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PubMed1995

Hypolipidemic agents alter hepatic mitochondrial respiration in vitro.

The direct effects of three different classes of structurally diverse hypolipidemic agents on respiration were studied in mitochondria isolated from donor Sprague-Dawley rats. Two classes of peroxisome proliferators (i.e. plasticizers and hypolipidemic hormones and drugs) and one class of peroxisome inhibitors (i.e. anti-psychotic drugs) were studied. The phthalate ester plasticizers dibutylphthalate, ethylhexanoic acid and di(2-ethylhexyl) adipate, the hypolipidemic hormones or drugs dehydro-epiandrosterone (DHEA), thyroxine (T4), triiodothyronine (T3), gemfibrozil, clofibrate and naphthoflavone, and the anti-psychotic drugs chlorpromazine, thioridazine and fluphenazine were studied. As the dose of the plasticizer dibutylphthalate increased from 8 to 200 mumol/l, there was a decrease (P < 0.05) in state 3 (+ADP) respiration and in the respiratory control ratio for both substrates tested. The anti-psychotic drug chlorpromazine decreased state 3 malate + pyruvate-supported respiration and increased state 3 succinate-supported respiration. As the concentration of all three anti-psychotic drugs increased, there was a linear increase in state 4 respiration (-ADP) and a decrease in the respiratory control ratio for both substrates tested. As the dose of the hypolipidemic agents DHEA, gemfibrozil and T4 increased, there was a linear reduction in state 3 malate + pyruvate-supported respiration. However, when succinate was used as the substrate to support respiration, only the thyroid hormones significantly decreased state 3 respiration. Gemfibrozil, T4 and T3 increased state 4 respiration, regardless of the substrate used. As the dose of clofibrate, gemfibrozil, and the thyroid hormones increased, there was a linear reduction in the respiratory control ratio for both substrates tested.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1989

Octavalent Pseudomonas aeruginosa O-polysaccharide-toxin A conjugate vaccine.

An octavalent Pseudomonas aeruginosa conjugate vaccine was synthesized by covalently coupling the O-polysaccharide (O-PS) moiety derived from lipopolysaccharides of Habs serotypes 1, 2, 3, 4, 5, 6, 11 and 12 to toxin A. Adipic acid dihydrazide was used as a spacer molecule to facilitate conjugation. The vaccine was composed of 37% (w/w) O-PS and 63% toxin A, devoid of enzymatic activity characteristic of toxin A, non-toxic for mice and guinea pigs, and non-pyrogenic. The vaccine elicited a significant rise in immunoglobulin G antibody levels to all serotypes of lipopolysaccharide contained in the vaccine and to toxin A. Serotypes 6, 10 and 11 were most immunogenic in mice whereas serotypes 1 and 5 engendered the lowest antibody response. Antitoxin A antibody was able to neutralize the cytotoxicity of toxin A. Immunization of mice with the vaccine conferred significant protection against subsequent challenge with all P. aeruginosa serotype strains contained in the vaccine.

Animals↗
PubMed1992

Assay for L-pipecolate oxidase activity in human liver: detection of enzyme deficiency in hyperpipecolic acidaemia.

A direct assay method is described for L-pipecolate oxidase. The assay uses NaHSO3 to trap the L-alpha-amino [3H]adipate delta-semialdehyde (AAS) formed as a direct reaction product of L-pipecolate oxidase from L-[3H]pipecolic acid. The adduct so formed was separated from the substrate on Dowex 50 (H+) column. The product was identified as [3H]AAS by amino acid analysis after breaking down the adduct by boiling under acidic conditions. The assay is simpler and more specific than fluorometric methods; it is also more sensitive, requiring at most 16 micrograms of liver peroxisome-enriched protein per assay. We have used this assay procedure to detect L-pipecolate oxidase in skin fibroblasts obtained from a control subject and from patients of hyperpipecolic acidaemia and Zellweger syndrome and found that this enzyme activity is present in the control, but absent or decreased in the patients with the peroxisomal disorders.

Humans↗
PubMed2004

Controlled degradation of hydrogels using multi-functional cross-linking molecules.

Hydrogels, chemically cross-linked or physically entangled, have found a number of applications as novel delivery vehicles of drugs and cells. However, the narrow ranges of degradation rates and mechanical strength currently available from many hydrogels limits their applications. We have hypothesized that utilization of multi-functional cross-linking molecules to form hydrogels could provide a wider range and tighter control over the degradation rates and mechanical stiffness of gels than bi-functional cross-linking molecules. To address the possibility, we isolated alpha-L-guluronate residues of sodium alginate, and oxidized them to prepare poly(aldehyde guluronate) (PAG). Hydrogels were formed with either poly(acrylamide-co-hydrazide) (PAH) as a multi-functional cross-linking molecule or adipic acid dihydrazide (AAD) as a bi-functional cross-linking molecule. The initial properties and degradation behavior of both PAG gel types were monitored. PAG/PAH hydrogels showed higher mechanical stiffness before degradation and degraded more slowly than PAG/AAD gels, at the same concentration of cross-linking functional groups. The enhanced mechanical stiffness and prolonged degradation behavior could be attributed to the multiple attachment points of PAH in the gel at the same concentration of functional groups. This approach to regulating gel properties with multifunctional cross-linking molecules could be broadly used in hydrogels.

Acrylic Resins↗
PubMed2004

Enhanced biological stability of collagen porous scaffolds by using amino acids as novel cross-linking bridges.

Collagen porous scaffolds have been widely employed as a dermal equivalent to induce fibroblasts infiltration and dermal regeneration. To eliminate the disadvantageous drawback of the fast degradation speed, a cross-linking method was adopted by using a water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDAC) and N-hydroxysuccinimide (NHS) in the presence of amino acids (glycin, glutamic acid or lysine), which function as cross-linking bridge between collagen molecular chains. In vitro assessment of the biological stability of the cross-linked collagen scaffolds found that the collagenase biodegradation degree was greatly decreased when lysine was added, resulting in a more biological stable scaffold. On the other hand, the biodegradation degree was accelerated compared with the purely cross-linked when glutamic acid was added, while less influenced by glycin addition. By comparing the biodegradation degree of the scaffolds added with amino acids and their model compounds, i.e. adipic acid and hexane diamine, the key factor influencing the biological stability was further investigated. The results indicated that the crucial factor is dependent on the ratio of amino groups to carboxyl groups in the cross-linking system. At optimal ratio the lowest biodegradation degree is achieved. Scanning electron microscopy measurements prove that the three-dimensional structure of the scaffolds was largely preserved. Preliminary in vitro culture of fibroblasts in the collagen scaffold cross-linked with EDAC/NHS in the presence of lysine has shown that the original good cytocompatibility of collagen was retained.

Amino Acids↗
PubMed2006

In situ cross-linkable hyaluronic acid hydrogels prevent post-operative abdominal adhesions in a rabbit model.

We studied the efficacy of an in situ cross-linked hyaluronic acid hydrogel (HAX) in preventing post-surgical peritoneal adhesions, using a rabbit sidewall defect-cecum abrasion model. Two cross-linkable precursors were prepared by modifying hyaluronic acid with adipic dihydrazide and aldehyde, respectively. The hydrogel precursors cross-linked to form a flexible hydrogel upon mixing. The hydrogel was biodegradable and provided a durable physical barrier, which was highly effective in reducing the formation of post-operative adhesions. Ten out of 12 animals in the untreated control group developed fibrous adhesions requiring sharp dissection, while only 2 out of 8 animals treated with HAX gels showed such adhesions, and those occurred in locations that were not covered by the hydrogel. We also studied means by which gel degradation time can be modulated by varying the precursor concentration and molecular weight.

Abdomen↗
PubMed2006

Biodegradation of poly(anhydride-esters) into non-steroidal anti-inflammatory drugs and their effect on Pseudomonas aeruginosa biofilms in vitro and on the foreign-body response in vivo.

The ability of poly(anhydride-esters) composed of non-steroidal anti-inflammatory drugs that biodegrade to salicylic acid (SA) and adipic acid to prevent colonization by Pseudomonas aeruginosa and their effects on the foreign-body response were studied in vitro and in vivo, respectively. Soluble SA in bacterial medium at concentrations up to 300 mg/L did not affect the growth rate or viability of P. aeruginosa, indicating that SA does not exhibit a direct toxicity effect on the bacterium. Batch degradation rates of the salicylate-based polymer in the presence of an actively growing bacterial culture only marginally (14%) increased relative to polymer degradation rates in sterile medium. Short-term (3h) bacterial adhesion studies in agitated batch systems indicated a 47% reduction in the rate of P. aeruginosa adhesion relative to a control polymer that does not release SA upon biodegradation. Long-term (3-day) biofilm accumulation studies indicated a dramatic reduction in biofilm formation on salicylate-based polymer versus controls. A recombinant P. aeruginosa pMHLAS, containing a fluorescent reporter gene prior to the las regulon, was employed to determine whether salicylate-based polymer prevents biofilm formation by the released SA inhibiting quorum sensing pathways. Long-term biofilm accumulation studies with P. aeruginosa pMHLAS insinuate that salicylate-based polymer prevents biofilm accumulation by inhibiting the las quorum sensing system. Furthermore, unlike control polymer, salicylate-based polymer implanted subcutaneously for a period of 4 weeks-resisted cell-mediated degradation and remained intact. Histological and immunohistochemical analysis indicated a reduction in overall encapsulation and paucity of macrophages in the area of the salicylate-based polymer implant.

Anti-Infective Agents↗
PubMed2007

Bone regeneration using hyaluronic acid-based hydrogel with bone morphogenic protein-2 and human mesenchymal stem cells.

Acrylated hyaluronic acid (HA) was used as a scaffold for bone morphogenic protein-2 (BMP-2) and human mesenchymal stem cells (hMSCs) for rat calvarial defect regeneration. HA was acrylated by two-step reactions: (1) introduction of an amine group using adipic acid dihydrazide (ADH); (2) acrylation by N-acryloxysuccinimide. Tetrathiolated poly(ethylene) glycol (PEG-SH(4)) was used as a cross-linker by a Michael-type addition reaction and the hydrogel was formed within 10min under physiological conditions. This hydrogel is degraded completely by 100U/ml hyaluronidase in vitro. hMSCs and/or BMP-2 was added during gelation. Cellular viability in vitro was increased up to 55% in the hydrogels with BMP-2 compared with the control. For in vivo calvarial defect regeneration, five different samples (i.e., control, hydrogel, hydrogel with BMP-2, hydrogel with MSCs, and hydrogel with BMP-2 and MSCs) were implanted for 4 weeks. The histological results demonstrated that the hydrogels with BMP-2 and MSCs had the highest expression of osteocalcin and mature bone formation with vascular markers, such as CD31 and vascular endothelial growth factors, compared with the other samples. This study demonstrated that HA base hydrogel can be used for cell and growth factor carriers for tissue regeneration.

Animals↗
PubMed2005

Estimation and projection of nitrous oxide (N2O) emissions from anthropogenic sources in Taiwan.

Taiwan is a densely populated and developed country with more than 97% of energy consumption supplied by imported fuels. Greenhouse gas emissions are thus becoming significant environmental issues in the country. Using the Intergovernmental Panel on Climate Change (IPCC) recommended methodologies, anthropogenic emissions of nitrous oxide (N2O) in Taiwan during 2000-2003 were estimated to be around 41 thousand metric tons annually. About 87% of N2O emissions come from agriculture, 7% from the energy sector, 3% from industrial processes sector, 3% from waste sector. On the basis of N2O emissions in 2000, projections for the year 2010 show that emissions were estimated to decline by about 6% mainly due to agricultural changes in response to the entry of WTO in 2002. In contrast to projections for the year 2020, N2O emissions were projected to grow by about 17%. This is based on the reasonable scenario that a new adipic acid/nitric acid plant will be probably started after 2010.

Agriculture↗
PubMed2005

Development and validation of a reversed-phase liquid chromatography method for the quantitative determination of carboxylic acids in industrial reaction mixtures.

Usually analysis of low molecular-mass carboxylic acids was performed by anion-exchange or ion-exclusion chromatographic methods. Reversed-phase liquid chromatography (RPLC) was evaluated in this work as an alternative method for the analysis of low molecular-mass aliphatic mono- and di-carboxylic acids (formic, acetic, propionic, butyric, valeric, caproic, succinic, glutaric and adipic) in aqueous media. The separation of the nine organic acids was optimised in 21 min on a high-density C18 column with an elution gradient made up of HClO4 aqueous solution 10(-3) mol L(-1) and acetonitrile. For the quantitation, external standard and standard addition methods were compared. Both methods gave similar results, so the most convenient method, external standard, was chosen for acids quantitation. Then the method had been validated and applied to the semi-quantitative analysis of formic and acetic acids and to the quantitative analysis of the others compounds in industrial reaction mixtures with concentrations ranging from 20 to 570 ppm.

Carboxylic Acids↗
PubMed2005

Determination of biogenic amines by capillary zone electrophoresis with conductometric detection.

A capillary zone electrophoresis (CZE) method with conductometric detection of biogenic amines (cadaverine, putrescine, agmatine, histamine, tryptamine and tyramine) is described. The optimised background electrolyte was the following: 15 mM histidine + 5 mM adipic acid + 1.5 mM sulphuric acid + 0.1 mM ethylenediaminotetraacetic acid + 0.1% hydroxyethylcellulose + 50% methanol. A clear separation of six biogenic amines from other components of acidic sample extract was achieved within 10 min. Method characteristics, i.e., linearity (0-100 micromol/ml), accuracy (recovery 86-107%), intra-assay repeatability (2-4%), and detection limit (2-5 micromol/l) were evaluated. Low laboriousness, sufficient sensitivity, speed of analysis, and low running cost are important attributes of this method. The developed method was successfully applied on the determination of biogenic amines in selected food samples.

Beer↗
PubMed2005

Nature and properties of ionomer assemblies. II.

The principle subject in the current paper is to summarize and characterize the ionomers based on polymers and copolymers such as polystyrene (PSt), polyisoprene (PIP), polybutadiene (PB), poly(styrene-b-isobutylene-b-styrene) (PSt-PIB-PSt), poly(butadiene-styrene) (PB-PSt), poly(ethylene terephthalate) (PET), poly(butylene adipate) (PBA), poly(butylene succinate) (PBSi), poly(dimethylcarbosiloxanes), polyurethane, etc. The self-assembly of ionomers, models concerning ionomer morphologies, physical and rheological properties of ionomer phase and percolation behavior of ionomers were discussed. The ionomer phase materials and dispersions have been characterized by differential scanning calorimetry (DSC), small-angle X-ray catering (SAXS), small-angle neutron scattering (SANS), Fourier transform infrared (FTIR) spectroscopy, transmission electron microscopy (TEM), etc. The wide range of compositions, molecular architectures, and morphologies present in ionomeric disperse systems are of great interest. The research is particularly devoted to the potential application of these materials and an understanding of the fundamental principles of the ionomers. They are extremely complex systems, sensitive to changes in structure and composition, and therefore not easily amenable to modeling and to the derivation of general patterns of behavior. The reviewed data indicate that a large number of parameters are important in influencing multiplet formation and clustering in random ionomers. Among these are the ion content, size of the polyion and counterion, dielectric constant of the host, T(g) of the polymer, rigidity or persistence length of the backbone, position of the ion pair relative to the backbone, steric constraints, amount and nature of added additive (plasticizer), thermal history, etc.

Polymers↗
PubMed2005

Development of a single unit extended release formulation for ZK 811 752, a weakly basic drug.

ZK 811 752, a potent candidate for the treatment of autoimmune diseases, demonstrated pH-dependent solubility. The resulting release from conventional matrix tablets decreased with increasing pH-values of the dissolution medium. The aim of this study was to overcome this problem and to achieve pH-independent drug release. Three different polymers were used as matrix formers, the partly water-soluble and poorly swellable mixture of polyvinylacetate/polyvinylpyrrolidone, the water-insoluble and almost unswellable ethylcellulose (EC) and the water-soluble and highly swellable hydroxypropyl methylcellulose (HPMC). To solve the problem of pH-dependent solubility different organic acids, such as fumaric, tartaric, adipic, glutaric and sorbic acid were added to the drug-polymer system. The addition of organic acids to all three matrix formers was found to maintain low pH-values within the tablets during release of ZK 811 752 in phosphate buffer pH 6.8. Thus, the micro-environmental conditions for the dissolution of the weakly basic drug were kept almost constant. An extended release matrix tablet for ZK 811 752 consisting of drug, polymer and organic acid providing the desired pH-independent drug release has been developed.

Chemistry, Pharmaceutical↗
PubMed2006

Canine retinal angioblasts are multipotent.

The purpose of this study was to culture and characterize endothelial cells and angioblasts, vascular precursors, from adult and neonatal dog retina and determine if angioblasts are committed to endothelial cell lineage or have the potential to be multipotent, i.e. express phenotypic characteristics of other vascular cell types. Endothelial cells were established from adult dog retina (ADREC) by the technique of Gitlin and D'Amore. For angioblasts, pieces of neonatal day 2 (P2) avascular peripheral retina were placed under coverslips until sufficient cells had explanted. All cells were maintained initially on hyaluronic acid (HA)/fibronectin (FN) substratum. Neonatal canine retinal angioblasts (NCRA) were maintained initially on retinal-derived growth factor with alpha-amino adipic acid to inhibit growth of Muller cells. Cell lines were characterized by enzyme histochemistry [menadione-dependent alpha glycerophosphate dehydrogenase (alphaGPDH), marker for angioblasts] and immunocytochemistry. Once characterized, cells were grown on FN, or collagens I or IV substrata and fed platelet-derived growth factor-BB (PDGF-BB) or fibroblast growth factor-2 (FGF-2). The phenotypic expression of a marker for endothelial cells [acetylated LDL (acLDL) uptake] or a marker for pericytes and smooth muscle cells, production of alpha smooth muscle actin (alphaSMA), was evaluated under those conditions. The canine retinal cell lines that were established had the following characteristics when maintained on serum and a retinal extract. Angioblasts had low expression of vWf and VEGF-R2 (two markers for canine endothelial cells), and very low uptake of acLDL but high expression of alphaGPDH and adenosine A2a receptors (A2aR) (two markers for canine angioblasts in vivo). ADREC had high expression of endothelial cell markers (vWf, VEGF-R2, and acLDL uptake) but minimal expression of alphaGPDH and A2aR. Both angioblasts and endothelial cells expressed CXCR4, a marker for hemangioblasts. Angioblasts grown on any of the substrata in the presence of FGF-2 had high uptake of acLDL and low expression of alphaSMA, while those grown in the presence of PDGF-BB had high expression of alphaSMA and low uptake of acLDL. In conclusion, angioblasts cultured from peripheral vascular retina have low expression of endothelial cell markers and high alphaGPDH and A2aR, markers for canine angioblasts in vivo. Angioblasts will internalize acLDL when maintained on FGF-2 and express alphaSMA when maintained on PDGF-BB, suggesting that they have the potential to become endothelial cells or pericytes, i.e. are multipotent.

Actins↗
PubMed2004

Commonly consumed and naturally occurring dietary substances affect biomarkers of oxidative stress and DNA damage in healthy rats.

The influence of black currant juice, Bowman-Birk protease inhibitor (BBI), kolaviron (a biflavonoid fraction of Garcinia kola seed), sugars, vitamin C and tert-butyl hydroperoxide on a wide range of biomarkers for oxidative stress, DNA damage and sugar or lipid metabolism has been investigated in male F 344 rats. The selected pro-oxidant control, tert-butyl hydroperoxide, significantly increased plasma and liver 2-amino-adipic semialdehyde (AAS), a marker of protein oxidation (p <0.05) whereas lipid oxidation assessed as malon dialdehyde (MDA) and DNA oxidation were not significantly increased. Feeding BBI also increased the level of oxidized protein in plasma and liver at the higher dose level (0.5%). No effect was observed at the lower dose level (0.25%), which even decreased lipid oxidation in plasma. BBI did not affect background levels of DNA strand breaks or oxidation (comets). In rats exposed to black currant juice, a statistically significant decrease in liver AAS and MDA was observed. This effect could not be explained by its content of sugars or of the known redox active constituent, vitamin C. The lowering effect of black currant juice on protein and lipid oxidation was similar in magnitude to that of the known liver protectant, kolaviron. In rats treated with kolaviron (200 mg/kg body weight), background AAS levels were significantly reduced in both plasma and liver whereas the effect on MDA only reached statistical significance in plasma. Kolaviron was the only extract tested which decreased oxidative damage to DNA in the liver. The erythrocyte antioxidant enzyme activities, catalase and glutathione peroxidase were decreased in rats treated with tert-butyl hydroperoxide (p <0.05) but were not affected by the other treatments. Black currant juice and sugars increased plasma triglyceride levels and black currant juice increased plasma cholesterol but neither of them nor any other treatment affected blood glucose, erythrocyte HbA1c or fructosamine. We conclude that markers of oxidative stress may be modified by several mechanisms after feeding rats with complex dietary factors and that both pro- and antioxidant effects may consequently be observed simultaneously after short-term feeding of antioxidant-rich foods, herb medicines, or known pro- and antioxidants.

Animals↗
PubMed2006

Gamma-hydroxybutyric acid stability and formation in blood and urine.

Gamma-hydroxybutyric acid (GHB) can cause problems in interpretation of toxicological findings due to its endogenous nature, significant production in tissues after death and potential formation in stored samples. Our study was designed to determine the influence of storage conditions on GHB levels and its possible in vitro formation in blood and urine in cases where no exogenous use of GHB or its precursors was suspected. The samples were prepared by validated method based on liquid-liquid reextraction with adipic acid internal standard and MSTFA derivatization and assayed on a GC-MS operating in EI SIM mode. The first part of the study was performed with pooled blood and urine samples obtained from living and deceased subjects stored with and without NaF (1% w/v) at 4 and -20 degrees C over 8 months. In ante-mortem samples (both blood and urine) no significant GHB production was found. After 4 months of storage, the substantial GHB rise up to 100 mg/Lwas observed in post-mortem blood stored at 4 degrees C without NaF with subsequent gradual decrease in following months. The inhibition of GHB production was apparent during storage in NaF treated frozen blood samples. In post-mortem urine only slight temporary GHB levels were ascertained (up to 8 mg/L). The second part of our study was aimed to analyse 20 individual post-mortem blood samples stored at 4 degrees C for 16-27 days between autopsy and analysis without preservation followed by storage at 4 degrees C with NaF for 4 months. The temporary GHB production with maximum of 28 mg/Lwas detected in some samples.

Animals↗
PubMed2006

Extractables/leachables from plastic tubing used in product manufacturing.

While the ability of packaging systems to contribute leached substances to finished drug products is well established, increasing interest is being focused on the potential contamination of drug substances by plastic materials encountered during their production. The direct contact of such plastic parts (such as tubing, gaskets, filters and temporary storage containers) with the drug substance at some point in its production raises the possibility that plastic-related contaminants (leachables) may be present in the finished drug product. In this study, eight tubing materials potentially encountered in pharmaceutical production facilities, including six silicone materials and two Santoprene materials, were characterized for their extractable substances by static extraction coupled with comprehensive chemical characterization of the resulting extracts. Based on the extractables profiles thus generated, target leachables were identified for each tubing material. The accumulation of these target leachables was studied by subjecting the tubing to dynamic flow, simulated use extractions. The primary organic extractables from the silicone tubing were a homologous series of silicone oligomers, with most of the tubings demonstrating a unique distribution of oligomers. Several of the silicone tubings also possessed extractable dioctyl phthalate and dioctyl adipate. The primary organic extractables from the Santoprene-type tubing included a number of phthalates, a series of alkyl phenols and decomposition products of Irganox-type antioxidants. Inorganic extractables associated with many of the tubings included Ca, Mg, Zn and B. In general, the levels of targeted leachables extracted from the tubing materials under simulated use (flow) conditions was much smaller than the total amount of these leachables in the tubing.

Drug Contamination↗