Pharmacognostic researches on Helichrysum armenium subsp. araxinum

In the present study, n-hexane, ethyl acetate, and ethanol extracts prepared from the above-ground parts of Helichrysum armenium subspecies araxinum, were investigated for their 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activities, tyrosinase and elastase enzyme inhibitions, and toxic effects on human glioblastoma multiforme (U87-MG) and human neuroblastoma (SH-SY5Y) cell lines. The correlation between these activities and the chemical profiles of the extracts was also investigated. The extracts were prepared by subjecting the plant material to two 24-hour macerations. The IC₅₀ values of extracts showing over 50 % enzyme inhibition were calculated. Cytotoxic effects of the extracts were determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) assay at concentrations of 100, 50, 25, and 12.5 μg/mL after 24, 48, and 72 hours of incubation. The chemical contents of the extracts were analysed by Ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) using 56 standard compounds. The enzyme affinities of the compounds were evaluated in silico using AutoDock 4.2 software. The ethanol extract (329.60 ± 0.45 mg TE/g DW) exhibited the highest scavenging effect against the ABTS radical, while the ethyl acetate extract (694.11 ± 1.98 mg TEAC/g) exhibited the highest scavenging effect against the DPPH radical. The highest inhibition on the tyrosinase enzyme was detected in the ethyl acetate extract with IC₅₀ = 156 ± 3.97 μg/mL, while the lowest inhibition was detected in the ethanol extract with IC₅₀ = 280 ± 2.94 μg/mL. The extracts did not exhibit significant inhibition on the elastase enzyme. At a concentration of 25 μg/mL in the U87-MG cell line, the n-hexane extract showed higher inhibition after 24 hours of incubation (92.1 %), while the ethyl acetate extract showed higher inhibition after 48 and 72 hours of incubation (71.3 % and 85.5 %, respectively). The ethanol extract showed significant inhibition of SH-SY5Y cells at all incubation times. The main compounds identified in the extracts were acacetin, quinic acid, astragalin, chlorogenic acid, apigenin, naringin, and luteolin. Acacetin, found in high amounts in all extracts, was detected most in the ethyl acetate extract (25.774 ± 0.41 mg/g). Docking studies exhibited that acacetin inhibits tyrosinase enzyme at an IC₅₀ value of 27.81 μM. This study is the first to determine the inhibition effects of this taxon on tyrosinase and elastase enzymes and on the U87-MG and SH-SY5Y cell lines. The plant's rich chemical profile and therapeutic potential indicate that it can be used as a source of natural raw materials for cosmetic and medical purposes.