592 Gut, 1992, 33, 592-596 Effect ofHelicobacterpylori infection on colloidal bismuth subcitrate concentration in gastric mucus Gut: first published as 10.1136/gut.33.5.592 on 1 May 1992. Downloaded from D J B Mufnoz, C Tasman-Jones, J Pybus Abstract bacterpylori. Movement ofa drug into mucus will Necropsy gastric mucus infected with Helico- be determined by its intragastric availability. bacter pylon has a reduced capacity to con- Availability will depend upon various factors centrate colloidal bismuth subcitrate when including the drug's behaviour in solution. compared with non-infected mucus. Mucus It is not known whether non-infected and mounted in a modified in vitro diffusion Helicobacter pylon infected mucus differ in their chamber was bathed with colloidal bismuth capacity to contain colloidal bismuth subcitrate subcitrate solutions at different concentra- and little is known on the behaviour of colloidal tions and pH levels. Bismuth was measured by bismuth subcitrate in solution. atomic absorption spectrophotometry to In this in vitro study we have examined the assess intramucus colloidal bismuth subcitrate effect of Helicobacter pylon' infection on the concentrations. Bismuth concentrations in capacity ofgastric mucus to concentrate colloidal non-infected mucus were higher than in bismuth subcitrate and the behaviour ofcolloidal Helicobacter pylori infected mucus at all bismuth subcitrate in acidic solutions. experimental colloidal bismuth subcitrate con- centrations and pH levels. Regardless of the infection status, the intramucus concentration Methods of colloidal bismuth subcitrate was dependent upon the concentration of the bathing solution SPECIMENS and independent of the pH and the mucus Mucus from 20 human stomachs was collected at thickness. Colloidal bismuth subcitrate solu- necropsy within 24 hours of death. All persons bility in saline solution varied with pH. (18-75 years) had died ofacute trauma or sudden http://gut.bmj.com/ and was least soluble in the pH range 1 1 to unexpected medical collapse. Each stomach was 3-25 and more soluble above and below this pH opened along the greater curvature and food range. This study suggests that Helicobacter contents were removed by washing with normal pylon infection is associated with physico- saline. chemical changes in the gastric mucus with a Mucus was harvested by gently scraping the reduction in its capacity to concentrate col- gastric mucosa with a glass slide and stored at loidal bismuth subcitrate. Such a reduction -20°C for later experiments. on September 23, 2021 by guest. Protected copyright. may compromise the attainment of optimum Immediately before harvesting, a gastric tissue colloidal bismuth subcitrate concentrations specimen was taken for Helicobacterpylori urease necessary for its bactericidal activity. detection by the camplobacter like organism test (CLO test, Delta West Limited, Western Aus- tralia).34 Gastric mucus plays an important role in A mucus sample was considered infected with mucosal protection.' Disruption in the Helicobacter pylori when the tissue specimen integrity of the blanket of mucus has been found showed positive urease activity within one hour in individuals with peptic ulcer.8"12 Helicobacter with the CLO test and non-infected when the pylori present in gastric mucus of the stomach tissue specimen showed no positive urease or duodenal gastric metaplasia is associated with activity within 24 hours. gastritis and peptic ulceration.'3 22 Helicobacter Department of pylori resides in gastric mucus but not within Medicine, School of epithelial cells.2"24 It may play a role in mucin COLLOIDAL BISMUTH SUBCITRATE Medicine, University of Auckland, Auckland, breakdown.25-27 CONCENTRATION IN GASTRIC MUCUS New Zealand Eradication of Helicobacter pylori by colloidal The modified in vitro diffusion chamber pre- D J B Munioz bismuth subcitrate is followed by histological viously used for ion exchange studies on mucus` C Tasman-Jones improvement of gastritis and healing of peptic was used to expose gastric mucus to colloidal Department of Clinical ulcer.242829 If infection recurs, ulcer relapse is bismuth subcitrate. Chemistry, Auckland frequent, particularly after one year.2430 31 Mucus (115 + 1 mg or 230 ± 1 mg, depending Hospital, Auckland, New A combination of colloidal bismuth subcitrate on mucus layer thickness) was held between two Zealand with one or two antibiotics - fQr fine gauze mesh discs Swiss J Pybus example, (Nybolt PA-22/17; is more Correspondence to: amoxycillin and/or metronidazole, Silk Bolting Cloth Mfg Co Ltd, Zurich, Switzer- Dr C Tasman-Jones, successful than monotherapy in eradicating land) separated by a plastic ring to form a layer Department of Medicine, School of Medicine, Helicobacterpylor 24323 400 ,im or 800 iim thick. The mucus containing University of Auckland, Ulcer relapse after colloidal bismuth sub- cassette was held in the chamber by two rigid Private Bag, Auckland, New Zealand citrate and other antibiotics may be the result mesh discs (Tetex Mono PES 130/TWL/C; Accepted for publication of a failure of the drug(s) to reach sufficient Swiss Silk Bolting Cloth Mfg Co Ltd, Zurich, 5 August 1991 intramucus concentrations to eradicate Helico- Switzerland). One side of the mucus was bathed Effect ofHelicobacter pylori infection on colloidal bismuth subcitrate concentration in gastric mucus 593 with saline solution adjusted to pH 6.0 and the different pH's. For each pH, one 60 ml test tube other side with filtered solutions (Filter Paper 1, containing 45 ml saline and one whole commer- Whatman International Ltd, Maidstone, cial tablet of colloidal bismuth subcitrate was England) of colloidal bismuth subcitrate (De- 'tumbled' in the Chiltern rotatory mixer (Chiltern Gut: first published as 10.1136/gut.33.5.592 on 1 May 1992. Downloaded from Nol®, Gist-Brocades, Delft, The Netherlands) Scientific, Germany) at a rate of 32 'tumbles' per at different concentrations (as bismuth) and minute. Disintegration was assessed visually and pH's as follows: halfand total disintegration times were recorded. Series 1: Colloidal bismuth subcitrate 93 To determine if the outer coating of the com- mg/l, pH 6.0, mucus layer 800 [im thick. mercial tablet affects disintegration, tablets were Series 2: Colloidal bismuth subcitrate 232 stripped of their coating and exposed to saline mg/l, pH 6-0, mucus layer 400 ,um thick. solutions under the same conditions as above. Series 3: Colloidal bismuth subcitrate 64 mg/l, pH 6.0, mucus layer 400 iim thick. Series 4: Colloidal bismuth subcitrate 64 STATISTICAL ANALYSIS mg/l, pH 1 5, mucus layer 400 [tm thick. Results for colloidal bismuth subcitrate concen- Series 5: Colloidal bismuth subcitrate 127 tration experiments were expressed as mean mg/l, pH 1 5, mucus layer 400 [im thick. (SD). A general linear approach to the analysis of The solutions were continuously circulated for variance37 was used to determine the statistical 90 minutes at a rate of 4 5 ml/min, room significance of differences in bismuth concen- temperature 2 1C. tration between non-infected and Helicobacter After each experiment the excess of colloidal pylori infected mucus. Significant mean and bismuth subcitrate solution was washed off with interaction effects were further investigated saline. The mucus was taken out of the chamber using Tukey's procedure to analyse differences and dried at 1 10°C for four hours. within pH's and mucus thickness. Type III sums The bismuth content of the samples was of squares are presented because the design was measured by atomic absorption (Graphite unbalanced. p Values of less than 0.05 were Furnace Atomic Absorption Spectrophotometer considered significant. SpectrAA-20; Melbourne, Australia) to assess the intramucus colloidal bismuth subcitrate concentrations. Results were expressed as [tg Results bismuth/g mucus on the assumption that all mucus samples contain 95% water.36 The possi- COLLOIDAL BISMUTH SUBCITRATE bility that Helicobacter pylon infection may CONCENTRATION IN GASTRIC MUCUS modify the water content has not been con- The Table shows mean (SD) and range bismuth http://gut.bmj.com/ sidered in this study. concentrations in non-infected and Helicobacter pylon infected gastric mucus. Bismuth concentrations were higher in non- COLLOIDAL BISMUTH SUBCITRATE SOLUBILITY infected than in Helicobacter pylon infected Normal saline solutions were adjusted to dif- mucus at all experimental colloidal bismuth ferent pH's (E512 Metrohm Herisau pH meter, subcitrate concentrations and pH levels (F(1,62)= Switzerland). Commercial tablets of colloidal 52.86, p=00001). on September 23, 2021 by guest. Protected copyright. bismuth subcitrate stripped of their outer coat- Regardless of the infection status, bismuth ing were ground to a homogeneous powder. concentrations varied in proportion to the con- Weighed samples of the powder were added to centration of the bathing solution and were no saline to give mixtures with the same weight/ different at mucus thickness 400 iim or 800 ,tm volume ratio. After colloidal bismuth subcitrate and at pH 1.5 or 6.0 (F(s,62)=l106, p=0.0389). was dissolved in the saline solution, samples In separate experiments, using unfiltered were obtained from the supernatant and colloidal bismuth subcitrate solutions and no fine analysed for bismuth content by atomic absorp- gauze mesh we found that a whitish precipitate tion spectrophotometry. Colloidal bismuth sub- was deposited on the mucus surface after the citrate solubility was estimated by calculating the experiments,