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PubMed2001

Alginate as a macroaffinity ligand and an additive for enhanced activity and thermostability of lipases.

Lipases are being employed increasingly for the synthesis of drug intermediates and pharmaceutically important molecules as well as for the resolution of racemic mixtures for obtaining physiologically active enantiomers. Alginate was used as a macroaffinity ligand to purify lipases from Chromobacterium viscosum, porcine pancreas and wheatgerm by employing the technique of affinity precipitation. In all the cases, the purified preparation showed a single band on SDS/PAGE. The process gave adequate yields of 87, 75 and 62% in the case of Chromobacterium, porcine and wheatgerm lipases respectively. Alginate was also found to activate the enzymes; this effect was most dramatic in the case of wheatgerm lipase, for which a 4-5-fold activity increase was observed. Furthermore, alginate was also found to protect the enzyme against thermoinactivation.

Alginates↗
PubMed2002

Growth and production of cholesterol oxidase by alginate-immobilized cells of Rhodococcus equi No. 23.

Rhodococcus equi No. 23 was immobilized in calcium alginate. No detrimental effect on the viability of the test organism was observed during the immobilization procedure. Approx. 98% of the cell population originally present in the alginate solution were immobilized in the gel beads. When the cells of an equal volume of the culture, obtained respectively at exponential phase (12 h preculture), late-exponential phase (20 h preculture) or stationary phase (36 h preculture) were immobilized, the gel beads prepared with the stationary-phase culture were found to contain the highest cell population [about 10(8) colony-forming units (CFU)/g of beads]. In addition, gel beads, prepared with late-exponential-phase culture, exhibited the highest production of cholesterol oxidase (CholOx) after 48 h of incubation. Increasing the bead mass from 3.5 to 14.0 g/100 ml of medium increased CholOx production. However, further increasing the bead mass resulted in a reduction of CholOx production. Furthermore, on the basis of a similar initial cell population, the alginate-immobilized cells of R. equi No. 23 produced a significantly higher amount of CholOx (P<0.05) than did the free cells.

Alginates↗
PubMed2002

Compartmentalization of the matrix formed by nucleus pulposus and annulus fibrosus cells in alginate gel.

Intervertebral disc cells cultured in alginate gel are capable of reforming in alginate, a matrix that consists of two compartments: a rim of metabolically active cell-associated matrix and a more abundant, but metabolically less active, further removed matrix. At any one age and in most species, the cell-associated matrix formed by a nucleus pulposus or annulus fibrosus cell cultured in this way is less abundant than that formed by an articular chondrocyte. In both the cell-associated matrix and further removed matrix, the ratio of aggrecan to collagen is significantly higher in the case of nucleus pulposus than of annulus fibrosus, a feature that also distinguishes the matrices of the nucleus pulposus and annulus fibrosus in vivo. Nucleus pulposus and annulus fibrosus cells from older donors show a decreased ability to reform a cell-associated matrix rich in aggrecan. There is, however, some evidence that gene therapy and/or exposure of the cells to defined stimulatory factors can help overcome some of these age-related limitations. This contention is supported by recent evidence that nucleus pulposus and annulus fibrosus cells from adult donors can be manipulated to form, using the recently developed alginate-recovered chondrocyte system, a resilient tissue that bears many of the characteristics of the tissue in which these cells reside in vivo.

Alginates↗
PubMed1990

The dimensional stability of self-disinfecting alginate impressions compared to various immersion regimes.

Alginate impressions of a master acrylic study model pair were made in order to assess the effect of various disinfection techniques on dimensional stability. Impressions were made using self-disinfecting alginate, traditional alginate which had been dipped or soaked in a disinfecting solution, and included was a control group which was not disinfected. Inter- and intra-arch linear measurements of the resultant study casts were made using a Reflex Metrograph. The small differences found for the variables measured were not statistically significant.

Alginates↗
PubMed1999

Consistency of alginate impression materials and their evaluation.

The apparent viscosity of 17 commercially available alginate impression materials was examined using a rheometer. Each material was mixed for 15 s at a temperature of 23 +/- 0.5 degrees C, using a specially designed alginate mixing instrument. The water powder ratio was determined according to the manufacturer's instruction. The mixed alginate paste was immediately transferred to the sample stage of the rheometer or to a commercial perforated metal tray. Over the same time scale adjusted according to the rheometer test, a discrimination test (subjective test) was performed by 16 young dentists (each with more than 4 years experience). The apparent viscosity of all materials rose as a function of time after mixing and most of the material had properties similar to pseudoplastic fluids. The relationship between the results of the subjective test and the experimental values (objective test) with the rheometer was expressed in a quadratic equation. The maximum value of the apparent viscosity derived from this result was 1.52 x 10(3) Pa.s.

Alginates↗
PubMed2002

In vitro development of bovine embryos encapsulated in sodium alginate.

The objective of this study was to evaluate the in vitro development of bovine embryos encapsulated in alginate. Day-4 embryos produced in vitro (n = 110) were encapsulated with 1.5% sodium alginate and co-cultured with oviduct cells. Unencapsulated embryos (n = 106) were used as controls. In vitro development rate to the blastocyst stage at Day 7 was similar between encapsulated, 42.7%, (47/110) and control. 34% (36/106). embryos. Although encapsulated embryos were able to hatch on Day 9, they did so in a lower proportion than controls (P < 0.05). In conclusion, alginate encapsulation of bovine embryos does not disturb the in vitro development up to the blastocyst stage but significantly reduces the hatching process.

Alginates↗
PubMed2002

Sequential treatment with calcium alginate dressings and hydrocolloid dressings accelerates pressure ulcer healing in older subjects: a multicenter randomized trial of sequential versus nonsequential treatment with hydrocolloid dressings alone.

OBJECTIVES: To compare the efficacy of a sequential strategy combining calcium alginate and hydrocolloid dressings treatment of grade III or IV pressure ulcers (PUs) and the efficacy of nonsequential strategy with hydrocolloids alone. DESIGN: An open, randomized, multicenter parallel-group trial. SETTING: Twenty geriatrics hospital wards. PARTICIPANTS: One hundred ten older patients with grade III or IV PUs. INTERVENTION: The control strategy consisted of applying hydrocolloid dressings (DuodermE) for 8 weeks; the sequential strategy consisted of applying combined calcium alginate dressings (UrgoSorb) for the first 4 weeks and hydrocolloid dressings (Algoplaque) for the next 4 weeks. MEASUREMENTS: PU surface areas were measured weekly by ulcer tracing. The endpoints were the mean absolute surface area reduction (SAR) during the 8-week study period and the number of patients achieving a 40 or more SAR (SAR40). RESULTS: Fifty-seven and 53 patients were randomly allocated to sequential and control strategies respectively. Baseline patient characteristics and PU ulcer features at inclusion were similar in the two groups. Mean +/- standard deviation SAR was significantly larger in the sequential treatment group (5.4 +/- 5.7 cm2 and 7.6 +/- 7.1 cm2 at 4 and 8 weeks) than in the control group (1.6 +/- 4.9 cm2 and 3.1 +/- 7.2 cm2, P< .001). In the sequential treatment group, 68.4 of the patients reached SAR40 at 4 weeks and 75.4 at 8 weeks, proportions significantly larger than in the control group (22.6 and 58.5, respectively, P< .0001). Dressing tolerance was good in both strategies. CONCLUSIONS: In grade III or IV PUs, treatment using first calcium alginate dressings and then hydrocolloid dressings promotes faster healing than treatment with hydrocolloid dressings alone.

Aged↗
PubMed1987

Composition of alginate synthesized during the growth cycle of Pseudomonas aeruginosa.

A mucoid P. aeruginosa isolated from the sputum of a cystic fibrosis patient was grown in batch culture on a complex medium. During the growth cycle the amount of alginate produced was estimated and its composition was determined by proton magnetic resonance (1H-n.m.r) spectroscopy. Exopolysaccharide production occurred mainly during the exponential phase of growth. The alginate samples isolated varied little in composition and were characterized by being highly acetylated, high mannuronate (0.83-0.93 mole fraction) polymers. Guluronate was present only within heteropolymeric regions of the polysaccharides which all displayed a complete absence of polyguluronate. Ca2+ ion supplementation of the medium was not observed to increase the levels of guluronate in the alginates produced.

Alginates↗
PubMed1992

AlgR2 is an ATP/GTP-dependent protein kinase involved in alginate synthesis by Pseudomonas aeruginosa.

The exopolysaccharide alginate is a major virulence factor in the pathogenicity of Pseudomonas aeruginosa infecting the lungs of cystic fibrosis (CF) patients. Alginate synthesis by P. aeruginosa is believed to occur in response to environmental signals present in the CF lung. Transcription of a critical alginate biosynthetic gene, algD, is triggered by environmental signals and is known to be controlled by regulatory proteins AlgR1, AlgR2, and AlgR3. AlgR1 is a member of the family of response regulators of the phosphorylation-dependent two-component bacterial signal transduction systems. In this report, we describe the characterization of AlgR2 as the kinase involved in phosphorylation of AlgR1. AlgR2, an 18-kDa soluble protein undergoes rapid autophosphorylation in the presence of either ATP or GTP and transfers the phosphate to AlgR1. AlgR2 retains high affinity for both ATP and GTP with an apparent Km of 137 and 249 nM, respectively, for phosphorylation by these two substrates. ADP and GDP exhibit competitive inhibition with an apparent Ki of 94 and 314 nM, respectively, during phosphorylation by ATP and 481 and 273 nM during phosphorylation by GTP. AlgR1 and AlgR2 can be isolated in the form of an 80-kDa complex that is capable of undergoing phosphorylation and intracomplex phosphotransfer in vitro. A 16-kDa AlgR2 analog, capable of autophosphorylation in the presence of ATP or GTP and transferring the phosphate moiety to AlgR1, has been characterized in Escherichia coli.

Adenosine Triphosphate↗
PubMed1995

The algT (algU) gene of Pseudomonas aeruginosa, a key regulator involved in alginate biosynthesis, encodes an alternative sigma factor (sigma E).

Chronic infection by alginate-producing (mucoid) Pseudomonas aeruginosa is the leading cause of mortality among cystic fibrosis (CF) patients. During the course of sustained infection, the production of an alginate capsule protects the bacteria and allows them to persist in the CF lung. One of the key regulators of alginate synthesis is the algT (algU) gene encoding a putative alternative sigma factor (sigma E). AlgT was hyperproduced and purified from Escherichia coli. The N-terminal sequence of the purified protein matched perfectly with that predicted from the DNA sequence. The purified protein, in the presence of E. coli RNA polymerase core enzyme, was able to initiate transcription of an algT promoter. Deletion of the -35 region of this promoter abolished this activity in vitro as well as in vivo. These data indicate that the algT gene encodes a sigma factor that is autoregulatory.

Alginates↗
PubMed2004

Helix induction in antimicrobial peptides by alginate in biofilms.

Bacterial exopolysaccharides provide protection against phagocytosis, opsonization, and dehydration and act as a major structural component of the extracellular matrix in biofilms. They contribute to biofilm-related resistance by acting as a diffusion barrier to positively charged antimicrobial agents including cationic antimicrobial peptides (CAPs). We previously created novel CAPs consisting of a nonamphipathic hydrophobic core flanked by Lys residues and containing a Trp residue in the hydrophobic segment as a fluorescent probe. Peptides of this type above a specific hydrophobicity threshold insert spontaneously into membranes and have antimicrobial activity against Gram-positive and Gram-negative bacteria at micromolar concentrations. Here we show that alginate, a polymer of beta-d-mannuronate and alpha-l-guluronate secreted by the cystic fibrosis pathogen Pseudomonas aeruginosa, induces an alpha-helical conformation detected by circular dichroism spectroscopy and blue shifts in Trp fluorescence maxima in peptides above the hydrophobicity threshold, changes typically observed upon association of such peptides with nonpolar (membrane) environments. Parallel effects were observed in the archetypical CAPs magainin II amide and cecropin P1. Fluorescence resonance energy transfer studies indicated that alginate induces peptide-peptide association only in peptides above the hydrophobicity threshold, suggesting that the hydrophilic alginate polymer behaves as an "auxiliary membrane" for the bacteria, demonstrating a unique protective role for biofilm matrices against CAPs.

Alginates↗
PubMed2001

Xenotransplantation of parathyroids in rats using barium-alginate and polyacrylic acid multilayer microcapsules.

The integrity and function of encapsulated parathyroid tissue following xenotransplantation is limited by oxygen and nutrition supply and capsule fibrosis. Since some of these factors depend on stability and biocompatibility of the coating material, multilayer microcapsules have been developed. Parathyroid tissue pieces and digested single cells from pigs were encapsulated in barium-alginate and in polyacrylic acid (PAA) multilayer capsules. After 7 days of culture the function of the encapsulated cells were assessed. Subsequently, in a part of the cultured microcapsules the viability was directly assessed whereas the other part was transplanted in dark animal [DA] rats for 30 days. After explantation viability and fibrotic reaction were examined. Single cells showed a significant increase in parathyroid hormone [PTH] secretion when exposed to medium low in calcium, whereas minced tissue pieces revealed necrosis without stimulatory responsiveness. Morphometry showed significantly better viability of single cells compared with minced tissue in vitro and in vivo. The fibrotic reaction against capsules with minced tissue was more pronounced than for capsules containing single cells. There was no difference between barium alginate and PAA capsules when containing minced tissue. In single cells, however, the fibrous tissue reaction differed significantly between barium alginate and PAA capsules. Encapsulated single cells of parathyroid tissue maintain detectable function and viability. In contrast minced tissue underwent necrosis and induced significantly more connective tissue reaction than single cells indicating an interrelationship between necrosis and fibrosis.

Acrylic Resins↗
PubMedDate not supplied

The effect of alginate on the invasion of cystic fibrosis respiratory epithelial cells by clinical isolates of Pseudomonas aeruginosa.

Chronic infection in the cystic fibrosis (CF) lung is characterized by Pseudomonas aeruginosa strains that overproduce the mucoid exopolysaccharide, alginate. Previous experiments have shown that long-term survival of P. aeruginosa in the CF lung may be facilitated by increased adherence and decreased invasion of respiratory epithelial cells. Therefore, mucoid and nonmucoid clinical isolates of P. aeruginosa were assayed for their ability to associate with and invade the CF respiratory epithelial cell line, CF/T43. Association assays and gentamicin exclusion assays demonstrated that mucoid P. aeruginosa associates with and invades CF/T43 cell monolayers significantly less than nonmucoid P. aeruginosa strains (P = .004, .02). Fluorescence microscopy invasion assays confirmed these results. The differences in association and invasion by the P. aeruginosa strains were not due to differences in lipopolysaccharide phenotype or cytotoxicity for CF/T43 respiratory epithelial cells. Exogenous bacterial alginate had no effect on the invasion of CF respiratory epithelia by a nonmucoid strain. Invasion assays with the wild-type P. aeruginosa strain PAO1 and isogenic algU and mucA mutant strains failed to show differences in invasion (P = .25). We conclude that (i) mucoid P. aeruginosa isolates associate with and invade CF/T43 respiratory epithelial cells with less efficiency than nonmucoid P. aeruginosa, (ii) these differences are not due to variations in lipopolysaccharide phenotype between strains, (iii) neither exogenous nor endogenous alginate affects the ability of P. aeruginosa to invade CF/T43 respiratory epithelial cells, and (iv) invasion of CF/T43 respiratory epithelial cells by a laboratory reference strain of P. aeruginosa does not appear to be regulated by AlgU.

Alginates↗
PubMedDate not supplied

Effect of lecithins on BCG-alginate-PLL microcapsule particle size and stability upon storage.

Alginate-PLL microcapsules containing BCG were prepared by emulsification/internal gelation of an alginate solution dispersed within a vegetable oil containing lecithins as emulsifiers. The lecithins studied were soy bean lecithin at 0.1, 0.5, 1 and 2% concentration; and dried egg yolk lecithin at 0.1, 0.25, 0.5 and 1%. The microcapsule particle size showed a dependence upon the amount and type of lecithin added to the oil. Increasing the emulsifying agent concentration was found to reduce particle size, from 50.9 microm obtained when lecithins were not used in the emulsification step, to 13.9 microm obtained when 1% dried egg yolk lecithin was employed. The encapsulated BCG was identified by the Difco TB stain set K, followed by observation under optical microscopy. Once prepared, microcapsules were freeze-dried using 5% trehalose as cryoprotectant in order to preserve their stability upon storage. The stability of the microcapsules was assayed over 12 months at room temperature, finding that alginate-PLL microcapsules were stable up to 6 months. Moreover, in the case of microcapsules prepared with lecithins, a significant increase in particle size was observed, from 16.9 microm at the beginning of the study to 25.2 microm at 12 months storage.

Alginates↗
PubMedDate not supplied

Interpenetrating polymer networks of alginate and polyethylene glycol for encapsulation of islets of Langerhans.

A mixture of alginate and polyethylene glycol acrylate was investigated as a system for the encapsulation of islets of Langerhans. This system showed dual crosslinkability: the alginate was ionically crosslinked by multivalent cations, and the PEG was covalently crosslinked by photoactivated free radical polymerization. The major advantage of the dually crosslinkable system was the chemical stability of the resultant gels due to the presence of covalent bonds that maintain the integrity of the gel as opposed to reversible ionic linkages that were the only mode of crosslinkage in previous generations of alginate-based encapsulation systems. The physical aspects of gelation of such alginate/PEG compositions were investigated. Diffusion of dextrans of known molecular weights through these gels was studied in order to shed light on the hydrogel porosity and permeability. In vitro viability and function tests demonstrated that these gels were biocompatible. Islets encapsulated in these systems were healthy and retained both viability and insulin secretory function.

Alginates↗
PubMedDate not supplied

Microencapsulation of oils using sodium alginate.

The feasibility of encapsulating wheatgerm oil and evening primrose oil using sodium alginate by the emulsification method was explored in this study. It is based on the chemical reaction between the water-soluble sodium alginate and polyvalent cation, calcium, to form the water-insoluble alginate. The factors investigated were the physical appearance of the microspheres, the amount of oil that could be encapsulated, the flow property, size distribution and mean size of the microspheres produced. Encapsulation efficiency and oil content of wheatgerm oil increased with an increase in oil load. The mean size of the microspheres increased sharply at a high oil load of 250% w/w. Photographs of microspheres taken showed that the microspheres were larger, spherical and had more vesicles within, as oil load increased. Encapsulation efficiency of evening primrose oil microspheres was similar to wheatgerm oil microspheres at the respective oil loads of 50, 250 and 350% w/w. The emulsification method developed was successfully applied to wheatgerm oil, a fixed oil, with a maximum encapsulation efficiency of approximately 88%. It was satisfactory for evening primrose oil, which also belongs to the family of fixed oils.

Alginates↗
PubMedDate not supplied

Protein immobilization in crosslinked alginate microparticles.

Oral administration of peptide and protein drugs requires their protection from the acidic and enzymatic degradation in the gastro-intestinal environment and their targeting to the absorption zone. For this purpose, an alginate microsystem, as a carrier of bovine serum albumin (BSA), as a model protein, was developed using a spray-drying technique. A hardening process with Ca2+ and chitosan (CS) provided a system with resistance to the gastro-intestinal barriers and of appropriate size for targeting to the Peyer's patches. The present work aims to evaluate the effects of the ratio of sodium alginate (Na-A) and BSA as well as the pH of the crosslinking medium on the microsystem properties. Microparticle morphological and dimensional characteristics did not change significantly with the formulation variables. BSA loading at a pH value less than the protein isoelectric point (pI) was higher than that at a pH similar to the pI owing to an electrostatic interaction between the charged protein and the polyanionic alginate. The maximum encapsulation efficiency was obtained at the highest Na-A/BSA ratio. Protein release in a simulated gastro-intestinal fluid was not affected by the preparative variables, but was controlled by the pH-dependent nature of the polymer material. Polyacrylamide gel electrophoresis (PAGE) demonstrated the stability of the protein to both the preparative conditions and the gastro-intestinal pH values.

Alginates↗
PubMed2005

Characteristics of vitamin C immobilized particles and sodium alginate beads containing immobilized particles.

This paper reports the properties of vitamin C encapsulated sodium alginate beads prepared by an alternative approach. The alternative encapsulation process mainly involves immobilization of vitamin C in hydrated zinc oxide layers and encapsulation of prepared immobilized particles in sodium alginate bead. The immobilization of vitamin C in hydrated zinc oxide layers was achieved by a coprecipitation process. Fourier transform infrared (FTIR) spectroscopy showed that the vitamin C was found to be stable after its immobilization. X-ray diffraction (XRD) studies revealed that anionic vitamin C molecules are adsorbed between positively charged zinc hydroxide layers with a 1:1 layer sequence, since well-defined change in basal spacing was observed. Well-defined change in surface morphology was observed by scanning electron microscopy (SEM) when vitamin C immobilized particles are encapsulated in sodium alginate bead. The biological activity of vitamin C was retained, even after its immobilization which was confirmed by 4-dihydroxy-L-phenylalanine (L-DOPA) oxidase inhibition and free radical scavenging activity studies. The release rate of vitamin C from immobilized particles and beads was sustained through an ion exchange process. A higher amount of stable vitamin C was recovered from the bead when compared to neat vitamin C itself.

Alginates↗