The objective of this work was to investigate the cytotoxicological effect of the extracts (hexane, ethyl acetate, and n-butanol) of Cotula cinerea and Salvia verbenaca in addition to the essential oil of Cotula cinerea. These plants are widely used in the Moroccan traditional folk medicine. The cytotoxic effect was explored against two cancer cell lines, Vero and RD, using the colorimetric MTT assay. The obtained results showed that the cytotoxicity differed according to the used extract with an efficient effect of Cotula cinerea extracts compared to Salvia verbenaca. A potent cytotoxicity was thus observed for the Cotula cinerea hexane extract which inhibited the growth of RD cell line at the lowest IC50 value (57.21±3.43 µg/mL). This was followed by the ethyl acetate extract and the essential oil with moderate effects against RD cell line and showed IC50 values of 187.52±6.27 µg/mL and 173.05±4.46 µg/mL, respectively. On the other hand, different results were obtained and Cotula cinerea essential oil was the most cytotoxic with the lowest IC50 value (72.72±2.18 µg/mL) against Vero cell line. In the same conditions, higher concentrations were needed in the case of Salvia verbenaca extracts. The results of this study showed thus that Cotula cinerea essential oil and hexane extract showed significant cytotoxic effects against RD and Vero cell lines, respectively, and could be considered as novel source of antitumor agents. This study is expected to be beneficial for clinical and traditional applications for Cotula cinerea as a remedy against cancer and opens new perspectives for further investigations on other types of cancer cell lines.

Background: Cotula cinerea belongs to the Asteraceae family and grows in desert areas such as Moroccan Sahara. The use of this plant in Moroccan traditional medicine prompted us to investigate its chemical composition, its acute oral toxicity, its analgesic and antioxidative activities. Methods: Extraction was conducted by steam distillation for essential oil and by maceration using solvents (hexane, ethyl acetate, n-butanol) for other non-volatile compounds. Quantitative analysis of total polyphenols, procyanidins and flavonoids was conducted through spectrophotometric assays. Qualitative phytochemical composition of the essential oil was investigated by GC/MS analysis. Acute oral toxicity was tested at a dose of 2000 mg/kg in mice. Central analgesic effect was assessed in rat using tail flick and hot plate models and the obtained results were compared to morphine. Antioxidant activity of the essential oil and the obtained extracts was evaluated through 2,2-diphenyl-1- picrylhydrazyl (DPPH°) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assays and the obtained results were compared to TROLOX. Results: The obtained results showed that the studied extracts contained significant amounts of total polyphenols, flavonoids and condensed tannins. The phytochemical composition of the essential oil was predominated by thujone, eucalyptol and santolinatriene. The results of the acute oral toxicity showed that the tested essential oil and extracts were not toxic even at the highest dose of 2000 mg/kg. Experiments on analgesic activity showed that the administered extracts have a central analgesic effect. The highest effect was observed with the n-butanol and ethyl acetate extracts for both tail-flick and hot plate tests. The antioxidant activity of the explored extracts showed higher scavenging activities of the studied samples compared to TROLOX. Conclusion: Our results indicate thus that C. cinerea could be considered as a source of various secondary metabolites including terpenoids and polyphenols. Exploration of its biological activities showed that the plant essential oil and extracts possessed antioxidant and analgesic effects. Based on the results of this study, it is likely that extracts of C. cinerea could open perspectives for its use for pain relief.

Rationale Cotula cinerea of the Asteraceae family is a traditional Moroccan plant with various biological activities such as analgesic, cytotoxic and antioxidant effects which are often related to the presence of secondary metabolites. The present work aims to screen and identify the main phytochemicals compounds of Cotula cinerea extracts. Methods A method was developed that coupled a rapid and simple ultra‐high‐performance liquid chromatography system with both photodiode array and high‐resolution time‐of‐flight mass spectrometry detectors (UHPLC‐PDA/TOF‐HRMS) for the identification of the main secondary metabolites of three investigated extracts (hexane, AcOEt and n‐BuOH). Results A total of 30 phytocomponents pertaining to phenolic compounds and terpenoids have been detected, identified and quantified. Among these were previously reported free and conjugated coumaric and caffeic acids along with free and conjugated flavones and flavonols with kaempferol, quercetin, luteolin and apigenin aglycones. In addition, sulfated flavonoids were identified in the investigated extracts and are reported in this work for the first time in Cotula cinerea. The obtained results have been discussed in relation to the biological activities of the corresponding extracts. Conclusions This study proposed a practical strategy for rapidly screening and identifying secondary metabolites of Cotula cinerea and provided new information on the phytochemicals of this Saharan plant. This work has therefore provided useful results for further pharmacological studies and the design of new drugs based on this species and will facilitate the utilization of Cotula cinerea in the clinic and its safety evaluation. It is also hoped that the information presented here might stimulate further studies that will possibly lead to development of therapeutic agents from this plant.