Magnesium sulphate has beneficial effects as an adjuvant during general anaesthesia for Caesarean section.

BACKGROUND: The use of low concentrations of volatile anaesthetics with avoidance of opioids may induce intraoperative awareness and adverse haemodynamic responses during Caesarean section. Magnesium is well known to reduce anaesthetic requirements and to block noxious stimuli. We investigated whether i.v. magnesium sulphate modulates anaesthetic depth and analgesic efficacy during Caesarean section. METHODS: Seventy-two patients undergoing Caesarean section were randomly assigned to receive i.v. saline (control group) or magnesium sulphate 30 mg kg(-1) bolus+10 mg kg(-1) h(-1) continuous infusion (Mg 30 group) or 45 mg kg(-1) bolus+15 mg kg(-1) h(-1) continuous infusion (Mg 45 group) after induction. Bispectral index (BIS) value, mean arterial pressure (MAP), and midazolam, fentanyl, and atracurium consumptions were recorded. RESULTS: BIS values [mean (sd)] at 7.5 and 10 min after surgery and before delivery in the control [64 (9), 66 (8), 67 (8), P<0.001] and the Mg 30 groups [62 (8), P<0.01; 64 (7), 63 (9), P<0.001] were higher than in the Mg 45 group [56 (8), 55 (8), 55 (7)]. MAP was greater in the control group (P<0.05) than in the Mg 30 and Mg 45 groups during the pre-delivery period. The magnesium groups required less midazolam (P<0.05), fentanyl (Mg 30, P<0.05; Mg 45, P<0.01), and atracurium (P<0.001) vs the control group. CONCLUSIONS: Preoperative i.v. magnesium sulphate attenuated BIS and arterial pressure increases during the pre-delivery period. Magnesium sulphate can be recommended as an adjuvant during general anaesthesia for Caesarean section to avoid perioperative awareness and pressor response resulting from inadequate anaesthesia, analgesia, or both.

Self-assembled "nanocubicle" as a carrier for peroral insulin delivery.

AIMS/HYPOTHESIS: A patient with (insulin-dependent) diabetes mellitus receives at least one subcutaneous insulin injection a day to maintain low serum glucose concentrations. Since patients' compliance with such dosage regimens is too low, the development of an oral formula is clearly attractive. We present the development of a liquid formula that can be easily dispersed in water to produce particles named "nanocubicles" which efficiently encapsulate insulin. METHODS: Fasted streptozotocin-induced diabetic rats were administered orally with particles encapsulating insulin, and particles without insulin or soluble insulin in water. Groups of rats were also injected soluble insulin in PBS for control. Blood glucose concentration and insulin concentration were measured 1, 2, 3, 4 and 6 h after the administration of the insulin formulas. RESULTS: In vitro experiments show that the particles can be taken up by the Caco-2 cells at a high ratio. The serum glucose concentration was controlled for more than 6 h after oral insulin administration but returned to the basal concentration in 3 h when 1 IU/kg of insulin was injected intravenously. CONCLUSION/INTERPRETATION: Our biocompatible and stable oral insulin formulation is easy to prepare and produces reproducible hypoglycaemic effects, therefore we anticipate clinical acceptance and utilization of this form of insulin therapy.

Structural characteristics of size-controlled self-aggregates of deoxycholic acid-modified chitosan and their application as a DNA delivery carrier.

Precise control of the size and structure is one critical design parameter of micellar systems for drug delivery applications. To control the size of self-aggregates, chitosan was depolymerized with various amounts of sodium nitrite, and hydrophobically modified with deoxycholic acid to form self-aggregates in aqueous media. Formation and physicochemical characteristics of size-controlled self-aggregates were investigated using dynamic light scattering, fluorescence spectroscopy, and computer simulation method. The size of self-aggregates varied in the range of 130-300 nm in diameter, and their structures were found to depend strongly on the molecular weight of chitosan ranging from 5 to 200 kDa. Due to the chain rigidity of chitosan molecule, the structure of self-aggregates was suggested to be a cylindrical bamboolike structure when the molecular weight of chitosan was larger than 40 kDa, which might form a very poor spherical form of a birdnestlike structure. To explore the potential applications of self-aggregates as a gene delivery carrier, complexes between chitosan self-aggregates and plasmid DNA were prepared and confirmed by measuring the fluorescence intensity of ethidium bromide and electrophoresis on agarose gels. The complex formation had strong dependency on the size and structure of chitosan self-aggregates and significantly influenced the transfection efficiency of COS-1 cells (up to a factor of 10). This approach to control the size and structure of chitosan-derived self-aggregates may find a wide range of applications in gene delivery as well as general drug delivery applications.

PDMS-based polyurethanes with MPEG grafts: mechanical properties, bacterial repellency, and release behavior of rifampicin.

PDMS-based polyurethanes (PUs) grafted with monomethoxy poly(ethylene glycol) (MPEG) were synthesized to develop a coating material for urinary catheters with a silicone surface for minimizing urinary tract infections. MPEG was grafted on PDMS-based PUs by two methods depending on the PU synthetic routes: esterification and allophanate reactions. It was confirmed from mechanical characterization that an increase of the hard segment amount enhanced the ultimate strength and Young's modulus, while reducing elongation at the end-points. The incorporation of MPEG in PDMS-based PUs induced a decrease in tensile strength and Young's modulus, and increased elongation at the break point due to its high flexibility. When hydrated in distilled water, mechanical properties of all PUs synthesized in this study deteriorated due to water absorption. It was evident from the bacterial adhesion test that PDMS-based PUs showed moderate resistance to adhesion of E. coli on their surfaces compared to Pellethane, while the incorporation of MPEG significantly enhanced repellency to bacteria, including E. coli and S. epidermidis. We also studied the release behavior of an antibiotic drug, rifampicin, from the polymeric devices fabricated by solvent evaporation. Although rifampicin is hydrophilic and soluble in pH 7.4 phosphate buffer, it showed a sustained release over 45 days from PDMS-based PUs with MPEG that were grafted on ethylene glycol residues by allophanate reaction. This release characteristic was predominantly influenced by a hydrogen bond interaction between the polymers and rifampicin, which was confirmed through an ATR-IR study. This may imply that the specific interaction is responsible for the delayed release. Considering the mechanical properties, morphologies of drug-incorporated polymeric matrices, and drug release behaviors, PDMS-based PU with MPEG that were grafted on ethylene glycol (a chain extender) residues by allophanate reaction showed better material properties for uretharal catheter coating pusposes in order to minimize urinary tract infections.

Optimization of lipid composition in cationic emulsion as in vitro and in vivo transfection agents.

PURPOSE: To enhance in vitro and in vivo transfection activity by optimizing lipid composition of cationic lipid emulsions. METHODS: Various emulsion formulations having different cationic lipids as emulsifiers, and additional helper lipids as co-emulsifiers, were prepared. The stability of the emulsion and its complex with DNA was investigated by measuring the particle size change in phosphate buffer saline (PBS) over a period of 20 days. The activity of the emulsions in transfecting pCMV-beta into COS-1 cells in the presence or absence of 80% serum was evaluated. We also evaluated in vivo transfection activity using intravenously administered pCMV-Luc+ as a reporter gene. RESULTS: Among the cationic emulsifiers, 1,2-dioleoyl-sn-glycero-3-trimethylammonium-propane (DOTAP) formed the most stable and efficient emulsion gene carrier. Addition of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) increased in vitro transfection activity, but slightly compromised the stability of the emulsion. The loss was compensated for by including small amounts of Tween 80 in the emulsion. The in vitro and in vivo transfection activities were also increased by adding Tween 80. Even though in vitro transfection activity of liposomes was high in the absence of serum, the transfection activity of emulsions was far greater than that of liposomes in the presence of serum and for in vivo applications. CONCLUSIONS: By including DOPE as an endosomolytic agent and Tween 80 as a stabilization agent, the cationic emulsion becomes a more potent gene carrier for in vitro and in vivo applications, especially in the presence of serum.

Oil components modulate physical characteristics and function of the natural oil emulsions as drug or gene delivery system.

Oil-in-water (o/w) type lipid emulsions were formulated by using 18 different natural oils and egg phosphatidylcholine (egg PC) to investigate how emulsion particle size and stability change with different oils. Cottonseed, linseed and evening primrose oils formed emulsions with very large and unstable particles. Squalene, light mineral oil and jojoba bean oil formed stable emulsions with small particles. The remaining natural oils formed moderately stable emulsions. Emulsions with smaller initial particle size were more stable than those with larger particles. The correlation between emulsion size made with different oils and two physical properties of the oils was also investigated. The o/w interfacial tension and particle size of the emulsion were inversely proportional. The effect of viscosity was less pronounced. To study how the oil component in the emulsion modulates the in vitro release characteristics of lipophilic drugs, three different emulsions loaded with two different drugs were prepared. Squalene, soybean oil and linseed oil emulsions represented the most, medium and the least stable systems, respectively. For the lipophilic drugs, release was the slowest from the most stable squalene emulsion, followed by soybean oil and then by linseed oil emulsions. Cationic emulsions were also prepared with the above three different oils as gene carriers. In vitro transfection activity was the highest for the most stable squalene emulsion followed by soybean oil and then by linseed oil emulsions. Even though the in vitro transfection activity of emulsions were lower than the liposome in the absence of serum, the activity of squalene emulsion, for instance, was ca. 30 times higher than that of liposome in the presence of 80% (v/v) serum. In conclusion, the choice of oil component in o/w emulsion is important in formulating emulsion-based drug or gene delivery systems.

Prophylactic efficacy of a new gentamicin-releasing urethral catheter in short-term catheterized rabbits.

OBJECTIVE: To describe an indwelling urethral catheter coated with gentamicin sulphate on the inner and outer surface of the catheter, and to evaluate the efficacy and safety of this catheter in preventing catheter-associated infections in rabbits. Materials and methods Sixty rabbits were divided equally into control and experimental groups which were then subdivided equally according to the duration of catheterization (1, 3 and 5 days). Silicone-treated latex catheters were used in the control group and gentamicin-releasing catheters in the experimental group. Urine samples and surface swabs from the catheter were cultured for bacteriological assessment, and the catheter surface examined by scanning electron microscopy to structurally analyse the biofilms. RESULTS: The gentamicin-releasing catheter reduced the incidence of bacteriuria (defined as > or = 100 c.f.u./mL) after both 3 and 5 days of catheterization (eight and 10 rabbits, respectively, for the control catheter, vs two and four rabbits for the gentamicin-releasing catheter, P < 0.05). The surfaces of the gentamicin-releasing catheter were colonized less often than those of the control catheter after both 3 and 5 days (eight and 10, respectively, for the control, vs one and four for the gentamicin-releasing catheter, P < 0.05). Scanning electron microscopy showed the formation of bacterial biofilm throughout the 3-day and 5-day control catheters, but deterioration of the bacterial biofilm was visible on the surface of the gentamicin-releasing catheters. CONCLUSION: This new gentamicin-releasing catheter produced an antibacterial barrier which inhibited catheter-associated urinary tract infection with no toxicity for at least 5 days. These in vivo studies suggest that this new catheter may be useful for controlling infection, with systemic and local safety, in patients undergoing short-term indwelling urethral catheterization.

In vivo gene transfer to the mouse nasal cavity mucosa using a stable cationic lipid emulsion.

We evaluate a new cationic emulsion as a mucosal gene carrier and elucidate the relationship between the transfection efficiency and the stability of the carrier/DNA complex. A cationic lipid emulsion was formulated with soybean oil and 1,2-dioleoyl-sn-glycero-3-trimethylammonium-propane (DOTAP) as major components and was used to transfer genes to the epithelial cells of the mouse nasal cavity via intranasal instillation. Correlation between the transfection efficiency and the stability of the carrier/DNA complex was investigated by measuring the carrier size changes and by observing the degree of DNA protection against DNase I digestion in the presence of heparin. The cationic emulsion showed at least 3 times better transfection activity than the liposomal carriers in nasal mucosae. The cationic emulsion was stable in the presence of heparin whereas the liposomal carriers became very unstable. Unlike DNA in liposome/DNA complexes, DNA in the emulsion/DNA complex was resistant to heparin exchange and DNase I digestion. The cationic emulsion was more effective in delivering DNA to nasal mucosae than commercially available liposomal carriers. The transfection activities of the lipid carriers in nasal cavity mucosae are in agreement with the stability of the lipid carriers and their complexes with DNA.

A cationic lipid emulsion/DNA complex as a physically stable and serum-resistant gene delivery system.

PURPOSE: To develop a non-viral gene delivery system in the form of an oil-in-water (o/w) lipid emulsion. METHOD: Cationic lipid emulsions were formulated with soybean oil, 1,2-dioleoyl-sn-glycero-3-trimethylammonium-propane (DOTAP) as a cationic emulsifier and other co-emulsifiers. The physical characteristics of the lipid emulsion and the emulsion/DNA complex were determined. The in vitro transfection efficiency of the emulsion/DNA complex was determined in the presence of up to 90% serum. RESULTS: The average droplet size and zeta potential of emulsions were ca. 180 nm and ca. +50 mV, respectively. Among the emulsions, a stable formulation was selected to form a complex with a plasmid DNA encoding chloramphenicol acetyltransferase. By increasing the ratio of emulsion to DNA. zeta-potential of the emulsion/DNA complex increased monotonously from negative to positive without any changes in the complex size. The complex was stable against DNase I digestion and an anionic poly-L-aspartic acid (PLAA). The complex delivered DNA into the cells successfully, and the transfection efficiency was not affected by complex formation time from 20 min to 2 h. More importantly, the cationic lipid emulsion facilitated the transfer of DNA in the presence of up to 90% serum. CONCLUSIONS: The cationic lipid emulsion/DNA complex has physical stability and serum resistant properties for gene transfer.

Protective immunity of microsphere-based mucosal vaccines against lethal intranasal challenge with Streptococcus pneumoniae.

Mucosal vaccination of capsular polysaccharide (PS) of Streptococcus pneumoniae and subsequent creation of the first line of immunological defense in mucosa were examined. Mucosal as well as systemic antibody responses to PS were evoked by peroral or intranasal immunization of BALB/c mice with PS-cholera toxin B subunit (CTB) conjugates entrapped in the alginate microspheres (AM). The bacterial colonization at the lung mucosa was most profoundly inhibited (<95%) by intranasal immunization with the naked conjugate (PS-CTB). The mice vaccinated orally with encapsulated conjugate [AM(PS-CTB)] showed significant reduction on the level of pneumococcal bacteremia (<99%). Eighty percent of the mice perorally immunized with AM (PS-CTB) were protected from lethal intranasal challenge with S. pneumoniae, whereas more than 60% of the mice in the other control groups died of infection. Our novel approach may prove to be important in the development of a mucosal vaccine that will provide protection of mucosal surfaces of host.

Peroral immunization of microencapsulated human VP8 in combination with cholera toxin induces intestinal antibody responses.

To develop an orally delivered subunit vaccine for rotavirus infection, a trypsin cleavage product of VP4, recombinant VP8*, was expressed in Escherichia coli. The recombinant VP8* (rVP8*), purified by affinity chromatography, was reactive against human rotavirus positive serum in Western-blot analysis. To further evaluate the immunogenicity of the oral-delivered rVP8*, it was encapsulated with alginate-microsphere and administered in combination with cholera toxin (CT) as a mucosal adjuvant perorally into mice. The ELISPOT assay showed that the number of rVP8*-specific IgG1 antibody secreting cells increased about 3-fold and about 2-fold in spleen and Peyer's patch, respectively as compared to non-immune mice. In addition, the number of rVP8*-specific IgA antibody secreting cells increased about 2-fold in Peyer's patch. Finally, rVP8*-specific IgA antibody response was significantly enhanced in the intestinal fluids from the mice immunized perorally with encapsulated rVP8* and CT. Taken together, these results indicate that rVP8* possessed proper immunogenicity and it would be potentially useful as a subunit vaccine against rotavirus-associated disease through peroral immunization.

Novel mucosal immunization with polysaccharide-protein conjugates entrapped in alginate microspheres.

A novel mucosal immunization was examined using biocompatible and biodegradable alginate microspheres containing a conjugate of polysaccharide antigen and cholera toxin B subunit (CTB). In order to prepare the alginate microspheres with diameters of less than 5 microm, a new diffusion-controlled interfacial gelation technique was developed. Also, in order to improve the mucosal immune response, a pneumococcal capsular polysaccharide type 19 (PS19) was conjugated to the CTB (PS19-CTB). This conjugate was subsequently encapsulated into the alginate microspheres. The loading content of PS19-CTB to the alginate microspheres was 60%. An in vitro sustained release pattern was observed with the antigen-loaded microspheres, showing 80% antigen release within one day. Mucosal and systemic immunities following oral immunization with the alginate microspheres were studied. Balb/c mice were immunized perorally three times at intervals of two weeks. Peroral immunization with 25 microg of PS19-CTB entrapped in the alginate microspheres evoked both the mucosal IgA and systemic IgM responses to PS19 in small intestine and in sera, respectively. The results suggest that both the mucosal and systemic antibody responses could be induced by oral administration of the PS19-CTB antigen entrapped in alginate microspheres.

Effect of surface properties on the antithrombogenicity of silk fibroin/S-carboxymethyl kerateine blend films.

Polymeric blends of silk fibroin (SF) and S-carboxymethyl kerateine (SCMK) were prepared by the solvent casting method to study the effect of surface properties on the antithrombogenicity. The films of SF/SCMK showed better antithrombogenic properties than SF or SCMK alone. Among them, the film containing 50 wt% SCMK showed the best antithrombogenicity. When the SF/SCMK films were treated with methanol, the antithrombogenicity of the films was scarcely affected except the SF-rich ones. The enhanced antithrombogenic properties were explained in terms of polarity of the surface. The blend films showed an enhancement of polar contribution to surface free energy (gamma(P)S and polar stabilization energy (I(SW)). SF-rich films showed high gamma(P)S and I(SW) values when treated with methanol. This change of surface properties was considered to be due to the fact that the conformational transition from random coil structure to beta-structure of proteins may have affected the surface properties, especially the polar properties.

Stable lipiodolized emulsions for hepatoma targeting and treatment by transcatheter arterial chemoembolization.

We attempted to develop lipiodolized emulsions that remain in the tumour for a long period, release drug in a sustained release pattern, and thus improve the conventional treatment of hepatocellular carcinoma (HCC) [1]. Polyoxyethylene derivatives of hydrogenated castor oil (HCO) were the most suitable emulsifiers in stabilizing emulsions containing Lipiodol as an oil phase. The length of ethylene oxide coupled to HCO rather than the hydrophilic-lipophilic balance (HLB) values was an important factor in preparing stable emulsions and in achieving sustained-release characteristics. When distilled water was replaced with Iopamiro, a heavy water soluble contrast medium with a specific gravity of 1.335, more stable lipiodolized emulsions with longer sustained release behaviour could be prepared with smaller amount of HCO. To study the in vivo stability of the w/o Lipiodol emulsion and the sustained-release characteristics of doxorubicin from the emulsion, the pharmacokinetic study was performed with normal dogs using transcatheter arterial chemoembolization technique. The area under the plasma concentration-time curve for the first eight hours (AUC0-8) and AUCtotal values of the stabilized emulsion were three to four times higher than those of the coarse emulsion prepared lacking HCO 60. From the in vitro and in vivo studies, Lipiodol based water in oil emulsion with HCO 60 containing doxorubicin showed higher stability and released doxorubicin in a sustained fashion.

Preparation of chitosan self-aggregates as a gene delivery system.

Hydrophobically modified chitosan containing 5.1 deoxycholic acid groups per 100 anhydroglucose units was synthesized by an EDC-mediated coupling reaction. Formation and characteristics of self-aggregates of hydrophobically modified chitosan were studied by fluorescence spectroscopy and dynamic light scattering method. The critical aggregation concentration (cac) of the self-aggregate was determined by measuring the fluorescence intensity of pyrene as a fluorescent probe. The cac value in PBS solution (pH 7.2) was 1.7x10(-2) mg/ml. Mean diameter of self-aggregates in PBS solution (pH 7.2) was 162 +/- 18 nm with an unimodal size distribution. Charge complex formation between self-aggregates and plasmid DNA was confirmed by electrophoresis on an agarose gel. Migration of DNA on an agarose gel was completely retarded above a charge ratio ( +/-) of 4/1 at pH 7.2. The free DNA dissociated from the complexes was observed by electrophoresis above pH 8.0 at a fixed charge ratio of 4/1. An efficient of COS-1 cells was achieved by self-aggregates/DNA complexes.

Electrically erodible polymer gel for controlled release of drugs.

New controlled drug-delivery systems are being explored to overcome the disadvantages of conventional dosage forms. For example, stimulated drug-delivery has been used to overcome the tolerance problems that occur with a constant delivery rate, to mimic the physiological pattern of hormonal concentration and to supply drugs on demand. Stimuli-sensitive polymers, which are potentially useful for pulsed drug delivery, experience changes in either their structure or their chemical properties in response to changes in environmental conditions. Environmental stimuli include temperature, pH, light (ultraviolet or visible), electric field or certain chemicals. Volume changes of stimuli-sensitive gel networks are particularly responsive to external stimuli, but swelling is slow to occur. As well as being useful in the controlled release of drugs, such systems also provide insight into intermolecular interactions. Here we report on a novel polymeric system, which rapidly changes from a solid state to solution in response to small electric currents, by disintegration of the solid polymer complex into two water-soluble polymers. We show that the modulated release of insulin, and by extension other macromolecules, can be achieved with this polymeric system.