Time-Related Evidence of Intestinal Oxidative Stress in Obstructive Jaundice-Induced Rats.

INTRODUCTION: Obstructive jaundice is known to affect intestinal permeability and facilitate bacterial translocation through related mechanisms. This study was conducted to evaluate the alterations concerning blood biochemistry and levels of several markers of oxidative stress (OS) in blood and intestinal mucosa caused by obstructive jaundice and how these fluctuate over time, in order to further explore the possibility of intervening in the OS path in future experiments. METHODS: A total of 54 albino Wistar rats were randomly divided into three groups (control, sham operated, and bile duct ligation) and sacrificed at specific time intervals (12 h and 2, 7, and 14 days). The intestinal barrier function was evaluated by measuring endotoxin levels in portal, aortic, and peripheral blood. Also, basic biochemical parameters were simultaneously measured in peripheral blood. Tissue samples collected from the terminal ileum were homogenized for determining the OS markers, lipid peroxidation, and protein-free radical-induced oxidation. RESULTS: We designed this experiment to examine the alterations in enteric mucosa primarily in relation to OS in a period of 14 days. During this time period, we investigated in specific time intervals not only OS fluctuations but also other liver function parameters, as well as CRP and endotoxin levels. The alterations were monitored in relation to time after bile duct ligation. CONCLUSION: Bile duct ligation in rats causes OS versus post-ligation time progression of the common bile duct. OS was increased by ∼50% compared to control/sham and peaked at 7 days and at least up to 14 days post-ligation. This phenomenon was accompanied with a deranging of liver function after ligation, as anticipated, but not in all measured parameters; biochemical and endotoxin levels followed the same pattern.

Detection of superoxide radical in all biological systems by Thin Layer Chromatography.

The study presents a new method that detects O2•-, via quantification of 2-hydroxyethidium (2-ΟΗ-Ε+) as low as ∼30 fmoles by High-Performance Thin Layer Chromatography (HPTLC). The method isolates 2-ΟΗ-Ε+ after its extraction by the anionic detergent SDS (at 18-fold higher than its CMC) together with certain organic/inorganic reagents, and its HPTLC-separation from di-ethidium (di-Ε+) and ethidium (Ε+). Quantification of 2-OH-E+ is based on its ex/em maxima at 290/540 nm, and of di-E+ and E+ at 295/545 nm. The major innovations of the present method are the development of protocols for (i) efficient extraction (by SDS) and (ii) sensitive quantification (by HPTLC) for 2-OH-E+ (as well as di-E+ and E+) from most biological systems (animals, plants, cells, subcellular compartments, fluids). The method extracts 2-ΟΗ-Ε+ (by neutralizing the strong binding between its quaternary N+ and negatively charged sites on phospholipids, DNA etc) together with free HE, while protects both from biological oxidases, and also extracts/quantifies total proteins (hydrophilic and hydrophobic) for expressing O2•- levels per protein quantity. The method also uses SDS (at 80-fold lower than its CMC) to extract/remove/wash 2-ΟΗ-Ε+ from cell/organelle exterior membrane sites, for more accurate internal content quantification. The new method is applied on indicative biological systems: (1) artificially stressed (mouse organs and liver mitochondria and nuclei, ±exposed to paraquat, a known O2•- generator), and (2) physiologically stressed (cauliflower plant, exposed to light/dark).

Oxidized lipid-associated protein damage in children and adolescents with type 1 diabetes mellitus: New diagnostic/prognostic clinical markers.

BACKGROUND: Type 1 diabetes mellitus (DM1), a chronic metabolic disorder of autoimmune origin, has been associated with oxidative stress (OS), which plays a central role in the onset, progression, and long-term complications of DM1. The markers of OS lipid peroxidation products, lipid hydroperoxides (LOOH), and also malondialdehyde (MDA) and thiobarbituric reactive substances (TBARS) that oxidatively modify proteins (Pr) (i.e., PrMDA and PrTBARS, respectively), have been associated with DM2, DM1, diabetic neuropathy, and microalbuminuria. OBJECTIVE/SUBJECTS: Here, we investigated LOOH, PrMDA and PrTBARS in 50 children and adolescents with DM1 and 21 controls. RESULTS: The novel OS marker PrTBARS was assessed for the first time in children and adolescents with DM1. LOOH and the pair PrMDA/PrTBARS, representing early and late peroxidation stages, respectively, are found to be significantly higher (130%, 50/90%, respectively, at p < 0.001) in patients with DM1 compared to controls. The studied OS parameters did not differ with age, age at diagnosis, sex, duration of DM1, presence of recent ketosis/ketoacidosis, or mode of treatment. CONCLUSIONS: We propose that LOOH, PrMDA and the new marker PrTBARS could serve as potential diagnostic clinical markers for identifying OS in children and adolescents with DM1, and may, perhaps, hold promise as a prognostic tool for future complications associated with the disease.

Oxidative Stress and NADPH Oxidase: Connecting Electromagnetic Fields, Cation Channels and Biological Effects.

Electromagnetic fields (EMFs) disrupt the electrochemical balance of biological membranes, thereby causing abnormal cation movement and deterioration of the function of membrane voltage-gated ion channels. These can trigger an increase of oxidative stress (OS) and the impairment of all cellular functions, including DNA damage and subsequent carcinogenesis. In this review we focus on the main mechanisms of OS generation by EMF-sensitized NADPH oxidase (NOX), the involved OS biochemistry, and the associated key biological effects.

Age-related aqueous humor (AH) and lens epithelial cell/capsule protein carbonylation and AH protein concentration in cataract patients who have pseudoexfoliative diseases.

Purpose: The aim of this study is to investigate the age-correlation of oxidative stress (OS, assessed by the accumulative OS damage marker protein carbonyls) in aqueous humour (AH; together with protein concentration) and lens epithelial cells plus capsule (LECs/capsule) in patients with cataract (CAT), and also suffering from pseudoexfoliation syndrome (PEX), primary open-angle glaucoma (POAG) and pseudoexfoliation glaucoma (PXG). Methods: AH samples from 78 male/female patients (21, 20, 19 and 18 with CAT, PEX, PXG, and POAG, respectively), and LECs/capsule samples from 104 male/female patients (34, 32, 18, and 20 with CAT, PEX, PXG and POAG, respectively) were collected during phacoemulsification CAT surgery. Average protein carbonyl concentrations were measured in patients grouped in 5-year age intervals (ranging from 56-60 to 86-90). The non-overlapping age ranges and numbers of the tested subjects did not allow comparative follow up studies for the tested diseases. Results: There is an age-dependent increase of protein carbonyls in AH (nmol mg-1 protein and ml-1), and in the order CAT

Ischemia-Reperfusion Injury of Sciatic Nerve in Rats: Protective Role of Combination of Vitamin C with E and Tissue Plasminogen Activator.

An ischemia/reperfusion injury of rat's sciatic nerve was experimentally developed. In this model, we measured the in vivo production of superoxide radical, as a marker of oxidative stress and the occludin expression as an indicator of blood-nerve barrier function and we examined potential protective innervations against these abnormalities. Right sciatic nerves of the animals underwent 3 h of ischemia followed by 7 days of reperfusion and were divided into three groups: ischemic, pretreated with vitamin C in conjunction with vitamin E and treated with tissue plasminogen activator. Compared to measurements from left sciatic nerves used as sham, the ischemic group showed significantly increased superoxide radical and reduced expression of occludin in western blot and immunohistochemistry. No such differences were detected between sham and nerves in the vitamin or tissue plasminogen activator groups. It is suggested that the experimental ischemia/reperfusion model was suitable for studying the relationship between oxidative state and blood-nerve barrier. The reversion of abnormalities by the applied neuroprotective agents might prove to be a clinically important finding in view of the implication of vascular supply derangement in various neuropathies in humans.

An accurate and sensitive Coomassie Brilliant Blue G-250-based assay for protein determination.

In this protocol we present a rapid and sensitive assay for the accurate determination of protein concentration. The assay is a modification of a previous method, and measures minimum 0.2 µg protein.

Role of oxidative stress in Sclerotial differentiation and aflatoxin B1 biosynthesis in Aspergillus flavus.

We show here that oxidative stress is involved in both sclerotial differentiation (SD) and aflatoxin B1 biosynthesis in Aspergillus flavus. Specifically, we observed that (i) oxidative stress regulates SD, as implied by its inhibition by antioxidant modulators of reactive oxygen species and thiol redox state, and that (ii) aflatoxin B1 biosynthesis and SD are comodulated by oxidative stress. However, aflatoxin B1 biosynthesis is inhibited by lower stress levels compared to SD, as shown by comparison to undifferentiated A. flavus. These same oxidative stress levels also characterize a mutant A. flavus strain, lacking the global regulatory gene veA. This mutant is unable to produce sclerotia and aflatoxin B1. (iii) Further, we show that hydrogen peroxide is the main modulator of A. flavus SD, as shown by its inhibition by both an irreversible inhibitor of catalase activity and a mimetic of superoxide dismutase activity. On the other hand, aflatoxin B1 biosynthesis is controlled by a wider array of oxidative stress factors, such as lipid hydroperoxide, superoxide, and hydroxyl and thiyl radicals.

DNA fragmentation induced by all-trans retinoic acid and its steroidal analogue EA-4 in C2 C12 mouse and HL-60 human leukemic cells in vitro.

We have recently shown that retinoic acid induces micronucleation mainly via chromosome breakage (Alakhras et al. Cancer Lett 2011; 306: 15-26). To further study retinoic acid clastogenicity and evaluate DNA damaging potential we investigated the ability of (a) all-trans retinoic acid and its steroidal analogue EA-4 to induce DNA fragmentation by using Comet assay under alkaline unwinding and neutral condition electrophoresis, and (b) the retinoids under study to induce small (0-1 kb) DNA fragments. Two cell lines, C2C12 mouse cells and HL-60 human leukemic cells were used in this study. We found that all-trans retinoic acid and its steroidal analogue EA-4 (a) provoke DNA migration due to DNA fragmentation as it is shown by the increased values of Comet parameters, and (b) induce significantly small-size fragmented genomic DNA as indicated by the quantification of necrotic/apoptotic small DNA segments in both cell systems. A different response between the two cell lines was observed in relation to retinoid ability to increase the percentage of DNA in the tail as well as break DNA in to small fragments. Our findings confirm the ability of retinoic acid to provoke micronucleation by disrupting DNA into fragments, among which small pieces of double-stranded DNA up to 1 kb are identified.

New Clinical Markers of Oxidized Lipid-Associated Protein Damage in Children and Adolescents with Obesity.

Obesity in children and adolescents has been associated with oxidative stress (OS). The lipid hydroperoxides (LOOH) and the malondialdehyde (MDA) and thiobarbituric reactive substances (TBARS) that oxidatively modify proteins (Pr) (i.e., PrMDA and PrTBARS, respectively) represent markers of OS-associated lipid peroxidation. We aimed to assess OS in children and adolescents with obesity using-for the first time-markers involved in the early and late lipid oxidation process. LOOH, PrMDA, and PrTBARS were investigated in 41 children and adolescents with obesity and 31 controls. Obesity was defined as BMI > 95% for age and sex. The PrMDA/PrTBARS pair, which reflects a late peroxidation stage, was found to be significantly high (39%/180%) in children and adolescents with obesity compared to controls (p < 0.001). Similarly, the early LOOH peroxidation stage marker was increased by 30%. The studied OS parameters were not influenced by sex or age. Our study introduces LOOH, PrTBARS, and PrMDA as markers for evaluating OS in children and adolescents with obesity. LOOH, PrTBARS, and PrMDA may also hold promise as prognostic markers for potential obesity-associated long-term complications.

Future of the Search for Life: Workshop Report.

The 2-week, virtual Future of the Search for Life science and engineering workshop brought together more than 100 scientists, engineers, and technologists in March and April 2022 to provide their expert opinion on the interconnections between life-detection science and technology. Participants identified the advances in measurement and sampling technologies they believed to be necessary to perform in situ searches for life elsewhere in our Solar System, 20 years or more in the future. Among suggested measurements for these searches, those pertaining to three potential indicators of life termed "dynamic disequilibrium," "catalysis," and "informational polymers" were identified as particularly promising avenues for further exploration. For these three indicators, small breakout groups of participants identified measurement needs and knowledge gaps, along with corresponding constraints on sample handling (acquisition and processing) approaches for a variety of environments on Enceladus, Europa, Mars, and Titan. Despite the diversity of these environments, sample processing approaches all tend to be more complex than those that have been implemented on missions or envisioned for mission concepts to date. The approaches considered by workshop breakout groups progress from nondestructive to destructive measurement techniques, and most involve the need for fluid (especially liquid) sample processing. Sample processing needs were identified as technology gaps. These gaps include technology and associated sampling strategies that allow the preservation of the thermal, mechanical, and chemical integrity of the samples upon acquisition; and to optimize the sample information obtained by operating suites of instruments on common samples. Crucially, the interplay between science-driven life-detection strategies and their technological implementation highlights the need for an unprecedented level of payload integration and extensive collaboration between scientists and engineers, starting from concept formulation through mission deployment of life-detection instruments and sample processing systems.

Intestinal mucosal proliferation, apoptosis and oxidative stress in patients with liver cirrhosis.

BACKGROUND: Intestinal mucosal barrier dysfunction in liver cirrhosis and its implicated mechanisms is of great clinical importance because it is associated with the development of serious complications from diverse organs through promotion of systemic endotoxemia. AIM: The present study was designed to investigate whether enterocytes' proliferation, apoptosis and intestinal oxidative stress are altered in the intestinal mucosa of patients with compensated and decompensated liver cirrhosis. MATERIAL AND METHODS: Twelve healthy controls (group A) and twenty four cirrhotic patients at a compensated (n = 12, group B) or decompensated condition (n = 12, group C) were subjected to duodenal biopsy. In intestinal specimens mucosal apoptotic and mitotic activity and their ratio were recorded by means of morphological assessment and mucosal lipid hydroperoxides were measured. Plasma endotoxin concentration, an index of gut barrier function, was also determined. RESULTS: Cirrhotic patients presented significantly higher serum endotoxin concentrations as compared to healthy controls (P < 0.001), whilst endotoxemia was higher in decompensated disease (P < 0.05 vs. compensated cirrhosis). Intestinal mucosal mitotic count was significantly lower in patients with compensated and decompensated cirrhosis compared to controls (P < 0.01, respectively), whilst a trend towards increased apoptosis was recorded. The mitotic/apoptotic ratio was significantly reduced in groups B (P < 0.05) and C (P < 0.01) as compared to controls. Intestinal lipid peroxidation was significantly increased in decompensated cirrhotics (P < 0.001 vs. groups A and B). CONCLUSIONS: The present study demonstrates for the first time that human liver cirrhosis is associated with decreased intestinal mucosal proliferation and proliferation/apoptosis ratio even at early stages of cirrhosis and increased intestinal oxidative stress in advanced liver disease.

Organic Catalytic Activity as a Method for Agnostic Life Detection.

An ideal life detection instrument would have high sensitivity but be insensitive to abiotic processes and would be capable of detecting life with alternate molecular structures. In this study, we propose that catalytic activity can be the basis of a nearly ideal life detection instrument. There are several advantages to catalysis as an agnostic life detection method. Demonstrating catalysis does not necessarily require culturing/growing the alien life and in fact may persist even in dead biomass for some time, and the amplification by catalysis is large even by minute amounts of catalysts and, hence, can be readily detected against abiotic background rates. In specific, we propose a hydrolytic catalysis detection instrument that could detect activity in samples of extraterrestrial organic material from unknown life. The instrument uses chromogenic assay-based detection of various hydrolytic catalytic activities, which are matched to corresponding artificial substrates having the same, chromogenic (preferably fluorescent) upon release, group; D- and L-enantiomers of these substrates can be used to also answer the question whether unknown life is chiral. Since catalysis is a time-proportional product-concentration amplification process, hydrolytic catalytic activity can be measured on a sample of even a minute size, and with instruments based on, for example, optofluidic chip technology.

Novel oxidized LDL-based clinical markers in peritoneal dialysis patients for atherosclerosis risk assessment.

Maintenance peritoneal dialysis (PD) is commonly associated with cardiovascular diseases (CVDs), whose risk is assessed via LDL-C. Nonetheless, oxidized LDL (oxLDL), as being a key component of atherosclerotic lesions, could be also associated with atherosclerosis and related CVDs. However, its predictive value for CVDs risk assessment is subject of research studies due to the lack of specific methods to measure oxLDL status from its individual lipid/protein components. In the present study, six novel oxLDL markers, representative of certain oxidative modifications on the LDL protein and lipid components, are measured in atherosclerosis-prone PD patients (39) versus those in chronic kidney disease patients (61) under hemodialysis (HD) and healthy controls (40). LDL from serum of PD, HD and control subjects were isolated and fractionated into cholesteryl esters, triglycerides, free cholesterol, phospholipids and apolipoprotein B100 (apoB100). Subsequently the oxLDL markers cholesteryl ester hydroperoxides (-OOH), triglyceride-OOH, free cholesterol-OOH, phospholipid-OOH, apoB100 malondialdehyde and apoB100 dityrosines were measured. LDL carotenoid levels and LDL particle serum concentration were also measured. The levels of all oxLDL lipid-OOH markers were significantly elevated in PD patients versus control, while the levels of cholesteryl ester-/triglyceride-/free cholesterol-OOH were significantly elevated in PD versus HD patients, regardless of patients' underlying medical conditions, sex, age, PD type, clinical biochemical markers and medication. It should be noted that all fractionated lipid-OOH levels were inversely correlated with LDL-P concentration, while LDL-P concentration was not correlated with LDL-C in PD patients. Moreover, LDL carotenoids were significantly lower in PD patients versus control. The increased levels of oxLDL status specific markers in both PD and HD patients (compared to control), support a potential prognostic value of oxLDL regarding CVD risk assessment in both patient groups. Lastly, the study introduces the oxLDL peroxidation markers free cholesterol-OOH and cholesteryl ester-OOH as complementary to LDL-P number, and as possible alternatives to LDL-C.

Quantification of superoxide radical production in 4 vital organs of rats subjected to hemorrhagic shock.

OBJECTIVE: The aim of this study was to measure the production of superoxide radical (O2-), a direct indicator of oxidative stress, in 4 vital organs of rats subjected to hemorrhagic shock. For this purpose, and for the first time, a new quantitative assay for the ex vivo measurement of O2- via an established 1:1 molar relationship between O2- and 2-OH-ethidium was used. The production of lipid hydroperoxides (LOOHs), a standard method of evaluation of oxidative stress, was also used for reasons of comparison. METHODS: Sixteen male Wistar rats were divided into 2 groups: sham and hemorrhagic shock, targeting to a mean arterial pressure of 30 to 40 mm Hg for 60 minutes. Three hours after resuscitation, tissues were collected for measurement of LOOHs and O2- production. RESULTS: Hemorrhagic shock induced increased production of LOOHs in the gut, liver, and lungs (P<.001), whereas the production of O2- was also increased in the gut (P<.001), liver (P<.001), and, to a lesser extent, in the lungs (P<.05). The oxidative load of the kidneys, as estimated by both techniques, remained unaffected. CONCLUSION: The results of this new O2- assay were comparable with the results of the established LOOHs method, and this assay proved to be accurate and sensitive in the detection and quantification of O2- production in all organs tested. Thus, the proposed direct measurement of O2- in critically ill patients often facing in extremis situations could be used as a prognostic tool and as a method to evaluate therapeutic interventions in the setting of emergency medicine.

Time progression and regional expression of brain oxidative stress induced by obstructive jaundice in rats.

BACKGROUND: Obstructive jaundice induces oxidative changes in the brain parenchyma and plays significant role in clinical manifestations of hepatic encephalopathy. We aim to study the progression of the brain oxidative status over time and the differences of its pattern over the hemispheres, the brainstem and the cerebellum. We use an experimental model in rats and measuring the oxidative stress (OS) specific biomarkers protein malondialdehyde (PrMDA) and protein carbonyls (PrC = O). RESULTS: Hyperbilirubinemia has been confirmed in all study groups as the result of common bile duct obstruction. We confirmed increase in both PrMDA and PrC = O biomarkers levels with different type of changes over time. We also confirmed that the oxidative process develops differently in each of the brain areas in study. CONCLUSIONS: The present study confirms the progressive increase in OS in all brain areas studied using markers indicative of cumulative protein modification.

Methods on LDL particle isolation, characterization, and component fractionation for the development of novel specific oxidized LDL status markers for atherosclerotic disease risk assessment.

The present study uses simple, innovative methods to isolate, characterize and fractionate LDL in its main components for the study of specific oxidations on them that characterize oxidized low-density lipoprotein (oxLDL) status, as it causatively relates to atherosclerosis-associated cardiovascular disease (CVD) risk assessment. These methods are: (a) A simple, relatively time-short, low cost protocol for LDL isolation, to avoid shortcomings of the currently employed ultracentrifugation and affinity chromatography methodologies. (b) LDL purity verification by apoB100 SDS-PAGE analysis and by LDL particle size determination; the latter and its serum concentration are determined in the present study by a simple method more clinically feasible as marker of CVD risk assessment than nuclear magnetic resonance. (c) A protocol for LDL fractionation, for the first time, into its main protein/lipid components (apoB100, phospholipids, triglycerides, free cholesterol, and cholesteryl esters), as well as into LDL carotenoid/tocopherol content. (d) Protocols for the measurement, for the first time, of indicative specific LDL component oxidative modifications (cholesteryl ester-OOH, triglyceride-OOH, free cholesterol-OOH, phospholipid-OOH, apoB100-MDA, and apoB100-DiTyr) out of the many (known/unknown/under development) that collectively define oxLDL status, which contrasts with the current non-specific oxLDL status evaluation methods. The indicative oxLDL status markers, selected in the present study on the basis of expressing early oxidative stress-induced oxidative effects on LDL, are studied for the first time on patients with end stage kidney disease on maintenance hemodialysis, selected as an indicative model for atherosclerosis associated diseases. Isolating LDL and fractionating its protein and main lipid components, as well as its antioxidant arsenal comprised of carotenoids and tocopherols, paves the way for future studies to investigate all possible oxidative modifications responsible for turning LDL to oxLDL in association to their possible escaping from LDL's internal antioxidant defense. This can lead to studies to identify those oxidative modifications of oxLDL (after their artificial generation on LDL), which are recognized by macrophages and convert them to foam cells, known to be responsible for the formation of atherosclerotic plaques that lead to the various CVDs.

Maternal Calorie Restriction Induces a Transcriptional Cytoprotective Response in Embryonic Liver Partially Dependent on Nrf2.

BACKGROUND: Calorie restriction is known to enhance Nrf2 signaling and longevity in adult mice, partially by reducing reactive oxygen species, but calorie restriction during pregnancy leads to intrauterine growth retardation. The latter is associated with fetal reprogramming leading to increased incidence of obesity, metabolic syndrome and diabetes in adult life. Transcription factor Nrf2 is a central regulator of the antioxidant response and its crosstalk with metabolic pathways is emerging. We hypothesized that the Nrf2 pathway is induced in embryos during calorie restriction in pregnant mothers. METHODS: From gestational day 10 up to day 16, 50% of the necessary mouse diet was provided to Nrf2 heterozygous pregnant females with fathers being of the same genotype. Embryos were harvested at the end of gestational day 16 and fetal liver was used for qRT-PCR and assessment of oxidative stress (OS). RESULTS: Intrauterine calorie restriction led to upregulation of mRNA expression of antioxidant genes (Nqo1, Gsta1, Gsta4) and of genes related to integrated stress response (Chac1, Ddit3) in WT embryos. The expression of a key gluconeogenic (G6pase) and two lipogenic genes (Acacb, Fasn) was repressed in calorie-restricted embryos. In Nrf2 knockout embryos, the induction of Nqo1 and Gsta1 genes was abrogated while that of Gsta4 was preserved, indicating an at least partially Nrf2-dependent induction of antioxidant genes after in utero calorie restriction. Measures of OS showed no difference (superoxide radical and malondialdehyde) or a small decrease (thiobarbituric reactive substances) in calorie-restricted WT embryos. CONCLUSIONS: Calorie restriction during pregnancy elicits the transcriptional induction of cytoprotective/antioxidant genes in the fetal liver, which is at least partially Nrf2-dependent, with a physiological significance that warrants further investigation.

Superoxide radical induces sclerotial differentiation in filamentous phytopathogenic fungi: a superoxide dismutase mimetics study.

This study shows that the superoxide radical (O(2) *( -)), a direct indicator of oxidative stress, is involved in the differentiation of the phytopathogenic filamentous fungi Rhizoctonia solani, Sclerotinia sclerotiorum, Sclerotium rolfsii and Sclerotinia minor, shown by using superoxide dismutase (SOD) mimetics to decrease their sclerotial differentiation. The production rate of O(2) *(-) and SOD levels in these fungi, as expected, were significantly lowered by the SOD mimetics, with concomitant decrease of the indirect indicator of oxidative stress, lipid peroxidation.

The Oxygen Release Instrument: Space Mission Reactive Oxygen Species Measurements for Habitability Characterization, Biosignature Preservation Potential Assessment, and Evaluation of Human Health Hazards.

We describe the design of an instrument, the OxR (for Oxygen Release), for the enzymatically specific and non-enzymatic detection and quantification of the reactive oxidant species (ROS), superoxide radicals (O2•-), and peroxides (O22-, e.g., H2O2) on the surface of Mars and Moon. The OxR instrument is designed to characterize planetary habitability, evaluate human health hazards, and identify sites with high biosignature preservation potential. The instrument can also be used for missions to the icy satellites of Saturn's Titan and Enceladus, and Jupiter's Europa. The principle of the OxR instrument is based on the conversion of (i) O2•- to O2 via its enzymatic dismutation (which also releases H2O2), and of (ii) H2O2 (free or released by the hydrolysis of peroxides and by the dismutation of O2•-) to O2 via enzymatic decomposition. At stages i and ii, released O2 is quantitatively detected by an O2 sensor and stoichiometrically converted to moles of O2•- and H2O2. A non-enzymatic alternative approach is also designed. These methods serve as the design basis for the construction of a new small-footprint instrument for specific oxidant detection. The minimum detection limit of the OxR instrument for O2•- and O22- in Mars, Lunar, and Titan regolith, and in Europa and Enceladus ice is projected to be 10 ppb. The methodology of the OxR instrument can be rapidly advanced to flight readiness by leveraging the Phoenix Wet Chemical Laboratory, or microfluidic sample processing technologies.