Application of Electron Paramagnetic Resonance (EPR) Oximetry to Monitor Oxygen in Wounds in Diabetic Models

A lack of oxygen is classically described as a major cause of impaired wound healing in diabetic patients. Even if the role of oxygen in the wound healing process is well recognized, measurement of oxygen levels in a wound remains challenging. The purpose of the present study was to assess the value of electron paramagnetic resonance (EPR) oximetry to monitor pO₂ in wounds during the healing process in diabetic mouse models. Kinetics of wound closure were carried out in streptozotocin (STZ)-treated and db/db mice. The pO₂ was followed repeatedly during the healing process by 1 GHz EPR spectroscopy with lithium phthalocyanine (LiPc) crystals used as oxygen sensor in two different wound models: a full-thickness excisional skin wound and a pedicled skin flap. Wound closure kinetics were dramatically slower in 12-week-old db/db compared to control (db/+) mice, whereas kinetics were not statistically different in STZ-treated compared to control mice. At the center of excisional wounds, measurements were highly influenced by atmospheric oxygen early in the healing process. In pedicled flaps, hypoxia was observed early after wounding. While reoxygenation occurred over time in db/+ mice, hypoxia was prolonged in the diabetic db/db model. This observation was consistent with impaired healing and microangiopathies observed using intravital microscopy. In conclusion, EPR oximetry using LiPc crystals as the oxygen sensor is an appropriate technique to follow wound oxygenation in acute and chronic wounds, in normal and diabetic animals. Nevertheless, the technique is limited for measurements in pedicled skin flaps and cannot be applied to excisional wounds in which diffusion of atmospheric oxygen significantly affects the measurements.     

A comparative evaluation of EPR and OxyLite oximetry using a random sampling of pO(2) in a murine tumor

Methods currently available for the measurement of oxygen concentrations (oximetry) in viable tissues differ widely from each other in their methodological basis and applicability. The goal of this study was to compare two novel methods, particulate-based electron paramagnetic resonance (EPR) and OxyLite oximetry, in an experimental tumor model. EPR oximetry uses implantable paramagnetic particulates, whereas OxyLite uses fluorescent probes affixed on a fiber-optic cable. C3H mice were transplanted with radiation-induced fibrosarcoma (RIF-1) tumors in their hind limbs. Lithium phthalocyanine (LiPc) microcrystals were used as EPR probes. The pO(2) measurements were taken from random locations at a depth of approximately 3 mm within the tumor either immediately or 48 h after implantation of LiPc. Both methods revealed significant hypoxia in the tumor. However, there were striking differences between the EPR and OxyLite readings. The differences were attributed to the volume of tissue under examination and the effect of needle invasion at the site of measurement. This study recognizes the unique benefits of EPR oximetry in terms of robustness, repeatability and minimal invasiveness.     

The effects of Efaproxyn (efaproxiral) on subcutaneous RIF-1 tumor oxygenation and enhancement of radiotherapy-mediated inhibition of tumor growth in mice

Efaproxiral, an allosteric modifier of hemoglobin, reduces hemoglobin-oxygen binding affinity, facilitating oxygen release from hemoglobin, which is likely to increase tissue pO(2). The purpose of this study was to determine the effect of efaproxiral on tumor oxygenation and growth inhibition of RIF-1 tumors that received X radiation (4 Gy) plus oxygen breathing compared to radiation plus oxygen plus efaproxiral daily for 5 days. Two lithium phthalocyanine (LiPc) deposits were implanted in RIF-1 tumors in C3H mice for tumor pO(2) measurements using EPR oximetry. Efaproxiral significantly increased tumor oxygenation by 8.4 to 43.4 mmHg within 5 days, with maximum increases at 22-31 min after treatment. Oxygen breathing alone did not affect tumor pO(2). Radiation plus oxygen plus efaproxiral produced tumor growth inhibition throughout the treatment duration, and inhibition was significantly different from radiation plus oxygen from day 3 to day 5. The results of this study provide unambiguous quantitative information on the effectiveness of efaproxiral to consistently and reproducibly increase tumor oxygenation over the course of 5 days of treatment, modeling the clinical use of efaproxiral. Also, based on the tumor growth inhibition, the study shows the efaproxiral-enhanced tumor oxygenation was radiobiologically significant. This is the first study to demonstrate the ability of efaproxiral to increase tumor oxygenation and to increase the tumor growth inhibition of radiotherapy over 5 days of treatment.     

A naphthalocyanine-based EPR probe for localized measurements of tissue oxygenation.

A new electron paramagnetic resonance (EPR) oximetry probe, based on a naphthalocyanine macrocycle, is reported to exhibit high oxygen sensitivity and favorable EPR characteristics for biological applications. The free radical probe, lithium naphthalocyanine (LiNc), is synthesized as fine microcrystalline powder with particle size less than 1 microm and high spin density. It exhibits a single sharp EPR peak, whose width varies linearly with oxygen partial pressure (pO2). The EPR spectrum is nonsaturable at typical microwave power levels (< 25 mW at X-band). These unique characteristics make this probe ideal for measuring oxygen concentration in biological tissues, in vivo. The peak-to-peak width under anoxic conditions is 0.51 G (at X-band), and it increases linearly with increase in oxygen partial pressure and reaches 26.0 G for 100% oxygen (760 mmHg), showing an oxygen sensitivity of 34 mG/mmHg. The probe responds to changes in pO2 quickly and reproducibly, thus enabling dynamic measurements of regional oxygenation in real time. The application of this probe for oximetry is demonstrated in an in vivo biological system. The changes in pO2 were monitored in the leg muscle tissue of a living mouse breathing room air and carbogen (95% oxygen + 5% CO2), alternatively. The mean pO2 measured with this probe in muscle tissues was consistent with values reported previously using other methods. Overall, the probe shows very desirable characteristics for localized measurements of tissue oxygenation.     

Injectable LiNc-BuO loaded microspheres as in vivo EPR oxygen sensors after co-implantation with tumor cells

Electron paramagnetic resonance (EPR) oximetry is a technique which allows accurate and repeatable oxygen measurements. We encapsulated a highly oxygen sensitive particulate EPR spin probe into microparticles to improve its dispersibility and, hence, facilitate the administration. These biocompatible, non-toxic microspheres contained 5–10 % (w/w) spin probe and had an oxygen sensitivity of 0.60±0.01 µT/mmHg. To evaluate the performance of the microparticles as oxygen sensors, they were co-implanted with syngeneic tumor cells in 2 different rat strains. Thus, tissue injury was avoided and the microparticles were distributed all over the tumor tissue. Dynamic changes of the intratumoral oxygen partial pressure during inhalation of 8 %, 21 %, or 100 % oxygen were monitored in vivo by EPR spectroscopy and quantified. Values were verified in vivo by invasive fluorometric measurements using Oxylite probes and ex vivo by pimonidazole adduct accumulation. There were no hints that the tumor physiology or tissue oxygenation had been altered by the microparticles. Hence, these microprobes offer great potential as oxygen sensors in preclinical research, not only for EPR spectroscopy but also for EPR imaging. For instance, the assessment of tissue oxygenation during therapeutic interventions might help understanding pathophysiological processes and lead to an individualized treatment planning or the use of formulations with hypoxia triggered release of active agents.     

Comparison of methods for measuring oxygen consumption in tumor cells in vitro

The oxygen consumption rate of tumor cells affects tumor oxygenation and response to therapies. Highly sensitive methods for determining cellular oxygen consumption are, therefore, needed to identify treatments that can modulate this parameter. We compared the performances of three different methods for measuring cellular oxygen consumption: electron paramagnetic resonance (EPR) oximetry, the Clark electrode, and the MitoXpress fluorescent assay. To compare the assays, we used K562 cells in the presence of rotenone and hydrocortisone, compounds that are known to inhibit the mitochondrial electron transport chain to different extents. The EPR method was the only one that could identify both rotenone and hydrocortisone as inhibitors of tumor cell oxygen consumption. The Clark electrode and the fluorescence assay demonstrated a significant decrease in cellular oxygen consumption after administration of the most potent inhibitor (rotenone) but failed to show any significant effect of hydrocortisone. EPR oximetry is, therefore, the most sensitive method for identifying inhibitors of oxygen consumption on cell assays, whereas the Clark electrode offers the unique opportunity to add external compounds during experiments and still shows great sensitivity in studying enzyme and chemical reactions that consume oxygen (non-cell assays). Finally, the MitoXpress fluorescent assay has the advantage of a high-sample throughput and low bulk requirements but at the cost of a lower sensitivity.     

Measurement of time-resolved oxygen concentration changes in photosynthetic systems by nitroxide-based EPR oximetry

The application of recent developments of EPR oximetry to photosynthetic systems is described and used to study rapid processes in isolated thylakoid membranes from spinach and in intact photoautotrophic soybean cells. Using the peak heights of 15N perdeuterated Tempone and two microwave power levels oxygen evolution and consumption were measured. The method measured time-resolved oxygen concentration changes in the micromolar range. Oxygen evolution was linearly proportionate to the chlorophyl concentration of thylakoid membrane over the range studied (0-2 mg/ml). Oxygen evolution associated with single turnover light pulses was consistent with the four state model. The time (t1/2) to reach equilibrium of oxygen concentrations after a single turnover pulse was 0.4-0.5 ms, indicating that the evolution of oxygen coupled to the S4-S0 transition may be shorter than reported previously. The time for equilibrium of oxygen after single turnover pulses in soybean cells was relatively long (400 ms), which suggests that there are significant barriers to the free diffusion of oxygen in this system. The method also was used to study oxygen consumption by the electron transport chain of photosystem I and photosystem II. We conclude that EPR oximetry can provide quantitative and time-resolved data on oxygen concentrations with a sensitivity that is useful for studies of such systems.     

Novel particulate spin probe for targeted determination of oxygen in cells and tissues

The synthesis and characterization of a new lithium octa-n-butoxy-substituted naphthalocyanine radical probe (LiNc-BuO) and its use in the determination of concentration of oxygen (oximetry) by electron paramagnetic resonance (EPR) spectroscopy are reported. The probe is synthesized as a needle-shaped microcrystalline particulate. The particulate shows a single-line EPR spectrum that is highly exchange-narrowed with a line-width of 210 mG. The EPR line-width is sensitive to molecular oxygen showing a linear relationship between the line-width and concentration of oxygen (pO(2)) with a sensitivity of 8.5 mG/mmHg. We studied a variety of physicochemical and biological properties of LiNc-BuO particulates to evaluate the suitability of the probe for in vivo oximetry. The probe is unaffected by biological oxidoreductants, stable in tissues for several months, and can be successfully internalized in cells. We used this probe to monitor changes in concentration of oxygen in the normal muscle and RIF-1 tumor tissue of mice as a function of tumor growth. The data showed a rapid decrease in the tumor pO(2) with increase of tumor volume. Human arterial smooth muscle cells, upon internalization of the LiNc-BuO probe, showed a marked oxygen gradient across the cell membrane. In summary, the newly synthesized octa-n-butoxy derivative of lithium naphthalocyanine has unique properties that are useful for determining oxygen concentration in chemical and biological systems by EPR spectroscopy and also for magnetic tagging of cells.     

In vivo electron paramagnetic resonance oximetry with particulate materials

Electron paramagnetic resonance (EPR) methods can be used to study tissue pO(2) (PtO(2)) in anesthetized or awake animals (EPR oximetry). The method takes advantage of the fact that some paramagnetic materials have an EPR linewidth that is sensitive to the pO(2) in which the material is located. This article provides an overview of the method of EPR oximetry using implanted particulate materials as the sensors of pO(2). Characteristics of these materials are described to help the reader understand the factors involved in choosing the optimum particulate material. Examples of biological studies are included that show how EPR oximetry may be used on both awake and anesthetized animals.     

Microfabricated teflon membranes for low-noise recordings of ion channels in planar lipid bilayers

We present a straightforward, accessible method for the fabrication of micropores with diameters from 2 to 800 micro m in films of amorphous Teflon (Teflon AF). Pores with diameters 相似文献   

Electron paramagnetic resonance highlights that the oxygen effect contributes to the radiosensitizing effect of paclitaxel

Background

Paclitaxel (PTX) is a potent anti-cancer chemotherapeutic agent and is widely used in the treatments of solid tumors, particularly of the breast and ovaries. An effective and safe micellar formulation of PTX was used to administer higher doses of PTX than Taxol® (the current commercialized drug). We hypothesize that PTX-loaded micelles (M-PTX) may enhance tumor radiosensitivity by increasing the tumor oxygenation (pO₂). Our goals were (i) to evaluate the contribution of the “oxygen effect” to the radiosensitizing effect of PTX; (ii) to demonstrate the therapeutic relevance of the combination of M-PTX and irradiation and (iii) to investigate the underlying mechanisms of the observed oxygen effect.

Methodology and Principal Findings

We used (PEG-p-(CL-co-TMC)) polymeric micelles to solubilize PTX. pO₂ was measured on TLT tumor-bearing mice treated with M-PTX (80 mg/kg) using electron paramagnetic resonance (EPR) oximetry. The regrowth delay following 10 Gy irradiation 24 h after M-PTX treatment was measured. The tumor perfusion was assessed by the patent blue staining. The oxygen consumption rate and the apoptosis were evaluated by EPR oximetry and the TUNEL assay, respectively. EPR oximetry experiments showed that M-PTX dramatically increases the pO₂ 24 h post treatment. Regrowth delay assays demonstrated a synergy between M-PTX and irradiation. M-PTX increased the tumor blood flow while cells treated with M-PTX consumed less oxygen and presented more apoptosis.

Conclusions

M-PTX improved the tumor oxygenation which leads to synergy between this treatment and irradiation. This increased pO₂ can be explained both by an increased blood flow and an inhibition of O₂ consumption.     

Regional transcranial oximetry with near infrared spectroscopy (NIRS) in comparison with measuring oxygen saturation in the jugular bulb in infants and children for monitoring cerebral oxygenation]

Using a dual channel near infrared (NIR) in vivo optical spectroscopy (INVOS) system (INVOS 3100A, Somanetics Corp. Troy, MI, USA) we investigated the relationship between jugular venous oxygen saturation (SjvO2) and regional cerebral oxygen saturation (rSO2) in 30 infants and children (mean age 4.5 years) with congenital heart disease undergoing cardiac catheterisation. The NIRS-SomaSensor (emitter and dual receiver probe) was applied at a standardised right fronto-temporal location (over the right frontal cortex) on the infant's head and covered with an adhesive flexible bandage. Using NIR light (730 and 810 nm) and two source-detector spacings (3 and 4 cm from the transmitter), percentage values of rSO2 were calculated from detected haemoglobin saturations. Simultaneously, jugular venous oxygen saturation (SjvO2) monitoring was performed via a catheter placed in the right internal jugular vein with its tip positioned in the jugular bulb, as verified by fluoroscopy. To compare the reliability of NIRS measurement characteristics, jugular venous blood was analysed for SjvO2 as a reference measure of global cerebral oxygenation, by co-oximetry (OSM3-Hemoximeter, Radiometer Copenhagen, Denmark). Other measured variables included pulse oximetry, arterial blood pressure, and venous and arterial oxygen saturations. Over a jugular venous oxygen saturation range of 31-83%, a significant positive linear correlation was found between rSO2 (NIRS measurement) and SjvO2 (jugular bulb oximetry) (r = 0.93, p < 0.001). No significant correlation was observed between rSO2 values and arterial blood saturation or pulse oximetry. The quantitative correlation between rSO2 (haemoglobin oxygenation in a small hemi-elliptical area of the brain) and reference SjvO2 measurement (method for monitoring global cerebral oxygenation) suggests that NIRS measurement with subtraction algorithm should identify predominantly intracranial saturation in the pediatric age group, and will tend to reflect global oxygenation under physiological conditions. Transcranial oximetry using dual receiving channel NIRS offers a noninvasive, real-time, reliable and practicable means of monitoring cerebral haemoglobin oxygenation changes infants and children with cyanotic and noncyanotic congenital heart disease.     

Tumor PO(2) changes during photodynamic therapy depend upon photosensitizer type and time after injection

In this study, the vascular and tissue oxygen changes induced by photodynamic therapy in the RIF-1 tumor were examined, using electron paramagnetic resonance (EPR) oximetry. Two photosensitizers, including verteporfin (BPD-MA in a lipid-based formulation) and aminolevulinic acid-induced protoporphyrin IX (ALA-PPIX), were investigated with optical irradiation, sufficient to induce sub-curative damage in the tumor tissue, and the transient changes in PO(2) and vascular perfusion were examined. A large increase in tissue oxygenation (from 3 up to 9.5 mmHg) was observed when treated with ALA-PPIX based photodynamic therapy, which lasted during the treatment and a small residual increase that returned back to baseline levels by 48 h after treatment. With verteporfin-based photodynamic therapy, one group of animals was irradiated 15 min after injection and exhibited a small decrease in oxygenation relative to pre-irradiation levels. The second group was irradiated at 3 h after injection and exhibited a large increase in the average PO(2), (from 3 to 15 mmHg) by the end of the treatment. These observations indicate that photodynamic therapy significantly increases tissue PO(2) under certain treatment conditions, with the potential cause being either increased local blood flow or decreased local oxygen metabolic consumption due to cellular damage.     

Closed loop control of oxgenation and ventilation

Closed loop control of oxygenation and ventilation during mechanical ventilatory support is essential for remote medical care in an austere environment. Closed loop control allows for expert systems to provide the current standard of care in the absence of on-site expertise. Ventilation may be controlled by simple systems incorporating patient height or by advanced systems incorporating measurements of end-tidal carbon dioxide (ETCO2) and pulmonary impedance. Oxygenation may be controlled by adjustments of inspired oxygen concentrations (FIO2) and positive end-expiratory pressure (PEEP) using pulse oximetry (SpO2) as the input. Control of oxygenation can prevent hypoxemia and has the potential to reduce oxygen requirements. A double closed loop system of oxygenation control including control of FIO2 via SpO2 and control of oxygen generation by a portable oxygen generator (POG) based on FIO2 and minute ventilation (VE) promises safety and efficiency. Remote control of ventilation and oxygenation is possible using existing technology.     

Impact of myo‐inositol trispyrophosphate (ITPP) on tumour oxygenation and response to irradiation in rodent tumour models

Tumour hypoxia is a well‐established factor of resistance in radiation therapy (RT). Myo‐inositol trispyrophosphate (ITPP) is an allosteric effector that reduces the oxygen‐binding affinity of haemoglobin and facilitates the release of oxygen by red blood cells. We investigated herein the oxygenation effect of ITPP in six tumour models and its radiosensitizing effect in two of these models. The evolution of tumour pO₂ upon ITPP administration was monitored on six models using 1.2 GHz Electron Paramagnetic Resonance (EPR) oximetry. The effect of ITPP on tumour perfusion was assessed by Hoechst staining and the oxygen consumption rate (OCR) in vitro was measured using 9.5 GHz EPR. The therapeutic effect of ITPP with and without RT was evaluated on rhabdomyosarcoma and 9L‐glioma rat models. ITPP enhanced tumour oxygenation in six models. The administration of 2 g/kg ITPP once daily for 2 days led to a tumour reoxygenation for at least 4 days. ITPP reduced the OCR in six cell lines but had no effect on tumour perfusion when tested on 9L‐gliomas. ITPP plus RT did not improve the outcome in rhabdomyosarcomas. In 9L‐gliomas, some of tumours receiving the combined treatment were cured while other tumours did not benefit from the treatment. ITPP increased oxygenation in six tumour models. A decrease in OCR could contribute to the decrease in tumour hypoxia. The association of RT with ITPP was beneficial for a few 9L‐gliomas but was absent in the rhabdomyosarcomas.     

Electron paramagnetic resonance oximetry as a quantitative method to measure cellular respiration: a consideration of oxygen diffusion interference

Electron paramagnetic resonance (EPR) oximetry is being widely used to measure the oxygen consumption of cells, mitochondria, and submitochondrial particles. However, further improvement of this technique, in terms of data analysis, is required to use it as a quantitative tool. Here, we present a new approach for quantitative analysis of cellular respiration using EPR oximetry. The course of oxygen consumption by cells in suspension has been observed to have three distinct zones: pO(2)-independent respiration at higher pO(2) ranges, pO(2)-dependent respiration at low pO(2) ranges, and a static equilibrium with no change in pO(2) at very low pO(2) values. The approach here enables one to comprehensively analyze all of the three zones together-where the progression of O(2) diffusion zones around each cell, their overlap within time, and their potential impact on the measured pO(2) data are considered. The obtained results agree with previously established methods such as high-resolution respirometry measurements. Additionally, it is also demonstrated how the diffusion limitations can depend on cell density and consumption rate. In conclusion, the new approach establishes a more accurate and meaningful model to evaluate the EPR oximetry data on cellular respiration to quantify related parameters using EPR oximetry.     

Endotoxin-induced liver hypoxia: defective oxygen delivery versus oxygen consumption.

In vivo EPR was used to investigate liver oxygenation in a hemodynamic model of septic shock in mice. Oxygen-sensitive material was introduced either (i) as a slurry of fine particles which localized at the liver sinusoids (pO2 = 44.39 +/- 5.13 mmHg) or (ii) as larger particles implanted directly into liver tissue to measure average pO2 across the lobule (pO2 = 4.56 +/- 1.28 mmHg). Endotoxin caused decreases in pO2 at both sites early (5-15 min) and at late time points (6 h after endotoxin; sinusoid = 11.22 +/- 2.48 mmHg; lobule = 1.16 +/- 0.42 mmHg). The overall pO2 changes observed were similar (74.56% versus 74.72%, respectively). Blood pressures decreased transiently between 5 and 15 min (12.88 +/- 8% decrease) and severely at 6 h (59 +/- 9% decrease) following endotoxin, despite volume replacement with saline. Liver and circulatory nitric oxide was elevated at these times. Liver oxygen extraction decreased from 44% in controls to only 15% following endotoxin, despite severe liver hypoxia. Arterial oxygen saturation, blood flow (hepatic artery), and cardiac output were unaffected. Pretreatment with l-NMMA failed to improve endotoxin-induced oxygen defects at either site, whereas interleukin-13 preserved oxygenation. These site-specific measurements of pO2 provide in vivo evidence that the principal cause of liver hypoxia during hypodynamic sepsis is reduced oxygen supply to the sinusoid and can be alleviated by maintaining sinusoidal perfusion.     

Does cerebral oxygen delivery limit incremental exercise performance?

Previous studies have suggested that a reduction in cerebral oxygen delivery may limit motor drive, particularly in hypoxic conditions, where oxygen transport is impaired. We hypothesized that raising end-tidal Pco(2) (Pet(CO(2))) during incremental exercise would increase cerebral blood flow (CBF) and oxygen delivery, thereby improving peak power output (W(peak)). Amateur cyclists performed two ramped exercise tests (25 W/min) in a counterbalanced order to compare the normal, poikilocapnic response against a clamped condition, in which Pet(CO(2)) was held at 50 Torr throughout exercise. Tests were performed in normoxia (barometric pressure = 630 mmHg, 1,650 m) and hypoxia (barometric pressure = 425 mmHg, 4,875 m) in a hypobaric chamber. An additional trial in hypoxia investigated effects of clamping at a lower Pet(CO(2)) (40 Torr) from ～75 to 100% W(peak) to reduce potential influences of respiratory acidosis and muscle fatigue imposed by clamping Pet(CO(2)) at 50 Torr. Metabolic gases, ventilation, middle cerebral artery CBF velocity (transcranial Doppler), forehead pulse oximetry, and cerebral (prefrontal) and muscle (vastus lateralis) hemoglobin oxygenation (near infrared spectroscopy) were monitored across trials. Clamping Pet(CO(2)) at 50 Torr in both normoxia (n = 9) and hypoxia (n = 11) elevated CBF velocity (～40%) and improved cerebral hemoglobin oxygenation (～15%), but decreased W(peak) (6%) and peak oxygen consumption (11%). Clamping at 40 Torr near maximal effort in hypoxia (n = 6) also improved cerebral oxygenation (～15%), but again limited W(peak) (5%). These findings demonstrate that increasing mass cerebral oxygen delivery via CO(2)-mediated vasodilation does not improve incremental exercise performance, at least when accompanied by respiratory acidosis.     

Synthesis and characterization of a perchlorotriphenylmethyl (trityl) triester radical: a potential sensor for superoxide and oxygen in biological systems

Synthesis and characterization of an inert perchlorotriphenylmethyl triester radical, PTM-TE, are reported. PTM-TE was prepared by a facile 3-step synthesis using Friedel-Crafts reaction of tetrachlorobenzene with chloroform followed by ethoxycarbonylation and subsequent oxidation. PTM-TE is paramagnetic and exhibits a single sharp EPR spectrum. In solution, the EPR linewidth of PTM-TE is highly sensitive to the dissolved oxygen content, thus enabling accurate measurement of oxygen concentration (oximetry). In addition, the radical also shows high reactivity towards superoxide. The ester radical has the potential for use as a high-sensitive probe for determination of oxygen concentration and superoxide in biological systems.     

A highly sensitive biocompatible spin probe for imaging of oxygen concentration in tissues

The development of an injectable probe formulation, consisting of perchlorotriphenylmethyl triester radical dissolved in hexafluorobenzene, for in vivo oximetry and imaging of oxygen concentration in tissues using electron paramagnetic resonance (EPR) imaging is reported. The probe was evaluated for its oxygen sensitivity, biostability, and distribution in a radiation-induced fibrosarcoma tumor transplanted into C3H mice. Some of the favorable features of the probe are: a single narrow EPR peak (anoxic linewidth, 41 microT), high solubility in hexafluorobenzene (>12 mM), large linewidth sensitivity to molecular oxygen ( approximately 1.8 microT/mmHg), good stability in tumor tissue (half-life: 3.3 h), absence of spin-spin broadening (up to 12 mM), and lack of power saturation effects (up to 200 mW). Three-dimensional spatial and spectral-spatial (spectroscopic) EPR imaging measurements were used to visualize the distribution of the probe, as well as to obtain spatially resolved pO(2) information in the mice tumor subjected to normoxic and hyperoxic treatments. The new probe should enable unique opportunities for measurement of the oxygen concentration in tumors using EPR methods.