13CO2 Breath Test Research Articles

Novel high resolution melt curve assay for the analysis of predominance of Helicobacter pylori clarithromycin resistance.

Clarithromycin resistance is the most common cause of Helicobacter pylori treatment failure and it is attributed to three point mutations, A2142G, A2142C and A2143G, within the 23S rRNA gene. We aimed to determine the prevalence of H. pylori clarithromycin resistance using a novel high resolution melt assay. A total of 151 stool samples were collected from treatment-naïve patients with general gastric discomfort who also performed 13CO2 breath tests. Stool antigen tests were also performed on 126 of the 151 stool samples collected. Bacterial DNA was extracted from the stool and analyzed by comparing it with four reference plasmids incorporating the three mutations and the wild type (WT) sequences. The melt assay detected 106 H. pylori positive samples, of which 54 had a WT sequence, and 52 had a point mutation associated with clarithromycin resistance, including A2142G in 10, A2142C in 13, A2143G in 18 and heterozygosity (multiple peaks) in 11. Compared with the gold standards (13CO2 breath and stool antigen tests), the melt assay had a sensitivity of 100% and 99% and a specificity of 82% and 78%, respectively. Therefore, our stool-based molecular assay is able to identify H. pylori infection and clarithromycin resistance. It could be used for screening prior to administration of clarithromycin eradication therapy.

The effect of resistant starches on fat oxidation in healthy adults as measured by a 13CO2-breath test

ABSTRACTA resistant starch (RS) mixture (MIX) consisting of fibre of potatoes (FP) and wrinkled pea starch (WPS), and high amylose maize starch (HAMS) were supplemented in adults to evaluate their effects on fat oxidation by means of a 13CO2-breath test. Sixteen subjects received a regular diet either without or with the supplementation of MIX and HAMS in randomised order. After administration of a [U-13C]algal lipid mixture, exhaled air was collected over 14 h in 0.5- and 1-h intervals. The 13C abundances were measured by nondispersive infrared spectroscopy. In comparison to the dry run (DR), supplementation with MIX and with HAMS increased the cumulative percentage dose recovery: (DR: 16.7 %, MIX: 16.9 %, HAMS: 18.0 %), but without statistical significance. The colonic degradation of MIX and HAMS to short-chain fatty acids tends to lower the formation of carbohydrate-derived acetyl-CoA and contributes to a postprandial lipid oxidation increase by using fat-derived acetyl-CoA as a compensatory fuel source.

Reproducibility of two 13CO2 breath tests dedicated to assess pancreatic exocrine function

The aim of this study was to check on the reproducibility of two breath tests intended to test the pancreatic exocrine function accomplished with 13C-mixed triglyceride (13C-MTG) or cornflakes naturally enriched in 13C (13C-CF). The 13CO2 content within breath samples was determined with isotope-selective non-dispersive infrared spectrometry. A 72-h monitoring performed in healthy subjects revealed that a statistically significant rise in breath 13CO2 occurs between the 1st and the 9th hour and between the 1st and the 24th hour after intake of a test meal containing 300 mg 13C-MTG (n=10) or 100 g 13C-CF (n=12), respectively. In another two groups of 12 healthy volunteers each, short-term reproducibility of the two tests was assessed with paired examinations taken at a median interval of two days, whereas paired examinations separated by a median of 20 days served for the medium-term reproducibility assessment. In the case of either test, the medium-term reproducibility was not any worse than the short-term one. The reproducibility of the 13C-CF breath test tended to be slightly worse than that of the 13C-MTG breath test: a least detectable difference in 6-h cumulative 13C breath excretion (which is expressed as the percentage of the administered dose of the substrate) amounted to 2.7 and 4.4 % (short-term reproducibility) and to 3.5 and 4.4 % (medium-term reproducibility) in the case of the 13C-MTG breath test and the 13C-CF breath test, respectively. It is concluded that both tests offer a satisfactory reproducibility for use within a clinical setting. In case the lipolytic and the amylolytic activity would be required to be examined in the same patient, the 13C-CF breath test can be executed on the next day following the 13C-MTG breath test, whereas reciprocally, a 1-day break is recommended before accomplishment of a 13C-MTG breath test following a 13C-CF breath test.

Quantitative determination of metabolization dynamics by a real-time 13CO2 breath test

We have developed a flow-through fast liver investigation packet (FLIP) to quantitatively measure the amount of exhaled 13CO2, as a result of liver metabolization processes. The FLIP system allows investigation of every single breath in real time. Line width variations due to interactions with other gas components disturb traditional measurements and limit their sensitivity to the ppm range. Detection of the complete breath volume and the spectrally fully resolved line shape allows sensitivity in the ppb range with a standard deviation of approximately 80 ppb, a prerequisite to quantitatively analyze liver metabolization processes.

13CO2 breath tests, a tool to assess intestinal and liver function in the ICU?

CO2 breath tests, based on nondispersive infrared (NDIR) measurement technology, offer the chance for a noninvasive online quantitation of physiological processes. With recent extensions of the NDIR measurement technology breath tests could be used in an ICU setting without any restriction. Since online tools to assess feed intolerance with gastric emptying and to quantify gradual changes in liver function are still missing, this review explores the potential of breath tests to fill this gap. Gastric emptying may be correctly quantified; however, the corresponding tests fail to capture aberrations in nutrient absorption, which appear to be linked to hyperglycemia and insulin resistance and may contribute to feed intolerance. The progression or ameloriation of liver injury could be assessed with breath tests, which cannot, however, distinguish between functional and macrocirculatory or microcirculatory disorders. Breath tests for liver function and gastric emptying taken alone may fail in their ability to specifically quantify the clinical disorders. However, invasive test extensions could help to characterize their validity and thus to identify additional, noninvasive measurements to assure and expand this domain.

Effect of alcohol consumption on the liver detoxication capacity as measured by [13C2]aminopyrine and L-[1-13C]phenylalanine breath tests

The aim of this study was to investigate the hepatic microsomal and cytosolic functions by using the 13CO2 breath test in healthy subjects either before or after consumption of red wine. Twelve adults received [13C2]aminopyrine and l-[1-13C]phenylalanine together with a standardised dinner. Expired air samples were taken over 6 h. After a wash-out period, the subjects consumed 0.4 ml ethanol per kg per day together with dinner over a 7.5-day period on average. Thereafter, 13C-tracer administration was repeated under identical conditions. The 13CO2 enrichments were measured by isotope ratio mass spectrometry. The mean cumulative percentage 13C-dose recovery after administration of [13C2]aminopyrine and l-[1-13C]phenylalanine either without or with red wine consumption amounted to 17.0±4.4 vs. 14.7±3.1% (p=0.170) and 14.0±2.8 vs. 11.5±3.9% (p=0.084), respectively. Moderate alcohol consumption does not induce significant short-term changes of the microsomal and the cytosolic function of the human liver in healthy subjects.

Effects of postgastric13C-acetate processing on measurement of gastric emptying: a systematic investigation in health

Uniform postgastric processing of the gastric emptying (GE) marker 13C-acetate (Ac) is an unverified assumption behind its widespread application to measure GE. This study assessed the postgastric processing of Ac administered by intraduodenal (i.d.) infusion simulating different physiological conditions. 13CO2 in breath was assessed in three groups of six volunteers after i.d. administration of A: Different caloric densities (0.75/1.5/3 kcal min(-1) in a 200 mL meal at constant 1 mg Ac min(-1) simulating a physiological range of nutrient delivery rates; B: different tracer delivery rates (0.5/1.0/2.5 mg Ac min(-1) simulating delayed, normal and increased GE; C1: a 500 mL meal resulting in same marker and caloric delivery compared to protocol A; C2: 50 mL water bolus injections of 12.5/25/50/100 mg Ac and C3 bolus injections of 50 mg Ac in 50/100/200 mL water in randomized order. A: 13CO2 excretion was independent of caloric load (P = 0.59). B: The dynamic of 13CO2 excretion was modulated by tracer elimination which was in turn dependent on the speed of tracer delivery, i.e. with faster deliveries resulting in lower 13CO2 recovery during infusion (P < 0.001). C: Increasing Ac doses resulted in decreased 13CO2 recovery (P < 0.001) over the first hour. 13CO2 recovery kinetics was independent of the volume delivered. This study shows 13C-acetate absorption and metabolism is independent of the volume and caloric delivery of test meals. The 'lag' in estimates of GE derived from 13CO2 breath tests is due to a postgastric, dose-dependent delay to 13CO2 elimination. This can be corrected for in analytical derivations of GE parameters based on 13C-acetate breath test measurements.

Effect of arabinoxylo-oligosaccharides on proximal gastrointestinal motility and digestion in healthy volunteers

Summary Background & aims Before evaluating the effect of arabinoxylo-oligosaccharides (AXOS), a newly proposed prebiotic, on colonic metabolism, we evaluated its influence on proximal gastrointestinal motility [gastric emptying (GE) and oro-caecal transit time (OCTT)] and digestion of proteins and lipids. Methods Ten healthy volunteers each performed 2 13CO2-breath tests for assessing protein digestion and 2 for lipid digestion, each in the absence and presence of 2.2 g AXOS. To each test meal, 14C-labelled octanoate and inulin-14C-carboxylic acid were added for simultaneous measurement of GE and OCTT, respectively. Breath samples were collected and analysed with isotope ratio mass spectrometry (13C) or liquid scintillation (14C). Results Addition of AXOS to the lipid test meal did not influence lipid digestion. However, GE but not OCTT of the lipid test meal was significantly accelerated by ingestion of AXOS (p = 0.028). In the presence of AXOS, the rate of protein digestion proceeded slower but the overall extent of protein digestion after 5 h was not significantly different. GE and OCTT of the protein digestion test meal remained unchanged. Conclusions Addition of a low dose of AXOS to a meal does not influence the supply of lipids and proteins to the colon. Therefore, possible effects of AXOS on the colonic metabolism should not be due to differences in proximal motility and digestion.

Effect of alcohol consumption on the liver detoxication capacity as measured by [13C]methacetin- and [methyl-13C]methionine-breath tests

The aim of this study was to investigate the hepatic microsomal and mitochondrial functions by using the 13CO2-breath test in healthy subjects either before or after the consumption of red wine. Fourteen adults received [13C]methacetin and [methyl-13C]methionine together with a standardised dinner. Expired air samples were taken over 6 h. After a wash-out period, the subjects consumed 0.4 ml ethanol/kg/day together with dinner over a 10-day period. Thereafter, 13C-tracer administration was repeated under identical conditions. The 13CO2-enrichments were measured by isotope ratio mass spectrometry. The mean cumulative percentage 13C-dose recovery (CPDR) after administration of [13C]methacetin and [methyl-13C]methionine either without or with red wine consumption amounted to 38.2±6.3 vs. 36.3±6.7% (p=0.363) and 9.5±3.3 vs. 8.8±2.5% (p=0.47), respectively. Moderate alcohol consumption does not induce significant short-term changes of the microsomal and the mitochondrial functions of the human liver in healthy subjects.

Quantitative Assessment of Effective Energy of Resistant Cornstarch Using 13CO2, H2, and CH4 Breath Tests

We calculated the effective energy of a resistant cornstarch (HB-450) by using 13CO2, H2, and CH4 breath tests to assess HB-450 digestion and fermentation in healthy female subjects. On the basis of the areas under the curves (AUCs) for the concentrations of 13CO2 and H2 following ingestion of 40 g of HB-450, 23.0±6.8 g was digested in the upper digestive tract and 3.6±2.1 g was fermented by intestinal flora. The effective energy of the 40 g dose was thus calculated as 2.5±0.8 kcal/g. These results indicate that 13CO2, H2, and CH4 breath tests are useful for measuring the effective energy of resistant cornstarch.

A new oral formulation for the release of sodium butyrate in the ileo-cecal region and colon

To develop a new formulation with hydroxy propyl methyl cellulose and Shellac coating for extended and selective delivery of butyrate in the ileo-caecal region and colon. One-gram sodium butyrate coated tablets containing 13C-butyrate were orally administered to 12 healthy subjects and 12 Crohn's disease patients and the rate of 13C-butyrate absorption was evaluated by 13CO2 breath test analysis for eight hours. Tauroursodeoxycholic acid (500 mg) was co-administered as a biomarker of oro-ileal transit time to determine also the site of release and absorption of butyrate by the time of its serum maximum concentration. The coated formulation delayed the 13C-butyrate release by 2-3 h with respect to the uncoated tablets. Sodium butyrate was delivered in the intestine of all subjects and a more variable transit time was found in Crohn's disease patients than in healthy subjects. The variability of the peak 13CO2 in the kinetic release of butyrate was explained by the inter-subject variability in transit time. However, the coating chosen ensured an efficient release of the active compound even in patients with a short transit time. Simultaneous evaluation of breath 13CO2 and tauroursodeoxycholic acid concentration-time curves has shown that the new oral formulation consistently releases sodium butyrate in the ileo-cecal region and colon both in healthy subjects and Crohn's disease patients with variable intestinal transit time. This formulation may be of therapeutic value in inflammatory bowel disease patients due to the appropriate release of the active compound.

The dispersion state of milk fat influences triglyceride metabolism in the rat

Milk fat, which has different structures in the various dairy products, is a major and controversial lipid source in the Western diet. However, information about the digestion fate of milk fat depending on its supramolecular structure for a given composition is scarce. In this study, 13CO2 breath tests were performed with fasted rats force-fed different dairy preparations of similar composition but differing in fat suprastructure in order to highlight differences of general lipid metabolism. Each preparation consisted of a NaCl solution, anhydrous milk fat labelled with a 13C mixed triacylglycerol, casein (as native phosphocaseinate powder with some lactose), and dipalmitoylphosphatidylcholine. Milk fat was either fed (i) unemulsified consecutively to the aqueous phase, or emulsified as (ii) coarse droplets of approximately 10 microm covered mainly with the phospholipid, or (iii-iv) fine droplets of approximately 1 microm covered mainly with casein, force-fed either in the liquid state or in a semi-crystallized state. 13C abundance in expired air samples was measured by isotope ratio mass spectrometry; results were expressed as 13C enrichment and were submitted to an ANOVA analysis. The 13CO2 excretion curves of the unemulsified preparation and the coarse emulsion were similar and presented a sharp peak, both significantly different from the fine emulsion curves characterized by a nearly linear cumulative recovery. The crystalline state of the fine emulsion droplets and the viscosity of these emulsions did not affect significantly their excretion curves. The lipid metabolization (indicated by the 13C recovery) was significantly slower for the fine droplets coated with casein than for the large droplets coated with the phospholipid and the unemulsified fat. For the latter, a single 13C peak rapidly appeared, while for small droplets coated with caseins, 13C excretion was continuous up to 6 h. Global lipid metabolism based on oxidation to CO2 was decreased with smaller compared to larger emulsified milk fat particles with different coatings. These data support the concept that dairy products with different fat suprastructures are digested and metabolized differently.

The ion channel ASIC1 contributes to visceral but not cutaneous mechanoreceptor function

Visceral mechanoreceptors are critical for perceived sensations and autonomic reflex control of gastrointestinal function. However, the molecular mechanisms underlying visceral mechanosensation remain poorly defined. Degenerin/epithelial Na+ channel (DEG/ENaC) family ion channels are candidate mechanosensory molecules, and we hypothesized that they influence visceral mechanosensation. We examined the influence of the DEG/ENaC channel ASIC1 on gastrointestinal mechanosensory function, on gastric emptying, and on fecal output. We also compared its role in gastrointestinal and somatic sensory function. To assess the role of ASIC1 we studied wild-type and ASIC1-/- mice. Reverse-transcription polymerase chain reaction (RT-PCR) and Western blot analysis determined expression of ASIC1 messenger RNA and protein in vagal and spinal sensory ganglia. Colonic, gastroesophageal, and cutaneous afferent fibers were characterized by functional subtype and their mechanical stimulus-response relationships were determined. Gastric emptying was determined by using a 13CO2 breath test. Behavioral tests assessed somatic mechanical and thermal sensitivity. ASIC1 was expressed in sensory ganglia and was lost after disruption of the ASIC1 gene. Loss of ASIC1 increased mechanosensitivity in all colonic and gastroesophageal mechanoreceptor subtypes. In addition, ASIC1-/- mice showed almost double the gastric emptying time of wild-type mice. In contrast, loss of ASIC1 did not affect function in any of the 5 types of cutaneous mechanoreceptors, nor did it affect paw withdrawal responses or fecal output. ASIC1 influences visceral but not cutaneous mechanoreceptor function, suggesting that different mechanisms underlie mechanosensory function in gut and skin. The role of ASIC1 is highlighted by prolonging gastric emptying of a meal in ASIC1-/- animals.

Whole-Body .BETA.-Oxidation of Docosahexaenoic Acid Is Decreased under n-3 Polyunsaturated Fatty Acid Insufficiency as Assessed by a 13CO2 Breath Test Using [U-13C]Fatty Acids

We applied a continuous 13CO2 breath test to examination of the effects of n-3 polyunsaturated fatty acid (PUFA) insufficiency on fatty acid β-oxidation. Fischer rats were divided into two groups, and were reared on two different regimens of diet for multiple successive generations. One diet was adequate with respect to n-3 PUFA and the other was insufficient in n-3 PUFA. Rats (the F18 generation) were given [U-13C]docosahexaenoic acid or [U-13C]palmitic acid by intravenous injection, and expired 13CO2 was continuously measured using a mass spectrometer. Fatty acid oxidation was evaluated by two parameters, the half-life of 13CO2 in the breath and cumulative 13CO2 recovery from the labeled fatty acids. When [U-13C] docosahexaenoic acid was administered, cumulative 13CO2 recovery was 40% lower in the n-3 PUFA-insufficient diet group than in the n-3 PUFA-adequate diet group. The half-life appeared to be the same for the two diet groups. In the case of [U-13C]palmitic acid, neither the half-life nor cumulative 13CO2 recovery significantly differed between the two diet groups. These results suggest that overall β-oxidation of DHA in a whole-body system is decreased under n-3 PUFA insufficiency. Our finding shows that the 13CO2 breath test can be used to assess n-3 PUFA insufficiency. The test provides a useful and invasive method for evaluation of fatty acid β-oxidation.

Cooking resistant starch reduces its prebiotic properties: Assessment with the 13CO2 and H2 breath test

Cooking resistant starch reduces its prebiotic properties: Assessment with the 13CO2 and H2 breath test

Towards an Inhalative 13C Breath Test Method

Customary 13CO2 breath tests—and also 15N urine tests—always start with an oral administration of a test substrate. The test person swallows a stable isotope labelled diagnostic agent. This technique has been used to study several pathophysiological changes in gastrointestinal organs. However, to study pathophysiological changes of the bronchial and lung epithelium, the inhalative administration of a stable isotope labelled agent appeared more suitable to us. [1-13C]Hexadecanol and [1-13C]glucose were chosen. Inhaled [1-13C]hexadecanol did not yield 13CO2 in the exhaled air, but [1-13C]glucose did. To study the practicability of the [1-13C]glucose method and the reproducibility of the results, 18 inhalation tests were performed with healthy subjects. In 6 self-tests, the optimum inhalative dose of [13C]glucose was determined to be 205 mg. Using the APS aerosol provocation system with the nebulizer ‘Medic Aid’ (Erich Jaeger Würzburg), a 25% aqueous solution was inhaled. Then, breath samples were collected at 15 min. intervals and analysed for 13CO2. 75–120min after the end of inhalation a well-reproducible maximum δ13C value of 6‰ over baseline (DOB) was detected for 12 healthy probands. Speculating that the pulmonary resorption of the [13C]glucose is the rate-limiting step of elimination, decompensations in the epithelium ought to be reflected in changed [1-13C]glucose resorption rates and changed 13CO2 output. Therefore, we speculate that the inhalation of suitable 13C-labelled substrates will pave the way for a new group of 13CO2 breath tests aiding investigations of specific pathophysiological changes in the pulmonary tract, such as inflammations of certain sections and decompensations of cell functions.

Stable Isotopes and 13CO2 Breath Tests for Investigating Gastrointestinal Functions

In medical investigation there is a need for non-invasive methods. Moreover, patients ask for easy methods that are simple to perform and medical doctors demand reliable techniques. With the advent of stable isotopes a new area of tracer technology became available. In gastroenterology, 13CO2 breath tests are used which fulfill all the conditions needed in modern clinical research and investigation.

13C Basal Abundance of Expired CO2 -Definition of Pre-Requisites for Kinetic Breath Tests

A sufficiently stable rate of 13CO2 exhalation is necessary when the diagnostic 13CO2 breath tests are performed in healthy subjects and patients. The aim of the research was to define prerequisite conditions for kinetic breath tests in order to ensure a stable 13CO2 background. A 3-part protocol was developed. Part I: a study of the one-day variation of 13CO2 abundance in expired CO2 confirmed that shifts of the basal 13C abundance in breath are inherent in nature. Part II: a study of the variations of 13C enrichment after the ingestion of different meals and beverages showed that ingestion of food items containing C4 plant sugars, such as maize, induces a significant increase in isotopic abundance. Part III: a new test breakfast containing rice grain cereal, milk and orange juice was tested. This test meal induces no significant change on the basal 13CO2 abundance in healthy subjects. This new finding allows to avoid the fasting period normally required prior to a breath test which is sometimes difficult for children and pregnant women.

Simple Sampling Method and Routine Gas Chromatography-Mass Spectrometry Analysis in 13C Breath Tests

The 13C-labeled CO2 breath tests are convenient methods for various uses. They can be used to measure gastric emptying rate of solids (1), detect fat malabsorption (2), and evaluate pancreatic lipase activity (3), exocrine pancreatic insufficiency (4), glucose adsorption and utilization (5), liver function control (6), and lactase deficiency (7) with appropriate 13C isotope-labeled compounds. In recent years, the 13C-urea breath test has been used to detect Helicobacter pylori (8). With an appropriate analytical background and the facilities required for the detection or quantification of labeled CO2, the 13CO2 breath test could be widely applied for diagnostic purposes in the medical practice. Although several infrared or mass spectrometric methods (9) are available, the transport of large volumes of gaseous samples to analytical centers as well as storage before measurement involves great problems. Therefore, we have developed a simple method that allows storage of these samples in minimal liquid volume. By this technique, the liberation of isotope-labeled CO2-containing gas occurs immediately before measurements. Because most medical centers are not equipped with expensive and complex isotope-ratio mass spectrometers but generally have a routine low-mass-resolution gas chromatography-mass spectrometry (GC-MS) apparatus, we have also used a simple detection method for 13CO2 by standard GC-MS techniques. Analysis requires only a few minutes, and the method is suitable for selection of 13CO2-containing samples. In our test experiments, we used an air …

Variations in Colonic H2 and CO2 Production As a Cause of Inadequate Diagnosis of Carbohydrate Maldigestion in Breath Tests

Background: Lactose maldigestion is usually diagnosed by means of the H2 breath test. When 13C-lactose is used as substrate, a 13CO2 breath test can be performed simultaneously. In an earlier publication we described the relation between both the H2 and 13CO2 exhalation in breath and the measured intestinal lactase activity after consumption of 13C-lactose. We found a discrepancy between both breath test results in 36% of the cases. To investigate the possible cause of these incongruous breath test results, we studied gas production from carbohydrate in the colon, using 13C-lactulose as a non-absorbable substrate. Methods: Experiments were performed in 21 subjects, by applying 5 different doses of 13C-lactulose. Repeatability studies were performed in six of these subjects, using 10 g substrate (three tests with 1-week intervals). Results: Both the H2 and the 13CO2 excretion in breath varied strongly interindividually and intraindividually after consumption of 13C-lactulose. In both cases no dose-response relation was observed. A significant positive linear relationship was found between H2 and 13CO2 exhalation (r = 0.45, P < 0.005). Extrapolation of these results to 13C-lactose breath tests indicates that the colonic contribution of 13CO2 production to the total 13CO2 excretion in breath varies but is on the average large enough to cause false-negative 13CO2 breath test results. Conclusions: Excretion in breath of 13CO2 produced in the colon during a 13C-lactulose breath test correlates with the breath H2 excretion. This could explain the occurrence of false-negative 13CO2 lactose breath tests when colonic gas production is high and false-negative lactose H2 breath test results when gas production is low. It can also explain the improved sensitivity of the combined H2/13CO2 lactose breath test compared with both breath tests alone.