INTRODUCTION:

Date palm (Phoenix dactylifera L), cultivated in the arid and semi-arid regions since the foot for its edible sweet fruits it is an important crop in south of Algeria. Dates have a high nutritional value due to its rich in carbohydrates (70–80%), Mineral salts, fibers, vitamins, fatty acids, amino acids and proteins [1].

The date fruit have shown to contain antibacterial and antioxidant activities [2], it is riches of gallic acid, o-coumaric acid, q-coumaric acid, ferulic acid, protocatechuic acid, sinapic acid and cinnamic acid [1]. These bioactive compounds which are synthesized by plants have various biological effects and cytotoxic properties [3,4].

Generally, there are four date fruit maturation stages khellal (Green stages), Besser (Yellow or orange stage), rutab (crisp stage), and tamr (full ripe stage). Many works have been done to valorize some Algeria date fruits at full ripe stage but the study of these fruits during ripening stage were limited to few work, Therefore, the objective of the present study was to determine the phytochemical compositions and characterize the antioxidant properties of date fruits because it is expected to increase the interest of the food industry in using at different maturation stage for human health products.

MATERIAL AND METHODS:

Plant material:

Date palm fruit (Ghars) was harvested at four maturation stages (Fig.1) from Ouargla region (Southeast of Algeria). Khellal stage was harvested in Joan, besser was harvested in August, routab in September and Tamr stage in November.

Preparation of crude extracts:

After the harvest, the date fruit was washed with tap water. The seeds were removed from date, the edible part was cut to small pieces with a sharp knife and dried in dark at room temperature. The samples (100g) of dried date at different maturity stages were extracted with 300ml methanol: water (4:1, v/v) at room temperature for 24 h. The extracts were filtered twice in a row using whatman No1. The alcohol was removed under vacuum at 40°C. The crude extract was kept in dark glass bottles at -4ºC until used. The storage conditions were the same for all extracts [5].

Total phenolic content (TPC):

The amounts of TPC in methanolic extracts were carried out with the Folin–Ciocalteu reagent using the colorimetric method [6]. 0.5mL of gallic acid standard or dilution extracts was mixed with 1.5mL of Folin-Ciocalteu reagent (10%) and 3mL of sodium carbonate solution (7.5%, w/v), the mixture was shaken and reacted in dark at room temperature for 30 min before measuring the absorbance at 765nm using a spectrophotometer. The results of TPC were expressed as mg gallic acid equivalents per 100g of dry weight (mg GAE/100g DW). All the measurements were taken in triplicate and the mean values were calculated.

Total flavonoid content (TFC):

The amounts of TFC in methanolic extracts were carried out with the aluminum chloride reagent using the colorimetric method [7]. A volume of 2 % AlCl3 ethanol solution was added to volume of extract or quercetin as standard. The mixture was shaken and reacted in dark at room temperature for 15 min before measuring the absorbance at 430nm using a spectrophotometer. The results of TFC were expressed as mg quercetin equivalents per 100g of dry weight (mg QE/100 g DW). All the measurements were taken in triplicate and the mean values were calculated.

Total tannin content (TTC):

The amounts of TTC in methanolic extracts were carried out using the colorimetric method [8]. 3mL of 4% ethanol vanillin solution and 1.5mL of conc. HCl were added to 0.4mL of extract or catechin as standard. The mixture was shaken and reacted in dark at room temperature for 15 min before measuring the absorbance of pink colour at 500nm using a spectrophotometer. The results of TTC were expressed as mg of catechin equivalents per 100g of dry weight (mg CE/100 g DW). All the measurements were taken in triplicate.

Ferric reducing powe (FRP):

The reducing power of the date extracts was estimated according to the method of Kumaran and Joel Karunakaran [9], with slight modifications. Each extract (1ml) was mixed with 2.5ml of phosphate buffer (0.2 mol/l, pH 6.6) and 2.5ml of potassium ferricyanide [K3Fe (CN)6] (1%). The mixture was incubated at 50◦C for 20 min. Then, 2.5ml of 10% trichloroacetic acid were added to the mixture. 2.5ml of solution was mixed with 2.5ml of distilled water and 0.5ml of 0.1% ferric chloride (FeCl3), and the absorbance was measured at 700nm. Increased absorbance of the reaction mixture indicates increased reducing power of the sample. Reducing power was expressed as millimolar (mM) ascorbic acid equivalents antioxidant capacity (AEAC). All analyses were carried out in triplicate.

Antioxidant Capacity by ABTS:

The ABTS assay was measured as described by Re et al [10] radical cation (ABTS^.+) was produced by reacting ABTS stock solution(7mM) with potassium per sulfate (2.45mM) in the dark at room temperature for 12–16 h. The ABTS^.+ solution was diluted with methanol (80 %) to an absorbance of 0.700±0.002 at 734nm. Diluted ABTS^.+ solution (975μL) was added to 25μL of sample or methanol for blank or Trolox solution as standard and the absorbance was recorded at 734nm. The results of samples were computed as Trolox equivalents (μM TE /g extract).

RESULTS AND DISCUSSION:

Total phenolic content (TPC):

The amount of total phenolic of Ghars date harvested at four successive stages of maturation was presented in Table 1. The results obtained from TPC assay demonstrate significant decreases of total phenolic content during the ripening of date fruits. The highest TPC was detected at Khellal stage with concentration of 980.54±22.31 mg GAE/100g Fw. The lowest concentration was found in Tamr stage (98.35±6.01mg QE/100g Fw). As fruit maturation progressed to Tamr stage the concentrations decreased, The decreases in TPC in date fruits were also reported in other work [11] and in other fruits; cherry [12] and peruvian camu camu CHIRINOS. The decrease in total phenolic level may be due to the oxidation of phenolic content by polyphenol oxidase [13].

Total flavonoid content (TFC):

The amounts of TFC showed that the highest level of flavonoids was found at Khellal stage (7.91±0.02 mg CE/100g Fw) then decreased progressively at Besser (6.16±0.07 mg CE/100 g Fw), Routab (4.39±0.16mg CE/100g Fw) and then at Tamr (2.24±0.03mg CE/100g Fw) stages. Our results are different to those reported by Odeh et al. [14] which found that flavonoids content increased with the stages of maturity. This difference should be attributed to the genetic variability.

Total tannin content (TTC):

The amount of total tannins was determined and the results show that Khellal stage was characterised by the highest level of tannin (683.76±25.26mg QE/ 100g DW) and the lowest value was found at tamr stage (8.75±0.28mg QE/100g DW). This result confirms the astringent taste of khellal stage. In another study, Elarem et al [2].reported that condensed tannins content ranged from 94.89±0.43 to 277.15±0.14mg CE/100g fw at Besser stage and ranged from 54.93±0.14 to 98.37±0.14 mg CE/100g fw at tamr stage. The difference between studies may be due to the genetic makeup, agricultural practices, or analytical procedures.

Ferric reducing power (FRP):

As it is shown in the table (2), antioxidant activity assayed by FRP showed that AEAC values varied from 17.95±0.42 to 220.92±2.60mM. Khellal stage exhibited the highest activity while tamr stage showed the minimum activity. The decreases of AEAC values in date fruit during the ripening was reported in other study of Elarem et al [11], in contrast Odeh et al they found that there was an increase during maturity in Madjhool, Barhi Baladi, Rotab, Zahidi, and Barhi Iraqi cultivars [14]. This difference may be due to the genetic variability. A highly positive relationship was found between AEAC values and total phenolic contents (R2 = 0.886). A strong correlation was also found between total phenolic and tannins of date cultivars in other reported studies [11,14].

Antioxidant Capacity by ABTS:

Comparing the four maturation stages we found that Khellal stage was characterized by the greatest level of TEAC (113.6±1.20 μMTE/g) While Routab stage by the less level (15.86±0.10μMTE/g) Our values are less than the values obtained by Al-Jasass et al [3] Who found that the values of TEAC varied from 341 to 1300𝜇mol TE/g dw. The variations in antioxidant activities of different date cultivars were expected due to variation in agroclimatic conditions, country of origin. In addition, maturity stage [15,16]. Highly positive relationship was found between TEAC values and TPC (R² = 0.998).

CONCLUSION:

The present paper showed that the date extracts demonstrated high phytochemicals composition and antioxidant activity potent during maturation. In that sense, it necessary to isolate and characterize this fruit at different maturation stage to identify their full composition and the exact compounds responsible for its bioactivity.

Tables:

Table 1. Total phenolic, flavonoid and Tannin contents of Ghars date fruit extract at different stages of maturity.

Stage	TPC (mg GAE/100 g DW)	TFC (mg QE/100 g DW)	TTC (mg CE/ 100 g DW)
Khellal	980.54±22.31	7.91±0.02	683.76±25.26
Beser	525.04±6.55	6.16±0.07	158.10±8.78
Routab	66.69±0.66	4.39±0.16	7.49±0.18
Tamr	98.35±6.01	2.24±0.03	8.75±0.28

Table 2. Results of total antioxidan capacity of Ghars date fruit extract at different stages of maturity.

Stage	AEAC (mM)	TEAC (μMTE/g)
Khellal	220.92±2.60	113.6±1.20
Besser	51.75±1.28	67.38±0.82
Routab	18.18±0.70	15.86±0.10
Tamr	17.95±0.42	17.11±0.15

Fig.1. Stages of maturation of date palm fruits (Ghars)

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Received on 21.05.2020 Modified on 16.06.2020

Asian J. Research Chem. 2020; 13(5):323-326.

DOI: 10.5958/0974-4150.2020.00062.0