Gas Chromatography–Mass Spectrometry Profiling and DPPH Radical-Scavenging Capacity of the N-Hexane Extract of Arnebia Linearifolia A.DC. (Boraginaceae): First Report of Shikonin and Its Esters in the Species

Abstract

The genus Arnebia (Boraginaceae) is a recognised source of shikonin-type naphthoquinones, but pharmacognostic attention has concentrated on a small number of slow-growing perennials, several of which are now threatened in the wild. Arnebia linearifolia A.DC., an annual with a wide native distribution across Iraq and South-West Asia, has not previously been chemically profiled. In this study the lipophilic constituents of A. linearifolia were recovered by cold maceration in n-hexane and characterised by gas chromatography–mass spectrometry (GC-MS), and the radical-scavenging capacity of the resulting extract was measured by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. Maceration of 20.0 g of dried plant powder in n-hexane (1:10 w/v, 24 h, 22–25 °C) gave 0.770 g of a lipophilic extract, a yield of 3.85% (w/w). GC-MS resolved twelve constituents accounting for 94.62% of the total integrated peak area. Fatty acids and their esters formed the largest fraction (47.10%; linoleic acid 18.65%, palmitic acid 14.20%, oleic acid 11.40%, ethyl palmitate 2.85%), followed by naphthoquinones (31.00%; shikonin 15.80%, acetylshikonin 9.75%, isobutyrylshikonin 5.45%), phytosterols (9.25%; β-sitosterol 7.10%, stigmasterol 2.15%), squalene (4.30%), α-tocopherol (1.72%) and tetracosane (1.25%). Combining the extraction yield with the peak-area data places the shikonin content of the dried plant material at approximately 6.08 mg g−1 and total naphthoquinone at approximately 11.93 mg g−1. The extract scavenged DPPH radicals in a concentration-dependent manner, from 18.42 ± 1.15% at 6.25 µg mL−1 to 91.15 ± 1.65% at 200 µg mL−1, giving a half-maximal inhibitory concentration (IC50) of 26.85 ± 1.12 µg mL−1 against 8.82 ± 0.35 µg mL−1 for ascorbic acid. These results constitute the first report of shikonin, acetylshikonin and isobutyrylshikonin in A. linearifolia and identify the species as a potentially sustainable annual source of shikonin-type naphthoquinones. Because these compounds are thermolabile and only semi-quantitatively determined by GC-MS, the reported proportions require confirmation by HPLC with photodiode-array detection.