Influence of the Sodium Salt of 3α,7α-Dihydroxy-12-Oxo-5β-Cholanate on Antimicrobial Activity of Ampicillin In Vitro

Ljiljana Suvajdžić, Slobodan Gigov, Aleksandar Rašković, Srđan Stojanović, Maja Bekut, Dubravka Milanov, Ivana Čanak, Svetlana Goločorbin-Kon, Momir Mikov

Abstract


Background: Multiple resistances to antibiotics are an emergent problem worldwide. Scientists intensively search for new substances with the antimicrobial potential or the mode to restore the activity of old-generation antibiotics. Ampicillin is the antibiotic with the expanded range of antimicrobial activity, but its use has decreased due to the poor absorption and highly developed resistance. In vivo studies showed that ampicillin has better absorption and bioavailability if combined with bile acid salts. The aim of this study was to examine antimicrobial effects of ampicillin alone and its combination with semisynthetic monoketocholic acid salt (MKH) in vitro.

Materials, Methods & Results: In this study, commercial preparation of ampicillin and sodium salt of 3α,7α-dihydroxy-12oxo-5β-cholanate were used. Their effects were evaluated on Escherichia coli (E. coli), Enterococcus faecalis (E. faecalis) and Enterococcus faecium (E. faecium), obtained from urine specimens of dogs with clinically manifested cystitis. The first two investigated strains were ampicillin-sensitive, while E. faecium was resistant to ampicillin. Modified macrodilution method according to Clinical and Laboratory Standards Institute Guidelines (M7-A8) was performed. Bacterial suspension equivalent to 0.5 McFarland was prepared in saline, compared to the standard (Biomerieux) ad oculi. The density was checked spectrophotometrically at a wavelength of 625 nm and adjusted if necessary to the desired absorbance from 0.08 to 0.1. The resultant suspension was diluted 1:100 and inoculated in test tubes. Number of bacteria was counted on Petri plates using dilutions from 10-3 to 10-7 in order to obtain valid and countable plates. One hundred microliters of appropriate dilutions were aseptically plated in triplicate onto nutrient agar. Plates were incubated on 37°C for 72 h, under aerobic conditions. The number of colony forming units (CFU) was determined by direct counting. As a valid for enumeration, we took plates with 30 to 300 CFU. Percentage of killed bacteria for ampicillin was from 69.33-95.19% for E. coli, 87.1296.92% for E. faecalis and 7.20-33.30% for E. faecium. Ampicillin applied in the combination with MKH killed 99.99% to 100% of E. coli, 94.59% to 99.91% of E. faecalis and 31.73% to 64.76% of E. faecium. Mean percentage of killed bacteria for ampicillin was 81.93% for E. coli, 91.64% for E. faecalis, and 18.13% for E. faecium, while in combination with MKH percentage was 99.96% for E. coli, 98.23% for E. faecalis and 47.54% for E. faecium.

Discussion: Results are presented as pharmacological minimal inhibitory concentration (MIC) values. Ampicillin was applied at the concentration higher than the therapeutic one, which could explain high MIC values for E. coli and E. faecalis. The combination of ampicillin with MKH showed the best improvement of antimicrobial effect on E. faecium (Δ = 29.41%), isolate that was resistant to ampicillin when applied alone. In all the investigated isolates, the combinations with MKH were more effective than ampicillin administered alone. It seems that MKH demonstrates a synergistic antimicrobial activity with ampicillin in vitro, which considerably decreases MIC values for all investigated isolates. These results implicate that MKH could restore the previous activity of ampicillin against some bacteria, which could be a significant benefit for clinical practice.


Keywords


antimicrobial effect; ampicillin; resistance; bile acid; in vitro.

Full Text:

PDF

References


Atanacković M., Posa M., Heinle H., Gojković-Bukarica L. & Cvejić J. 2009. Solubilization of resveratrol in micellar solutions of different bile acids. Colloids and Surfaces B: Biointerfaces. 72(1): 148-154.

CLSI. 2009. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically. Approved Standard M7-A8. 8th edn. CLSI. Wayne, PA. USA. 29(2): 16-17.

Damborg P., Top J., Hendrickx A.P., Dawson S., Willems R.J. & Guardabassi L. 2009. Dogs Are a Reservoir of Ampicillin-Resistant Enterococcus faecium Lineages Associated with Human Infections. Applied and Environmental Microbiology. 75(8): 2360-2365.

De Campos T., Pisseti C., Werlang G.O., Lopes G.V., Kich J.D. & Cardoso M. 2015. Antimicrobial Resistance of Enterococcus Isolated from Pre-Chill Swine Carcasses. Acta Scientiae Veterinariae. 43: 1259.

De Souze Lopez E., Maciel W.C., Albuquerque A.H., Machado D.N., Bezerra W.G.A., Vasconcelos R.H., Lima B.P., Gonçalves G.A.M. & Teixeira R.S.C. 2015. Prevalence and Antimicrobial Resistance Profile of Enterobacteria Isolated from Psittaciformes of Illegal Wildlife Trade. Acta Scientiae Veterinariae. 43: 1313.

Golocorbin-Kon S., Mikov M., Arafat M., Lepojevic Z., Mikov I., Sahman-Zaimovic M. & Tomic Z. 2009. Cefotaxime pharmacokinetics after oral application in the form of 3alpha, 7alpha-dihydroxy- 12-keto- 5beta- cholanate microvesicles in rat. European Journal of Drug Metabolism and Pharmacokinetics. 34(1): 31-36.

Gómez-Zavaglia A., Kociubinski G., Pérez P., Disalvo E. & De Antoni G. 2002. Effect of bile on the lipid composition and surface properties of bifidobacteria. Journal of Applied Microbiology. 93(5): 794-799.

Krstić T., Suvajdžić Lj., Stojanović S., Crvenković-Lozanov Z., Dejanović J., Čabarkapa I., Velhner M. & Stefanović V. 2016. Antimicrobial Activity of Sour Cherry. Agro Food Industry Hi Tech. 27(1): 56-58.

Krstić T., Suvajdžić Lj., Stojanović S., Petrović T., Bekut M., Ilić N. & Suvajdžić Z. 2015. Antimicrobial activity of blackberry juice from Serbia on animal pathogens. In: Proceedings of First International Symposium of Veterinary Medicine ISVM2015 (Vrdnik, Serbia). pp.462-467.

Krstić T., Suvajdžić Lj., Stojanović S., Velhner M., Milanov D., Bojić G. & Ilić N. 2014. Different antimicrobial effects of raspberry depending on the method of active components isolation. Food and Feed Research. 41(2): 125-130.

Lalić-Popović M., Vasović V., Milijašević B., Goločorbin-Kon S., Al-Salami H. & Mikov M. 2013. Deoxycholic Acid as a Modifier of the Permeation of Gliclazide through the Blood Brain Barrier of a Rat. Journal of Diabetes Research. (17 i): 598603. [doi: 10.1155/2013/598603].

Mikov M., Kevrešan S., Kuhajda K., Jakovljević V. & Vasović V. 2004. 3α, 7α-dihydroxy-12-oxo-5β-cholanate as blood-brain barrier permeator. Polish Journal of Pharmacology. 56(3): 367-371.

Mikov M., Kuhajda K. & Kandrač J. 2003. Savremeni pravci farmakološke primene žučnih kiselina. Medicinski Pregled. 56(5-6): 237-242.

Mikov M., Raskovic A., Jakovljevic E., Dudvarski D. & Fawcett P. 2005. Influence of the bile salt sodium 3α, 7α dihydrooxy-12-oxo-5β-cholanate on ampicilin pharmacokinetics in rats. Asian Journal of Drug Metabolism and Pharmacokinetics. 5(3): 197-200.

Miljkovic D., Kuhajda K. & Hranisavljevic J. 1996. Selective C-12 oxidation of cholic acid. Journal of Chemical Research. 2(2): 106-107.

Mrestani Y., Bretschneider B., Härtl A. & Neubert R.H. 2003. In vitro and in vivo studies of cefpirom using bile salts as absorption enhancers. Journal of Pharmacy and Pharmacology. 55(12): 1601-1606.

Murakami T., Sasaki Y., Yamayo R. & Yata N. 1984. Effect of bile salts on the rectal absorption of sodium ampicillin in rats. Chemical and Pharmaceutical Bulletin. 32(5): 1948-1955.

Posa M. & Kuhajda K. 2010. Hydrophobicity and haemolytic potential of oxo derivatives of cholic, deoxycholic and chenodeoxycholic acids. Steroids. 75(6): 424-431.

Sabo A., Tomić Z., Rašković A. & Stanulović M. 2014. Antibakterijski lekovi (sa ostalim antiinfektivnim lekovima). 4th edn. Novi Sad: Alfagraf, p.59.

Stojančević M., Pavlović N., Goločorbin-Kon S. & Mikov M. 2013. Application of bile acids in drug formulation and delivery. Frontiers in Life Science. 7(3-4): 112-122.

Suvajdzic L., Leovac V., Joksovic M., Bogdanovic G., Kojic V., Vujic N., Mrdja T. & Kocic B. 2013. Antimicrobial Activity of Copper (II) Complex with 1,2-bis [(1,3-diphenylpyrazol-4-yl)methyl] Diaminoethane. Acta Scientiae Veterinariae. 41: 1123.

Suvajdzic L., Stojakovic N., Mikov M., Satara S.S., Skrbic R., Vidic B., Dankuc D. & Suvajdzic Z. 2015. Influence of Bile Acids on Rat Gut Microflora Deterioration Induced by Oral Ampicillin Treatment. Acta Scientiae Veterinariae. 43: 1282.

Todorović D., Velhner M., Milanov D., Vidanović D., Suvajdžić Lj. & Krnjaić D. 2015. Characterization of tetracycline resistance of Salmonella enterica subspecies enterica serovar infantis isolated from poultry in the northern part of Serbia. Acta Veterinaria-Beograd. 65(4): 548-556.

Vasović V., Vukmirović S., Mikov M., Mikov I., Budakov Z., Stilinović N. & Milijašević B. 2014. Influence of bile acid derivates on morphine analgesic effect in mice. Vojnosanitski Pregled. 71(8): 767-771.

Velhner M., Suvajdžić Lj., Petrović J. & Šeperanda M. 2012. Antimicrobial resistance of Escherichia coli in wild animals. Arhiv veterinarske medicine. 5(2): 35-44.

Waar K., Van der Mei H.C., Harmsen J.M., Degener J.E. & Busscher H.J. 2002. Adhesion to bile drain materials and physicochemical surface properties of Enterococcus faecalis strains grown in the presence of bile. Applied and Environmental Microbiology. 68(8): 3855-3858.

Yang L., Zhang H., Mikov M. & Tucker I.G. 2009. Physicochemical and biological characterization of monoketoholic acid, a novel permeability enhancer. Molecular Pharmaceutics. 6(2): 448-456.




DOI: https://doi.org/10.22456/1679-9216.81166

Copyright (c) 2018 Ljiljana Suvajdžić, Slobodan Gigov, Aleksandar Rašković, Srđan Stojanović, Maja Bekut, Dubravka Milanov, Ivana Čanak, Svetlana Goločorbin-Kon, Momir Mikov

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.