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Validation of sterilizing grade filtration.

Validation consideration of sterilizing grade filters, namely 0.2 micron, changed when FDA voiced concerns about the validity of Bacterial Challenge tests performed in the past. Such validation exercises are nowadays considered to be filter qualification. Filter validation requires more thorough analysis, especially Bacterial Challenge testing with the actual drug product under process conditions. To do so, viability testing is a necessity to determine the Bacterial Challenge test methodology. Additionally to these two compulsory tests, other evaluations like extractable, adsorption and chemical compatibility tests should be considered. PDA Technical Report # 26, Sterilizing Filtration of Liquids, describes all parameters and aspects required for the comprehensive validation of filters. The report is a most helpful tool for validation of liquid filters used in the biopharmaceutical industry. It sets the cornerstones of validation requirements and other filtration considerations.

Drug Industry↗

[Method for developing a controlled process. Study of the possible regulation of rifamycin B biosynthesis].

A procedure for developing a controlled process for biosynthesis of secondary metabolites is described with reference to rifamycin B as an example. The response of the antibiotic producing culture to changed concentrations of the main nutrients in the initial medium was determined. Mathematical processing of the experimental findings with design of the experiment resulted in defining nutrients such as ammonium sulfate and cornsteep liquor useful for further development of the controlled process.

Ammonium Sulfate↗

Production of vitamins, coenzymes and related biochemicals by biotechnological processes.

Vitamins and related biofactors belong to those few chemicals with a direct positive appeal to people. There is indeed a large need for extra vitamins, other than those derived from plant and animal food sources, due to unbalanced food habits or processing, food shortage or disease. Added vitamins are now either prepared chemically or biotechnologically via fermentation or bioconversion processes. Several vitamins and related biofactors are now only or mainly produced chemically (vitamin A, cholecalciferol (D3), tocopherol (E), vitamin K2, thiamine (B1), niacin (PP or B3), pantothenic acid (B5), pyridoxine (B6), biotin (H or B8), folic acid (B9) or via extraction processes (beta-carotene or provitamin A, provitamin D3, tocopherol, vitamin F-group). However, for several of these compounds microbiological or algal methods also exist or are rapidly emerging. Others are produced practically exclusively via fermentation (ergosterol or provitamin D2, riboflavin (B2), cyanocobalamin (B12), orotic acid (B13), vitamin F-group, ATP, nucleosides, coenzymes, etc. or via microalgal culture (beta-carotene, E, F). Both chemical and microbial processes are run industrially for vitamin B2 while vitamin C (ascorbic acid) is produced via a combination of chemical reactions and fermentation processes. A survey is given here of the current state of vitamin production, with emphasis on developments and strategies for improved biotechnological production and its significance, as compared to existing chemical processes. The screening or construction of vitamin hyperproducing microbial strains is a difficult task; pathway elucidation and metabolic (de)regulation need further study; r-DNA technology has only recently been introduced; improved fermentation processes and immobilised biocatalysts bioconversions for the synthesis of chiral vitamin compounds or intermediates or derivatives are gaining importance; the recovery and purification of these vitamin compounds from their fermentation broths remains equally complex.

Biotechnology↗

Gene expression profiles for detecting and distinguishing potential endocrine-disrupting compounds in environmental samples.

Industrial and municipal processes may produce and release endocrine-disrupting compounds (EDCs) into the environment, but the exact nature of their effects is difficult to investigate. EDCs typically exert their effect by affecting gene expression aberrantly. To determine if gene expression profiles could be used to detect and distinguish estrogenic EDCs, an estrogen receptor positive human breast cancer cell line (MCF-7) was exposed to known estrogenic compounds, suspected EDCs, and extracts from three effluent samples. A set of specifically estrogen-regulated genes was identified by microarray analysis. Nine estrogen up-regulated genes (IGFBP4, HSPA8, B4GALT1, XBP1, KRT8, GTPBP4, HNRPAB, SLC2A1, and CALM1) and two estrogen down-regulated genes (ID2 and ZNF217) were consistently detectable in response to estrogen and other estrogenic compounds. Gene expression patterns in cells that were exposed to effluent sample extracts were compared to gene expression patterns in cells that were exposed to known endocrines. Using this technique, two of the effluent samples were shown to have estrogenic activity. This approach could easily be extended to screen for other types of receptor-mediated endocrine disruption. For example, cells expressing androgen or aryl hydrocarbon receptors could be used in gene expression profiling assays to detect androgenic effects or for the presence of bioactive aromatic hydrocarbons. Gene expression profiling is emerging as a sensitive and specific method to screen complex samples for endocrine disrupting activity.

Androgens↗

[Obtaining beta-galactosidase using whey].

The search of a microorganism with the ability to produce the enzyme beta-galactosidase was undertaken according with the requirements of the market, in economical, technological and sanitary terms. The process consisted of recovery and use of the effluent from milk and cheese used to feed pigs, producing at the same time different types of contamination; once investigated and adjusted the technological variables to produce the enzyme, and selected the most convenient microorganism for such purpose the acting upon the extraction, conservation and purification of the product were adjusted. Comparative results of conversion were obtained with different test, at laboratory scale and industrial plants, in similar conditions to those obtained with importation products.

Animals↗

Degradation of polyethoxylated nonylphenols in a sewage treatment plant. Quantitative analysis by isotopic dilution-HRGC/MS.

Polyethoxylated alkylphenols (APnEO, where n is the number of ethylene oxide molecules), are non-ionic surfactants widely used for domestic and industrial purposes. Most of APnEO are polyethoxylated nonylphenols (NPnEO). NPnEO are widespread environmental pollutants with relatively low toxicity for mammals and higher toxicity for aquatic organisms. In addition, they have been described as endocrine disrupters in recent publications. One of the main problems related to these surfactants is their uncomplete degradation, even in the most effective sewage treatment plants. Usually, the final products, more toxic and resistant to biological degradation than NPnEO, are nonylphenol (NP), monoethoxylated nonylphenol (NP1EO), diethoxylated nonylphenol (NP2EO), nonylphenoxy acetic acid (NP1EC), and nonylphenoxyethoxy acetic acid (NP2EC). In this paper, the degradation of NPnEO was studied in the different processes of a sewage treatment plant. For this purpose, NP, NP1EO and NP2EO were analysed in composite samples collected at different points along the plant (influent, pre-treatment effluent, primary effluent, plant effluent). Analyses were carried out by isotopic dilution-HRGC/MS, using available labelled nonylphenols (13C6-NP, 13C6-NP1EO, 13C6-NP2EO) as internal standards. Extraction of NPnEO from aqueous samples, previous to analysis, was performed by the Likens-Nickerson method (simultaneous steam distillation/solvent extraction, SDE).

Biodegradation, Environmental↗

Removal of PAHs from highly contaminated soils found at prior manufactured gas operations.

Removal of PAHs from highly contaminated soil found at a manufactured gas site was evaluated using solvent washing with mixed solvents. The following solvents were considered as water miscible co-solvents in mixed solvents: ethanol, 2-propanol, acetone, and 1-pentanol. In batch solvent extraction of soil, ethanol and 2-propanol were selected as primary components of mixed solvents in addition to 1-pentanol. Using ternary solutions containing either ethanol or 2-propanol with a volume fraction of 1-pentanol ranging from 5 to 25% and a water volume fraction ranging from 5 to 30%, ethanol was more effective than 2-propanol in extracting PAHs from soil. A solvent mixture of 5% 1-pentanol, 10% water and 85% ethanol was selected as the extraction solvent. Using a 1g:4ml soil:solvent extraction ratio, extraction kinetics showed that from 65 to 90% of the extractable PAHs were removed within an hour of contact between soil and solvent. Using this 1g:4ml extraction ratio, PAHs were removed in a three-stage cross-current solvent washing process where the same batch of soil was extracted with clean solvent for 1h in each stage. PAH removals in three-stage cross-current solvent washing were comparable to PAH removals obtained with Soxhlet extraction.

2-Propanol↗

Natural and anthropogenic hydrocarbon inputs to sediments of Patos Lagoon Estuary, Brazil.

The Patos Lagoon Estuary, southern Brazil, is an area of environmental interest not only because of tourism, but also because of the presence of the second major port of Brazil, with the related industrial and shipping activities. Thus, potential hydrocarbon pollution was examined in this study. Sediment samples were collected at 10 sites in the estuary, extracted, and analyzed by GC-FID and GC-MS for composition and concentration of the following organic geochemical markers: normal and isoprenoid alkanes, petroleum biomarkers, linear alkylbenzenes (LABs), and polycyclic aromatic hydrocarbons (PAHs). The total concentrations varied from 1.1 to 129.6 microg g(-1) for aliphatic hydrocarbons, from 17.8 to 4510.6 ng g(-1) for petroleum biomarkers, from 3.2 to 1601.9 ng g(-1) for LABs, and from 37.7 to 11,779.9 ng g(-1) for PAHs. Natural hydrocarbons were mainly derived from planktonic inputs due to a usual development of blooms in the estuary. Terrestrial plant wax compounds prevailed at sites located far from Rio Grande City and subject to stronger currents. Anthropogenic hydrocarbons are related to combustion/pyrolysis processes of fossil fuel, release of unburned oil products and domestic/industrial waste outfalls. Anthropogenic hydrocarbon inputs were more apparent at sites associated with industrial discharges (petroleum distributor and refinery), shipping activities (dry docking), and sewage outfalls (sewage). The overall concentrations of anthropogenic hydrocarbons revealed moderate to high hydrocarbon pollution in the study area.

Alkanes↗

[Symptoms and functional disorders of the respiratory system caused by exposure to tea dust].

A case report is presented of a female worker of a tee packing factory, with a 5 years history of tea dust exposition and symptoms of reversible airway obstruction and early allergic alveolitis. The results of spirometric and lung mechanics demonstrated reversible airway obstruction with co-existent restrictive changes, lowering of CO diffusing capacity (DLc0). Bronchial provocation tests with tea dust were carried out thrice and all were positive. The authors believe that the etiological factor of above mentioned changes were Aspergillus niger spores found in tea dust. Tea dust and the patient's sputum cultured for fungi revealed massive growth of A. niger. Serological tests revealed additionally, antibodies to A. niger and tea dust extracts. The authors discuss the possible mechanisms of airway disturbances in this patient.

Adult↗

Effect of phosphate concentration on the production of dextransucrase by Leuconostoc mesenteroides NRRL B512F.

Leuconostoc mesenteroides NRRL B512F is the main strain used in industrial fermentations to produce dextransucrase and dextran. This process has been studied since the Second World War, when it was used as blood plasma expander. A study about the effect of phosphate concentration on cell propagation in a semicontinuous shake-flask culture is described in this work. Dextransucrase is obtained by fermentation of the Leuconostoc mesenteroides NRRL B512F in the presence of sucrose as substrate, a nitrogen source (corn liquor or yeast extract) and minerals. Phosphate is currently used in order to buffer the culture medium. Cell propagation can be done through a repeated batch culture, where dilution in a fresh medium is made with relatively short periods. The standard medium for dextransucrase production is prepared using 0.1 M of K(2)HPO(4). In this work the level of phosphate was increased to 0.3 M, and an increase on biomass and on the enzyme activity was found when phosphate enriched medium was used. Higher phosphate buffer concentration was also able to keep the pH values above 5.0 during the entire process, avoiding enzyme denaturation.

Bioreactors↗

Laccase: new functions for an old enzyme.

Laccases occur widely in fungi; they have been characterized less frequently in higher plants. Here we have focused on more recent reports on the occurrence of laccase and its functions in physiological development and industrial utility. The reports of molecular weights, pH optima, and substrate specificity are extremely diverse. Conclusive proof of the occurrence of laccase in a tissue must demonstrate that the enzyme be able to oxidize quinol with concomitant uptake of oxygen. Laccase is involved in the pigmentation process of fungal spores, the regeneration of tobacco protoplasts, as fungal virulence factors, and in lignification of cell walls and delignification during white rot of wood. Commercially, laccases have been used to delignify woody tissues, produce ethanol, and to distinguish between morphine and codeine. A very wide variety of bioremediation processes employ laccase in order to protect the environment from damage caused by industrial effluents. Research in recent years has been intense, much of it elicited by the wide diversity of laccases, their utility and their very interesting enzymology.

Animals↗

Monitoring a complex medium fermentation with sample-sample two-dimensional FT-NIR correlation spectroscopy.

The possibility of monitoring state transitions during the time course of a fermentation through the analysis of the on-line collected near-infrared (NIR) spectra with sample-sample two-dimensional correlation spectroscopy (SS-2DCoS) was investigated. SS-2DCoS has proved to be useful for extracting process information directly from the spectra collected on-line. The complexity of the system studied prevented the extraction of concentration profiles, but nevertheless, the application of SS-2DCoS enables the identification of fermentation state transitions due to metabolic and morphological changes of the microorganism.

Cells, Cultured↗

Treatability of chromite ore processing waste by leaching.

Developing treatment and disposal strategies and health-based clean-up standards for chromium containing wastes continues to be an important environmental regulatory issue because of the opposing solubility and toxicity characteristics of chromium species under diverse environmental conditions. In this study, leaching characteristics of total Cr and Cr(VI) were investigated using laboratory column studies. The data obtained from the experimental studies were analysed to assess the treatability of chromite ore processing waste (COPW) by leaching and to identify the leaching strategies that enhance mass removal rates of chromium species. COPW used for laboratory soil column studies was obtained from an industrial plant producing sodium chromate in Mersin, Turkey. Laboratory investigations involved chemical characterisation of waste material and column studies. For waste characterisation, U.S. EPA toxicity characterisation leaching procedure (TCLP) was performed on COPW to determine the concentrations of metal species in the TCLP extract. For column studies, various laboratory columns containing plain COPW material, 1:1 COPW/reducing agent (elemental iron or manure) mixture and different type soils (sand, loam and clay) overlain by COPW were subjected to leaching tests using acidic, neutral and alkaline influent water to determine Cr mass leaching efficiencies. Based on the TCLP analyses, COPW is classified as hazardous waste. As a result of comparing the leaching efficiency data from twelve leaching columns, the maximum removal of total Cr was achieved by leaching COPW/manure mixture using acidic (pH 4.78) influent water. The highest Cr(VI) leaching efficiency was achieved in the columns of plain COPW and COPW/manure mixture using highly alkaline (pH 12.0) influent water. The least effective leaching efficiency for both total Cr and Cr (VI) was obtained by leaching plain COPW with neutral (pH 7.0) influent water. Land-disposal of the treated COPW material by mixing with clayey soils seems to be a viable alternative.

Chromium↗

Effect of enzymatic treatment of extracted sunflower proteins on solubility, amino acid composition, and surface activity.

Industrial proteins from agriculture of either animal or vegetable origin, including their peptide derivatives, are of great importance, from the qualitative and quantitative point of view, in food formulations (emulsions and foams). A fundamental understanding of the physical, chemical, and functional properties of these proteins is essential if the performance of proteins in foods is to be improved and if underutilized proteins, such as plant proteins (and their hydrolysates and peptides derivatives), are to be increasingly used in traditional and new processed food products (safe, high-quality, health foods with good nutritional value). In this contribution we have determined the main physicochemical characteristics (solubility, composition, and analysis of amino acids) of a sunflower protein isolate (SPI) and its hydrolysates with low (5.62%), medium (23.5%), and high (46.3%) degrees of hydrolysis. The hydrolysates were obtained by enzymatic treatment with Alcalase 2.4 L for DH 5.62 and 23.5% and with Alcalase 2.4 L and Flavorzyme 1000 MG sequentially for DH 46.3%. The protein concentration dependence on surface pressure (surface pressure isotherm), a measure of the surface activity of the products (SPI and its hydrolysates), was obtained by tensiometry. We have observed that the degree of hydrolysis has an effect on solubility, composition, and content of the amino acids of the SPI and its hydrolysates. The superficial activity and the adsorption efficiency were also affected by the degree of hydrolysis.

Adsorption↗

Colloidal interactions between Langmuir-Blodgett bitumen films and fine solid particles.

In oil sand processing, accumulation of surface-active compounds at various interfaces imposes a significant impact on bitumen recovery and bitumen froth cleaning (i.e., froth treatment) by altering the interfacial properties and colloidal interactions among various oil sand components. In the present study, bitumen films were prepared at toluene/water interfaces using a Langmuir-Blodgett (LB) upstroke deposition technique. The surface of the prepared LB bitumen films was found to be hydrophobic, comprised of wormlike aggregates containing a relatively high content of oxygen, sulfur, and nitrogen, indicating an accumulation of surface-active compounds in the films. Using an atomic force microscope, colloidal interactions between the LB bitumen films and fine solids (model silica particles and clay particles chosen directly from an oil sand tailing stream) were measured in industrial plant process water and compared with those measured in simple electrolyte solutions of controlled pH and divalent cation concentrations. The results show a stronger long-range repulsive force and weaker adhesion force in solutions of higher pH and lower divalent cation concentration. In plant process water, a moderate long-range repulsive force and weak adhesion were measured despite its high electrolyte content. These findings provide more insight into the mechanisms of bitumen extraction and froth treatment.

Journal Article↗

Engineering of an oenological Saccharomyces cerevisiae strain with pectinolytic activity and its effect on wine.

A pectinolytic industrial yeast strain of Saccharomyces cerevisiae was generated containing the S. cerevisiae endopolygalacturonase gene (PGU1) constitutively expressed under the control of the 3-phosphoglycerate kinase gene (PGK1) promoter. The new strain contains DNA derived exclusively from yeast and expresses a high polygalacturonic acid hydrolyzing activity. Yeast transformation was carried out by an integrative process targeting a dispensable upstream region of the acetolactate synthase locus (ILV2), which determines sulfometuron methyl resistance. Microvinification assays were performed on white and red musts with the transformed UCLMS-1M strain and with the same strain untransformed. It was found that the changes in the pectic polysaccharide contents did not directly affect the taste or flavor of the wine. From the data reported, it is deduced that the chief advantage of using the modified strain is that it improves the yield of must/wine extraction, while it also positively affects some variables relating to appearance.

DNA, Fungal↗

Solid-phase extraction in malondialdehyde analysis.

Malondialdehyde (MDA) has been widely used as an index of lipoperoxidation in biological and medical sciences as well as in the food industry. A solid-phase extraction (SPE) of the condensation product of the MDA with 2-thiobarbituric acid (TBA) was developed using LiChrolut C18ec, 200 mg (Merck, Darmstadt, Germany), as a SPE cartridge and methanol as an eluent for sample pretreatment before HPLC analysis. The samples of blood plasma, platelet concentrates, or erythrocyte membranes (ghosts) were deproteinized by acetonitrile in the presence of sodium hydroxide prior to the reaction with TBA. The reaction mixture was processed using SPE. The SPE extracts (800 microL of methanol) were put to dryness and after dissolution with 100 microliters of mobile phase, 50 microliters was analyzed by RP-HPLC with fluorescence detection (excitation at 514 nm, emission at 556 nm). The mean MDA concentration in plasmas of 32 healthy donors was 0.37 +/- 0.25 mumol/L and the mean MDA concentration in normal ghosts was 8.3 +/- 4.1 pmol/microgram of protein content. In the case of a patient with a severe form of beta-thalassemia, the concentration of plasma MDA was raised to 1.22 mumol/L and the amount of MDA in erythrocytal ghosts was raised to 21.05 pmol/microgram of protein content. MDA concentration in platelet concentrates (six bags) in the first day of storage was 0.46 +/- 0.18 mumol/L and in the fifth day of storage was 0.55 +/- 0.44 mumol/L.

Blood Platelets↗

Fish and shellfish upgrading, traceability.

Recognition of the limited biological resources and the increasing environmental pollution has emphasised the need for better utilisation of by-products from the fisheries. Currently, the seafood industry is dependent on the processing of the few selected fish and shellfish species that are highly popular with consumers but, from economic and nutritional points of view, it is essential to utilise the entire catch. In this review, we will focus on recent developments and innovations in the field of underutilised marine species and marine by-product upgrading and, more precisely, on two aspects of the bioconversion of wastes from marine organisms, i.e. extraction of enzymes and preparation of protein hydrolysates. We will deal with the question of accurate determination of fish species at the various steps of processing. Methods of genetic identification applicable to fresh fish samples and to derived products will be described.

Animals↗