The measurement of redox potential, or redox for short, is relatively simple in concept, requiring only a reference electrode and working electrode through which the balance of oxidants and reductants in any media can be measured via voltage (mV). However, historically there has been no reliable method to measure it due to biofouling (blockage of the electrode surface by proteins in biologic media), and consequently no rapid, reliable method for evaluating redox species and redox balance in metabolic and oxidative stress. Because of the inability to provide this direct measure of oxidative stress, current standards for evaluating the severity of these derangements in conditions such as sepsis and shock include only secondary measures of oxidative and metabolic stress, such as central venous oxygen saturation (SvO2) and serum lactate measurements. These values are currently followed not only as indicators of the severity of the initial insult, but also as measures of the success of therapeutic interventions. Further, recent international guidelines define therapeutic endpoints based on the value of SvO2, with saturations less than 70% indicating an ongoing metabolic and physiologic deficit necessitating an escalation of support.
Though these measures are the current standard, they have numerous limitations as they reflect only crude, averaged measures of the metabolic and clinical state, and provide only indirect measurements for directly relevant clinical data: cardiac output, oxygen metabolism, oxygen debt and overall oxidative stress. Even recent advances in regional tissue oxygenation measures, such as Near-Infrared Spectroscopy (NIRS) monitoring, can only measure regional oxygen saturations or serve as a non-invasive measure of SvO2 that reports an oxygen saturation that may or may not be clinically relevant, but has no ability to reflect the true redox environment or oxidative stress present.
Furthermore, while there is much debate regarding antioxidant therapy in septic shock and critical illness, blood redox measurements can be useful in guiding such a therapeutic approach in the future. Pre-clinical studies have demonstrated the benefits of antioxidant therapy, such as the use of ascorbic acid in both attenuating the development of multiple organ dysfunction syndrome (MODS), and reducing the proinflammatory and procoagulant states that induce organ injury such as lung vascular injury in sepsis or after other insults such as trauma, cardiac arrest, or burns. There has been renewed interest in the role of Vitamin C, along with other antioxidants, in protecting against oxidative stress in ischemia/reperfusion injury and sepsis, including its ability to mitigate organ injury in these states. As speculated by multiple investigators, resuscitation fluids containing targeted anti-oxidant therapy may improve the ability to support patients with sepsis and protect against multiple organ dysfunction.
To accurately and reproducibly make electrochemical (redox) measurements in complex biologic fluid such as blood and plasma, the electrode must be able to exchange electrons, H+ ions, and oxygen free radicals with the redox species in solution. If the electrode surface is somehow modified or contaminated (e.g., by protein adsorption), then the electrode can lose its effectiveness as a redox sensor. The impact of biofouling on the electrochemical measurement will depend on the degree of modification and the redox species being detected. It is well known that when a conducting metal electrode is exposed to protein, adsorption will take place. To circumvent or reduce the impact of protein adsorption on the electrochemical activity of redox active molecules at the electrode surface, prior solutions have been to modify the electrode surface with, e.g., a pyridinethiol or polymer film (e.g., polyethylene glycol). Electrode modification, however, can impede electron exchange at the electrode surface and subsequently bias the results due to selective partitioning/exclusion of certain analyte species.
Another factor that becomes important, particularly with the potentiometric measurement of blood redox potential, is the reference electrode. In a potentiometric sensor, a high impedance voltmeter measures the potential of the indicator electrode with respect to the reference electrode.
The potential of the reference electrode stays constant during and between samples for the measured potential to reflect what takes place at the nanoporous electrode in solution. A common reference electrode to use is a standard commercial silver/silver chloride reference electrode (saturated potassium chloride (KCl)) with a porous Vycor frit. By fixing the concentration of potassium chloride, the potential of the reference electrode then becomes fixed. A fitted reference electrode, however, can also become clogged from the proteins/enzymes/cells in blood giving rise to a significant junction potential, and extreme cases, a sluggish, variable response. Because the contribution of the junction potential to the measured cell potential is not known, it is impossible to know the potential of the indicating electrode to any degree of accuracy.
The need for measuring redox at a direct measure of metabolic and oxidative stress is clear; however, there has been little progress in the field of redox measurements in complex biologic media. As noted above, the problem is that traditional planar (flat) metal electrodes cannot reliably or accurately make these measurements as they are fouled and rendered unresponsive in complex protein containing biologic solutions. Because of this, accurate measures for POC testing can be difficult. Furthermore, biofouling can prevent the development of a redox potential device capable of continuous or semi-continuous measurements of blood, urine, exhaled breath condensate, or tissue interstituium.