Materials and Methods
- Chelators. The outer membrane of gram-negative bacteria acts as a permeability barrier for the cell. It is responsible for preventing molecules such as antimicrobials, detergents, and dyes from reaching the cytoplasmic membrane.17 Magnesium ions serve to stabilize the lipopolysaccharide layer of the outer membrane. Chelating agents, such as ethylenediaminetetraacetic acid (EDTA), bind magnesium ions in the lipopolysaccharide layer and produce cells with increased susceptibility to antimicrobial molecules and detergents.17 When E coli or Bacteroides were exposed to cell wall–degrading enzymes such as lysozyme, no loss of optical density was observed; however, when they were combined with a chelator, a drop in density occurred rapidly. This is due to the fact that gram-negative bacteria have a protective outer membrane that is stabilized by metal ions, and a chelator is required to remove it (Figure 1).
- Enzymes. We selected lysozyme, which is found in tears and other secretions. It is responsible for breaking down the polysaccharide walls of many kinds of bacteria. Lysozyme is known to lyse gram-positive bacteria but has difficulty accessing the cell wall of gram-negative organisms.18 A chelator such as EDTA or citrate can be combined with lysozyme to effectively kill E coli and Bacteroides (Figure 1).
- Peptides. Nisin was selected as an antibacterial peptide that exhibits a broad spectrum of inhibitory activity against many colonic bacterial species. Kordel and Sahl17 showed that E coli exhibited nisin sensitivity.9,19-21 Nisin at 1 mg/mL could reduce and slow the growth of Bacteroides fragilis as seen in Figure 2.
- Essential oils. Essential oils include but are not limited to carvarcol; thymol; and oils of ginger, cinnamon, mint, onion, black cumin, oregano, thyme, clove, garlic, eucalyptus, lavender, leleshwa, lemon, lemon myrtle, neem, cilantro, tea tree, and peppermint. The composition, structure, and functional groups of the oils play an important role in determining their antimicrobial activity.22 Usually compounds with phenolic groups are most effective.9,12 Among these, the oils of clove, oregano, rosemary, thyme, sage, and vanillin have been found to be most consistently effective against microorganisms. Oregano, clove, cinnamon, and citral are effective against both groups of bacteria.23 Some nonphenolic constituents of oils, such as allyl isothiocyanate, are quite effective against gram-negative bacteria (Figure 3).
Figures 5A and 5B.
Figure 5. Enumerations of Bacillus subtilis in the absence (plate A) and presence (plate B) of the antimicrobial formulation 1, the plates were diluted and plated at 1x106 after 6 hours of growth in nutrient media. Unlike Escherichia coli, B subtilis was not reduced by the antimicrobial formulation at the same concentration.
- Posserud I, Stotzer PO, Björnsson ES, Abrahamsson H, Simrén M. Small intestinal bacterial overgrowth in patients with irritable bowel syndrome. Gut. 2007;56(6):802-808.
- Corazza GR, Menozzi MG, Strocchi A, et al. The diagnosis of small bowel bacterial overgrowth. Reliability of jejunal culture and inadequacy of breath hydrogen testing. Gastroenterology. 1990;98(2):302-309.
- Sutherland JB, Bridges BM, Heinze TM. Comparison of the effects of antimicrobial agents from three different classes on metabolism of isoflavonoids by colonic microflora using Etest strips. Curr Microbiol. 2012;64(1):60-65
- Chu GM, Lee SJ, Jeong HS, Lee SS. Efficacy of probiotics from anaerobic microflora with prebiotics on growth performance and noxious gas emission in growing pigs. Anim Sci J. 2011;82(2):282-290.
- Lin HC. Small intestinal bacterial overgrowth: a framework for understanding irritable bowel syndrome. JAMA. 2004;292(7):852-858.
- Riordan SM, McIver CJ, Wakefield D, Bolin TD, Duncombe VM, Thomas MC. Small intestinal bacterial overgrowth in the symptomatic elderly. Am J Gastroenterol. 1997;92(1):47-51.
- Virally-Monod M, Tielmans D, Kevorkian JP, et al. Chronic diarrhoea and diabetes mellitus: prevalence of small intestinal bacterial overgrowth. Diabetes Metab. 1998;24(6):530-536.
- Pimentel M, Chow EJ, Lin HC. Eradication of small intestinal bacterial overgrowth reduces symptoms of irritable bowel syndrome. Am J Gastroenterol. 2000;95(12):3503-3506.
- Castiglione F, Rispo A, Di Girolamo E, et al. Antibiotic treatment of small bowel bacterial overgrowth in patients with Crohn's disease. Aliment Pharmacol Ther. 2003;18(11-12):1107-1112.
- Delves-Broughton J. Nisin and its application as a food preservative. Int J Dairy Technol. 1990;43(3):73-76.
- Lauritano EC, Gabrielli M, Scarpellini E, et al. Small intestinal bacterial overgrowth recurrence after antibiotic therapy. Am J Gastroenterol. 2008;103(8):2031-2035.
- Dorman HJ, Deans SG. Antimicrobial agents from plants: antibacterial activity of plant volatile oils. Appl Microbiol. 2000;88(2):308-316.
- Marino M, Bersani C, Comi G. Impedance measurements to study the antimicrobial activity of essential oils from Lamiaceae and Compositae. Int J Food Microbiol. 2001;67(3):187-195.
- Vertzoni M, Dressman J, Butler J, Hempenstall J, Reppas C. Simulation of fasting gastric conditions and its importance for the in vivo dissolution of lipophilic compounds. Eur J Pharm Biopharm. 2005;60(3):413-417.
- Worsøe J, Fynne L, Gregersen T, et al. Gastric transit and small intestinal transit time and motility assessed by a magnet tracking system. BMC Gastroenterology. 2011 Dec 29;11:145.
- Karamanolis G, Theofanidou I, Yiasemidou M, Giannoulis E, Triantafyllou K, Ladas SD. A glass of water immediately increases gastric pH in healthy subjects. Dig Dis Sci. 2008;53(12):3128-3132.
- Kordel M, Sahl HG. Susceptibility of bacterial, eukaryotic, and artificial membranes to the disruptive action of the cationic peptides Pep 5 and nisin. FEMS Microbiol Lett. 1986;34(2):139-144.
- Neidhardt FC, ed. Escherichia coli and Salmonella typhimurium: Cellular and Molecular Biology. Washington, DC: American Society for Microbiology; 1987.
- Hughey VL, Johnson EA. Antimicrobial activity of lysozyme against bacteria involved in food spoilage and food-borne disease. Appl Environ Microbiol. 1987;53(9):2165-2170.
- Barefoot SF, Klaenhammer TR. Detection and activity of lactacin B, a bacteriocin produced by Lactobacillus acidophilus. Appl Environ Microbiol. 1983;45(6):1808-1815.
- Liu W, Hansen JN. Some chemical and physical properties of nisin, a small-protein antibiotic produced by Lactococcus lactis. Appl Environ Microbiol. 1990;56(8):2551-2558.
- Tomczykowa M, Tomczyk M, Jakoniuk P, Tryniszewska E. Antimicrobial and antifungal activities of the extracts and essential oils of Bidens tripartita. Folia Histochem Cytobiol. 2008;46(3):389-393.
- Skandamis P, Koutsoumanis K, Fasseas K, Nychas GJ. Inhibition of oregano essential oil and EDTA on Escherichia coli O157:H7. Ital J Food Sci. 2001;13(1):65-75.
- Quigley EM, Quera R. Small intestinal bacterial overgrowth: roles of antibiotics, prebiotics, and probiotics. Gastroenterology. 2006;130(2 Suppl 1):S7-S90.