The Effects of Lyophilization on the Physico-Chemical Stability of Sirolimus Liposomes

Purpose: The major limitation in the widespread use of liposome drug delivery system is its instability. Lyophilization is a promising approach to ensure the long-term stability of liposomes. The aim of this study was to prepare sirolimus-loaded liposomes, study their stability and investigate the effect of lyophilization either in the presence or in the absence of lyoprotectant on liposome properties. Methods: Two types of multi-lamellar liposomes, conventional and fusogenic, containing sirolimus were prepared by modified thin film hydration method with different ratio of dipalmitoylphosphatidylcholine (DPPC), cholesterol and dioleoylphosphoethanolamine (DOPE), and were lyophilized with or without dextrose as lyoprotectant. Chemical stability investigation was performed at 4°C and 25°C until 6 months using a validated HPLC method. Physical stability was studied with determination of particle size (PS) and encapsulation efficiency (EE %) of formulations through 6 months. Results: Chemical stability test at 4°C and 25°C until 6 months showed that drug content of liposomes decreased 8.4% and 20.2% respectively. Initial mean EE % and PS were 72.8 % and 582 nm respectively. After 6 months mean EE % for suspended form, lyophilized without lyoprotectant and lyophilized with lyoprotectant were 54.8 %, 62.3% and 67.1 % at 4°C and 48.2%, 60.4 % and 66.8 % at 25°C respectively. Corresponding data for mean PS were 8229 nm, 2397 nm and 688nm at 4°C and 9362 nm, 1944 nm and 737 nm at 25°C respectively. Conclusion: It is concluded that lyophilization with and without dextrose could increase shelf life of liposome and dextrose has lyoprotectant effect that stabilized liposomes in the lyophilization process

Effects of polyethylene glycols on intestinal efflux pump expression and activity in Caco-2 cells

The present study was planned to investigate the influence of polyethylene glycols (PEGs) on the activity and expression of P-glycoprotein (P-gp). Sub-toxic concentrations of PEGs in Caco-2 cells were determined using the MTT test assay. Then the measurement of Rhodamine-123 (Rho-123) uptake, a P-gp fluorescence substrate, in Caco-2 cells confronting PEG 400 (1% and 2% w/v), PEG 4000 (2% and 4% w/v), PEG 6000 (2% and 4% w/v), PEG 10000 (2% and 4% w/v), PEG 15000 (1% and 2% w/v), and PEG 35000 (2% and 4% w/v) overnight was taken to elucidate whether non-toxic concentrations of PEGs are able to impact P-gp activity. Furthermore, western blotting was carried out to investigate P-gp protein expression. The results showed that PEG 400 at concentrations of 1% (w/v) and 2% (w/v) and PEG 6000 at the concentration of 4% (w/v) are notably capable of blocking P-gp. Based on the obtained results it is concluded that the mentioned excipients could be used to obstruct P-gp efflux transporter in order to increase the bioavailability of co-administered substrate drug.O presente estudo foi planejado para investigar a influência de polietileno glicóis sobre a atividade e expressão da P- glicoproteína (P-gp) . Concentrações sub-tóxicas de PGPs e em células Caco-2 foram determinadas por meio do ensaio de MTT. Em seguida, efetuou-se a a medida de captura de Rodamina-123 (Rho-123), um substrato fluorescente de P-gp, em células Caco-2, confrontando com PEG 400 (1% e 2% m/v), PEG 4000 (2% e 4% m/v) e PEG 6000 (2% e 4% m /v), PEG 10000 (2% e 4% w/v), PEG 15000 (1% e 2% m/v), e PEG 35000 (2% e 4% m/v). Essa medida foi efetuada durante a noite, para saber se as concentrações não tóxicas de excipientes são capazes de influenciar a actividade da P-gp. Além disso, realizou-se o western blotting para investigar a expressão da proteína P-gp. Os resultados mostraram que o PEG 400, nas concentrações de 1% (m/v) e 2% (m/v), e PEG 6000, na concentração de 4% (m/v) são capazes de bloquear P-gp. Com base nos resultados conclui-se que os excipientes mencionados poderiam ser utilizados para obstruir o efluxo por P-gp, a fim de aumentar a biodisponibilidade de do fármaco co-administrado

The effect of protocol for disinfection of extracted teeth recommended by center for disease control (CDC) on microhardness of enamel and dentin

Background: According to the guideline of the United States center for disease control (CDC), the extracted teeth should be sterilized by autoclaving or storage in 10% formalin before using for educational or research purposes. The objective of this study was to evaluate the effect of this protocol on microhardness of dentin and enamel. Material and Methods: Thirty extracted single-root teeth were used in this study. The crowns were resected, and the roots were longitudinally sectioned into two halves. The Vickers microhardness (VHN) of specimens was measured on polished canal dentin and buccal enamel surfaces. The crowns were randomly divided into three groups (n=10). Group 1 and 2 were sterilized using autoclave and formalin, respectively while group 3 (control) was stored in synthetic tissue fluid. The root halves were also randomly divided into 3 groups (n=20) which were treated as mentioned above for crown samples. Following sterilization, VHN of samples was measured again. ANOVA and paired samples t-tests were used to analyze the data. Results: Autoclaving caused a significant reduction in microhardness of dentin ( P <0.001, 12.04% decreases in VHN). However, there were no significant differences for before and after sterilization within other groups. Conclusions: Based on the results of this study, the CDC protocol is recommended in studies related to enamel microhardness. However, Autoclaving is not an appropriate sterilization method in studies related to dentin microhardness. In these studies, two-week immersion in 10% formalin is recommended

Application of lactobionic acid and nonionic surfactants as solubilizing agents for parenteral formulation of clarithromycin

Purpose: The purpose of this paper is to enhance the solubility of clarithromycin (CLR) using nonionic surfactants and some type of acids for preparation of the new formulations. Methods: Myrj 52 and chremophor (2.5 and 5% w/v) were used in two concentrations. To investigate solubility, the formulations were shaken for 48 hours at room temperature. For stability test, lyophilized samples were maintained in refrigerator at 4° C, and in oven at 40° C. Drug analysis was performed by reverse phase high performance liquid chromatography (HPLC) with ultraviolet detection. Results: Solubility tests indicated that lactobionic acid was the most effective to increase clarithromycin solubility and chremophor showed higher enhancing effect than myrj 52 on CLR solubility. The stability tests results also confirmed that shelf-lives of all formulations have been the equivalent to 24 months. Conclusion: On the whole, formulations described in this article may be very suitable for industrial-scale manufacturing and clinical application

A study on enhanced intestinal permeability of clarithromycin nanoparticles

The main objective of the present study was to determine the permeability of clarithromycin (CLA)-PLGA nanoparticles using single-pass intestinal perfusion technique in rats. Clarithromycin nanoparticles were prepared by nano-precipitation according to the modified quasi emulsion solvent diffusion technique and evaluated for their physicochemical characteristics. Permeability coefficients (Peff) in anaesthetized rats were determined at 3 different concentrations. Drug solution or suspensions in PBS was perfused through a cannulated jejunal segment and samples were taken from outlet tubing at different time points up to 90 min. Microbiological assay of CLA and phenol red in the samples were analyzed using an agar well diffusion procedure and HPLC method respectively. The average particle size of prepared nanoparticles was 305 ± 134 nm. The mean Peff of CLA solution in concentrations of 150, 250 and 400 µg/mL was found to be 1.20 (±0.32) ×10-3, 9.62 (±0.46) ×10-4, and 1.36 (±0.95) ×10-3 cm/sec, respectively. The corresponding values for the same concentration of nanoparticles were found to be 2.74 (±0.73) ×10-3, 2.45 (±0.88) ×10-3, and 3.68 (±0.46) ×10-3 cm/s, respectively. The two-tailed Student’s t-test showed that the intestinal permeability of CLA nanoparticle suspensions in prepared concentrations were significantly increased in comparison with its solution.O objetivo principal do presente estudo foi determinar a permeabilidade de nanopartículas de claritromicina (CLA)-PLGA, utilizando a técnica de perfusão intestinal de passo único em ratos. As nanopartículas de claritromicina foram preparadas por nanoprecipitação, de acordo com a técnica modificada de difusão de solvente quase-emulsão, e suas características físico-químicas avaliadas. Os coeficientes de permeabilidade (Peff) em ratos anestesiados foram determinados em três concentrações diferentes. A solução, ou suspensões, do fármaco em PBS foi perfundida através do segmento de jejuno canulado e as amostras foram tomadas do tubo externo em diferentes tempos até 90 minutos. Os ensaios microbiológico de CLA e de vermelho de fenol das amostras foram realizados, utilizando-se o procedimento de difusão em poço de ágar e de CLAE, respectivamente. O tamanho médio das partículas das nanopartículas preparadas foi de 305 ± 134 nm. O Peff médio da solução de CLA em concentrações de 150, 250 and 400 µg/mL foi de 1.20(±0.32)×10-3, 9.62(±0.46)]×10-4 e de 1.36(±0.95)×10-3 cm/s, respectivamente. O valor correspondente para a mesma concentração de nanopartículas foi de 2.74 (±0.73)×10-3, 2.45(±0.88)×10-3 e de 3.68 (±0.46)×10-3 cm/s, respectivamente. O teste t de Student com duas variáveis mostrou que a permeabilidade intestinal das suspensões de nanopartículas de CLA nas concentrações preparadas foram significativamente aumentadas em comparação com sua solução

Investigating the Effect of Basic Amino Acids and Glucosamine on the Solubility of Ibuprofen and Piroxicam

Purpose: Poor aqueous solubility hampers the development of several compounds as pharmacological agents. Hence, preparing novel formulations with augmented absorption is a challenge in pharmaceutical industries. In this paper, we have examined the effect of basic amino acids including arginine (ARG), lysine (LYS), and glucosamine (GlucN) on the solubility of ibuprofen (IBU) and piroxicam (PXM) as drugs with limited solubility. We have also studied the effect of the dissolution media with the pH values 1.2 to 7.4. Methods: The saturation shake-flask method was used for solubility studies in the presence of amino acids. Briefly, buffer solutions containing different concentrations of amino acids were prepared. Then, an excess amount of each drug with these buffers was shaken to reach equilibrium. After 48 hours, the upper phase was separated, and solubility was calculated by reading their UV-Vis absorbance. Results: The results illustrated that amino acids increased solubility of both drugs with different ratios, which were pH and concentration-dependent. Solubility improved as the amount of amino acids went up, and this upward pattern was more robust with ARG than LYS. The presence of GlucN in citrate buffer significantly enhanced IBU solubility. The solubility of PXM in accompany of GlucN in water did not change significantly while in citrate buffer solubility enhanced specially at pH 6. Conclusion: Overall, GlucN in citrate buffer and ARG in phosphate buffer could be introduced as the most suitable media for IBU and PXM solubility improvement, respectively

Studies on Dissolution Enhancement of Prednisolone, a Poorly Water-Soluble Drug by Solid Dispersion Technique

Introduction: Prednisolone is a class II substance according to the Biopharmaceutics Classification System. It is a poorly water soluble agent. The aim of the present study was to improve dissolution rate of a poorly water-soluble drug, prednisolone, by a solid dispersion technique. Methods: Solid dispersion of prednisolone was prepared with PEG 6000 or different carbohydrates such as lactose and dextrin with various ratios of the drug to carrier i.e., 1:10, 1:20 and 1:40. Solid dispersions were prepared by coevaporation method. The evaluation of the properties of the dispersions was performed using dissolution studies, Fourier-transform infrared spectroscopy and x-ray powder diffractometery. Results: The results indicated that lactose is suitable carriers to enhance the in vitro dissolution rate of prednisolone. The data from the x-ray diffraction showed that the drug was still detectable in its solid state in all solid dispersions except solid dispersions prepared by dextrin as carrier. The results from infrared spectroscopy showed no well-defined drug–carrier interactions for coevaporates. Conclusion: Solid dispersion of a poorly water-soluble drug, prednisolone may alleviate the problems of delayed and inconsistent rate of dissolution of the drug

Stomach-Specific Drug Delivery of Clarithromycin Using aSemi Interpenetrating Polymeric Network Hydrogel Made ofMontmorillonite and Chitosan: Synthesis, Characterization and InVitro Drug Release Study

Purpose: In this study, we aimed to prepare an extended drug delivery formulation ofclarithromycin (CAM) based on a semi-interpenetrating polymer network (semi-IPN) hydrogel.Methods: Synthesis of semi-IPN hydrogel nanocomposite made of chitosan (CS), acrylicacid (AA), acrylamide (AAm), polyvinylpyrrolidone (PVP), and montmorillonite (MMT) wasperformed by free radical graft copolymerization method. Swelling kinetic studies were done inacidic buffer solutions of hydrochloric acid (pH = 1.2), acetate (pH = 4), and also distilled water.Also, the effects of MMT on the swelling kinetic, thermal stability, and mechanical strengthof the hydrogels were evaluated. Moreover, in vitro release behavior of CAM and its releasekinetics from hydrogels were studied in a hydrochloric acid buffer solution.Results: Fourier transform infrared spectroscopy (FTIR) results revealed that synthesis of semi-IPN superabsorbent nanocomposite and CAM incorporation into hydrogel was performed,successfully. Introducing MMT into hydrogel network not only improved its thermal stabilitybut also increased mechanical strength of the final hydrogel product. Also, in comparisonwith neat hydrogel (1270 g/g), hydrogel nanocomposite containing 13 wt% MMT exhibitedgreater equilibrium swelling capacity (1568 g/g) with lower swelling rate. In vitro drug releaseexperiments showed that CS-g-poly(AA-co-AAm)/PVP/MMT/CAM formulation possesses asustained release character over extended period of time compared with CS-g-poly(AA-co-AAm)/PVP/CAM formulation.Conclusion: In the presence of MMT, the effective life time of drug is prolonged, demonstrating asustained release property. The reason is that interlinked porous channels within superabsorbentnanocomposite network hinder penetration of aqueous solutions into hydrogel and subsequentlycause a slower drug release