacteriocin-treated cells was found to be ruptured which was further confirmed by Fourier Transform Infrared (FTIR) analysis suggesting interaction of bacteriocin with phospholipids in cell membrane. It showed broad host-range and inhibited the growth of Lactobacillus delbrueckii NRRL B-4525, L. plantarum NRRL B-4496, L. acidophilus NRRL B-4495, Enterococcus hirae LD3, Weissella confusa LM85, Staphylococcus aureus, Salmonella typhi ATCC 13311, Serratia marcescens ATCC 27137, Pseudomonas aeruginosa ATCC 27853, Proteus vulgaris ATCC 29905, Haloferax larsenii HA1, HA3, HA8, HA9 and HA10. These properties suggested a new bacteriocin from soil isolate P. pentosaceus LB44 which may offers possible applications in food-safety and therapeutics.

Activity-guided separation and characterization of new halocin HA3 from fermented broth of Haloferax larsenii HA3.

Haloferax larsenii HA3 was able to grow optimally in HS medium containing 15% NaCl, at pH 7.2 and 42 °C in aerobic conditions. Strain HA3 was found to be round shape, Gram-negative, catalase-positive, sensitive to bile acid, and resistant to chloramphenicol, and could not utilize arginine. The lipid profile revealed the presence of glycerol diether moiety (GDEM) suggesting Haloarchaea characteristics. Phylogenetic analysis based on 16S rRNA gene sequence similarities showed that it was closely related to H. larsenii ZJ206. Interestingly, strain HA3 was found to produce halocin HA3 which was purified using ultrafiltration and chromatography. It was found to be stable up to 80 °C, pH 2.0-10.0, organic solvents, surfactants, and detergents tested. However, the activity of halocin HA3 was completely reduced in the presence of proteinase K and trypsin. It was found to be halocidal against H. larsenii HA10, rupturing cell boundary and leading to cell death. The molecular weight of halocin HA3 was found to be ~13 kDa and MALDI-TOF MS/MS analysis suggested no homology with known halocins. The N-terminal ten amino-acid residues, NH2MNLGIILETN-COOH, suggested a new/novel halocin. These properties of halocin HA3 may be applicable for control of Haloarchaea in environments and salted foods.

Polyaniline-Stabilized Intertwined Network-like Ferrocene/Graphene Nanoarchitecture for Supercapacitor Application.

The present work highlights the effective H-π interaction between metallocenes (ferrocene; Fc) and graphene and their stabilization in the presence of polyaniline (PANI) through π-π interactions. The PANI-stabilized Fc@graphene nanocomposite (FcGA) resembled an intertwined network-like morphology with high surface area and porosity, which could make it a potential candidate for energy-storage applications. The relative interactions between the components were assessed through theoretical (DFT) calculations. The specific capacitance calculated from galvanostatic charging/discharging indicated that the PANI-stabilized ternary nanocomposite exhibited a maximum specific capacitance of 960 F g- at an energy density of 85 Wh Kg-1 and a current density of 1 A g- . Furthermore, electrochemical impedance spectroscopy (EIS) analysis confirmed the low internal resistance of the as-prepared nanocomposites, which showed improved charge-transfer properties of graphene after incorporation of Fc and stabilization with PANI. Additionally, all electrodes were found to be stable up to 5000 cycles with a specific capacitance retention of 86 %, thus demonstrating the good reversibility and durability of the electrode material.