Spectral characterisation of red pigment in Italian-type dry-cured ham. Increasing lipophilicity druing processing and maturation

Spectroscopic studies of Parma ham during processing revealed a gradual transformation of muscle myoglobin, initiated by salting and continuing during ageing. Electron spin resonance spectra did, however, conclusively show that the pigment in dry-cured Parma ham at no stage is a nitrosyl complex of ferrous myoglobin as found in brine-cured ham and Spanish Serrano hams. Both near-infra red reflectance spectra of sliced ham and UV/visible absorption spectra of extract of hams, obtained with aqueous buffer or acetone, showed the presence of different red pigments at varying processing stages for both solvents. Especially, the pigment extracted with aqueous buffer exhibited unique spectral features different from those of well-known myoglobin derivatives. At the end of processing, the pigment(s) becomes less water extractable, while the fraction of red pigment(s) extractable with acetone/water (75%/25%) increases throughout the processing time up to full maturation at 18 months. The chemical identity of the 6th ligand of myoglobin could not be conclusively established, but possible candidates are discussed. The partition of the pigment(s) between pentane and acetone/water showed a strong preference for pentane, suggesting that only the heme moiety is present in the acetone/water extract, and that Parma ham pigment is gradually transformed from a myoglobin derivative into a non-protein heme complex, which was found to be thermally stable in acetone/water solutio

Oxidation of Native and Modified Hemoglobin and Myoglobin by Sodium Nitrate. Effect of Inositol Hexaphosphate

Native and modified hemoglobin, myoglobin and a and phemoglobin subunits were oxidized by sodium nitrite at pH 6. The experiments were carried out under oxy and deoxy conditions with and without inositol hexaphosphate (IHP). It is shown (a) that under oxy condition low concentration of IHP inhibits the oxidation of native hemoglobin only. However, high concentration of IHP inhibits the oxidation of both myoglobin and modified hemoglobin (digested or 0-93-SH groups blocked). This inhibition is partially counteracted by high oxygen pressure, (b) Under deoxy condition the oxidation rates of all hemeproteins studied are significantly faster than that of native hemoglobin. IHP inhibits the oxidation of all except the myoglobin and hemoglobin subunits. It is concluded that although the IHP inhibitory effect on hemoglobin oxidation by nitrite can be explained by the shift of the R↔T conformational equilibrium towards T conformation, some other structural changes such as alteration in molecular surface charges must occur to account for the effect of IHP on the oxidation of hemeproteins devoid of heme-heme interaction

An investigation into the feasibility of myoglobin-based single-electron transistors

Myoglobin single-electron transistors were investigated using nanometer- gap platinum electrodes fabricated by electromigration at cryogenic temperatures. Apomyoglobin (myoglobin without heme group) was used as a reference. The results suggest single electron transport is mediated by resonant tunneling with the electronic and vibrational levels of the heme group in a single protein. They also represent a proof-of-principle that proteins with redox centers across nanometer-gap electrodes can be utilized to fabricate single-electron transistors. The protein orientation and conformation may significantly affect the conductance of these devices. Future improvements in device reproducibility and yield will require control of these factors

Development of methods for capillary isoelectric focusing of dairy proteins : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Chemistry at Massey University

Capillary Isoelectric Focusing (CIEF) is a high-resolution technique which can be applied to the separation and characterisation of complex biological mixtures such as dairy proteins. Although dairy proteins are commonly analysed by traditional gel electrophoresis techniques including 2-Dimensional PAGE, CIEF offers the advantages of reduced analysis times, the ability to handle smaller sample volumes and increased sensitivity with improved separation efficiencies. Several methods for capillary isoelectric focusing of dairy proteins have been developed herein. For the analysis of soluble whey proteins methods that can be used with either UV or mass spectrometry (MS) detection have been set up. For MS detection a coaxial sheath flow interface in conjunction with electrospray ionisation has been utilised. For analysis of the inherently insoluble casein proteins with UV detection denaturing and reducing agents have been introduced into the system. Results have shown very close similarities to those obtained by IEF gels

Catalytic Reduction of Bisulfite by Myoglobin/Surfactant Films

The voltammetry of bisulfite at a film formed with myoglobin was studied in aqueous solutions. A broad wave was observed for the reduction of bisulfite. Using controlled potential electrolysis, the reduction at potentials positive of the FeII/FeIwave formed dithionite exclusively. As the potential approached the region for the FeII/FeI reduction, bisulfite was reduced primarily to HS−. Even at the negative potentials, some dithionite was still formed, which could then be electrochemically reduced to thiosulfate. Analysis of the formation of HS−, dithionite and thiosulfate during the electrolysis was consistent with the parallel formation of HS−and dithionite, the latter of which was reduced to thiosulfate. Thiosulfate was verified by chemical analysis of the products from controlled potential electrolysis of the solution, and dithionite was observed spectroscopically using spectroelectro−chemistry

2H and 13C NMR studies on the temperature-dependent water and protein dynamics in hydrated elastin, myoglobin and collagen

2H NMR spin-lattice relaxation and line-shape analyses are performed to study the temperature-dependent dynamics of water in the hydration shells of myoglobin, elastin, and collagen

Top-Down Mass Analysis of Protein Tyrosine Nitration: Comparison of Electron Capture Dissociation with “Slow-Heating” Tandem Mass Spectrometry Methods

Tyrosine nitration in proteins is an important post-translational modification (PTM) linked to various pathological conditions. When multiple potential sites of nitration exist, tandem mass spectrometry (MS/MS) methods provide unique tools to locate the nitro-tyrosine(s) precisely. Electron capture dissociation (ECD) is a powerful MS/MS method, different in its mechanisms to the “slow-heating” threshold fragmentation methods, such as collision-induced dissociation (CID) and infrared multiphoton dissociation (IRMPD). Generally, ECD provides more homogeneous cleavage of the protein backbone and preserves labile PTMs. However recent studies in our laboratory demonstrated that ECD of doubly charged nitrated peptides is inhibited by the large electron affinity of the nitro group, while CID efficiency remains unaffected by nitration. Here, we have investigated the efficiency of ECD versus CID and IRMPD for top-down MS/MS analysis of multiply charged intact nitrated protein ions of myoglobin, lysozyme, and cytochrome c in a commercial Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer. CID and IRMPD produced more cleavages in the vicinity of the sites of nitration than ECD. However the total number of ECD fragments was greater than those from CID or IRMPD, and many ECD fragments contained the site(s) of nitration. We conclude that ECD can be used in the top-down analysis of nitrated proteins, but precise localization of the sites of nitration may require either of the “slow-heating” methods

Cooking patties from a frozen state, endpoint temperature, and post-cookery chilling affect internal and external color and cooking losses in ground beef patties

To determine the effects of cooking state (frozen vs. thawed), endpoint temperature (65.5 vs. 73.9°C), and post-cookery chilling on color of ground beef patties, 85% coarse-ground beef was purchased and ground through a 9.5-mm plate, formed into 115-g patties, and crust frozen before 4 patties were vacuum packaged and stored at -10 °C. Packages were either thawed in a water bath for 2 h prior to cooking or cooked directly from frozen. Within each package, patties were weighed before being cooked to their assigned temperature (65.5 or 73.9oC) and either allowed to cool at room temperature on paper plates or placed in a plastic baggie and submerged in an ice water bath. Patty temperature was monitored at 0, 1, 5, 10, 15, and 30 min post-cooking, and patties were reweighed to calculate cook loss percentage before external and internal instrumental color (L*, a*,and b*) was measured on each patty. Patties cooked from frozen, to 73.9°C, or cooled at room temperature had greater (P \u3c 0.05) cooking losses than those cooked from a thawed state, to 65.5°C, or cooled in an ice bath, respectively. External color of patties cooked from a thawed state was lighter (greater L*; P \u3c 0.05), redder (greater a*; P \u3c 0.05), and more yellow (greater b*; P \u3c 0.05) than those cooked from frozen. Moreover, L*, a*, and b* values were greater (P \u3c 0.05) for the surface of patties cooked to 65.5 than 73.9°C, whereas L*, a*, and b* values were greater (P \u3c 0.05) externally for patties cooled in an ice bath than those cooled at room temperature. Internally, patties cooked from frozen, cooked to 65.5°C, or cooled in an ice bath were lighter (P \u3c 0.05) than those cooked from a thawed state, cooked to 73.9°C, or cooled at room temperature, respectively. Patties cooked to 65.5°C from a thawed state had the greatest (P \u3c 0.05) internal a* and b* values, whereas frozen patties cooked to 73.9°C had the least red and yellow (P \u3c 0.05) internal color. Moreover, thawed patties cooked and chilled in an ice bath were redder (P \u3c 0.05) internally than other cooking state × cooling method combinations. It was expected that cooking to 65.5°C would result in redder internal cooked color, but persistent redness was also observed when patties were cooked from a thawed, rather than frozen, state and when cooled in an ice bath