Perfluorodecalin-soluble fluorescent dyes for the monitoring of circulating nanocapsules with intravital fluorescence microscopy.

Perfluorodecalin (PFD) is an established artificial oxygen carrier due to its physical capability to solve the respiratory gases oxygen and carbon dioxide. PFD-filled poly(n-butyl-cyanoacrylate) (PACA) nanocapsules are already discussed as effective artificial oxygen carriers, and their principal suitability for intravenous administration had been shown. To further elucidate their action in vivo, it is imperative to characterise their preclinical safety and particularly their biodistribution. For these purposes, intravital fluorescence microscopy would display an attractive technique in order to monitor the PACA nanocapsules in vivo, but unfortunately, it is impossible to stain the PACA nanocapsules with a fluorescent dye fulfilling special criteria required for in vivo microscopy. In order to develop such a dye, a long-chained fluorinated thiol was used to modify a BODIPY derivative that is a highly fluorescent organic compound belonging to the difluoro-boraindacene family, as well as to functionalise mesoscopic systems, such as CdSe/ZnS-quantum dots and gold nanoparticles. Furthermore, a functionalisation of porphyrin derivatives was investigated by placing divalent ions in the centre of these systems. Due to the high solubility of all synthesised dyes in PFD, it should be possible to stain PFD-filled particles in general. However, only the functionalised BODIPY derivative was suitable for in vivo monitoring of the PFD-filled PACA nanocapsules.

In vivo visualisation of nanoparticle entry into central nervous system tissue.

Because the potential neurotoxicity of nanoparticles is a significant issue, characterisation of nanoparticle entry into the brain is essential. Here, we describe an in vivo confocal neuroimaging method (ICON) of visualising the entry of fluorescent particles into the parenchyma of the central nervous system (CNS) in live animals using the retina as a model. Rats received intravenous injections of fluorescence-labelled polybutyl cyanoacrylate nanoparticles that had been synthesised by a standard miniemulsion polymerisation process. We performed live recording with ICON from before and up to 9 days after particle injection and took photomicrographs of the retina. In addition, selective retrograde labelling of the retinal ganglion cells was achieved by stereotaxic injection of a fluorescent dye into the superior colliculus. Using ICON, we observed vascular kinetics of nanoparticles (wash-in within seconds), their passage to the retina parenchyma (within minutes) and their distribution (mainly cellular) under in vivo conditions. For the detection of cell loss--which is important for the evaluation of toxic effects--in another experiment, we semi-quantitatively analysed the selectively labelled retinal neurons. Our results suggest that the dye per se does not lead to neuronal death. With ICON, it is possible to study nanoparticle kinetics in the retina as a model of the blood-brain barrier. Imaging data can be acquired within seconds after the injection, and the long-term fate of cellular uptake can be followed for many days to study the cellular/extracellular distribution of the nanoparticles. ICON is thus an effective and meaningful tool to investigate nanoparticle/CNS interactions.

Pharmacokinetic analysis and optimization of hydroxycamptothecin-loaded nanoparticles for liver targeting.

In this paper, a pharmacokinetic model to describe the tissue distribution process of nanoparticles was established. To test the possibility of the model, nanoparticles composed of poly(butylcyanoacrylate) and hydroxypropyl-ß-cyclodextrins (HP-ß-CD) was prepared with a poorly soluble anticancer drug, hydroxycamptothecin (HCPT). Characteristics were determined including particle's size, morphology and in vitro release. The tissue distribution of nanoparticles was also studied. Further, mathematical equation was fitted to the curve of drug concentration-time in liver of hydroxycamptothecin-loaded nanoparticles and the pharmacokinetic parameters of liver were obtained. The effectiveness of hydroxycamptothecin-loaded nanoparticles for liver targeting was evaluated. The results showed that nanoparticles composed of poly(butylcyanoacrylate) and hydroxypropyl-ß-cyclodextrins (HP-ß-CD) exhibited enhanced liver targeting in rats after i.v. injection. More importantly, the pharmacokinetic parameters (transport constant from blood to target organ KT, drug release rate from nanoparticles Kr and drug elimination constant in target organ Ke) provided some quantitative measure of liver distribution and were useful guidelines for development of targeted drug delivery systems.

Opsonization, biodistribution, cellular uptake and apoptosis study of PEGylated PBCA nanoparticle as potential drug delivery carrier.

PURPOSE: For nanocarrier-based targeted delivery systems, preventing phagocytosis for prolong circulation half life is a crucial task. PEGylated poly(n-butylcyano acrylate) (PBCA) NP has proven a promising approach for drug delivery, but an easy and reliable method of PEGylation of PBCA has faced a major bottleneck. METHODS: PEGylated PBCA NPs containing docetaxel (DTX) by modified anionic polymerization reaction in aqueous acidic media containing amine functional PEG were made as an single step PEGylation method. In vitro colloidal stability studies using salt aggregation method and antiopsonization property of prepared NPs using mouse macrophage cell line RAW264 were performed. In vitro performance of anticancer activity of prepared formulations was checked on MCF7 cell line. NPs were radiolabeled with 99mTc and intravenously administered to study blood clearance and biodistribution in mice model. RESULTS: These formulations very effectively prevented phagocytosis and found excellent carrier for drug delivery purpose. In vivo studies display long circulation half life of PBCA-PEG20 NP in comparison to other formulations tested. CONCLUSIONS: The PEGylated PBCA formulation can work as a novel tool for drug delivery which can prevent RES uptake and prolong circulation half life.

Efficacy of intravenous amphotericin B-polybutylcyanoacrylate nanoparticles against cryptococcal meningitis in mice.

Amphotericin B deoxycholate (AmB), a classic antifungal drug, remains the initial treatment of choice for deep fungal infections, but it is not appropriate for treatment of cryptococcal meningitis due to its inability to pass through the blood-brain barrier (BBB). We examined the efficacy of amphotericin B-polybutylcyanoacrylate nanoparticles (AmB-PBCA-NPs) modified with polysorbate 80 that had a mean particle diameter less than 100 nanometers (69.0 ± 28.6 nm). AmB-PBCA-NPs were detected in the brain 30 minutes after systemic administration into BALB/c mice and had a higher concentration than systemically administered AmB liposome (AmB-L, P < 0.05); AmB was not detected in the brain. Following infection for 24 hours and then 7 days of treatment, the survival rate of mice in the AmB-PBCA-NP group (80%) was significantly higher than that of the AmB (0%) or AmB-L (60%) treatment groups. Fungal load was also lower when assessed by colony-forming unit counts obtained after plating infected brain tissue (P < 0.05). Our study indicates that AmB-PBCA-NPs with polysorbate 80 coating have the capacity to transport AmB across the BBB and is an efficient treatment against cryptococcal meningitis in a mouse model.

Quinoline-n-butylcyanoacrylate-based nanoparticles for brain targeting for the diagnosis of Alzheimer's disease.

A survey of research activity on nanoparticles (NPs) based on polymeric devices that could cross the blood-brain barrier (BBB) is given along with the presentation of our own data on the development of NPs of n-butyl-2-cyanoacrylate (BCA) for brain delivery to aid the early diagnosis of Alzheimer's disease (AD), a neurodegenerative disorder of the elderly people, the most prevalent form of dementia. Typical data are presented on in vivo detection of amyloid peptides (A beta) (amyloid plaques) that are used as targets for developing the biological markers for the diagnosis of AD. In order to develop efficient in vivo probes, polymeric n-butyl-2-cyanoacrylate (PBCA) NPs have been prepared and encapsulated with the radio-labeled amyloid affinity drug (125)I-clioquinol (CQ, 5-chloro-7-iodo-8-hydroxyquinoline) to improve the transport to brain and amyloid plaque retention of (125)I-CQ using the NPs of PBCA. The (125)I-CQ discriminately binds to the AD post-mortem brain tissue homogenates versus control. (125)I-CQ-PBCA NPs labeled the A beta plaques from the AD human post-mortem frontal cortical sections on paraffin-fixed slides. Storage phosphor imaging verified preferential uptake by AD brain sections compared to cortical control sections. The (125)I-CQ-PBCA NPs crossed the BBB in wild type mouse, giving an increased brain uptake measured in terms of % ID/g i.e., injected dose compared to (125)I-CQ. Brain retention of (125)I-CQ-PBCA NPs was significantly increased in the AD transgenic mice (APP/PS1) and in mice injected with aggregated A beta 42 peptide versus age-matched wild type controls. The results of this study are verified by in vivo storage phosphor imaging and validated by histopathological staining of plaques and select metal ions, viz. Fe(2+) and Cu(2+). The (125)I-CQ-PBCA NPs had more efficient brain entry and rapid clearance in normal mice and enhanced the retention in AD mouse brain demonstrating the ideal in vivo imaging characteristics. The (125)I-CQ-PBCA NPs exhibited specificity for A beta plaques both in vitro and in vivo. This combination offered radio-iodinated CQ-PBCA NPs as the promising delivery vehicle for in vivo single photon emission tomography (SPECT) ((123)I) or PET ((124)I) amyloid imaging agent. The importance of the topic in relation to brain delivery and other similar type of work published in this area are covered to highlight the importance of this research to medical disciplines.

The first step into the brain: uptake of NIO-PBCA nanoparticles by endothelial cells in vitro and in vivo, and direct evidence for their blood-brain barrier permeation.

By using fluorescent polysorbate 80 coated poly(n-butylcyanoacrylate) (PBCA) nanoparticles in an in vivo study, direct evidence was found for the presence of nanoparticles entering the brain and retina of rats. The nanoparticles, prepared with a miniemulsion process, were labeled in situ with a fluorescent dye and coated with polysorbate 80. After preparation the particle size, zeta potential, and the molecular weight distribution were determined. BMEC cells were used as an in vitro model for the BBB. The cells showed significant uptake of the particles, but no transcytosis could be observed in vitro. After applying the particles to the animals at two concentrations, cryosections of the brains and retinas were prepared. Regarding the sections of the rats that received the lower dose, co-localization of the applied fluorescent particles and the stained endothelial cells could be detected in the brain and retina, indicating particle internalization in the endothelial cells. Applied at higher doses, the particles could be detected within the brain and retina with few co-localized signals, suggesting passage through the blood-brain and blood-retina barriers.

Delivery of nanoparticles to the brain detected by fluorescence microscopy.

This study aimed to explore and extend the application potential of poly(n-butylcyano-acrylate) (PBCA) nanoparticles to cross the blood-brain barrier (BBB) and to deliver their content into the central nervous system. PBCA particles were prepared by a new and efficient mini-emulsion method with excellent yield and reproducibility. These nanoparticles were loaded with 1.5% (w/v) fluorescein-isothio-cyanate-dextran (FITC-dextran), 1.5% rhodamine-123 or 7.3% doxorubicin. Particles were characterized by dynamic light scattering determining particle size, polydispersity index and zeta-potential. They were coated with 10% w/v polysorbate 80 and administered to rats. Cryosections of the brain were prepared and time-dependent distribution of fluorescence was studied. After the administration of polysorbate 80-coated particles by carotic injection, fluorescence could first be detected in capillary lumina with a progressive shift to capillary endothelial cells at 30min and a rather evenly spread distribution across the brain tissue at 60min after administration. 60min after administration into the tail vein, fluorescent particles could be assigned to endothelial cells, whereas after 2h a rather evenly spread distribution across the brain tissue was seen. These observations indicate that surface-coated PBCA nanoparticles are able to cross the blood-brain barrier and to serve as a drug-delivery system to the central nervous system.

Cellular uptake behavior of unfunctionalized and functionalized PBCA particles prepared in a miniemulsion.

Fluorescent dye labeled unfunctionalized and functionalized poly(n-butylcyanoacrylate) nanoparticles were prepared using a miniemulsion technique. Amino acid and methoxyPEG functionalization could be introduced by using aqueous solutions as an initiator for the anionic polymerization in the heterophase. All the particles prepared had sizes smaller than 250 nm and negative zeta-potentials. The molar mass distribution of the polymer was dependent on the acid used as the continuous phase and the initiator solution applied. Cells of three lines (HeLa, Jurkat and mesenchymal stem cells) were incubated with the particles. The molar mass of the polymer determined the onset and extent of apoptosis, and the total uptake was determined by the size and functionalization of the particles. Different uptake kinetics were obtained with HeLa and Jurkat cells after incubation with the same particle batch. The intracellular particle distribution, visualized by confocal laser scanning microscopy, did not show significant differences for either of the cell lines or particle batches.

Biochemical and histopathological findings of N-butyl-2-cyanoacrylate in oral surgery: an experimental study.

OBJECTIVES: The increasing use of cyanoacrylates in dentistry, particularly as an adhesive and sealing glue, has raised concerns regarding its potential toxicity in humans. Several different forms of these compounds including methyl- (MCA), ethyl- (ECA), isobutyl-, isohexyl-, and octyl CA have been developed to eliminate tissue toxicity. N-butyl-2-cyanoacrylate is becoming an increasingly popular method for wound closure under low tension. Despite their increasing use, pharmacologic effects of these substances on liver and kidney functions are not widely known. The objective of the present study was to investigate possible immediate and long-term systemic effects of N-butyl-2-cyanoacrylate in oral surgery. STUDY DESIGN: Ten male Wistar rats weighing 220 to 270 g were used in the study. Straight incisions were made to the buccal mucosa of the animals. N-butyl-2-cyanoacrylate adhesive (Indermil) was applied and wounds were closed primarily. Blood specimens were taken periodically from the vena cava of the animals before the surgical procedure and 2, 14, 21, and 65 days after the surgical procedure. The blood specimens of those taken before the application of the adhesive were defined as the control group; blood specimens that were taken 2, 14, 21, and 65 days from the application were defined as study group. The stored plasma samples were analyzed for blood urea nitrogen (BUN), creatinine (CRE), alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBI), total protein (TP), albumin (ALB), and amylase (AML). In addition to biochemical parameters, histopathological examination was performed. Blood parameter values of the control and study groups were statistically compared with the Duncan test (P < .05). RESULTS: There were no significant differences in the values of BUN, CRE, ALT, AST, TBI, TP, ALB, and AML between the control and at 2, 14, 21, and 65 days. CONCLUSION: The present study shows that N-butyl-2-cyanoacrylate is a suitable adhesive applicable in oral surgery.

[Effect of diameter of adriamycin polybutylcyanoacrylate nanoparticles on liver targeting].

OBJECTIVE: To observe the different tissue distributions of the adriamycin polybutylcyanoacrylate nanoparticle (ADM-PBCA-NP) in the mice body after the injection via lateral tail vein, and to study the liver targeting effects of ADM-PBCA-NP in different diameters on normal mice livers. METHODS: One hundred and eighty mice were randomly divided into 6 groups with 30 mice in each group: non-conjugated free ADM (Group 1); (22.3+/-6.2) nm in diameter ADM-PBCA-NP group (Group 2); (48.6+/-9.2) nm ADM-PBCA-NP group (Group 3); (101.9+/-20.3) nm ADM-PBCA-NP group (Group 4); (143.5+/-23.5) nm ADM-PBCA-NP group (Group 5), and (194.2+/-28.4) nm ADM-PBCA-NP group (Group 6). A single dose of either conjugated or free adriamycin equaled 2 mg/kg of body weight was delivered via the tail vein. Five mice in each trail were sacrificed at 5, 15, 30 minutes, 1, 5 and 12 hours after the injection, respectively. The adriamycin concentrations in the collected livers, kidneys, spleens, hearts, lungs and plasma were demonstrated using a high performance liquid chromatography with fluorescence detector. RESULTS: Compared with that of the control group, adriamycin was hardly detected in the heart muscles of the treatment groups (P<0.05). The nanoparticle-conjugated adriamycin was cleaned up quickly from the kidney tissues. The adriamycin concentrations of the mice liver and spleen in the experimental groups was significantly higher than those in the control group, except for the group with the nanoparticles diameters of (22.3+/-6.2) nm (P<0.05). The ADM-PBCA-NP in (101.9+/-20.3) nm diameter had the highest liver distribution, and the second highest adriamycin distribution in the liver was the group of (143.5+/-23.5) nm diameter (P<0.05). Adriamycin was released slowly in the liver during the detection period in the experimental groups. ADM-PBCA-NP in (22.3+/-6.2) nm diameter was not distributed in the tissues of the livers, kidneys, hearts, spleens, and lungs. CONCLUSION: ADM-PBCA-NP with a 100 - 150 nm diameter range has the best liver targeting with slow medicine release. It also decreases the medicine distribution in the heart and other organs. In the treatment of liver cancer, the polybutylcyanoacrylate nanoparticle system has a good liver targeting ability, which increases the anticancer activity and markedly decreases the toxicity of adriamycin.

Polybutylcyanoacrylate nanoparticles as a carrier for mitomycin C in rabbits bearing VX2-liver tumor.

Polybutylcyanoacrylate nanoparticles (PBCA-NP) were prepared by addition of the monomer to an aqueous phase containing dextran 70 and loaded with mitomycin C (MMC-PBCA-NP), a highly effective anticancer drug. When injected into mice, MMC-PBCA-NP accumulated more in the liver than did free MMC. In contrast, MMC-PBCA-NP accumulated less in the heart and kidney than did free MMC. However, when MMC-PBCA-NP and MMC were administered to rabbits bearing VX2 cells implanted into the liver, the antiproliferative effects on the tumor cells of both drugs were similar. Histological analyses indicated that organization of myocardial filaments was disrupted and vacuolization was observed in the MMC treated group, whereas MMC-PBCA-NP treatment did not appear to damage the host's heart. Hydropic degeneration of renal tubular epithelia was observed in the MMC treated group and not in the control group, whereas MMC-PBCA-NP treatment did not appear to damage the host's kidney.

The in vitro stability of air-filled polybutylcyanoacrylate microparticles.

Different methods of manufacturing permitted the production of air-filled PBCA microparticles (af-pbca-mp) with different physical properties such as size and wall thickness. These differences led to distinctions with respect to mechanical stability and, at the same time, to different levels of biochemical stability when incubated in biofluids. Microparticles, designed as they are to be mechanically more stable (composed of larger nanoparticles resulting in thicker shell wall, no surface hydrolysis), persist longer under in vitro conditions in biofluids such as serum, plasma and whole blood than do the more fragile ones. It was possible when using the measurement of ultrasound attenuation to characterize af-pbca-mp degradation with respect to the disappearance of the ultrasound properties of the particles and therefore to find out how long different formulations can be expected to be active as contrast agents under simulated in vivo conditions. The present examination showed that using either serum, plasma or whole blood leads to results with the same tendencies in terms of the stability and durability of af-pbca-mp in the media, mimicking in vivo conditions. It was thus possible to validate successfully the use of either serum or plasma as substitutes for whole blood. Further studies dealing with the in vitro in vivo correlation will be needed to find out if the situation in this in vitro assay corresponds to the situation in the body.

Acute and chronic swine rete arteriovenous malformation models: effect of ethiodol and glacial acetic acid on penetration, dispersion, and injection force of N-butyl 2-cyanoacrylate.

BACKGROUND AND PURPOSE: Liquid embolic agents are increasingly gaining importance in the embolization of cerebral arteriovenous malformations (AVMs). Currently, the most commonly used agent is N-butyl 2-cyanoacrylate (NBCA). Various NBCA mixtures, arterial hypotension, and Valsalva maneuver (increased positive end-expiratory pressure) during the injection of the acrylate have been used to address hemodynamic and architectural variations of an AVM; however, the precise in vivo polymerization, distribution, and kinetics of NBCA mixtures are unknown. We investigated the effect of different acrylate/Lipiodol mixtures and the addition of glacial acetic acid (GAA) on the penetration, dispersion, and injection force of NBCA. METHODS: A swine rete AVM model that has been described elsewhere was used for the embolization. In one subgroup of animals, embolization was performed immediately after construction of the AVM model. In a second subgroup, a chronic AVM model was used. GAA was added to the NBCA mixture to decrease the pH value of the solution and prolong the polymerization time. The addition of GAA allowed us to reduce the amount of Lipiodol, thereby reducing the viscosity of the mixture. A total of 30 swine were used for both the acute (n = 23) and chronic (n = 7) subgroups. The following mixtures of Lipiodol/NBCA and GAA (% vol/%vol + microL) were used for embolization: 80/20 + 0; 50/50 + 0; 50/50 + 5; 50/50 + 10; and 50/50 + 20. A total of six retia per mixture were used for the analysis. Glue injection pressure profiles were recorded in each experiment. High-resolution radiographic images obtained from the harvested retia were used to correlate the dispersion and depth of glue penetration with the AVM hemodynamics. The effect of different amounts of GAA on the glue dispersion and depth of penetration of the mixtures was also studied. RESULTS: Using the same pressure gradients, less viscous NBCA + GAA mixtures led to a deeper nidal penetration. The addition of 20 microL of GAA resulted in a three times higher penetration and dispersion of the NBCA mixture that was more homogenous. CONCLUSION: The viscosity of the liquid embolic agent used is an important limiting factor for an AVM embolization. Reducing the amount of Lipiodol improves nidus penetration. Quicker polymerization can be overcome by adding GAA, which reduces the pH of the mixture.

Body distribution of polysorbate-80 and doxorubicin-loaded [14C]poly(butyl cyanoacrylate) nanoparticles after i.v. administration in rats.

Previously it was shown that poly(butyl cyanoacrylate) (PBCA) nanoparticles coated with polysorbate 80 are able to cross the blood-brain barrier (BBB) after i.v. administration. The objective of the present study was to investigate the influence of polysorbate 80 and doxorubicin-loading on the body distribution in rats. The biodistribution profile and brain concentration of (14)C-radiolabeled PBCA nanoparticles, polysorbate 80 coated (14)C-PBCA nanoparticles, and doxorubicin-loaded (14)C-PBCA nanoparticles were determined by radioactivity counting after i.v. administration in rats. The (14)C-PBCA nanoparticles showed a significant accumulation in the organs of the reticuloendothelial system (RES). Polysorbate 80 coating of the (14)C-PBCA nanoparticles decreased this accumulation to about 40% after 1 h post injection. The brain concentration was increased about 2-fold after polysorbate 80-coating at this time point. The presence of doxorubicin in this preparation, however, decreased the brain concentration to levels similar to uncoated particles, probably caused by the positive charge of this compound. After longer time periods after injection the differences between the three preparations decreased.

[Antiamnesic activity of nerve growth factor adsorbed on poly(butyl) cyanoacrylate nanoparticles coated with polysorbate-80].

The antiamnesic activity of nerve growth factor (NGF) in various medicinal forms [aqueous NGF solution with and without polysorbate-80 (PS-80) additives, NGF adsorbed on poly(butyl) cyanoacrylate (PBCA) nanoparticles with and without PS-80 coating] has been studied in rats with model amnesia induced by scopolamine (2 mg/kg, s.c.). The antiamnesic activity was evaluated by the ability of subsequently introduced drugs (5 microg NGF per animal, i.p.) to inhibit the amnesic action of scopolamine as manifested by retrieval of the passive avoidance reflex (PAR) learned using the Lafayette Instruments USP system 30 min after drug injection. The PAR memory trace was evaluated as an increase in the latent time before visiting the dark compartment 24 h after drug injection. In the untreated amnesia control group, the scopolamine amnesia was manifested by the absence of any increase in the PAR latent time. In the form of an aqueous solution, NGF did not inhibit the scopolamine-induced amnesia. NGF adsorbed on uncoated PBCA and in the form of solution with PS-80 produced an antiamnesic action manifested by a reliable increase in the PAR latent time as compared to that in the untreated control group. NGF adsorbed on PBCA nanoparticles coated with PS-80 not only exhibited a significant antiamnesic effect, but even stimulated the cognitive function and increased the PAR latent time above the value in the learned control group.

Improved synthesis, antibacterial activity and potential carcinogenicity of 5-amino-1 ,2,4-thiadiazol-3(2H)-one.

This paper reports on the preparation of 5-amino-1,2,4-thiadiazol-3(2H)-one, a sulfur-containing analogue of cytosine with the -CH=CH- group between the positions 5 and 6 of the pyrimidine ring replaced by the divalent sulfur (-S-). Improved procedures for the preparation of thiobiuret, some of its methyl derivatives and 5-amino-1,2,4-thiadiazol-3(2H)-one are documented. Thiobiuret and its N-methyl derivatives were obtained by addition of hydrogen sulfide to the respective 1-cyanoureas. Subsequent oxidation of thiobiuret with hydrogen peroxide in alkaline medium produced 5-amino-1,2,4-thiadiazol-3(2H)-one. This substance was traced back converted to the starting thiobiuret by reaction with cysteine hydrochloride. Alkaline degradation of thiadiazol led to the formation of 1-cyanourea isolated as its silver salt. An investigation of the thiadiazol biological activities has shown that it inhibits the growth of E. coil by 10% at 8.5 microM concentrations, but exhibited no cytostatic activity in L1210, HeLa S3 and HL-60 cell lines. Potential carcinogenicity of the prepared compounds was determined by a DC polarographic method. While the values of the parameter of carcinogenicity for all intermediates indicate only marginal carcinogenic potential, the value of the parameter of carcinogenicity for the thiadiazole indicates possible carcinogenicity of this compound.

Influence of polymerization technique and experimental variables on the particle properties and release kinetics of methotrexate from poly(butylcyanoacrylate) nanoparticles.

Poly(butylcyanoacrylate) nanoparticles were prepared by dispersion polymerization (DP) and emulsion polymerization (EP) of n-butyl cyanoacrylate monomer. The particles were characterized by infrared spectroscopy, differential scanning calorimetry, X-ray diffractometry and transmission electron microscopy. Particle properties such as size and zeta potential were determined for nanoparticles prepared by DP and EP techniques and compared. EP technique resulted in a low particle size compared to the DP. A high zeta potential was observed for nanoparticles prepared by the DP method. Incorporation of methotrexate resulted in a decrease in zeta potential in both types of nanoparticles, the decrease being greater in DP nanoparticles. Effect of experimental variables such as monomer concentration, polymerization time and temperature on drug entrapment and particle size was studied. Both types of nanoparticles showed an increase in drug entrapment with increased monomer concentrations. Variable polymerization time did not influence the drug entrapment of EP nanoparticles. Polymerization at 60 +/- 2 degrees C resulted in a decrease of drug entrapment and a great increase in the particle size of both types of nanoparticles. In vitro drug release studies showed a comparatively high release of methotrexate from DP nanoparticles suggesting the channelizing effect of dextran chains incorporated into nanoparticles during polymerization. Though the release profiles of nanoparticles appeared similar, a significant difference in release rates was found for DP and EP nanoparticles in 0.1 mol L(-1) HCl and pH 7.4 phosphate buffer (p < 0.01). Drug release data indicate that the release of methotrexate from DP and EP nanoparticles followed Fickian diffusion in 0.1 mol L(-1) HCl, while the mechanism was found anomalous in pH 7.4 phosphate buffer. An effort was also made to critically correlate the properties of nanoparticles synthesized by the above two techniques, and emphasize the importance of these characteristics in targeted drug delivery.

Physicochemical characterisation of cationic polybutylcyanoacrylate-nanoparticles by fluorescence correlation spectroscopy.

The aim of this study was to compare different physical and chemical methods with fluorescence correlation spectroscopy (FCS) in order to characterise cationic acrylate nanoparticles (NP), which can deliver oligonucleotides (ON) into mammalian cells. These positively charged nanoparticles were prepared from diethylaminoethyl dextran (DEAE-dextran) and poly(n-butyl-2-cyanoacrylate) (PBCA). NP consists of PBCA oligochains with an average size of PBCA 9 mer and were formed by entrapping DEAE-dextran and dextran 70,000 in high amounts into the particle matrix. The oligochain length of PBCA was investigated by mass-spectroscopy (MALDI TOF). The molecular weight of a particle with d = 108 nm was estimated to be approximately 3.6 x 10(8) Da. The mean size of the nanoparticles were in a range of dh = 130-140 nm, as determined independently by FCS and dynamic light scattering. Atomic force microscopy and scanning electron microscopy images confirm this size range. Furthermore, the particle mass of the PBCA-NP was estimated by FCS measurements. For this approach two new methods for fluorescence labelling of cationic particles were developed. Fluorescent labelled dextran 70,000 was entrapped into the particle matrix; in addition, the derivatisation of hydroxyl groups of the NP was achieved with 5-([4,6-dichlorotriazin-2-yl]amino) fluorescein (DTAF). ON can be localised in a complex with the NP by dual-colour fluorescence cross correlation spectroscopy measurements. The zetapotential of the unloaded NP was positively charged with about +39 mV and decreased down to -40 mV on addition of excess ON. After centrifugation quantification of the ON loading onto the particles by strong anion exchange high performance liquid chromatography (SAX HPLC) and FCS showed that approximately 20 microg ON per 100 g NP was adsorbed. The FCS measurements of the ON adsorption in situ was found to be much higher with approximately 95 microg ON per 100 g NP.

[Human recombinant interferon-alpha 2a polybutylcyanoacrylate sustained release lyophilized nanospheres for liver-targeting].

OBJECTIVE: To evaluate the liver-targeted and sustained release characteristics of human recombinant interferan-alpha 2a polybutylcyanoacrylate lyophilized nanospheres (rIFN alpha 2a-PBCA-NS). METHODS: Emulsification/polymerization method was used to prepare the nanospheres which was lyophilized to obtain rIFN alpha 2a-PBCA-NS, and cell pathological embarrassment (CPE) was used to determine the rIFN alpha 2a concentration in the preparations and biological samples. The in vivo distribution and pharmacokinetics in liver after intravenous injection (i.v.) of rIFN alpha 2a-PBCA-NS and rIFN alpha 2a in mice were evaluated. RESULTS: The rate of entrapment of the nanospheres was (56.2 +/- 4.3)%; the particle diameter, span of rIFN alpha 2a-PBCA-NS were (108 +/- 37) nm and 0.55, respectively, and their biological activity had little change after being stored at 3-5 degrees C for 6 months. The distribution ratio of rIFN alpha 2a after i.v. in mice liver increased from 13.1% (for rIFN alpha 2a) to 50.6% (for rIFN alpha 2a-PBCA-NS), and the mean retain time from 1.41 h (for rIFN alpha 2a) to 8.35 h (for rIFN alpha 2a-PBCA-NS). Of the distribution parameters (te for rIFN alpha 2a, re and te for rIFN alpha 2a-PBCA-NS) in 5 main organs, the largest ones were in liver and both re and te for rIFN alpha 2a-PBCA-NS in liver larger than one. CONCLUSION: The rIFN alpha 2a-PBCA-NS showed obviously liver-targeting and sustained release characteristics.