Development of mesophasic microreservoir-based transdermal drug delivery system of propranolol.

The mesophasic microreservoir comprises lyotrophic liquid crystals. The liquid crystals were prepared of Brij-35, cetosteryl alcohol and propranolol and evaluated for parameters viz. anisotropy, size and size distribution and drug entrapment efficiency. Subsequent to this liquid crystals based transdermal drug delivery system (TDS) was prepared by incorporating liquid crystals in previously prepared matrix based transdermal patch and evaluated for stability studies like temperature, humidity and aging. The system was also studied for tensile strength, moisture content, water vapor transmission, drug content, anisotropy and In vitro drug release studies.

Development of hemoglobin aquasomes from spherical hydroxyapatite cores precipitated in the presence of half-generation poly(amidoamine) dendrimer.

Spherical hydroxyapatite cores were prepared by using carboxylic acid terminated half-generation poly(amidoamine) (PAMAM) dendrimer as templates or crystal modifiers. The hydroxyapatite cores were characterized by infrared spectroscopy (IR), X-ray diffraction (XRD) and transmission electron microscopy (TEM). The spherical core formation depended on phosphate saturation, pH of the simulated body fluid (SBF) and rate of crystal growth. Hydroxyapatite so formed was amorphous and a mixture of various calcium phosphates. Ca/P ratio determination which showed phosphate rich apatite formation. Hydroxyapatite ores were coated with a sugar layer followed by hemoglobin to obtain aquasomes. Aquasomes were characterized for size, hemoglobin loading, oxygen-binding characteristics and storage stability. The nanometric sized aquasome formulation could load approximately 13.7 mg hemoglobin per g of core and retained oxygen-affinity and cooperativity and stability for at least 30 days. Formulation efficacy was tested in albino rats and indicated its potential utility as blood-substitute.

Dendrimer grafts for delivery of 5-fluorouracil.

Polyamidoamine (PAMAM) dendrimers were prepared by linking methyl methacrylate and ethylenediamine successively on an amine core. Surface modification of PAMAM dendrimer was done by fatty acid grafting converting them to a unimolecular micellar system (Dendrimer grafts). IR, 1H NMR, 13C NMR studies confirmed the structure. The drug 5-fluorouracil (5-FU) was entrapped in dendrimer grafts. The effects of various solvents (ethanol, dichloromethane, tetrahydrofuran), pH and ionic strength on solubilization of 5-FU were determined. Phospholipid was further coated on the dendrimer grafts. The product was lyophilized and obtained as yellowish-white powder. Average particle size was ca. 375 nm as determined by Malvern's Mastersizer 4. Drug loading was ca. 53% by weight. Stability studies were conducted for 1 month at room temperature and 40 degrees C, where the systems were relatively stable. Release rate was sustained across cellulose tubing in PBS. In vivo studies were performed in albino rats and pharmacokinetic parameters and bioavailability were determined from the plasma profile of 5-FU. The phospholipid coated dendrimer graft formulation was found to be more effective orally than free drug. The lymphatic uptake was also increased indicating absorption of the developed formulation through the lymphatic route.

Concanavalin-A conjugated fine-multiple emulsion loaded with 6-mercaptopurine.

Fine-multiple (water-in-oil-in-water) emulsions were prepared by two-step emulsification using sonication. They were coated with concanavalin-A (Con-A) by three methods. The one involving covalent coupling of Con-A to the multiple emulsion incorporated anchor was better compared with lipid derivatized Con-A anchoring or the glutaraldehyde-based cross-linking method, as shown by the faster rate of dextran-induced aggregation. The selected multiple emulsions were characterized by physical properties such as droplet size, encapsulation efficiency, and zeta potential. Stability parameters such as droplet size, creaming, leakage, and aggregation as a function of relative turbidity were monitored over a 1-month period, which revealed good stability of the formulations. The release profile of 6-mercaptopurine followed zero-order kinetics. Pharmacokinetic studies showed an increase in half-life and bioavailability from multiple emulsion formulations administered intravenously. There was prolonged retention of drug in various tissues of rats when treated with Con-A-coated multiple emulsion as compared with uncoated one. Our study demonstrates the suitability of fine-multiple emulsion for intravenous administration and the potential for prolonged retention of drugs and targeting in biological systems.

Liposphere based lipoprotein-mimetic delivery system for 6-mercaptopurine.

Long-circulating lipospheres containing 6-mercaptopurine (6-MP) were prepared by solidification of warm microemulsion at low temperature. Palmitoyl PEG was incorporated in the system to confer stealth-type nature. The size of lipospheres was in the range of 60-70 nm and was inversely proportional to sonication time. The size range was attained after 8 h. of sonication. The entrapped 6-MP contained 0.12 mmol/mole of lipid. The coating efficiency of 63-71% was attained. The zeta potential substantially decreased after PEG coating, however, the lipospheres were stable due to steric repulsion and exhibited no aggregation. The release of 6-MP was found to be 18-25% of administered dose in 24 h. and followed a mixed profile for stealth lipospheres. The percent dose remaining in plasma was found to be high even after 24 h as compared to control, indicating an increase in circulation time of lipospheres. Tissue accumulation of drug correlated with the pharmacokinetic behavior of lipospheres. The system seems to be an ideal carrier for anticancer drug delivery.

High-entrapment liposomes for 6-Mercaptopurine--a prodrug approach.

Low entrapment of drugs into liposomes is a serious challenge in their commercial application. 6-Mercaptopurine (6-MP), an antineoplastic agent, is such a drug with low entrapment efficiency (EE). We devised their lipophilic derivatization as a means of enhancing EE by covalently coupling 6-MP with glyceryl monostearate (GMS) via a succinic anhydride spacer. This prodrug had an improved partition coefficient value of 25.16 compared to 1.22 for free drug, confirming higher lipophilicity. A hydrolysis rate study of prodrug indicated 2.90%, 12.5%, 24.1%, and 25.1% hydrolysis in phosphate buffered saline (PBS) (pH 7.4) and 10%, 20%, and 30% serum, respectively. Liposomes of phosphatidylcholine (PC)/sphingomyelin, cholesterol, and dicetyl phosphate bearing drug or prodrug were prepared by shaking by hand and sonication methods. The EE was found to increase from 1.92% for free drug to 91.8% for drug-conjugate. An in vitro cell line toxicity study on L1210 leukemia cells showed improved performance of liposome-encapsulated drug-conjugate compared to free drug. The plasma drug level profile following administration of free drug and the liposomal formulation containing prodrug (HE liposome) manifested a higher sustained level of the latter, which was further improved in case of sphingomyelin-containing liposomes (STHE liposome). The pharmacokinetic parameters revealed an increase in half-life, from 61 min to 120 min for the HE liposomes and 296 min for the STHE liposomes. Therefore, increased entrapment was made possible through lipophilic derivatization, and it was subsequently tested in vivo.

Lipoprotein-mimicking biovectorized systems for methotrexate delivery.

The present investigation reports a new family of lipid nanoparticles biomimetic of lipoproteins, lipoprotein-mimicking biovectorized systems (LMBVs), for the delivery of methotrexate. LMBVs were prepared by microemulsion congealing technique, and the composition and process were optimized. The palmitoylpolyethylene glycol 4000 (p-PEG 4000) was anchored on LMBVs as apoprotein analogue. Various formulations were prepared and characterized for size and polydispersity, zeta potential, drug entrapment efficiency, anchoring efficiency, release kinetics, pharmacokinetics and tissue distribution. The size was 70-76 nm and the polydispersity was 0.09-0.18. The zeta potential was -63.2 which was reduced to -19.3 after p-PEG 4000 coating. Entrapment efficiency varied from 22.6% to 30.2%. Anchoring efficiency was 74.0 +/- 3.9%. The drug release showed zero-order profile. Their circulation half-life was enhanced and exhibited capability to accumulate in tissues for longer periods. The composition of lipidic part of LMBVs and p-PEG 4000 anchoring impart similarity with natural lipoproteins in terms of in vivo behaviour. This new drug carrier named LMBVs, holds promise for targeting and systemic controlled release, which may prove effective in the treatment of various types of cancer.

Lipid grafts of egg-box complex: a new supramolecular biovector for 5-fluorouracil delivery.

An attempt was made to improve the pharmacokinetic behaviour of 5-fluorouracil (5-FU) by incorporating it into lipoprotein imitating synthetic carrier 'supramolecular biovector (SMBV)' which is an important prerequisite for achieving its better therapeutic performance against cancer. The polysaccharide core of SMBVs was prepared by ionotropic gelation technique by cross-linking polyguluronate units in the alginate molecules with calcium ions to form so called 'egg-box structure'. The formulation and process variables were optimized to obtain particles of nanometer size range. Hydrophobization was carried out by fatty-acylation on the surface followed by phospholipid coating. Palmitoyl polyethylene glycol (p-PEG) was anchored to impart stealth behaviour. The scanning electron microscopy showed discrete spheres of average diameter 748 nm. Polydispersity was estimated to be 0.37. Overall zeta potential was -21.3 mV. The drug loading capacity and encapsulation efficiency was found to be 10.0% and 97.9%, respectively. The release from drug solution (AP) followed zero-order kinetics. Higuchi release pattern was obtained for egg-box complex cores (AP1) while first-order pattern was followed for fatty acylated (AP2) and lipid coated cores before (AP3) and after p-PEG anchoring (AP4). The amount of drug liberated in 24 h was in the order AP > AP1 > AP2 > AP4 > AP3. The release pattern obtained was a combined effect of drug diffusion through egg-box matrix as well as partitioning in hydrophobic layer and p-PEG layer around the SMBV. The stability study showed negligible leakage and no appreciable change in particle size upon storage at different temperatures which is an indication of good stability of SMBV formulation. The plasma clearance data revealed increase in circulation half-life of drug and bioavailability. Tissue distribution data obtained was a result of competitive uptake of formulations from tissue macrophages and lymphatics depending upon its surface characteristics and residence period in vascular system. The enhanced delivery of drug to lymphatics and improvement in its half-life render SMBVs useful for control of metastasis and tumour growth.

Multiple emulsions containing rifampicin.

A stable multiple emulsion containing rifampicin in the internal aqueous phase was prepared by the incorporation of additives in both aqueous and oily phases. The formulation and process variables were optimized for primary and secondary emulsification. Drug release studies were performed on selected multiple emulsions to observe the effect of dilution. The release data were fitted to the Higuchi equation for slab and spherical geometry and effective diffusion coefficients were calculated. Stability studies for three months revealed good stability of the multiple emulsions with respect to creaming, phase separation, viscosity change, drug leakage, change in droplet size upon storage and exposure to osmotic and shear stress. The in vivo studies performed with stable multiple emulsions administered orally in human volunteers showed prolonged plasma drug levels. The multiple emulsions were found suitable for an improved tuberculosis therapy.

Proniosome based transdermal delivery of levonorgestrel for effective contraception.

A proniosome based transdermal drug delivery system of levonorgestrel (LN) was developed and extensively characterized both in vitro and in vivo. The proniosomal structure was liquid crystalline-compact niosomes hybrid which could be converted into niosomes upon hydration. The system was evaluated in vitro for drug loading, rate of hydration (spontaneity), vesicle size, polydispersity, entrapment efficiency and drug diffusion across rat skin. The effect of composition of formulation, amount of drug, type of Spans, alcohols and sonication time on transdermal permeation profile was observed. The stability studies were performed at 4 degrees C and at room temperature. The biological assay for progestational activity included endometrial assay and inhibition with the formation of corpora lutea. The study demonstrated the utility of proniosomal transdermal patch bearing levonorgestrel for effective contraception.

Iontophoretic transdermal delivery of salbutamol: methyl-orange ion-pair.

The influence of pulsed current duty cycle, addition of electrically neutral species and pH on iontophoretic transdermal delivery of an organic ion-pair salbutamol:methyl orange through excised rabbit abdominal skin is described. It was compared with salbutamol sulphate and salbutamol:sulphate ion-pair. It was found that there exists an optimum duty cycle range (1:3-1:4) at which iontophoretic flux of the ion-pair was at a maximum because of partition-electric current synchronization. Addition of mannitol decreases the flux of the ion-pair showing the important role of electroosmosis in ion-pair transport. The extremes of pH decrease the flux rates.

Effects of drug concentration in inner aqueous phase and additives in oleaginous phase on release and bioavailability of isoniazid from multiple emulsion.

The effects of drug concentration in internal aqueous phase of stabilized w/o/w emulsion and additives in oleaginous phase on release characteristics of isoniazid were investigated. The release was significantly effected by both of the formulation variables. The release was enhanced initially with increasing concentration of drug in internal aqueous phase followed by a steady release at high concentration of isoniazid. The release declined substantially in the presence of aluminum tristearate, cetostearyl alcohol, and cholesterol, and it increased with egg lecithin and oleic acid in oily phase. The bioavailability was increased with a multiple-emulsion formulation.

A stable multiple emulsion system bearing isoniazid: preparation and characterization.

Multiple emulsions with an oily liquid membrane (w/o/w) bearing isoniazid were prepared by an improved 2 x 2 step emulsification technique. Both of the interfaces of the liquid membrane were stabilized by using microcrystalline cellulose (MCC) in external as well as internal aqueous phases. The emulsions were characterized for droplet size, percent formation of multiple emulsion, release rate effect of Tween-80 in external phase, phase volume ratio on release, and stability during aging at various storage conditions. The droplet size was small and yield of multiple emulsion was fairly good. The increasing concentration of MCC in either internal or external phase increased the droplet size. The system holds promise in tuberculosis therapy.

Lectin-functionalized multiple emulsions for improved cancer therapy.

Fine-multiple emulsions bearing 6-mercaptopurine (6-MP) in internal aqueous phase were prepared by two step emulsification using sonication technique. It was coated with Concanavalin-A (Con-A) using carbodiimide method to obtain lectin-functionalized multiple emulsions. The Con-A coated multiple emulsion was characterized for antitumour activity on murine leukemia cell line L-1210 in vitro and compared with uncoated multiple emulsion and free drug. An increased uptake and cytotoxicity were observed for Con-A coated multiple emulsion in vitro. The IC50 was decreased upto 4-fold with Con-A coated emulsion. In vivo antitumour activity was seen by recording survival times of mice injected with L-1210 cells i.v. or i.p. The mean survival time was found to increase upon treatment with Con-A coated multiple emulsion. The tumour cell count in the peritoneal cavity was decreased significantly when animal was treated by i.p. route while there was no significant difference when it was treated by i.v. route. The normal peritoneal cells remained unaltered in number and blood parameters were also restored on treatment to tumour bearing mice. The formulation was found to be effective for the treatment of cancer.

Proliposome-based transdermal delivery of levonorgestrel.

Mesophasic proliposomal system for levonorgestrel was developed and evaluated both in vitro and in vivo. The vesicles were mostly unilamellar, however, few vesicles were multilamellar which budded off spontaneously upon hydration. The release of drug from this system adhered to zero order kinetics. The effect of alcohols and volatile oils on transdermal flux was investigated. The flux was found to be the highest for alcohol, and followed by that for lemon oil. The in vivo studies indicate the requirement for a loading dose, since, a significant lag phase was observed before the therapeutic levels were reached. This system was, however, superior to the PEG-based ointment system which was employed as the control formulation. The results demonstrate the potential of proliposomal system for efficacious transdermal delivery of hydrophobic drugs.

Targeting of multiple emulsions to the lungs.

The conventional W/O/W emulsions are readily taken up by the reticuloendothelial system (RES) so that it is necessary to develop either stealth type or tissue specific multiple emulsion for effective targeting. A multiple emulsion system W/O/W containing rifampicin as encapsulant (W/O/W) was prepared. The droplet size was kept small to study targeting independent of passive embolism of larger droplets in the lung capillaries. It was characterised in vitro for drug release through a treated cellophane membrane. A prolonged first order drug release was observed. The W/O/W multiple emulsion system was coated with an o-palmitoyl derivative of mannan (mol.wt. 20,000) to assess its toxicity in vitro and its distribution behaviour in vivo. The multiple emulsion was found to be non toxic at 0.2 ml formulation/10(6) cells level in presence or absence of serum. An intravenous injection of the polysaccharide coated multiple emulsion was given and the tissue distribution of the drug after 1 h and 24 h was investigated. A significant enhancement in lung uptake and a decreased internalisation by spleen was noticed.

Self assembling nanostructures for sustained ophthalmic drug delivery.

Nanopseudolatex was prepared for sustained ophthalmic delivery. It was prepared by a solvent injection method without the use of any stabilizer. The acrylate copolymers Eudragit RLPM and RSPM were used as carrier materials. Their formation was confirmed by electron microscopy. The system was extensively characterized for particle size, viscosity, sedimentation volume, encapsulation efficiency, in vitro release profile and pharmacodynamic properties. The particle size was found to be stirring rate dependent above the injection rate of 1 ml/min. Only at slow stirring rates it was temperature dependent. There was no sedimentation for about 60 d. The viscosity was 23-27 cps which was particle size dependent. The results show an excellent encapsulation efficiency of about 94-98%. Release studies showed slow and consistent drug release from the formulation. However, Eudragit RSPM showed comparatively longer release than Eudragit RLPM nanosuspensions. Using this method we have developed a new formulation of diclofenac diethyl ammonium which showed increased therapeutic efficacy as compared with eye drops.