A trout (Oncorhynchus mykiss) perfusion model approach to elucidate the role of blood removal for lipid oxidation and colour changes in ice-stored fish muscle

Whole body saline-perfused rainbow trout (Oncorhynchus mykiss) was ice-stored for 4 weeks and compared with unwashed/washed minces from unbled and bled trout in terms of rancid odour, peroxide value (PV), thiobarbituric acid reactive substances (TBARS) and redness loss. Muscle from saline-perfused fish, which had 72% less total haem, was deficient in rancid odour during the whole storage, while bled (54% less haem) and unbled samples developed rancid odour already after ~4 and 2 days; higher intensity without bleeding. PV/TBARS also developed in the order unbled > bled > perfused samples; however, PV/TBARS were not as completely prevented as rancid odour after perfusion. Saline washing (3 × 3 volumes) of unbled mince removed 84% haem and yielded the second most stable sample while saline washing (1 × 1 volumes) destabilised unbled mince, despite 64% haem removal. Concurrent antioxidant removal during washing of minces obviously counteracted the effect of blood removal and washing fish mince with small volumes of solution should be used with great care.     

The cytochrome c content of various poultry meats

The cytochrome c content and total haem pigments in the breast and thigh meat of broiler chickens, hens out of lay, turkeys, geese, duckling (Peking), and muscovy ducks were determined. The levels of cytochrome c in the breast and thigh meat were: for chicken 11.4 and 35.5; for hens 14.0 and 84.2; for turkeys 13.1 and 47.4; for geese 141.0 and 107.8; for duckling 111.2 and 149.9 and for muscovy ducks 81.5 and 97.3 μg per g of tissue, respectively. The correlation coefficient between cytochrome c and total haem pigments concentration was r=0.98 for breast, and r=0.85 for thigh meat. The authors postulate that the cytochrome c may play a noticeable role in the colour of fresh and processed poultry meat, but further investigations are needed to confirm this.     

Effects of Various Viscosity Enhancers and pH on Separating Haem from Porcine Red Blood Cells

Haem derived from animal blood is a valuable source of organic iron which may be used as a supplement in foods. The purpose of this study was to examine the effects of sodium carboxymethyl cellulose (NaCMC), propylene glycol alginate (PGA) and sodium alginate (SA) as viscosity enhancers on the separation of haem from porcine red blood cells, and on the distribution of iron between the haem and non-haem fractions of the extracts. Iron determinations were carried out spectrophotometrically on the supernatants obtained by centrifuging the cell suspensions and on two successive washings of the cells. In the first washings the total extracted iron content using NaCMC was highest when a high-viscosity grade was used; there were no significant differences (P>0·05) between the results obtained with this material and those obtained using PGA or SA. There were also no significant differences (P>0·05) among the three treatments in terms of non-haem content. The absorbances at 540nm of the first washings were lower when the washing was carried out at pH 1·4 rather than at pH 1·0 or 2·0. There were no significant differences associated with changes in the viscosity enhancer in the absorbances of the second washings, but the absorbances at pH 1·4 were generally higher than those at pH 1·0 or 2·0. Enhancement of the absorbances on the addition of 1,10-o-phenanthroline indicated that the extracted iron was present in the ferrous form.     

1H-NMR study of bovine myoglobin autoxidation. Influence of muscle type and time post mortem

¹H NMR spectroscopy was used to determine first order rate constants at four temperatures (300, 304, 308, 312 K) and activation energies of the autoxidation reaction for oxymyoglobin. the haeminic pigment was purified from two bovine muscles with different colour stabilities ( psoas major (PM) and longissimus lumborum (LL)) at 2h (day 0) and 192 h (day 8) post mortem. to characterize this autoxidation reaction, we have focused attention on the time-temperature dependent disappearance of the Val-E11 methyl group signal. This study showed that, whatever the time post mortem , although the myoglobin autoxidation rate was greater for PM than for LL muscle, the activation energies were similar. It was also worth noting that, in the range 300–312 K, the average ratio of autoxidation rate constants between day 8 and day 0 was near 1.6 for the two muscles studied. It is reasonable to think that oxidative processes developed during 8 days meat storage have led to a structural change within the cavity of the heme pocket of the myoglobins. Moreover, only one orientation of the porphyrin within the heme pocket was noted for the two muscles studied.     

Comparison of the chemical properties of iron and cobalt porphyrins and corrins

Density functional calculations have been used to compare various geometric, electronic and functional properties of iron and cobalt porphyrin (Por) and corrin (Cor) species. The investigation is focussed on octahedral M(II/III) complexes (where M is the metal) with two axial imidazole ligands (as a model of b and c type cytochromes) or with one imidazole and one methyl ligand (as a model of methylcobalamin). However, we have also studied some five-coordinate M(II) complexes with an imidazole ligand and four-coordinate M(I/II) complexes without any axial ligands as models of other intermediates in the reaction cycle of coenzyme B12. The central cavity of the corrin ring is smaller than that of porphine. We show that the cavity of corrin is close to ideal for low-spin Co(III), Co(II), and Co(I) with the axial ligands encountered in biology, whereas the cavity in porphine is better suited for intermediate-spin states. Therefore, the low-spin state of Co is strongly favoured in complexes with corrins, whereas there is a small energy difference between the various spin states in iron porphyrin species. There are no clear differences for the reduction potentials of the octahedral complexes, but [Co(I)Cor] is more easily formed (by at least 40 kJ mole(-1)) than [Fe(I)Por]. Cobalt and corrin form a strong Cobond;C bond that is more stable against hydrolysis than iron and porphine. Finally, Fe(II/III) gives a much lower reorganisation energy than Co(II/III); this is owing to the occupied d(z2) orbital in Co(II). Altogether, these results give some clues about how nature has chosen the tetrapyrrole rings and their central metal ion.     

Polygonum viviparum L. inhibits the lipopolysaccharide‐induced inflammatory response in RAW264.7 macrophages through haem oxygenase‐1 induction and activation of the Nrf2 pathway

BACKGROUND: Polygonum viviparum L. (PV) is a member of the family Polygonaceae and is widely distributed in high‐elevation areas. It is used as a folk remedy to treat inflammation‐related diseases. This study was focused on the anti‐inflammatory response of PV against lipopolysaccharide (LPS)‐induced inflammation in RAW264.7 macrophages. RESULTS: Treatment with PV did not cause cytotoxicity at 0–50 µg mL^?1 in RAW264.7 macrophages, and the IC₅₀ value was 270 µg mL^?1. PV inhibited LPS‐stimulated nitric oxide (NO), prostaglandin (PG)E₂, interleukin (IL)‐1β and tumour necrosis factor (TNF)‐α release and inducible NO synthase (iNOS) and cyclooxygenase (COX)‐2 protein expression. In addition, PV suppressed the LPS‐induced p65 expression of nuclear factor (NF)‐κB, which is associated with the inhibition of IκB‐α degradation. These results suggest that, among mechanisms of the anti‐inflammatory response, PV inhibits the production of NO and these cytokines by down‐regulating iNOS and COX‐2 gene expression. Furthermore, PV can induce haem oxygenase (HO)‐1 protein expression through nuclear factor E2‐related factor 2 (Nrf2) activation. A specific inhibitor of HO‐1, zinc(II) protoporphyrin IX, inhibited the suppression of iNOS and COX‐2 expression by PV. CONCLUSION: These results suggest that PV possesses anti‐inflammatory actions in macrophages and works through a novel mechanism involving Nrf2 actions and HO‐1. Thus PV could be considered for application as a potential therapeutic approach for inflammation‐associated disorders. © 2012 Society of Chemical Industry     

Multi-haem cytochromes in Shewanella oneidensis MR-1: structures,functions and opportunities

Multi-haem cytochromes are employed by a range of microorganisms to transport electrons over distances of up to tens of nanometres. Perhaps the most spectacular utilization of these proteins is in the reduction of extracellular solid substrates, including electrodes and insoluble mineral oxides of Fe(III) and Mn(III/IV), by species of Shewanella and Geobacter. However, multi-haem cytochromes are found in numerous and phylogenetically diverse prokaryotes where they participate in electron transfer and redox catalysis that contributes to biogeochemical cycling of N, S and Fe on the global scale. These properties of multi-haem cytochromes have attracted much interest and contributed to advances in bioenergy applications and bioremediation of contaminated soils. Looking forward, there are opportunities to engage multi-haem cytochromes for biological photovoltaic cells, microbial electrosynthesis and developing bespoke molecular devices. As a consequence, it is timely to review our present understanding of these proteins and we do this here with a focus on the multitude of functionally diverse multi-haem cytochromes in Shewanella oneidensis MR-1. We draw on findings from experimental and computational approaches which ideally complement each other in the study of these systems: computational methods can interpret experimentally determined properties in terms of molecular structure to cast light on the relation between structure and function. We show how this synergy has contributed to our understanding of multi-haem cytochromes and can be expected to continue to do so for greater insight into natural processes and their informed exploitation in biotechnologies.     

Process optimisation of haemoglobin hydrolysis by complex proteases to produce haem-enriched peptides and its iron uptake property evaluation by Caco-2 cell model

In this study, haemoglobin was hydrolysed by compound protease to produce haem-enriched peptides in order to improve the bio-absorptivity of haem iron. The hydrolysis process was optimised through an orthogonal experiment. The molecular weight, solubility and absorptivity of haem-enriched peptides were also investigated. The optimal hydrolysis process was papain/alkaline protease ratio 6:4, enzyme content 9000 U g⁻¹, hydrolysis at 50 °C and pH 8 conditions for 4 h. After hydrolysis at this condition, the haem released amount and haem/peptide ratio were 89.6% and 28.2%, respectively. The molecular weight of the haem-enriched peptides was mainly distributed in 3–14.4 kD, and the solubility increased significantly with pH increasing. Caco-2 cell absorption experiment demonstrated that the iron absorption of haem-enriched peptides was significantly higher (50.25 ng ferritin per mg protein) than haemin chloride, haemoglobin and collagen plus haemin chloride complex. This study provides a very promising way for the development of iron supplement products.     

同步电动机失步原因分析、产生的危害及保护措施

本文主要对同步电动机产生失步的几种原因进行了详细的分析,对由此而产生的多各进行了详细的论述,并针对不同的失步情况提出了不同的保护措施。     

Design and characterisation of an artificial DNA-binding cytochrome

We aim to design novel proteins that link specific biochemical binding events, such as DNA recognition, with electron transfer functionality. We want these proteins to form the basis of new molecules that can be used for templated assembly of conducting cofactors or for thermodynamically linking DNA binding with cofactor chemistry for nanodevice applications. The first examples of our new proteins recruit the DNA-binding basic helix region of the leucine zipper protein GCN4. This basic helix region was attached to the N and C termini of cytochrome b(562) (cyt b(562)) to produce new, monomeric, multifunctional polypeptides. We have fully characterised the DNA and haem-binding properties of these proteins, which is a prerequisite for future application of the new molecules. Attachment of a single basic helix of GCN4 to either the N or C terminus of the cytochrome does not result in specific DNA binding but the presence of DNA-binding domains at both termini converts the cytochrome into a specific DNA-binding protein. Upon binding haem, this chimeric protein attains the spectral characteristics of wild-type cyt b(562). The three forms of the protein, apo, oxidised holo and reduced holo, all bind the designed (ATGAcgATGA) target DNA sequence with a dissociation constant, K(D), of approximately 90 nM. The protein has a lower affinity (K(D) ca. 370 nM) for the wild-type GCN4 recognition sequence (ATGAcTCAT). The presence of only half the consensus DNA sequence (ATGAcgGGCC) shifts the K(D) value to more than 2500 nM and the chimera does not bind specifically to DNA sequences with no target recognition sites. Ultracentrifugation revealed that the holoprotein-DNA complex is formed with a 1:1 stoichiometry, which indicates that a higher-order protein aggregate is not responsible for DNA binding. Mutagenesis of a loop linking helices 2 and 3 of the cytochrome results in a chimera with a haem-dependent DNA binding affinity. This is the first demonstration that binding of a haem group to a designed monomeric protein can allosterically modulate the DNA binding affinity.