In vitro dual activity of Aloe marlothii roots and its chemical constituents against Plasmodium falciparum asexual and sexual stage parasites

Abstract

Ethnopharmacological relevance: Aloe marlothii A.Berger (Xanthorrhoeaceae) is indigenous to southern African countries where its aqueous preparations are used in traditional medicine to treat several ailments including hypertension, respiratory infections, venereal diseases, chest pain, sore throat and malaria.

Aim of the study: The aims of this study were as follows: (i) isolate and identify the antiplasmodial active compounds in A. marlothii roots. As the water extract was previously inactive, the dichloromethane:methanol (DCM:MeOH) (1:1) was used, (ii) examine the activity of the isolated compounds against Plasmodium falciparum asexual blood stage (ABS) parasites as well as for transmission-blocking activity against gametocytes and gametes, and (iii) to use in silico tools to predict the target(s) of the active molecules.

Materials and methods: The crude DCM:MeOH (1:1) extract of A. marlothii roots was fractionated on a reverse phase C8 column, using a positive pressure solid-phase extraction (ppSPE) workstation to produce seven fractions. The resulting fractions and the crude DCM:MeOH extract were tested in vitro against P. falciparum (NF54) ABS parasites using the malaria SYBR Green I based-fluorescence assay. Flash silica chromatography and mass-directed preparative high-performance liquid chromatography were utilised to isolate the active compounds. The isolated compounds were evaluated in vitro against P. falciparum asexual (NF54 and K1 strains) and sexual (gametocytes and gametes) stage parasites. Molecular docking was then used for the in silico prediction of targets for the isolated active compounds in P. falciparum.

Results: The crude extract and two SPE fractions displayed good antiplasmodial activity with >97% and 100% inhibition of ABS parasites proliferation at 10 and 20 μg/mL, respectively. Following UPLC-MS analysis of these active fractions, a targeted purification resulted in the isolation of six compounds identified as aloesaponol I (1), aloesaponarin I (2), aloesaponol IV (3), β-sorigenin-1-O-methylether (4), emodin (5), and chrysophanol (6). Aloesaponarin I (2) was the most bioactive, compared to other isolated constituents, against P. falciparum ABS parasites exhibiting equipotency against the drug-sensitive (NF54) (IC₅₀ = 1.54 μg/mL (5 μM)) and multidrug-resistant (K1) (IC₅₀ = 1.58 μg/mL (5 μM)) strains. Aloesaponol IV (3) showed pronounced activity against late-stage (>90% stage IV/V) gametocytes (IC₅₀ = 6.53 μg/mL (22.6 μM)) demonstrating a 3-fold selective potency towards these sexual stages compared to asexual forms of the parasite (IC₅₀ = 19.77 ± 6.835 μg/mL (68 μM)). Transmission-blocking potential of aloesaponol IV (3) was validated by in vitro inhibition of exflagellation of male gametes (94% inhibition at 20 μg/mL). In silico studies identified β-hematin and DNA topoisomerase II as potential biological targets of compounds 2 and 3, respectively.

Conclusion: The findings from our study substantiate the traditional use of A. marlothii to treat malaria. To our knowledge, this study has provided the first report on the isolation and identification of antiplasmodial compounds from A. marlothii roots. Furthermore, our study has provided the first report on the transmission-blocking potential of one of the compounds from the genus Aloe, motivating for the investigation of other species within this genus for their potential P. falciparum transmission-blocking activity.

Keywords: Aloe marlothii; Anthraquinones; Antiplasmodial; Gametocidal; Gametocytocidal; Plasmodium falciparum; Transmission-blocking.