Dual Anti-Inflammatory Activities of COX-2/5-LOX Driven by Kratom Alkaloid Extracts in Lipopolysaccharide-induced RAW 264.7 Cells

Abstract

Cyclooxygenase (COX) and lipoxygenase (LOX) enzymes play a crucial role in the production of pro-inflammatory eicosanoids, including prostaglandins and leukotrienes (LTs) via arachidonic acid (AA) pathways. Non-steroidal anti-inflammatory drugs (NSAIDs) typically work by inhibiting COX enzymes (COX-1, COX-2) to alleviate inflammatory responses in our bodies. However, the use of these selective COX inhibitors results in an upregulation of the AA pathway. This condition stimulates the LOX enzymes to increase LT production, exacerbating the severity of the disorders. In this study, the alkaloid extract derived from the leaf of Mitragyna speciosa (Kratom) demonstrated a dual inhibitory effect on COX-2/5-LOX enzymes in lipopolysaccharides (LPS)-induced RAW 264.7 macrophage cells. The alkaloid extract containing ~ 46% mitragynine inhibited COX-2 and 5-LOX activity at concentrations of less than 25 ppm with no toxicity to the cells. Above 25 ppm, the alkaloid extract exhibited toxicity to the cells (e.g,, ~ 46% viability at 50 ppm) and only inhibited COX-2 activity. In contrast, the Kratom crude extract containing ~ 5% mitragynine did not inhibit COX-2 or 5-LOX activity in LPS-induced RAW 264.7 macrophage cells at more than 25 ppm and did not exhibit toxicity to the cells even at 100 ppm. The alkaloid compounds in the Kratom leaf are likely responsible for this activity, as the alkaloid extract containing these biomolecules suppressed reactive oxygen species (ROS), nitric oxide (NO), inducible nitric oxide synthase (iNOS), and pro-inflammatory cytokines like tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). Molecular studies also suggested a strong binding affinity of Kratom alkaloids to the active sites of COX-2 and 5-LOX. The dual inhibitory activity of the Kratom alkaloids against COX-2 and 5-LOX provides insights into their potential as safer NSAIDs.