Swelling behavior and controlled release of theophylline and sulfamethoxazole drugs in beta-lactoglobulin protein gels obtained by phase separation in water/ethanol mixture

Physically cross-linked beta-lactoglobulin (BLG) protein gels containing theophylline and sulfamethoxazole low molecular weight drugs were prepared in 50% ethanol solution at pH 8 and two protein concentrations (6 and 7% (w/v)). Swelling behavior of cylindrical gels showed that, irrespective of the hydrated or dehydrated state of the gel, the rate of swelling was the highest in water. When the gels were exposed to water, they first showed a swelling phase in which their weight increased 3 and 30 times for hydrated and dehydrated gels, respectively, due to absorption of water, followed by a dissolution phase. The absorption of solvent was however considerably reduced when the gels were exposed to aqueous buffer solutions. The release behavior of both theophylline and sulfamethoxazole drugs from BLG gels was achieved in a time window ranging from 6 to 24 h. The drug release depended mainly on the solubility of the drugs and the physical state of the gel (hydrated or dry form). Analysis of drug release profiles using the model of Peppas showed that diffusion through hydrated gels was governed by a Fickian process whereas diffusion through dehydrated gels was governed partly by the swelling capacities of the gel but also by the structural rearrangements inside the network occurring during dehydration step. By a judicious selection of protein concentration, hydrated or dehydrated gel state, drug release may be modulated to be engineered suitable for pharmaceutical as well as cosmetics and food applications.     

Preparation and characterization of N-(2-carboxybenzyl)chitosan as a potential pH-sensitive hydrogel for drug delivery

A novel water-soluble chitosan derivative [N-(2-carboxybenzyl)chitosan, CBCS] was synthesized. The chemical structure of CBCS was characterized by FTIR, (1)H NMR and UV spectroscopies. The degree of substitution (DS) of N-2-carboxybenzyl was determined by colloid titration. In different pH buffer solutions, the swelling characteristics of hydrogels based on CBCS (CBCSG) prepared by crosslinking with glutaraldehyde have been studied. Results showed that the swelling ratio (SR) of CBCSG decreased with an increase of the amount of glutaraldehyde, and that CBCSG swelled more significantly in alkaline solution than in acidic medium, showing the lowest SR at pH5.0. The SR of CBCSG increased with the raising of the DS of the N-2-carboxybenzyl group in alkaline solution, but no significant change was observed in an acidic environment. CBCSG showed swelling reversibility when alternately soaked in pH1.0 and 7.4 buffer solutions. Release profiles of fluorouracil (5-FU), a poorly water-soluble drug, from CBCSG were studied under both simulated gastric and intestinal pHconditions. The release was much quicker in pH7.4 buffer than in pH1.0 solution. Results indicated that CBCS could be a potential pH-sensitive carrier for colon-specific drug delivery system.     

Design and <Emphasis Type="Italic">In Vitro</Emphasis> Evaluation of Capsaicin Transdermal Controlled Release Cubic Phase Gels

The purpose of this study was to design and investigate the transdermal controlled release cubic phase gels containing capsaicin using glycerol monooleate (MO), propylene glycol (1,2-propanediol, PG), and water. Three types of cubic phase gels were designed based on the ternary phase diagram of the MO–PG–water system, and their internal structures were confirmed by polarizing light microscopy (PLM) and small-angle X-ray scattering (SAXS). Release results showed the cubic phase gels could provide a sustained system for capsaicin, while the initial water content in the gels was the major factor affecting the release rate. Release kinetics was determined to fit Higuchi’s square-root equation indicating that the release was under diffusion control. The calculated diffusion exponent showed the release from cubic phase gels was anomalous transport. The unique structure of the cubic phases, capsaicin distributed in the lipid bilayers, and cubic phase gel swelling contributed to the release mechanism. The cubic phase gel may be an interesting application for transdermal delivery system of capsaicin in alleviating the post-incision pain.     

Galactosylated nanostructured lipid carriers for delivery of 5-FU to hepatocellular carcinoma

The aim of the present study was to design a targeted delivery system of 5-fluorouracil (5-FU) for hepatocellular carcinoma (HCC). Lactobionic acid (LB) was conjugated to stearyl amine (SA) by a chemical reaction. The nanostructured lipid carriers (NLCs), containing LB conjugate, lecithin, glyceryl monostearate, oil [oleic acid (OA) or Labrafac 5 or 10%], and 5-FU, were dissolved in alcohol/acetone, the oil phase was added to the aqueous phase containing Tween 80 or Solutol(?) HS15 (0.25 or 0.5%), and NLCs were prepared by an emulsification-solvent diffusion method. Physical properties and drug release were studied in NLCs. The thiazolyl blue tetrazolium bromide assay was used to study the cytotoxicity of NLCs on HepG(2) cells, and the cellular uptake of NLCs was determined by flow cytometry. Fourier transform infrared spectroscopy and (1)H-NMR spectra confirmed the successful conjugation of LB and SA. The optimized NLCs consisted of 0.5% Solutol HS15 and 10% OA oil. The particle size of these nanoparticles was 139.2 nm, with a zeta potential of -18 mV, loading efficiency of 34.2%, release efficiency after 2 hours of the release test was 72.6%, and crystallinity was 0.63%. The galactosylated NLCs of 5-FU were cytotoxic on the HepG(2) cell line in a half concentration of 5-FU and seems promising in reducing 5-FU dose in HCC.     

PEGylation of bovine serum albumin using click chemistry for the application as drug carriers

Monomethyl poly(ethylene glycol) (mPEG)-modified bovine serum albumin (BSA) conjugates (BSA-mPEG) were obtained by the mild Cu(I)-mediated cycloaddition reaction of azided BSA (BSA-N(3) ) and alkyne-terminated mPEG. The structure and characteristics of BSA-mPEG conjugates were thoroughly investigated. There were about two PEG chains conjugated onto each BSA molecule as determined by matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) analysis. The intrinsic nonspecific binding ability of BSA was used for adsorption and sustained release of both rifampicn and 5-fluorouracil (5-FU). The helical structures of BSA were preserved to a large extent after modification and drug adsorption on BSA was confirmed via circular dichroism spectroscopy. Drugs adsorbed onto the conjugated formulation to a lesser extent than on BSA due to mPEG modification. The in vitro release of both rifampicin and 5-FU, however, indicated that BSA-mPEG can function as a drug carrier. Overall, the click reaction provided a convenient tool for the pegylation of BSA. The biological activity of the BSA-mPEG conjugates, including the drug transportation capacity and biocompatibility, were largely retained.     

Galactosylated nanostructured lipid carriers for delivery of 5-FU to hepatocellular carcinoma

The aim of the present study was to design a targeted delivery system of 5-fluorouracil (5-FU) for hepatocellular carcinoma (HCC). Lactobionic acid (LB) was conjugated to stearyl amine (SA) by a chemical reaction. The nanostructured lipid carriers (NLCs), containing LB conjugate, lecithin, glyceryl monostearate, oil [oleic acid (OA) or Labrafac 5 or 10%], and 5-FU, were dissolved in alcohol/acetone, the oil phase was added to the aqueous phase containing Tween 80 or Solutol^® HS15 (0.25 or 0.5%), and NLCs were prepared by an emulsification-solvent diffusion method. Physical properties and drug release were studied in NLCs. The thiazolyl blue tetrazolium bromide assay was used to study the cytotoxicity of NLCs on HepG₂ cells, and the cellular uptake of NLCs was determined by flow cytometry. Fourier transform infrared spectroscopy and ¹H-NMR spectra confirmed the successful conjugation of LB and SA. The optimized NLCs consisted of 0.5% Solutol HS15 and 10% OA oil. The particle size of these nanoparticles was 139.2 nm, with a zeta potential of –18 mV, loading efficiency of 34.2%, release efficiency after 2 hours of the release test was 72.6%, and crystallinity was 0.63%. The galactosylated NLCs of 5-FU were cytotoxic on the HepG₂ cell line in a half concentration of 5-FU and seems promising in reducing 5-FU dose in HCC.     

5-Fluorouracil Nanoparticles Inhibit Hepatocellular Carcinoma via Activation of the p53 Pathway in the Orthotopic Transplant Mouse Model

Biodegradable polymer nanoparticle drug delivery systems provide targeted drug delivery, improved pharmacokinetic and biodistribution, enhanced drug stability and fewer side effects. These drug delivery systems are widely used for delivering cytotoxic agents. In the present study, we synthesized GC/5-FU nanoparticles by combining galactosylated chitosan (GC) material with 5-FU, and tested its effect on liver cancer in vitro and in vivo. The in vitro anti-cancer effects of this sustained release system were both dose- and time-dependent, and demonstrated higher cytotoxicity against hepatic cancer cells than against other cell types. The distribution of GC/5-FU in vivo revealed the greatest accumulation in hepatic cancer tissues. GC/5-FU significantly inhibited tumor growth in an orthotropic liver cancer mouse model, resulting in a significant reduction in tumor weight and increased survival time in comparison to 5-FU alone. Flow cytometry and TUNEL assays in hepatic cancer cells showed that GC/5-FU was associated with higher rates of G0–G1 arrest and apoptosis than 5-FU. Analysis of apoptosis pathways indicated that GC/5-FU upregulates p53 expression at both protein and mRNA levels. This in turn lowers Bcl-2/Bax expression resulting in mitochondrial release of cytochrome C into the cytosol with subsequent caspase-3 activation. Upregulation of caspase-3 expression decreased poly ADP-ribose polymerase 1 (PARP-1) at mRNA and protein levels, further promoting apoptosis. These findings indicate that sustained release of GC/5-FU nanoparticles are more effective at targeting hepatic cancer cells than 5-FU monotherapy in the mouse orthotropic liver cancer mouse model.     

Preparation and characterization of konjac glucomannan–poly(acrylic acid) IPN hydrogels for controlled release

In this paper, we reported the synthesis and properties of interpenetrating polymer network (IPN) hydrogel systems designed for colon targeted drug delivery. The gels were composed of konjac glucomannan (KGM) and cross-linked poly(acrylic acid) (PAA) by N,N-methylene-bis-(acrylamide) (MBAAm). It was possible to modulate the swelling degree of the gels. And the swelling ratio has sensitive respondence to the environmental pH value variation. The degradation tests show that the hydrogels retain the enzymatic degradation character of KGM. In vitro release of model drug VB₁₂ was studied in the presence of Cellulase E0240 in pH 7.4 phosphate buffer at 37 °C. The accumulative release percent of the model drug reached 85.6% after 48 h and the drug release was controlled by the swelling and the degradation of the hydrogels. The results indicated that the IPN hydrogels can be exploited as potential carriers for colon-specific drug delivery.     

The Biodegradation of Zein <Emphasis Type="Italic">In Vitro</Emphasis> and <Emphasis Type="Italic">In Vivo</Emphasis> and its Application in Implants

A unique polymer-based sustained-release implant drug delivery system was prepared by using biocompatible and biodegradable Zein as the skeleton meterial. After preparing Zein colloids, the Zein-loaded implant rods were formulated by injection molding followed by evaporating the solvent, and being coated with poly(lactic-co-glycolic) acid (PLGA) solution. Drug release kinetics was examined by using Fluorouracil (5-FU) as model drug. Nearly zero-order release was achieved for the model drugs for a period of 0–25 days when the implants were incubated in distilled water at 37°C. And then the degradation kinetics of the rods in vivo and in vitro were evaluated, which indicated that Zein could be absorbed by body and has good degradation property. The effects of different ratios of Zein/5-FU and the rods’ diameter on drug release were studied, respectively. The plasma concentration of 5-FU in the implants were determined by HPLC after implanting a single dose of the implants in rats. All data were subsequently processed by using the computer program 3P97, and the values were showed as follows: the area under the plasma concentration–time curve (AUC) value was 321.88 (μg/ml) × day, and the mean residence time (MRT) value was 23.05 days. The sustained-release implants of Zein/5-FU were successfully formulated. The uniqueness of the article is that Zein has been used as a skeleton material in implant delivery system for the first time and zero-order release kinetics has been obtained successfully.     

Pectin-coated chitosan microgels crosslinked on superhydrophobic surfaces for 5-fluorouracil encapsulation

5-Fluorouracil (5-FU)-loaded chitosan microgels for oral and topical chemotherapy were prepared applying a superhydrophobic surface-based encapsulation technology. Drug-loaded chitosan dispersions were cross-linked and then coated with drug-free chitosan or pectin layers at the solid-air interface in a highly efficient and environment-friendly way. The size of the microgels (with diameters of ca. 280 and 557 μm for the chitosan seeds and pectin-coated microgels respectively) was the lowest obtained until now using similar biomimetic methodologies. The microgels were characterized regarding 5-FU release profiles in vitro in aqueous media covering the pH range of the gastrointestinal tract, and cytotoxicity against two cancer cell lines sensitive to 5-FU. Owing to their control of 5-FU release in acidic medium, calcium pectinate-coated microgels can be considered as suitable for oral administration. Growth inhibition of cancer cells by 5-FU was greater when incorporated to chitosan microgels; these being potentially useful for treatment of skin and colorectal tumors.     

Formulation and characterization of nanoliposomal 5-fluorouracil for cancer nanotherapy

A scalable and safe method was developed to prepare nanoliposome carriers for the entrapment and delivery of 5-fluorouracil (5-FU). The carrier systems were composed of endogenously occurring dipalmitoylphosphatidylcholine (DPPC), negatively charged dicetylphosphate (DCP), cholesterol (CHOL) and glycerol (3%, v/v). Nanoliposomes were prepared by the heating method in which no harmful chemical or procedure is involved. Results indicated fast and reproducible formation of non-toxic liposomes that possess high entrapment efficiency (up to 96.9%) and vesicle size range of ca. 530–620?nm. Transmission electron and optical micrographs of the 5-FU liposomes revealed that they were spherical and some were multilayered. There was an increase in the release rate of 5-FU from the liposomes prepared with a high ratio of drug:lipid. The release data showed that the highest release rates were obtained for nanoliposomes containing 5-FU with the drug concentration of 500?mM and that it followed the diffusion model. Nanoliposome preparation method introduced here has the potential of large-scale manufacture of safe and efficient carriers of 5-FU.     

Development of hydrocortisone succinic acid/and 5-fluorouracil/chitosan microcapsules for oral and topical drug deliveries

Recently, we demonstrated the safety use of calendula oil/chitosan microcapsules as a carrier for both oral and topical deliveries. We also reported the improved biological activity towards skin cells and Staphylococcus aureus of phyllanthin containing chitosan microcapsules. However, the possibility of both oral and topical applications was still necessary to be further studied. Here we investigated that both oral and topical applications of chitosan-based microcapsules were tested using hydrocortisone succinic acid (HSA) and 5-fluorouracil (5-FU), respectively. The drug loading efficiency, particle size, surface morphology and chemical compositions of both drug loaded microcapsules were confirmed by UV-vis spectrophotometer, particle size analyzer, scanning electron microscope and Fourier transform infrared spectroscopy. The in vitro release studies revealed that both HSA and 5-FU could be released form chitosan microcapsules. The mean adrenocorticotropic hormone concentration in HSA loaded microcapsule mice plasma was detected to be lower than that of water control. One hundred micrograms per milliliter of 5-FU containing microcapsules exhibited a stronger growth inhibition towards skin keratinocytes than that of free 5-FU. In vitro drug delivery model demonstrated the delivery of 5-FU from microcapsule treated textiles into nude mice skin. Further uses of the drug loaded microcapsules may provide an efficiency deliverable tool for both oral and topical applications.     

Optimization of an Aqueous Tablet-Coating Process Containing Carboxymethylated Cassia fistula Gum

The present investigation was aimed at developing and optimizing a simple aqueous tablet-coating formulation and its process. 5-Fluorouracil (5-FU) was used to ascertain the relative lipophilic/hydrophilic behavior of the coating system. Optimization was performed by evaluating the adhesive force strength and cohesive force strength of the tablet coat using a texture analyzer. The in vitro release of 5-FU was found to decrease with an increase in (tablet surface-coat) adhesive force strength. The (tablet-tablet) cohesive force strength was reduced by the addition of magnesium silicate to the coating solution. The addition of magnesium silicate (0.2% w/v) to the carboxymethyl Cassia fistula gum-chitosan (CCG-CH) coating surface significantly inhibited the release of 5-FU possibly due to an increase in the hydrophobic character of the coated tablet surface. This was possible by coating cohesive force strength reduction coating compositions (CCG-CH (70:30) and 0.3% magnesium silicate). Further, the FTIR-ATR and DSC analyses suggested the pivotal role of magnesium silicate in modifying the release of 5-FU from CCG-CH-coated tablets due to hydrogen bonding of its Si-O-Si or Mg-O groups with -OH moieties of CCG-CH.     

Comparative label-free LC-MS/MS analysis of colorectal adenocarcinoma and metastatic cells treated with 5-fluorouracil

A label-free mass spectrometric strategy was used to examine the effect of 5-fluorouracil (5-FU) on the primary and metastatic colon carcinoma cell lines, SW480 and SW620, with and without treatment. 5-FU is the most common chemotherapeutic treatment for colon cancer. Pooled biological replicates were analyzed by nanoLC-MS/MS and protein quantification was determined via spectral counting. Phenotypic and proteomic changes were evident and often similar in both cell lines. The SW620 cells were more resistant to 5-FU treatment, with an IC(50) 2.7-fold higher than that for SW480. In addition, both cell lines showed pronounced abundance changes in pathways relating to antioxidative stress response and cell adhesion remodeling due to 5-FU treatment. For example, the detoxification enzyme NQO1 was increased with treatment in both cell lines, while disparate members of the peroxiredoxin family, PRDX2 or PRDX5 and PRDX6, were elevated with 5-FU exposure in either SW480 or SW620, respectively. Cell adhesion-associated proteins CTNNB1 and RhoA showed decreased expression with 5-FU treatment in both cell lines. The differential quantitative response in the proteomes of these patient-matched cell lines to drug treatment underscores the subtle molecular differences separating primary and metastatic cancer cells.     

Combinations of 5-fluorouracil with UCN-01 or staurosporine

The action of 5-Fluorouracil (5-FU) is mediated by inhibition of thymidylate synthase (TS), which is regulated by cell cycle proteins controlled by protein phosphorylation. We studied the effects of staurosporine and its analogue UCN-01, inhibitors of protein kinase C (PKC) on 5-FU cytotoxicity in Lovo colon cancer cells. Each drug contributes equally to the cell cycle effects of the 5-FU combinations. In sequential drug administration, the cell cycle distribution was determined by the first drug. Simultaneous 5-FU combinations induced additive effects in induction of apoptosis. When staurosporine was used as the second drug, induction of apoptosis was 2-fold higher than the sum of both drugs alone. Based on induction of apoptosis 5-FU addition prior to the PKC inhibitors seemed preferable.     

Matrix metalloproteinase-8 mediates the unfavorable systemic impact of local irradiation on pharmacokinetics of anti-cancer drug 5-Fluorouracil

Concurrent chemoradiation with 5-fluorouracil (5-FU) is widely accepted for cancer treatment. However, the interactions between radiation and 5-FU remain unclear. Here, we evaluated the influence of local irradiation on the pharmacokinetics of 5-FU in rats. The single-fraction radiation was delivered to the whole pelvic fields of Sprague-Dawley rats after computerized tomography-based planning. 5-FU at 100 mg/kg was prescribed 24 hours after radiation. A high-performance liquid chromatography system was used to measure 5-FU in the blood. Matrix metalloproteinase-8 (MMP-8) inhibitor I was administered to examine whether or not RT modulation of 5-FU pharmacokinetic parameters could be blocked. Compared with sham-irradiated controls, whole pelvic irradiation reduced the area under the concentration versus time curve (AUC) of 5-FU in plasma and, in contrast, increased in bile with a radiation dose-dependent manner. Based on protein array analysis, the amount of plasma MMP-8 was increased by whole pelvic irradiation (2.8-fold by 0.5 Gy and 5.3-fold by 2 Gy) in comparison with controls. Pretreatment with MMP-8 inhibitor reversed the effect of irradiation on AUC of 5-FU in plasma. Our findings first indicate that local irradiation modulate the systemic pharmacokinetics of 5-FU through stimulating the release of MMP-8. The pharmacokinetics of 5-FU during concurrent chemoradiaiton therapy should be rechecked and the optimal 5-FU dose should be reevaluated, and adjusted if necessary, during CCRT.     

A novel 5-fluorouracil prodrug using hydroxyethyl starch as a macromolecular carrier for sustained release

The objective of this study was to develop a sustained-release drug delivery system for 5-fluorouracil (5-FU) to improve its short half-life. 5-Fluorouracil-1-acetic acid (FUAC) was prepared and then conjugated to hydroxyethyl starch (HES) through ester bonds. The conjugates were relatively stable in acidic buffer solution at pH 5.8 and slowly released FUAC but became more sensitive to hydrolysis with an increase in the pH and temperature. The conjugates were degraded to FUAC both in human and rat plasma with half-time life of 20.4 h and 24.6 h, respectively. Both 5-FU and FUAC were released in a rat liver homogenate following a 12 h incubation of the conjugates. The pharmacokinetic behavior was evaluated in rats after intravenous injection of 5-FU, FUAC and the conjugates. The drug release data in vitro and in vivo indicated that HES is a promising carrier for the sustained-release of antitumor drugs.     

Photon transmission study on swelling of kappa-carrageenan gels prepared in various concentrations

κ-Carrageenan gels prepared with various carrageenan concentrations in pure water were completely dried and then swelled in pure water. Photon transmission measurements were performed using a UV-Vis (UVV) spectrometer during the swelling of κ-carrageenan gels. Transmitted photon intensity, I_tr, increased exponentially as swelling time is increased for all gel samples. The behaviour of I_tr was interpreted by Monte-Carlo Simulation. The increase in I_tr was quantified by employing Li-Tanaka equation, from which time constants τ₁ and collective diffusion coefficients, D_o were determined for the gels in various carrageenan concentrations. Gravimetric and volumetric measurements were also carried out during swelling of gels. It is observed that gel with high carrageenan content possess more double helices and more lattice dislocations and swell slower than gels with low carrageenan content which may contain less double helices and less lattice imperfections. Increase in I_tr was interpreted by the homogeneous distribution of double helices in the carrageenan gel system.