DOI: 10.17632/4mwv9cwjmv.1
title: Antioxidant, anti-clotting and anti-platelet aggregation activities of polyphenols extracts recovered from olive mill wastewater of Olea europea L. cv. Arbequina

description: Olive mill waste water, a significant environmental issue in Mediterranean countries, contains polyphenols and flavonoids that are beneficial for health, protecting against harmful oxidants species linked to numerous cardiovascular disorders like arterial thrombosis, atherosclerosis, and thrombo-embolic diseases. 
The study aims to assess the antioxidant and antithrombotic activities of polyphenols extracts recovred from deffated olive mill waste water of Olea europea L var. Arbequina, which is obtained from a modern olive oil mill located in Setif wilaya (Algeria), by methanolic and ethanolic aqueous solution (80%). Initially, pH, water amount, dry matter percent, total polyphenols and total flavonoids were determined. Polyphenols profiles were determined by HPLC analysis. The antioxidant activities of hydro-alcoholic extracts were evalueted using DPPH radical scavenging, ABTS radical scavenging and reducing power assays. The anti-clotting effects of extracts, on plasma poor platlets (PPP), were determined using activated partial thromboplastin time (APTT), prothrombin time (PT) and fibrinogen quantitation assays, and their anti-platelet aggregation activity on plasma rich platlets (PRP) was assessd using agregometry.
The olive mill wastewater is a slightly acidic medium (pH= 4.56). The amounts of water and dry matter are 89.1 ± 0.12%, and 10.9 ± 0.51%, respectively. The total polyphenols of methanolic and ethanolic extracts are 157.9 ± 0,05 and 137.8 ± 0,03 mg GAE/g dried extract, respectively. The total flavonoids represent approximatively 1/3 of total polyphenols with a ratio of 28.70% for methanolic and 29.53% for ethanolic extracts. HPLC analysis of both extracts gave 20 peaks corresponding to different biophenols. The constituent’s identity was established by comparing the retention times of these peaks with those of standards. Depending of the type of extracts, it appears that they have have significant antioxidants activities with an IC50 ≥ 26.41 ± 0.31, ≥ 12.83 ± 0.49, and A0.5 ≥ 14.12 ± 1.78 for DPPH, ABTS, and reducing power assays, respectively. 
For anticoagulant activities, methanolic and ethanolic extracts were able to extend the APTT and PT times in a dose dependent manner. In fact, the clotting times are ≥ 57,31 ± 0,69 s, and ≥ 16,40 ± 0,49 s for APTT and PT, respectively, which are higher than Nadroparin time uses as standard anticoagulant. Both extracts do not have pro-fibrinolytic properties, which is demonstrated by the normal amount of fibrinogen in the assay medium. For anti-platelet aggregation activity, the methanolic and ethanolic extracts at a dose of 5000 µg in terms of dried extract inhibit the platelets aggregation, in presence of arachidonic acid as an inducer, with a percentages of 92 and 96% respectively.
The olive mill wastewater, pollutant, is a important rich source of bioactive compounds particularly polyphenols wich make it a good candidate for the development of antioxidant and anti-thrombosis drugs