الفهرس 
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Abstract
Researchers are developing increasing number of new chemical entities which have promising pharmacological response. Unfortunately, many of these compounds fail to continue the way through the pipelines to reach the market development stage. The main reason of this failure is attributed to erratic dissolution rate and poor solubility of these compounds. In addition, nearly 40% of marketed oral drugs are poorly water soluble .The solubility of the drugs is directly proportional to drug dissolution rate, and so decreasing solubility resulted in obvious decreasing in dissolution rate with subsequent decrease bioavailability and therapeutic efficacy of the drugs. The problem becomes even greater if the solubility and permeability problems are associated with presystemic metabolism.
Various methods have been developed to modify the bioavailability of these drugs by enhancing their dissolution rate, solubility and permeability. These include microionization, solid dispersion technique, inclusion complexation, nanosuspension, hydrotrophy and micellar solubilization. The benefit will become even greater for drugs if the selected carrier and/or delivery systems have the ability to enhance the membrane permeability of drugs.
Recently, lipid based carrier systems have attracted increasing attention to improve the objective of modern drug therapy. Lipid based carrier systems include, microemulsifying systems, nanoemulsions, self-emulsifying formulations (SEF) and self micro/nanoemulsifying systems, liquid crystalline system and vesicular based system such as cubosomes. The interesting criteria of SEF systems include their ability to undergo self dispersion in addition to their tendency for self assembly forming viscous
Abstract 2
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
systems of various physical characteristics ranging from liquid crystalline to gel phase systems. Accordingly the main objective of this thesis was to study the effect of phase transition of SEDDS which occurs upon dilution on the rate of release and/or oral absorption of selected drugs.
The proceeding sections summarize the studies performed to achieve this objective.
1- Self-dispersing self-assembling systems for controlled oral delivery of gliclazide
a. Development of a suitable method for analysis of the drug.
UV spectrophotometric method was employed for determination of gliclazide in different pH, 0.1N HCL and PH 6.8. The quantification was conducted at 229 nm. The method was linear in the range of 6-18 g/ml in both pH. The validation results indicated both inter-day and intraday accuracy and precision. The LOQ was 4.4g/ml at PH 1.2 and was equal to4.8 g/ml at phosphate buffer.
b. Pseudoternary phase diagram.
This diagram was constructed to determine the composition of tested formulations of gliclazide. The phase diagram was constructed using peceol as oily phase to enhance permeation. Gelucire 50/13 with hydrophilic transcutol being adopted as surfactant and co-surfactant, respectively. Two different phase diagrams were constructed by using transcutol and gelucire at different ratio of 1:1 and 1:2 w/w. These phase diagrams show 4 different zones include microemulsion, liquid crystal, gel and coarse emulsion. The area occupied by each zone depended on the relative proportions of Transcutol to Gelucire. Thus increasing the proportions of Gelucire in the
Abstract 3
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
mixture is expected to widen the areas of higher viscosity (LC and gel phases) with a reduction of the liquid zone (coarse emulsion phase)
C. Transmission electron microscopy (TEM) The microemulsion system was subjected for TEM analysis using transmission electron microscopy. Transmission electron micrographs of microemulsion phase revealed the spherical nature of the microemulsion systems. The average droplet size of the ME containing Gelucire and Transcutol (surfactant/co-surfactant) at 1:1 molar ratio was 55+33nm. For systems containing surfactant/co-surfactant mixture at 2:1, the average droplet size was 46+10nm.
d. In vitro drug release study
The dissolution rate of the gliclazide was researched from its unprocessed powder and after incorporation into different formulations. The release of gliclazide was monitored using continuous pH variation method in which the formulations were mounted in simulated gastric pH for 2 hours at the end of which the pH was adjusted to intestinal conditions. This design provided a mimic for the in vivo conditions. The pure gliclazide showed slow dissolution in acid phase with only 34% of the dose being dissolved within 2 hours this can be attributed to the hydrophobic nature of gliclazide .Changing the pH to the intestinal conditions resulted in significant increase in the dissolution rate with total amount dissolved exceeding 75% after 3 hours to exceed 98% after 4 hours (2 hours in each phase). This can be explained on the base of the dependence of the dissolution rate of gliclazide on the pH of the dissolution medium. The release profiles of gliclazide from phase transition formulations showed a dependence on the composition and physical form of the tested formulation. ME containing Transcutol and Gelucire at 1:1 molar ratio (ME1) liberating more than 75% of the drug in
Abstract 4
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
the first 30 minutes and more than 86% in the first hour. While (ME2) showed slower rate of gliclazide release compared to ME1. This can be attributed to the apparent increase in the viscosity of ME after increasing the concentration of Gelucire 50/13. The enhancement of drug dissolution from microemulsion system is expected and can be attributed to the massive increase in the surface area of ME due to small droplet size.
For liquid crystal systems there were significant reduction in the release rate compared to the corresponding microemulsion formulation. The LC1 liberated 29.6% in the acidic phase. Switching to the intestinal phase there was an abrupt increase in the amount of gliclazide dissolved to reach 71.5 after 1 hour in the intestinal phase. For the LC2 which contained higher proportions of Gelucire 50/13, the release rate underwent further reduction The overall release was significantly lower than that of the corresponding ME and was even lower than LC1 .With respect to the gel phase systems, there was significant reduction in gliclazide release to liberate only 21.3% and 16.7% for gel 1 and gel 2 in the acid phase. These formulations provided gradual liberation of gliclazide in the intestinal phase to reach 97% and 82%, respectively at the end of intestinal phase. The reduction in drug release rate from liquid crystalline and gel phase systems can be explained on the base that increasing the water content of the phase transition systems resulted in more viscous formulations with subsequent reduction in the rate of drug release. This explains the recorded reduction of gliclazide release from the gel phases as well and supports the recorded rank order in each system.
E. Investigation of the hypoglycemic effect of gliclazide
The in vitro results indicated better control on release rate from formulations containing Peceol in presence of 2:1 weight ratio Gelucire 50/13 with Transcutol. The hypoglycemic effect was monitored after oral
Abstract 5
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
administration of 10 mg/kg of unprocessed gliclazide suspension in water or its formulations which contain higher concentration of gelucire 50/13. Oral administration of unprocessed gliclazide to diabetic rats resulted in a gradual decrease in blood glucose concentration to reach a maximum glucose reduction level of 112.8 mg/dl after 3 hours. After this level the rate of glucose reduction decreased progressively. This profile produced an AUC of 801.4 mg. h/dl. Administration of gliclazide ME reduced blood glucose level to reach a maximum of 152.6 mg/dl after 3 hours to produce a profile with AUC value of 1253 mg. h/dl. This trend of increased efficacy after inclusion in ME system can be attributed to the intestinal membrane fluidizing effect of the ME components which can subsequently increase the extent of drug absorption.
Administration of gliclazide in the form of LC or gel resulted in a delay in the drug absorption with the maximum reduction of blood glucose being attained after 8 and 9 hours for LC and gel, respectively. Interestingly, this delay in the Tmax was not associated with reduction in the efficacy which is shown by recording maximum reduction in glucose levels of 149.8 and 144.5 mg/dl, respectively. The reduction of blood glucose was maintained for longer time after administration of LC or gel, compared with the recorded profiles of the control or ME. This was reflected as significant increase in the AUC after administration of LC or gel form compared with either the control or ME system.
The recorded in vivo data were correlated to the in vitro release results. This employed Pearson`s correlation which reflected good correlation between the release data and recorded blood glucose reduction in cases of ME, LC and gel. There was poor correlation between drug release from the unprocessed gliclazide and the blood glucose reduction.
Abstract 6
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
2. The effect of phase transition systems on the dissolution of cinnarzine
a. Development of a suitable method for analysis of the drug.
The assay utilized the UV spectrophotometer. Stock solution containing 1mg/ml was prepared in ethanol before suitable dilution to prepare the standard solution in 0.1N HCL. The absorbance values were recorded at 254nm and plotted as a function of concentration to draft the calibration graphs.
b. Preparation and evaluation of phase transition formulations of cinnarzine
Pseudoternary phase diagram was constructed of peceol, transcutol and gelucire44/14 which were used as the oily phase, co-surfactant and surfactant, respectively, Gelucire 44/14 with hydrophilic transcutol being adopted as surfactant /co-surfactant at ratio of 1:1w/w. This phase diagram shows microemulsion (ME), liquid crystal (LC), gel and coarse emulsion (EM) zones which were developed based on water content. These phases occupied 36, 23, 2.5, and 39.5% of the total area of phase diagram, respectively. This diagram was used to determine the composition of phase transition systems of cinnarzine. The microemulsion system was subjected for TEM analysis using transmission electron microscopy. The micrograph pictures revealed spherical ME droplets. The average droplet size was 135 + 34.6 nm. The high values of the standard deviation prove absence of homogeneity in distribution of particle size.
The release of cinnarzine was researched using 0.1N HCl as a release medium for 8 hours to provide the necessary sink environment. The unprocessed cinnarzine exhibited rapid dissolution in acidic pH with 28.8%
Abstract 7
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
dissolving in the first five minutes (Q5) before liberating 97.2% of the dose after the first hour. This high dissolution can be explained on the base of pH dependency of cinnarzine solubility with lower pH favoring drug dissolution. Changing the pH to the intestinal conditions resulted in precipitation of the drug with significant decrease in the dissolution rate. The release profiles of cinnarzine from phase transition formulations showed a dependence on the composition and physical form of the researched system. (SMEDDS) formula underwent fast release of cinnarzine as noticed from the Q5 value which reached 90.7% of dose to record 99.1% release efficiency. The release rate was significantly higher than the unprocessed drug. The improvement of cinnarzine dissolution from SMEDDS is due to solubilizing power of the components of SMEDDS. Peceol based ME showed rapid dissolution of cinnarzine (Q5 = 84% and Q 60 = 98%). The efficacy of ME in this respect correlates with its ability to provide fast surface area for drug release based on its nano-architecture.
LC showed slow liberation of cinnarzine compared with the tested liquid preparations. The LC liberated only 9.14% in the first five minutes and dissolved a total of 35.3 in the first hour. LC was able to deliver 94.5% within 8 hours. The release profile of cinnarzine from LC was dissimilar to that obtained from pure drug (F2 = 17%) and ME (F2= 5).
Further reduction in the release was achieved from gel phase to deliver only 28.89% in the first hour and 89% at the end of the study (8 hours). This provided an overall release efficiency of 55.5%. This is due to viscosity increase which can be intensified by water dilution. Incorporation of carboxy methyl cellulose (CMC) into gel systems resulted in significant reduction in release rate of cinnarzine from such formulations. For formulation containing 0.25% CMC, it liberated only a total of 9.9% in the
Abstract 8
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
first five minutes to produce a release efficiency of 43.2% after 8 hours this can be explained on the base of addition of CMC resulted in increasing the viscosity of gel systems with subsequent reduction in release rate of the drug. For F6 and F7 which contained higher concentration of CMC the release rate underwent further reduction to record an overall release efficiency of 38.3% and 31.5%, respectively. This can be explained that drug release rate decreases by increasing viscosity of gel formulations which inversely proportional to concentration of CMC added.
To investigate the effect of solid self dispersing systems Avicel/aerosol mixture (9:1) was employed for this purpose. Loading the LC and gel systems on the solid surface modulated the release pattern and changed the nature of the formulation from sustained release to immediate release. This change in release pattern is related to loading of the formulation on the large surface area of solid surface. This will provide fast dispersion of the formulation in the form of larger surface area for faster cinnarzine release. Accordingly, if sustained release is required, it will be better to load the formulation into capsules rather than loading on a solid surface. Similar finding was reported after adsorption of the same drug onto solid surface followed by tablet compression.
3. Cubosomes for enhancing intestinal absorption of fexofenadine hydrochloride: In situ and in vivo investigation
a. Development of a suitable method for analysis of the drug.
Analysis of in vitro release samples employed UV spectrophotometry but quantification of FEX-HCl in the in situ intestinal perfusion samples was accomplished using high pressure liquid chromatography (HPLC).
Abstract 9
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
The UV spectrophotometric analysis was successfully utilized for FEX-HCl quantification in drug release studies employing 0.001N HCl, PBS (pH 6.8) and PBS (pH 7.4) as release media. The absorbance values were recorded at 218nm. The calibration graphs were constructed in the corresponding release media and were linear in the range of 10-20 g/ml.
The HPLC method eluted FEX-HCl after 4.84 and 4.78 minutes after injecting FEX-HCl solutions in phosphate buffered saline, pH 6.8 and 7.4, respectively. The standard calibration graphs were linear in the concentration range of 0.5-20 g/ml for drug solutions prepared in both pH values. b. Preparation and characterization of the tested cubosome formulations Cubosomes are nanostructured systems self-assembled from amphiphilic lipids as peceol in water with the aid of suitable stabilizers as ploxomer 407. The composition of the selected cubosomal formulations included peceol and Poloxamer407 at a weight ratio of either 8:2 or 7:3, respectively. FEX-HCl-loaded cubosomes were prepared through controlled hydration of peceol/poloxamer407 mixture using homogenization technique. Morphological analysis of the liquid crystalline cubosomes applied TEM analysis using transmission electron microscope. For cubosomes incorporating lower poloxamer concentration (F1), TEM micrographs revealed spherical nanostructures free from any aggregates. Increasing poloxomer concentration in F2 cubosomes resulted in creation of spherical and polygonal structures arranged in honeycomb organization. The mean particle size values for the captured nanostructures were calculated to be 106+37 nm and 98+52 nm for cubosomes containing lower and higher poloxamer concentrations, respectively. Particle size analysis of cubosomes were assessed utilizing dynamic light scattering technique (DLS).The recorded average particle size values
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Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
were 162.1 nm (SD= 46.8) and 112.2 nm (SD= 7) for F1 and F2 cubosomes This correlates to the recorded particle size rank measured from TEM micrographs.
In vitro release studies of FEX-HCl encapsulated cubosomes revealed sustained drug release over 8 hours in 0.001N HCL. This pattern was recorded for both F1 and F2 cubosomes. FEX-HCl release from cubosomes formulation selected for in situ intestinal permeability investigation was monitored at pH values 6.8 and 7.4 for 2 hours. FEX-HCl release from F2 cubosomes revealed pH dependent release pattern. This was evidenced from the significantly reduced release efficiency values at pH values 6.8 and 7.4 compared to that recorded using 0.001N HCl as release medium. This indicates the pH dependent solubility of FEX-HCl. The release kinetics of FEX-HCl from cubosomal formulations followed Higuchi model suggesting matrix diffusion-based release
In situ rabbit intestinal diffusion technique was selected to monitor the efficacy of cubosomes for intestinal absorption enhancement. Perfusion of simple aqueous solution of FEX-HCl showed incomplete drug absorption from the tested intestinal segments due to its poor permeation through biological membranes. This was reflected from calculated high L95% which reflected site dependant absorption. Incorporation of FEX-HCl into cubosomes increased the intestinal absorption of the drug compared with the corresponding aqueous solution. The enhancement was recorded as a trend in case of the duodenum and ileum segments with significant increase in drug absorption being recorded in the jejunum and colon segments. The ability of cubosomes to enhance the intestinal permeability depends on their components capacity to inhibit P-gp efflux transporters and fluidize intestinal membrane. The increase in the absorptive clearance was associated with
Abstract 11
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
noticeable increase in the contribution of the transcellular pathway in drug absorption from all segments. This can be due to cubosomal components to inhibit P-gp efflux transporters which minimize transcellular pathway.
Correlation analysis of the intestinal absorption data to the released amount of FEX-HCl at the same time points was employed using person’s correlation. The computed results reflected no in vitro-in situ correlation which was shown from the recorded non-significant Pearson’s correlation coefficient “r” values.
The anti-inflammatory effect of FEX-HCl cubosomes was researched to verify the recorded enhanced in situ intestinal absorption using carrageenan provoked paw edema technique. Carrageenan induced edema formation was monitored after administration of plain water (negative control), FEX-HCl simple aqueous dispersion or cubosomal formulation. The negative control rats produced typical edema formation curve which was reached the maximum after 3 hours. Administration of a single dose of FEX-HCl suspension resulted in non significant. The recorded non-significant effect of FEX-HCl aqueous dispersion correlates with the recorded in situ intestinal perfusion data which reflected its incomplete absorption from the GIT. This implies poor bioavailability of FEX-HCl which classified as poorly permeable drug. Poor permeability of FEX-HCl was also assisted by P-gp efflux. Administration of FEX-HCl cubosomal dispersion resulted in significant reduction in edema formation compared with the negative control group or that treated with simple suspension. The recorded enhancement in the anti-inflammatory effect correlates with the recorded increase in the intestinal permeability from cubosomes. This can suggest that cubosomal dispersion is able to enhance drug absorption of poorly permeable drugs. Alternative mechanisms have been suggested for enhanced oral
Abstract 12
Pharmaceutical Technology department, College of Pharmacy, University of Tanta, Tanta, Egypt
bioavailability from cubosomes. Being colloidal dispersion, cubosomes can traverse the intestine via lymphatic pathway bypassing the negative effects of pre-systemic disposition. The effect of colloidal nature of cubosomes is further magnified by the nature of the primary components. peceol is believed to inhibit P-gp efflux and augment trans-lymphatic transport of drug. Poloxamer can also contribute by permeabilization of intestinal membrane with additional inhibitory effect on CYP 450 enzymes. These factors combine to explain the recorded increase in the in vivo efficacy of FEX-HCl after administration in cubosomal formulation.