TITLE …………………… 13 Journal homepage: www.fia.usv.ro/fiajournal Journal of Faculty of Food Engineering, Ştefan cel Mare University of Suceava, Romania Volume XVIII, Issue 1 - 2019, pag. 13 - 17 (IMIDAZOLE-5-YL)YLIDENTHIAZOLIDONES: A NEW CLASS OF ANTIMICROBIAL AGENTS FOR FOOD INDUSTRY *Vitaliy CHORNOUS 1 , Alina GROZAV 1 , Victoria ANTONIYCHUK 1 , Nina YAKOVYCHUK 2 , Mykhaylo VOVK 3 1 Department of Medicinal and Pharmaceutical Chemistry, Bucovina State Medical University, 2 Teatralna Sq., Chernivtsi, 58002, Ukraine chornous.vitalij@bsmu.edu.ua 2 Department of Microbiology and Virology, Bucovina State Medical University, 2 Teatralna Sq., Chernivtsi, 58002, Ukraine 3Department of Mechanics of Organic Reactions, Institute of Organic Chemistry National Academy of Science of Ukraine, 2 Murmanska St., Kyiv, 02660, Ukraine *Corresponding author Received 23th January, 2019 accepted 28th March 2019 Abstract: An antimicrobial and antifungal activity of new derivatives of (imidazole-5- yl)ylidenthiazolidinones and its dependence on the substance structure and the type of substitutes in the imidazole and thiazole nucleus have been investigated. It was found that the sulphur of thiazoli- done section is the key factor that governs antimicrobial activity of the compounds. High antibacterial and antifungal activity against pathogenic microbes S. aureus ATCC25923, E. coli ATCC25922 and fungi C. albicans ATCC885-653 was determined for some representatives of the compounds series. Since the evaluated acute toxicity of the compounds is quite low, they can be recommended for further systematic investigation as possible prospective disinfection agents for food industry. Keywords: organic synthesis; imidazoles; thiazolidinones; antimicrobial and antifungal activity; foodstuff treatment and production 1. Introduction A risk of microbial and/or fungal contami- nation is one of important problems en- dangering foodstuff processing, production and storage at every stage of manufactur- ing. That is why, extensive efforts are made in various directions to safeguard the foodstuff and raw materials from primary and/or secondary bacterial/fungi contami- nation or to ensure that the pathogenic ob- jects would not be able to grow and devel- op. Strict observation of all sanitary and hy- gienic requirements is considered as one of the most effective actions protecting the food materials from pathogenic germs. Disinfection chemicals are known as inex- pensive and very effective solution to re- duce or exterminate unwanted microorgan- isms or yeast-like fungi such as E. coli, S. aureus and C. albicans at every stage of food processing technologies [1]. A number of chemical agents of different nature are used for this purpose: phenolic compounds, chloroorganic compounds, alcohols, aldehydes, oxygen-containing and oxidizing substances, various mixtures of quaternary ammonium salts, guanidine derivatives etc. [2]. However, many of them are either low efficient or toxic, cor- rosive to the equipment parts or destructive for the foodstuff components. Moreover, it usually takes shorter and shorter time now for the pathogenic germs to develop strong resistance against regular antibiotics or dis- infectants [3], which pushes new investiga- http://www.fia.usv.ro/fiajournal mailto:chornous.vitalij@bsmu.edu.ua Food and Environment Safety - Journal of Faculty of Food Engineering, Ştefan cel Mare University - Suceava Volume XVIII, Issue 1 – 2019 Vitaliy CHORNOUS, Alina GROZAV, Victoria ANTONIYCHUK, Nina YAKOVYCHUK, Mykhaylo VOVK (Imidazole-5- yl)ylidenthiazolidones: a nex class of antimicrobial agents for food industry, Food and Environment Safety, Volume XVIII, Issue 1 – 2019, pag. 13 – 17 14 tions directed onto finding of more effec- tive chemical disinfectants. That is why it is strongly topical problem to investigate and develop new stable dis- infection compounds with prolonged activ- ity, low toxicity and other adhere effects. It is known that 1,2,4-substituted 5- formylimidazoles can be considered as in- teresting candidates for this category. They have already been used widely to synthe- size various biologically active compo- nents [4-6] and it has been reported [7] that some derivatives of thiazolidone exhibit distinct antimicrobial and antifungal activi- ty. That is why a deeper investigation of antimicrobial potential of imidazol- ylmethylenthiazolidones seems topical and promising. 2. Experimental All target 5-[(1-aryl-1N-imidazole-5- yl)methylen]-1,3-thiazolidine-2-thio-4- ones (Fig. 1, structure 1), 5-[(1-aryl-1N- imidazole-5-yl)methylen]-1,3-thiazolidine- 2-thio-4-ones (Fig.1, structure 2) and 5-[(1- aryl-1N-imidazole-5-yl)methyl]-1,3- thiazolidine-2-thio-4-ones (Fig.1, structure 3) were synthesized by the condensation of available 1-arylimidazole-5-carbaldehydes with rhodanine or thiazolidine-2,4-diones under the basic catalysis conditions [8]. General structures of the synthesized com- pounds are shown in Fig. 1. N N N S O S R R R H 1 2 3 N N N S O O R R R H 1 2 3 N N N S O O R R R H 1 2 3 1 (a-f) 2 (a-f) 3 (a-f) Fig. 1. Structural composition of three classes of antimicrobial/antifungal agents synthesized and investi- gated in this work. Substitutes R 1 , R 2 and R 3 were combined as shown in Table 1. The degree of bioactivity of all the sub- stances has been tested on the museum strains of S. aureus ATCC25923, E. coli ATCC25922 and pathogenic fungi C. albi- cans ATCC885-653 by the double serial dilution method [9]. According to this method, the disposable 96-hole pads and the 8-channel titrator were used to evaluate an antimicrobial activity of the compounds libraries. The clinical strains mentioned above are known to cause usual invasions in humans and were chosen as the test- cultures to evaluate disinfection potential of the substances synthesized in this pro- ject. The clear bacterial colonies were cultivated in the meat-peptone broth for 24 hours at 37±1 0C until the number of the cells reached 105 CFU/ml. The clear fungi colo- nies of C. albicans were cultivated in Saburo nutritional agar for 7 days at 30±1 0C until the suspension concentration reached 10 CFU/ml. Then concentration of all solutions was standardized to 0.5 McFarland units by visual comparison. All compounds under investigation were used to prepare a series of solutions with con- centrations ranged from 500 μg/ml to 7.8 μg/ml and then the minimum inhibition concentration was found after 24 hours of Food and Environment Safety - Journal of Faculty of Food Engineering, Ştefan cel Mare University - Suceava Volume XVIII, Issue 1 – 2019 Vitaliy CHORNOUS, Alina GROZAV, Victoria ANTONIYCHUK, Nina YAKOVYCHUK, Mykhaylo VOVK (Imidazole-5- yl)ylidenthiazolidones: a nex class of antimicrobial agents for food industry, Food and Environment Safety, Volume XVIII, Issue 1 – 2019, pag. 13 – 17 15 cultivation in case of bacteria or after 48- 72 hours in case of fungi. The minimum bacteriostatic (MBsC), minimum bacteri- cide (MBcC), minimum fungi static (MFsC) and minimum fungicide concen- trations (MFcC) were determined as the lowest solution concentration of a given compound that results in the distinct op- pression of the test-cultures growth (static concentrations) or extermination of the colonies (MBcC and MFcC). 3. Results and discussion It was found that antimicrobial activity of imidazolylmethylenthiazolidones against various types of pathogenic germs depends significantly on the compound structure and composition (see Table 1). Table 1. Antimicrobial activity of the 1-3 compound series (μg/ml) № R1 R2 R3 S. aureus ATCC25923 E. coli АТСС25922 C. albicans АТСС885-653 MBsC MBcC MBsC MBcC MFsC MFcC 1 a CH3 H Cl 250 250 250 250 31.3 31.3 1 b C6H5 H Cl 3.9 3.9 31.3 62.5 7.81 7.81 1 c 4-ClC6H4 H Cl 15.6 31.3 7.81 15.6 31.3 31.3 1 d CH3 H SCH2CO2 H 31.3 31.3 31.3 125 15.6 31.3 1 e 4-ClC6H4 H SCH2CO2 H 250 250 250 1000 15.6 125 1 f 4-CH3C6H4 Cl Cl 7.8 7.8 250 250 31.3 62.5 Averaged activity amongst 1 а-f - 95.7 - 283.9 - 48.2 2 а C6H5 H Cl 62.5 125 250 500 125 250 2 b 2-CH3C6H4 H Cl 125 250 250 500 62.5 125 2 c 3-CH3C6H4 H Cl 125 250 250 500 125 250 2 d 4-CH3C6H4 H Cl 125 250 250 500 62.5 125 2 e 4- CH3OC6H4 H Cl 500 1000 500 1000 250 500 2 f 1-C10H7 H Cl 62.5 125 125 500 125 250 Averaged activity amongst 2 а-f - 333.3 - 500 - 250 3 а C6H5 H Cl 62.5 125 250 500 125 250 3 b 2-CH3C6H4 H Cl 125 250 250 500 62.5 125 3 c 3-CH3C6H4 H Cl 125 250 250 500 125 250 3 d 4-CH3C6H4 H Cl 125 250 250 500 62.5 125 Averaged activity amongst 3 а-d - 218.8 - 500 - 187.5 Total averaged activity amongst 1-3 series - 215.6 - 450.2 - 158.7 *Control *7.8 *15.6 *7.8 *15.6 **3.9 **1.9 *Furacilinum (furacin or nitrofurazone) and **Clotrimazole were used in the control experiments as the reference antimicr o- bial and antifungal compounds correspondingly. It should be noted that comparatively high antimicrobial and antifungal activities have been registered for all the compounds in- volved in this work. The total averaged MBcC for the entire series in case of S. aureus was 215.6 μg/ml, in case of E. coli – 450.2 μg/ml while MFcC was even lower – 158. 7 μg/ml. In our opinion, such a high antifungal efficiency can be caused by im- idazole nucleus present in all compounds of the series 1-3. Since bactericide and fungicide indexes are more important than Food and Environment Safety - Journal of Faculty of Food Engineering, Ştefan cel Mare University - Suceava Volume XVIII, Issue 1 – 2019 Vitaliy CHORNOUS, Alina GROZAV, Victoria ANTONIYCHUK, Nina YAKOVYCHUK, Mykhaylo VOVK (Imidazole-5- yl)ylidenthiazolidones: a nex class of antimicrobial agents for food industry, Food and Environment Safety, Volume XVIII, Issue 1 – 2019, pag. 13 – 17 16 corresponding ‘static’ concentrations, we skipped the former parameters in the fur- ther analysis. A dependence of antimicrobial activity on the structure of thiazolidone section can easily be understood from comparison be- tween subseries 1 (containing the exocyclic atom of sulphur) and subseries 2 (contain- ing the exocyclic atom of oxygen). An in- fluence of reduction of the methylene fragment to methyl on antimicrobial activi- ty can also be understood from comparison between subseries 2 and 3. For instance, it can be noted that the series 1 and 2 have same structural composition while antimicrobial and antifungal activi- ties of the sulphur-containing series 1 are higher than those of the oxygen-containing series 2. An average MBcC and MFcC for the set 1 are (μg/ml): 95.7 (S. aureus), 283.9 (E. coli) and 48.2 (C. albicans) cor- respondingly. Similar values for the set 2 are sufficiently higher that proves lower activity of these compounds: 333.3 (S. au- reus), 500 (E. coli) and 250 μg/ml (C. albi- cans). Transformation of the methylene structural bridge in the set 2 to the methyl one (set 3) results in some increase of averaged anti- microbial and antifungal activity of the compounds and the corresponding values of MBcC and MFcC decrease down to 218.8, 500 and 187.5 μg/ml (see Table 1). Therefore, it is obvious that the compounds 1a-f reveal the highest germ-suppressing activity and should be analyzed further to find the most effective representative. As seen from the data of Table 1, the struc- ture of the imidazole part of a compound is the key factor influencing germ- suppressing activity. For instance, both an- timicrobial and antifungal activities are increasing after exchange of a substitute in the first position from methyl (1a) to phe- nolic (1b) or 5-chlorophenolic (1c). On the other hand, a substitute in the sec- ond position of imidazole cycle also seems influential. When the polar atom of chlo- rine is inserted in the R2 position, the value of MBcC improves to 7.8 μg/ml in case of S. aureus while MFcS reaches 62.5 μg/ml. In contrary, this substitute makes no effect on the value of MBcC in case of E. coli (see comparison between 1a and 1f). Fig. 2. Comparison of minimal bactericide (MBcC) and fungicide (MFcC) concentrations (mg/ml) for some representatives of the subseries 1 and the control compounds. Food and Environment Safety - Journal of Faculty of Food Engineering, Ştefan cel Mare University - Suceava Volume XVIII, Issue 1 – 2019 Vitaliy CHORNOUS, Alina GROZAV, Victoria ANTONIYCHUK, Nina YAKOVYCHUK, Mykhaylo VOVK (Imidazole-5- yl)ylidenthiazolidones: a nex class of antimicrobial agents for food industry, Food and Environment Safety, Volume XVIII, Issue 1 – 2019, pag. 13 – 17 17 It is also interesting to analyze an influence of a substitute to the fourth position of im- idazole cycle (R3). As seen from the com- parison between the compounds 1a and 1d, both MBcC and MFsC values get im- proved for all three types of the germs when chlorine is substituted with the hy- drophilic fragment of thioglycolic acid. When there is the aryl fragment in the first position of imidazole fragment (1c and 1e), substitution of chlorine in the fourth posi- tion with the thioglycolic acid fragment results in a serious decrease of antigerm activity especially against S. aureus and C. albicans (compare corresponding values of MBsC, MBcC and MFcC for 1c and 1e). It is interesting to note that the change of MFsC in this case is opposite and it is im- proving because of the above-mentioned substitution. The above discussion is visualized graph- ically in Fig. 2 for better comprehension of comparative bactericide and fungicide ac- tivities of some representatives of the sub- series 1 and the corresponding control compounds. Finally, toxicity of all the synthesized compounds has been evaluated using the computer simulation software GUSAR Online [10]. The toxicity of the entire class of the synthesized compounds has been found low: Rat oral LD50 = 1027 mg/kg; Rat SC LD50 = 1366 mg/kg. 4. Conclusion A wide spectrum of bactericide and fungi- cide or static activities has been found for some representatives of the investigated compounds. MBcC and MFcC values were close or sometimes better than those of the control agents. Since predicted toxicity was appropriately low, (imidazole-5-yl) ylidenthiazolidones can be used in further search for the low toxic bacterio- and fun- gicide agents for food industry. 5. References [1]. G. WIRTANEN, S. SALO, Disinfection in Food Processing – Efficiency Testing of Disinfect- ants., Rev. Environ. Sci. and Biotech., 2(2–4): 293– 306 (2003). [2]. G. MCDONNELL, A. D. RUSSELL, Antisep- tics and Disinfectants: Activity, Action, and Re- sistance. Clin. Microbiol., Rev., 12(1): 147–179 (1999). [3]. A. J. MCBAIN, P. GILBERT, D. G. ALLI- SON, Biofilms and Biocides: Are there Implica- tions for Antibiotic Resistance? Rev. Environ. Sci. and Biotech., 2(2–4): 141–146 (2003). [4]. V. A. CHORNOUS, M. K. BRATENKO, M. V. VOVK, Polyfunctionalimidazoles: I. Synthesis of 1-substituted 4-chloro-1H-imidazole-5- carbaldehydes by Wilsmeier-Haack reaction. Rus. J. Org. Chem., 45(8): 1210–1213 (2009). [5]. V. A. CHORNOUS, A. N. GROZAV, L. D. TODORIKO, M. V. VOVK, Synthesis and Biolog- ical Activity of 4-Chloro-1H-Imidazole-5- Carbaldehyde Thiosemicarbazones. Pharm. Chem. J., 47(10): 524–526 (2014). [6]. V. A. CHORNOUS, A. A. PALAMAR, I. N. YAREMII, M. V. VOVK, Synthesis and antioxi- dant activity of [(1-aryl-5-formylimidazol-4- yl)thio]acetic acids. Pharm. Chem. J., 47(2): 96–98 (2013). [7]. N. C. DESAI, A. M. DODIYA, P. N. SHIHORA, A clubbed quinazolinone and 4- thiazolidinone as potential antimicrobial agents. Med. Chem. Res., 21(8): 1577–1586 (2012). [8]. V. O. CHORNOUS, O. YA. MELNYK, O. M. HLIEBOV, M. V. TYKHONENKO, L. M. SHEREMETA, O. K. YAROSH, et al., The synthe- sis, hypoglycemic activity and acute toxicity of new imidazole-thiazolidines hybrid structures. J. Org. Pharm. Chem., 14(1): 46-52 (2016). [9]. S. N. KULIKOV, R. Z. HAYRULLIN, S. A. LISOVSKAYA, Antimycotic activity of chitozan with various molecular weights and its influence on the cell morphology in some yeast-like fungi. Prob- lem. Medic. Mycol., 12(2): 32-36 (2010) (In Rus- sian). [10]. http://pharmaexpert.ru/GUSAR/acutoxpredict. html (Accessed Jan. 05, 2019). http://pharmaexpert.ru/GUSAR/acutoxpredict.html http://pharmaexpert.ru/GUSAR/acutoxpredict.html 1. Introduction It should be noted that comparatively high antimicrobial and antifungal activities have been registered for all the compounds involved in this work. The total averaged MBcC for the entire series in case of S. aureus was 215.6 μg/ml, in case of E. coli... A dependence of antimicrobial activity on the structure of thiazolidone section can easily be understood from comparison between subseries 1 (containing the exocyclic atom of sulphur) and subseries 2 (containing the exocyclic atom of oxygen). An influ... For instance, it can be noted that the series 1 and 2 have same structural composition while antimicrobial and antifungal activities of the sulphur-containing series 1 are higher than those of the oxygen-containing series 2. An average MBcC and MFcC f... Transformation of the methylene structural bridge in the set 2 to the methyl one (set 3) results in some increase of averaged antimicrobial and antifungal activity of the compounds and the corresponding values of MBcC and MFcC decrease down to 218.8, ... Therefore, it is obvious that the compounds 1a-f reveal the highest germ-suppressing activity and should be analyzed further to find the most effective representative. As seen from the data of Table 1, the structure of the imidazole part of a compound is the key factor influencing germ-suppressing activity. For instance, both antimicrobial and antifungal activities are increasing after exchange of a substitute in th... On the other hand, a substitute in the second position of imidazole cycle also seems influential. When the polar atom of chlorine is inserted in the R2 position, the value of MBcC improves to 7.8 μg/ml in case of S. aureus while MFcS reaches 62.5 μg/m... The above discussion is visualized graphically in Fig. 2 for better comprehension of comparative bactericide and fungicide activities of some representatives of the subseries 1 and the corresponding control compounds. Finally, toxicity of all the synthesized compounds has been evaluated using the computer simulation software GUSAR Online [10]. The toxicity of the entire class of the synthesized compounds has been found low: Rat oral LD50 = 1027 mg/kg; Rat SC LD50 =... 4. Conclusion