Anti-inflammatory and Antimicrobial Activity of Isoflavonoids

Ing. Jana Hummelová

Doctoral thesis

Abstract:

In this study, 19 structurally related isoflavonoids were tested for their possible anti-inflammatory and antimicrobial properties. The first part of the research was focused on the assessment of anti-inflammatory activity of the isoflavonoids expressed as an inhibition of cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) enzymes in vitro. None of the tested compounds was active at least half as much as indomethacin, a standard COX inhibitor. The results showed that genistein inhibited the activity of COX-2 enzyme by 23.6 %, 39.9 % and 25.5 % in concentrations 50, 100, and 150 µM, respectively. In contrast, the most of the isoflavonoid structures exhibited 5-LOX inhibitory effect, whereas the most promising results were observed in biochanin A and formononetin, which inhibited the leukotrienes B4 (LTB4) production by 88.2 % and 78.9 % at 100 μM. For comparison, zileuton, a generally used inhibitor of 5-LOX enzyme, inhibited the LTB4 production by 95.4 % at the same concentration. In the second part, the in vitro antibacterial activity of isoflavonoids against nine Gram-positive and Gram-negative bacteria was determined using the broth microdilution method, and the results were expressed as minimum inhibitory concentrations (MICs). Demethyltexasin, hydroxydaidzein, biochanin A, demethylretusin and genistein produced significant antibacterial activity (MICs ≥ 16 µg ml-1). The most effective compound, demethyltexasin, was subsequently tested for its growth-inhibitory effect against Staphylococcus aureus, and it exhibited significant antistaphylococcal effects against various standard strains and clinical isolates, including methicillin and tetracycline resistant ones with the MICs ranging from 16 to 128 μg ml-1. Based on its inhibitory activity against S. aureus resistant infections, which are often hard to treat due to increasing resistance, especially to β-lactams, demethyltexasin was tested for its possible antistaphylococcal combinatory effects with antibiotics amoxicillin and oxacillin using checkerboard method. For comparison, common therapeutically used combination of amoxicillin/clavulanic acid was tested. Demethyltexasin showed strong synergistic interactions against most of S. aureus strains when combined with amoxicillin [sum of fractional inhibitory concentrations (∑ FIC) 0.257–0.461] and oxacillin (∑ FIC 0.109–0.484). When oxacillin was combined with demethyltexasin, resistance to this antibiotic was overcome in many cases. Moreover, antibiotic/demethyltexasin combinations were effective mainly against methicillin-resistant S. aureus (MRSA); whereas the commonly used drug amoxicillin/clavulanic acid was effective only against sensitive strains. The results indicated demethyltexasin as a compound able to act synergistically with β-lactams. In addition, some combinations are effective against MRSA and decrease staphylococcal resistance. Demethyltexasin seems to be a possible candidate for further research focused on antistaphylococcal drug development, especially against antibiotic-resistant strains. Additionally, biochanin A proved certain dual inhibitory effect against both 5-LOX activity and against some bacteria strains. These activities should be verified by future experiments, focused on its possible dual effect in vivo. As biochanin A is widely distributed in plants and food, mainly from Leguminosae family, the dual effect can be used in future as synergistic and/or concomitant treatment of infections and inflammation.

Keywords

isoflavones, structure-activity relationship, MICs, FICs, 5-lipoxygenase

Language used: English

Date on which the thesis was submitted / produced: 9. 11. 2018

Thesis defence

• Supervisor: prof. Ing. Ladislav Kokoška, Ph.D.

Czech University of Life Sciences Prague

Faculty of Tropical AgriSciences

Doctoral programme / field:
Agricultural Specialization / Tropical and Subtropical Agriculture