Abstract

Malaria remains one of the most persistent infectious diseases worldwide, and the spread of parasite resistance to chloroquine, sulfadoxine-pyrimethamine, and even artemisinin derivatives has renewed interest in plant-derived antimalarial leads. A widely used in vitro strategy for screening such leads targets the hemozoin-formation pathway, in which free ferriprotoporphyrin IX (heme), liberated during hemoglobin digestion by the intraerythrocytic parasite, is bio-mineralized into the inert crystalline pigment hemozoin. Its synthetic analogue, β-hematin, is used as a convenient surrogate target in semiquantitative colorimetric assays. This review compares ten in vitro studies (2014–2024) from our research group that examined the β-hematin-inhibitory activity of crude water and ethanolic extracts, preparative HPLC fractions, and pure phytochemical isolates obtained from Artemisia annua, Inula viscosa, grape ( Vitis vinifera) peels and seeds, pomegranate (Punica granatum ), turmeric ( Curcuma longa ), dietary flavonoid standards (morin, quercetin, catechin, quercitrin), guava ( Psidium guajava ), tea ( Camellia sinensis ), wild sage (Salvia officinalis) and dwarf nettle ( Urtica urens). Across these studies, the extracts and isolates were evaluated using the Deharo semi-quantitative β-hematin inhibition assay, complemented by HPLC-PDA and UPLC/HPLC-ESI-MS profiling that identified flavonols, flavones, flavan-3-ols, curcuminoids, phenolic acids, ellagitannins and diterpenes as the principal marker constituents. Pomegranate leaves, black grape peels, wild-sage bicarbonate extracts and Inula viscosa crude extract emerged as the most potent preparations, rivalling or exceeding chloroquine at pharmacologically relevant concentrations, while Cameroon-sourced Artemisia annua, curcumin and dwarf nettle showed intermediate-tomodest activity and several matrices (white grape, pomegranate juice, green tea prepared in water, and nettle roots/stems) were essentially inactive. Sodium bicarbonate pretreatment consistently enhanced and stabilized activity, most strikingly converting an inactive greentea water infusion into one comparable to chloroquine. Mechanistically, the reviewed data support a model in which polyphenolic hydroxyl groups engage the propionate side chains of free heme through hydrogen bonding, while aromatic rings stack against the porphyrin macrocycle, together blocking the hemozoin crystal growth face and precipitating oxidative parasite death. The consistent superiority of crude or semi-purified extracts over several single isolates further points to synergistic phytochemical interactions. Because β-hematin inhibition is a preliminary biochemical screen and not a validated surrogate for antiplasmodial efficacy, the rankings and mechanistic conclusions offered here should be read as hypothesis-generating rather than confirmatory.

Keywords

antimalarial activity, hemozoin, flavonoids, polyphenols, crude plant extracts; HPLC-PDA, LC-MS, sodium bicarbonate, natural products