PHYTOCHEMICAL CONSTITUENTS AND ANTIOXIDANT ACTIVITY OF THE SEEDS OF CUCUMEROPSIS EDULIS (CUCURBITACEAE) FROM MOMO IN GABON

This paper describes results of phytochemical analyses and antioxidant activity of the seeds of Cucumeropsis edulis . For phytochemical analyses, extraction was performed with the same powder using different solvents: Acetone the first, ethyl acetate the second and methanol the last. Aqueous extract was used for evaluation of antioxidant activity. The results obtained show the abundance of polyphenols, gallic tannins and alkaloids in all extracts. Reducing compounds are present in ethyl acetate and acetonic extracts and leucoanthocyans are present in methanolic extract only. The results (IC 50 = 2249± 87,03 µg/mL; AAI = 0,022 ± 0,001) of evaluation of antioxidant activity are very low. coumarins, flavones, flavanones and flavanols are absent. These results suggest that the polyphenols of this seeds are in majority gallic tannins. Despite the low antioxidant activity attributed to the absence of many groups of flavonoids and catechic tannins, the use of seeds of Cucumeropsis edulis in the diet or herbal medicine could protect populations against many diseases.

This paper describes results of phytochemical analyses and antioxidant activity of the seeds of Cucumeropsis edulis. For phytochemical analyses, extraction was performed with the same powder using different solvents: Acetone the first, ethyl acetate the second and methanol the last. Aqueous extract was used for evaluation of antioxidant activity. The results obtained show the abundance of polyphenols, gallic tannins and alkaloids in all extracts. Reducing compounds are present in ethyl acetate and acetonic extracts and leucoanthocyans are present in methanolic extract only. The results (IC50 = 2249± 87,03 µg/mL; AAI = 0,022 ± 0,001) of evaluation of antioxidant activity are very low.

…………………………………………………………………………………………………….... Introduction:-
Cucumeropsis edulis (Cucurbitaceae) commonly called cucumber for many people in Africa (Savadogo et al., 2011; pl@nte use, 2020) is a plant used for food, medicine and other purposes. Its leaves are lobed and convenient to purge infants (Raponda and Sillans, 1995). The fruits are cylindrical and contain many small, flat and oval seeds (Savadogo et al., 2011;Raponda and Sillans, 1995). These seeds, when roasted and crushed, are used to prepare many dishes and to extract vegetable oil (Raponda and Sillans, 1995;Adodo et al., 2015;Ndola, 2015). Consumption of vegetables is recommended for the good health of human beings. Indeed, vegetables contain essentials elements needed by organism such as secondary metabolites which are many biological properties: anticancer, antidiabetic, antihypertensive, anti-inflammatory, analgesic, antioxidant, etc. (Yété et al.,2015;Mangambu et al.,2014;Adaramoye et al.,2005). Studies into free radicals have shown that foods rich in antioxidants can prevent to cardiovascular diseases, cancers and neurodegenerative diseases (Narenda et al.,2010). Despite the consumption of food prepared by the seeds of Cucumeropsis edulis, chemical studies are not available in Gabon. The interest of this study is to determine the phytochemical composition and the antioxidant activity of these seeds and then to evaluate its therapeutic potential.
The mixture of 30 g of dried powder of seeds and 150 mL of distilled water was heated in reflux for 1 h. After cooling at room temperature and filtration through Whatman No. 1 filter paper, the aqueous extract (Mefouet et al., 2021) was lyophilized and stored in the refrigerator at 4°C until evaluation of antioxidant activity.

Phytochemical analyses
Phytochemical screening is a qualitative analysis, based on staining or precipitation reactions. It permits identification of secondary metabolites. The analyses were released using standard methods with small modifications. The extracts were tested for the presence of polyphenols, flavonoids, tannins, alkaloids, saponins, coumarins, reducing compounds, sterols and triterpenes. Flavonoids 1 mL of 1% ammonia solution was added to 4 mL of filtrate. Yellow coloration indicated the presence of flavonoids (Mahdjouba, 2015). For differentiate flavonoids, the mixture of 5 mL of filtrate, 5 mL of hydrochloric ethanolic solution, 1 mL of isoamylic alcohol and some magnesium chips was prepared. Appearance of orange pink coloration indicated the presence of flavones. Flavanones were detected by purplish pink coloration and red coloration indicated the presence of flavanols. The cyanidin reaction without magnesium and after heating for 10 minutes in the water bath showed a cherry-red color indicating the presence of leucoanthocyans (Daira et al., 2016).

Sterols and triterpenoids
To 2 mL of filtrate were added a few drops of concentrated sulfuric acid. Observation of a purple coloration indicated the presence of triterpenes and green coloration for sterols (Daira et al., 2016).
Coumarins 3 mL of 10% sodium hydroxide solution was added to 2 mL of filtrate. After shaking the mixture, the appearance of a yellow color indicated the presence of coumarins (Daira et al., 2016).

Saponins
The mixture of 3 mL of distilled water and 2 mL of filtrate was shaken vigorously for 15 seconds. The observation of persistent foam for 20 minutes indicated the presence of saponins (Ano et al., 2018).

Reducing compounds
The mixture of 1 mL of Fehling's liqueur and 2 mL of filtrate was heated in a water bath for 15 minutes. The formation of a brick-red precipitate indicated the presence of reducing compounds (Ano et al., 2018).

Evaluation of the antioxidant activity
The antioxidant activity index (AAI) was determinate according to the radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) method described by Scherer and Godoy (Scherer and Godoy, 2009;Nsi et al., 2019). Standard DPPH solution containing 10 mg of DPPH was prepared in 100 mL of ethanol. Different concentrations of extracts were prepared in ethanol ranging from 0.781 µg/mL to 100 µg/mL. 100 µL of standard DPPH solution was mixed with 100 µL of each extract solution. Absorbance was measured at 517 nm after 15 min incubation at room temperature in the dark. Ascorbic acid (vitamin C) was used as reference. Measures were released in triplicate. The scavenging activity of DPPH radical was calculated by using the following equation: RSA is the percentage of free radical scavenging activity, Abs(control) the absorbance of DPPH radical + ethanol and Abs(sample) the absorbance of DPPH radical + sample extract or standard. The antioxidant activity is expressed as IC50 which is the measure of concentration in µg/mL of extract that inhibits 50 % of DPPH radicals. The antioxidant activity index was calculated using the following formula:

Results and Discussions:-
The results of the phytochemical characterization of the crude extracts of the seeds of Cucumeropsis edulis are given in Table 1. These results show the presence of polyphenols, alkaloids, triterpenes and gallic tannins in all extracts. Leucoanthocyans are present in the methanolic extract and reducing compounds in the ethyl acetate and acetonic extracts. Contrary to the results obtained in Benin (Yété et al., 2015), coumarins, catechic tannins and sterols are absent in all extracts like flavones, flavanones and flavanols. This difference could be explained by environmental, climatic and seasonal factors (Yété et al., 2015;Daddona et al., 1976). The absence of reducing compounds in methanolic extract shows that the quantity of these compounds is low in the seeds. All reducing compounds were then extracted by ethyl acetate and acetone. The absence of many groups of flavonoids suggests that polyphenols of this extract are in majority gallic tannins.  (Yété et al., 2015). Finally, leucoanthocyanes have a diuretic (antihypertensive) action (Yété et al., 2015). The presence of these phytochemicals in the seeds of Cucumeropsis edulis extracts could prevent risks against pathologies such as cardiovascular diseases, hypertension, diabetes and cancer.

(++) abundant; (+) present; (-) absent
The results of the evaluation of the antioxidant activity by the anti-free radical activity index (IAA) method are given in Table 2. The index of the anti-free radical activity is 0.022 ± 0.001. According to this index, the antioxidant activity can be considered low (IAA < 0.5), moderate (0.5 < IAA <1), high (1 < IAA < 2) and very high (IAA> 2) (Scherer and Godoy, 2009). The antioxidant activity of seeds of Cucumeropsis edulis is then low despite the abundance of polyphenols. These results agree with those described in the literature (Yété et al., 2015). The low antioxidant activity is in according to the phytochemical screening through the absence of many groups of flavonoids. Similarly, the low antioxidant activity could be explained by the absence of condensed tannins (Ba et al., 2010). indeed, catechic tannins have a stronger antioxidant activity than gallic tannins.

Conclusion:-
The phytochemical characterization of the extracts of Cucumeropsis edulis seeds identified polyphenols, alkaloids, gallic tannins, reducing compounds, triterpenes and leucoanthocyanins. These compounds have known to have many biological properties. However, saponins, catechic tannins, sterols, coumarins, flavones, flavanones and flavanols are absent. These results suggest that the polyphenols of this seeds are in majority gallic tannins. Despite the low antioxidant activity attributed to the absence of many groups of flavonoids and catechic tannins, the use of seeds of Cucumeropsis edulis in the diet or herbal medicine could protect populations against many diseases.