Development of vegetable extracts by microencapsulation

The microencapsulation of essential oils offers protection against oxidation and evaporation, and allows the concurrent utilization of several vegetable extracts. Complex coacervation methods have been described for essential oils. Even though microencapsulation involves wrapping the essential oils in shells, some difficulties arise in the process of stabilizing the essential oils: oil may be lost by evaporation and partial dissolution in the water-gelatin phase and this will vary with the type of essential oil being encapsulated. In order to investigate the efficacy of the gelatin-polyphosphate methods we analysed their essential oil microcapsules peppermint and rosemary, in particular their granulometric size distribution, oil content (%) and encapsulation yield (%). In addition the essential oils were analysed by GC before and after microencapsulation so as to investigate the loss of their components during the process.     

Microencapsulation in yeast cells

A method for encapsulating high concentrations of essential oils into bakers' yeast (Saccharomyces cerevisiae) is described. The process involves mixing an aqueous suspension of yeast and an essential oil, which allows the oil to pass freely through the cell wall and membrane and remain passively within the cell. Oil droplets sequestered within the cell were clearly visible using confocal microscopy. Transmission electron microscopy demonstrated that the cell wall and membrane remain intact during the process. Cells quickly lost viability during the process and it appeared unnecessary for the cells to be viable for the process to occur. Encapsulated oil was recovered from the cells using a water/ethanol extraction procedure and analysed by gas chromatography. No significant differences were noted between encapsulated and unencapsulated oil profiles. The rate of permeation of oil into the yeast cells was found to increase significantly at higher temperatures due to the phase transition of the lipid membrane. The rates at which different essential oils permeated the cell varied considerably due to variations in terpene chemistry. The encapsulation of straight chain hydrocarbons highlighted the effects of molecular size, shape and the presence of hydroxl groups on the process. The process occurs by passive diffusion as a result of hydrophobic flavour components partitioning into the cell membrane and intracellular lipid. This paper briefly reviews the patented literature and reports some of the initial observations of the transport mechanisms involved during the accumulation of essential oils by yeast cells.     

Diminished kidney function and nephrocalcinosis in rats fed a magnesium-deficient diet

Microcapsules have been used as drug delivery systems in the pharmaceutical field for sustained or controlled release of drug, and for artificial cells and organs. Biodegradable polymers, especially polylactic acid, have been widely used in this field. In this study, an attempt was made to develop a new method to prepare polylactic acid microcapsules for drug delivery. The biodegradable polylactic acid microcapsules were made by the phase separation process: two types of polylactic acid, poly[(D,L)lactic acid] and poly[(L)lactic acid] were combined as the membrane material. Because of the difference of the crystal properties of the two polymers, the aggregation which happens frequently in the phase separation process was prevented. As a model drug, Ciprofloxacin was encapsulated in the polylactic acid microcapsules.     

Biocompatibility and immunology in the encapsulation of islets of Langerhans (bioartificial pancreas)

Successful transplantation of encapsulated islets (bioartificial pancreas) would circumvent problems of islet availability and rejection in the treatment of insulin-dependent diabetes with biological organ replacement. Alginates are widely used as a hydrogel matrix or membrane for immunoprotected transplantation. A major problem in the use of diffusion-based devices is the biocompatibility of the material used. The foreign body reaction after implantation of empty microcapsules into different compartments in rats, dogs and pigs is evaluated in this article. However, biocompatibility of the bioartificial pancreas has three different aspects: reaction of the entrapped islet to the encapsulation technique and material; reaction of the recipient against the incorporated device ( = foreign body reaction); and finally the reaction of the recipient against the encapsulated islet ( = immunology of bioartificial pancreas). It is obvious from different experiments that even if foreign body reactions (reactions against material) are almost abolished the recipient may react against material released from the encapsulated islet. In conclusion, transplantation of encapsulated islets induces various morphological reactions (i.e. inflammation and fibrosis) as a result of a variety of donor and recipient related factors. Therefore, the use of an adequate animal model that reflects the human situation is essential for progress in the development of a bioartificial pancreas.     

Antilisterial activity of pediocin AcH in model food systems in the presence of an emulsifier or encapsulated within liposomes

The listericidal activity of pediocin AcH was evaluated in slurries (5, 10, or 25% in dH2O) of nonfat dry milk, butterfat, beef muscle tissue, or beef tallow. Slurries were inoculated with Listeria monocytogenes (2-strain mixture; 2.5 x 10(6) cfu/ml) and then with pediocin AcH (30,000 arbitrary units (AU) per ml of slurry). Although pediocin activity was reduced in slurries, sufficient pediocin remained to decrease the listeriae population. For all slurries tested, the greatest decrease in counts of Listeria (1.2-1.8 log10 cfu decrease) and decrease in pediocin activity (12-54% recovery of original activity) occurred within 1.5 min of addition of pediocin to slurries. Thereafter, counts of Listeria did not change appreciably, but pediocin activity continued to decrease in most treatments for up to 60 min. In general, greater activity was recovered from: (i) slurries of lower (5%) compared to higher (25%) concentrations of food; and (ii) dairy- compared to meat-based slurries. Next, pediocin AcH was encapsulated within phosphatidyl-choline-based liposomes before addition to slurries (10%), or was used unencapsulated in slurries (10%) containing the emulsifier Tween 80. Greater pediocin activity (29-62% increase; average over all concentrations) was recovered from slurries containing encapsulated compared to free pediocin AcH. Likewise, greater pediocin activity was recovered from slurries containing an emulsifier (4-90% increase; average over all concentrations) compared to otherwise similar slurries without Tween 80. The additional recovery of pediocin activity afforded by liposomes or Tween 80 underscores the potential for direct application of biopreservatives to provide another hurdle for L. monocytogenes in foods.     

Survey of disease of cultural shrimp in Taiwan

A water-in-oil-in-water (w/o/w) technique, sometimes known as in-water drying method, was used to prepare microcapsules consisting of polylactic acid and poly(lactide-co-glycolide). The influence of shear force to produce an initial water-in-oil (w/o) emulsion on the characteristics of microcapsules and protein release was investigated. Bovine serum albumin (BSA) was used as the model protein drug for encapsulation. The initial w/o emulsion was prepared by a Polytron homogenizer. The shear rate was varied from 11 to 23 krpm to produce w/o emulsions with different shear forces. This study revealed pronounced effects of shear force on the characteristics of microcapsules and release profiles of BSA. Depending on the degree of the shear applied, the inner structure of microcapsules showed very different morphology, which was responsible for different release patterns. A low shear produced microcapsules with a high initial burst release of BSA, whereas microcapsules using a high shear exhibited a controlled release of protein without any initial burst release. Also, at a given shear force, a variation in polymer composition of microcapsules was found to be effective in controlling the release characteristics of protein. Thus, the homogenization technique should be carefully considered in designing microcapsules with desirable release profiles of proteins and an adequate period of protein delivery.     

Protective effect of liposome encapsulation on paclitaxel developmental toxicity in the rat

Paclitaxel is an anticancer drug that has demonstrated severe embryotoxicity in chicks. This embryotoxicity is reduced by liposome encapsulation of the drug. The current study was designed to evaluate the potential of liposome encapsulation for reducing paclitaxel embryotoxicity in rats. Wistar rats were treated with paclitaxel on day 8 of pregnancy (plug = day 0) at doses of 0.67, 2.0, or 10.0 mg/kg intravenously. The same doses of paclitaxel encapsulated in liposomes were administered intravenously to other groups of animals. Control animals were given blank liposomes. Free paclitaxel produced maternal and embryotoxicity at 10.0 mg/kg with three of seven dams dying and resorption of all embryos in surviving dams. Liposome encapsulation at 10.0 mg/kg was not associated with maternal death and there were live fetuses on evaluation at term, although litter size was reduced and malformations occurred in surviving fetuses. At 2.0 mg/kg free paclitaxel, fetal weight was decreased and resorptions increased. Liposome encapsulation at 2.0 mg/kg produced litter results similar to those obtained in control animals given empty liposomes. Malformations were prominent at 2.0 mg/kg free paclitaxel and at 10.0 mg/kg paclitaxel in liposomes and included exencephaly/anencephaly, ventral wall defects, facial clefts, anophthalmia, diaphragmatic hernia, and defects of the kidney, cardiovascular system, and tail. Liposome encapsulation appeared to shift the developmental response to paclitaxel such that 10 mg/kg encapsulated drug produced effects similar to 2.0 mg/kg free drug. These results may have implications for drug delivery of therapeutic agents used during human pregnancy.     

Granulocyte colony-stimulating factor activates a 72-kDa isoform of STAT3 in human neutrophils

A series of dextran molecular weight markers were encapsulated in decylamine carboxymethylcellulose microcapsules to serve as probes of capsule retentivity. The capsules were prepared by allowing microdrops of aqueous sodium carboxymethylcellulose to fall into aqueous decylamine acetate solution. Salt exchange reaction at the droplet pseudointerface resulted in self-assembling films which essentially instantaneously enclosed the droplets. Concentrations of anionic polymer were varied in the range from 1-3%. Chromophore-bearing dextrans were incorporated into these capsules by blending the dextrans with the carboxymethylcellulose prior to the encapsulation step. Four dextrans of differing (light scattering) molecular weights were used: 2 x 10(6), 6 x 10(5), 7 x 10(4), and 1.9 x 10(4) amu. The mass balance of dextran retained in the capsules, released on washing the capsules or which escaped encapsulation was determined spectrophotometrically. To measure total dextran in a population of washed capsules, the capsules were lysed in a 0.3 M solution of sodium chloride. To monitor dextran release, washed capsules were suspended in water and dextran concentration in the supernatant was measured. Encapsulation efficiency exceeded 80% for high molecular weight dextran but was lower with the smaller dextrans.     

Fatty acids in 80 brands of edible vegetable oil and in 14 brands of mayonnaise (author's transl)

The fatty acid patterns have been defined in 80 commonly used table oils chosen at random and of 14 brands of mayonnaise commercially available in West Germany. In addition, sterols of the various mayonnaises were analysed. The oils could be grouped into 8 different categories: 12 oliver oils, 8 maize oils, 14 soya oils, 5 linseed oils, 22 sunflower oils, 3 safflower oils, 5 groundnut oils, and 8 rapeseed oils. As a result of the analyses, it is in almost every case a question of a pure variety of non-animal oils. The range of the values obtained for the composition of the fatty acids for each variety, which can be considerable, represents a modern complement to previous analyses. The fatty acid pattern of 3 additional commerical table oils suggest an amalgamation with fats of other types, among which are 2 cases of mixture with rapeseed oils. For mayonnaise, soya oil is used almost exclusively as a fat component. The presence of sunflower oil was only confirmed in one brand of mayonnaise. The concentrations of cholesterol which were determined lie between 17.0 and 72.3 mg (average 53 mg) per 100 g mayonnaise, so that even in the case of persons which present an elevated risk of atherosclerosis there can be no objection to the consumption of mayonnaise. The nutritive-physiological significance of these various oils for human nutrition is thoroughly discussed. A possible health hazard is the sale of vegetable oils rich in erucic acid permitted in the West German Republic (proportion of erucic acid in the 8 samples examined: 223-53%). These vegetable oils, as is shown by the additionally indicated trade names with the corresponding supplementary specifications are overwhelmingly labelled as vegetable oil (Pflanzen?l) or table oil (Speise?l). The purchase prices indicated and paid by us for the oils examined are subject to considerable fluctuations even for oils of the same quality.     

Molten globule intermediate of recombinant human growth hormone: stabilization with surfactants

We demonstrate that a surfactant-stabilized molten globule intermediate exists for recombinant human growth hormone (rhGH), is very hydrophobic, and tends to form aggregates. Characterization of this intermediate included equilibrium denaturation measured by electron paramagnetic resonance (EPR) and CD spectroscopy, assessment of aggregation during refolding, and fluorescence studies of its binding to the hydrophobic probe, 1-anilinonapthalene-8-sulfonate (1,8-ANS). We have found that at 4.5 M guanidinium hydrochloride (GuHCl), a molten globule intermediate of rhGH is stabilized and results in significant aggregation upon refolding. This intermediate is populated by the addition of the nonionic surfactant, Tween. This surfactant also reduces the extent of aggregation during refolding of rhGH from 4.5 M GuHCl. Overall, our studies reveal that rhGH forms a molten globule-like intermediate during folding and this intermediate self-associates. This self-association is reduced upon formation of a Tween-rhGH complex. Tween also binds to the native protein. Thus, nonionic surfactants such as Tween may act like molecular chaperones in facilitating protein folding while not altering the native conformation.     

Projection structure of frog rhodopsin in two crystal forms

Rhodopsin is the G protein-coupled receptor that upon light activation triggers the visual transduction cascade. Rod cell outer segment disc membranes were isolated from dark-adapted frog retinas and were extracted with Tween detergents to obtain two-dimensional rhodopsin crystals for electron crystallography. When Tween 80 was used, tubular structures with a p2 lattice (a = 32 A, b = 83 A, gamma = 91 degrees) were formed. The use of a Tween 80/Tween 20 mixture favored the formation of larger p22(1)2(1) lattices (a = 40 A, b = 146 A, gamma = 90 degrees). Micrographs from frozen hydrated frog rhodopsin crystals were processed, and projection structures to 7-A resolution for the p22(1)2(1) form and to 6-A resolution for the p2 form were calculated. The maps of frog rhodopsin in both crystal forms are very similar to the 9-A map obtained previously for bovine rhodopsin and show that the arrangement of the helices is the same. In a tentative topographic model, helices 4, 6, and 7 are nearly perpendicular to the plane of the membrane. In the higher-resolution projection maps of frog rhodopsin, helix 5 looks more tilted than it appeared previously. The quality of the two frog rhodopsin crystals suggests that they would be suitable to obtain a three-dimensional structure in which all helices would be resolved.     

An EPR dosimetry method for rapid scanning of children following a radiation accident using deciduous teeth

The purpose of this study was to investigate the formulation and delivery of a protein in a pressurized metered-dose inhaler (pMDI) containing HFA 134a as the propellant for aerosol delivery. Ethanol and surfactants, including polyoxyethylene 10 oleyl ether (Brij 97), polyoxyethylene 20 oleyl ether (Brij 98), polyoxyethylene sorbitan monooleate (Tween 80) and Aerosol OT (AOT), were investigated as formulation adjuvants to improve the dose delivery characteristics of the model protein (bovine serum albumin) containing pMDI formulations. The aqueous solution of a surfactant and protein was lyophilized to obtain a solid carrier system of the protein. Readily dispersible suspensions were obtained by suspending this solid carrier system in HFA 134a with ethanol as a dispersing aid. The formulations containing Tween 80 resulted in the highest respirable fraction. This study suggested a potential formulation containing a lyophilized complex of surfactant and protein readily dispersible in HFA 134a for delivering a therapeutic protein to the respiratory tract by inhalation.     

Enhancement of diltiazem bioavailability by traces of surfactants in rats after neomycin-induced malabsorption syndrome

Bioavailability of diltiazem was investigated in rats with the neomycin induced malabsorption syndrome in presence of surfactants. Tween 80, sodium lauroyl sulphate and lecithin were added under their critical miscellar concentration levels. Increases of diltiazem bioavailability (in the range of 5-29%) promoted by additions of the surfactants were observed.     

pH-dependent microspheres from modified soybean protein hydrolysate

Soybean hydrolysate is a hydrophilic mixture of amino acids and low molecular peptides which are soluble over the whole pH range. Chemical modification of soybean hydrolysate with aromatic acyl chlorides resulted in a product that yielded pH-dependent microspheres. Investigation into the physiochemical properties of the microsphere forming material indicated that acylation had alerted the hydrophobic/hydrophilic ratio as evidenced by an increased column retention time on reverse phase HPLC. This was further confirmed by analysis of the amino acid composition of the modified material. The data indicated that the hydrophobic/hydrophilic ratio and low molecular were critical factors in the formation of this supramolecular complex. An estimation based on sedimentation rate revealed an average molecular weight of these microspheres as 10(7)-10(8) Daltons. Light scattering experiments indicated that the microspheres have discrete chambers in the interior. Included among the properties of the microspheres that have been determined are the pH range of the phase transition, size distribution and zeta-potential. The physiochemical characteristics of the microspheres prepared from modified soybean protein are similar to the microsphere forming material produced by thermal condensation of amino acids. Formation of microspheres in solution containing either porcine insulin or salmon calcitonin resulted in the encapsulation of nearly 60% of the dissolved proteins. Oral gavage of encapsulated porcine insulin or salmon calcitonin into the stomach of rats resulted in significant titers of either protein in the systematic circulation.     

Newcastle disease oil emulsion vaccines prepared with animal, vegetable, and synthetic oils

Animal, vegetable, and synthetic oils were tested as potential replacements for mineral oil in Newcastle disease oil emulsion vaccines. Emulsifying surfactants of seed oil origin comprised 10% of the the oil phase that was used to prepare water-in-oil emulsion vaccines that contained a final concentration of 20% aqueous antigen. The hemagglutination inhibition responses of chickens inoculated with 46 of the newly formulated oil vaccines were, in most cases, not significantly different from those of control chickens inoculated with mineral oil vaccine. Tissue reactions associated with animal, vegetable, and synthetic oil vaccines were less severe than those associated with mineral oil vaccines. Viscosity of the mineral oil formulations ranged from 1/2 to 3 1/2 times that of the mineral oil control vaccines. These findings indicate that any of several oils may be more suitable than mineral oil for preparation of immune adjuvants for poultry vaccines.     

Influence of interfacial rheological properties of mixed emulsifier films on the stability of water-in-oil-in-water emulsions

The purpose of this study was to investigate the influence of mixtures of the emulsifiers Span 80, 83 and 85 and Tween 80 on multiple emulsion stability. An oscillatory ring-surface rheometer was used to measure interfacial elasticity at the oil-aqueous interface. Multiple emulsions were prepared via a two-step emulsification process and stability was evaluated by investigation of drug transport from freshly prepared and eight-day-old emulsions by use of a dialysis method. Photomicrography and droplet-size analysis of multiple emulsions were also conducted. Spans 80 and 83 were appreciably elastic (683.10+/-29.13 mNm(-1) and 1128.09+/-14.81 mNm(-1), respectively at 5% w/v) when present at the mineral oil-aqueous interface whereas Span 85 and Tween 80 were not (11.10+/-3.88 mNm(-1) (5% w/v) and 0 (0.1-5% w/v) respectively). The interfacial elasticities of Spans 80 and 83 decreased in the presence of Tween 80 in the aqueous phase; this was attributed to co-adsorption of Tween 80 at the interface or aqueous-phase solubilization of the Spans within mixed micelles, or both. Drug-transport studies indicated that drug release on storage was lower from water-in-oil-in-water (w/o/w) emulsions prepared with 5% w/v Span 80 or 83 and 0.1% w/v Tween 80 than from emulsions prepared with 5% w/v Spans 80 or 83 and 1% w/v Tween 80. Photomicrography and droplet-size analysis indicated the same trend-emulsions containing a higher percentage of Tween 80 were less stable. The relatively stable w/o/w emulsions (e.g. 5% w/v Spans 80 or 83 and 0.1% w/v Tween 80) contained a large number of multiple droplets for up to eight weeks of storage whereas the relatively unstable w/o/w emulsions (e.g., 5% w/v Span 85 and 0.1% w/v Tween 80 and 5% w/v Spans and 1% w/v Tween 80) contained mostly simple droplets after only one week of storage. The mean volume/weight droplet size decreased on storage with breakdown of these w/o/w emulsions to simple oil-in-water emulsions. There was a positive correlation between the interfacial elasticity and emulsion stability data. Mixed emulsifiers giving higher film strength, as quantified by interfacial elasticity measurements, resulted in more stable w/o/w emulsions.     

Effects of dietary vegetable oils on behavior and drug responses in mice

Previously, we noted significant differences in the behavioral patterns of mice fed safflower oil with a very low alpha-linolenate/linoleate ratio and perilla oil with a high alpha-linolenate/linoleate ratio from mothers to offsprings. In this report, we compared the behavior and drug responses in mice fed diets containing six different vegetable oils-corn, rapeseed, soybean, safflower, perilla and a mixture of perilla and safflower oils- for a relatively short period: 8 months after weaning. Soybean oil is a component of most conventional diets and was used as a control. The alpha-linolenate/linoleate ratios of the oils appeared to affect the locomotor activities in a wheel cage: the activity decreased in the order of safflower, the mixture (1:1) and the perilla oil groups. However, the rapeseed oil group exhibited much higher locomotor activity than that expected from the alpha-linolenate/linoleate ratio. Additionally, the rapeseed oil group exhibited unusual behavior patterns, including higher ambulation and rearing activities, faster acquisition of the water maze task and slower habituation behavior as compared with the control group. Susceptibility to pentobarbital anesthesia tended to be higher in the rapeseed oil group. The differences in the alpha-linolenate/linoleate ratios of these oils alone do not account for the observed differences in the behavioral patterns among the six dietary groups. Although we cannot exclude the possibility that the observed behavioral anomaly is due to the unique fatty acid composition of rapeseed oil, we speculate that a factor(s) other than fatty acids in rapeseed oil affected nervous system functions.     

Development of injectable microcapsules for use in the treatment of narcotic addiction

Injectible microcapsules containing narcotic antagonists have been prepared with dl-poly (lactic acid) as the coating material. The encapsulation technology has developed to the point that high yields of less than 180 mu capsules can be prepared routinely. Such capsules with an initial payload of 50 wt. % naltrexone pamoate provide 60-90% antagonism to the action of morphine 28 days after injection into mice as a peanut oil/aluminum monostearate suspension at a dose level of 40 miligrams naltrexone pamoate/ kg. mouse.     

Liposomes containing boronophenylalanine for boron neutron capture therapy

In the present work, liposomes loaded with Boronophenylalanine (BPA), with or without stabilization, were formulated for the application in boron neutron capture therapy. BPA was encapsulated into liposomes as a complex with fructose, but also as a free drug in two different pH buffers. The influence of critical variables (cholesterol content, drug:lipid molar ratio, osmotic stress of liposomes containing hyperosmotic drug solution) on liposome morphology and drug content was evaluated. The drug content and dissolution profile of different BPA loaded liposomes were also studied. The physical stability of liposomes in terms of changes in the size distribution in different osmotic pressure buffers and the chemical oxidation of phospholipids during storing conditions were investigated. The encapsulation efficiencies of all formulations were always satisfactory, being between 20-48%; even when the liposomes were exposed to high osmotic stress, the particle size was below 200 nm. The BPA-fructose complex loaded liposomes showed a slower drug release profile.     

Improving stability and release kinetics of microencapsulated tetanus toxoid by co-encapsulation of additives

PURPOSE: Tetanus toxoid (Ttxd) encapsulated in polyester microspheres (MS) for single injection immunization have so far given pulsatile in vitro release and strong immune response in animals, but no boosting effect. This has been ascribed to insufficient toxoid stability within the MS exposed to in vivo conditions over a prolonged time period. This study examined the effect of co-encapsulated putative stabilizing additives. METHODS: Two different Ttxd were encapsulated in poly(D,L-lactic-co-glycolic acid) (PLGA 50:50) and poly(D,L-lactic acid) (PLA) MS by spray-drying. The influence of co-encapsulated additives on toxoid stability, loading in and release from the MS, was studied by fluorimetry and ELISA. RESULTS: Co-encapsulated albumin, trehalose and gamma-hydroxypropyl cyclodextrin all improved the toxoid encapsulation efficiency in PLGA 50:50 MS. Albumin increased the encapsulation efficiency of antigenic Ttxd by one to two orders of magnitude. Further, with albumin or a mixture of albumin and trehalose ELISA responsive Ttxd was released over 1-2 months following a pulsatile pattern. CONCLUSIONS: Optimized Ttxd containing MS may be valuable for a single-dose vaccine delivery system.