Encapsulation of Lipophilic Bioactive Molecules by Microchannel Emulsification

Currently, much effort is being invested in novel formulations of bioactive molecules, such as emulsions, for pharmaceutical, food, and cosmetic applications. Therefore, methods to produce emulsions with controlled-size droplets of uniform size distribution have been developed. On this concern, a microfluidic device called the microchannel (MC) was used in this work for emulsification. This is a novel method for producing monodispersed emulsion droplets with very narrow droplet size distribution and low energy input, due to the spontaneous droplet generation basically driven by the interfacial tension, unlike other conventional emulsification processes. This technology provides the formulation of oil-in-water (O/W) emulsions containing lipophilic active molecules with increased bioavailability, which may be readily absorbed by the human body. MC emulsification enables the preparation of highly monodispersed O/W emulsions, which may be applied as enhancer on active molecules delivery systems, as well as in foodstuff. In this study, formulations of O/W emulsions loaded with bioactive molecules, such as β-carotene and γ-oryzanol, were prepared by the MC emulsification process. Refined soybean oil containing the dissolved lipophilic molecule and either sugar ester or gelatin solution (1 wt.%) were used as the dispersed and continuous phases, respectively. The emulsification process conducted using the asymmetric straight-through MC plate enabled the production of monodispersed O/W emulsions, resulting in β-carotene-loaded O/W emulsions with average droplet size (d _av) of 27.6 μm and coefficient of variation (CV) of 2.3% and γ-oryzanol-loaded droplets with d _av of 28.8 μm and CV of 3.8%. The highly monodisperse β-carotene-loaded droplets were physically stable throughout the storage period observed, resulting in droplets with d _av 28.2 μm and CV of 2.9% after 4 months storage in darkness at 5 °C. Single micrometer-sized monodisperse emulsions loaded with β-carotene were successfully formulated using the grooved MC emulsification, resulting in droplets with d _av of 9.1 μm and CV of 6.2%. This work was funded by The Ministry of Agriculture, Forestry and Fisheries of Japan, through the Food Nanotechnology Project, and the Japan Society for the Promotion of Science.     

Preparation and characterizations of self-assembled PEGylated gelatin nanoparticles

The PEGylated gelatin nanoparticles were prepared by self-assembling method and characterized. The gelatin drug carrier was proposed as a targeting drug delivery system with the hypothesis that the gelatin carrier could be degraded by the matrix metalloprotease (MMP) and release the anticancer drug loaded inside carriers around the cancer site. The gelatin nanoparticles proposed in this study were composed of deoxycholic acid (DOCA), monomethoxy polyethylene glycol (MPEG), and gelatin. The carboxyl groups of DOCA and carboxylated MPEG were coupled with amine group of gelatin by dichlorohexylcarbodiimide (DCC) method. One molecule of gelatin coupled with 205 molecules of MPEG and 275 molecules of DOCA. The synthesized gelatin/DOCA/MPEG conjugates (GDM) were ultrasonicated to produce self-assembled nanoparticles. DOCA acted as the hydrophobic core, thereby aggregating gelatin molecules and hydrophilic MPEG chains located at the surface of the nanoparticles. The concentration of GDM, intensity of sonication, sonication time and temperature, all affected to control the particle size in the ultrasonication. The optimum condition was obtained as 1.0 mg/mL of GDM, 28 W for sonication intensity, 3 min of sonication time, and room temperature. The size distribution of particle was found to be 100–1000 nm in this condition. The particles which had a broad size distribution were filtered by 0.2 μm membrane. The product yield of particles having below 200 nm of size was about 30%. After filtration, an average diameter of GDM nanoparticle was 176 nm (155–200 nm).     

Agarose–gelatin conjugate for adherent cell-enclosing capsules

Spherical capsules were prepared by extruding aqueous agarose–gelation conjugate solution into co-flowing liquid paraffin at 38°C and cooling the resultant emulsion. Capsule diameter was controlled between 40 and 250 μm by changing the velocity of the liquid paraffin. Adherent Crandall–Reese feline kidney cells enclosed in conjugate capsules of 141 ± 23 μm diam. had a higher degree of proliferation than those in unmodified agarose capsules. Mitochondrial activity, detected for cell-enclosing conjugate capsules normalized against unit volume of gel, was about double that of unmodified agarose capsules over 28 days. These results demonstrated the feasibility of agarose–gelatin conjugate as a material of cell-enclosing capsules.     

Small microcapsules of crystal proteins and spores of Bacillus thuringiensis by an emulsification/internal gelation method

This paper describes a microencapsulation process of a spore crystal aggregate produced by Bacillus thuringiensis var. kurstaki HD-1. The methodology is based on the emulsification/internal gelation method, and was implemented to produce microcapsules of small diameter (<10 μm) with the capacity to protect the spore crystal aggregate from extreme ultraviolet radiation. The diameter of microcapsules was in the range of 3.1 ± 0.2–6.8 ± 0.4 μm, which is considered adequate for biological control purposes. The protective effect of the alginate coat was verified by the remaining 60 ± 2% and 40 ± 1% of spore viability and protein activity, respectively, after UV-B radiation of 236 J, and with bioassays with Spodoptera frugiperda. It is expected that the protective effect of the alginate coat will improve the effectiveness of the Bt-HD1 formulated as small diameter microcapsules, and their yield, once they are released into the environment, will also be improved.     

Development of Spray Dried Liposomal Dry Powder Inhaler of Dapsone

This investigation was undertaken to evaluate practical feasibility of site specific pulmonary delivery of liposomal encapsulated Dapsone (DS) dry powder inhaler for prolonged drug retention in lungs as an effective alternative in prevention of Pneumocystis carinii pneumonia (PCP) associated with immunocompromised patients. DS encapsulated liposomes were prepared by thin film evaporation technique and resultant liposomal dispersion was passed through high pressure homogenizer. DS nano-liposomes (NLs) were separated by ultra centrifugation and characterized. NLs were dispersed in phosphate buffer saline (PBS) pH 7.4 containing different carriers like lactose, sucrose, and hydrolyzed gelatin, and 15% l-leucine as antiadherent. The resultant dispersion was spray dried and spray dried formulation were characterized to ascertain its performance. In vitro pulmonary deposition was assessed using Andersen Cascade Impactor as per USP. NLs were found to have average size of 137 ± 15 nm, 95.17 ± 3.43% drug entrapment, and zeta potential of 0.8314 ± 0.0827 mV. Hydrolyzed gelatin based formulation was found to have low density, good flowability, particle size of 7.9 ± 1.1 μm, maximum fine particle fraction (FPF) of 75.6 ± 1.6%, mean mass aerodynamic diameter (MMAD) 2.2 ± 0.1 μm, and geometric standard deviation (GSD) 2.3 ± 0.1. Developed formulations were found to have in vitro prolonged drug release up to 16 h, and obeys Higuchi's Controlled Release model. The investigation provides a practical approach for direct delivery of DS encapsulated in NLs for site specific controlled and prolonged release behavior at the site of action and hence, may play a promising role in prevention of PCP.     

Preparation of a Water-in-oil-in-water (W/O/W) Type Microcapsules by a Single-droplet-drying Method and Change in Encapsulation Efficiency of a Hydrophilic Substance during Storage

Microcapsules of a water-in-oil-in-water (W/O/W) emulsion, which contained a hydrophilic substance, 1,3,6,8-pyrenetetrasulfonic acid tetrasodium salt (PTSA), in its inner aqueous phase, was prepared by hot-air-drying or freeze-drying the emulsion using a single-droplet-drying method. Pullulan, maltodextrin, or gum arabic was used as a wall material, and the oily phase was tricaprylin, oleic acid, olive oil, or a mixture of tricaprylin and olive oil. An encapsulation efficiency higher than 0.95 was reached except for the microcapsules prepared using gum arabic and oleic acid. The hot-air-dried microcapsules were generally more stable than the freeze-dried microcapsules at 37°C and various relative humidities. The stability was higher for the microcapsules with tricaprylin as the oily phase than for the microcapsules with oleic acid. The higher stability of the microcapsules with tricaprylin would be ascribed to the lower partition coefficient of PTSA to the oily phase. There was a tendency for the stability to be higher at lower relative humidity for both the hot-air- and freeze-dried microcapsules. The volumetric fraction of olive oil in its mixture with tricaprylin did not significantly affect either the encapsulation efficiency or the stability of the hot-air-dried microcapsules.     

Activated sludge encapsulation in gellan gum microbeads for gasoline biodegradation

A two-phase dispersion technique, termed emulsification–internal gelation, is proposed for encapsulation of activated sludge in gellan gum microbeads. The influence of emulsion parameters on size distribution of microbeads was investigated. Mean diameter of microbeads varied within a range of 34–265 µm as a descending function of emulsion stirring rate (1,000–5,000 rpm), emulsification time (10–40 min), and emulsifier concentration (0–0.1% w/w), and as an ascending function of disperse phase volume fraction (0.08–0.25). Encapsulated sludge expressed a high biodegradation activity compared with non-encapsulated sludge cultures even at 4.4 times lower level of overall biomass loading. Over 90% of gasoline at an initial concentration of 35 and 70 mg l^–1 was removed by both encapsulated and non-encapsulated sludge cultures in sealed serum bottles within 7 days. Encapsulation of activated sludge in gellan gum microbeads enhanced the biological activity of microbial populations in the removal of gasoline hydrocarbons. The results of this study demonstrated the feasibility of the production of size-controlled gellan gum-encapsulated sludge microbeads and their use in the biodegradation of gasoline.     

膜材与制备过程对血红蛋白微胶囊粒径和包埋率的影响

以单甲氧基聚乙二醇聚乳酸共聚物（PELA）为膜材用复乳溶剂扩散法制备了包含牛血红蛋白（BHb）的微胶囊,微胶囊中BHb的P₅₀和Hill系数分别为3 466 Pa和2.4左右,接近于天然BHb的生物活性。研究发现膜材种类对BHb微胶囊包埋率和粒径的影响最大,使用MPEG2000为亲水性嵌段的PELA共聚物时,包埋率最高,达到90％以上,粒径为3～5 μm左右;随着膜材浓度的增大,微胶囊包埋率和粒径均增加;随着外水相NaCl浓度的增大,微胶囊包埋率升高、粒径减小;随着外水相稳定剂PVA浓度的增大,微胶囊粒径减小,包埋率先升高后降低,在较低浓度下（10 g/L、20 g/L）包埋率较高;初乳化搅拌速率的增大,有利于包埋率的提高,但对粒径影响不大;复乳化搅拌速率的影响较复杂,当复乳液体积较大时,复乳化搅拌速率对微胶囊制备的影响规律性不明显。当固定膜材和初乳化搅拌速率时,包埋率和粒径之间存在着类似抛物线的关系,包埋率随着粒径的减小而降低。     

Effect of Thermal Treatment on the Physicochemical Properties of Emulsion Stabilized by Gelatin from Black Tilapia (Oreochromis mossambicus) Skin

This study investigated the influence of thermal treatment (30 °C to 110 °C, 30 min) on the physicochemical and rheological properties of an emulsion stabilized by black tilapia (Oreochromis mossambicus) skin at pH 4. The protein pattern of tilapia gelatin did not have any significant difference after the gelatin was heated within a temperature range of 30 °C to 70 °C. However, at 90 °C and 110 °C, denaturation occurred where α-, β- and γ-chains of the gelatin were degraded, leading to a concomitant increase in low molecular peptides. The emulsion stability was investigated through a particle size analyzer, zeta potential, microscopic observation and creaming index. The gelatin emulsion was physically stable at 30 °C to 70 °C with a mean droplet size of less than 13 μm. When the heating temperature was increased to 90 °C and 110 °C, the emulsion showed a pronounced increase in droplet size due to coalescence. The gelatin emulsion heated at 90 °C and 110 °C also displayed instability against creaming after storage at room temperature for 7 days. As the heating temperature increased, the gelatin emulsion exhibited a decrease in apparent viscosity and the flow behavior changed from shear thinning to Newtonian. The rheological data also showed that the storage modulus (G′) of emulsion became more frequency dependent as the heating temperature increased, indicating weak droplet interactions. The tilapia gelatin emulsion was physically unstable when subjected to thermal treatment above 70 °C. The data reported in this study provides useful insight into the formulation of acidic food emulsions that require thermal treatment.

     

Formation of nano-hydroxyapatite in gelatin droplets and the resulting porous composite microspheres

A one-pot strategy was first presented in this paper to synthesize gelatin/hydroxyapatite (HAP) composite microspheres in a water-in-oil (W/O) emulsion. Using gelatin droplets as microreactors and colloid protective medium, needle-like nano-HAP crystals (5 nm x 60-100 nm) in form of clusters were homogeneously and orderly precipitated within gelatin matrix. The results of scanning electron microscopy (SEM) revealed that the as-prepared microspheres with an average diameter of 7.5 microm displayed a narrow particle size distribution, a high dispersity and a naturally porous structure. This microsphere material is expected to have a great potential for both controlled drug release and faster bone in-growth in bone tissue engineering.     

Preparation of a water-in-oil-in-water (W/O/W) type microcapsules by a single-droplet-drying method and change in encapsulation efficiency of a hydrophilic substance during storage

Microcapsules of a water-in-oil-in-water (W/O/W) emulsion, which contained a hydrophilic substance, 1,3,6,8-pyrenetetrasulfonic acid tetrasodium salt (PTSA), in its inner aqueous phase, was prepared by hot-air-drying or freeze-drying the emulsion using a single-droplet-drying method. Pullulan, maltodextrin, or gum arabic was used as a wall material, and the oily phase was tricaprylin, oleic acid, olive oil, or a mixture of tricaprylin and olive oil. An encapsulation efficiency higher than 0.95 was reached except for the microcapsules prepared using gum arabic and oleic acid. The hot-air-dried microcapsules were generally more stable than the freeze-dried microcapsules at 37 degrees C and various relative humidities. The stability was higher for the microcapsules with tricaprylin as the oily phase than for the microcapsules with oleic acid. The higher stability of the microcapsules with tricaprylin would be ascribed to the lower partition coefficient of PTSA to the oily phase. There was a tendency for the stability to be higher at lower relative humidity for both the hot-air- and freeze-dried microcapsules. The volumetric fraction of olive oil in its mixture with tricaprylin did not significantly affect either the encapsulation efficiency or the stability of the hot-air-dried microcapsules.     

Hardly water-soluble drug-loaded gelatin nanoparticles sustaining a slow release: preparation by novel single-step O/W/O emulsion accompanying solvent diffusion

Bioprocess and Biosystems Engineering - Paclitaxel (PTX)-loaded gelatin nanoparticles (NPs) were prepared, for the first time, by novel O/W/O emulsion with a single-step emulsion process...     

Stability of CoQ10-Loaded Oil-in-Water (O/W) Emulsion: Effect of Carrier Oil and Emulsifier Type

Co-enzyme Q10 (CoQ10), a lipophilic compound that widely used in the food and pharmaceutical products was formulated in a κ-carrageenan coated oil-in-water (O/W) emulsion. In this work, we examined the solubility of CoQ10 in different carrier oils and effects of emulsifier type on the formation and stability of CoQ10-loaded O/W emulsion. Nine vegetable oils and four types of emulsifiers were used. CoQ10 was found significantly (p?<?0.05) more soluble in medium chain oils (coconut oil and palm kernel oil) as compared to other vegetable oils. The O/W emulsions were then prepared with 10 % (w/w) coconut oil and palm kernel oil containing 200 g CoQ10/L oil stabilized by 1 % (w/v) emulsifiers (sucrose laurate (SEL), sodium stearoyl lactate (SSL), polyglycerol ester (PE), or Tween 80 (Tw 80)) in 1 % (w/v) κ-carrageenan aqueous solution. Particle size distribution and physical stability of the emulsions were monitored. The droplet sizes (surface weighted mean diameter, D[3,2]) of fresh O/W emulsion in the range of 2.79 to 5.83 μm were observed. Irrespective of the oil used, results indicated that complexes of SSL/κ-carrageenan provided the most stable CoQ10-loaded O/W emulsion with smaller and narrower particle size distribution. Both macroscopic and microscopic observations showed that O/W emulsion stabilized by SSL/κ-carrageenan is the only emulsion that exhibited no sign of coalescence, flocculation, and phase separation throughout the storage period observed.     

Permeation and Distribution of Ferulic Acid and Its α-Cyclodextrin Complex from Different Formulations in Hairless Rat Skin

Ferulic acid (FA) is a natural product that occurs in seeds of many plants where it is generally located in the bran. This compound is a multifunctional ingredient endowed with antioxidative, radical scavenging, sunscreening and antibacterial actions. The aim of this study was to analyse the ferulic acid cutaneous permeation and distribution, through and into the skin layers, from different cosmetic vehicles, an O/W emulsion (pH 6.0) and two gel-type formulations at different pH levels (6.0 and 7.4), containing FA alone or an inclusion complex with α-cyclodextrin (CD-FA). In vitro permeation studies were performed in vertical diffusion cells using hairless rat excised skin. At appropriate intervals of time, the amount of permeated sunscreen/radical scavenger was evaluated by high-performance liquid chromatography (HPLC). At the end of experiments, treated skin samples were sectioned with a cryomicrotome and the FA content of the individual slices was analysed by HPLC. FA-containing formulations, O/W emulsion, gels A and B, originated FA fluxes of 8.48 ± 2.31, 8.38 ± 0.89 and 5.72 ± 0.50 μg/cm² h, respectively, thus suggesting the pH influence on FA percutaneous permeation. The use of the inclusion complex, CD-FA, determined in all cases a decrease of FA transdermal permeation while no influence of pH was observed. Gel-type formulations containing FA ensured higher sunscreen storage in the superficial layers if compared with O/W emulsion. When FA was included in α-cyclodextrin, FA amount retained into skin layers decreased markedly.     

Ethyl Cellulose Microcapsules for Protecting and Controlled Release of Folic Acid

Ethyl cellulose microcapsules were developed for use as a drug-delivery device for protecting folic acid from release and degradation in the undesirable environmental conditions of the stomach, whilst allowing its release in the intestinal tract to make it available for absorption. The controlled release folic acid-loaded ethyl cellulose microcapsules were prepared by oil-in-oil emulsion solvent evaporation using a mixed solvent system, consisting of a 9:1 (v/v) ratio of acetone:methanol and light liquid paraffin as the dispersed and continuous phase. Span 80 was used as the surfactant to stabilize the emulsion. Scanning electron microscopy revealed that the microcapsules had a spherical shape. However, the particulate properties and in vitro release profile depended on the concentrations of the ethyl cellulose, Span 80 emulsifier, sucrose (pore inducer), and folic acid. The average diameter of the microcapsules increased from 300 to 448 μm, whilst the folic acid release rate decreased from 52% to 40%, as the ethyl cellulose concentration was increased from 2.5% to 7.5% (w/v). Increasing the Span 80 concentration from 1% to 4% (v/v) decreased the average diameter of microcapsules from 300 to 141 μm and increased the folic acid release rate from 52% to 79%. The addition of 2.5–7.5% (w/v) of sucrose improved the folic acid release from the microcapsules. The entrapment efficiency was improved from 64% to 88% when the initial folic acid concentration was increased from 1 to 3 mg/ml.     

Preparation of stimulus responsive multiple emulsions by membrane emulsification using con a as biochemical sensor

In this work, a novel strategy for the controlled fabrication of biomolecular stimulus responsive water-in-oil-in-water (W/O/W) multiple emulsion using the membrane emulsification process was investigated. The emulsions interface was functionalized with a biomolecule able to function as a receptor for a target compound. The interaction between the biomolecular receptor and target stimulus activated the release of bioactive molecules contained within the structured emulsion. A glucose sensitive emulsion was investigated as a model study case. Concanavalin A (Con A) was used as the biomolecular glucose sensor. Various physicochemical strategies for stimulus responsive materials formulation are available in literature, but the preparation of biomolecule-responsive emulsions has been explored for the first time in this paper. The development of novel drug delivery systems requires advanced and highly precise techniques to obtain their particular properties and targeting requirements. The present study has proven the flexibility and suitability of membrane emulsification for the preparation of stable and functional multiple emulsions containing Con A as interfacial biomolecular receptor able to activate the release of a bioactive molecule as a consequence of interaction with the glucose target molecule. The influence of emulsion interfacial composition and membrane emulsification operating conditions on droplets stability and functional properties have been investigated. The release of the bioactive molecule as a function of glucose stimulus and its concentration has been demonstrated.     

Measurement of particle diameter of <Emphasis Type="Italic">Lactobacillus acidophilus</Emphasis> microcapsule by spray drying and analysis on its microstructure

Lactobacillus acidophilus, as a probiotic, is widely used in many functional food products. Microencapsulation not only increases the survival rate of L. acidophilus during storage and extends the shelf-life of its products, but also optimal size microcapsule makes L. acidophilus have an excellent dispersability in final products. In this paper, L. acidophilus was microencapsulated using spray drying (inlet air temperature of 170°C; outlet air temperature of 85–90°C). The wall materials used in this study were β-cyclodextrin and acacia gum in the proportion of 9:1 (w/w), and microcapsules were prepared at four levels of wall materials (15, 20, 25 and 30% [w/v]) with a core material concentration of 6% (v/v). The microcapsule diameters were measured by Malvern’s Mastersizer-2000 particle size analyzer. The results showed that the particle diameters of microcapsule were mostly within 6.607 μm and 60.256 μm and varied with 2.884–120.226 μm (the standard smaller microcapsule designated as <350 μm). Through comparison of microcapsule size and uniformity with different concentration of wall materials, we concluded that the optimal concentration of wall material was 20% (w/v), which gave microcapsule with a relatively uniform size (averaging 22.153 μm), and the number of surviving encapsulated L. acidophilus was 1.50 × 10⁹ c.f.u./ml. After 8 weeks storage at 4°C, the live bacterial number was above 10⁷ c.f.u./ml, compared with unencapsulated L. acidophilus, 10⁴–10⁵ c.f.u./ml. Through the observation of scanning electron microscopy, we found that the shapes of microcapsule were round and oval, and L. acidophilus cells located in the centre of microcapsule.     

Injectable Polysaccharide Microcapsules for Prolonged Release of Minocycline for the Treatment of Periodontitis

Injectable polysaccharide microcapsules holding minocycline were fabricated from alginate and chitosan for the treatment of periodontitis. The microcapsules were examined for the release and degradation of minocycline, as well as antimicrobial activity. The microcapsules were biodegradable and released minocycline between 10 and 1000 μg ml⁻¹, which was higher than the usual therapeutic concentration (1–5 μg ml⁻¹), for up to 7 days. These microcapsules showed a statistically significant suppression of pathogenic bacteria, such as Prevotella intermedia causing periodontitis. The microcapsules are thus potentially useful for drug delivery for the treatment of periodontitis.     

Reproduction and larval development of the gastropod molluskTegula rustica in Peter the Great Bay, Sea of Japan

The reproduction and larval development of the prosobranch gastropod molluskTegula rustica were studied under laboratory conditions. In Vostok Bay (Peter the Great Bay), the reproduction of theTegula takes place in August, when the water temperature is 19–20°C. Under laboratory conditions, the spawning of females was stimulated by adding a sperm emulsion to a vessel containing adult females. We observed asynchronous and intermittent release of gametes. The egg cell is 190 μm in diameter. Fertilization is external, and the course of development includes a lecithotrophic pelagic larva. Complete development, from fertilization to metamorphosis, takes 7 days in a laboratory culture. The larvae settle when the shell size across the first whorl is 220–230 μm and the total shell whorl is about 90°. The shape of the veliger shell is elongated mitriform and the velum is rounded, made up of a single lobe. The sculpture of the protoconch is irregularly ribbed.     

CHO immobilization in alginate/poly-l-lysine microcapsules: an understanding of potential and limitations

Microencapsulation offers a unique potential for high cell density, high productivity mammalian cell cultures. However, for successful exploitation there is the need for microcapsules of defined size, properties and mechanical stability. Four types of alginate/poly-l-Lysine microcapsules, containing recombinant CHO cells, have been investigated: (a) 800 μm liquid core microcapsules, (b) 500 μm liquid core microcapsules, (c) 880 μm liquid core microcapsules with a double PLL membrane and (d) 740 μm semi-liquid core microcapsules. With encapsulated cells a reduced growth rate was observed, however this was accompanied by a 2–3 fold higher specific production rate of the recombinant protein. Interestingly, the maximal intracapsular cell concentration was only 8.7 × 10⁷ cell mL^-1, corresponding to a colonization of 20% of the microcapsule volume. The low level of colonization is unlikely to be due to diffusional limitations since reduction of microcapsule size had no effect. Measurement of cell leaching and mechanical properties showed that liquid core microcapsules are not suitable for continuous long-term cultures (>1 month). By contrast semi-liquid core microcapsules were stable over long periods with a constant level of cell colonization (ϕ = 3%). This indicates that the alginate in the core plays a predominant role in determining the level of microcapsule colonization. This was confirmed by experiments showing reduced growth rates of batch suspension cultures of CHO cells in medium containing dissolved alginate. Removal of this alginate would therefore be expected to increase microcapsule colonization.