Breath biomarkers and non-alcoholic fatty liver disease: preliminary observations.

Breath biomarkers have the potential to offer information that is similar to conventional clinical tests or they are entirely unique. Preliminary data support the use of breath biomarkers in the study of liver disease, in particular non-alcoholic fatty liver disease (NAFLD). It was evaluated whether breath ethanol, ethane, sulfur compounds and acetone would be associated with hepatic histopathology amongst morbidly obese patients presenting for bariatric surgery. Breath samples were collected during a preoperative visit and compared with liver biopsies obtained during the surgery. A Student's two-tailed t-test was used to compare differences between the two groups. Linear regression was used to analyse associations between the concentrations of breath molecules and independent predictor variables. It was found that breath ethanol, ethane and acetone can be useful biomarkers in patients with NAFLD. In particular, breath ethanol can be associated with hepatic steatosis, and breath acetone can be associated with non-alcoholic steatohepatitis.

Abnormal exhaled ethane concentrations in scleroderma.

Scleroderma (systemic sclerosis) is a chronic multisystem autoimmune disease in which oxidative stress is suspected to play a role in the pathophysiology. Therefore, it was postulated that patients with scleroderma would have abnormally high breath ethane concentrations, which is a volatile product of free-radical-mediated lipid peroxidation, compared with a group of controls. There was a significant difference (p<0.05) between the mean exhaled ethane concentration of 5.27 pmol ml(-1) CO(2) (SEM=0.76) in the scleroderma patients (n=36) versus the mean exhaled concentration of 2.72 pmol ml(-1) CO(2) (SEM=0.71) in a group of healthy controls (n=21). Within the scleroderma group, those subjects taking a calcium channel blocker had lower ethane concentrations compared with patients who were not taking these drugs (p=0.05). There was a significant inverse association between lung diffusion capacity for carbon monoxide (per cent of predicted) and ethane concentration (b=-2.8, p=0.026, CI=-5.2 to -0.35). These data support the presence of increased oxidative stress among patients with scleroderma that is detected by measuring breath ethane concentrations.

Patterns and significance of exhaled-breath biomarkers in lung transplant recipients with acute allograft rejection.

BACKGROUND: Obliterative bronchiolitis (OB) remains one of the leading causes of death in lung transplant recipients after 2 years, and acute rejection (AR) of lung allograft is a major risk factor for OB. Treatment of AR may reduce the incidence of OB, although diagnosis of AR often requires bronchoscopic lung biopsy. In this study, we evaluated the utility of exhaled-breath biomarkers for the non-invasive diagnosis of AR. METHODS: We obtained breath samples from 44 consecutive lung transplant recipients who attended ambulatory follow-up visits for the Johns Hopkins Lung Transplant Program. Bronchoscopy within 7 days of their breath samples showed histopathology in 21 of these patients, and we included them in our analysis. We measured hydrocarbon markers of pro-oxidant events (ethane and 1-pentane), isoprene, acetone, and sulfur-containing compounds (hydrogen sulfide and carbonyl sulfide) in exhaled breath and compared their levels to the lung histopathology, graded as stable (non-rejection) or AR. None of the study subjects were diagnosed with OB or infection at the time of the clinical bronchoscopy. RESULTS: We found no significant difference in exhaled levels of hydrocarbons, acetone, or hydrogen sulfide between the stable and AR groups. However, we did find significant increase in exhaled carbonyl sulfide (COS) levels in AR subjects compared with stable subjects. We also observed a trend in 7 of 8 patients who had serial sets of breath and histopathology data that supported a role for COS as a breath biomarker of AR. CONCLUSIONS: This study demonstrated elevations in exhaled COS levels in subjects with AR compared with stable subjects, suggesting a diagnostic role for this non-invasive biomarker. Further exploration of breath analysis in lung transplant recipients is warranted to complement fiberoptic bronchoscopy and obviate the need for this procedure in some patients.

Obesity and female gender increase breath ethanol concentration: potential implications for the pathogenesis of nonalcoholic steatohepatitis.

OBJECTIVES: Similarities between histological features of alcoholic hepatitis and obesity-related liver disease suggest a common pathogenic mechanism. Because intestinal bacteria can produce ethanol, it is conceivable that intestinally derived alcohol may contribute to fatty liver disease. An indirect way of measuring endogenous ethanol is to measure the breath ethanol concentration. In a previous study in ob/ob mice, breath ethanol decreased with a course of non-absorbable antibiotics, suggesting that the ethanol is derived from intestinal bacterial flora. The aims of this study were 1) to determine whether alcohol can be detected in the breath of human subjects, and 2) to assess whether there is any correlation between ethanol and obesity in patients with nonalcoholic steatohepatits (NASH) and control subjects without known liver disease. METHODS: Breath ethanol concentration was determined in 21 patients with biopsy-proven NASH and in 10 control subjects by gas chromatography. An abnormal breath ethanol level was defined as two standard deviations above the mean value of the breath ethanol of lean controls. RESULTS: Minute quantities of ethanol were detected in the breath of human subjects who had not consumed alcohol in the recent past. Patients who were obese were more likely to have higher breath ethanol concentrations. Women also had higher breath alcohol than men. However, there was no difference between patients with NASH and controls. Severity of liver disease, as evidenced by cirrhosis, did not influence the breath ethanol concentration. CONCLUSIONS: Higher breath ethanol concentrations are observed in obese subjects than in leaner ones. It is possible that intestinally derived ethanol may contribute to the pathogenesis of NASH.

Breath ethane as a marker of reactive oxygen species during manipulation of diet and oxygen tension in rats.

Breath ethane, O2 consumption, and CO2 production were analyzed in 24-mo-old female Fischer 344 rats that had been fed continuously ad libitum (AL) or restricted 30% of AL level (DR) diets since 6 wk of age. Rats were placed in a glass chamber that was first flushed with air, then with a gas mixture containing 12% O2. After equilibration, a sample of the outflow was collected in gas sampling bags for subsequent analyses of ethane and CO2. The O2 and CO2 levels were also directly monitored in the outflow of the chamber. O2 consumption and CO2 production increased for DR rats. Hypoxia decreased O2 consumption and CO2 production for the AL-fed and DR rats. These changes reflect changes in metabolic rate due to diet and PO2. A significant decrease in ethane generation was found in DR rats compared with AL-fed rats. Under normoxic conditions, breath ethane decreased from 2.20 to 1.61 pmol ethane/ml CO2. During hypoxia the levels of ethane generation increased, resulting in a DR-associated decrease in ethane from 2.60 to 1.90 pmol ethane/ml CO2. These results support the hypothesis that DR reduces the level of oxidative stress.

Ethane: a marker of lipid peroxidation during cardiopulmonary bypass in humans.

The goals of this study were to (1) determine the utility of quantification of ethane as a marker of ischemia-reperfusion during human cardiopulmonary bypass (CPB); and (2) determine, using an animal model for this surgical procedure, whether the mode of surgical approach produced increases the quantity of exhaled ethane. Human CPB was initiated following standard anesthetic and monitoring regimens. Samples of gas were collected at baseline and at multiple defined time points throughout the studies. Ethane was determined using cryogenic concentration and gas chromatography. Sternotomy increased exhaled ethane compared to baseline (p < .007; 5.8 +/- 1.7 vs. 3.0 +/- 0.7 nmol/m2 x min); ethane returned to baseline levels prior to the initiation of CPB. Aortic unclamping produced ethane elevation (p < .05; 2.3 +/- 0.8 vs. 1.5 +/- 0.4 nmol/m2 x min) with the levels being related to a lower cardiac index and a higher systemic vascular resistance post aortic unclamping. Termination of CPB significantly increased ethane levels compared to baseline (p < .002; 4.8 +/- 1.7 vs. 3.0 +/- 0.7 nmol/m2 x min). Independent variables that correlated with increased ethane measurements included a higher arterial blood pH on bypass and the change in hemoglobin pre- and post-CPB. Electrocautery, but not scalpel, incision of the porcine abdominal wall increased ethane levels significantly (p < .02). These results indicate that exhaled ethane may be a valuable marker of lipid peroxidation during and following CPB.

Clinical application of breath biomarkers of oxidative stress status.

Isolation and quantification of volatile breath biomarkers indicative of relevant alterations in clinical status has required development of new techniques and applications of existing analytical chemical methods. The most significant obstacles to successful application of this type of sample have been reduction in required sample volume permitting replicate analysis (an absolute requirement for all clinical studies), separation of the analyte(s) of interest from background molecules, water vapor and other molecules with similar physical properties, introduction of automation in analysis and the use of selective detection systems (electron impact mass spectrometry, flame photometric, thermionic detectors), and automated sample collection from the human subject. Advances in adsorption technology and trace gas analysis have permitted rapid progress in this area of clinical chemistry.

Effect of dietary patterns on measures of lipid peroxidation: results from a randomized clinical trial.

BACKGROUND: Free radical-mediated oxidative damage to lipids is thought to be an important process in the pathogenesis of atherosclerosis. Although previous studies have demonstrated a beneficial impact of antioxidant vitamin supplements on lipid peroxidation, the effect of dietary patterns on lipid peroxidation is unknown. METHODS AND RESULTS: During the 3-week run-in period of a randomized trial, 123 healthy individuals were fed a control diet, low in fruits, vegetables, and dairy products, with 37% of calories from fat. Participants were then randomized to consume for 8 weeks: (1) the control diet, (2) a diet rich in fruits and vegetables but otherwise similar to the control diet, and (3) a combination diet rich in fruits, vegetables, and low-fat dairy products and reduced in fat. Serum oxygen radical-absorbing capacity, malondialdehyde (an in vitro measure of lipid peroxidation), and breath ethane (an in vivo measure of lipid peroxidation) were measured at the end of run-in and intervention periods. Between run-in and intervention, mean (95% CI) change in oxygen radical-absorbing capacity (U/mL) was -35 (-93, 13) in the control diet, 26 (-15, 67) in the fruits and vegetables diet (P=0.06 compared with control), and 19 (-22, 54) in the combination diet (P=0.10 compared with control). Median (interquartile range) change in ethane was 0.84 (0.10, 1.59) in the control diet, 0.02 (-0.61, 0.83) in the fruits and vegetables diet (P=0.04 compared with control), and -1.00 (-1.97, 0.25) in the combination diet (P=0.005 compared with control). Change in malondialdehyde did not differ between diets. CONCLUSIONS: This study demonstrates that modification of diet can favorably affect serum antioxidant capacity and protect against lipid peroxidation.

Induction of apoptosis by particulate matter: role of TNF-alpha and MAPK.

Particulate matter (PM) is a major by-product from the combustion of fossil fuels. The biological target of inhaled PM is the pulmonary epithelium and resident macrophages. In this study, we demonstrate that cultured macrophages (RAW 264.7 cells) exposed continously to a well-defined model of PM [benzo[a]pyrene adsorbed on carbon black (CB+BaP)] exhibit a time-dependent expression and release of the cytokine tumor necrosis factor-alpha (TNF-alpha). CB+BaP also evoked programmed cell death or apoptosis in cultured macrophages as assessed by genomic DNA-laddering assays. The CB+BaP-induced apoptosis was inhibited when macrophages were treated with CB+BaP in the presence of a neutralizing antibody to TNF-alpha, suggesting that TNF-alpha plays an important role in mediating CB+BaP-induced apoptosis in macrophages. Interestingly, neither untreated carbon black nor benzo[a]pyrene alone induced apoptosis or caused the release of TNF-alpha in RAW 264.7 cells. Moreover, we observed that TNF-alpha activates mitogen-activated protein kinase (MAPK) activity, the extracellular signal-regulated kinases p42/p44, in a time-dependent manner. RAW 264.7 cells treated with PD-098059, a selective inhibitor of MAPK kinase activity, did not exhibit CB+BaP-induced apoptosis and TNF-alpha secretion. Furthermore, cells treated with the MAPK kinase inhibitor did not undergo TNF-alpha-induced apoptosis. Taken together, our data suggest that TNF-alpha mediates PM-induced apoptosis and that the MAPK pathway may play an important role in regulating this pathway.

Possible antioxidant effect of vitamin A supplementation in premature infants.

BACKGROUND: Increased lipid peroxidation caused by oxygen free radicals is thought to be one of the common pathogenetic mechanisms for the so-called oxygen radical diseases of prematurity. Since in vitro studies have shown that various forms of vitamin A can exert antioxidant effects that are more potent than those of vitamin E (treatment with which has been ineffective in these diseases), the purpose of this prospective, controlled study was to determine whether administration of supplemental vitamin A to premature infants deficient in this vitamin would have an antioxidant effect in vivo. METHODS: Fourteen infants (1181 +/- 35 g; gestational age 29 +/- 0.04 weeks) with a serum retinol concentration at 7 +/- 2 days of age in the deficient range, lower than 0.7 mumol/l (< 20 micrograms/dl), were enrolled in the study. Infants were randomized to receive the standard amount of vitamin A or standard plus supplemental (2.6 mumol/l [2500 IU] orally each day) vitamin A, beginning at 1 week of age. Antioxidant effects of supplementation were assessed by a decrease in lipid peroxidation, quantified by the ethane content of expired air. RESULTS: Three weeks after study enrollment, total daily vitamin A intake in the infants receiving supplements was 4.565 +/- 0.236 mumol (4354 +/- 225 IU) versus 1.879 +/- 0.317 mumol/l (1792 +/- 302 IU) in infants receiving standard amounts of the vitamin. In spite of the difference in intake of vitamin A, 3 weeks after study enrollment, serum retinol concentrations did not differ between the infants given supplements and those receiving standard amounts of vitamin A, 0.70 +/- 0.21 versus 0.66 +/- 0.07 mumol/l (20 +/- 6 micrograms/dl versus 19 +/- 2 micrograms/dl, respectively). In the infants receiving supplemental vitamin A, breath ethane values declined from baseline values. There was an inverse correlation between the number of weeks of supplementation and breath ethane values, whereas there was no significant correlation between the duration of the study and breath ethane values in the infants not given supplements. CONCLUSIONS: Our data suggest that supplementation with vitamin A in a small group of vitamin A-deficient preterm infants was associated with an antioxidant effect. Although no immediate clinical benefits were associated with supplementation, the data provide the rationale for future investigations of possible antioxidant effects of (larger amounts?) of vitamin A in higher risk premature infants born with subnormal serum retinol concentrations.

Association between cigarette smoking and lipid peroxidation in a controlled feeding study.

BACKGROUND: Cigarette smoke may promote atherogenesis by producing oxygen-derived free radicals that damage lipids. However, evidence in support of this hypothesis is inconsistent because most studies did not control for aspects of diet (antioxidants and lipid substrate) that may confound the association between smoking and measures of lipid peroxidation. METHODS AND RESULTS: The relationships between cigarette smoking and two measures of lipid peroxidation, breath ethane (an in vivo assay) and thiobarbituric acid-reactive substances (TBARS, an in vitro assay), were examined in 123 adults (11% of whom were smokers) participating in a controlled feeding study. After 3 weeks of controlled feeding on a common diet (36% total fat, 14% saturated fats, 6% polyunsaturated fats, and 12% monounsaturated fats), breath and fasting serum samples were collected for measurement of ethane and TBARS, respectively. Baseline characteristics of smokers and nonsmokers were similar, including several indices related to diet and nutritional status (albumin, cholesterol, body mass index, and oxygen radical-absorbing capacity). Cigarette smokers had significantly higher breath ethane (8.88 versus 1.71 pmol/L; P<.0001) and TBARS (24.0 versus 20.7 micromol/mL; P=.008) than nonsmokers. The interval between breath collection and the time the last cigarette was smoked was significantly and inversely correlated with breath ethane. Neither measure of lipid peroxidation was associated with measures of serum cholesterol or albumin, body mass index, or serum oxygen radical-absorbing capacity. CONCLUSIONS: Cigarette smokers have higher rates of in vivo and in vitro lipid peroxidation. These results support the hypothesis that the atherogenic effects of smoking are mediated in part by free radical damage to lipids.

Prooxidant effects of maternal smoking and formula in newborn infants.

BACKGROUND: The purpose of this study was to use the breath ethane test to determine if either maternal cigarette smoking, formula, and/or deficiency of the antioxidant nutrients vitamins A and E was associated with oxidant stress in newborn infants. The rationale for this study was: (1) our observation that cigarette smoking was a source of oxidant stress in pregnant women, suggesting that it could be a source of oxidant stress for infants exposed in utero; (2) formula was predicted to be prooxidant compared to colostrum, which contains several compounds with antioxidant activity in vitro; and (3) deficiencies of vitamins A and E have been shown to promote oxidant stress in experimental animals. METHODS: Breath ethane, a volatile alkane produced by peroxide of n-3 fatty acids, was utilized as an index of oxidant stress status. Forty-five healthy full-term infants of the women mentioned above were studied at 18-24 h of age, after four to six feedings of breast milk (colostrum) or caseinbased infant formula. Relationships between infant breath ethane, maternal smoking, mode of infant nutrition, and serum concentrations of the antioxidant vitamins A and E of infants were examined. RESULTS: The breath ethane of the entire group of infants whose mothers smoked (n = 19) was increased compared to values of infants whose mothers did not smoke (n = 26): 97 +/- 16 versus 43 +/- 9 pmol/kg/min, p < 0.03. When infants of mothers who smoked were eliminated from the analysis in order to study effects of nutrition alone, formula appeared to be prooxidant compared to breast milk. Breath ethane of formula-fed infants (n = 16) was 62 +/- 13 versus 13 +/- 4 pmol/kg/min for breast-fed infants (n = 10), p < 0.04. For the group as a whole, there was no correlation between infant breath ethane and serum concentrations of vitamins A and E. CONCLUSIONS: Exposure to maternal smoking in utero is prooxidant in newborn infants. Formula also has a prooxidant effect compared to colostrum in newborn infants not exposed to maternal smoking in utero. Further investigations will be necessary to explore the clinical consequences of these observations.

Breath isoprene: temporal changes in respiratory output after exposure to ozone.

Isoprene is a major hydrocarbon found in human breath. This study was conducted to evaluate whether respiratory isoprene output could serve as a monitor for ozone exposure. Healthy young adult subjects (n = 10) underwent chamber exposure on separate days to filtered air and to a variable concentration of ozone. Exposures had durations of 130 min that included alternate periods of rest and light treadmill exercise; breath was sampled pre- and postexposure. For six subjects, breath was resampled 19 +/- 1 h postexposure. Breath samples were concentrated cryogenically and analyzed by capillary gas chromatography. Isoprene output immediately postexposure was significantly reduced by ozone or filtered air (17 and 19%, respectively). These results suggest that exercise alone reduces isoprene levels in breath without an additive ozone effect. However, in the six subjects restudied 19 +/- 1 h postexposure to ozone, breath isoprene concentrations were now increased above the preexposure output by 99% (P < 0.01) and exceeded the 51% increase in output of isoprene at this time point after filtered-air exposure (P < 0.01). Therefore, breath isoprene is proposed as a noninvasive marker of a physiological response to oxidant-induced injury to epithelial membranes and fluid linings of the lower respiratory tract by ozone.

Breath ethane generation during clinical total body irradiation as a marker of oxygen-free-radical-mediated lipid peroxidation: a case study.

Total body irradiation (TBI) is used therapeutically for treatment of leukemias and other malignancies of the hemopoietic system. Ionizing radiation produces oxygen free radicals that contribute to cytotoxicity. Breath collected from one patient undergoing therapeutic TBI showed measurable changes in levels of ethane during treatment. Breath ethane is a marker of lipid peroxidation of n-3 fatty acids. The TBI treatment involved 4 days of irradiation. The largest changes in breath ethane occurred on Day 2. The increased levels of breath ethane on Day 2 were correlated to clinical manifestations of toxicity. The correlation of the onset of gastrointestinal side effects with higher levels of breath ethane suggests that breath ethane may be a clinically useful measure of the toxicity of various TBI fractionation treatment protocols currently in use at different medical centers. The levels of breath ethane on the other days of treatment were lower, suggesting that the oxidative-antioxidative balance of the patient may be important in protection against free radical mediated injury. These results for a single patient suggest that breath ethane may be a promising approach to elucidate the role of antioxidants in clinical TBI and should be extended for verification to a larger volunteer patient population.

Visceral lipid peroxidation occurs at reperfusion after supraceliac aortic cross-clamping.

PURPOSE: Recently, we have reported that lipid peroxidation specific to oxygen free radical-mediated injury increased immediately after reperfusion of human liver allografts. However, in the human liver transplantation setting it was impossible to disassociate the contributions to lipid peroxidation caused by the warm and cold ischemic phases from those caused by reperfusion. Therefore we now have studied lipid peroxidation at reperfusion after supraceliac aortic cross-clamping in patients with normal livers. METHODS: Ethane, a noninvasive biomaker of lipid peroxidation, was measured in exhaled breath of patients before and during cross-clamping of the thoracic aorta and at sequential time intervals after visceral reperfusion. RESULTS: Approximately a two-fold transient increase in the ethane level was observed at around 15 minutes after reperfusion in those patients whose aortas were cross-clamped for more than 18 minutes. CONCLUSIONS: These results indicate that free radical-mediated lipid peroxidation occurs at reperfusion of warm ischemic viscera in the clinical setting of aortic repair. This observation supports the hypothesis that substantial lipid peroxidation occurs when tissues are subjected to cold or warm ischemia followed by reperfusion.

Skin elicitation threshold of ethylbutyl thiourea and mercaptobenzothiazole with relative leaching from sensitizing products.

We studied 3 contact sensitizers present in rubber products, ethylbutyl thiourea (EBT), 2-mercaptobenzothiazole (MBT) and 2,2-dithio-bis-benzothiazole (MBTS), to relate the amount of sensitizer eliciting allergic contact dermatitis to the quantity leaching from a product into various biological fluids: normal saline, human plasma and 3 synthetic sweat solutions of pH 5.5 to 7.5. To determine the amount of sensitizer remaining after leaching, Soxhlet extraction with acetonitrile was subsequently performed. High-performance liquid chromatography was used for chemical analysis. 12 MBT-sensitive patients were patch tested with serial dilutions of MBT and MBTS in petrolatum. A Latin Square design was used in statistical analysis of variance of the patch test results. Large amounts of thioureas leached from 2 rubber articles eliciting thiourea dermatitis, the literature suggesting that these would have been well above the elicitation threshold. Leaching of MBTS was relatively greater than MBT into most media, whereas MBT was a more potent elicitor than MBTS at equivalent concentrations. The lowest eliciting concentration of MBT in 1 subject was 0.01%. Such information should prove helpful to manufacturers in designed products that do not release allergens sufficiently to cause reactions in consumers.

Induction of heat-shock gene expression in postischemic pig liver depends on superoxide generation.

BACKGROUND/AIMS: Both hemorrhagic and cardiogenic shock are associated with hepatic shock gene expression at resuscitation. This study investigated the potential role of intravascular superoxide anion as a proximal trigger of heat shock protein gene expression. METHODS: Preanesthetized pigs were subjected to 120 m of total warm hepatic ischemia. The survival model consisted of warm, total hepatic ischemia and reperfusion (with active portal-systemic bypass) followed by reperfusion and survival for 3 days. Serial hepatic biopsy samples were evaluated for the expression of heat shock protein 72 (HSP-72) messenger RNA (mRNA) by Northern and Western analysis and by in situ RNA hybridization. The possible role of intravascular O2- as a mediator of heat shock response was evaluated by its specific inhibition by the intravenous infusion of recombinant human superoxide dismutase (SOD). RESULTS: Ischemia for 120 minutes followed by 60 minutes of reperfusion caused accumulation of HSP-72 mRNA. Transcripts were localized to hepatocytes. HSP-72 mRNA was detected neither following ischemia alone nor when SOD was infused for 15 minutes at reperfusion. Three days later, transcripts were not detectable, but HSP-72 protein accumulated irrespective of SOD administration. CONCLUSIONS: Warm hepatic ischemia induces the hepatocyte expression of HSP-72 at reperfusion by a mechanism that is dependent upon the superoxide anion, probably generated intravascularly. However, the transient dismutation of superoxide is insufficient to suppress subsequent accumulation of HSP-72.

Evidence for free radical-mediated lipid peroxidation at reperfusion of human orthotopic liver transplants.

BACKGROUND: Generation of toxic oxygen metabolites at reperfusion may contribute to the injury sustained as a consequence of harvest and ischemic preservation of organ allografts. Because there is a paucity of evidence that this mechanism is operative in human beings, we measured the generation of ethane into the exhaled breath as a biomarker of free radical-mediated lipid peroxidation in human liver transplantation. METHODS: A novel technique that increased the previous standard of sensitivity 100-fold was used to measure picomole quantities of ethane in exhaled breath of eight recipients undergoing human orthotopic liver transplantation. RESULTS: Ethane production correlated closely with the specific events of liver transplantation including the initial reperfusion of the allografts. In every case a twofold to threefold increase in ethane production was superimposed on a stable baseline immediately after reestablishment of portal vein blood flow through the donor liver. CONCLUSIONS: Ethane production was interpreted as evidence of hepatic lipid peroxidation, presumably mediated by toxic metabolites of oxygen occurring at reperfusion. This noninvasive approach allowed localization of the time point at which lipid peroxidation occurred and may facilitate quantification of lipid peroxidation mediated by free radicals and other toxic oxygen metabolites during operation.

Breath ethane: a specific indicator of free-radical-mediated lipid peroxidation following reperfusion of the ischemic liver.

A major component of the organ injury mediated by toxic oxidants, such as seen following reperfusion of the ischemic liver, is due to the peroxidation of polyunsaturated fatty acids, especially of cell membranes. We utilized the measurement of exhaled breath ethane, a metabolic product unique to oxidant-mediated lipid peroxidation, as a noninvasive indicator of this process in swine liver subjected to warm ischemia/reperfusion. Under rigorously controlled anesthesia conditions, pig livers were subjected to 2 h of warm total ischemia, followed by reperfusion in situ. Expired air was collected and its ethane content quantitated by a novel gas chromatographic technique. The time course of breath ethane generation correlated closely with the appearance of hepatocellular injury as measured by impairment of Factor VII generation and other measures of liver integrity. Moreover, the administration of the specific superoxide free radical scavenger, superoxide dismutase (SOD), significantly attenuated both the elaboration of ethane and the hepatocellular injury. These findings not only provide confirmation of the previously reported link between hepatocellular injury by free radicals generated at reperfusion, but also establish the use of expired breath ethane analysis as a sensitive, specific, and noninvasive indicator of the injury process in real time.

Use of physical chemistry and in vivo exposure to investigate the toxicity of formaldehyde bound to carbonaceous particles in the murine lung.

Knowledge about the health effects of exposure to formaldehyde associated with automotive emissions is of pivotal importance in the risk assessment of this agent. Mobile sources emit many combustion-derived pollutants, including formaldehyde, in association with respirable carbon particles. Because it is hydrophilic, most of the inhaled formaldehyde is absorbed in the upper respiratory tract. However, if the organic vapor is adsorbed on respirable particles, formaldehyde may be deposited in the deep lung with the inhaled particles and may be available to interact adversely with cells along the lung parenchyma. On the respiratory surface, the alveolar macrophage phagocytic system plays the pivotal role in defending the lung against infectious agents. Susceptibility to respiratory infections is a relevant and sensitive indicator of the adverse effects of air pollution because acute and chronic exposures to a variety of air pollutants have been shown to decrease pulmonary antibacterial defenses. The goal of this research was to investigate whether exposure to formaldehyde decreases resistance to respiratory infections through dysfunctions of the alveolar macrophage phagocytic system. The study also explored whether interactions between formaldehyde and respirable carbon black particles alter susceptibility to respiratory infections and impairment of alveolar macrophage phagocytosis by delivering adsorbed formaldehyde to the deep lung with the inhaled particles. A carbon black, Regal GR, was used in these studies as a surrogate for the carbonaceous core of Diesel particulate matter. This material was selected to represent the worst-case scenario because the carbon black was expected to adsorb formaldehyde strongly. To accomplish this goal, mice were exposed to formaldehyde and to carbon black and formaldehyde combinations; increased susceptibility to respiratory infections was quantified by alveolar macrophage-dependent intrapulmonary killing of Staphylococcus aureus after an inhalation challenge with the bacterium. The salient findings of the bactericidal studies are as follows: Fifteen parts per million (ppm)* formaldehyde impaired the intrapulmonary killing of S. aureus when exposure followed the bacterial challenge. One ppm formaldehyde impaired the intrapulmonary killing of S. aureus when exposure preceded and was continued after the bacterial challenge. Coexposures to target concentrations of 3.5 mg/m3 carbon black and 2.5 ppm formaldehyde, or 10 mg/m3 carbon black and 5 ppm formaldehyde after the bacterial challenge had no effect on the intrapulmonary killing of S. aureus. Preexposure for four hours per day for four days to target concentrations of 3.5 mg/m3 carbon black and 2.5 ppm formaldehyde had no effect on the intrapulmonary killing of S. aureus when the assay was performed one day after the cessation of exposure.(ABSTRACT TRUNCATED AT 400 WORDS)