Albert Tangerman

Measurement and biological significance of the volatile sulfur compounds hydrogen sulfide, methanethiol and dimethyl sulfide in various biological matrices.

This review deals with the measurement of the volatile sulfur compounds hydrogen sulfide, methanethiol and dimethyl sulfide in various biological matrices of rats and humans (blood, serum, tissues, urine, breath, feces and flatus). Hydrogen sulfide and methanethiol both contain the active thiol (-SH) group and appear in the free gaseous form, in the acid-labile form and in the dithiothreitol-labile form. Dimethyl sulfide is a neutral molecule and exists only in the free form. The foul odor of these sulfur volatiles is a striking characteristic and plays a major role in bad breath, feces and flatus. Because sulfur is a biologically active element, the biological significance of the sulfur volatiles are also highlighted. Despite its highly toxic properties, hydrogen sulfide has been lately recommended to become the third gasotransmitter, next to nitric oxide and carbon monoxide, based on high concentration found in healthy tissues, such as blood and brain. However, there is much doubt about the reliability of the assay methods used. Many artifacts in the sulfide assays exist. The methods to detect the various forms of hydrogen sulfide are critically reviewed and compared with findings of our group. Recent findings that free gaseous hydrogen sulfide is absent in whole blood urged the need to revisit its role as a blood-borne signaling molecule.

Extra-oral halitosis: an overview

Halitosis can be subdivided into intra-oral and extra-oral halitosis, depending on the place where it originates. Most reports now agree that the most frequent sources of halitosis exist within the oral cavity and include bacterial reservoirs such as the dorsum of the tongue, saliva and periodontal pockets, where anaerobic bacteria degrade sulfur-containing amino acids to produce the foul smelling volatile sulfur compounds (VSCs), especially hydrogen sulfide (H(2)S) and methyl mercaptan (CH(3)SH). Tongue coating is considered to be the most important source of VSCs. Oral malodor can now be treated effectively. Special attention in this overview is given to extra-oral halitosis. Extra-oral halitosis can be subdivided into non-blood-borne halitosis, such as halitosis from the upper respiratory tract including the nose and from the lower respiratory tract, and blood-borne halitosis. The majority of patients with extra-oral halitosis have blood-borne halitosis. Blood-borne halitosis is also frequently caused by odorous VSCs, in particular dimethyl sulfide (CH3SCH3). Extra-oral halitosis, covering about 5-10% of all cases of halitosis, might be a manifestation of a serious disease for which treatment is much more complicated than for intra-oral halitosis. It is therefore of utmost importance to differentiate between intra-oral and extra-oral halitosis. Differences between intra-oral and extra-oral halitosis are discussed extensively. The importance of applying odor characterization of various odorants in halitosis research is also highlighted in this article. The use of the odor index, odor threshold values and simulation of bad breath samples is explained.

Determination of volatile sulphur compounds in air at the parts per trillion level by tenax trapping and gas chromatography

A gas chromatographic technique is described for measuring various volatile sulphur compounds (H2S, COS, CS2, thiols, sulphides, disulphides) in air at the parts per trillion level. The sulphur compounds are trapped and concentrated at -196 degrees C in a small glass tube filled with the porous polymer Tenax GC. The excess of water in the air samples is pretrapped by calcium chloride, which drying agent does not adsorb any of the sulphur volatiles. The Tenax trap tube fits exactly in the injection port of the gas chromatograph, where the adsorbed sulphur compounds are liberated at 200 degrees C directly into the carrier gas stream and are transferred to the gas chromatographic column. The sulphur compounds are then assayed by means of a flame photometric detector. The Tenax trap tubes can be stored at -196 degrees C for more than 1 week without any loss of sulphur volatiles.

A new sensitive assay for measuring volatile sulphur compounds in human breath by Tenax trapping and gas chromatography and its application in liver cirrhosis

A new analytical technique is described for measuring volatile sulphur compounds in human breath. The sulphur compounds were trapped and concentrated onto Tenax GC and then assayed by gas chromatography, using a specific sulphur detector. The detection limit amounts to about 0.2 ng/l (0.1 ppb). Among the sulphur volatiles, dimethylsulphide and methanethiol were quantitatively analysed in 100 ml of breath of 20 normal subjects and 35 cirrhotic patients. Dimethylsulphide in the breath of cirrhotics (113.4 +/- 31.9 ng/l, mean +/- SEM) was significantly elevated (p less than 0.05) compared with normals (21.1 +/- 1.7 ng/l). The concentration of methanethiol in the breath of normals and of most cirrhotics was less than 1 ng/l. In only five cirrhotics could methanethiol be detected in 100 ml of breath (3-23 ng/l). Dimethyldisulphide and hydrogen sulphide were not present in detectable amounts in the breath of normals. In cirrhotics, dimethyldisulphide was detected in a few cases. Ethanethiol was absent in the breath of both normals and cirrhotics.

Analysis of conjugated and unconjugated bile acids in serum and jejunal fluid of normal subjects

A reliable method is described for the determination of conjugated and unconjugated bile acids in serum and jejunal fluid. Bile acids are extracted using reverse-phase octadecylsilane bonded silica cartridges and are separated into their unconjugated and conjugated fractions using the lipophilic anion exchanger diethylaminohydroxypropyl Sephadex LH-20 (DEAP-LH-20). The conjugated fraction can be separated into a glycine and a taurine fraction, using the same anion exchanger. The bile acids are measured using a hydroxysteroid dehydrogenase-fluorimetric assay for serum and a hydroxysteroid dehydrogenase-photometric assay for jejunal fluid. The normal fasting serum value of total 3 alpha-hydroxy bile acids amounts to 3.5 +/- 2.8 mumol/l (mean +/- SD, range 1.4-10.8, n = 22). The corresponding unconjugated bile acid fraction amounts to 39.9 +/- 11.2% (range 20.7-64.6%) of total bile acids. The concentration of conjugated bile acids became significantly elevated 30, and 60 min after a standard meal, whereas that of unconjugated bile acids remained unchanged. In jejunal fluid only conjugated bile acids are found, as well in fasting subjects as postprandial, 30 or 60 min after a standard meal.

Effect of bile salt binding or protease inactivation on plasma cholecystokinin and gallbladder responses to bombesin.

BACKGROUND/AIMS Bombesin-stimulated plasma cholecystokinin levels decrease after an initial increase despite continuous infusion of bombesin. The aim of this study was to determine if a feedback mechanism, mediated by bile salts or proteolytic enzymes, is responsible for this decline. METHODS Bombesin (1.0 ng.kg-1.min-1) was infused into volunteers for 180 minutes on separate occasions. Cholestyramine, colestipol, camostate, or saline were perfused intraduodenally during the second hour of the tests. Cholestyramine was also administered without infusion of bombesin. RESULTS Colestipol and cholestyramine, dependent on their bile salt-binding capacity, markedly enhanced (P < 0.05) bombesin-stimulated plasma cholecystokinin from 2.1 +/- 0.5 pmol/L to 6.4 +/- 2.2 pmol/L and 12.1 +/- 3.3 pmol/L (P < 0.05 vs. colestipol), respectively, and further decreased gallbladder volume (P < 0.05) from 9.4 +/- 1.6 mL to 2.0 +/- 0.4 mL and 2.2 +/- 0.5 mL, respectively. The protease inhibitor camostate had no effect. Bile salt precipitation also enhanced plasma pancreatic polypeptide responses (P < 0.01) but did not alter gastrin responses. Plasma cholecystokinin responses to cholestyramine without bombesin infusion varied considerably, but increments were highly correlated to decreases in gallbladder volume (r = 0.91; P < 0.005). CONCLUSIONS Bile salt sequestration but not protease inactivation enhances plasma cholecystokinin and gallbladder responses to bombesin infusion in humans.

Unconjugated serum bile acids as a marker of small intestinal bacterial overgrowth

Abstract. Non‐invasive methods to detect small intestinal bacterial overgrowth often lack specificity in patients who have undergone an ileal resection or have an accelerated intestinal transit. Since elevated serum unconjugated bile acid levels have been found in patients with clinical signs of bacterial overgrowth, we studied the clinical value of unconjugated serum bile acids as a marker of small intestinal bacterial overgrowth. Patients with culture‐proven bacterial overgrowth had significantly elevated fasting unconjugated serum bile acid levels (median and range: 4.5; 1.4–21.5 μmol l‐1) as compared to healthy subjects (0.9; 0.3–1.7 μmol l‐1, P < 0.005), to persons with an accelerated intestinal transit (1.0; 0.3–1.9 μmol l‐1, P < 0.005) and to persons who have undergone an ileal resection (2.1; 0.7–3.6 μmol l‐1P < 0.005). The same was true 30 and 60 min after ingestion of a Lundh meal. Serum unconjugated bile acid levels above 4 μmol l‐1 were found in eight of 10 patients with culture‐proven small intestinal bacterial overgrowth whereas serum levels above 4 μmol l‐1 were found in none of the patients from the three control groups. These results suggest that determination of unconjugated serum bile acids is of clinical value in the evaluation of patients suspected of small intestine bacterial overgrowth.

Halitosis and Helicobacter pylori infection

There is disagreement about a possible relationship between Helicobacter pylori (H. pylori) infection and objective halitosis, as established by volatile sulfur compounds (VSCs) in the breath. Many studies related to H. pylori used self-reported halitosis, a subjective and unreliable method to detect halitosis. In this study a possible relation between H. pylori and halitosis was evaluated, using an objective method (gas chromatography, GC) to detect the VSCs, responsible for the halitosis. The levels of the VSCs hydrogen sulfide (H(2)S), methyl mercaptan (MM) and dimethyl sulfide (DMS) were measured in mouth breath and in stomach air of 11 H. pylori positive patients and of 38 H. pylori negative patients, all with gastric pathology. Halitosis was also established by organoleptic scoring (OLS) of mouth-breath. The levels of H(2)S, MM and DMS in the mouth-breath and stomach air of the H. pylori positive patients did not differ significantly from those of the H. pylori negative patients. OLS of the mouth-breath resulted in 9 patients with halitosis, 1 out of the H. pylori positive group and 8 out of the H. pylori negative group, which is not statistically different. The concentrations of the VSCs in stomach air were in nearly all cases below the thresholds of objectionability of the various VSCs, indicating that halitosis does not originate in the stomach. The patients with gastric pathology were also compared with control patients without gastric pathology and with normal volunteers. No significant differences in VSCs in mouth breath were observed between these groups. Thus, in this study no association between halitosis and H. pylori infection was found. Halitosis, as established by GC and OLS, nearly always originates within the oral cavity and seldom or never within the stomach.

Comparison of different treatment modalities for oral halitosis

Abstract Objectives. To assess the effects on intra-oral halitosis by a mouth rinse containing zinc acetate (0.3%) and chlorhexidine diacetate (0.025%) with and without adjunct tongue scraping. Materials and methods. Twenty-one subjects without a diagnosis of periodontitis were randomized in a cross-over clinical trial. Organoleptic scores (OLS) were assessed to define intra-oral halitosis by total volatile sulfur compound (T-VSC) measurements and by gas chromatography. Results. Twenty-one subjects with a mean age of 45.7 years (SD: ±13.3, range: 21–66). The OLS were significantly lower following active rinse combined with tongue scraping (p < 0.001) at all time points. Immediately after, at 30 min, and at day 14, the T-VSC values were lower in the active rinse sequence than in the negative rinse sequence (p < 0.001, p < 0.001 and p < 0.05, respectively). At 30 min and at day 14, the hydrogen sulfide (H2S) and methyl mercaptan (MM) values were lower in the active rinse sequence compared to the inactive rinse sequence (p < 0.001). The inactive rinse sequence with tongue scraping reduced T-VSC at 30 min (p < 0.001) but not at 14 days. Similar reductions in T-VSC, H2S and MM were found in the active rinse sequence with or without tongue scraping. Conclusion. The use of a tongue scraper did not provide additional benefits to the active mouth rinse, but reduced OLS and tongue coating index.

Volatile sulphur compounds in morning breath of human volunteers

OBJECTIVE morning breath contains elevated concentrations of volatile sulphur components (VSCs). Therefore, morning breath is recognised as a surrogate target for interventions on breath quality. Nevertheless, factors influencing morning breath are poorly understood. Our aim was to evaluate concentrations of VSC at the time of awakening. METHODS a procedure was developed to collect breath samples at home. Intra- and inter-person variations were determined in two small studies based on measurements of hydrogen sulphide, methyl mercaptan and dimethyl sulphide in healthy volunteers. RESULTS highest levels of VSC were found directly after waking up, followed by a significant decline afterward. Considerable day-to-day variation was found, but could not be linked to dietary intake. A significantly higher concentration of H(2)S and CH(3)SH was observed in the group of female subjects compared to males. CONCLUSIONS when morning breath is used as a target for interventions, breath collected at the time of or shortly after waking up is preferred over breath collected later during the morning. Gender plays an important role in VSC levels, and should be taken into account.