Integrated in vivo Anti-Inflammatory, Antioxidant-Related, and Preliminary Acute Oral Toxicity Responses to Gracilaria gracilis Extract

Inflammation and oxidative stress reinforce one another and are important targets in the development of marine-derived pharmacological candidates. This study evaluated the in vivo anti-inflammatory and antioxidant-related effects of an ethanol extract of BRIN-authenticated Gracilaria gracilis and included a separate preliminary single-dose oral toxicity assessment. Dried seaweed (300 g) was macerated with 96% ethanol in three 24-h cycles at a 1:3 (w/v) ratio, producing 24.5 g extract (8.17%, w/w). Qualitative screening detected flavonoids, triterpenoids, steroids, alkaloids, and glycosides. Untargeted LC-HRMS profiling of the same extract generated 325 ion features and 51 candidate annotations distributed across five broad metabolite classes. In the efficacy arm, 25 male Wistar rats received vehicle, diclofenac sodium (10 mg/kg), or extract at 100, 200, or 400 mg/kg (n=5/group) before carrageenan induction. Four-hour paw-response inhibition was 53.7%, 69.1%, and 73.6%, respectively, compared with 76.9% for diclofenac. Serum thiobarbituric acid-reactive substances (TBARS) declined dose-dependently from 90.3±13.5 µM MDA equivalents in the vehicle group to 60.2±8.4, 44.8±5.8, and 37.9±4.5 µM. In the preliminary toxicity arm, male Wistar rats received vehicle or a single oral dose of 300, 2000, or 5000 mg/kg and were followed for 14 days. Group-level day-15 data showed increased AST and urea at the upper doses, accompanied by mild descriptive liver and kidney alterations. Because clinical-observation logs, diagnostic-kit details, and standardized blinded histopathological scoring were incomplete, these findings are interpreted as preliminary toxicity signals and cannot establish an LD50, a formal hazard category, or safety up to 5000 mg/kg. The results provide exploratory in vivo proof of concept for G. gracilis while identifying the need for quantitative standardization, mechanism-focused studies, and confirmatory safety evaluation.