Griess Assay Evaluation of Nitric Oxide Scavenging by a Multi-Extract Emulgel for Topical Steroid Withdrawal

Topical steroid withdrawal (TSW) is an iatrogenic dermatosis following prolonged topical corticosteroid use, with limited non-steroidal alternatives. This study developed a multi-extract emulgel containing Andrographis paniculata, Tinospora cordifolia, Curcuma longa, and Piper nigrum and screened its nitric oxide (NO)-scavenging activity. Standardized ethanolic extracts were incorporated into eight emulgels (F1–F8) using a 2³ full factorial design, varying A. paniculata (3–5%), T. cordifolia (4–6%), and C. longa (4–6%) with P. nigrum fixed at 10%. Physical characterization showed homogeneous oil-in-water emulgels with pH 5.26–5.90, spreadability 6.7–6.8 cm, and viscosity 12,560–17,120 cP. IC₅₀ values for all formulations exceeded the highest tested concentration (100 µg/mL) without reaching the concentration-response curve plateau, and are reported as > 100 µg/mL compared to 18.48 µg/mL for ascorbic acid. Descriptive ranking placed F4 first, followed by F7 and F8. Exploratory factorial analysis indicated that C. longa levels and the A. paniculata × T. cordifolia interaction drove IC₅₀ variation, showing that higher extract loading did not enhance activity. Because this cell-free assay reflects only direct chemical NO scavenging, findings are strictly preliminary and descriptive. Future validation requires independent runs across an expanded concentration range capturing the full upper plateau. F4 is tentatively proposed as the lead formulation for confirmatory testing and subsequent stability, cell-based, and in vivo studies.