Quantification of amino acids in fermentation media by isocratic HPLC analysis of their α-hydroxy acid derivatives

In this paper we describe a novel method for quantification of amino acids. First, α-hydroxy acid derivatives of amino acids were formed after reaction with dinitrogen trioxide by the van Slyke reaction. Second, the α-hydroxy acid derivatives were separated on an Aminex HPX-87H column (Bio-Rad) eluted isocratically with 5 mM H(2)SO(4) and quantified by refractive index detection. We were able to measure the reaction products of 13 of the 20 classical amino acids: glycine, l-alanine, l-valine, l-leucine, l-isoleucine, l-methionine, l-serine, l-threonine, l-asparagine, l-glutamine, l-aspartic acid, l-glutamic acid, and l-proline. We obtained linear relationships between the product peak areas and initial amino acid concentration, whereby the concentrations of these amino acids could be quantified on the basis of the quantification of their products. The method can be used to analyze amino acids in parallel with other small molecules, such as sugars or short chain fatty acids, and was used for parallel quantification of glycine, l-alanine, or l-glutamic acid, and glucose uptake in cultures of the heterotrophic dinoflagellate Crypthecodinium cohnii . The method can also be used to quantify other amines, as demonstrated by detection of Tris (2-amino-2-(hydroxymethyl)propane-1,3-diol).     

Comments by a Councilman

This article presents results from the first statistically significant study of traffic forecasts in transportation infrastructure projects. The sample used is the largest of its kind, covering 210 projects in 14 nations worth U.S.$59 billion. The study shows with very high statistical significance that forecasters generally do a poor job of estimating the demand for transportation infrastructure projects. For 9 out of 10 rail projects, passenger forecasts are overestimated; the average overestima-tion is 106%. For half of all road projects, the difference between actual and forecasted traffic is more than ±20%. The result is substantial financial risks, which are typically ignored or downplayed by planners and decision makers to the detriment of social and economic welfare. Our data also show that forecasts have not become more accurate over the 30-year period studied, despite claims to the contrary by forecasters. The causes of inaccuracy in forecasts are different for rail and road projects, with political causes playing a larger role for rail than for road. The cure is transparency, accountability, and new forecasting methods. The challenge is to change the governance structures for forecasting and project development. Our article shows how planners may help achieve this.

This article was substantially reproduced in the article "Inaccuracy in Traffic Forecasts", published in 2006 in Transport Reviews [Bent Flyvbjerg, Mette Skamris Holm, and Søren L. Buhl, Transport Reviews, Vol. 26, Issue 1, 2006, pp. 1–24 (retracted)] and is the subject of a Notice of Redundant Publication in Journal of the American Planning Association, Vol. 78, Issue 3, page 352, 2012. doi:http://dx.doi.org/10.1080/01944363.2012.719432     

Studies on the antioxidative activity of red pigments in Italian-type dry-cured ham

Aqueous phosphate buffer extracts and acetone/water extracts of pigments from Parma ham were assessed as antioxidants by (1) electron spin resonance spectroscopy using a spin probing technique to evaluate their efficiencies as scavengers of free radicals, and (2) by electrochemical measurement of oxygen depletion rate in an aqueous methyl linoleate emulsion to evaluate their efficiencies as chain-breaking antioxidant, and using both methods, compared with the effect of apomyoglobin and nitrosylmyoglobin. Aqueous phosphate extracts and acetone/water extracts of Parma ham pigment both scavenged a semi-stable nitroxide radical (Fremy's salt), and both extracts reduced the rate of oxygen consumption for lipid peroxidation (initiated by metmyoglobin) very efficiently. For apomyoglobin no antioxidative capacity was observed, and the heme moiety of the pigment(s) of Parma ham were concluded to have antioxidative properties. The more lipophilic pigment, as extracted by acetone/water, had the most significant effect, and its ability to inhibit lipid oxidation was further tested in a model food system based on cooked pork. The lipid oxidation was increasingly inhibited by increasing additions from 0.12 ppm to 0.24 ppm Parma ham pigment, and the pigment protected -tocopherol against degradation in a concentration dependent manner.     

Cultivation and characterization of pterygium as an ex vivo study model for disease and therapy

Purpose

Development of ex vivo model to study pathogenesis, inflammation and treatment modalities for pterygium.

Storage affects the phenolic profiles and antioxidant activities of cherries (Prunus avium L) on human low‐density lipoproteins

Four sweet cherry cultivars (cvs), Burlat, Saco, Summit and Van, were analysed at harvest and after storage at 2 and 15 °C for 30 and 6 days respectively. Phenolic profiles in methanolic extracts of freeze‐dried samples of the fresh and differently stored cherries were quantified by high‐performance liquid chromatography. Hydroxycinnamates dominated in all samples and represented 60–74% by weight of the phenols in the fresh and stored samples of the cvs Saco, Summit and Van, and 45% by weight of the phenols in the cv Burlat samples, which were richer in anthocyanins. The relative and total levels of hydroxycinnamates, anthocyanins, flavonols and flavan‐3‐ols varied among cultivars and during storage. Storage at 15 °C increased the phenol levels, particularly the cyanidin‐3‐rutinoside concentration. Cold storage induced decreased total phenol levels in the cvs Summit and Van but increased total phenol levels in the cvs Burlat and Saco. Phenolic cherry extracts inhibited low‐density lipoprotein oxidation in vitro in a dose‐dependent manner. Extracts of freshly harvested cherries exhibited significantly higher antioxidant activities than extracts of stored samples. The cv Summit samples had the highest antioxidant activity. Differences in the antioxidant effects of the cherry samples were positively correlated with their levels of p‐coumaroylquinic acid (p < 0.1) but negatively correlated with their cyanidin‐3‐rutinoside levels (p < 0.05). Copyright © 2004 Society of Chemical Industry     

The microbial ecology of processing equipment in different fish industries-analysis of the microflora during processing and following cleaning and disinfection

The microflora adhering to the processing equipment during production and after cleaning and disinfecting procedures was identified in four different processing plants. A total of 1009 microorganisms was isolated from various-agar plates and identified. A stepwise procedure using simple phenotypic tests was used to identify the isolates and proved a fast way to group a large collection of microorganisms. Pseudomonas, Neisseriaceae, Enterobactericeae, Coryneform, Acinetobacter and lactic acid bacteria dominated the microflora of cold-smoked salmon plants, whereas the microflora in a plant processing semi-preserved herring consisted of Pseudomonas, Alcaligenes and Enterobactericeae. Psychrobacter, Staphylococcus and yeasts were found in a caviar processing plant. Overall, many microorganisms that are often isolated from fish were also isolated from the fish processing plants. However, some selection depending on processing parameters occurred, since halo- and osmo-tolerant organisms dominated in the caviar processing. After cleaning and disinfection, yeasts, Pseudomonas, Neisseriaceae and Alcaligenes remained in smokehouses, yeasts and Pseudomonas in the herring plant and Pseudomonas, Staphylococcus and yeasts in the caviar plant. The dominant adhering organisms after cleaning and disinfection were pseudomonads and yeasts independently of the microflora during processing. Knowledge of the adhering microflora is essential in the Good Hygienic Practises programme of food processing plants, as the development and design of improved cleaning and disinfecting procedures should target the microorganisms persisting and potentially contaminating the product.     

Water properties and structure of pork sausages as affected by high-pressure processing and addition of carrot fibre

The effects of high-pressure processing (HPP) and addition of carrot fibre on pork sausages have been studied using NMR T? relaxometry and measurements of water-binding capacity (WBC) by centrifugation. Significant effects of temperature (raw, 40, 50, or 60 °C), holding time (1s, 3, 6, or 9 min), and addition of carrot fibre on the distribution and mobility of water were found. However, the effect of carrot fibre could not be explained by structural changes in the sausages when examined by confocal laser scanning microscopy (CLSM). Correlations between T? relaxation measurements and WBC determined by centrifugation revealed that T? relaxation times were able to explain more than 90% of the variation in WBC for both non-pressure and pressure-treated sausages. However, only 49% of the variation was explained for pressure-treated sausages with carrot fibre, indicating that combining addition of fibre and high pressure treatment causes non-coherent changes in T? NMR relaxation times.     

Recognition and uptake of free and nanoparticle-bound betalactoglobulin--a food allergen--by human monocytes

Scope : To improve our understanding of the interaction of food allergens with cells of the immune system, the endocytosis by human monocytes of bovine β‐lactoglobulin (BLG) and ovomucoid (OM) – two major food allergens – and human serum albumin (HSA) was studied. Methods and results : BLG was covalently conjugated to dextran‐coated magnetic nanoparticles (MNPs) without affecting its structure and immunoreactivity. BLG‐conjugated MNPs were taken up by human monocytes much more efficiently than non‐conjugated MNPs, allowing easy magnetic separation of cells that had adsorbed the allergen. BLG, OM, and HSA were conjugated to MNPs also labeled with a fluorescent probe. The uptake of these materials by human monocytes was monitored through flow cytometry, and compared with fluorescent MNPs and the free fluorescently labeled proteins, confirming higher uptake of the BLG‐conjugated MNPs versus non‐conjugated MNPs. OM but not HSA conjugation to particles enhanced uptake of the MNPs. Confocal microscopy provided direct evidence of the actual internalization of BLG–MNP conjugates into the cytoplasm. Conclusions : These results contribute to the current understanding of the interaction between food allergens and antigen‐presenting cells, and demonstrate that the BLG is readily endocytosed by monocytes both as the single protein and as a conjugate.