New perspectives on perfluorochemical ecotoxicology: inhibition and induction of an efflux transporter in the marine mussel, Mytilus californianus

The toxicological effects of perfluoroalkyl acids on the p-glycoprotein (p-gp) cellular efflux transporter were investigated using the marine mussel Mytilus californianus as a model system. Four of the perfluoroalkyl acids studied exhibit chemosensitizing behavior, significantly inhibiting p-gp transporter activity. The inhibitory potency is maximal for the longer chain acids perfluorononanoate (PFNA) and perfluorodecanoate (PFDA), with average IC50 values of 4.8 and 7.1 microM, respectively. Results indicate that PFNA inhibits p-gp by an indirect mechanism, and this inhibition is reversible and accompanied by a rapid loss of PFNA from the tissue. In addition, PFNA induces expression of the p-gp transporter after a 2-h exposure, a stress response that may result in a metabolic cost to the organism. Given that most organisms, including humans, share efflux transporters as a first line of defense against toxicants, the results of this study may have broader implications for the ecotoxicology of perfluoroalkyl acids.     

Deuterium enhancement in H3+ in pre-stellar cores

Deuterium enhancement of monodeuterated species has been recognized for more than 30 years as a result of chemical fractionation that results from the difference in zero-point energies of deuterated and hydrogenated molecules. The key reaction is the deuteron exchange in the reaction between HD, the reservoir of deuterium in dark interstellar clouds, and the H3+ molecular ion, leading to the production of H2D+ molecule, and the low temperature in dark interstellar clouds favours this production. Furthermore, the presence of multiply deuterated species have incited our group to proceed further and consider the subsequent reaction of H2D+ with HD, leading to D2H+, which can further react with HD to produce D3+. In pre-stellar cores, where CO was found to be depleted, this production should be increased as CO would normally destroy H3+. The first model including D2H+ and D3+ predicted that these molecules should be as abundant as H2D+. The first detection of the D2H+ was made possible by the recent laboratory measurement for the frequency of the fundamental line of para-D2H+. Here, we present observations of H2D+ and D2H+ towards a sample of dark clouds and pre-stellar cores and show how the distribution of ortho-H2D+ (1(1,0)-1(1,1)) can trace the deuterium factory in pre-stellar cores. We also present how future instrumentation will improve our knowledge concerning the deuterium enhancement of H3+.     

Radical scavenging by dietary flavonoids. A kinetic study of antioxidant efficiencies

The rate of scavenging of peroxyl radicals and of diphenylpicrylhydrazyl radicals by flavanones, flavones, flavanols and flavonols commonly occurring in foods was found, when determined by ESR spin trapping and by stopped-flow spectroscopy, respectively, to be the highest for the most reducing of the flavonoids. Among 12 flavonoids investigated a threshold potential of approximately +0.4 V seems to exist, above which the flavonoids become ineffective radical scavengers, as determined by ESR spectroscopy using 5,5-dimethyl-1-pyrroline-N-oxide as spin trap. Scavenging of diphenylpicrylhydrazyl radicals showed (pseudo) first order kinetics for excess of flavonoids, and the reaction half-life could also be determined for the less effective flavonoids. For kaempferol and eriodictyol, flavonoids with very similar reduction potentials for their phenoxyl radicals, rate constants for the bimolecular scavenging could be determined in methanol and had the value (7.0 ±0.7)×10² s⁻¹ M⁻¹ and 33±1 s⁻¹ M⁻¹ at 25°C, respectively, showing that other factors than the driving force, such as hydrophilic/lipophilic balance are important, as was further confirmed in an oxygen consumption assay based on a linoleic acid emulsion, where decreasing antioxidant efficiency followed the order: (+)-catechin>(±)-taxifolin≥luteolin>kaempferol>quercetin>>naringenin. Received: 5 November 1999     

Reviews

Olav Arnfinn Laudal; Ragni Piene (eds.), The Legacy of Niels Henrik Abel. The Abel Bicentennial, reviewed by Henrik Kragh Sørensen Michael Weichenhan, “Ergo perit coelum…” Die Supernova des Jahres 1572 und die Überwindung der aristotelischen Kosmologie, Boethius. Texte und Abhandlungen zur Geschichte der Mathematik und der Naturwissenschaften, vol. 49, reviewed by Charlotte Methuen     

The Biochemical Assessment of Mitochondrial Respiratory Chain Disorders

Mitochondrial respiratory chain (MRC) disorders are a complex group of diseases whose diagnosis requires a multidisciplinary approach in which the biochemical investigations play an important role. Initial investigations include metabolite analysis in both blood and urine and the measurement of lactate, pyruvate and amino acid levels, as well as urine organic acids. Recently, hormone-like cytokines, such as fibroblast growth factor-21 (FGF-21), have also been used as a means of assessing evidence of MRC dysfunction, although work is still required to confirm their diagnostic utility and reliability. The assessment of evidence of oxidative stress may also be an important parameter to consider in the diagnosis of MRC function in view of its association with mitochondrial dysfunction. At present, due to the lack of reliable biomarkers available for assessing evidence of MRC dysfunction, the spectrophotometric determination of MRC enzyme activities in skeletal muscle or tissue from the disease-presenting organ is considered the ‘Gold Standard’ biochemical method to provide evidence of MRC dysfunction. The purpose of this review is to outline a number of biochemical methods that may provide diagnostic evidence of MRC dysfunction in patients.     

Production and oxidative stability of a human milk fat substitute produced from lard by enzyme technology in a pilot packed-bed reactor

Residence time and time of production were investigated during the enzymatic production of a specific structured lipid/human milk fat substitute (SL-HMFS), on a kg scale, made from lard and soybean oil fatty acids, using a packed-bed reactor and short path distillation. There were no effects of residence time or time of production on C18:2 and C18:3 incorporation or on acyl migration in the sn-2 position. In addition, the SL-HMFS was compared to commercial human milk fat substitute (HMFS) regarding fatty acid composition, content of antioxidants and oxidative stability. Fats were stored at 60 °C for four days and the oxidative stability evaluated by analysis of peroxide value (PV) and volatile secondary oxidation products. SL-HMFS had a lower oxidative stability than did commercial HMFS products or lard, probably due to a lower level of tocopherol in SL-HMFS.     

Interlaboratory study on lipid oxidation during accelerated storage trials with rapeseed and sunflower oil analyzed by conjugated dienes as primary oxidation products

Accelerated storage tests are frequently used to assess the oxidative stability of foods and related systems due to its reproducibility. Various methods and experimental conditions are used to measure lipid oxidation. Differences between laboratories make it necessary to determine the repeatability and reproducibility of oxidation tests performed under the same conditions. The objective of the present interlaboratory study was to evaluate the outcome of a storage test for two different bulk oils, sunflower oil (SFO) and rapeseed oil (RSO), during a period of 9 weeks at 20°C, 30°C, 40°C, and 60°C. Sixteen laboratories were provided with bottled oils and conducted the storage tests according to a detailed protocol. Lipid oxidation was monitored by the formation of conjugated dienes (CD) and the activation energy (E_a) was determined for comparative purposes and statistically evaluated. An increase in CD formation was observed for both oils when the storage temperature was increased in all laboratories. The E_a,1 ranged from 47.9 to 73.3 kJ mol⁻¹ in RSO and from 27.8 to 62.6 kJ mol⁻¹ in SFO, with average values of 58.2 and 46.8 kJ mol⁻¹, respectively. The reproducibility coefficients were 10.9% and 18.2% for RSO and SFO, respectively. Practical applications: In order to compare results on oxidative stability of foods derived from different studies, the reproducibility of storage tests and methods employed to evaluate the oxidation level should be considered. This study provides fundamental data on the reproducibility of lipid oxidation under accelerated storage conditions and defines important parameters to be considered for the conduction of experiments.     

Fast Self-Assembly Dynamics of a β-Sheet Peptide Soft Material

Peptide-based hydrogels are promising biocompatible materials for wound healing, drug delivery, and tissue engineering applications. The physical properties of these nanostructured materials depend strongly on the morphology of the gel network. However, the self-assembly mechanism of the peptides that leads to a distinct network morphology is still a subject of ongoing debate, since complete assembly pathways have not yet been resolved. To unravel the dynamics of the hierarchical self-assembly process of the model β-sheet forming peptide KFE8 (Ac-FKFEFKFE-NH₂)_, high-speed atomic force microscopy (HS-AFM) in liquid is used. It is demonstrated that a fast-growing network, based on small fibrillar aggregates, is formed at a solid–liquid interface, while in bulk solution, a distinct, more prolonged nanotube network emerges from intermediate helical ribbons. Moreover, the transformation between these morphologies has been visualized. It is expected that this new in situ and in real-time methodology will set the path for the in-depth unravelling of the dynamics of other peptide-based self-assembled soft materials, as well as gaining advanced insights into the formation of fibers involved in protein misfolding diseases.     

Selective and Sustainable Production of Sub-terminal Hydroxy Fatty Acids by a Self-Sufficient CYP102 Enzyme from Bacillus Amyloliquefaciens

Enzymatic hydroxylation of fatty acids by Cytochrome P450s (CYPs) offers an eco-friendly route to hydroxy fatty acids (HFAs), high-value oleochemicals with various applications in materials industry and with potential as bioactive compounds. However, instability and poor regioselectivity of CYPs are their main drawbacks. A newly discovered self-sufficient CYP102 enzyme, BAMF0695 from Bacillus amyloliquefaciens DSM 7, exhibits preference for hydroxylation of sub-terminal positions (ω-1, ω-2, and ω-3) of fatty acids. Our studies show that BAMF0695 has a broad temperature optimum (over 70 % of maximal enzymatic activity retained between 20 to 50 °C) and is highly thermostable (T₅₀ >50 °C), affording excellent adaptive compatibility for bioprocesses. We further demonstrate that BAMF0695 can utilize renewable microalgae lipid as a substrate feedstock for HFA production. Moreover, through extensive site-directed and site-saturation mutagenesis, we isolated variants with high regioselectivity, a rare property for CYPs that usually generate complex regioisomer mixtures. BAMF0695 mutants were able to generate a single HFA regiosiomer (ω-1 or ω-2) with selectivities from 75 % up to 91 %, using C12 to C18 fatty acids. Overall, our results demonstrate the potential of a recent CYP and its variants for sustainable and green production of high-value HFAs.