Bioactive substance contents and therapeutic potential for skin inflammation of an herbal gel containing Derris reticulata and Glycyrrhiza glabra

Abstract

Context: Derris reticulata Craib. and Glycyrrhiza glabra L., of the Fabaceae, have been used as active components in Thai herbal formulas for the treatment of fever and skin diseases.

Objective: To evaluate the physicochemical and pharmacological properties of the developed herbal gel formulation containing the combined extract from D. reticulata stem wood and G. glabra root (RGF).

Materials and methods: The potential of the herbal gel formulation containing RGF (8% w/w) as the active ingredient was studied by evaluating the anti-inflammatory, antioxidant, and anti-Staphylococcus aureus activities using quantitative reverse transcription-polymerase chain reaction assay, spectrophotometric method, and broth microdilution technique, respectively. The reference standards for the biological testing included Nω-nitro-L-arginine (L-NA), ascorbic acid, catechin, and penicillin G. The stability study of the RGF herbal gel was performed by a heating-cooling test (at 45 °C for 24 h and at 4 °C for 24 h/1 cycle; for 6 cycles), and the bioactive marker compounds in the herbal gel were investigated by the HPLC technique.

Results: RGF showed promising pharmacological effects, particularly on its anti-inflammatory property (IC₅₀ 73.86 µg/mL), compared to L-NA (IC₅₀ 47.10 µg/mL). The RGF-containing gel demonstrated anti-inflammatory (IC₅₀ 3.59 mg/mL) and free radical scavenging effects (IC₅₀ 0.05-4.39 mg/mL), whereas it had no anti-S. aureus activity (MIC > 10 mg/mL). The active ingredient in the developed herbal gel significantly inhibited lipopolysaccharide-induced nitric oxide production by downregulating iNOS mRNA levels. The contents of the bioactive markers in the RGF gel (lupinifolin and glabridin) did not change significantly after stability testing.

Discussion and conclusions: The RGF-containing gel has potential to be further developed as an herbal product for the treatment of skin inflammation.

Keywords: Dermatitis; folk medicines; herbal medicines; hydrogel; traditional Thai medicine.

Figure 1.

The developed gel formulation at day 0; A: RGF (8%w/w) gel and B: Gel base.

Figure 2.

HPLC chromatogram of the combined standards glabridin and lupinifolin. Detection: VWD (wavelength; 281 nm at 0–6 min, 254 nm at 6.1–20 min). Mobile phase: Isocratic system (methanol:15% acetic acid in DI water, 80:20 v/v).

Figure 3.

Anti-inflammatory effect of RGF gel, gel base, standard indomethacin gel, and standard L-NA on lipopolysaccharide (LPS)-induced nitric oxide (NO) production in murine macrophage RAW264.7 cells: A herbal gel at day 0, B herbal gel after accelerated testing, C gel base at day 0, D gel base after accelerated testing, E std. indomethacin gel, and F std. L-NA. IC₅₀ was determined by non-linear regression analysis in Prism software.

Figure 4.

Inhibition of nitric oxide production of different concentrations of the extract (6.25-200 µg/mL) on RAW 264.7 cells stimulated by lipopolysaccharides (LPS). The viability was investigated using MTT assay (A). The nitric oxide levels in the cell culture supernatant were measured by nitrite assay (B). Values are representative of three independent experiments performed in triplicate and are expressed as mean ± SD (error bars). **** significant difference between treatment and control group (^****p < 0.0001).

Figure 5.

Inhibitory effects of the extract on LPS-stimulated iNOS expression in RAW 264.7 cells. The expression levels of the gene are shown after normalization with the housekeeping actin gene β-actin. Values are representative of three independent experiments performed in triplicate and are expressed as mean ± SD (error bars). ***, **** significant difference between treatment and control group (p < 0.001 and 0.0001, respectively).

Figure 6.

RGF-containing herbal gel; A: at day 0 and B: after the heating-cooling test.