Chemical Composition Analysis Using HPLC-UV and Inhibitory Activity of Thai Herbal Formulation on Proinflammatory Mediators

Herbal medicines are widely used to treat various physical disorders. One such remedy, Tri-Phon-That, is composed of rhizomes of three herbs: Zingiber zerumbet, Zingiber montanum, and Cymbopogon nardus. Traditionally, it is used to alleviate fever, swelling, bruising, and muscle pain-conditions associated with inflammation. This study aimed to evaluate the effects of Tri-Phon-That remedy and its individual ingredients on the production of nitric oxide (NO), interleukin-6(IL-6), and prostaglandin E2 (PGE-2). Additionally, the chemical compositions of the remedy and its plant extracts were analyzed. The results showed that the Tri-Phon-That extract strongly inhibited NO, IL-6, and PGE-2 production, with IC50 values of 5.35 ± 0.77, 7.48 ± 0.26, and 24.75 ± 0.33 μg/mL, respectively. Among the individual ingredients, Z. montanum exhibited the most potent anti-inflammatory activity, followed by Z. zerumbet, which effectively inhibited nitric oxide and IL-6 but had no effect on PGE-2. Chemical analysis using high-performance liquid chromatography revealed that the Tri-Phon-That extract contained zerumbone and compound D or (E)-4-(3’,4’-dimethoxyphenyl) but -3-en-1-ol at concentrations of 13.31 ± 0.87 and 3.16 ± 0.17 % w/w, respectively. These compounds were primarily found in Z. zerumbet and Z. montanum with concentrations of 36.05 ± 1.67 and 6.97 ± 0.28 % w/w, respectively. In contrast, no chemical constituents were found in C. nardus, which correlates with its weaker anti-inflammatory activity. In conclusion, Tri-Phon-That extract strongly inhibits pro-inflammatory cytokines, making it a promising treatment for acute inflammation. HIGHLIGHTS The finding of this study showed that the Tri-Phon-That remedy, a Thai herbal preparation, suppressed pro-inflammatory mediators such as nitric oxide, IL-6, and PGE-2. The inhibitory effect of the Tri-Phon-That extract on inflammatory mediators was comparable to that of its individual plant constituents. The Tri-Phon-That extract, analyzed in this study using HPLC validation methods, contained D and zerumbone as its main components. GRAPHICAL ABSTRACT