With the emergence of antimalarial drug resistance, the search for new treatment options is urgently needed. New drugs are also needed to be used as com-bination with the standard antimalarial drugs, particularly artemisinin-based combination (Sinclair et al. 2009). In the present study, the antimalarial activity of GIE, an indigenous medicinal plant, and functional food in Thailand, in combination with DHA on mice infected with PbANKA was reported. The mice treated with 50, 100, and 200 mg/kg of GIE showed significant 32%, 50%, and 65% inhibition, respectively, compared to untreated control. It has been described that in vivo antimalarial activity of plant extract in which > 30% inhibition makes this extract to be considered active (Krettli et al. 2009). Hence, GIE can be classified as active antimalarial activity and in agreement with the previous report (Ounjaijean et al. 2021a). This can be explained by the fact that antimalarial activity of GIE could have resulted from single or in the combined action of the bioactive metabolites such as phenols, flavones, alkaloids, anthraquinones, quinones, tannin, and triterpene saponins (Dunkhunthod et al. 2021; Khan et al. 2019; Rasoanaivo et al. 2011). Gymnemic acids, major active compounds of GIE, might play a vital role in exerting antimalarial activity (Kanetkar et al. 2007; Sahu et al. 1996; Saiki et al. 2020). The possible antimalarial mechanisms might be through antioxidant, intercalate with parasite DNA, inhibiting the fatty acid and protein biosynthesis of the parasite, elevating erythrocyte oxidation, immunomodulatory, inhibition of parasite invasion, or by other unknown mechanisms (Fidock et al. 2004; Trang et al. 2021).
We also tested GIE in combination with DHA on mice infected with PbANKA. The combination treatment ratio of 60/40 (DHA/GIE) was more effective than the others with 88.95% inhibition, and synergism was observed in this ratio. Significant anti-malarial activity of GIE in combination with DHA was found when compared to either GIE or DHA monotherapies. It is noteworthy that DHA has been proposed several mechanisms of action, including the production of free radicals or reactive metabolites and inhibition of nutrient flow to the parasite by interfering membrane transport properties (Dai et al. 2021; Guo 2016; Tilley et al. 2016). Moreover, DHA is an inhibitor of SERCA (SE sarco/endoplasmic reticulum Ca2+ - ATPase) of parasite and is a suitable target for artemisinin and its derivatives (Lu et al. 2015). GIE might be associated with different metabolic pathways, and as expected, the combination of GIE and DHA showed the best synergy with beneficial results. However, more work is needed before any firm conclusion.
Loss of BW is one manifestation of malaria infection in mice, and BW change of mice is a parameter for evaluating the antimalarial activity of extract (Basir et al. 2012). PbANKA infected mice treated with DHA/GIE combination at the doses of ratios 60/40, 40/60, 20/80, and 0/100 showed a significant increase in BW compared to the untreated control and DHA monotherapy suggested the effect of GIE in preventing malaria-related BW loss. This could be attributed to the presence of compounds in GIE that affect appetite leading to protect BW loss during malaria infection (Pinent et al. 2017). However, the activity of GIE in the combination ratio of 80/20 was not strong enough to significantly prevent BW loss.
Additionally, reduction of PCV during malaria infection is also considered. The PCV of untreated control was reduced because of PbANKA infection leading to rapid hemolysis (Khobjai et al. 2014; Zhu et al. 2015). The absence of significant PCV reduction among DHA/GIE combination-treated mice at the doses of ratios 60/40, 40/60, 20/80, and 0/100 may indicate the protective effect of GIE on PCV reduction during malaria infection. This could be due to activating erythropoietin and sustaining the availability of new erythrocytes production in the bone marrow. This finding agrees with the previous studies that showed the protective effect of GIE on PCV reduction in rodent malaria (Ounjaijean et al. 2021b).
MST is another parameter to evaluate the antimalarial activity of plant extracts. In this study, all doses of DHA/GIE combination significantly prolonged MST compared to untreated control, especially at the ratio of 60/40. These further supplements the evidence on inhibition of PbANKA resulting in a reduced overall parasite pathology on the experimental mice. A plant extract that can prolong MST of infected mice compared to untreated control is considered active (Oliveira et al. 2009). Hence, GIE, in combination with DHA, is regarded as active for antimalarial activity against PbANKA.
All results in this study provided the first evidence-based antimalarial activity with the synergistic effect of GIE in combination with DHA against PbANKA infected mice. Moreover, the combination between DHA and GIE also showed protective effects on BW loss and PCV reduction induced by malaria infection with prolonged MST. This study recommends GIE as an alternative antimalarial substance for using with standard antimalarial drugs such as DHA in the future. However, modes of action and possible mechanisms of GIE and its combination with DHA in malarial treatment should be further investigated.