Optimization of protein encapsulation in PLGA nanoparticles

Influences of process parameters were investigated on the efficiency of encapsulation of bovine serum albumin (BSA) in poly(dl-lactic-co-glycolic acid) (PLGA) nanoparticles produced by w₁/o/w₂ (water-in-oil-in-water) double emulsion-solvent evaporation method. According to a 5-factorial 3-level Box-Behnken type experimental design aqueous solution of BSA was emulsified in an immiscible organic phase composed of dichloromethane and various quantities of dissolved PLGA to get water-in-oil (w₁/o) emulsion. This latter was then dispersed in a second aqueous phase (w₂) containing poly-vinyl-alcohol (PVA) surfactant as an emulsifier/stabilising agent. PLGA nanoparticles with encapsulated BSA were obtained by evaporating the dichloromethane from the w₁/o droplets. Encapsulation efficiency was determined as the weight ratio of BSA remained in the PLGA nanoparticles relative to the total weight of BSA used in the process. By statistical evaluation of the experimental results an equation was proposed to predict the encapsulation efficiency as a function of five process variables. Two optimization procedures were carried out to increase the efficiency of encapsulation, with and without constraints referring to the required mean particle size. Correlation was found between the latter and the achievable maximal encapsulation efficiency under optimal process conditions.     

Influence of Surfactant on the Characteristics of W1/O/W2-Microparticles

The water-in-oil-in water (W₁/O/W₂) double emulsion evaporation technique is widely used when the microencapsulation of soluble agents like naloxone HCl is intended. The present work shows the effect of HLB emulsifiers added to phase O on microsphere morphology, size, release, drug encapsulation efficiency. The addition of sorbitan ester to first emulsion (W₁/O) and the HLB of the surfactant have an important effect on the characteristics of poly-lactide-co-glycolide (PLGA) microparticles (MP). This MP with sorbitan esters added were smaller and released the hydrophilic drug, naloxone, with no-significant difference at pH 5 versus pH 7.5 (phosphate medium). This is an important fact when long-drug release is considered since it is known that PLGA degradation leads to media acidification. The HLB value had an important effect on drug loading. Sorbitan monooleate led to the highest naloxone loading. Because of its low HLB (4.3), it is most suitable for stabilizing the W₁/O emulsion, which is fundamental for the successful entrapment of a hydrophilic compound in MP prepared by double emulsion technique. Finally, drug solubility in the MP matrixes cannot be considered as a predictive parameter for drug encapsulation. Both surfactants increased the naloxone solubility in the polymer PLGA and only sorbitan monooleate increased the drug entrapment.     

Preparation,optimization, and characterization of simvastatin nanoparticles by electrospraying: An artificial neural networks study

The purpose of this study was to determine major factors impacting the size of simvastatin (SIM)‐loaded poly(d , l ‐lactic‐co‐glycolide) (PLGA) nanoparticles (NPs) that was prepared using electrospraying. Three variables including concentration of polymer and salt as well as solvent flow rate were used as input variables. Size of NPs was considered as output variable. For the first time, our findings using a systematic and experimental approach, showed the importance of salt concentration as the dominant factor determining the size with a sharp and reverse effect. Optimum formulation (i.e., flow rate 0.08 mL h^?1, polymer concentration 0.7 w/v %, and salt concentration 0.8 mM) was then evaluated for aqueous solubility, encapsulation efficiency, particle size, in vitro drug release pattern and cytotoxicity. A very appreciable encapsulation efficiency (90.3%) as well as sustained release profile, considerable enhancement in aqueous solubility (～5.8 fold) and high IC₅₀ (>600 µM of SIM‐loaded PLGA NPs) indicated PLGA as a promising nanocarrier for SIM. The optimum formulation had particle size, zeta potential value, polydispersity index (PDI) and drug loading of 166 nm, +3 mV, 0.62 and 9%, respectively. © 2016 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2016 , 133, 43602.     

Poly(hydroxybutyrate-co-hydroxyvalerate) microspheres loaded with atrazine herbicide: screening of conditions for preparation,physico-chemical characterization,and in vitro release studies

Due to their widespread use in agriculture as well as in urban areas, agricultural chemicals are globally some of the most commonly encountered substances in waters. The objective of this study is to develop (including preparation and characterization) a new modified release system for the herbicide atrazine, employing poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) microspheres. The microspheres were prepared by the emulsification/solvent evaporation method, emulsifying an organic phase (atrazine and PHBV dissolved in chloroform) into an aqueous phase containing polyvinyl alcohol (PVA) as surfactant, under stirring, and then evaporating the solvent. A 2^4-1 fractional factorial design, investigating the influence of four variables at two levels, was performed to obtain formulations with optimized association efficiencies. There was a greater dependence of association efficiency on PVA concentration (negative) and the mass of polymer (positive) with lesser influence of both stirring speed and organic phase volume. The size of the particles was assessed using scanning electron microscopy, which showed that the particles were rough-surfaced spheres. The results obtained are promising, since the formulations presented encapsulation efficiency near 25% and the release kinetics profile of atrazine was altered when it was encapsulated in the microparticles, indicating that these systems may be efficient in reducing the environmental impact caused by the herbicide, hence making it safer to use.     

Assessment of formulation parameters needed for successful vitamin C entrapped polycaprolactone nanoparticles

Vitamin C (Vit.C)-entrapped polycaprolactone (PCL) nanoparticles (Vit.C–PCNs) were prepared by encapsulation of Vit.C into PCL-based nanoparticles (PCNs) which were prepared using double emulsion method with two steps. First, the inner aqueous phase (W₁) was added to dichloromethane solution containing PCL with homogenization to form primary emulsion (W₁/O) which was emulsified with the outer aqueous phase (W₂) containing polyvinyl alcohol as stabilizer to attain the double emulsion (W₁/O/W₂). Versatile parameters were investigated to reach to the most successful formulation for Vit.C–PCNs, such as time, effect of speed of homogenization on drug encapsulation efficiency, etc.     

Liposome Microencapsulation for the Surface Modification and Improved Entrapment of Cytochrome c for Targeted Delivery

In this study, we established a procedure based on the microencapsulation vesicle (MCV) method for preparing surface‐modified liposomes, using polyethylene glycol (PEG) and a site‐directed ligand, with high entrapment efficiency of cytochrome c (Cyt c). For preparing a water‐in‐oil (W/O) emulsion, egg phosphatidylcholine and cholesterol were dissolved in organic solvents (O phase) and emulsified by sonication with aqueous solution of Cyt c (W₁). Although the dispersion stability of the W₁/O emulsion was low when n‐hexane was used to dissolve the lipids in the O phase, it was substantially improved by using mixed solvents consisting of n‐hexane and other organic solvents, such as ethanol and dichloromethane (DCM). The W₁/O emulsion was then added to another water phase (W₂) to prepare the W₁/O/W₂ emulsion. PEG‐ and/or ligand‐modified lipids were introduced into the W₂ phase as external emulsifiers not only for stabilizing the W₁/O/W₂ emulsion but also for modifying the surface of liposomes obtained later. After solvent evaporation and extrusion for downsizing the liposomes, approximately 50% of Cyt c was encapsulated in the liposomes when the mixed solvent consisting of n‐hexane and DCM at a volume ratio of 75/25 was used in the O phase. Finally, the fluorescence‐labeled liposomes, with a peptide ligand having affinity to the vasculature in adipose tissue, were prepared by the MCV method and intravenously injected into mice. Confocal microscopy showed the substantial accumulation of these liposomes in the adipose tissue vessels. Taken together, the MCV technique, along with solvent optimization, could be useful for generating surface‐modified liposomes with high drug entrapment efficiency for targeted delivery.     

Lipid Vesicle Preparation Using W/O/W Multiple Emulsions Via Solvent Evaporation: The Effect of Emulsifiers on the Entrapment Yield of Hydrophilic Materials

We studied the effects of emulsifiers on the entrapment yields of a hydrophilic material, calcein, in lipid vesicles formed using the multiple emulsion method. The primary emulsion (W₁/O) was prepared via sonication while the secondary emulsification that produced the W₁/O/W₂ multiple emulsions was achieved using the microchannel (MC) emulsification technique. The emulsifiers used in the continuous (W₂) phase were Tween® 80, Pluronic® F68, and bovine serum albumin (BSA). Lipid vesicles formed via solvent evaporation of the multiple emulsion droplets had an average diameter of about 180 nm, similar to the size of the water droplets in the primary W₁/O emulsions. The entrapment yields of calcein increased with decreasing concentrations of Tween® 80 but decreased with decreasing concentrations of Pluronic® F68 and BSA. The effects of type and concentration of emulsifier were considered in relation to three possible mechanisms: (i) destabilization/solubilization of lipid bilayers by emulsifiers, (ii) reversed-micellar transport of W₁ contents from internal to external water phases through the O phase, and (iii) release of inner water (W₁) contents into the continuous (W₂) phase via the instability of W₁/O/W₂ and leakage of W₁ contents. Using the food grade emulsifier Tween® 80 at a low concentration of about 0.1 or 0.05 wt%, calcein as a model hydrophilic material could be efficiently entrapped (ca. 80%) in homogenously dispersed lipid vesicles.     

PLGA Biodegradable Nanoparticles Containing Perphenazine or Chlorpromazine Hydrochloride: Effect of Formulation and Release

In our study, poly(dl-lactide-co-glycolide) (PLGA) nanoparticles loaded with perphenazine (PPH) and chlorpromazine hydrochloride (CPZ-HCl) were formulated by emulsion solvent evaporation technique. The effect of various processing variables, including PLGA concentration, theoretical drug loading, poly(vinyl alcohol) (PVA) concentration and the power of sonication were assessed systematically to obtain higher encapsulation efficiency and to minimize the nanoparticles size. By the optimization formulation process, the nanoparticles were obtained in submicron size from 325.5 ± 32.4 to 374.3 ± 10.1 nm for nanoparticles loaded with PPH and CPZ-HCl, respectively. Nanoparticles observed by scanning electron microscopy (SEM) presented smooth surface and spherical shape. The encapsulation efficiency of nanoparticles loaded with PPH and CPZ-HCl were 83.9% and 71.0%, respectively. The drug loading were 51.1% and 39.4% for PPH and CPZ-HCl, respectively. Lyophilized nanoparticles with different PLGA concentration 0.8%, 1.3% and 1.6% (w/v) in formulation process were evaluated for in vitro release in phosphate buffered saline (pH = 7.4) by using dialysis bags. The release profile for both drugs have shown that the rate of PPH and CPZ-HCl release were dependent on a size and amount of drugs in the nanoparticles.     

Emblicanin Rich Emblica officinalis Encapsulated Double Emulsion and its Antioxidant Stability during Storage

Emblicanin rich water‐soluble extract of Emblica officinalis (EEO) is encapsulated in the inner phase of double emulsion (DE) by using emulsifiers in different phases at different concentrations. The effects of other variables like homogenization speed, salt and herbal concentration are also investigated on various phases of DE to obtain a stable matrix. Finally, optimized EEO encapsulated DE has 2% w/w NaCl and 50% w/w EEO in inner (W₁) phase, 4% w/w polyglycerol polyricinoleate (PGPR) in middle oil‐phase and 2% w/w low‐methoxy‐pectin and reverse osmosis water in outer (W₂) phase. Ultra‐Turrax high shear homogenizer is employed to prepare primary emulsion (W₁/O) at 20 000 rpm and DE (W₁/O in W₂) at 12 000 rpm. The EEO encapsulated DE has been characterized for encapsulation efficiency (>90%), viscosity (0.715 ± 0.18 Pa s), sedimentation stability, zeta potential (?32.17 ±1.17 mV), and particle size. Light and confocal laser microscopy are used for elaborating the microscopic structure of EEO encapsulated DEs. DE has shown storage stability up to 42 days and protect antioxidant activities as compared to control (herbal extract was not encapsulated in the inner phase). The present study demonstrates that the optimized DE matrix can be used to protect the bioactive properties of EEO for its use in functional food formulation.     

Influence of process conditions on the mean size of PLGA nanoparticles

Investigation was carried out to determine the influence of process conditions on the volume mean size of PLGA nanoparticles intended as drug delivery vehicle for an injection formula of protein type therapeutic agent. Nanoparticles were produced by double emulsion-solvent evaporation method to encapsulate bovine serum albumin into the PLGA matrix material. At first aqueous BSA solution was dispersed by sonication into an organic phase composed of dichloromethane as solvent and dissolved PLGA. This first emulsion was dispersed in a second aqueous phase containing PVA emulsifier. Solid nanoparticles were obtained by evaporating the dichloromethane from the droplets. Size distribution of particles was determined by dynamic light scattering. It was found that volume mean particle size was influenced by several process variables, such as PVA concentration in the external phase, PLGA concentration in the organic phase, BSA concentration in the inner phase, volume ratio of the external and intermediate phases, and the time of sonication during the second emulsification. To elucidate their effects a 5-factorial 3-level experimental design and statistical analysis were carried out. Relationship between the mean particle size and process parameters was proposed.     

Preparation and characterization of miglitol-loaded Poly (d,l-lactide-co-glycolide) microparticles using high pressure homogenization-solvent evaporation method

Sustained release Miglitol-loaded poly (d, l-lactide-co-glycolide) (PLGA) microparticles were prepared using high pressure homogenization-solvent evaporation method. 2³ full factorial design was employed to study effect of independent variables (X₁-Polymer amount; X₂-Surfactant concentration and X₃-Homogenization Pressure) on percent encapsulation efficiency (%EE) as response. The microparticles produced were characterized for particle size, morphology, % EE, drug polymer compatibility and in vitro drug release. An average particle size of Miglitol-loaded PLGA microparticles was 230.1?nm and found almost spherical with smooth surface. % EE ranged from 58.7%?±?2.11 to 86.5%?±?0.24 depending on the polymer amount, surfactant concentration and homogenization pressure. An absence of chemical interaction between drug-polymer and reduction in % crystallinity of drug was confirmed by FTIR and X-ray diffraction analysis respectively. In vitro release studies showed a sustained release of Miglitol from microparticles up to 12?hrs.     

Preparation and characterization of a poly(vinyl alcohol)/tetraethoxysilane ultrafiltration membrane by a sol–gel method

Using poly(vinyl alcohol) (PVA) with highly hydrophilic properties as membrane material and poly(ethylene glycol) (PEG) as an additive, we prepared PVA/tetraethoxysilane (TEOS) ultrafiltration (UF) membranes with good antifouling properties by a sol–gel method. The PVA/TEOS UF membranes were characterized by X‐ray diffraction patterns, Fourier transform infrared spectroscopy, scanning electron microscopy, and static contact angle of measurement of water. The hybridization of TEOS to PVA for preparing the PVA/TEOS UF membranes achieved the required permeation performance and good antifouling behaviors. The morphology and permeation performance of the PVA/TEOS membranes varied with the different TEOS loadings and PEG contents. The pure water fluxes (J_W) increased and the rejections (Rs) decreased with increasing TEOS loading and PEG content. The PVA/TEOS UF membrane with a PVA/TEOS/PEG/H₂O composition mass ratio of 10/3/4/83 in the dope solution had a J_W of 66.5 L m^?2 h^?1 and an R of 60.3% when we filtered it with 300 ppm of bovine serum albumin aqueous solution at an operational pressure difference of 0.1 MPa. In addition, the filtration and backwashing experiment proved that the PVA/TEOS membranes possessed good long‐term antifouling abilities. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 130: 4066–4074, 2013     

Preparation of magnetic poly(lactic‐co‐glycolic acid) microspheres featuring monodispersity and controllable particle size using a microchannel device

Poly(lactic‐co‐glycolic acid) (PLGA) microspheres prepared using a traditional solvent evaporation or double emulsification method are usually polydisperse with an uncontrollable particle size distribution, which brings about poor application performance. In our research, monodisperse magnetic PLGA microspheres were prepared using a microchannel device based on a water‐in‐oil‐in‐water composite emulsion. The composite emulsion was formed by injecting a dichloromethane–gelatin water‐in‐oil emulsion into a microchannel together with an external water phase, i.e. poly(vinyl alcohol) (PVA) aqueous solution. Mean particle size control of the microspheres was executed using the osmotic pressure difference between internal and external aqueous phases caused by regulating NaCl concentration in PVA aqueous phase. It is found that monodisperse magnetic PLGA microspheres with high magnetic responsiveness can be successfully prepared combining the microchannel device with composite emulsion method. Mean particle size of the microspheres with coefficient of variation value below 4.72% is controllable from 123 to 203 µm depending on the osmotic pressure. The resulting samples have pyknotic and smooth surfaces, as well as spherical appearance. These monodisperse magnetic PLGA microspheres with good superparamagnetism and magnetic mobility have potential use as drug carriers for uniform release and magnetic targeting hyperthermia in biological fields. © 2015 Society of Chemical Industry     

Influence of Poly(L-lactic acid) Molecular Weight on the Encapsulation Efficiency of Urea in Microcapsule Using a Simple Solvent Evaporation Technique

Poly(L-lactic acid) microencapsulated urea was prepared in water-in-oil-in-water (W₁/O/W₂) system by the solvent evaporation technique. The influence of poly(L-lactic acid) molecular weight on the percent loading, encapsulation efficiency, and the microcapsule morphology was studied using poly(L-lactic acid) having different number average molecular weights (M_n). Using the higher M_n, the smoother shell with complete encapsulation microcapsules was formed. Moreover, the percent loading and encapsulation efficiency of urea also increased with the poly(L-lactic acid) molecular weight. At 80,000 g/mol of poly(L-lactic acid), the obtained microcapsule gave the highest both percent loading (32%) and encapsulation efficiency (56%). The urea control release study of the prepared microcapsules was implemented by in vitro testing. The encapsulated urea was gradually released from the microcapsules, approximately 53, 29, and 22% of poly(L-lactic acid) at 3,000, 30,000, and 80,000 g/mol, respectively, for a month. These results presented the possibility of the prepared poly(L-lactic acid) microcapsules-encapsulated urea for urea control release that could be utilized in agricultural applications.     

Fabrication of cation exchange membrane from polyvinyl alcohol using lignin sulfonic acid: Applications in diffusion dialysis process for alkali recovery

We report polyvinyl alcohol (PVA)-based hybrid membranes composed of salt of lignin sulfonic acid (LSA) and tetraethyl orthosilicate. The concentration of LSA with respect to PVA varied from 10% to 40%. The hybrid membranes showed water uptake (W_U) in the range of 122–210%, ion exchange capacities in the range of 0.32–0.75 mmol g^?1, dialysis coefficient (U_OH) from 0.0068 to 0.0119 m h^?1, and selectivity (S) from 15 to 26. The hybrid membranes also showed thermal and mechanical stability.     

Stability of multiple emulsions under shear stress

Multiple liquid emulsions of the water in oil in water (W₁/O/W₂) type are used in a variety of consumer or technical applications, for instance in the encapsulation of certain active ingredients. The encapsulation process and release mechanisms of the inner phase of the carrier drops are important in order to properly process and formulate such liquid-liquid systems. In this work the stability and breakage of multiple W₁/O/W₂ emulsions under mechanical shear stress are investigated for emulsions with different surfactants and surfactant concentrations of the internal emulsion. Stressing the emulsions in a mechanical stirring process is compared to the membrane emulsification process. The membrane emulsification process results in higher encapsulation efficiencies than the stirring process. The emulsion droplets were subjected to shear stress below and above the critical capillary number for drop breakup. The results show that stable inner emulsions with sufficient surfactant concentrations increase the overall encapsulation efficiency for multiple emulsions subjected to shear stress, although the effect is not prominent. The depletion of the carrier oil droplets could be achieved for Ca numbers below the critical limit, reducing the encapsulation efficiency below 10 %. This shows that even a low shear stress can result in content release from the internal droplet phase. The experimental emulsion release study is supported by a numerical simulation of drop deformation and break-up under shear stress.     

A transdermal diltiazem hydrochloride delivery device using multi-walled carbon nanotube/poly(vinyl alcohol) composites

Membranes with high strength and elasticity are of great demand in patch therapy. Similar membranes have been developed by combining carboxy-functionalized multiwalled carbon nanotube (c-MWCNT) with different poly(vinyl alcohol) (PVA) as potential diltiazem delivery device through aqueous mixing. High molecular weight PVA (PVA_H) produced stronger interaction with c-MWCNT than low molecular weight PVA (PVA_L) preferably at low concentration. Positive changes in favor of PVA_H in infrared and solid state ¹³C nuclear magnetic resonance spectroscopy, wide angle X-ray scattering, thermal stability, morphology and dry and wet mechanical properties clearly demonstrate that. Fibrillar c-MWCNT array at 1 wt.% in PVA_H (PVA_H/1) has drastically improved PVA crystalline cell dimension, tensile strength (201%) and elongation (196%) than neat PVA_H whereas the similar improvement is much less (100% and 185%) in PVA_L (PVA_L/1) due to globular morphology. Instead, c-MWCNT performed better at 0.5 wt.% in PVA_L (PVA_L/0.5). The kinetic data reflects better encapsulation and slower release by PVA_H (5.87%) than PVA_L (10.17%) due to greater interfacial interaction.     

Synthesis and characterization of star-shaped poly (lactide-co-glycolide) and its drug-loaded microspheres

The star-shaped poly (lactide-co-glycolide) (PLGA) was synthesized via the ring-opening polymerization of d,l-lactide and glycolide, with pentaerythritol as a multifunctional initiator and stannous 2-ethyl hexanoate as a catalyst. The structures of these polymers were characterized by ¹³C-NMR spectroscopy, while the molecular weight and polydispersity index (PDI) were determined by gel permeation chromatography (GPC). The glass transition temperature (T _g) of copolymer was determined by differential scanning calorimetry (DSC). Bovine serum albumin (BSA) loaded microspheres were fabricated using star-shaped PLGA by a W/O/W double emulsion solvent evaporation method. The results of characterization demonstrated that the particle size of the PLGA microspheres were about 80–150 μm, the maximum loading capacity and encapsulation efficiency of BSA-loaded microspheres were 67.51 μg/(mg microspheres) and 78.39%, respectively, which were better than linear PLGA. The in vitro release profiles of BSA in phosphate buffer saline (PBS) lasted for 37 h. Drug release profiles can be affected by polymer molecular weight and the ratio of polymer to drug. The maximum release percentage was 80%.     

膜乳化法与复乳法结合制备粒径均一的载溶菌酶微胶囊

采用微孔膜乳化法与复乳法结合制备粒径均一可控的以聚乳酸和聚(乳酸-羟基乙酸)共聚物为膜材的载溶菌酶微胶囊,粒径分布系数CV(Coefficient of Variation)为14.04%,远低于机械搅拌法制备的微囊的CV(76.54%). 分别加入内水相添加剂PVA, PEG400, HP-b-CD,使溶菌酶的包埋率从无添加剂时的68.1%分别增大到86.6%, 89.0%和94.1%. 添加剂降低了溶菌酶的突释. PEG400, PEG6000, HP-b-CD的加入降低了溶菌酶的释放速率,而PVP或PVA的加入则加快了溶菌酶的释放. 溶菌酶在油水界面上的吸附变性是失活的主要原因. 在酶液中加入PEG400, PEG6000, PVP, HP-b-CD可有效地避免由于油水界面造成的溶菌酶活性的损失.