Cytochrome flow injection analysis

With a method in hand for routinely constructing cytochrome c oxidase modified electrodes that exhibited direct electron transfer between the electrode and the oxidase, amperometry was used to detect reduced cytochrome c in solution at the oxidase-modified electrodes in a flow injection analysis format [69]. The dialysis cell was equipped with a wall jet inlet to direct cytochrome c solution past the oxidase-modified electrodes. Figure 12 shows the current response for three sequential reduced cytochrome c injections. Control experiments conducted at bilayer modified electrodes containing no oxidase showed current responses that are about 2% of those shown in Figure 12. This response may be due to changes in electrode capacitance and/or cytochrome c reacting at bilayer defect sites on the electrode. QCM measurements showed that no cytochrome c incorporated into the bilayer. However, cytochrome c was electrostatically held at the surface of the bilayer membrane at lower ionic strength [69]. 

Figure 12 Flow injection analysis of reduced cytochrome c reacting at a cytochrome c oxidase modified electrode. Three sequential 83 jiL injections of 100 xM reduced cytochrome c. Applied potential is 472 mV vs NHE, solution contains 0.1 M phosphate buffer, pH 7.4, and the electrode area is 0.2 cm. ...

Identical injections containing approximately 860 finol of the cytochrome c digest were analyzed and the results are shown in Figure 4. The high sensitivity of the analysis is evident from the excellent response observed with this subnanomole analysis with all combinations of detector and flow cell. Comparison... 

Figures. High sensitivity analysis of Cytochrome c tryptic digests. Samples were analyzed on the 2mm ID column using a 5 1 flow cell. Amounts injected were (A) 860 fmol, (B) 172 finol and (C) a derivatization blank with reagent amount equivalent to the analysis in (A) and five times the reagent injected in (B). Major peaks present in the blank are labeled with an asterisk ( ).

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