OPTIMIZATION AND EVALUATION OF BIOACTIVE CHARACTERSTICS OF COMMERCIAL RUMEX ABYSSINCUS (MEKMEKO) TEA in Ethiopia

The physicochemical parameters, mineral composition, and nutraceutical properties of commercial “mekmeko” teas powder were analyzed in the present study. The pH of samples was slightly acidic (3.74 to 4.15), titratable acidity was low (0.094 to 0.155%) and sensory attributes (over all acceptability) were wide variable (5.9 - 6.69). The optimized selected commercial “mekmeko” tea obtained pH(4.05), titrable acidity(0.069%) ,acceptable sensory results (6.69), acceptable highest content of zinc (0.069 mg·L−1), magnesium (2.12 mg·L−1), potassium (62.34 mg·L−1),iron (0.28 mg·L−1) and calcium (17.19 mg·L−1). The nutrceticual property of optimized commercial “mekmeko” tea obtained acceptable highest content in total phenols (211.38 mg·L−1), total flavonoids (62.98 mg·L−1), total catechins (409.67 mg·L−1), and total anthocyanins (581.97 mg·L−1). Optimized “Mekmeko” teas showed the highest antioxidant capacity levels in DPPH (718.41 μmolTE·L−1), ABTS (607.62μmolTE·L−1), and FRAP (953.81μmolTE·L−1) assays. The results obtained in the present work give information to consumers for choosing flavored-colored ready-to-drink “mekmeko” tea based on the physicochemical, nutritional and nutraceutical properties.


Introduction
A spice may be defined as "any dried, fragrant, aromatic, or pungent vegetable or plant substance, in the whole, broken, or ground form, that contributes flavor, whose primary function in food is seasoning rather than nutrition, and that may contribute enjoyment or spiciness to foods or beverages" Although spices have been important for centuries in food preparation throughout the world, we see considerable difference in the patterns of spice use among cultures and countries. The indigenous knowledge and use of plants as spices and condiments (culinary use) is as old as the history of mankind [5][6][7][8][9] Achinewu et al. [10] pointed out that plants used as Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: 9 November 2020 doi:10.20944/preprints202011.0281.v1 spices and condiments are usually aromatic and pungent. Iwu [13]; Billing and Sherman [18] had also reported that spices and condiments owe these properties to the presence of varying types of essential oils. Many authors also mentioned the antiseptic and the preservative properties of these plants to the possession of the essential oils such as what is reported in the work of Macmillan [7]. Dziezak [11] again indicated the rich presence of essential oils and oleoresins determine the aromatic, flavoring, coloring and pungent properties of spices and condiments.. ( [14][15] Ethiopians have a long tradition of using spices and condiments for culinary as well as medicinal purposes. Here spices are important ingredients of foods and beverages of many ethnic cultures that are usually consumed in relatively large quantities and therefore can have significant health effects on a large sector of the Ethiopian population . [48][49][50] Ethiopia as an ancient country has its own unique tastes and cultural inclinations. One that stands out is the Ethiopians love of spices. Almost all the food and drinks taken by Ethiopians in their daily routine consists of one or more spices, herbs, etc. Ethiopia and condiments has a long history in the world. A piece of the root (rhizome) of Rumex abyssinicus is dried, chopped and boiled in water and is added to flavor tea or can be drunk alone as tea. This is either to provide flavor or for its medicinal purpose. [34] Amare Getahun [34] wrote about culinary use of R. abyssinicus. He explains that the cooked tuber of the plant is used to make a dye that gives yellow coloration to the clarified butter or other spice preparations. Rhizome of R. abyssinicus is pounded and added to tea or coffee and drunk. This is done to treat headache .Similar results have been described in the work of Tesfaye.
[37] But here the part of the plant used to treat headache is different from what is reported by the local community in the study area. In the study area the bulb (rhizome) is used to relieve the problem but in Tesfaye [37], the leaf is used instead. The mode of administration of the remedy is also different. Mathewos and Agize [43] mentioned the medicinal value of R. abyssinicus and its refreshment in humans. Health benefits of "Mekmeko" that contains phytochemical to reduce blood sugar, anti diabetics, prevent blood pressure& Cancer, has anti diarrhea and anti microbial effect and also rich minerals and vitamins . [44] Even though spices produced in Ethiopia, there are no processing technologies; value additions and small scale processing industries are not yet available. Therefore the market demand at locally is minimum and the awareness of this spice to use is minimum because of un known its function .This implies most of organic spices in Ethiopia underutilized and its productivity is low because of lack technologies i.e. value addition.
In this study would optimize the mixing ingredients and evaluated the physicochemical, nuterucetical and sensory attributes

Sample collection and preparation
Rumex abyssinicus, spices additives ('Kerefa', 'Kirunfud'& 'Hel') sample were collected from local market and in the highlands and low lands of Amhara region. After collecting, "Mekmeko' and other spice additives samples were washed and dried. Then dried samples were milled as tea standard of course size as powder form.

Tea formulation and processing
Different formulations were used for 'Mekmeko' tea making ( Table 1). The control tea(CST) was made using for comparable to the commercial one and 'Mekmeko'( 100%, 75%,50%,25%) with that of other spice additives mixture . The blending samples were stored and sealed polyethylene bags at room temperature until they would analyze.

Determination of Mineral Composition
Mineral analysis was done based on AOAC, 2008 methods using an Agilent Atomic Absorption 240FS spectrometer (Santa Clara, United States). Briefly, 5 mL of 3 M.HCl was added to 50 mL of tea sample and digested-evaporated at boiling temperature until 20 mL of sample will obtain.
Afterwards, samples was filtered and used for mineral analysis. The content of total flavonoids will evaluate based on the reaction of aluminum chloride. Briefly, 0.2 mL of phenolic extract will place in 3.5 mL of distilled water, followed by 0.15 mL of 5% NaNO2. After 5 min, 0.15 mL of 10% AlCl3 will added, and 5 min later, 1.0 mL of 1 M.NaOH will add. Reaction will leave for 15 min, and finally, the absorbance of sample will measure at 510 nm. Catechin will use as the standard (0 to 200 mg·L−1), and the results will express as milligrams of catechin equivalents per liter of sample (mgCatE·L−1).

Determination of Nutraceutical Properties
The

Sensory Evaluation of Tea Samples
Sensory evaluation was carried out using a 15 main panelist to assess the organoleptic attributes of the tea samples. The organoleptic attributes assessed were; the taste, the aroma, the flavor, the color, and the overall acceptability. The panelists were trained and selected randomly from the students of the university from Food process engineering department. They were made to carry out the organoleptic assessment under controlled environment to avoid biased results The panelists will be instructed to rate the breads based on 7-point hedonic scale ranging from 1=liked extremely to 7=disliked extremely.

Statistical Analysis
For the analysis of the data, a one-way analysis of variance (ANOVA) was performed using SPSS software version 16.0. When significant deference's was observed, a mean comparison was made by the Tukey multiple range procedure (p≤0.05).

Physicochemical Parameters
The pH and titratable acidity are analytically determined in separate ways, and each has its own particular impact on food quality [16]. Titratable acidity is a better predictor of the impact of acid content on flavor of food, and the pH is a better predictor of the ability of a microorganism to grow in a specific food [17]. On the other hand, sensory attributes are the first notable characteristic of a food and often predetermines our expectation. Natural and synthetic colors play several roles in foods and beverages. There were statistical differences between tea samples in physicochemical parameters ( Table 2). The values of pH in tea samples were slightly acidic and ranged from 3.74 to 4.15 while titratable acidity ranged from 0.0094 to 0.155 %. ( Table 2) The results of the present study show higher pH values than data reported by Lunkes and Hashizume [19], whom analyzed 11 commercial ready-to-drink flavored teas available in Brazil market and found pH values ranging from 2.89 to 3.41. In addition, the titratable acidity reported by these authors was higher than our observations, since they found values ranging from 0.193 to 0.325%. The differences in the pH and titratable acidity values between these authors and our study are attributed to the fact that commercial available ready-to-drink teas contain citric acid as acidifier and sodium citrate as pH stabilizer to extend the shelf life of product. Therefore the selected optimized mekmeko tea in treatment two (75% mekmeko tea) had pH value, 4.05 and Titrable acidity, 0.069.

Mineral Composition
Minerals are crucial for the interaction between genetic and physiological factors. If a dietary deficiency of these elements exists, it will lead to physiological and structural abnormalities that are preventable and which may be reversed by administration of the element. Calcium, magnesium, sodium, potassium, iron, and zinc are essential elements, while copper and manganese are trace elements [21].
There were statistical differences between tea samples as in minor and as in major elements (  A wide range of selected micronutrient in made tea has been found in literature. The difference in micronutrient content of made teas could be attributed to the tea produced in different growing areas of the world having high variation in micronutrient contents, which resulted in differential micronutrient uptake by tea plants [22]. There is no literature available for comparison with the current report as there are no studies on the evaluation of mineral content of commercially readyto-drink teas. Nevertheless, several studies have evaluated the mineral composition of prepared tea infusions. Reto et al. [23], prepared tea infusions by boiling 1.5 g of green tea in 250 mL of boiling water for 10 min obtaining levels of calcium (1.9 to 3.5 mg·L−1), potassium (92 to 15 mg·L−1), sodium (35 to 69 mg·L−1), and iron (0.02 to 0.12 mg·L−1). These values were lower than the mekmeko teas analyzed in the present study, but the content of manganese (0.5 to 1.9 mg·L−1) reported by these authors was higher than all our samples.

Nutraceutical Properties
Phenolic compounds are receiving interest because of the recognition of their antioxidant properties, their abundance in the human diet, and their probable prevention of several diseases Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: 9 November 2020 doi:10.20944/preprints202011.0281.v1 associated with oxidative stress [25]. In addition, it is of great interest from consumers, nutritional experts, and food science researchers to know the antioxidant capacity of the foods that we consume [26].
There were statistical differences between tea samples in all phenolic evaluations ( all samples were especially rich in total catechins as these compounds are the major phenolic group in tea infusions, accounting for 40% of total water-soluble solids in tea infusions [4]. The content of total catechins was higher than the content of total phenols, and although this behavior is very rare, it has been reported previously in some legumes by Xu and Chang [27] and Zia-Ul-Haq et al. [28]. The explanation of this behavior is that acid treatment of vanillin-HCl method has an effect on depolymerization of catechins, and they are quantified in more quantity than the phenolics quantified by Folin-Ciocalteu method where phenolics are not depolymerized. Other studies have reported higher values than ours, and this is because teas are prepared directly from tea product at laboratory scale without the thermal process (sterilization) and dilution that is carried on the commercial industry of ready-to-drink tea.
Although the polyphenols content in food label is not mandatory in the Official Mexican Standard for food labeling (NOM-051-SCFI/SSA1-2010), all the analyzed teas reported the content of these compounds in their labels, and this is may be to make an impact on the consumers about the functional food ingredients in the product.

Sensory evaluation
The commercial mekmeko tea among the treatments just after preparation were awarded sensory score of Tate, flavor, color and overall acceptability by the panelist were showed in below table.
The sensory scores of treatment 2&3 were significantly different with the advancement of Even though treatments 2&3 were statically not significant difference but the panelist were selected treatment "2" that is 75% of mekmeko ingredient mixed with 25% of spices. The sensory scores are given below in Table 5. Formulation and optimized of mixing flavored ingredients with mekmeko tea In the optimization of "mekmeko" tea beverage formulation mainly the quantity of spices in equal amount( "Kurifund", "Hel", "Kerefa") in mekmeko tea was optimized based on sensory evaluation. The sensory evaluation was based on four parameter color, taste, flavor and over all acceptability. For the optimization process different ratio of "mekmeko" with equal amount of spices (100%, 75%,50% 25% mekmeko) were employed. The average score obtained by each ratio are given in above table 5, in which the Treatment No. 2 (75% of "mekmeko" and 25% equal amount of spices( "Kerefa", "Kurufund", "Hell") obtained the best scores.

Conclusions
Because of the increasing interest of consumers in the consumption of mekmeko tea powder teas with different flavors, it is necessary to know about their physicochemical and nutrition characteristics. There were differences between commercial "mekmeko" teas powder samples in physicochemical parameters, mineral composition, and nutraceutical properties. The pH values 13 of tea samples were slightly acidic with values around pH 4.05; titratable acidity was low with values near to 0.1% in the selected treatment (75% mekmeko tea). the highest content of iron, zinc, magnesium, potassium, and calcium obtained in selected treatment.. In addition, the highest content in total phenols, total flavonoids, total catechins, and total anthocyanins were obtained in selected treatment (2) i.e 75% "mekmko" ingredients and showed the highest antioxidant capacity in DPPH, ABTS, and FRAP assays.. Finally, the results obtained in the present work give information to consumers for choosing commercial "mekmeko" tea powder based on the physicochemical, nutritional, and nutraceutical properties.