Quantitative analysis of anti-inflammatory activity of orengedokuto: importance of combination of flavonoids in inhibition of PGE₂ production in mouse macrophage-like cell line J774.1

Orengedokuto is a Kampo formula which has been used for removing “heat” and “poison” to treat inflammation, hypertension, gastrointestinal disorders, and liver and cerebrovascular diseases. In this report, we quantitatively analyzed the anti-inflammatory effect of the component crude drugs of orengedokuto and their constituents, using inhibition of prostaglandin E₂ (PGE₂) production in the murine macrophage-like cell line J774.1. First, we compared PGE₂ production inhibitory activities of extracts of combinations of the component crude drugs, which showed that the activity could be ascribed to Scutellaria Root. Next, as baicalin (1), one of the major constituents of Scutellaria Root, did not show any activity, and baicalein (2), the aglycon of 1, showed only weak activity (IC₅₀ 92 μM), a hot-water extract of Scutellaria Root was fractionated under the guidance of the activity to give wogonin (3) (IC₅₀ 28 μM), 6-methoxywogonin (4) (IC₅₀ 7.2 μM) and oroxylin A (5) (IC₅₀ 45 μM) from the most active fraction. However, the activities of these compounds at concentrations equivalent to those in the extract were weaker than that of the extract, and none of these compounds alone could explain the activity of the extract. Therefore, we examined the activity of combinations of compounds 2–5. Comparison of all combinations of the four compounds in a ratio which is the same as in the extract revealed that wogonin (3) had an essential role in the activity, and a combination of baicalein (2) and wogonin (3), together with 6-methoxywogonin (4) or oroxylin A (5), was necessary to show activity equivalent to that of the extract.