# Comparative Study of _Plantago media_ Extracts in the Treatment of _Acanthamoeba_ sp. Trophozoites
## 4 Discussion
The mechanism behind the antiprotozoan, exactly anti-leishmanial activity of acetoside has been found to be the inhibition of an arginase enzyme, leading to the inhibition of polyamine biosynthesis that is important to infectivity, and has been attributed to the catechol groups in this compound (two adjacent hydroxyl groups on each of the two phenyl rings) [28]. In our studies, the correlation between IC\({}_{50}\) values and groups of phenolic secondary metabolites was highest for phenylethanolamine glycosides and acetoside (R\({}^{2}\) 0.6248 and 0.6131, respectively). Although the inhibition of arginase has not been mentioned among modes of anti-acanthamoeba action [30], it is possible due to the ubiquitous occurrence of that enzyme [56]. The brews of green tea (_Camelia sinensis_), known for the high content of catechin derivatives possessing catechol and pyrogallol groups (e.g., EGCG--epigallocatechin 3-gallate), have exhibited pronounced anti-acanthamoeba activity [50]. The authors have hypothesized that the activity could be due to the inhibition of DHFR (dihydrofolate reductase) and serine protease and metalloproteinase enzymes resulting in the disruption of DNA synthesis and cytolysis of _Acanthamoeba_ trophozoites, respectively [50]. When it comes to all the _Plantago media_ extracts used, it can be seen that the lowest IC\({}_{50}\), and thus the highest biological activity, have leaf extracts, both from ground plants (IC\({}_{50}\) from 0.95 to 1.80 mg/mL) and obtained by biotechnological methods in vitro (3.30 to 5.50 mg/mL). A similar tendency was observed in the case of extracts from the leaves of shoot cultures of _E. campestre_ and _E. planum_, for which IC\({}_{50}\) was estimated to be lower than that for extracts from roots obtained in vitro [51]. As it turned out, this activity did not correlate with the content of polyphenols, which, for example, for _Eryngium planum_, was similar in roots and shoots obtained from micropropagated plants or with the content of triterpene saponins, which was impressively higher in the case of in-vitro-derived roots [55]. If we consider extracts from. _C. japonica_, IC\({}_{50}\) was the lowest for extracts from leaves of shoots regenerated in vitro than from shoots of plants from natural sites, callus, or fruits [55]. The anti-acanthomoebic activity of the extracts clearly correlated with the content of pentacyclic triterpenoids [55]. Considering the anti-acanthoamoeba activity of _L. flos-cuculi_ extracts, 80% methanol fractions from the herb had a higher IC\({}_{50}\) value than those from roots or even biotechnology obtained callus cells [54]. This relationship is probably not conditioned by the amount of main cedysteroids, because, in the case of this species, the content of the sum of polypodine B and 20-hydroxycedysone in the herb and roots is similar, while these compounds are not synthesized by callus cells [57]. Among aerial part extracts from plant species belonging to the _Asteraceae_ family (_C. bella_, _C. daghestanica_, _R. pulchrum_, and _T. vulgare_), the lowest IC\({}_{50}\) was calculated for _C. bella_, _C. daghestanica_, and _T. vulgare_, while IC\({}_{50}\) for _R. pulchrum_ was very high (>10 mg/mL). The amoebicidal effect of extracts may be attributed mainly to the sesquiterpene lactones, which are major compounds of four species from _Asteraceae_. The most common sesquiterpene lactones isolated from the investigated plants were guaianolides: cyanopicrin, janerin, and chlorojanerin [53]. On the other hand, the acanthamoeba activity of _Origanum syriucum_ and _O. laevisatum_ extracts seemed to be very low, even at a concentration of 32 mg/mL [58]. The above-discussed data on the anti-acanthamoeba activity of various plant species are compiled in the Table 5.