A variety of drugs that bind to iron have significant reductions in absorption when coadministered with iron compounds. Cimetidine has a structure that would ...
Digestive Diseases and Sciences, Vol. 38, No. 5 (May 1993), pp. 950-954
Ferrous Sulfate Reduces Cimetidine Absorption N O R M A N R.C. C A M P B E L L , MD, B R I A N B. H A S I N O F F , PhD, J O N A T H A N B. M E D D I N G S , MD, W I L L I A M D. A N D E R S O N , S U S A N R O B E R T S O N , and K A R L G R A N B E R G
A variety o f drugs that bind to iron have significant reductions in absorption when coadministered with iron compounds. Cimetidine has a structure that would suggest strong binding to iron ions. In vitro experiments were performed to examine a variety o f characteristics o f the binding o f iron to cimetidine. Further studies were conducted to determine the effect of concurrent administration o f ferrous sulfate on cimetidine absorption in an in vivo isolated perfused rat jejunal model o f drug absorption. The dose o f cimetidine was chosen to represent a human dose o f 300 rag, while the ferrous sulfate doses were chosen to represent 150- and 300-mg doses. The higher ferrous sulfate dose completely inhibited cimetidine absorption (P < 0.01), while the lower dose o f ferrous sulfate caused a 63% reduction in cimetidine absorption (P < O.05). In vitro iron in its ferrous form rapidly oxidizes to the ferric form. The ferric form o f iron binds to cimetidine and may be the cause o f the decreased cimetidine absorption. Care should be taken in prescribing iron supplements with cimetidine. KEY WORDS: drug interaction; drug absorption; iron; histamine-2 blockers; complex formation; peptic ulcer disease.
Cimetidine, a histamine-2 receptor antagonist is one of the most frequently prescribed drugs (1). The structure of cimetidine suggests strong binding to iron ions. Several drugs that bind to iron have reductions in absorption and clinical effectiveness when concurrently ingested with iron supplements (2-12). Iron replacement therapy is relatively common and is often necessary in peptic ulcer disease when bleeding has occurred. As cimetidine is also indicated in peptic ulcer disease, concurrent therapy will occur frequently. This study was conducted to examine the characteristics of cimetidine binding to the ferrous and ferric forms of iron and to Manuscript received January 16, 1992; accepted July 15, 1992. From the Cardiovascular Research Group, Gastrointestinal Research Group, Faculty of Medicine, The University of Calgary, Calgary, Alberta, Canada; and Faculty of Pharmacy, University of Manitoba, Winnipeg, Manitoba, Canada. Address for reprint requests: Dr. N. Campbell, Department of Medicine, Faculty of Medicine, The University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta, Canada, T2N 4N1.
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determine the effect of this binding on the absorption of cimetidine in the rat gastrointestinal tract.
MATERIALS AND METHODS
Cimetidine absorption was assessed using a standard rat model of drug absorption (13). The experiments were approved by the Animal Care Committee of the University of Calgary. Nine male Sprague-Dawley rats weighing 232-469 g were used in the control group and in each of the experimental groups. The rats were anesthetized using intraperitoneal urethane at approximately 1.25 g/kg. Through a midline incision immediately distal to the ligament of Treitz, a jejunal segment approximately 25 cm in length was isolated with sutures. Catheters were inserted into the segment proximally and distally and tied in place. The segments were perfused by 20 ml of recycling buffer at 37~ The initial buffer consisted of 280 mM sodium chloride and 20 mM Bis Tris propane pH 6.5. After 30 min, another 20 ml of the buffer was peffused, which contained cimetidine 4.24 mM, trace amounts of [3H]cimetidine, trace amounts of [14C]polyethylene glycol 4000 with oi without ferrous sulfate. The doses of ferrous Digestive Diseases and Sciences, Vol. 38, No. 5 (May 1993)
0163-2116/93/0500-0950507.00/09 1993PlenumPublishingCorporation
FERROUS SULFATE AND CIMETIDINE ABSORPTION sulfate were 6.73 mM and 3.36 mM and were chosen to approximate a 300-mg and a 150-mg human dose on a milligram per kilogram basis for the rat assuming the total dose was in the bowel at all times. The dose of cimetidine in the rat was chosen to represent a human dose of 300 mg. Samples of luminal contents were collected at 10-min intervals for 1 hr and analyzed in triplicate in a scintillation counter. Cimetidine concentrations were calculated by multiplying the original cimetidine concentration by the dpmff dpmo of 3H and were corrected for water absorption and secretion by multiplying the cimetidine concentration by dpm0/dpmx of [14C]polyethylene glycol (dpmo is the disintegrations per minute of 3H or 14C at the start of the experiment and dprn~ is the disintegrations per minute at a given timex). Absorption rates were calculated from the slope of the linear regression line derived from the cimetidine concentration versus time data between 10 and 60 min, were corrected for length of the bowel, and are expressed in concentration (nanomolar) per centimeter per minute. The stability of cimetidine was assessed prior to perfusion and following perfusion in bowel segments from two animals in each group by high-pressure liquid chromatography (HPLC). The HPLC system consisted of a Bio-Rad HPLC pump (model 1330), and a Bio-Rad Biosil ODS 10 reverse-phase column heated to 35~ The mobile phase consisted of 400 ml 50 mM potassium phosphate pH 5.6 buffer and 600 ml methanol per liter of mobile phase. One-minute fractions of HPLC solvent were collected and analyzed by scintillation counting. Cimetidine eluted in 3- to 7-min fractions with the mobile phase run at 1.0 ml/min. In order to characterize the binding of cimetidine to iron, a number of spectrophotometric experiments were carried out on a Shimadzu UV 260 double-beam spectrophotometer with its cell compartment maintained at 25~ Experiments were carried out initially in 50 mM Bis Tris propane pH 6.5 and later in methanol to avoid the formation of interfering ferric hydroxide complexes. The stoichiometry of the iron-cimetidine complex was determined by adding increasing concentrations of cimetidine to the ferric form of iron. All data are expressed as mean -+ standard deviation unless stated otherwise. The differences in cimetidine absorption with and without ferrous sulfate were analyzed using ANOVA. Further statistical analysis was performed to examine changes in cimetidine absorption between the control rats and experimental rats at each ferrous sulfate concentration by independent two-tailed Student's t tests. The urethane, Bis Tris propane, Bis Tris, ferrous sulfate (for in vivo studies), and cimetidine were supplied by the Sigma Chemical Company (St. Louis, Missouri). The [14C]polyethylene glycol 4000 was supplied by New England Nuclear Products (Mississauga, Canada) and the [3H]cimetidine was purchased from Amersham Canada Limited (Oakville, Canada). For chemical studies the anhydrous ferric chloride was obtained from BDH Inc. (Toronto, Canada), FeC13 9 6HzO from J.T. Baker Chemical Company (Phillipsburg, New Jersey), and ferrous Digestive Diseases and Sciences, Vol. 38, No. 5 (May 1993)
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Fig 1. The rates of cimetidine absorption (X -+ SEM) in the absence and in the presence of two concentrations of ferrous sulfate in isolated perfused rat jejunal loops. Both ferrous sulfate concentrations resulted in significant reductions in cimetidine absorption.
sulfate from Fisher Scientific Company (Fair Lawn, New Jersey). RESULTS The average rates of cimetidine absorption in the p r e s e n c e and absence of ferrous sulfate can be seen in Figure 1. Cimetidine absorption w a s reduced ( A N O V A , P < 0.05) 63% b y 3.36 m M ferrous sulfate (5.3 ___ 3.4 v s 2.0 __. 2.5, P < 0.05) and completely inhibited b y 6.73 m M ferrous sulfate (5.3 ___ 3.4 v s 0.0 ___ 2.8, P < 0.01). T h e stability of cimetidine w a s a s s e s s e d b y H P L C . Prior to perfusion 91.5 ___ 3.5% of the injected radioactivity eluted in a single p e a k in the fractions collected b e t w e e n 3 and 7 min and following perfusion without an animal for 1 hr, 90% eluted in these fractions. In two animals perfused with buffer containing cimetidine but no iron, 85.2% of the injected radioactivity eluted in the 3- to 7-min fractions at the end of the 1-hr perfusion. In two animals perfused with 1.84 m M ferrous sulfate as well as cimetidine 83.4% of the injected radioactivity eluted in the 3- to 7-min peaks, while in the animals p e r f u s e d with 3.67 m M ferrous sulfate 83.3% of the radioactivity eluted in the 3- to 7-min fractions. S p e c t r o p h o t o m e t r i c e x p e r i m e n t s in which FeCI 3 and cimetidine w e r e p r e m i x e d together at p H 6.5 or initially under acidic conditions and added to p H 6.5 Bis Tris buffer did not exhibit a n y significant spectral b a n d s in the visible region different than those seen with FeC13 alone. The ferric ion f o r m s strongly absorbing ferric hydroxide c o m p l e x e s . In order to
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tool e i m e t i d i n e / m o l Fe a+ 3. Spectrophotometric e ctl ,hotometric titration of 0.51 mM FeC13 by cimetidine le at 366 ~6 1nm carried out in methanol solution. soh The linear least squares .ar calculated ~lcl ted intersection of the two sl straight line portions is at aI mole r : ra ratio of cimetidine to Fe 3+ of 1.9 1. consistent with the formation of a Fe3+(cimetidine)2 complex. Fig
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prevent this interfering ferric hydroxide formation, complex formation was looked for in methanol. A solution containing anhydrous FeC13 in methanol was titrated with cimetidine in methanol, which resulted in the spectral changes shown in Figure 2 (bottom). A new peak at 305 nm is seen due to the formation of a Fea+-cimetidine complex. Upon the addition of larger amounts of cimetidine, the isosbestic point originally at 335 nm is gradually lost, indicating that at low cimetidine-to-iron ratios there are only two absorbing species in solution, whereas at higher ratios there are more than two absorbing species present. This result is consistent with the formation of higher cimetidine complexes at higher mole ratios of cimetidine to Fe 3+. The difference spectrum shown in Figure 2 (top) was obtained by incubating FeSO 4 and cimetidine together for 3.5 hr in aqueous pH 6.5 Bis Tris buffer at 25~ to allow the ferrous ion to air oxidize to
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ferric iron and then spectrally subtracting controls containing FeSO 4 alone and cimetidine alone. The peak found at 305 nm corresponds closely to the peak found for the Fe3+-cimetidine complex in methanol. This result indicates that Fe3+-cimeti dine complex formation occurs after mixing FeSO 4 and cimetidine at physiological pH. In order to characterize the stoichiometry of the Fe3+--cimetidine complex, the titration of FeCI 3 with cimetidine was carried out in methanol (Figure 3). The two linear portions give a sharp intersection at a mole ratio of 1.91 ___ 0.03 mol cimetidine per mole of Fe 3§ This indicates a complex of the form of Fe3+(cimetidine)2 is formed. Since Fe 3§ usually forms six coordinate complexes, it is likely cimetidine is acting as a tridentate ligand. Both imidazole and thioether groups would be expected to strongly coordinate Fe 3+. One of the secondary amine groups on cimetidine could also function as a coordinating site as well. DISCUSSION Ferrous sulfate reduces the absorption of cimetidine in a dose-dependent manner in the rat. Ingestion of cimetidine with ferrous sulfate could have deleterious effects on the treated patients. The efficacy of cimetidine is related to systemic bioavailability; therefore, if cimetidine absorption is decreased, acid secretion will increase. The increased Digestive Diseases and Sciences, Vol. 38, No. 5 (May 1993)
FERROUS SULFATE AND CIMETIDINE ABSORPTION gastric acidity could result in continued dyspeptic symptoms, slower healing, and continued blood loss. Cimetidine and other agents that increase gas: tric pH reduce iron absorption (14-16). Continued blood loss and reduced iron absorption may lead to a poor response to iron therapy. Human studies are required to determine the significance of these observations. However, care should be taken in the prescription of cimetidine with concurrent ferrous sulfate therapy. Peptic ulcer disease resulting in gastric or duodenal ulcers is very common in the Western world, affecting about one in 10 persons (17), and cimetidine is one of the most frequently prescribed medications for peptic ulcer disease. Iron deficiency occurs commonly in peptic ulcer disease due to blood loss and is generally treated with oral ferrous sulfate (18). This suggests that concurrent therapy with ferrous sulfate and cimetidine will occur often and the iron-cimetidine interaction could be common. The mechanism by which ferrous sulfate reduces cimetidine absorption may be due to the formation of iron-cimetidine complexes. The ferrous (2+) form of iron rapidly oxidizes to the ferric form (3 +) in vitro under pH conditions similar to those in the small intestine (4-6). For many compounds, the binding of ferric iron is much stronger than that of the ferrous form (7). The structure of cimetidine suggests strong binding to iron. The reaction below is suggested for the binding of cimetidine to iron in methanol. Fe 2+ ~ Fe 3+ + 2(cimetidine) ~ Fe3+(cimetidine)2 There was difficulty demonstrating binding of ionic forms of iron to cimetidine in aqueous solutions. This is likely due to the formation of ferric hydroxide complexes. It is possible that the formation of a ferric hydroxide-cimetidine complex is responsible for the reduction in cimetidine absorption. This complex would be difficult to detect due to the strong absorbance associated with the ferric hydroxide complex. The relationship of complex formation in methanol to the in vivo drug interaction is not clear, and the data obtained with methanol cannot be directly extrapolated to aqueous solutions. Iron can catalyze oxidation and reduction reactions, irreversibly altering drugs (7); however, there was no significant decrease in the stability of cimetidine in the presence of ferrous sulfate under the tested conditions. Digestive Diseases and Sciences, Vol. 38, No. 5 (May 1993)
An inspection of the histamine-2 receptor blocker structure s suggests strong binding to famotidine, as well as cimefidjne. Ranitidine binds very weakly to ionicforms of iron even in methanol (B.B. Hasinoff, unpublished observations). The mechanism of the cimetidine-iron interaction is likely the formation of iron--cimetidine complexes. Concurrent ingestion of iron supplements with cimetidine and famotidine could reduce efficacy and be associated with treatment failures. Human studies examining the bioavailability and efficacy of histamine-2 receptor blockers in the presence of iron therapy are warranted to determine the clinical significance o f histamine-2 blocker-iron interactions. ACKNOWLEDGMENT
We would like to thank Heather Arcari for her expert secretarial assistance. REFERENCES 1. La Piana Simonsen L: Top two hundred drugs of 1989. Pharm Times 56(4):56-64, 1990 2. Campbell NRC, Paddock V, Sundaram R: Alteration of methyldopa absorption, metabolism and blood pressure control caused by ferrous sulfate and ferrous gluconate. Clin Pharmacol Ther 43:381-386, 1988 3. Campbell NRC, Hasinoff B: Ferrous sulfate reduces levodopa bioavailability: Chelation as a possibie mechanism. Clin Pharmacol Ther 45:220-225, 1989 4. Campbell NRc, Hasinoff B, Campbell RRA: Ferrous sulfate reduces methyldopa absorption: Methyldopa:iron complex formation as a likely mechanism. Clin Invest Med 13:329332, 1990 5. Campbell NRC, Rankine D, Goodridge AE, Hasinoff B B , Kara M: Sinemet-ferrous sulfate interaction in patients with Parkinson's disease. Br J Clin Pharmacol 30:599-605, 1990 6. Campbell RRA, Hasinoff B, Chernenko G, Barrowman J, Campbell NRC: The effect of ferrous sulfate and pH on 1-dopa absorption. Can J Physiol Pharmacol 68:603-607, 1990 7. Campbell NRC, Hasinoff BB: Iron-drug complex formation. A potentially common cause of drug-drug interactions. Br J Clin Pharmacol 31:251-255, 1991 8. Campbell NRC, Kara M, Hasinoff B, McKay DW: Norfloxacin interaction with antacids and minerals. Br J Clin Pharmacol 33:115-116, 1992 9. Kara M, Hasinoff B, McKay DW, CamPbell NRC: Clinical and chemical interactions between iron and eiprofloxacin. Br J Clin Pharmacol 31:257-261, 1991 10. Neuvonen PJ, Gothoni G, Hackman R, Bjorksten K: Interference of iron with the absorption of tetracyclines in man. Br Med J 4:532-534, 1970 11. Osman MA, Patel RB, Schuna A, Sundstrom WR, Welling PG: Reduction in oral penicillamine absorption by food, antacid and ferrous sulfate. Clin Pharmacol Ther 33:465470, 1983 12. Polk RE, Healy DP, Sahai J, Drwal L, Racht E: Effect of
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CAMPBELL ET AL ferrous sulfate and multivitamins with zinc on absorption of ciprofloxacin in normal volunteers. Antimicrob Agents Chemother 33:1841-1844, 1989 13. Meddings JD, Westergaard H: Intestinal glucose transport using perfused rat jejunum in vivo: Model analysis and derivation of corrected kinetic constants. Clin Sci 76(4):104113, 1989 14. Brunton LL: Agents for control of gastric acidity and treatment of peptic ulcers. Chapter 37. In Goodman and Gilman's The Pharmacological Basis of Therapeutics, 8th ed. New York, Pergamon Press, 1990, pp 897-913, 1990 15. Golubov J, Flanagan P, Adams P: Inhibition of iron absorp-
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tion by omeprazole in rat model. Dig Dis Sci 36:405-408, 1991 16. Skikne BS, Lynch SR, Cook JD: Role of gastric acid in food iron absorption. Gastroenterology 81:1068-1071, 1981 17. Lin JI-I: Pharmacokinetic and pharmacodynamic properties of histamine H2-receptor antagonists. Clin Pharmacokin 20:2118-2236, 1991 18. Hillman RS: Drugs effective in iron-deficiency and other hypochromic anemias, Chapter 54. In Goodman and Gilman's The Pharmacological Basis of Therapeutics, 8th ed. New York, Pergamon Press, 1990, pp 1277-1310, 1990
Digestive Diseases and Sciences, VoL 38, No. 5 (May 1993)
