Three strains of Spirulina platensis isolated from different locations showed capability of synthesizing poly(3-hydroxybutyrate) [P(3HB)] under nitrogen-starved conditions with a maximum accumulation of up to 10 wt.% of the cell dry weight (CDW) under mixotrophic culture conditions. Intracellular degradation (mobilization) of P(3HB) granules by S. platensis was initiated by the restoration of nitrogen source. This mobilization process was affected by both illumination and culture pH. The mobilization of P(3HB) was better under illumination (80% degradation) than in dark conditions (40% degradation) over a period of 4 days. Alkaline conditions (pH 10-11) were optimal for both biosynthesis and mobilization of P(3HB) at which 90% of the accumulated P(3HB) was mobilized. Transmission electron microscopy (TEM) revealed that the mobilization of P(3HB) involved changes in granule quantity and morphology. The P(3HB) granules became irregular in shape and the boundary region was less defined. In contrast to bacteria, in S. platensis the intracellular mobilization of P(3HB) seems to be faster than the biosynthesis process. This is because in cyanobacteria chlorosis delays the P(3HB) accumulation process.
A moderately halophilic bacterium isolated from fermenting shrimp paste, Salinivibrio sp. M318 was found capable of using fish sauce and mixtures of waste fish oil and glycerol as nitrogen and carbon sources, respectively, for poly(3-hydroxybutyrate) (PHB) production. A cell dry weight (CDW) of up to 10 g/L and PHB content of 51.7 wt% were obtained after 48 h of cultivation in flask experiment. Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] was synthesized when 1,4-butanediol, γ-butyrolactone, or sodium 4-hydroxybutyrate was added as precursors to the culture medium. The biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] was achieved by supplying precursors such as sodium valerate, sodium propionate, and sodium heptanoate. Salinivibrio sp. M318 was able to accumulate the above mentioned PHAs during the growth phase. High CDW of 69.1 g/L and PHB content of 51.5 wt% were obtained by strain Salinivibrio sp. M318 after 78 h of cultivation in fed-batch culture. The results demonstrate Salinivibrio sp. M318 to be a promising wild-type bacterium for the production of PHA from aquaculture residues.
Among the various types of polyhydroxyalkanoate (PHA), poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)] has a high potential to serve as commercial bioplastic due to its striking resemblance to petroleum-based plastics. In this study, five different genotypes of Cupriavidusnecator transformants harbouring the phaCBP-M-CPF4 gene (including PHB¯4/pBBR1-CBP-M-CPF4) were developed to evaluate the efficiency of 3HHx monomer incorporation. The fraction of 3-hydroxyhexanoate (3HHx) monomer that was incorporated into the PHA synthesized by these C. necator transformants using palm oil as the sole carbon source, was examined. Overall, co-expression of enoyl-CoA hydratase gene (phaJ1) from Pseudomonas aeruginosa, along with PHA synthase (PhaC), increased the 3HHx composition in the PHA copolymer. The differences in the enzyme activities of β-ketothiolase (PhaACn) and NADPH-dependent acetoacetyl-CoA reductase (PhaBCn) of the C. necator mutant hosts used in this study, were observed to alter the 3HHx composition and molecular weight of the PHA copolymer produced. The 3HHx fractions in the P(3HB-co-3HHx) produced by these C. necator transformants ranged between 1 and 18 mol%, while the weight-average molecular weight ranged from 0.7 × 106 to 1.8 × 106 Da. PhaCBP-M-CPF4 displayed a typical initial lag-phase and a relatively low synthase activity in the in vitro enzyme assay, which is thought to be the reason for the higher molecular weights of PHA obtained in this study.
A combination of palm olein (POl) and fructose was used as carbon source for the biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by recombinant Cupriavidus necator Re2058/pCB113. Cultures grown using 5g/L PO alone as carbon source produced cell dry weight (CDW) of 5.13g/L, 67% PHA/CDW and accumulated a copolymer containing 27mol% 3HHx in shake flask cultures. When cultures were grown in 5g/L fructose alone as the carbon source they produced CDW of 2.32g/L, 11% PHA/CDW and accumulated only poly(3-hydroxybutyrate) [P(3HB)] homopolymer. When the cells were cultured in 5g/L POl in combination with 7g/L fructose, CDW of 7.41g/L and 80% PHA/CDW was obtained with 17mol% 3HHx monomer fraction. Biosynthesis was carried out using a 13L fermenter to study the accumulation of 3HHx monomer fraction in the bacterial cells at different time point. The molecular weights of P(3HB-co-3HHx) with 4-15mol% 3HHx monomer were in the range between 5.47-6.85×105Da, which were at least two fold higher than previously reported values. Interestingly, the increase in Mwof the copolymer along with the increase in 3HHx molar fraction was observed. The viscoelastic property of the copolymer further confirmed the increase in Mw.
Polyhydroxyalkanoate (PHA) synthase, PhaC, is a key enzyme in the biosynthesis of PHA, a type of bioplastics with huge potential to replace petroleum-based plastics. While two structures have been determined, the exact mechanism remains unclear partly due to the absence of a tunnel for product passage. A model of the class I PhaC from Aquitalea sp. USM4, characterised with Km of 394 μM and kcat of 476 s-1 on 3-(R)-hydroxybutyryl-CoA, revealed a three-branched channel at the dimeric interface. Two of them are opened to the solvent and are expected to serve as the putative routes for substrate entrance and product exit, while the third is elongated in the class II PhaC1 model from Pseudomonas aeruginosa, indicating a role in accommodating the hydroxyalkanoate (HA) moiety of a HA-CoA substrate. Docking of the two tetrahedral intermediates, formed during the transfer of the growing PHA chain from the catalytic Cys to a new molecule of substrate and back to Cys, suggests a common elongation mechanism requiring the HA moiety of the ligand to rotate ~180°. Substrate specificity is determined in part by a bulky Phe/Tyr/Trp residue in the third branch in class I, which is conserved as Ala in class II to create room for longer substrates.
Polyhydroxyalkanoates (PHAs) are biodegradable polyesters produced by microorganisms, under unbalanced growth conditions, as a carbon storage compound. PHAs are composed of various monomers such as 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HHx). Silk fibroin (SF) derived from Bombyx mori cocoons, is a widely studied protein polymer commonly used for biomaterial applications. In this study, non-woven electrospun films comprising a copolymer of 3HB and 3HHx [P(3HB-co-3HHx)], SF and their blends were prepared by electrospinning technique. The growth and osteogenic differentiation of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) were studied using different types of fabricated electrospun films. The differentiation study revealed that electrospun P(3HB-co-3HHx)/SF film supports the differentiation of hUC-MSCs into the osteogenic lineage, confirmed by histological analysis using Alizarin Red staining, energy dispersive X-ray (EDX) and quantitative real-time PCR analysis (qPCR). Electrospun P(3HB-co-3HHx)/SF film up-regulated the expression of osteogenic marker genes, alkaline phosphatase (ALP) and osteocalcin (OCN), by 1.6-fold and 2.8-fold respectively, after 21 days of osteogenic induction. In conclusion, proliferation and osteogenic differentiation of hUC-MSCs were enhanced through the blending of P(3HB-co-3HHx) and SF. The results from this study suggest that electrospun P(3HB-co-3HHx)/SF film is a promising biomaterial for bone tissue engineering.
Polyhydroxyalkanoates (PHAs) are biopolyesters synthesized by microorganisms as intracellular energy reservoirs under stressful environmental conditions. PHA synthase (PhaC) is the key enzyme responsible for PHA biosynthesis, but the importance of its N- and C-terminal ends still remains elusive. Six plasmid constructs expressing truncation variants of Aquitalea sp. USM4 PhaC (PhaC1As) were generated and heterologously expressed in Cupriavidus necator PHB-4. Removal of the first six residues at the N-terminus enabled the modulation of PHA composition without altering the PHA content in cells. Meanwhile, deletion of 13 amino acids from the C-terminus greatly affected the catalytic activity of PhaC1As, retaining only 1.1-7.4% of the total activity. Truncation(s) at the N- and/or C-terminus of PhaC1As gradually diminished the incorporation of comonomer units, and revealed that the N-terminal region is essential for PhaC1As dimerization whereas the C-terminal region is required for stabilization. Notably, transmission electron microscopy analysis showed that PhaC modification affected the morphology of intracellular PHA granules, which until now is only known to be regulated by phasins. This study provided substantial evidence and highlighted the significance of both the N- and C-termini of PhaC1As in regulating intracellular granule morphology, activity, substrate specificity, dimerization and stability of the synthase.
The rapid acceleration of industrialization and urbanization has exacerbated water pollution, which is primarily caused by the presence of highly toxic, non-biodegradable contaminants in industrial waste and effluents. In response to this urgent issue, a novel nanobiocomposite film with titanium dioxide (TiO2) loaded onto a poly(3-hydroxybutyrate-co-18 mol% 3-hydroxyhexanoate) (18PHBH) matrix was developed to serve as an effective dual-function material with photocatalytic and antibacterial properties. Through Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR), Diffuse reflectance ultraviolet-visible (DRUV-Vis), Scanning Electron Microscope (SEM), and X-ray diffraction (XRD) analyses, the physicochemical properties of the TiO2/Gly/18PHBH nanobiocomposite film were exhaustively characterized, revealing effective TiO2 loading and uniform distribution on the film's surface. The film exhibited extraordinary photocatalytic degradation of methylene blue (MB) dye, with the 5TiO2/Gly/18PHBH film demonstrating the greatest efficiency. In addition, antibacterial testing revealed that the film was effective against 99.8 % of Staphylococcus aureus and 96.9 % of Pseudomonas aeruginosa. These results demonstrate the potential of polyhydroxyalkanoate-based films as exceptional nanoparticle matrices and position the 5TiO2/Gly/18PHBH film as a versatile candidate for applications in photocatalysis and antibacterial interventions, providing innovative solutions to critical environmental challenges.
Cassava starch has acquired many attentions owing to its ability to be developed as thermoplastic cassava starch (TPCS) where it can be obtained in low cost, making it to be one of alternatives to substitute petroleum-based plastic. An attempt was made to investigate the thermal, mechanical and moisture absorption properties of thermoplastic cassava starch blending with beeswax (TPCS-BW) fabricated using hot moulding compression method in the range of beeswax loading from 0, 2.5, 5 to 10 wt%. Addition of beeswax has significantly reduced tensile strength, elongation and flexural strength while improving tensile modulus and flexural modulus until 5 wt% beeswax. Incorporation of 10 wt% beeswax has successfully produced the lowest value of moisture absorption and water solubility among the bio-composite which might be attributed to the beeswax's hydrophobic properties in improving water barrier of the TPCS-BW bio-composite. Furthermore, the addition of beeswax resulted in the appearance of irregular and rough fractured surface. Meanwhile, fourier transform infrared (FT-IR) spectroscopy presented that incorporation of beeswax in the mixture has considerably improve hydrogen bonding of blends indicating good interaction between starch and beeswax. Hence, beeswax with an appropriate loading value able to improve the functional properties of TPCS-BW bio-composite.
Chitosan-deep eutectic solvent (DES) beads were prepared from chitosan and DESs. The DESs used were choline chloride-urea (DES A) and choline chloride-glycerol (DES B). Both chitosan-DES beads were used to remove malachite green (MG) dye from an aqueous solution. The optimum pH for chitosan-DES A was recorded at pH 8.0 while optimum pH for chitosan-DES B was pH 9.0. The maximum adsorption capacity obtained for chitosan-DES A and chitosan-DES B were 6.54 mg/g and 8.64 mg/g, respectively. The optimum conditions for both chitosan-DES beads to remove MG were 0.08 g of adsorbent and 20 min of agitation time. Five kinetic models were applied to analyse the data and the results showed that the pseudo-second-order and intraparticle diffusion model fitted best with R2 > 0.999. For the adsorption capacity, results show that the Freundlich and Langmuir adsorption isotherms fitted well with chitosan-DES A and chitosan-DES B, respectively. The maximum adsorption capacities (qmax) obtained from chitosan-DES A and chitosan-DES B were 1.43 mg/g and 17.86 mg/g, respectively. Desorption indicated good performance in practical applications.
In this study, chitosan/porous carbon composite (C-PC) modified in 1-Allyl-3-methyl imidazolium bromide [AMIM][Br] under airtight condition was prepared for the removal of Acid Blue-25 dye (AB-25) from aqueous medium. For comparison of adsorption efficiency of C-PC, chitosan-activated carbon composite (C-AC) was also prepared in 1% acetic acid. The adsorbents were characterised using SEM, EDX, XRD, BET, TGA and FTIR. The micrograph of C-PC revealed cavities and slightly rough surfaces dominated with similar sized and irregular shaped stone-like materials which differ from the precursors' micrograph. BET analysis revealed the domination of mesopores on the C-PC and C-AC surfaces, as the hydroxyl and amino group on C-PC are the main active sites for AB-25 dye uptake. The dye was better adsorbed onto C-PC at pH 2 and C-AC at pH 4. The adsorption capacity obtained for C-PC, C-AC, activated carbon (AC) and chitosan (CH) using Langmuir isotherm model are 3333.33 mg/g, 909.90 mg/g, 909.09 mg/g and 833.33 mg/g, respectively. The experimental data are well described by Langmuir and Fruendlich isotherms for adsorption of the dye onto C-PC, AC and CH. C-AC fitted into Langmuir isotherm only. The kinetics of the adsorption fitted into pseudo-second order indicating the possibility of chemical interactions in the adsorption process.
The presence of dyes in the aquatic environment as a result of anthropogenic activities, especially textile industries, is a critical environmental challenge that hinders the availability of potable water. Different wastewater treatment approaches have been used to remediate dyes in aquatic environments; however, most of these approaches are limited by factors ranging from high cost to the incomplete removal of the dyes and contaminants. Thus, the use of adsorption as a water treatment technology to remove dyes and other contaminants has been widely investigated using different adsorbents. This study evaluated the significance of chitosan as a viable adsorbent for removing dyes from water treatment. We summarised the literature and research results obtained between 2009 and 2020 regarding the adsorption of dyes onto chitosan and modified chitosan-based adsorbents prepared through physical and chemical processing, including crosslinking impregnation, grafting, and membrane preparation. Furthermore, we demonstrated the effects of various chitosan-based materials and modifications; they all improve the properties of chitosan by promoting the adsorption of dyes. Hence, the application of chitosan-based materials with various modifications should be considered a cutting-edge approach for the remediation of dyes and other contaminants in aquatic environments toward the global aim of making potable water globally available.
The aggressive search for unique materials in recent years has put forward chitosan and modified-chitosan as materials with unique structural and morphological characteristics for various important applications. Just as imidazolium-based ionic liquids are the commonly applied ionic liquids (ILs) type for chitosan modifications for various applications, their further modifications into beads for enhancing their properties is now gaining most attention. However, most of the currently prepared imidazolium ILs modified-chitosan beads are not in nano size due to preparation difficulties. In response to this and referencing the research works in the literature, the possible breakthrough directions including synthesis routes, and physical and mechanical transformation processes are proposed. These procedures are expected to provide certain theoretical and empirical basis, as well as technical guide for developing nano-micro size chitosan beads using imidazolium based ILs.
Adsorption efficiency of a duo-material blend featuring the fabrication of modified chitosan adsorbents (powder (C-emimAc), bead (CB-emimAc) and sponge (CS-emimAc)) for the removal of Cd(II) from aqueous solution was investigated. The chitosan@activated carbon (Ch/AC) blend was developed in a green ionic solvent, 1-ethyl-3-methyl imidazolium acetate (EmimAc) and its characteristics was examined using FTIR, SEM, EDX, BET and TGA. The possible mechanism of interaction between the composites and Cd(II) was also predicted using the density functional theory (DFT) analysis. The interactions of various blend forms (C-emimAc, CB-emimAc and CS-emimAc) with Cd(II) gave better adsorption at pH 6. The composites also present excellent chemical stability in both acidic and basic conditions. The monolayer adsorption capacities obtained (under the condition 20 mg/L [Cd], adsorbent dosage 5 mg, contact time 1 h) for the CB-emimAc (84.75 mg/g) > C-emimAc (72.99 mg/g) > CS-emimAc (55.25 mg/g), as this was supported by their order of increasing BET surface area (CB-emimAc (120.1 m2/g) > C-emimAc (67.4 m2/g) > CS-emimAc (35.3 m2/g)). The feasible adsorption interactions between Cd(II) and Ch/AC occurs through the O-H and N-H groups of the composites, as supported by DFT analysis in which an electrostatic interactions was predicted as the dominant force. The interaction energy (-1309.35 eV) calculated via DFT shows that the Ch/AC with amino (-NH) and hydroxyl (-OH) groups are more effective with four significant electrostatic interactions with the Cd(II) ion. The various form of Ch/AC composites developed in EmimAc possess good adsorption capacity and stability for the adsorption Cd(II).
Several water and wastewater technologies have been implored for the removal of dyes during wastewater treatments; however; different types have been reportedly found in surface and groundwater systems. Hence, there is a need to investigate other water treatment technologies for the complete remediation of dyes in aquatic environments. In this study, novel chitosan-based polymer inclusion membranes (PIMs) were synthesized for the removal of malachite green dye (MG) which is a recalcitrant of great concern in water. Two types of PIMs were synthesized in this study, the first PIM (PIMs-A) was composed of chitosan, bis-(2-ethylhexyl) phosphate (B2EHP), and dioctyl phthalate (DOP). While, the second PIMs (PIMs-B) were composed of chitosan, Aliquat 336, and DOP. The physico-thermal stability of the PIMs was investigated using Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and thermogravimetric analysis (TGA), both PIMs demonstrated good stability with a weak intermolecular force of attraction amongst the various components of the membranes. The effects of the initial concentration of MG, pH of the MG solution, stripping solution, and time were investigated. At optimum conditions, both membranes (PIM-A and B) recorded the highest efficiencies of 96 % and 98 % at pH 4 and initial contaminants concentration of 50 mg/L, respectively. Finally, both PIMs were used for the removal of MG in different environmental samples (river water, seawater, and tap water) with an average removal efficiency of 90 %. Thus, the investigated PIMs can be considered a potential suitable technique for the removal of dyes and other contaminants from aquatic matrices.
Globally, people suffering from bone disorders are steadily increasing and bone tissue engineering is an advanced approach to treating fractured and defected bone tissues. In this study, we have prepared polymeric nanocomposite by free-radical polymerization from sodium alginate, hydroxyapatite, and silica with different GO amounts. The porous scaffolds were fabricated using the freeze drying technique. The structural, morphological, mechanical, and wetting investigation was conducted by Fourier-transform infrared spectroscopy, X-ray diffraction, scanning electron microscope, universal tensile machine, and water contact angle characterization techniques. The swelling, biodegradation, and water retention were also studied. The biological studies were performed (cell viability, cell adherence, proliferation, and mineralization) against osteoblast cell lines. Scaffolds have exhibited different pore morphology SAG-1 (pore size = 414.61 ± 56 μm and porosity = 81.45 ± 2.17 %) and SAG-4 (pore size = 195.97 ± 82 μm and porosity = 53.82 ± 2.45 %). They have different mechanical behavior as SAG-1 has the least compression strength and compression modulus 2.14 ± 2.35 and 16.51 ± 1.27 MPa. However, SAG-4 has maximum compression strength and compression modulus 13.67 ± 2.63 and 96.16 ± 1.97 MPa with wetting behavior 80.70° and 58.70°, respectively. Similarly, SAG-1 exhibited the least and SAG-4 presented maximum apatite mineral formation, cell adherence, cell viability, and cell proliferation against mouse pre-osteoblast cell lines. The increased GO amount provides different multifunctional materials with different characteristics. Hence, the fabricated scaffolds could be potential scaffold materials to treat and regenerate fracture bone tissues in bone tissue engineering.
Dicranopteris linearis (DL) is a fern in the Gleicheniaceae family, locally known as resam by the Malay community. It has numerous pharmacological benefits, with antiulcer and gastroprotective properties. Peptic ulcer is a chronic and recurring disease that significantly impacts morbidity and mortality, affecting nearly 20 % of the world's population. Despite the effectiveness of peptic ulcer drugs, there is no perfect treatment for the ailment. Encapsulation is an advanced technique that can treat peptic ulcers by incorporating natural sources. This work aims to encapsulate DL extract using different types of cellulose particles by the solvent displacement technique for peptic ulcer medication. The extract was encapsulated using methyl cellulose (MC), ethyl cellulose (EC), and a blend of ethyl methyl cellulose through a dialysis cellulose membrane tube and freeze-dried to yield a suspension of the encapsulated DL extracts. The microencapsulated methyl cellulose chloroform extract (MCCH) has a considerably greater level of total phenolic (84.53 ± 6.44 mg GAE/g), total flavonoid (84.53 ± 0.54 mg GAE/g), and antioxidant activity (86.40 ± 0.63 %). MCCH has the highest percentage of antimicrobial activity against Escherichia coli (2.42 ± 107 × 0.70 CFU/mL), Bacillus subtilis (5.21 ± 107 × 0.90 CFU/mL), and Shigella flexneri (1.25 ± 107 × 0.66 CFU/mL), as well as the highest urease inhibitory activity (50.0 ± 0.21 %). The MCCH particle size was estimated to be 3.347 ± 0.078 μm in diameter. It has been proven that DL elements were successfully encapsulated in the methyl cellulose polymer in the presence of calcium (Ca). Fourier transform infrared (FTIR) analysis indicated significant results, where the peak belonging to the CO stretch of the carbonyl groups of methyl cellulose (MC) shifted from 1638.46 cm-1 in the spectrum of pure MC to 1639.10 cm-1 in the spectrum of the MCCH extract. The shift in the wavenumbers was due to the interactions between the phytochemicals in the chloroform extract and the MC matrix in the microcapsules. Dissolution studies in simulated gastric fluid (SGF) and model fitting of encapsulated chloroform extracts showed that MCCH has the highest EC50 of 6.73 ± 0.27 mg/mL with R2 = 0.971 fitted by the Korsmeyer-Peppas model, indicating diffusion as the mechanism of release.
In this present study, isolation, characterization and protective effect of sulfated polysaccharide (SP) isolated from the brown algae Padina gymnospora was investigated. SP was isolated and characterized through FT-IR, 1H NMR, TGA, GC-MS and CHN analysis. The molecular weight of SP was found to be 16 kDa. The isolated SP contains 29.4 ± 0.35% of sulfate, 27 ± 0.11% of fucose, 0.05 ± 0.12% of protein, respectively. Furthermore, SP exhibits its excellent radical scavenging effects were evaluated by DPPH, ABTS radical scavenging and reducing power assays. Moreover, pretreatment with SP significantly mitigates H2O2 induced cytotoxicity in L-929 cells in a dose dependent manner. Furthermore, SP pretreatment ameliorates oxidative stress induced apoptosis and DNA damage, alleviates the generation of intracellular reactive oxygen species (ROS) and restores mitochondrial membrane potential (MMP) in L-929 cells through its antioxidant potential. Together, these results suggest that SP can be exploited as a natural antioxidant in the food and pharmaceutical industries.
Dielectric spectroscopy is employed to study the relaxation phenomena in natural polyhydroxyalkanoates (PHAs) upon temperature and frequency variations. Effects of PHAs molecular structure on the relaxation, arising from the differences in monomeric composition, are investigated under identical conditions in a frequency range of 10-2-106 Hz, and at different temperatures. All PHA samples showed different dielectric response at different temperature. Primary α-relaxation signals are observed at temperature corresponding to the glass transition temperature. On the other hand, secondary β- and γ-relaxations are detected at low temperatures, and attributed to local motions of polar groups and small segments of the polymer chain. The dielectric properties of representative PHA samples are compared and discussed.
Diabetic retinopathy (DR) is one of the chronic complications of diabetes. It includes retinal blood vessels' damage. If untreated, it leads to loss of vision. The existing treatment strategies for DR are expensive, invasive, and need expertise during administration. Hence, there is a need to develop a non-invasive topical formulation that can penetrate deep to the posterior segment of retina and treat the damaged retinal vessels. In addition, it should also provide sustained release. In recent years, novel drug delivery systems (NDDS) have been explored for treating DR and found successful. In this study, chitosan (CS) modified 5-Fluorouracil Nanostructured Lipid Carriers (CS-5-FU-NLCs) were prepared by modified melt emulsification-ultrasonication method and optimized by Box-Behnken Design. The size, polydispersity index, zeta potential and entrapment efficiency of CS-5-FU-NLCs were 163.2 ± 2.3 nm, 0.28 ± 1.52, 21.4 ± 0.5 mV and 85.0 ± 0.2 %, respectively. The in vitro drug release and ex vivo permeation study confirmed higher and sustained drug release in CS-5-FU-NLCs as compared to 5-FU solution. HET-CAM Model ensured the non-irritant nature of CS-5-FU-NLCs. In vivo ocular studies of CS-5-FU-NLCs confirmed antiangiogenic effect of 5-FU by CAM model and diabetic retinopathy induced rat model, indicating successful delivery of 5-FU to the retina.