Screening of chemical content specific to Hypericum species growing in different parts of Turkey by LC-MS/MS and method validation; investigation of their biological activities and chemometric evaluation

In this study, chemical and biological screening of 16 Hypericum species (H. triquetrifolium, H. scabrum, H. ternatum, H. perforatum, H. retusum, H. empetrifolium subsp. empetrifolium, H. helianthemoides, H. linarioides, H. orientale, H. pruinatum, H. lysimachioides var. spathulatum, H. hyssopifolium var. elongatum, H. confertum subsp. confertum, H. armenum, H. pseudolaeve, H. lydium) collected from 50 different sites were studied. Some of the studied Hypericum species were endemic and most of their parameters were not studied before. Within this context, their fatty acid, essential oil, aromatic and phenolic contents, antioxidant, cytotoxic, anticholinesterase, urease and tyrosinase inhibitory activities were investigated. Plants have been used in the treatment of various diseases since ancient times. In our country, which has a rich flora, plants used to treat various diseases are being studied chemically and biologically. Hypericum species belong to the family Clusiaceae (Guttiferae, Hypericaceae) and are represented by about 500 species in the world. In Turkey there are 97 species of this genus and 48 of them are endemic. Extracts from Hypericum species is commonly used in the treatment of many diseases in folk medicine; depression, stomach disorders (gastritis, ulcers), as an appetizer, jaundice, (as an antiseptic), external injuries as an inflammatory desiccant (with ointment or in olive oil), as an antioxidant additive in molasses, athlete's foot, gingivitis (by mouthwash), as an expectorant, sinusitis, intestinal inflammation, hemorrhagic fever, as fever-reducing. Hypericum species contain a large number of bioactive compounds. The main ones are; hypericin and pseudohypericin, known as the naftodiantron group compounds, hyperforine and adhyperforine flavonoids known as fluoroglucinol derivatives, xanthones and procyanidin compounds. Hypericum species exhibit various medicinal properties, both in terms of chemical content and biological activity. There are a lot of studies in literature about Hypericum species both in the world and in our country. However, the studies in the literature are not so comprehensive and detailed. In this study, ethanol and petroleum ether extracts of root and aerial parts of 16 Hypericum species mostly collected from Eastern Turkey were prepared. Qualitative and quantitative determination of compounds specific to Hypericum species in their ethanol extracts such as hesperidin, protocatechuic acid, chlorogenic acid, luteolin-7-glycoside, hyperoside, rutin, apigetrin, quercitrin, astragalin, quercetin, luteolin, , pseudohypercine and hyericine were determined by LC-MS/MS was performed and the developed method was validated. Fatty acid composition of petroleum ether extracts were qualitatively and quantitatively determined by GC-MS. The aroma contents of these species were determined by Headspace-GC-MS/FID system. In addition, volatile oils obtained by hydrodistillation using Clevenger aparatus were analyzed by GC-MS and compared with the aroma content. In addition to chemical studies, the cytotoxic activities of essential oils of the studied species against healthy fibroblast, cancerous breast (MCF-7) and cancerous colon (HT-29) cell lines have been investigated. Besides, the antioxidant, anticholinesterase activities and urease and tyrosinase inhibitory activities of essential oils and ethanol extracts of aerial and root parts of all species were investigated. Finally, the results of chemical studies were analyzed by chemometric techniques such as PCA and HCA. In general, when looking at the results of volatile oil analysis performed by GC-MS, it is noteworthy that in many of the samples studied, the α-pinene compound was found to be the major component, even if not major, at a significant level. It was also found that all the species studied were rich in monoterpenes. Trans-β-ocimene, α-pinene, caryophyllene, humulene, germacrene D, β-selinene, caryophyllene oxide and α-cadinol monoterpenes were predominant in Hypericum species. When we look at the results of LC-MS/MS, it can be said that in general, all species studied contain all the compounds studied and that they may be the source species for some components. Specifically, it was observed that Hypericum species contained high amounts of hyperoside, quercitrin, rutin and chlorogenic acid. It can be noted that, Hypericum species have high antioxidant potential in CUPRAC, DPPH free radical scavenging activity and ABTS cation radical scavenging activity. Furthermore, volatile oil samples of Hypericum species seem to have high anticholinesterase enzyme activity potential. It was also determined that antitryrosinase enzyme activities of essential oil samples of H. scabrum, H. retusum and H. linarioides species were quite high. Additionally, it is noteworthy that the volatile oil obtained from H. lydium showed low toxicity on living cells and high cytotoxic effect on MCF-7 and HT-29 cell lines.