1. Introduction
Date palm (
Phoenix dactylifera L.) is cultivated in tropical and subtropical region in North Africa & Middle Eastern countries and play an important nutritional, social and economic role for peoples of these regions [
1]. In contrast to the most fruits, date fruit maturation goes through four stages, Kimri, Khalal, Rutab and Tamer, and generally it harvested in Tamer stage [
2]. The ripe date fruit consists mainly of three parts flesh (pulp) (85.7%), seed (14.25% and skin (0.05%) and consumed fresh, dried or frozen. The flesh is rich source of carbohydrates, dietary fiber, vitamins, mineral and energy with low content of lipid and protein, depending on ripening stage [3-6]. Both date fruit and seed were reported to an excellent source of polyphenols, flavonoids, sterols, glycosides, lycopene and anthocyanin [7-10]. Several studies had shown that date fruits and seed had protective effect against obesity, hyperlipidemic, diabetic, inflammatory, fatty liver, cardiovascular disease [11-15] .
Saudi Arabia is one of the leading country in date production and export of dates, with annual production exceeding 1.5 million tons from around 500 different date varieties that represent 17.0% of world production [
16].Ruziez variety is mainly grown in Al-Ahsa region, Eastern part of the kingdom and represent 69.0% of Ruziez trees in the kingdom with around 34 thousand palm trees [
17]. Compared to the Deglet Noor, khalas , Sukkari and Ajwa varieties, Ruziez is less popular and consider second grade variety. Therefore, its utilized in of date syrup industry, that generate a huge waste or by-product such as the date pomace, a mix of seed, skin and part of pulp. The agro-industrial waste is known to be harmful to the environment by increase the production methane and carbon dioxide gases and polluting water and soil. The developed countries spend billions of dollars to treat or decrease the risk of agricultural and food waste that harm to human, animals and environment. The date -industry wastes or by-products is an excellent source of fiber and bioactive compounds that can be utilized as functional foods ingredient which is beneficial to human health [
18,
19]. The selection of the solvent polarity and efficient of extraction methods is very important for extraction of bioactive compounds from plants source, because of its structural complexity and diversity [
20]. The beneficial effect of the phenolic compounds with antioxidant activity on the tissues, depend on its concertation in food and their bioaccessibility and bio-availability after ingestion [
21]. These bioactive compounds are bound to dietary fiber, proteins, and carbohydrates within food matrix, therefore it had to be hydrolyzed and released by the enzymes and make them bioavailable for absorption in the human intestine [22-24]. The Ruziez date pomace, the by-product of date syrup industry is used in animal feeding or disposed in landfill. However, to the best of our knowledge, the date syrup industry waste or by-product has not been extensively. Therefore, the purpose of this investigation was to evaluate the proximate composition and total polyphenolic compounds and antioxidant activity of Ruziez date pomace using different solvent system and to investigate the impact of in vitro gastrointestinal digestion on the bioavailability and stability of phenolic components and antioxidant activity of date by-products.
4. Discussion
The moisture, fat and protein, content of the Ruziez date pomace were comparable to that of Manify, Sifri and Selag date cultivars press cake [
35], but lower than that those of the Iranian date pomace [
36]. In contrast, the carbohydrates content was higher than that reported by [
36] probably due to the removal of date seed in this study, that is known to be higher fat content [
37]. The minerals content of the date pomace was comparable with that of Ruziez date fruit [
38], but lower than those of Barhee, Lulu and khulas date pomace [
39]. Furthermore, the Ruziez date pomace powder is found to be rich in total fiber with the majority being insoluble since most of the soluble fiber was washed off during date syrup preparation. The total fiber of the Ruziez date pomace is higher than those found in date pomace of Barhee, Lulu and khulas by- product of syrup/dibs production [
39]. These variations could be attributed to differences in genetic makeup, growing conditions, soil mineral content, fertilizers, irrigation, maturity stage and the processing methods [
2]. These results indicate that Ruziez pomace could be considered an excellent source of dietary fiber. Studies have shown the functionality and physiological impacts of dietary fibers, with Soluble fibers enhancing the viscosity that have positive effect on decreasing plasma lipid and glycemic response, while the Insoluble fibers, that are particularly abundant in date pomace enhance fecal volume and decrease intestinal transit [
40].
In This study, pure methanol and ethanol as solvents showed superior extraction efficiencies of total polyphenols and flavonoids with varying antioxidant activity from Ruziez date pomace, probably due the presence of various phenolic compounds with varying polarity and chemical properties that may or may not be soluble in a particular solvent [
41]. Mohammed et al., [
42], reported that the composition of the solvents and polarity had a major effect in extraction of phenolic compounds due to interactions between the polar sites of the antioxidant compounds and the solvent. Similar to our study Sultan et al., [
42] found that the methanol extracts of medicinal plants had the highest content TPC and TFC with high antioxidant activity followed by ethanol. Several studies had shown that methanol was superior in extracting polyphenolic compounds from vegetable and fruits compared to aqueous extracts that showed low extraction efficiency [
43,
44]. The antioxidant activity data showed similar pattern to the phenolic and flavonoids compounds, with the methanol solvent showing a higher efficiency than the other solvents. In contrast, several studies investigated the effect of different solvent system on phenolic compounds and antioxidant capacity of different date varieties such as Ajwa date [
9], Tantboucht dates [
45], and wild date palm [
46], and reported that 70% acetone and, 50% methanol were the most effective solvents. Despite the difficulty of finding a single solvent to extract all phenolic compounds, the majority of the phenolic components of the plant samples can be extracted using a mixture (50–70%) of organic solvent and water, and the more polar the solvent, the greater the antioxidant activity of the extract [
47].
The results of recovery index agree of antioxidants contents and activity agree with those reported by Gullon et al. [
23] who reported decrease in TPC and TFC recovery index during gastric and intestinal digestion of pomegranate peel. Similar trends were observed during in vitro gastrointestinal digestion of carob flour [
33] and tomato [
48]. In general, the recovery of TFC was higher than those of TPC indicating the release bound flavonoids compounds from the date pomace matrix probably due to the small particle size that increase the surface area and enzymes action that may results in breaking the bounds between phenolic compounds and fiber, lipid, proteins and sugars. Takanawa and Hirota, [
49] reported that flavonoids attached to high molecular weight molecules compounds such proteins and carbohydrates could be liberated by digestive enzyme action, resulting in a large rise in their concentrations following gastric digestion. The low bioaccessibility in the stomach stage could be attributed to the limited solubility in gastric or to rapid destruction of a portion of the bioactive compounds due to the combination of pH and enzymatic actions [
50]. The recovery index of the antioxidant mainly DPPH and ABTS showed same pattern of phenolic compounds, with the mouth stage digestion showing the highest recovery while intestinal showing the lowest. The mouth digestion stage increased DPPH and ABTS recovery index by 130.2% and 109.2%, whereas gastric digestion decreased DPPH and ABTS recovery by 48.3 and 59.1%, and further decrease were observed during intestinal digestion to 31.8 and 31.3% respectively. The increase in the release of antioxidant compounds in gastric digestion from date pomace matrix could be attributed to enzyme hydrolysis the glycosylic bond between the phenolic compound with antioxidant activities and fiber and sugars.
The bioaccessibility is referring to the available phenolic compounds such as polyphenol and flavonoids for absorption in intestine that released from the food matrix by digestion enzymes. However, studies had shown that some polyphenolic compounds such as anthocyanin glucosides were also absorbed rapidly into blood stream [
51]. Fig.2b. Show the bioaccessibility indexes of the Ruziez date pomace powder TPC, TFC and IC
50 DPPH, and ABTS after gastrointestinal digestion. Polyphenol bioaccessibility was significantly increased in after intestinal digestion compared to the gastric stage, whereas, flavonoids showed is no differences between the two digestion stages. The results showed that bioaccessibility of polyphenol and flavonoid after gastric digestion were 23.07 and 23.57% while in the intestinal digestion the last stage were 35.22% and 23.57% respectively. Antioxidants bioaccessibility data mainly, IC
50 DPPH and ABTS after gastric stage digestion were 17.0 and 16.2% while after intestine stage were 19.98 and 17.32% respectively. The bioaccessibility of the date pomace phenolic compounds (35.22%) is in agreement with that of pomegranate peel 35.9% [
23], but lower than those of apples 55%, carbo flour 81.0%, cinnamon beverage 79.8.0%, and date pits 78.45 and apple bagasse 91.58% [
32,
33,
52,
53] .The flavonoids bioaccessibility is lower than those pomegranate peel and carbo flour [
23,
33].The low phenolic compounds bioaccessiblity of date pomace could be attributed to high fiber content and/or inability of enzymes to release from the bound compounds from the matrix or due to the trapping during digestion by increasing viscosity of the fiber matrix in upper intestine [
54].
The results of bioactive compounds stability in pomace during in vitro gastrointestinal digestion were in agreement with li et al. [
55] who reported that TPC and TFC of rice-tatary buckwheat composite were significantly increased after mouth digestion compared to undigested samples. In contrast, an increase the release of polyphenol compounds and flavonoids after the final stage of intestinal digestion compared to gastric stage were reported in cooked green lentil, red and yellow quinoa, cooked pulses, cooked green lentil and lentil [56-58]. The in vitro gastrointestinal digestion finding suggest that amylase enzyme may promote the release polyphenols and flavonoids from matrix that is complexed with fiber and sugars by glcosidic bond, whereas the decrease in release in intestinal stage could be attributed to high pH that promote complex formation with protein, lipids and fiber, that hinder the phenolic compounds extractability.
The results of stability of antioxidant activity in pomace during in vitro gastrointestinal digestion were similar to TPC and TFC data which were higher in chyme insoluble fraction compared to the soluble fraction. This finding agree with that of Gullon et al., [
23] who reported that pomegranate peel phenolic compounds and antioxidants activities were higher after mouth digestion compared to gastric and intestinal stages, they also observed that the chyme insoluble fraction (pellet) contain more bioactive compounds compared to chyme soluble fraction. Gunathilake et al., [
50] observed that the gastric stage has a significant affected the antioxidant capacity of selected edible green leaves following simulated gastrointestinal digestion and dialysis. The antioxidant activity is known to be related to the amount of phenolic and flavonoids present. However, the antioxidant properties of these phenolic compounds may change due to chemical transformations during gastrointestinal digestion. The chemical structure of phenolic, pH, and interactions between phenolic and dietary fiber, or proteins and other dietary compounds released during digestion are known to affect polyphenol solubility and availability, as well as antioxidant activity in the intestinal [
59].
The findings of correlation between TPC, TFC, and antioxidant activity agree with that of Carbonell-Capella et al. [
60] who found a strong correlation between polyphenolics and antioxidant activity of beverages based on exotic fruits mixed with oat fruits before and after in vitro digestion. Similarly, Chen et al. [
61] reported a high association between antioxidant activity and TPC of 33 fruits after an in vitro gastrointestinal digestion. In same way, Kriaa et al., [
47], observed an excellent correlation between the total antioxidant capacity and the total phenolic and flavonoid contents of the methanol extract of three date palm leaves.
The increase in phenolic compounds after digestion as shown by the increase number of the peaks could be due to release the bound compounds to proteins or fiber in the original matrix by enzymatic digestion and pH change. Similarly, Mosele et al., [
62] reported an increase number of the compounds (peaks) after in vitro gastrointestinal digestion compared to the initial sample or undigested sample of pomegranate products (juice, pulp and peel extract. Chandrasekara and Shahidi, [
63] observed increase in phenolic compounds were released after in the gastric and intestinal digestion stages and colonic fermentation of five grains varieties, they suggested that those compounds may be absorbed or exert their antioxidant effects in the small intestine due to their solubility in the digest. Further study would be carried out to identify, quantitate and determine the fate of these phenolic compounds during in vitro gastrointestinal digestion stages, using the standard of these phenolic compounds instead of using the National Institute of Standard and Technology Spectral Library and Wiley GC/MS Library
Conclusions
The results of study revealed that Ruziez date pomace is rich source of fiber and bioactive compounds such polyphenols and flavonoids. In vitro gastrointestinal digestion process showed the release of the bioactive compounds from the pomace matrix compared to undigested sample that are bioaccessible to exert antioxidant activities and prevent of oxidative stress in intestine that promote health benefits effects. These results showed the possible application of the Ruziez date pomace as the potential bioactive source ingredient in the food industry, in formulation of functional foods to promote health life.