Pullulan Acetate Nanoparticles Research Articles

Hyalgan-decorated-ferulic acid-loaded pullulan acetate nanoparticles against gastrointestinal cancer cell lines

Hyalgan-decorated-ferulic acid-loaded pullulan acetate nanoparticles against gastrointestinal cancer cell lines

Formulation and Characterisation of Pullulan Acetate Nanoparticles Loaded with 5-Fluorouracil

This study was geared to analyze the preparation methods of pullulan acetate-based nanoparticles loaded with 5-fluorouracil, as well as the potential of pullulan as a biopolymer matrix for obtaining nanoparticles applied in the delivery of anticancer drugs (5-FU). Various methods were used to produce pullulan acetate-based nanoparticles loaded with 5-FU, including nanoprecipitation, modified nanoprecipitation, and double emulsion. Pullulan was previously chemically modified with acetic anhydride, dimethylformamide and pyridine, and yielded pullulan acetate. Pullulan was made using the Aureobasidium pullulans strain through a fermentation procedure. UV-Vis Spectro-photometric and dynamic light scattering (DLS) methods were used to assess entrapment effectiveness, size, and polydispersity index (PDI) of pullulan acetate-based nanoparticles loaded with 5-FU. Based on the properties of the nanoparticles obtained, the optimum preparation method was chosen. The maximum entrapment efficiency was found in pullulan acetate nanoparticles loaded with 5-FU generated by a double emulsion method. The mean hydrodynamic size and PDI of all nanoparticles were adequate. The best formulation showed faster 5-FU release profile in acid phosphate-buffered saline (pH 5) than in phosphate-buffered saline pH 7.4. According to the findings, pullulan derivatives have a great potential for producing nanoparticles that might be used to deliver anticancer medicines.

Self-Assembled Pullulan Acetate Nanoparticles for pH-Dependent Controlled Drug Delivery Application

Self-Assembled Pullulan Acetate Nanoparticles for pH-Dependent Controlled Drug Delivery Application

The study of establishment of an in vivo tumor model by three-dimensional cells culture systems methods and evaluation of antitumor effect of biotin-conjugated pullulan acetate nanoparticles

In this study, three-dimensional (3D) hydrogels were used for human hepatocellular carcinoma (HepG2) cells culture systems in vitro and establishment of an in vivo xenografted tumor model. Based on our previous work on the biotin-conjugated pullulan acetate nanoparticles (Bio-PA NPs) as anticancer drug carriers, we further studied the anti-tumor effect of the NPs in two-dimensional (2D) and 3D cell culture system. When embedded in 3D hydrogels, HepG2 cells formed tumor spheroids and the cytoplasmic actin microfilamentrates were rearranged over a period of 7 days. In vitro cytotoxicity results indicated that HepG2 cells in 3D hydrogels were more resistant to Bio-PA NPs treatments compared to the 2D system. The tumor formation rate of in vivo xenografted tumor model using 3D culture systems method was 98.2%, which was significantly higher than that using of 2D cultured cells (76.4%). Then we injected the 3D HepG2 cells systems in the right anterior axillary of female Balb/c nude mice, and evaluate the in vivo anti-tumor efficacy of Bio-PA NPs. In summary, these results suggested that HepG2 cells in 3D hydrogel system has shown the potential to provide an in vitro and in vivo model and for the evaluation of Bio-PA NPs.

Uptake and transport of pullulan acetate nanoparticles in the BeWo b30 placental barrier cell model.

IntroductionNanomedicine has shown a great potential in perinatal medicine because of its characteristics of sustained, controlled release and targeting ability; on the other hand, it may also lead to unexpected toxicities such as embryotoxicity and even malformation after crossing the placental barrier, but data concerning transplacental transport are scarce. Pullulan acetate (PA) nanoparticles (NPs) are a promising nanocarrier derived from natural polysaccharide; however, their transplacental transport ability and mechanism are unknown.Materials and methodsIn this study, fluorescein isothiocyanate (FITC) conjugated PA (PA-FITC) was synthesized. PA-FITC NPs were characterized by dynamic light scattering, transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The cytotoxicity of PA-FITC NPs at concentrations of 15, 30, 60, 125, 250, 500, 1,000 and 2,000 μg/mL was studied by cell counting kit-8. The human chorionic gonadotrophin (HCG) cytokine assay was conducted to evaluate the biological function of BeWo b30 cells. Endocytic mechanisms of PA-FITC NPs were investigated via fluorescence analysis. The monolayer properties were characterized by TEM, tight junction staining, transepithelial electrical resistance and fluorescein sodium transportation. The transport ability was measured in the cell based transwell model by confocal imaging and SEM.ResultsPA-FITC NPs were almost spherical shape with a size range of 200–300 nm. Cell viability of BeWo b30 cells was up to 100% in all groups. The concentrations of HCG increased with increasing numbers of cells and culture time, which showed the good biological function of BeWo b30 cells. PA-FITC NPs were rapidly endocytosed through caveolae-mediated endocytosis and pinocytosis, with uptake inhibition rates with nystatin (NY) and colchicines (Col) of 55% and 51% respectively. BeWo b30 cell monolayer was formed over 5 days. PA-FITC NPs were found in the cytoplasm of cells on the transwell membranes; while some NPs were found in the basolateral (fetal) compartment over 24 h.ConclusionIn summary, PA-FITC NPs are nontoxic, can cross the blood-placental barrier, and show mainly internalization to BeWo b30 cells through caveolae-mediated endocytosis and pinocytosis pathways, major via the former pathway. The results could benefit the adjustment and control of the transplacental transport of nanomedicines.

Controlled Release of Ursodeoxycholic Acid from Pullulan Acetate Nanoparticles to Modulate Glutamate-Induced Excitotoxicity in PC-12 Cells

The neuroprotective effects of the ursodeoxycholic acid- (UDCA-) loaded pullulan acetate (PA) (UDCA-PA) nanospheres stabilized by poly(vinyl alcohol) (PVA) were identified by in vitro study. The UDCA-PA nanospheres were constructed by nanoemulsion process. The UDCA-PA nanospheres were analyzed using Fourier transform-infrared spectroscopy (FT-IR) and transmission electron microscopy (TEM). Then, the UDCA-PA nanospheres were used to treat PC-12 neuronal cells, which were formerly triggered by glutamate-induced excitotoxicity. As a result, the cells treated with the UDCA-PA nanospheres showed higher survival rate against glutamate-induced excitotoxicity. Furthermore, the UDCA-PA nanospheres decreased immunoreactivity of Annexin V, a membrane marker for apoptotic cells, in PC-12 with glutamate-induced injury. Particularly, the UDCA-PA nanospheres decreased the level of apoptosis-related proteins such as caspase-3. Taken together, the UDCA-PA nanospheres increased neuroprotective effects against glutamate-induced neuronal damage via inhibition of apoptosis at low concentration.

Synthesis, Characterization and In Vivo Efficacy of Biotin-Conjugated Pullulan Acetate Nanoparticles as a Novel Anticancer Drug Carrier.

In this study, biotin grafted pullulan acetate (Bio-PA) copolymer were synthesized and Bio-PA nanoparticles (Bio-PA NPs) were prepared to be applied as a targeted and prolonged delivery system for anticancer drugs. Bio-PA NPs were fabricated by two strategies, dialysis method and solvent diffusion method. Bio-PA NPs were spherical shaped and had a smaller size and higher positive zeta potential which were fabricated by dialysis method. And the content of biotin on the surface of Bio-PA NPs prepared by dialysis was higher than that of by solvent diffusion method. Epirubicin (EPI), as a model drug, was encapsulated into Bio-PA NPs by dialysis method with a drug encapsulation efficiency of 79.8±3.0%. To study the effect of various cell uptake inhibitors on the uptake of Bio-PA NPs, human liver carcinoma cells (HepG2), human breast adenocarcinoma cells (MCF-7) and human emborynic kidney cells (293T) were preincubated with the inhibitors individually. The results showed that the clathrin-mediated endocytosis and the cell macropinocytosis were involved in the cellular internalizations of Bio-PA NPs in three cells, but the biotin mediated endocytosis was only observed in the biotin-overexpressed HepG2 and MCF-7 cells. After intravenous injection to HepG2-bearing nude mice, Bio-PA NPs achieved the enhanced antitumor effects and the increased distributions in the tumor. In summary, Bio-PA NPs, being used as a novel carrier for EPI, displayed significantly enhanced anticancer activities, indicating their potential applications in cancer chemotherapy.

Effect of inhibitors of endocytosis and NF-kB signal pathway on folate-conjugated nanoparticle endocytosis by rat Kupffer cells.

The regular accumulation of nanoparticles in the liver makes them hepatotoxic and decreases the circulation time, thus reducing their therapeutic effect. Resolving this problem will be significant in improving bioavailability and reducing side effects. In this study, we reduced the phagocytosis of epirubicin (EPI)-loaded folic acid-conjugated pullulan acetate (FPA/EPI) nanoparticles by Kupffer cells (KCs) through internalization and nuclear factor kappa B (NF-kB) signal pathway inhibitors, thus allowing development of FPA/EPI nanoparticles as a nanodrug delivery system (NDDS) based on our previous study. FPA/EPI nanoparticles were prepared by the dialysis method. Rat KCs were preincubated with the following individual or compound inhibitors: chlorpromazine (CPZ), nystatin (NY), colchicine (Col), amiloride (AMR), and pyrrolidine dithiocarbamate (PDTC). Dose- and time-dependent cellular uptake effects of inhibitors on FPA/EPI nanoparticles were determined through fluorometry. The cytokine levels of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), and IL-6 were tested in culture supernatants by bead-based multiplex flow cytometry. The uptake study demonstrated that inhibitors had an obvious inhibitory effect (P<0.05 or P<0.01), with NY, AMR and Col all showing time-dependent inhibitory effects. PDTC + NY had the strongest inhibitory effect, with an uptake rate of 14.62%. The levels of the three proinflammatory cytokines were changed significantly by the compound inhibitors. TNF-α was significantly inhibited (P<0.05 or P<0.01), but IL-1β and IL-6 showed smaller decreases. These results suggested that clathrin- and caveolae-mediated endocytosis were the main routes via which nanoparticles entered KCs and that the NF-kB signal pathway was very important too. In summary, multiple mechanisms, including clathrin- and caveolae-mediated endocytosis, contribute to cytokine production in macrophages following exposure to folic acid-conjugated pullulan acetate nanoparticles. Thus, the endocytosis inhibition strategy has great potential for improving therapy and reducing toxicity of an NDDS in the treatment of cancer.

Cytotoxicity and Cellular Internalization of Epirubicin-Loaded Folate-Conjugated Pullulan Acetate Nanoparticles in Cancer Cells

Cytotoxicity and Cellular Internalization of Epirubicin-Loaded Folate-Conjugated Pullulan Acetate Nanoparticles in Cancer Cells

Curcumin loaded on pullulan acetate nanoparticles protects the liver from damage induced by DEN

Curcumin loaded nanoparticles protect liver from damage induced by DEN.

Statistical modeling of pullulan production and its application in pullulan acetate nanoparticles synthesis

This work focused on the optimization of exo-polysaccharide, pullulan production by exploiting cassava bagasse, an agricultural solid waste residue by solid state fermentation and its application in the preparation of pullulan acetate nanoparticles. Statistical approach was investigated to maximize the pullulan production using C/N ratio, initial pH, NaCl and ZnSO4·5H2O. The optimum conditions for maximum yield of pullulan (39.42±0.62mg/gds) were found to be: C/N=25.94, initial pH=5.5 and NaCl=0.55g/L. Using the optimized medium variables, the production of pullulan was investigated in lab scale solid state fermentation. The pullulan produced was characterized by thermo gravimetric and XRD analysis. Also pullulan acetate nanoparticles were synthesized from chemical modification of pullulan and the average particle size of nanoparticles was examined by zeta particle sizer.

Stability, Pharmacokinetics, Biodistribution and Safety Assessment of Folate-Conjugated Pullulan Acetate Nanoparticles as Cervical Cancer Targeted Drug Carriers.

It is recognized that the stability and journey in the body of nanoparticles are important issues for drug formulations. In this study, we prepared folate-conjugated pullulan acetate nanoparticles (FPANs) and epirubicin loaded FPANs (FPA/EPI) using dialysis method. The storage stability of FPANs and FPA/EPI at 4 degrees C could be up to 3 months. Using folate receptor overexpressed Hela cells, dose dependent cellular uptake and receptor-mediated endocytosis of FPA/EPI were confirmed. From the in vivo pharmacokinetics test, compared to free EPI, half-life time (t½) of FPA/EPI was extended 1.57 times and the area under-the-curve (AUC) increased 3.95 times as well. In addition, biodistribution data showed that, EPI concentration in tumor in FPA/EPI group was 2.01 times higher than that in free EPI group after 96 h; The concentration of drug in liver treated by FPA/EPI was 5.7-11.6 times, while in heart, kidney, especially in stomach and intestine were much lower than those in free EPI group from 24 to 96 h. Furthermore, blank FPANs showed no apparent acute toxicity at dose up to 125 mg/kg. All results suggested that FPA/EPI showed a promising potential on treating cervical carcinoma and its metastatic hepatocellular carcinoma in future because of the high stability, less toxicity and tumor targeting.

Combination of paclitaxel- and retinoic acid-incorporated nanoparticles for the treatment of CT-26 colon carcinoma

The aim of this study was to evaluate the antitumor effect of combinatorial targeted therapy with paclitaxel and all-trans retinoic acid (ATRA) nanoparticles in vitro. Paclitaxel-incorporated pullulan acetate (PA) nanoparticles were prepared by the nanoprecipitation-solvent evaporation method. ATRA-incorporated nanoparticles were prepared by dialysis using a methoxy poly(ethylene glycol)-grafted chitosan (ChitoPEG) copolymer. Particle sizes of paclitaxel-incorporated nanoparticles and ATRA-incorporated nanoparticles were about 160 nm and 60 nm, respectively. Nanoparticles were reconstituted in various aqueous media such as deionized water, phosphate-buffered saline, and fetal bovine serum-supplemented cell culture media. The combination of paclitaxel + ATRA (10 + 10 μg/mL) delivered by nanoparticles showed a synergistic antiproliferative effect against CT26 cells that was not observed with other combinations. Furthermore, the activity of MMP-2, a key enzyme in tumor cell invasion, was significantly decreased in cells treated with the combination of paclitaxel and ATRA while other combinations and single agents did not significantly affect its activity. A matrigel assay supported these results, indicating that paclitaxel/ATRA combination nanoparticles are effective for the inhibition of the invasion of tumor cells. The results of the present study suggest that combination treatment with paclitaxel and ATRA could be an effective treatment for the inhibition of tumor cell proliferation and invasion, and that nanoparticles are promising candidates for antitumor drug delivery.

Stability and in vivo evaluation of pullulan acetate as a drug nanocarrier

To develop pullulan acetate nanoparticles (PANs) as a drug nanocarrier, pullulan acetate (PA) was synthesized and characterized. Its acetylation degree determined by the proton nuclear magnetic resonance (1H NMR) was 2.6. PANs were prepared by the solvent diffusion method and characterized by transmission electron microscope (TEM), size distribution, and ζ potential techniques. PANs had nearly spherical shape with a size range of 200–450 nm and low ζ potentials both in distilled water and in 10% FBS. The storage stability of PANs was observed in distilled water. PANs were stored for at least 2 months with no significant size and ζ potential changes. The safety of PANs was studied through single dose toxicity test in mice, and the result showed that PANs were well tolerated at the dose of 200 mg/kg in mice. Epirubicin-loaded PANs (PA/EPI) were also prepared and characterized in this study. Moreover, the in vivo pharmacokinetics of PA/EPI was investigated. Compared with the free EPI group, the PA/EPI group exhibited higher plasma drug concentration, longer half-life time (t1/2) and the larger area under the curve (AUC). All results suggested that PANs were stable, safe, and showed a promising potential on improving the bioavailability of the loaded drug of the encapsulated drug.

Preparation of folate-modified pullulan acetate nanoparticles for tumor-targeted drug delivery

The purpose of this work was to develop a novel nano-carrier with targeting property to tumor. In this study, pullulan acetate (PA) was synthesized by the acetylation of pullulan to simplify the preparation technique of nanoparticles. Folic acid (FA) was conjugated to PA in order to improve the cancer-targeting activity. The products were characterized by proton nuclear magnetic resonance (1H NMR) spectroscopy. Epirubicin-loaded nanoparticles were prepared by a solvent diffusion method. The loading efficiencies and EPI content increased with the amount of triethylamine (TEA) increasing in some degree. FPA nanoparticles could incorporate more epirubicin than PA nanoparticles. The folate-modified PA nanoparticles (FPA/EPI NPs) exhibited faster drug release than PA nanoparticles (PA/EPI NPs) in vitro. Confocal image analysis and flow cytometry test revealed that FPA/EPI NPs exhibited a greater extent of cellular uptake than PA/EPI NPs against KB cells over-expressing folate receptors on the surface. FPA/EPI NPs also showed higher cytotoxicity than PA/EPI NPs. The cytotoxic effect of FPA/EPI NPs to KB cells was inhibited by an excess amount of folic acid, suggesting that the binding and/or uptake were mediated by the folate receptor.

Pullulan acetate nanoparticles prepared by solvent diffusion method for epirubicin chemotherapy

Pullulan acetate (PA) was synthesized by the reaction of pullulan with acetic anhydride in the presence of pyridine. PA was characterized by Fourier transform infrared (FT-IR) and proton nuclear magnetic resonance ( 1H NMR). A solvent diffusion method was employed in the current work to prepare PA nanoparticles. This technique had some advantages compared with other methods. The particle size increased from 185.7 nm to 423.0 nm with the degree of acetylation increasing from 2.71 to 3.0. Drug-loaded PA nanopaticles were prepared for controlled release of epirubicin (EPI). The drug entrapment and drug content increased with the degree substitution of PA increasing. EPI was released from the nanoparticles in a biphasic profile with a fast release rate in the first 10 h followed by a slow release in vitro. A higher cytotoxicity against KB cells was found for EPI-loaded PA nanoparticles in comparison with free EPI. Confocal laser scanning microscopy (CLSM) observations indicate that EPI-loaded nanoparticles were internalized and released in the cytoplasmic compartment.

Ionic strength-sensitive pullulan acetate nanoparticles (PAN) for intratumoral administration of radioisotope: Ionic strength-dependent aggregation behavior and 99mTechnetium retention property

In order to design an effective intratumoral radioisotope carrier, a self-assembled nanoparticle evidencing ionic strength (IS)-sensitivity from a polysaccharide derivative (pullulan acetate nanoparticle (PAN)) was prepared via dialysis. The PAN had a spherical shape in a range of size of 50–130 nm and a low critical aggregation concentration (CAC) (<8 μg/mL). With increases in the IS of the dialysis media (IS dia), the CAC of PAN was reduced gradually and the rigidity of the hydrophobic core in PAN was increased. This suggests that the property of PAN was altered more hydrophobically at high IS values. The stabilities of PANs prepared from various IS dia were also monitored with changes in the turbidity and particle size in different IS solutions. In the case of PAN prepared at an IS dia = 0.0, the turbidity was dramatically reduced with increasing IS due to the facilitation of aggregation between the particles, whereas in the other cases, these changes were negligible. This finding indicates that PAN prepared in distilled water (IS = 0.0) can be readily injected as the consequence of its nano-size, and accumulates quickly, then remains in the tumor site for a considerable period (IS = 0.15). In order to closely estimate the potential of PAN as a radioisotope carrier, the radioisotope labeling efficiency of PAN with no chelating agents was evaluated. PAN evidenced a high degree of 99mTechnetium ( 99mTc) labeling efficiency (approximately 98%). The percentage retention rate (%RR) of the 99mTc-labeled PAN was significantly longer than that of the free 99mTc ( p < 0.05), due largely to PAN's IS-sensitivity. In conclusion, PAN may constitute a new approach to the achievement of maximal radioisotope efficiency with regard to intratumoral administration.

Self-assembled nanoparticles of hydrophobically-modified polysaccharide bearing vitamin H as a targeted anti-cancer drug delivery system

Vitamin H (biotin) was incorporated into a hydrophobically modified polysaccharide, pullulan acetate (PA), in order to improve the cancer-targeting activity and internalization of self-assembled nanoparticles. The biotinylated pullulan acetate (BPA) nanoparticles were prepared by a diafiltration method and the mean diameter was approximately 100 nm. Three samples of biotinylated pullulan acetate (BPA), comprising 7 (BPA 1), 20 (BPA 2), and 39 (BPA 3) vitamin H groups per 100 anhydroglucose units of PA, were synthesized. The critical aggregation concentrations (CAC) of the BPA nanoparticles in distilled water were 3.1×10 −3, 4.3×10 −3 and 6.8×10 −3 mg/ml for BPA 1, BPA 2, and BPA 3, respectively. Adriamycin (ADR) was loaded into the BPA nanoparticles as a model drug. The loading efficiencies and ADR content in the BPA nanoparticles decreased with increasing vitamin H content due to a lower hydrophobicity. The RITC-labeled BPA nanoparticles exhibited very strong adsorption to the HepG2 cells, while the RITC-labeled PA nanoparticles did not show any significant interaction. The degree of the interaction increased with increasing vitamin H content. Confocal laser microscopy also revealed that internalization of the BPA nanoparticles into the cancer cells depended on the vitamin H content.