Microbial fermentation-based synthesis of nano-curcumin suggesting the role of pullulan in nano-formulation.

Curcumin is a multitargeting nutraceutical with numerous health benefits, however, its efficacy is limited due to poor aqueous solubility and reduced bioavailability. While nano-formulation has emerged as an alternative to encounter such issues, it often involves use of toxic solvents. Microbial synthesis may be an innovative solution to address this lacuna. Present study, for the first time, reports exploitation of Aureobasidium pullulans RBF4A3 for production of nano-curcumin. For this purpose, Aureobasidium pullulans RBF4A3 was inoculated in YPD media along with curcumin (0.1 mg/mL) and incubated for 24 h, 48 h, and 72 h. Subsequently, residual sugar, biomass, EPS concentration, curcumin concentration, and curcumin nanoparticle size were measured. As a result, nano-curcumin with an average particle size of 31.63 nm and enhanced aqueous solubility was obtained after 72 h. Further, investigations suggested that pullulan, a reducing polysaccharide, played a significant role in curcumin nano-formulation. Pullulan-mediated nano-curcumin formulation, with an average particle size of 24 nm was achieved with conversion rate of around 59.19 %, suggesting improved aqueous solubility. Additionally, the anti-oxidant assay of the resulting nano-curcumin was around 53.7 % per µg. Moreover, kinetics and thermodynamic studies of pullulan-based nano-curcumin revealed that it followed first-order kinetics and was favored by elevated temperature for efficient bio-conversion. Also, various physico-chemical investigations like FT-IR, NMR, and XRD reveal that pullulan backbone remains intact while forming curcumin nanoparticle. This study may open up new avenues for synthesizing nano-polyphenols through a completely green and solvent free process with plausible diverse applications.

Enhanced encapsulation efficiency and controlled release of co-encapsulated Bacillus coagulans spores and vitamin B9 in gellan/κ-carrageenan/chitosan tri-composite hydrogel.

The current study, for the first time, attempts to co-encapsulate Bacillus coagulans spores as probiotics and vitamin B9 in the polysaccharide-based matrix for their targeted delivery. Instead of vegetative cells, probiotic spores were chosen owing to their higher stability. The matrix, tri-composite hydrogel, was synthesized from gellan, κ-carrageenan, and chitosan through self-assembly devoid of chemical cross-linkers. Hence, it was found suitable for application in the co-encapsulation of bioactive compounds. The synthesized hydrogel showed remarkable encapsulation efficiency for folic acid and probiotic spores, both individually and in combination. At acidic pH, loaded hydrogel exhibited 28.42 % and 45.14 % release of spores and folic acid, respectively, which was comparatively lower than the trends observed under neutral and alkaline pH. These results were correlated with the release pattern observed during in vitro digestibility studies. Moreover, spore conversion to vegetative cells and its high colonization were observed in the simulated intestinal phase. Therefore, the matrix maintained viability and stability of co-encapsulated folic acid and bacterial spores in gastric pH while they were slowly released in the intestinal phase. These promising findings pave the way to develop a natural matrix for co-encapsulating various bioactive compounds and probiotics.

Microbial Polysaccharide-Based Nanoformulations for Nutraceutical Delivery.

In recent times, nutrition and diet have become prominent health paradigms due to sedentary lifestyle disorders. Preventive health care strategies are becoming increasingly popular instead of treating and managing diseases. A nutraceutical is an innovative concept that offers additional health benefits beyond its fundamental nutritional value. These nutraceuticals have the potential to reduce the exorbitant use of synthetic drugs because the modern medicine approach of treating diseases with high-tech, expensive supplements, and long-term consequences aggravates consumers. However, most nutraceuticals are plant-derived, making them susceptible to degradation and prone to chemical instability, poor solubility, unpleasant taste, and bioactivity loss before absorption to the targeted site. To counteract this problem, the bioavailability of these labile compounds can be maximized by encapsulating them in protective nanocarriers. It is crucial that nanoencapsulation technologies convert bioactive compounds into forms that can be easily combined with functional foods and beverages without adversely affecting their organoleptic properties. In recent years, nanoformulations using food-grade materials, such as polysaccharides, proteins, lipids, etc., have received considerable attention. Among them, microbial polysaccharides are biocompatible, nontoxic, and nonimmunogenic, and most of them are US-FDA approved and can undergo tailored modifications. The nanoformulation of microbial polysaccharide is a relatively new frontier which has several advantages over existing systems. The present article, for the first time, comprehensively reviews microbial polysaccharides-based nanodelivery systems for nutraceuticals and discusses various techno-commercial aspects of these nanotechnological preparations. Moreover, this has also attempted to draw a future research perspective in this area.

Isolation, purification, and characterization of a novel exopolysaccharide isolated from marine bacteria Brevibacillus borstelensis M42.

Marine microbes produce polysaccharides with unique physicochemical and functional properties that help them survive in harsh marine environments. However, only a handful of marine exopolysaccharides (EPSs) have been reported to date. The present study explored the seashore of Visakhapatnam, India, to report a novel exopolysaccharide designated as Br42 produced by Brevibacillus borstelensis M42. The isolate was identified through morphological, biochemical, phylogenetic, and genome sequencing analysis. The studies on fermentation kinetics revealed that EPS Br42 was a primary metabolite with a maximum production of 1.88 ± 0.02 g/L after 60 h when production broth was fortified with 2% glucose. Additionally, EPS Br42 was found to be a heteropolysaccharide consisting of glucose and galacturonic acid with a molecular weight of about 286 kDa. Interestingly, this molecule possesses industrially relevant functional properties such as water-holding (510 ± 0.35%), oil-holding (374 ± 0.12% for coconut oil and 384 ± 0.35% for olive oil), and swelling capacities (146.6 ± 5.75%). EPS Br42 could form an emulsion that was stable at a wide pH range for about 72 h and, in fact, performed better as compared to Span 20, a commercially used synthetic emulsifier. Moreover, this EPS was also found to be heat stable and exhibited non-Newtonian pseudoplastic behavior. These physicochemical and functional properties of polysaccharides suggest that the EPS Br42 has potential for multifarious industrial applications as an emulsifier, stabilizer, viscosifier, and binding agent.

A review on the biosynthesis of metal and metal salt nanoparticles by microbes.

Metal nanoparticles have received great attention from researchers across the world because of a plethora of applications in agriculture and the biomedical field as antioxidants and antimicrobial compounds. Over the past few years, green nanotechnology has emerged as a significant approach for the synthesis and fabrication of metal nanoparticles. This green route employs various reducing and stabilizing agents from biological resources for the synthesis of nanoparticles. The present article aims to review the progress made in recent years on nanoparticle biosynthesis by microbes. These microbial resources include bacteria, fungi, yeast, algae and viruses. This review mainly focuses on the biosynthesis of the most commonly studied metal and metal salt nanoparticles such as silver, gold, platinum, palladium, copper, cadmium, titanium oxide, zinc oxide and cadmium sulphide. These nanoparticles can be used in pharmaceutical products as antimicrobial and anti-biofilm agents, targeted delivery of anticancer drugs, water electrolysis, waste water treatment, biosensors, biocatalysis, crop protection against pathogens, degradation of dyes etc. This review will discuss in detail various microbial modes of nanoparticles synthesis and the mechanism of their synthesis by various bioreducing agents such as enzymes, peptides, proteins, electron shuttle quinones and exopolysaccharides. A thorough understanding of the molecular mechanism of biosynthesis is the need of the hour to develop a technology for large scale production of bio-mediated nanoparticles. The present review also discusses the advantages of various microbial approaches in nanoparticles synthesis and lacuna involved in such processes. This review also highlights the recent milestones achieved on large scale production and future perspectives of nanoparticles.

Microbial glycolipoprotein-capped silver nanoparticles as emerging antibacterial agents against cholera.

BACKGROUND: With the increased number of cholera outbreaks and emergence of multidrug resistance in Vibrio cholerae strains it has become necessary for the scientific community to devise and develop novel therapeutic approaches against cholera. Recent studies have indicated plausibility of therapeutic application of metal nano-materials. Among these, silver nanoparticles (AgNPs) have emerged as a potential antimicrobial agent to combat infectious diseases. At present nanoparticles are mostly produced using physical or chemical techniques which are toxic and hazardous. Thus exploitation of microbial systems could be a green eco-friendly approach for the synthesis of nanoparticles having similar or even better antimicrobial activity and biocompatibility. Hence, it would be worth to explore the possibility of utilization of microbial silver nanoparticles and their conjugates as potential novel therapeutic agent against infectious diseases like cholera. RESULTS: The present study attempted utilization of Ochrobactrum rhizosphaerae for the production of AgNPs and focused on investigating their role as antimicrobial agents against cholera. Later the exopolymer, purified from the culture supernatant, was used for the synthesis of spherical shaped AgNPs of around 10 nm size. Further the exopolymer was characterized as glycolipoprotein (GLP). Antibacterial activity of the novel GLP-AgNPs conjugate was evaluated by minimum inhibitory concentration, XTT reduction assay, scanning electron microscopy (SEM) and growth curve analysis. SEM studies revealed that AgNPs treatment resulted in intracellular contents leakage and cell lysis. CONCLUSION: The potential of microbially synthesized nanoparticles, as novel therapeutic agents, is still relatively less explored. In fact, the present study first time demonstrated that a glycolipoprotein secreted by the O. rhizosphaerae strain can be exploited for production of AgNPs which can further be employed to treat infectious diseases. Although this type of polymer has been obtained earlier from marine fungi and bacteria, none of these reports have studied the role of this polymer in AgNPs synthesis and its application in cholera therapy. Interestingly, the microbial GLP-capped AgNPs exhibited antibacterial activity against V. cholerae comparable to ciprofloxacin. Thus the present study may open up new avenues for development of novel therapeutic agents for treatment of infectious diseases. Graphical abstract Development of novel therapeutic agents for treatment of cholera.

Facile and rapid thermo-regulated biomineralization of gold by pullulan and study of its thermodynamic parameters.

A novel method for the production of gold nanoparticles (AuNPs) using pullulan as reducing and stabilizing agent has been developed. Quasi-spherical shaped AuNPs in the range of 50-100 nm were produced at three different temperature regimes 80°C, 90°C and 100°C as characterized using UV-vis spectrophotometer, TEM and DLS. Study of reaction kinetics and thermodynamic parameters indicated that the reaction between pullulan and chloroauric acid for AuNPs formation followed first order reaction kinetics and higher temperature was favorable for the synthesis of smaller sized AuNPs. FT-IR data analyses, provided an insight towards the mechanism of gold nanoparticle formation which suggested that, the free CH2OH groups of pullulan molecule were oxidized to carboxylate ions resulted in formation of AuNPs whereas the basic skeletal structure of pullulan remained unaltered. This study may open up new avenues for synthesis of tailor made biogenic AuNPs with possible application in biomedical field.

Cost effective production of pullulan from agri-industrial residues using response surface methodology.

Response surface methodology was used to develop an economically feasible process for the fermentative production of pullulan using agri-industrial residues, jaggery, de-oiled jatropha seed cake (DOJSC) and corn steep liquor (CSL), as sole media components. A second order polynomial model was obtained using central composite design to understand the effects of interactions among these substrates on pullulan biosynthesis. Results indicated that, lower concentrations of CSL and DOJSC and higher concentrations of jaggery favoured pullulan production. The optimal nutrient composition (18% jaggery, 3% DOJSC and 0.97% CSL) as suggested by the model resulted in production of 66.25 g/L pullulan with a productivity of 0.92 g/Lh. Analysis of raw material cost component for pullulan production suggested that sole utilization of agri-residues may lead to development of cost effective process for pullulan production.

Development of suitable solvent system for downstream processing of biopolymer pullulan using response surface methodology.

Downstream processing is an important aspect of all biotechnological processes and has significant implications on quality and yield of the final product. Several solvents were examined for their efficacy on pullulan precipitation from fermentation broth. Interactions among four selected solvents and their effect on pullulan yield were studied using response surface methodology. A polynomial model was developed using D-optimal design and three contour plots were generated by performing 20 different experiments and the model was validated by performing optimization experiments. The results indicated that lower concentration of ethanol in combination with the other three solvents has resulted in higher yield of polymer from fermentation broth and the optimized solvent system was able to recover 1.44 times more pullulan as compared to the conventional ethanolic precipitation method. These observations may help in enhancing efficiency of pullulan recovery from fermentation broth and also result in reduced cost of production for the final product.

Biosynthesis of gold and silver nanoparticles using a novel marine strain of Stenotrophomonas.

The present study aims at exploiting marine microbial diversity for biosynthesis of metal nanoparticles and also investigates role of microbial proteins in the process of bio-mineralization of gold and silver. This is the first report for concurrent production of gold and silver nanoparticles (AuNPs and AgNPs) by extracellular secretion of a novel strain of Stenotrophomonas, isolated from Indian marine origin. This novel strain has faster rate kinetics for AgNPs synthesis than any other organism reported earlier. The nanoparticles were further characterized using UV-vis spectrophotometer, TEM, DLS and EDAX confirming their size ranging from 10-50 nm and 40-60 nm in dimensions for AuNPs and AgNPs, respectively. TEM analysis indicated formation of multi-shaped nanoparticles with heterogeneous size distribution in both the cases. Finally, the SDS-PAGE analysis of extracellular media supernatant suggested a potential involvement of certain low molecular weight secretory proteins in AuNPs and AgNPs biosynthesis.

Utilization of corn steep liquor for biosynthesis of pullulan, an important exopolysaccharide.

Five different agricultural wastes viz. rice bran oil cake, soya bean oil cake, cotton seed oil cake, mustard seed oil cake and corn steep liquor (CSL) were evaluated for their use as nutrient along with 15% (w/v) glucose as carbon source for biosynthesis of pullulan using Aureobasidium pullulans RBF 4A3. Among the selected agricultural wastes, CSL was found to be the best and supported production of 77.92gL(-1) pullulan under un-optimized conditions. Single point optimization technique resulted in increase in 18% pullulan (88.59gL(-1)) production. The process was successfully validated in a 7-L fermenter and a process economic analysis has suggested that use of CSL as nutrient may result in 3-fold reduction of cost of raw materials for pullulan production as compared to a process where conventional nitrogen sources were used. These observations may be helpful in development of a cost effective process for pullulan production.

Exploitation of marine bacteria for production of gold nanoparticles.

BACKGROUND: Gold nanoparticles (AuNPs) have found wide range of applications in electronics, biomedical engineering, and chemistry owing to their exceptional opto-electrical properties. Biological synthesis of gold nanoparticles by using plant extracts and microbes have received profound interest in recent times owing to their potential to produce nanoparticles with varied shape, size and morphology. Marine microorganisms are unique to tolerate high salt concentration and can evade toxicity of different metal ions. However, these marine microbes are not sufficiently explored for their capability of metal nanoparticle synthesis. Although, marine water is one of the richest sources of gold in the nature, however, there is no significant publication regarding utilization of marine micro-organisms to produce gold nanoparticles. Therefore, there might be a possibility of exploring marine bacteria as nanofactories for AuNP biosynthesis. RESULTS: In the present study, marine bacteria are exploited towards their capability of gold nanoparticles (AuNPs) production. Stable, monodisperse AuNP formation with around 10 nm dimension occur upon exposure of HAuCl(4) solution to whole cells of a novel strain of Marinobacter pelagius, as characterized by polyphasic taxonomy. Nanoparticles synthesized are characterized by Transmission electron microscopy, Dynamic light scattering and UV-visible spectroscopy. CONCLUSION: The potential of marine organisms in biosynthesis of AuNPs are still relatively unexplored. Although, there are few reports of gold nanoparticles production using marine sponges and sea weeds however, there is no report on the production of gold nanoparticles using marine bacteria. The present work highlighted the possibility of using the marine bacterial strain of Marinobacter pelagius to achieve a fast rate of nanoparticles synthesis which may be of high interest for future process development of AuNPs. This is the first report of AuNP synthesis by marine bacteria.

Deoiledjatropha seed cake is a useful nutrient for pullulan production.

BACKGROUND: Ever increasing demand for fossil fuels is a major factor for rapid depletion of these non-renewable energy resources, which has enhanced the interest of finding out alternative sources of energy. In recent years jatropha seed oil has been used extensively for production of bio-diesel and has shown significant potential to replace petroleum fuels at least partially. De-oiled jatropha seed cake (DOJSC) which comprises of approximately 55 to 65% of the biomass is a byproduct of bio-diesel industry. DOJSC contains toxic components like phorbol esters which restricts its utilization as animal feed. Thus along with the enhancement of biodiesel production from jatropha, there is an associated problem of handling this toxic byproduct. Utilization of DOJSC as a feed stock for production of biochemicals may be an attractive solution to the problem.Pullulan is an industrially important polysaccharide with several potential applications in food, pharmaceuticals and cosmetic industries. However, the major bottleneck for commercial utilization of pullulan is its high cost. A cost effective process for pullulan production may be developed using DOJSC as sole nutrient source which will in turn also help in utilization of the byproduct of bio-diesel industry. RESULTS: In the present study, DOJSC has been used as a nutrient for production of pullulan, in place of conventional nutrients like yeast extract and peptone. Process optimization was done in shake flasks, and under optimized conditions (8% DOJSC, 15% dextrose, 28°C temperature, 200 rpm, 5% inoculum, 6.0 pH) 83.98 g/L pullulan was obtained. The process was further validated in a 5 L laboratory scale fermenter. CONCLUSION: This is the first report of using DOJSC as nutrient for production of an exopolysaccharide. Successful use of DOJSC as nutrient will help in finding significant application of this toxic byproduct of biodiesel industry. This in turn also have a significant impact on cost reduction and may lead to development of a cost effective green technology for pullulan production.