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Chemistry Thesis and Dissertations

2020-11-12

PHYSICOCHEMICAL ANALYSIS AND

DETERMINATION OF SELECTED

METALS IN DIFFERENT BRAND OF

BOTTLED MANGO JUICE IN BAHIDAR

CITY, ETHIOPIA

TSEHAY, EBESTIE

http://hdl.handle.net/123456789/11603

Downloaded from DSpace Repository, DSpace Institution's institutional repository

BAHIR DAR UNIVERSITY

COLLEGE OF SCIENCE

DEPARTMENT OF CHEMISTRY `

MASTER THESIS ON

PHYSICOCHEMICAL ANALYSIS AND DETERMINATION OF SELECTED METAL S

IN DIFFERENT BRAND OF BOTTLED MANGO JUICE IN BAHIDAR CITY,

ETHIOPIA

BY

EBESTIE TSEHAY

ADVISOR

TIHITINNA ASMELLASH (PhD)

August, 2020

Bahir Dar, Ethiopia

PHYSICOCHEMICAL ANALYSIS AND DETERMINATION OF SELECTED METALSIN

DIFFERENT BRAND OF BOTTLED MANGO JUICE IN BAHIDAR CITY, ETHIOPIA

A THESIS SUBMITTED TO COLLEGE OF SCIENCE POST GRADUATE PROGRAMIN

PARTIAL FULFILLMENT OF THE DEGREE OF MASTER OF SCIENCE IN ANALYTICAL

CHEMISTRY

BY

EBESTIE TSEHAY

ADVISOR

TIHITINNA ASMELLASH (PhD)

August,, 2020

Bahir Dar, Ethiopia

APPROVAL SHEET

This thesis titled on €Physicochemical analysis and Determination of selected metals in

different brand of bottled mango juice in Bahirdar city, Ethiopia• by Ebestie Tsehay

approved for degree ofmasters ofScience inAnalytical Chemistry to the Graduate Program in

Department ofChemistry,College ofScience, Bahir DarUniversity.

Board of Examiners

Atinafu Guadie (PhD)

Name of External Examiner Signature Date

MearegeAmare(PhD)

Name of Internal Examiner Signature Date

Minaleshewa Atlabachew(PhD)

Name ofChairperson Signature Date

DECLARATION

This is to certify that the thesisentitled• Physicochemical analysis and Determination of

selected metals in different brand of bottled mango juice in Bahirdar city , Ethiopia•

submitted in fulfillment of the requirements for the award of the degree of master of sciences in

analyticalto the graduate program ofCollege ofSciences, Bahir Dar university is an authentic

work conducted by Ebestie Tsehayunder the supervision of Tihitinna Asmellash (PhD).

EbestieTsehay ______________ ______________

Student Signature Date

Tihitinna Asmellash (PhD) ______________ ______________

Advisor Signature Date

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ACKNOWLEDGEMENT

First and foremost, I want to thank God for helping me during all difficult times of my life and

giving methe strength to dothethesiswork.

My second deepest acknowledgement goes to my advisor Dr. Tihitinna Asmellash for effective

and intellectual guidance, support in fruitful ideas, consistent follow-up, provision of necessary

materials for the research, encouragement and supportthroughout the work devoting her

valuable time and effort, and in general interest on mythesis.

I am also grateful to the Collage of Science and department of chemistry forgive me thechance.

Finally, I would also like to thank my families whoencouraged and prayed for me throughout

the time of my life.

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TABLES OF CONTENT page

ACKNOWLEDGEMENT...............................................................................................................i

LIST OF TABLES..........................................................................................................................v

LIST OF FIGURES.......................................................................................................................vi

LIST OF ABBREVIATION AND ACRONYM..........................................................................vii

ABSTRACT.................................................................................................................................viii

1. INTRODUCTION...................................................................................................................1

1.1 Background of the Study.................................................................................................. 1

1.2 Statement of the Problem................................................................................................. 2

1.3 Objectives of the Study....................................................................................................2

1.3.1 General Objective........................................................................................................2

1.3.2 Specific Objective.......................................................................................................2

1.4 Significance of the study.................................................................................................. 2

2. LITRATURE REVIEW ..........................................................................................................3

2.1 Fruit.................................................................................................................................. 3

2.2 Mango fruits.....................................................................................................................3

2.3 Mango production in Ethiopia..........................................................................................4

2.4 Nutritional and health benefits.........................................................................................4

2.5 Quality of Fruit Juice........................................................................................................6

2.5.1 Physico-chemical Parameters......................................................................................6

2.5.1.1 pH..........................................................................................................................6

2.5.1.2 Electricalconductivity...........................................................................................6

2.5.1.3 Total Dissolved solids (TDS)................................................................................7

2.5.1.4 Acidity ...................................................................................................................7

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2.5.2 Metals..........................................................................................................................8

2.5.2.1 Macronutrients......................................................................................................8

2.5.2.2 Heavy metal...........................................................................................................9

2.5.2.3 Levels of Heavy Metals in Fruit Juices...............................................................15

3. MATERIALS AND METHODS..........................................................................................16

3.1 Juice sampling................................................................................................................16

3.2 Instruments and Equipment............................................................................................16

3.3 Chemicals and Reagents................................................................................................. 16

3.4 Sample Preparation and Analysis...................................................................................17

3.4.1 Determination of metals............................................................................................17

3.4.1.1 Digestion of Juice sample...................................................................................17

3.4.1.2 Instrument working conditions............................................................................17

3.4.1.3 Instrument calibrations........................................................................................18

3.4.1.4 Method performance and validation................................................................... 18

3.4.2 Physicochemical determination.................................................................................20

3.4.2.1 pH determination................................................................................................. 20

3.4.2.2 Determination of Total Dissolved Solids (TDS) and Electrical Conductivity....20

3.4.2.3 Determining Acidity............................................................................................21

3.5 Statistical Analysis.........................................................................................................21

4. RESULTS AND DISCUSSION............................................................................................22

4.1 Optimizations for Metal Content Determination...........................................................22

4.2 Instrument calibration....................................................................................................23

4.3 Levels of Metals in the four brands of mango juice.......................................................27

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4.4 Physicochemical Analysis..............................................................................................34

4.4.1 pH..............................................................................................................................35

4.4.2 Total dissolves solids of juice samples......................................................................35

4.4.3 Electric Conductivity (EC) of the samples................................................................36

4.4.4 Acidity value.............................................................................................................36

4.4.5 Correlation matrix among pH, EC, TDS, and TA physicochemical parameters......37

5. CONCLUSION AND RECOMMENDATION....................................................................39

5.1 Conclusion......................................................................................................................39

5.2 Recommendation............................................................................................................40

REFERENCES.............................................................................................................................41

APPENDIX................................................................................................................................... 51

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LIST OF TABLES

Table 1: Nutritive value of mango per 100g...................................................................................5

Table 2: Dietary intake of trace elements in the human body and acceptable limit in fruit juice15

Table 3: Operating conditions of ICP-OES..................................................................................18

Table 4: Optimization of Parameters for Wet Digestion of Juice Sample................................... 22

Table 5:Working standards and correlation coefficients of the calibration curves for

determinations of metals in mango juice sample...........................................................24

Table 6: RSD result of metalanalysisfor mango juice samples................................................25

Table 7: Method detection and quantification limits (mg/L) for Mango Juice............................26

Table 8: The Mean ± SD concentrations (mg/L) and significance value of metals were found in

mango juice sample........................................................................................................27

Table 9: Comparison of study metals concentration (mg/L) in mango juice with the reported

values.............................................................................................................................32

Table 10: pH, EC, TDS and Acidity results in juice samples from four brands...........................34

Table 11: Comparison of some physicochemical parameter in mango juice with Literature

Values............................................................................................................................37

Table 12: Correlation matrix among the physicochemical parameters of mango juices samples38

Table 13: Optimization of Parameters for Wet Digestion of Juice Sample by using HNO3 ........51

Table 14: Optimization of Parameters for Wet Digestion of Juice Sample by using HNO3 and

HCl................................................................................................................................. 52

Table 15: One way ANOVA test value for Physical parameter of four mango juice brand........53

Table 16: Post Hoc Tests for multiple comparison of mango juice..............................................54

Table 17: One way ANOVA test value for metal analysis of Ca, Cr, Cu, Fe, Mg, Mn, and Zn.. 57

Table 18: Tukey HSD multiple comparisons test for metal analysis of Ca, Cr, Cu, Fe, Mg, Mn,

and Zn............................................................................................................................59

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LIST OF FIGURES

Figure 1:Physiological roles of Mn (Li and Yang, 2018).............................................................11

Figure 2: Health effect of cadmium in humans (Sharma et al., 2015)..........................................14

Figure 3: Calibration curve of intensity verses concentration for Mn, Zn, Cd, and Pb................23

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LIST OF ABBREVIATION AND ACRONYM

ANOVA‚‚‚‚‚‚‚‚... Analysis of Variances

FAO ‚‚‚‚‚‚‚‚‚‚ Food and agricultural organization

ICP-OES‚‚‚‚‚‚‚‚ Inductively Coupled Plasma OpticalEmission Spectroscopy

SD ‚‚‚‚‚‚‚‚‚‚...

TA‚‚‚‚‚‚‚‚‚‚...

Standard deviation

Titrated acid

TDS ‚‚‚‚‚‚‚‚‚‚ Total dissolved solid

WHO ‚‚‚‚‚‚‚‚‚.. World Health Organization

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ABSTRACT

Mango juice has health and nutritional benefits-issue of uniformity. However, it may also

contain highlevel ofmetals or additives, causinga health problem and affect the quality of the

juice. Therefore, the mainpurpose of this studywas to assess the physicochemical

characteristicsand determine the metal contentof mango juicein Bahir Dar City. Four brands

of mango juice samples(3D, Prigat, Rani, and Texas)were collected and analyzed for

physicochemical( pH, TDS , EC and acidity value), essential metal(Mg, Ca, Cr, Ni, Mn, Zn, Fe

, andCu) and non-essential metal(Cd and Pb). Metalswereanalyzed by ICP-OES, andacidity

valuewas estimated bytitration. Statistical analysis(ANOVA) was alsodone for physicochemical

parameterand metal analysisof among four brandsof juices.Theobserved results ofquality

parameters of studiedmango juices werefound to have a pH range of 3.24€ 3.93, TDS of 571.6

€ 2088 mg/L, Electrical conductivity of 867.3€ 1374 µs/cm and acidity of 0.24€ 0.33 %, and

Metal content of calcium (0.40€ 69.50 mg/L), magnesium (6.1€ 36.4mg/L), chromium( 0.14

€ 0.16mg/L), manganese( 0.22€ 0.86mg/L), nickel (0.28mg/L) in texas brand but not detect

in other brand, iron ( 0.92€ 9.00 mg /L), copper (0.34€ 0.62mg/L), zinc ( 0.41€ 9.80 mg/L),

cadmium (0.01 € 0.02 mg/L ) in the three brand3D, Prigat and Rani respectively, and lead

doesnot detected inall samples. This study indicates the valuesof physicalparameters and

acidity were closeto literature valuewhereas, the obtained resultsof mostmetal (Mn, Cr, Ni,

Cu, Fe and Zn were aboveliterature value. The results of this studycan be taken as an

indication for the need of control in packing juice.

Keywords: mangojuice; physicochemical;essential metal; toxic metal

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1. INTRODUCTION

1.1 Background of the Study

The manufacture of juices from fruits and vegetables is older than agriculture(Bateset al.,

2001). Fruit drinks are popularly used in most urban households.Historically, the utilizationof

fruit juices began with consumption of orange juice, as a source of vitamin C to prevent scurvy.

However, today markets are flooded with a large variety of juicessuch as:mango, apple, guava,

litchi, grape, pineapple(Gupta and Gupta, 2008). The main reason for increased consumption is

changing lifestyles and rising level of health consciousness among consumers. Theyhave

health benefit and are well consumed for their nutritivevalue, mineral and vitamin contents

(Bikila Wedajo, 2019). These functional components are protective against degenerative and

chronic diseases such as cancer and cardiovascular diseases(Dragstedet al., 1993). for these

reasonrecent dietary guidelines have recommended high fruitconsumption(Ackahet al., 2014).

Mango juice is considered as the most preferred non-alcoholic beverage worldwide to all age

groups(Amin et al., 2018). They have high nutritive value,vitamins, carotenoids, minerals, and

metals(Islamet al., 2015, Tantratianet al., 2018). Phytochemical & antioxidantcomponents of

Mango Juice are so beneficial to prevent many diseases along with cancerand heart disease

(Khanam et al., 2018). They have been considered as a delicious, nutritious popular drink.

However, apart from their advantage, they should have acceptable physicochemical

characteristics andfree fromchemical contaminateor toxic metals. Therefore, the main aim of

this work was to assess the physicochemical qualityand the metal contentof mango juices

collected from super market.

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1.2 Statementof the Problem

Mango juice is considered as the most preferred non-alcoholic beverage worldwide to allor any

age groups. It is an important source of macronutrientssuch as vitamins and minerals.Due to

that the consumption of mango juice continues to increase. The quality of juice depends on the

growing conditions,the processing and storageprocedures.Therefore, it is quite important to

know the physicochemical qualityand metal contentof the increasingly popularfruit juices

available in the market.Hence, determination of these parameters of some commercially packed

mango juices will help consumers to know the present scenarioor the condition ofprocessed

juice.

1.3 Objectivesof the Study

1.3.1 General Objective

The general objective of this study was to determine thephysicochemicalcharacteristicsand

Metal contentsof bottledmangojuice in fourbrands.

1.3.2 Specific Objective

ðØ To determinepH, EC,TDS, and Acidity values in four brandsof mango juice

ðØ To establish/develop a working procedure for the digestion of mango juice sample

ðØ To determine the levels of some essentialmetal (Mg, Ca, Cr, Ni, Mn, Zn, Fe, andCu) and

non-essential metal (Cd and Pb) in mangojuice sample collected from bahir dar super

market.

ðØ To compare the variation of data betweenbrands

1.4 Significance of the study

This study helps to have better understanding and awareness aboutquality of mango juice in the

market, andmoreover the studywill be used as baseline informationfor future researches on

safety of fruit juice. It helps to aware people about the health risks that possibly associated with

consuming ofpackedfruit juice.

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2. LITRATURE REVIEW

2.1 Fruit

The term ƒfruit„ is applied to acritical stagewith in the reproduction of botanical species

throughout the plant kingdom. Botanically, fruit is a plant organ, the principal biological purpose

which is to protect and eventually nourish the seed(s) as part of the natural plant propagation

(Bateset al., 2001).They are made up of chiefly cellulose, hemi-cellulose and pectin substances

that give them their texture and firmness.It is a greatimportance in the diet because of the

presence of vitamins and mineral salts. In addition, they contain water, calcium, iron, sulphur

and potash(Ihesinachi and Eresiya, 2014).The constituents of biological active substances in

fruit give beneficial effects on human health such as antioxidants, anticarcinogens, antimutagens

and antibacterial(Elmaet al., 2019).

2.2 Mango fruits

Mango (Mangifera indica L) is one of the 73 genera of the family Anacardiaceae and order

Sapindales(Mhamed and Ahmed, 2015).There are several species of genusMangiferathat bear

edible fruit. The majority of trees that are commonly known as mango belong to the specie,

Mangifera indica. Mangiferaoriginates from tropical Asia, with larger number of species found

in Borneo, Java, Sumatra, and the Malay Peninsula. The most cultivatedMangifera species, M.

indica (mango), originatesfrom India and Myanmar(Bally, 2006).

Mango is also known as the ƒking of fruit„ due to itsconsumptionin most parts of the world and

its numerous health benefits(Chataet al., 2018).Theycontainsome important nutrients for good

health such as fibre and vitamin C, and alsoaid the digestion of food inhuman body system

(Olaniyan and Obajemihi, 2014).They arean excellent source of fibre and bioactive compounds

such as provitamina carotenoids, vitamin C and phenolic that can easily be degraded during

processing and storage. Fruits and vegetables, being rich sources of nutrients, have been of

interest due to their potential health benefits in preventing several chronic diseases(Slavin and

Lloyd, 2012).

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2.3 Mango production in Ethiopia

Ethiopia is agro-ecologically diverse and has a total area of 1.13 million km2. Many parts

of the countryare suitable for growing temperate, sub-tropical or tropical fruits. For example,

substantial areas in thesouthern and south-westernparts of the country receive sufficient

rainfall to support fruits adapted to therespective climatic conditions.In addition, there are

also many rivers and streams which could be used to grow various fruits(Faris, 2016). The

climatic and soil conditions of Ethiopia allow cultivation of a wide range of horticultural crops.

The country has a vast potential for production of fresh fruit and vegetable varieties for domestic

and export markets for Negibourere countrieslike Djibouti, Somalia and the Middle East(Abebe

and Bizayehu, 2019). The major fruits produced and exported are banana, citrus fruits, mango,

avocado, papaya and grape fruits(Zeberga, 2010).

Ethiopia has a potential irrigable area of 3.5 million ha with net irrigation area of about 1.61

million ha, of which currently only 4.6 % is utilized(Honja, 2014). Mango is produced mainly in

Harari region, west and east Oromia, Southern Nations, Nationalities, and People's Region

(SNNPR) and Amhara(Bezu et al., 2015). More than 47 thousand hectares of land is under fruit

crops in Ethiopia. Mangoes contributed about 12.61% of the area allocated for fruit production

and tookup 12.78% of fruit production with comparedto other fruits growing in the country and

the annualconsumption of mango by the processing plant at full production capacity is 8.6 tones

which is only 1.8% of the current production of mango(Girmaet al., 2016). However, less than

2% of the produce is exported(Joosten, 2007). But, according to CSA (2013) cropping season

mangoes contributed about 14.21% of the area of land allocated for fruit production and holds

14.55% of quintals of fruits produced in the country(Honja, 2014).

2.4 Nutritional and health benefits

Mangos are an extremely nutritious and contain carbohydrates, proteins, fats,minerals,vitamins

(table 1).It has been well documented that mango fruits are an important source of

micronutrients, vitamins and other phytochemicals. Moreover, provide energy,dietary fibre,

carbohydrates, proteins, fats and phenolic compounds(Parvez, 2016).which area vital to normal

human growth, development andhealth. Thisnutrient plays a crucial role in human nutritionand

health. For instance, deficiency in vitamin A can lead to reversible night blindness and

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keratinization of normal mucous tissue of the eye, lungs skin and other ectodermic tissues. Lack

of vitamin B1 can cause beriberi (edema and heart hypertrophy). Again deficient in vitamin C

which is a vitamin for humans and primates results in scurvy(Ampomah-Nkansah, 2015).

Studies have attributed regular consumption of mango prevent certain type of cancer, better iron

assimilation, and high immune response. In addition, it containchemical componentsincluding

carotenoid, anthocyanin,chlorophyll and phenols that contribute to theoverall nutritional

properties of the fruit(Jolayemi, 2019).

Table1: Nutritive value of mango per 100g

Nutrients Ripe mango Unripe mango

Protein (g) 0.6 0.7

Fat(g) 0.4 0.1

Minerals (g) 0.4 0.4

Fiber (g) 0.7 1.2

Carbohydrates (g) 16.9 10.1

Energy (kcal) 74 44

Vitamin C (mg) 16 3

Total carotene (mcg) 2,210 90

Beta carotene (mcg) 1,990 -

Potassium (mg) 205 83

Sodium (mg) 26 43

Calcium (mg) 14 10

Iron (mg) 1.3 0.33

Phosphorous (mg) 16 19

Source:(Ravani and Joshi, 2013)

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2.5 Quality of Fruit Juice

Food quality is one of the most importantfactors fordifferentiatetheconsumer„s perception and

acceptance, attraction to, and purchase of the product. Fruitjuices are a highly attracted, tasty

food, and usually have exceptional nutritional qualities. However, they can be a potential source

of toxic elements, some of themhaving an accumulative effect or leading to nutritional problems

due to the low concentrations of essential elements, justifying the control of mineral composition

in juice(Deheleanet al., 2016). Several factors need to beconsidered forassessingthequality of

fruit juice. The composition of a fruit juice depends on the variety, origin and growing conditions

of the fruit, its quality and the processing and storage procedures(Ndife et al., 2013).Apart from

nutritive value of the juice, it should have acceptable organoleptic and physicochemical

characteristics aswell as free fromchemicalcontaminants.

2.5.1 Physico-chemical Parameters

2.5.1.1 pH

pH is a measure of the active acidity which is defined as the logarithm of the reciprocal of

hydrogen ion concentration in gram per liter (Ganapathyet al., 2019). They give also

information about food stability and preservation, and it can be used to retard microbial spoilage

that could happen in the presence of some pathogens such as bacteria, molds, and yeasts.This is

usually best growing rate in the pH range of 6.5-7.5. In consideration of the fact that almost all of

the pathogenic agents and most of deterioration bacteria cannot grow at pH < 4.5 (Karastogianni

et al., 2016).

2.5.1.2 Electrical conductivity

Electrical conductivity or specific conductance is a property of (food)material that measures

ability to conduct an electric current. Ohmic heating process is influenced, in a number of ways,

by electrical conductivity of the food material(Lamsal and Jindal, 2014).

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2.5.1.3 Total Dissolved solids(TDS)

Total Dissolve Solid (TDS) is one of the more important quality factors for most fruit juices.

Dissolved solids refer to any minerals, salts, metals, cations or anions dissolved in water, and

they are related to both sugar and fruit acid. They are main contributors of pectin, glycosidic

material, salt of metals and electrolytes sodium, potassium, calcium etc. TDS content is

significantly influenced by the combined effect of storage of maturity and ripening conditioned

are more important quality factors formost fruit juices (Tasnimet al., 2010).

2.5.1.4 Acidity

Acids are added to beverages and compose a…avor pro†le giving the beverage a distinctive taste

and provide a tartness and tangy taste that helps to balance the sweetness of sugar present in the

beverage; they are key factors in the taste of the beverage. Such as: Phosphoric acid is added to

cola drinks to impart tartness, reduce growth of bacteria and fungi, and improve shelf-life. Citric

acid, a substance naturally occurring in citrus drinks and added to many others, imparts a tangy

…avor and functions as a preservative(Reddyet al., 2016). Fruit juice is composed mainly of

organic acids and sugars, which are used as the main index of maturity and one of the major

analytical measures of flavor quality (Karadeniz, 2004). Organic acids are naturally found in

vegetables and fruits and maybe formed during processes like fermentation or may be added into

food during the manufacturing process. The organic acid composition of fruits is also of interest

due to its impact on the sensory properties. Citrus fruits, one of the important fruit crop groups,

are consumed mostly as fresh or as juice because of their nutritional value and special flavor.

Consumption of citrus juice is found to be beneficial in preventing coronary diseases and chronic

asthma (Abd-Ghafaret al., 2010).

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2.5.2 Metals

2.5.2.1 Macronutrients

Plant nutrients are classified as macronutrients and micronutrients. €Macronutrient• refers to

elements needed in large amounts, and €micronutrient• signifies nutrients that are essential to

plants but are needed only in small amounts. The micronutrients are Fe, Zn, Mn, B, Cu, Mo, Cl,

and Ni (Zekri and Obreza, 2003). Macronutrients basically perform a counter role in various

metabolic processes of plants and human beings, and are therefore required in large quantities for

their survival .On the basis of their functions; macronutrients have been classified into two

groups: primary macronutrients, i.e., N,P, and K, and secondary macronutrients, i.e., Ca, Mg,

and S(Tripathi et al., 2014).

Calcium is an essential macro elementof the human body. It is majordeposited in the bones and

teeth, where it provides structure and mechanical strength(Krupa-Kozak and Drabi‡ska,2016).

Inadequate intakes of calcium have been associated with increased risks of osteoporosis,

nephrolithiasis (kidney stones), colorectal cancer; hypertension disorders have treatments but no

cures (WHO, 2009). Owing to a lack of compelling evidence for the role of calcium as a single

contributory element in relation to these diseases, estimates of calcium requirement have been

made on the basis of bone health outcomes, with the goal of optimizing bone mineral density.

Calcium is unique among nutrients, in that the body„s reserve is also functional: increasing bone

mass is linearly related to reduction in fracture risk (WHO, 2009).

The major source of calcium in the diet is milk and milk products, providing over 40% of

calcium intake in adults, followedby cereals and cereal products providing 30% Main dietary

sources(Theobald, 2005) . Vegetables and fruits, cereals, dairy products, cereals are the rich

sourcesof calcium (Hanet al., 2003). Calcium was found to be higher in Mango juice, Multiunit

juice and pulpy orange juice and it is of important because of its role in teeth, bone, muscular

system and heart function (Brody, 1994).

Magnesium is the fourth most common mineral in thehuman body after calcium, sodium, and

potassium and is the second most common intracellular cation after potassium. It is involved in

more than 300 enzymatic reactions in the body, participating in the metabolism of glucids, lipids,

proteins, and nucleicacids, in the synthesis of H2 transporters(Laireset al., 2004) .Within the

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frame of a 70 kg individual, there is an average of 25grams of Mg in reserve with 53%in bone,

27 %in muscle, 19% in soft tissues, and less than 1% in the serum. Although serum Mg

concentration (SMC) is tightly controlled with a normal serum value of 75ˆ 95 mmol/L, some

research indicated that serum levels less than 85 mmol/L should be considered deficient

(Schwalfenberg and Genuis, 2017).

Primary magnesium deficit originates from two etiological mechanisms: deficiency and

depletion. Deficiency is due to insufficient intake, and depletion is due to deregulation of factors

controlling magnesium status such as intestinal magnesium hypo absorption, urinary leakage,

reduced magnesium bone uptake and mobilization, hyperglucocorticism, insulin-resistance and

adrenergic hypo receptivity. Secondary magnesium deficit results from various pathologies and

treatments: non-insulin dependent Diabetes mellitus, alcoholism, or ingestion of

hypermagnesuric diuretics, etc.(Laires et al., 2004). Mg plays an important role in protein

synthesis.Green leafy vegetables, fruits, cereals, legumes, sea foods arethe richest sources

of Mg (Welshet al., 1992).

2.5.2.2 Heavy metal

Heavy metals have density at least five times heavier than that of water and they are stable

elements that cannot be metabolized by the body(Bedassaet al., 2017).And are not

biodegradable and have the potential for accumulation in the different body organs leading to

unwanted side effects(Verma et al., 2016). They are found naturally in the earth and become

concentrated as a result of human activities. Common sources are mining and industrial wastes;

vehicle emissions; lead-acid batteries; fertilizers, paints and treated woods(Rajeswari and

Sailaja, 2014).

Heavy metalsareclassified as essentialand non-essential. Essential heavy metals and their roles

have been recently documented(Izahet al., 2016). They are beneficial to human health and other

living things. However, essential heavy metals can be toxic to living things when the

concentration exceeds the tolerable limit for the organisms. Non-essential heavy metals could be

toxic to cells of thebody even at low concentrations(Pb, Cd and As) (Anastácio et al.,

2018).Whereascopper, iron, chromium and nickel are essential metals sincethey play an

important role inbiological systems(Abd-Allah and Ismail, 2017).

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Copper is one of the essential elements that has been found to be important in our biological

systems that acts as a cofactor in more than 30 types of enzymes(Yahyaet al., 2016).Theyactas

a prosthetic group in several key enzymes and is thus essential for the structure and function of

thebone marrow and nervous system(Jaiser and Winston, 2010). Cu deficiencyhas been linked

to a variety to clinical signs, including pale coat, poor sheep fleece quality, anemia, spontaneous

fractures, poor capillary integrity, myocardial degeneration, hypomyelinization of the spinal

cord, impaired reproductive performance, decreased resistanceto infectious disease, diarrhoea

and generalized ill-health(Hefnawy and El-Khaiat, 2015).

Although copper homeostasis plays an important role in the prevention of copper toxicity,

exposure to excessive levels of copper can result in a number of adverse health effects including

liver and kidney damage, anaemia, immune toxicity, and developmental toxicity. Many of these

effects are consistent with oxidative damage to membranes or macromolecules. Copper can bind

to the sulfhydryl groups of several enzymes, such as glucose-6 phosphatase and glutathione

reductase, thus interferingwith their protection of cells from free radical damage(AKAKI et al.,

2018).

Manganese is an essential nutrient necessary for a variety of metabolic functions including

those involved in normal human development, activation of certain metalloenzymes, energy

metabolism,immunological system function, nervous system function, reproductive hormone

function, and in antioxidant enzymes that protect cells from damage due to free radicalsspecies

(Avila et al., 2013). It is a component of arginase, pyruvate carboxylase and superoxide

dismutase and plays a role as co-factor of certain enzyme systems.

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Figure1:Physiological roles of Mn(Li and Yang, 2018)

Dietary consumption is the primary route of Mn intake for majority of people. Drinking water

contains Mn levels ranging from 1mg/L up to 2 mg/L depending on the locations and

contamination. Inhuman daily diets, rice, nuts (hazelnuts, almonds, andpecans), whole grains

(wheat germ, oats, and bran)and legumes contain the highest levels of Mn, leafygreen

vegetables, tea, chocolate and seafood (clamsand mussels) are also abundant in Mn(Chenet al.,

2019). Manganese deficiency resulting from poor diet alcoholism and malabsorption causes

dwarfism, hypogonadism anddermatitis. Its presence in environment and consequent uptake by

humans causes pulmonary manifestation, fever, chills and gastroenteritis(Chataet al., 2018).

Zinc is an essential traceelementfor all forms of life(Bhowmik et al., 2010), it is essential for

growth and development, testicular maturation, neurological function, wound healing and

immune competence(Christianson, 1991). Zn is known to serve as the active center of

about 300 enzymes(Yanagisawa, 2004). The best food sources of zinc include meat based

products, especially the most abundant meat, such as chicken, lamb, beef, rabbit meat, oysters,

scallops, blackfish, animal liver, and so on. Othersources of Zinc include, mushrooms, day

�1�2

lily flowers, edible fungus, cabbage, black sesame, black rice, dates, hazelnut, ebony and other

vegetables, food crops and fruit(Devi et al., 2014).

Zinc is such a critical element in human health that evena small de†ciency is adisaster. Lack of

zinc leads toanorexia,loss of appetite, smell and taste failure, andother symptoms in humans

and may affect the immunesystem, triggering arteriosclerosis and anemia(Chasapiset al.,

2012).

Iron is an essential nutrient for humans and has a considerable impact on several physiological

processes such as production of oxidative energy, transport and storage of oxygen(Pynaert et al.,

2005). As a micronutrientit is a component of many enzymes which are involved in diverse

biochemical processes including respiration, photosynthesis and N2 fixation (Abbaset al., 2015).

Additionally Fe is an indispensable mineral needed for the production of hemoglobin, asa

cofactor for several enzymes necessary for proper functioning of brain, and for the immune

system and muscle(Beard, 2001). Furthermore, iron is used to help produce the connective

tissues in our body, some of the neurotransmitters in our brain, and to maintain the immune

system(Casiday and Frey, 1998).

Chromium is anubiquitous metal, occurring inwater, soiland biological systems.(Lewicki et

al., 2014). It occurs in almost all oxidation states ranging from-2 to +6. But in environment Cr is

mostly stable in trivalent and hexavalent form(Shekhawatet al., 2015). Hexavalent chromium

(Cr6+) is the second most stable form and a strong oxidising agent, especially in acidic media. It

is bound to oxygen as chromate (CrO42ˆ ) or dichromate (Cr2O7

2ˆ ) with a strong oxidative

capacity. This form of Cr crosses biological membranes easily, reacting with protein components

and nucleic acids inside the cell while being deoxygenated to Cr3+. The reaction with genetic

matter provides for thecarcinogenic properties of Cr6+ (Pechovaand Pavlata, 2007). Human

exposure to sufficiently high chromium concentrations would result in potential harm through its

toxic, genotoxic and carcinogeniceffects.Chromium is one of eight metals in the top 50 toxic

substances in the world in the data issued by the Agency for Toxic Substances and Disease

Registry (ATSDR), and WHO has classified chromium as carcinogenic to human beings

(Achmad and Auerkari, 2017) . Release of Crcompounds to the environmentis mainly due to

electroplating, leather tanning, metal finishing, corrosion control and pigment manufacturing

industries (Liuet al., 2011).

�1�3

Nickel is one of the essential elements found in abundance in the earth„s crust occurring at an

average concentration of about 75‰g/g(Poonkothai and Vijayavathi, 2012),however, excessive

amounts of Ni can be toxic(Nishidaet al., 2015). Higher concentration of nickel in human body

maycause respiratory diseases, birth defects, heart diseases and lung failure(Khan et al., 2016).

And may cause gastrointestinal and cardiovascular disorder, liver damage, and carcinogenic

effect (Tan et al., 2018).Although Ni is a recognized essential mineral nutrient element for

higher plants, its agricultural and biological significance is poorly understood. This is largely

because of the low levels thought to be needed by plants (about 1ˆ 100 ng g-1 dry weight) in

relation to the relative abundance of Ni in essentially all soils (.5 kg ha-1) (Bai et al., 2006). Ni

is usually accumulated in vegetative part of the plant body and it is mobile through the

plant structure, translocate and concentrated in the leaves but after ageningness of the

leaves it is moved to the seeds for accumulation (Cataldoet al., 1978).According to

ATSDR (1999),Lower content of Ni in vegetables and fruits sourcescan leadto increased

blood sugarlevel, hypertension and deficient growth in human but on the other hand the

increaseduptake of Ni in fruitsand vegetables can reduced the blood glucose level, difficulty

in breathing, nausea etc.

Cadmium is typically a metal of the 20th century, even though large amounts of this by-product

of zinc production have been emitted by non-ferrous smelters during the 19th century. Currently,

Cd is mainly used in rechargeable batteries and for the production of special alloys. Although

emissions in the environment have markedly declinedIn most industrializedcountries,They

remains a source of concern for industrial workers andfor populations living in polluted areas,

especially in less developed countries(Bernard, 2008). Cadmium is widely used in industrial

processes, e.g.: as an anticorrosive agent, as a stabilizer in PVC products, as a colour pigment, a

neutron-absorber in nuclear power plants, and in the fabrication of nickel-cadmium batteries

(Godtet al., 2006). It has toxic effects on many organs and tissues especially on kidneys, bones,

and lungs(Yanget al., 2018).

Cadmiumaccumulates in the human body affecting negatively several organs: liver,kidney,

lung, bones, placenta, brain and the central nervous system andother damages that have been

observed include reproductive, and development toxicity, hepatic, haematological and

immunological effects(Slezakova et al., 2014) . Cadmium is present at low levels in most foods,

�1�4

with commodities such as cereals, fruits, vegetables, meat and fish. These foodstuffs are minor

contributors to overall intake of cadmium, as they are eaten in relatively small amounts, and it is

unlikely to exceed the allowed dailyintake (ADI) forcadmium (Hassanet al., 2014).

Figure2: Health effect of cadmium in humans(Sharmaet al., 2015)

Lead is considered as one of the most hazards and cumulative environmental pollutants that

affect all biological systems through exposure to air, water, and food sources(Assi et al.,

2016).Such poisoning occurs from different kinds of human �related activities such as painting of

home, smoking �related activities,leaded petrol, contaminated food, and drinking water;

smelting; and especially from the industries, which have been carrying out manufacturing

processes(Al Muktadir et al., 2019). It is primarily the causative agent of disorders of the central

and peripheral nervous systems in humans(MurtiŠ et al., 2014).Lead toxicity is a particularly

insidious hazard with the potential of causing irreversible health effects. It is known to interfere

with a number of body functions and it is primarily affecting the central nervous, hematopoietic,

hepatic and renal systemproducing serious disorders(Flora et al., 2012). Acute toxicity is

related to occupational exposure and is quite uncommon. Chronic toxicity on the other hand is

much more common and occurs at blood lead levels of about 40ˆ 60 ug/dL. It can be much more

severe if not treated in time and is characterized by persistent vomiting, encephalopathy,

lethargy, delirium, convulsions and coma(Flora et al., 2012, Flora et al., 2006). Leadwas not

�1�5

dominant in all juice and nectar samples.Levels of lead in fruits and vegetables generally are

stringently regulated in the European Union (EU) by Fruit Juice Directive 2001/112/EC&

2009/106/EC. Additional source of lead in the diet is from food containers containing lead, e.g.

storage in lead-soldered cans, ceramic vessels with lead glazes and leaded crystal glass. Finally,

the past use of lead as a material for water pipes(Hassanet al., 2014).

2.5.2.3 Levels of Heavy Metals in Fruit Juices

Dietary exposure to heavy metals, namely Cd, Pb, Zn and Cu has been identified as a risk to

human health through the consumption of vegetable crops. At higher concentrations, they may

be toxic to the biota and could disturb the biochemical process and cause hazards(Ihesinachi and

Eresiya, 2014). Many metals such as arsenic (As), cadmium (Cd), chromium (Cr), nickel (Ni)

and their compounds may be mutagenic at levels above the maximum permissible limits.

Mutagens are capable of causing mutations in the DNA of an organism above the natural

background level. These mutations cause cancer. Some metals such as As, Cd and Pb can

volatilize during high temperature processing. These metals will convert to oxides and condense

as fine particulates(Smith, 1995).

Table2: Dietaryintake of trace elements in the humanbodyand acceptable limit in fruit juice

No. Essential trace elements Diet mg/day WHO/FAO (2010)

Acceptable Limit

(mg/kg)in fruit juice

1 Chromium 0.05ˆ 0.1 -

2 Iron 15.0 0.80

3 Zinc 8.0- 15.0 99.40

4 Nickel 0.4 0.14

5 Manganese 2.2- 8.8 0.30

6 Cobalt 0.3 2.00

7 Copper 3.2 0.05ˆ 0.5

8 Cd - 0.10

Source:(Anwaret al., 2014).

�1�6

3. MATERIALS AND METHODS

3.1 Juice sampling

Four types ofbottled mango juicesamples (Prigat, 3D, Raniand Texas) were collected from

supermarket found in Bahir dar city, Amhara region, Ethiopia. A total of 12 samples were

collected. For each type of juice three samples were collected and placed in analytical laboratory

for further treatment and analysis.

3.2 Instruments and Equipment

ICP-OES (Perkin Elmer Optima 8000),pH meter(25 CW MICROPROCESSOR), conductivity

meter (YSI PRO 30), Fumehood (Envair England), hot plate, beaker (50 and 100 ml), Funnel,

Whatman filter paper, volumetric flask (25 mL), measuring cylinder (10, 25 and 50 mL),

Burette,conical flaskand Erlenmeyerflask were used.

3.3 Chemicals and Reagents

All the reagents and chemicals used in this study wereof analytical grade. Hydrochloricacid

(36%-UNI CHEM), Nitric acid (69%-OXFORD LAB CHEM), NaOH (Alphax chemical

industry, India), phenolphthaleinIndicator,Standardsolutions of each metal (Mg, Ca, Fe, Cu,

Mn, Zn, Cr,Ni, Cd, andPb) wereused.Distilled waterand deionized waterwas used throughout

the experimentfor sample preparationand rinsing of the apparatus.

�1�7

3.4 SamplePreparation and Analysis

3.4.1 Determination of metals

3.4.1.1 Digestion of Juice sample

Digestion procedure was carried out according to Musa and Parmeshwal, (2018) with slight

modification. In order to analyze metals present in juice samples efficiently, different

procedures for sample digestion were carriedout and the one that consumed smaller reagent

volume, temperature with smaller digestion time and producedcolor less andclear solutions with

no residue and suspended matter was selected for the routine digestion of the samples. Based on

the selection optimizing procedure a 5 mL of Mango juice sample was accurately measured and

transferred in to 50 mL conical flask and added with 3 mL of (HNO3 ˆ HCl) acid mixture in a

volume ratio of 1:1. The mixture was heated to 90 ºC on hot plate, and the digestion continued

until no brown fumes evolved and solution becomes clear and colourless. The sample solution

was then cooled, diluted andfiltered with filter paper in toa 25 mL ofvolumetric flask with

distilled water.

Blank sample was prepared by taking 5mLdistilled water and adding 3mL of (HNO3 ˆ HCl)

mixture in it. Placed it on the hot plate and heated to 90ºC in it which will give a clearand

colorlesssolution. Then the solution was cooled filtered and diluted it up to a mark of 25 mL

volumetric flask.

3.4.1.2 Instrument working conditions

All metal analysis were carried out on an ICP-OES (Perkin Elmer Optima 8000).The instrument

was operated underthe conditions shown in Table 3.

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Table3: Operating conditions of ICP-OES

No Parameters Value

1 Plasma flow 8 min-1

2 Auxiliary flow 0.2 min-1

3 Nebulizer flow 0.7 min-1

4 RF power 1500 W

5 Pump flow rate 1 mL min-1

3.4.1.3 Instrument calibrations

The instrument used for analysis, ICP-OES, was calibrated using working standards for each

metal of interest. Triplicate readings were taken for each working standard solutions. The

correlationcoefficientsrangeobtained for the calibration curves for Mg,Ca, Cr,Mn, Fe, Ni, Cu,

Zn, Cd, and Pb were0.985-0.999, it confirming avery good positive correlation. Finally, the

procedure was used for the determination of elements in the sample solutions and digested blank.

3.4.1.4 Method performance and validation

ICP-OES is method of choice for analysis of heavy metals in food and pharmaceutical products

because of its low detection limits and its high degree of selectivity (Tanet al., 2018). Before

being used for quantitative analysis of metals ( Mg , Ca, Cr, Mn, Fe ,Cu, Zn ,Ni, Cd , and Pb)

in juice sample, ICP-OES was validated by determining some analytical parameters, namely

linearity , sensitivity ,precision, and accuracy.

�1�9

Determination of Detection and Quantification Limits

Method of detection limitis defined as the minimum concentration of analyte that can be

measured. In other words, it is the lowest analyte concentration that can be distinguished from

statistical variations in a blank, which usually correspond to the signal of blank three times the

standard deviation of the blank(Gerenfeset al., 2019). In this study, the detection limit of each

element was calculated as three times the standard deviation of the blank which is summarized in

(Table 6).Mathematically given by:

MDL=3‹ (1)

Where, ‹ standard deviation; MDL is method detection limit

Limit of quantification is the lowest concentration of the analyte that can be measured in the

sample. The limit of quantification is the same as the concentration that gives a signal 10 times

the standard deviation of the blank (Tanet al., 2018). The quantification limit of each element

was calculated as ten times the standard deviation of the blank mathematically given by:

�= �1�0 (2)

Where, ‹ standard deviation; MQL is method of quantification limit

Recovery Test

Recovery is one of the most commonly used techniques utilized for validation of the analytical

results and evaluating how far the method is acceptable for itsintended purpose. To determine

the efficiency of the method, a recovery study was carried out. This was done by spikinga

known concentrationof the metalsof interest(Mg, Ca, Cu, Cr, Mn, Zn, Fe, Ni, Cd and Pb)in to

juice samples prior to digestion and analyzed using after pretreatment and digestion(Ajai et al.,

2014). Spike recoveries were calculatedAccordingto the following formula:

�% �="�

�× �1�0�0�% �(�3�)

�2�0

Relative standard deviation (RSD)

Relative standard deviation is the parameter of choice for expressing precision of methods.

Precision is typically evaluated by measuring the values of relative standard deviation (RSD) of a

set of data. It is an analytical procedure expresses the closeness of agreement (degree of scatter)

between a series of measurements obtained from multiple sampling of the same homogenous

sample under the prescribed conditions(Tanet al., 2018). Relative standard deviation (RSD) can

be expressed as a percentage:

�= ��1�0�0�% �(�4�)

Where, is standard deviation; is mean

3.4.2 Physicochemicaldetermination

3.4.2.1 pH determination

pH meter (25 CW MICROPROCESSOR) was used for analysis the pH of mango juice sample.

Prior to using, it was calibrated using buffer solutionsacidic (4) and neutral (7).After calibration,

100ml of sample was taken in the beaker andfinally, the electrode ofthe pH meter was

immersed in a glass beaker containing the samples and reading were obtained from photo

detector of the pH meter. Where the reading was taken and recorded immediately for each

sample. The meter was rinsed with distilled water after and before each measurement. Each

sample was analyzed in triplicate reading in order to be sure that the recorded readings are

reliable.

3.4.2.2 Determination of Total Dissolved Solids (TDS) and ElectricalConductivity

Electrical conductivity and total dissolved solids (TDS) was analyzed using portable digital

conductivity meter (YSI PRO 30). Before measurement, the multi meter was calibrated using

Distilled water. 500mL of juice sample was taken in a glass tube and the meter was immersed in

a glass tube containing the sample and read directly on scale of sector. Then the meter gives the

accurate result of electric conductivity and TDS of every juice samples in unit µs/cm and mg/L

respectively. Between every sample reading the electrodes were continuously rinsed with

distilled water.

�2�1

3.4.2.3 Determining Acidity

Acidity wasestimated by titration with 0.1 M NaOH using phenolphthalein as indicator.10ml of

juice sample was taken in a 250 ml beaker and added 50 ml water into the beaker. It was mixed

properly. Then 3 drops ofphenolphthalein indicator was added to the juice water solution. The

solution was titrated by the standard 0.1 M NaOH(Islam et al., 2015). The burette reading was

recorded.The results were calculated as anhydrous citric acid (%).

�% �=�×

!��× �1�0�0�% �(�5�)

Where,acidity factor = 0.0064

3.5 Statistical Analysis

Data analysis was performed by usingthe statistical software packages SPSS 21(IBM Corp,

USA). Each dataset consisted of a matrix, in which the columns represented the individual

concentration of Mango juicesamples, and the rows consist of the concentration in mg/L of all

the detected metals determined. One-way analysis of variance (ANOVA) was used to test for the

presence of significant differences in the mean concentrations of metals, pH value, TDS value,

E.C value and acidityamong the four Brands.Differences were considered significant when p<

0.05. Samples were digested intriplicate and also analyzed in triplicate.Karl Pearson„s

correlation coefficients were determined for analysis of correlation among thesomeparameters

and origin software was usedto drawingthe calibration curve of standard solution.

�2�2

4. RESULTSAND DISCUSSION

4.1 Optimizations for Metal Content Determination

In optimizing the digestion procedure, different procedures for the samples were tried and as can

be seen in (Table 4) alternative, consumed smaller reagent (acid) volume, smaller digestion time

and a colorless solution with no residue was obtained. On this basis total reagent volume 6 mL

(3mL HNO3 and 3mL HCl), at a temperature of 90oC for 45 min were chosen as the working

conditions. The blank solutions were prepared by digesting the mixture of reagents following the

same digestion procedure as the samples and diluted with distilled water.

Table4: Optimization of Parameters for Wet Digestion ofJuice Sample

HNO3 (mL) HCl (mL) Total Temp.(oC) Time (min) Observation(Result)

1 0 1 105 45 Yellow

2 0 2 105 45 Yellow

4 0 4 105 45 Yellow

4 0 4 105 60 Yellow

1 1 2 105 60 Yellow

1 2 3 105 60 Yellow

2 1 3 105 60 Yellow

3 2 5 105 60 Light Yellow

3 3 6 105 60 Colorless and

clear

3 3 6 90 60 Colorless and

Clear

3 3 6 90 45 Colorless and

clear

3 3 6 90 30 Lightly yellow

3 3 6 80 45 light yellow

3 3 6 80 60 Light yellow

Bold = Optimized Conditions

�2�3

Therefore, the procedure with total of 6mL reagents volume (3 mL HNO3 and 3mL of HCl),

heating at 90oC and 45 min digestion time was selected for this study.

4.2 Instrument calibration

Calibration curve for Standard Solutions

Calibration curves were used to understand the instrumental response to the metal analyzed and

predict the concentration in an unknown sample. Accordingly, a set of standard solutions were

prepared at various concentrations with a range that includes the unknown of the metal

concentration. In this regard, Calibration curves were prepared for each metal to determine their

concentration in the mango juice sample .A series of standards were prepared from the stock

standard solutions of 1000 mg/L by diluting with distilled water for each metal and a calibration

curve was plotted for each metal to determine their concentrations in juice samples. The

calibration curves forsomestudied metals are shown in Figure.

�-�2 �0 �2 �4 �6 �8 �1�0 �1�2 �1 �4 �1�6

�0�.�0

�2�.�0�x�1�0�7

�4�.�0�x�1�0�7

�6�.�0�x�1�0�7

�8�.�0�x�1�0�7 �M �n

�y� �=�(�3�.�1� �x� �1�0�5 �)� �+�(�5�.�9� �x� �1�0�6 �)�x

�R�2 �=� �0�.�9�9�9�5�3

�I�n�t�e

�n�s�

i�t�y

�C �o�n �c�e�n �t�r�a�t�i�o �n � �(�m �g�/�L �)

�-�2 �0 �2 �4 �6 �8 �1 �0 �1 �2 �1 �4 �1 �6

�0�.�0

�5�.�0�x�1�0�5

�1�.�0�x�1�0�6

�1�.�5�x�1�0�6

�2�.�0�x�1�0�6

�Z �n

�y� �=�(�1�.�8� �x� �1�0�4�)� �+�(�1�.�3� �x� �1�0�5�)�x

�R�2 �=� �0�.�9�9�8�0�7

�I�n�t�e

�n�s�

i�t�y

�C �o�n �c�e�n �t�r�a�t�i�o �n � �(�m �g�/�L �)

�-�2 �0 �2 �4 �6 �8 �1 �0 �1 �2 �1 �4 �1 �6

�0

�1 �x�1�0�6

�2 �x�1�0�6

�3 �x�1�0�6

�4 �x�1�0�6

�5 �x�1�0�6

�C �d

�y� �=�(�8�.�7� �x� �1�0�4�)� �+�(�3�.�7� �x� �1�0�5 �)�x

�R�2 �=� �0�.�9�9�1�4�5

�I�n�t�e

�n�s�

i�t�y

�C �o�n �c�e�n �t�r�a�t�i�o �n� �(�m �g�/�L �)�-�2 �0 �2 �4 �6 �8 �1 �0 �1 �2 �1 �4 �1 �6

�0 �.�0

�5�.�0 �x�1�0�4

�1�.�0 �x�1�0�5

�1�.�5 �x�1�0�5

�2�.�0 �x�1�0�5

�2�.�5 �x�1�0�5

�3�.�0 �x�1�0�5

�P�b

�y� �=�(�7�.�9� �x� �1�0�3�)� �+�(�2� �x� �1�0�4 �)�x

�R�2 �=� �0�.�9�8�5�5�5

�I�n�t�e

�n�s�

i�t�y

�C �o�n �c�e�n �t�r�a�t�i�o �n� �(�m �g�/�L �)

Figure3: Calibrationcurve of intensity verses concentration for Mn, Zn, Cd, and Pb

�2�4

The abovefigure shows that the linear relationship wasestablished forall regressionequations

with acceptable coefficient of determination (R2) values

Table 5: Working standards and correlation coefficients of thecalibration curves for

determinations of metals in mango juice sample

Metals Concentration of standard solutions(mg/L) Correlation coefficientof

calibration curves

Mg 0.002, 0.02, 0.2,2, 8, 14, 20 0.9932

Ca 0.002, 0.02, 0.2,2, 8, 14, 20 0.9985

Cr 0.002, 0.02, 0.2,2, 8, 14, 20 0.987

Mn 0.002,0.02,0.2,2, 8, 14,20 0.9995

Ni 0.002,0.02,0.2,2, 8, 14,20 0.99353

Fe 0.002,0.02,0.2,2, 8, 14 ,20 0.9987

Zn 0.002, 0.02, 0.2, 2, 8, 14 ,20 0.9987

Cu 0.002, 0.02, 0.2, 2, 8, 14 ,20 0.9995

Cd 0.002, 0.02, 0.2, 2, 8, 14 ,20 0.99145

Pb 0.002, 0.02, 0.2, 2, 8, 14 ,20 0.9855

Based on the above data, the intensity of each standard solution for eachtoxic and essential

metal was measured by using ICP-OES. The obtained squared correlation coefficient (R2) values

of calibration curves ranged between 0.985and 0.9995. These high square correlation coefficient

values demonstrate that there is a linearcorrelation between the intensity and metal

concentrations.

�2�5

RSD value

The RSD values for precision were shown in Table 6. The resulted %RSD values were below

10%. It indicates that the measured data for metal determination in juice samples was highly

precise, and itcan be stated that the ICP-OES method was shows good precision for

determination metal in juice.

Table6: RSD resultof metalanalysisfor mango juice samples

Metal RSD

(%)

Mg 0.3

Ca 0.37

Cr 0.71

Mn 0.87

Fe 0.62

Ni 0.36

Cu 0.18

Zn 0.13

Cd 4.08

Pb 2.92

�2�6

Detection and Quantification Limits

As observed from the table 6, theMDL and MQL values for each metal (Mg, Ca, Cu, Cr, Mn,

Ni, Zn, Fe, Cd, and Pb) were greater than the IDL values. Theseindicated that the selected

analytical method was good or ICP-OES Instrument able to detect the minimum concentration of

analyte.

Table7: Method detection and quantification limits (mg/L) for Mango Juice

Metal MDL MQL IDL(mg/L)

Mg 0.014 0.046 0.0016

Ca 0.03 0.1 0.01

Cr 0.009 0.03 0.0071

Mn 0.003 0.01 0.0014

Fe 0.009 0.03 0.0046

Ni 0.06 0.2 0.015

Cu 0.03 0.1 0.0097

Zn 0.006 0.02 0.0059

Cd 0.006 0.02 0.0027

Pb 0.063 0.21 0.042

Generally,Evaluation ofanalytical method parameters including linearity, sensitivity, precision,

and accuracy showed acceptable results. Therefore,the method can be successfully used for

determination of Mg, Ca ,Cr, Mn , Ni , Fe , Cu, Zn, Cd ,and Pb in mango juice sample.

�2�7

4.3 Levels of Metals in the four brands of mango juice

The mean concentration of the studied metals in the samples at each Brand is given in table7.

According to the analysis of the results metal content of the mango juice was different from one

brand toanother. Form the Ten metals, seven(Mg, Ca, Cr,Mn, Fe, Zn, and Cu) were detected in

four brands. However, Pb was not detected in allsamples,Ni was detected only in Texasbrand,

andCd was detected inall brandsexcept Texas.The mean concentration ofmetals for this study

were as follow, Ca>>Mg>>Fe> Zn >Cu> Mn>Cr > Cd forthe three brands(3D, Prigat, and

Rani), andMg>>Zn>>Fe>Mn >Ca> Cu>Ni > Cr for Texas brand.

Table8: The Mean± SD concentrations (mg/L) and significance valueof metals were found in

mango juice sample

Metal 3D Prigat Rani Texas

Mg 6.10a ± 0.10 8.57b ±0.64 6.83a ±0.26 36.40d±1.06

Ca 69.50a ± 0.70 54.50 b ±5.01 44.10 c ± 2.10 0.40d ±0.01

Cr 0.16 a ± 0.01 0.15a ±0.01 0.14b ± 0.01 0.16a± 0.00

Mn 0.23a± 0.01 0.27b± 0.00 0.22a± 0.01 0.86c±0.00

Ni ND ND ND 0.28 ± 0.00

Fe 1.58a± 0.09 1.37b± 0.10 0.92c ± 0.06 9.00d ± 0.07

Cu 0.62a±0.02 0.39b± 0.00 0.40b ± 0.01 0.34c ± 0.03

Zn 0.88a± 0.03 0.49b ± 0.02 0.41b ± 0.03 9.80c ± 0.20

Cd 0.02a± 0.00 0.02a± 0.00 0.01b ± 0.00 ND

Pb ND ND ND ND

Means followed bythe different letterin a row are significantly different (Turkey„s testP < 0.05). Tukey HSDa

Magnesium

Magnesium is required to regulate sugar metabolism, energy production, cell membrane

permeability, and muscle and nerve conduction(Deheleanet al., 2016). It plays a vital role in the

absorption of Ca and together combines for strengthening the bone to avoid occurrence of

osteoporosis (Swamiet al., 2012).In this studythe highest concentration Mgwas recorded in

Texas brand and the smallestconcentration wasfound in 3D brand.Whencompared to the result

of the current study to the literature, the value of Mg concentration in the texas brand were in

�2�8

agreement with those reported in Fruit juice 13.07ˆ 140.42 mg/LDehelean and Magdas,(2013).

The Permissible Levels of Mg in Fruit Juice Samples were < 8mg/L that could be reportedby

FAO/WHO (1992).However, themagnesium concentration in Prigat (8.57 mg/L) and Texas

(36.40mg/L) were abovethose limits. Magnesium sulphate is usually added to juiceas a

preservative(Deheleanet al., 2016). This could be a reason for high concentration of Mg in the

two type of juice.One-way ANOVA test showed that the meanconcentrations of Magnesium

weresignificant difference (P < 0.05) among four brands.

Calcium

Calcium ismajor found in our bones and teeth.It also regulates cell membrane permeability to

control nerve impulse transmission and muscle contraction, and it is important for blood clotting,

and it regulates hormonal secretion and cell division (Deheleanet al., 2016). The calcium

contents of the analyzed juices ranged from 0.40to 69.5 mg/L. Mean comparison of calcium

contents in juices sample characterized in increasing order of the metal level (mg/L)

0.40<44.10<54.50< 69.50 for Texas <Rani < Prigat< 3D juicerespectively. Suleiman and

Gaila, (2013) reportedthat, 0.44mg/L concentration of Ca in mango juice sample taken from

Nigeria were comparablewith texas (0.40mg/L) juice in this study. The observed values of Ca in

three juices (Prigat, Rani and 3D) are mostly in good agreement with the reported average

concentration range of 21.54- 338.35 mg/Lby Deheleanand Magdas,(2013).

Deheleanet al., (2016)also reportedthat, Ca concentrationwas0.42 mg/L to 107.40 mg/L in

commercial juices sampletaken from Romanian,this result agreed withpresentstudy.Calcium

ascorbate or calciumchloride isaddedin the production processof juice to prevent enzymatic

browningor to enrich the products in vitamin C, prevent changes in the smell and as antioxidants

and acidity regulators(Deheleanet al., 2016). Thesemay be a reason forpresenceof higher

calcium in three juices sample (3D, Prigatand Rani).

One-way ANOVA test showed that, the mean concentrations of calcium was significant

difference (P < 0.05) among four brands.

�2�9

Chromium

Chromium is commonly found in foodstuffs, as it enhances the action of insulin and functions in

mammalian glucose metabolism which is very vital to humans(Eneji et al., 2015). Previously, it

was thought that toxic effects of chromium are seldom seen; recently however, the safetyof the

dosage forms ofchromium hasbeen questioned. It is important to be aware that individual

patients with type2 diabetes mellitus might have increased risk of hypoglycaemic episodes with

accumulation of chromium, and also it can cause damage to the liver, kidney, nose, lungs; and

possible asthma attack(Kleefstraet al., 2004). The concentration of Cr in all sampleswas3D

(0.16 mg/L), Prigat (0.15 mg/L), Rani (0.14mg/L) and Texas (0.16mg/L). The chromium levels

observed inthis studywere higherthan thechromium levels reportedby Ajai et al., (2014), and

Abdel-Rahmanet al., (2019). Franciscoet al., (2015) reportedthat, Packing materials used for

juice contain 0.2% of Crtherefore;it could be one reason for the high concentration of Cr

detection in this study.One-way ANOVA test showed, there was significant difference (P <

0.05) among the mean concentrations of Cr in the Juice samples. However, the difference was

not significancebetween threesamples (3D,Prigat, andTexas)from the turkey analysis.

Manganese

Manganese is an essential trace element that plays a role in bone mineralization, protein and

energy metabolism, metabolic regulation, cellular protection from damaging free radical species

and the formation of glycosaminoglycans(Coleset al., 2012). Insufficient of Mn in the body can

result in anaemia while its accumulation in humans results to gastrointestinal disorder, cancer,

respiratory problem (Blaurock, 2009). It was detected in four brands with a concentrationof 3D

(0.23), Prigat (0.27), Rani (0.22) and Texas (0.86mg/L).except Texas brand, theresult of this

study agreed with previous study reported by Enejiet al., (2015),andwerecomparable to those

reported byAbdel-Rahmanet al., (2019), with in mangojuiceMn concentrationwas0.24to 0.32

mg/L. The result from oneway ANOVA revealed that, except the values between 3D and Rani,

the Mn valued observed has significance difference between the brands.

�3�0

Iron

The average concentrationof Femetal in the samples ranged from 0.92ˆ 9.00 mg/L. Fe being 4th

abundantly elementfound on the earth crust,and it is an importancein the entire human

biochemistry.For instance, process of hemoglobin (blood pigment) generation entirely depends

upon iron contentsof the body (Akhtaret al., 2015), and it was a core component of the red

blood cells which is present in most foods and beverages. Its deficiency can cause anemia (Eneji

et al., 2015b). Its concentration was observed lowest in RaniJuiceand highest in Texas sample.

Based on the average of samples, their concentration shows Rani< Prigat < 3D< Texas with

respective concentration of 0.92, 1.37,1.58, and 9.00mg/L. Except the result obtained for Texas

sample, all result obtained in the present study are significantly lower than the result obtained by

Ajai et al.,(2014).The Permissible Levels of Fe in Fruit Juice samplesset byFAO/WHO (1992)

were 15mg/L. However, theconcentration of Fe in all sampleswas below those limits.The

ANOVA testshowed, Fe concentration aresignificantly differences among the samples.

Nickel

Nickel is an element that occurs in the environment only at very low levels and is essential in

small doses, but it can be dangerous when the maximum tolerable amounts are exceeded

(Adepoju-Bello et al., 2012). This can cause various kinds of cancer on different sites of the

body. It is found in small quantities in many foodstuffs (0.001ˆ 0.01 mg/kg) and in higher

concentrations in foodstuffs such as grains, nuts, cocoa products and seeds (up to 0.8 mg/kg)

(Eneji et al., 2015b). In this studyNi was detected only in the Texas brand with a concentration

of 0.28mg/L. The result was abovethoseprevious studiesby Ackahet al, (2014), Deheleanet

al., (2016)with reportednickel level in mango juice 0.055, 0.155mg/L respectively. According

to the Companhia Siderúrgica Nacional (National Steel Company of Brazil, CSN), which

is responsible for the production of steel sheets used in the manufacture of cans, theproportion

of Ni in the steel is 0.15%(Franciscoet al., 2015). Therefore, the highest Ni content observed

for these samples could arise from contamination either from the water used in fruit juices

processing or from packing materials.

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Copper

The mean comparison of copper contents in different juicesobserved0.34< 0.39 < 0.40 < 0.62

(mg/L) for Texas, Prigat,Rani and3D juice respectively. The results agreed with previous study

by Ajai et al., (2014) with reported copper level in mango juice 0.4 mg/L.However,it is lower

than the concentration reported by Hassan et al., (2014) ranging 5.2 to 13.64 mg/kg in

commercial mango juice inEgypt.And theseresults are in parallel with those recorded byFarid

and Enani,(2010)with 0.317.8-0.500 ppmof Cu in fruit juice. Cu is an essential micronutrient

involved in a number of biological processes needed to sustain life. However, it can be toxic

when present in excess(Bedassaet al., 2017).Presence of copper in food products might be

correlated with its migration Cu ion from food contact material. However thereare chances of

copper contamination from copper pipelines of potable water supplies used for food preparation

and dilution in case of fresh fruit juices(Akhtaret al., 2015).

The ANOVA test showed,themean concentrationof Cu wassignificantly differences among the

samples.

Zinc

Zinc is a crucial component of the manyvital enzymes. The highest concentration of zinc was

found in Texas Brand (9.80 mg/L), while the lowest, was found in Rani Brand (0.41mg/L). Its

concentration in Prigat brand was 0.49 mg/L and 0.88 mg/L in 3D.The highestconcentrations of

zinc were observed in texas samples which may reflect contamination due tofood processing or

agricultural contamination (Enejiet al., 2015). Except the result obtained from Texas juice ,the

concentrations of Zn from this study were found to be below the maximum permissible limit of 5

mg/L set byFAO/WHO (1992). Zinc deficiency affects about two billion people in the

developing countries and it is associated with growth retardation, delayed sexual maturation,

infection susceptibility, and diarrhea in children (Ackahet al, 2014).

The ANOVA test showed,the mean concentration of Zn wassignificantly differences among the

samples. However, no significance difference between Prigat and Rani Samples from turkey

analysis.

�3�2

Cadmium

Cadmium is highly toxic and responsible for several cases of poisoning through food. Small

quantities of Cd cause adverse changes in the arteries of human kidney. It replaces Zn

biochemicallyand causes high blood pressures andkidney damage(Rajappaet al., 2010). Due

to the high toxicity of Cd, it is of public health interest that these metalsis quantified in

beverages and foodstuffs(Bingöl et al., 2010). For instance, cadmium intake in relatively high

amounts can be detrimental to human health. Over a long period of intake, cadmium may

accumulate in the kidney and liver because of its long biological half-life and may lead to kidney

damage(Adepoju-Bello et al., 2012).

Generally, In the present studythe level of Cd was detected in the three samples with a

concentration of 0.02mg/L, 0.02mg/L, and 0.01mg/L in 3D , Prigat, and Rani respectively.The

concentration Cd obtained is lower than the result reported by Suleman & Gaile, (2013) in

mango juice .but higher than the result reported byAbdel-Rahmanet al., (2019). Eventhough,

the result can be taken as an indication forthe need of control in packing juice.

Lead

In this study Pb were not detected in all samples. These results are comparable to those reported

by Hassanet al., (2014),Ajai et al., (2014), and Musa and Parmeshwal, (2018). Lead (Pb)

accumulates in the brain leading to plumbism. In children it leads to lower IQ, short attention

span, hyperactivity and mental deterioration. Loss of memory and weakness of joints have been

reported in adults(Ihesinachi and Eresiya, 2014).

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Table 9: Comparison of study metals concentration (mg/L) in mango juice with the reported

values

Metal Present study (Suleiman

and Gaila,

2013)

(Ajai et

al., 2014)

(Akhtar et

al., 2015)

(Dehelean

et al.,

2016)

(Musa and

Parmeshwal,

2018)

Mg 6.1- 36.4 0.42 - - 29.02 -

Ca 0.40- 69.5 0.44 - - 20.98 63.87

Cr 0.14- 0.16 <0.002 ND - 0.01 -

Mn 0.22- 0.86 0.66 0.00 0.095 0.246 -

Ni 0.28 <0.05 - 0.055 0.155 -

Fe 0.92- 9.00 8.40 9.00 0.885 0.8 ND

Cu 0.34- 0.62 <0.001 0.40 0.341 0.151 ND

Zn 0.41- 9.8 <0.08 0.90 0.372 0.245 ND

Cd 0.01- 0.02 0.14 ND 0.025 0.0005 ND

Pb ND < 0.004 ND 0.166 0.003 ND

ND: none detectedand-: not reported

The concentrations of Cr, Mn, Ni, Cu, and Znanalyzed inthis study,were found above the

values reported in literature for the mango juices (Table 8).the variation between the obtained

result, andreported in literature might be the variability of used rawmaterials in the fruit juices

production, packingquality, andby different manufacturing processes applied. Becausefruit

juices are coming from different countries, the metals content variesin soil composition where

the fruits were grown. In addition, packing quality represents another factor that influences metal

content in fruit juices(Dehelean and Magdas, 2013).

�3�4

Statistically, the differences in the mean values among the eight metals for each brands of the

mango juice sample aregreater than would be expected by chance; and therefore, here is a

statistically significant difference (P = <0.05).Variations in the mean levels of metals between

the samples were tested using one-way ANOVA. The results of metal concentration indicated

that significant differences were obtained (p < 0.05) at 95% confidence levelsfor Ca, Mg, Cr, Fe,

Mn, Zn, Cu, and Cd in mango juice samples. This significance difference and increase in

elemental concentrations may be resulted from the raw materials and water used in the juices

production, the conditions of plant growing such as levels of toxic metals in soil and irrigation

water, the environmental contamination (fertilizers and pesticides), the industrial processing, and

contamination from containers(Abdel-Rahmanet al., 2019, Hassanet al., 2014).

4.4 Physicochemical Analysis

Table 10 shows results for pH,EC, TDS, andTA in the four brands and their comparison, the

results are expressed as means ± SD for four Samples and P-value was calculated using One-

Way ANOVA Tukey HSDa among juices.

Table10: pH, EC, TDS and Acidity resultsin juice samples from four brands

Parameters 3D Prigat Rani Texas

pH 3.55b±0.08 3.93a±0.15 3.24d±0.01 3.67c±0.01

EC (µs/cm) 2088.30a±35.50 867.3b±2.08 929.00c±7.81 1118.60d± 18.87

TDS (mg/L) 1374.00a±20.29 571.60b±1.53 600.00b±2.08 741.00c±11.78

TA (%)

(as citric acid)

0.33a±0.001 0.26a±0.01 0.24a±0.03 0.30a±0.04

Values followed by different letters within each row and each brand indicate significance difference for eachparameter atP < 0.05.

�3�5

4.4.1 pH

Fruit juices have a low pH because they arecomparatively rich in organic acid. In this study the

pH varied from 3.24 to 3.93(Fig 13). The highest pH (3.93) was found in Prigat Brands and

followed by Texas, 3D, and Rani. According to Standard Organization of Nigeria (1997) and

FAO/WHO (1992) Permissible Levels of pH in Fruit Juice Samples was 1.4ˆ 4.0. Therefore, the

pH values from the present study (3.24-3.93) werewithin the permissible range. The results also

agreed with the previously reported results byBateset al., (2001), and Khanamet al., (2018)

with pH value ranged from 3.3.-4.1 in different brands of mangojuices. The pH value of 3 to 4

may give juice a good potential to inhibit the growth of pathogenic bacteria(Onyekwelu, 2017).

The ANOVA (p<0.05, n=3) test showed pH value are significantlydifferences among the

samples. This might be due to raw material, recipe or the ingredients used in the juice(Tasnimet

al., 2010).

4.4.2 Total dissolves solidsof juice samples

The TDS content is significantly influenced by solid materials dissolved in water in the juice.

The TDS values of the juices observed during the present study ranging from 571-1374 mg/L

were the highest value was observed in 3D (1374 mg/L)followed by Texas (741 mg/L).The TDS

Values are in good agreement with reportedvalue (Maryam.et al. 2012) in the average value of

523-2084 mg/L. however, this study was higher than the reported TDS value of Anwaret al.,

(2013).A high concentration of dissolved solids is usually not a health hazard. However, a very

low concentration of TDS has been found in juice, which is undesirable to many people

(Jahagirdaret al., 2015).

The result from oneway ANOVA revealed that, except the values between Prigat and Rani, the

TDS valued observed has significance difference between the brands.This could be because of

the variety, origin, growing conditions of the fruit, chemical (additives) and manufacturing

process(Amin et al., 2018).

�3�6

4.4.3 Electric Conductivity (EC) of the samples

Electrical Conductivity indicates theionic speciesfound in foodssuchassalts and acids act as

electrolytes that allow current to pass through thefoods. The EC and ionicconcentrationare

found closely related(Ganapathyet al., 2019), means the juice that has thehighest ECis with

high ionic species.In these studiesthe lowest count of electric conductivity was found inPrigat

mango juice (867.3‰s/cm) and the highest was found in 3D mango juice (2088‰s/cm).

Therefore, 3Dcontains highest ionic species than other brands. Anwar et al., (2013) reported

that the electrical conductivity of different brands of mango juicevaried from 1.017-1.917

ms/cm, the resultsfrom the current studywas close tothis literature.

The result showed any significant difference in the values of electrical conductivity (P <

0.05).These variations may be attributed to the variety, origin, growing conditions of the fruit

and chemical (additives) added by different companies(Maryamet al.,2012).

4.4.4 Acidity value

The total acidity of fruit juices was due to the presence of a mixture of organic acids, whose

composition varies depending on the fruit„s nature and Chemicalused that were added during

processing(Amin et al., 2018). Organic acids isimportance for the characteristics and nutritive

value of fruit juices and deliberated individual originality among beverages(Tasnim et al.,

2010). The results indicated that, the highestacidity value (0.33%) was found in 3D Brands

followed by Texas,PrigatandRani. Islamet al., (2015)reportedacidity valueof mango juices

ranged from 0.192% to 0.384%and FAO (2005), Bates et al., (2001) reported that, the

recommended values of acidity formango juices were0.25ˆ 0.5%. The acidity value depend on

the type of fruits, their ripening stage and the season of the year, inappropriate storage of fruit

and packing materials of fruit juices (Nongaet al., 2014,Aminet al., 2018). Therefore, the

results ofthis studywerewithin recommended value. This may indicate that the fruits used in the

preparation of the juices in this study were at the stateof fully maturity and ripening(Nongaet

al., 2014).TheTA values reported in this study were in agreement with those reportedby Ndife

et al., (2013). In the present study, there was no significant variation in the totalTA between

brandsas shown in Table10.

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Table 11: Comparisonof some physicochemical parameterin mango juice with LiteratureValues

Parameter Present study Maryamet

al. , 2012

(Nongaet

al., 2014)

(Amin et al., 2018) (Khanamet al., 2018)

pH 3.24-3.93 2.62-4.75 3.3ˆ 4.9 3.55-3.80 3.3-4.1

EC (‰s/cm) 867.3-2088 218-2451 - - -

TDS(mg/L) 571.6- 1374 523-2088 - - -

TA % 0.24- 0.33 0.1ˆ 0.4 0.21-0.24 0.16-0.39

- Not studied

4.4.5 Correlation matrix among pH, EC, TDS, and TA physicochemical

parameters

Correlation matrix among the parameters wasdetermined by Karl Pearson„s correlation

coefficient (Table12) with their significant level (P > 0.05). Based on theliteraturesdata, if

correlation coefficient is 1.0, there is complete dependency, if it is negative, both are said to be

correlated inopposite directionHowever, if it is positive, it correlated the samedirection

(Khanamet al., 2018).

�3�8

Table12: Correlation matrix among the physicochemical parameters of mango juices samples

Correlation between some physicochemical parameters

Parameter„s pH TDS E.C TA

pH 1

TDS -0.112 1

EC -0.1267 0.9998 1

TA -0.534 0.706 0.718 1

From the Table12 , it was found that total pH and total TDS werenegativecorrelated(r = -

0.112) anda significant positive correlation has been observed betweenTA and TDS (0.71) and

betweentotal TA and total E.C (0.72) andalso negative correlation withpH (r = -0.534).

Therefore, the negative correlationbetween pH and acidityis relevantbecause pH isthe

reciprocal logarithm of hydrogen ion concentration means pH decrease with increasing

concentration of hydrogen ionsor anincrease in acidity or vice versa. According toKhanamet

al., (2018), the value of acidity in mango juicenegative correlatedwith the value of pH.And the

results showed that the electrical conductivity and total dissolved solids were related at a very

high level (r=0.9998), this indicates all dissolved solid can be able to conduct electric current

and contains more ionic species.Similarly, Maryam et al., (2012) reportedthat, a positive

correlation between EC and TDS content (r= 0.98) for fruit based analysis.Therefore, our result

wascomparable with those reported value.

Generally,the currentstudies showed theEC increased witha decreasein pH or an increase in

acidity or vice versathis supported byGanapathyet al., (2019).

�3�9

5. CONCLUSION AND RECOMMENDATION

5.1 Conclusion

From the data presented inthis study, it can be concluded that the nutritional quality has a

varianceamong brands. The basic physicochemicalparameters such as pH,TDS, Electrical

conductivity, acid value, and metal content(Mg, Ca, Cr, Mn, Ni, Fe, Cu, Zn, Cd, and Pb)were

analyzedin mango juice sample. The optimized wet digestion method for mango juiceanalysis

was found effective for all of the metals. This was revealed by theexcellent RSD value(below

ten)obtained which were foundin the acceptable range for theanalyzed metals.

From the results of metal analysis, it can be stated that the studied mango juices are free fromPb,

However, the level of copper,manganese, Iron, chromium, zinc, magnesium and calcium

presented in all Mango juice samples. In this study, most metal presented in Mango juice

samples wasabove the report data value.Therefore, there is need to monitor further the levelsof

toxic metals in the fruit juices as a result of the toxic and bio accumulative potential of these

heavy metalsin human organs. Fromsome basicquality of physicochemical parameter likepH,

TDS, ElectricConductivity (EC),and acidity value was maintained within limit.Except the

acidity value, the results of physicochemical parameterand metal contentsindicated that

significant differences were obtained (p <0.05) at 95% confidence levels.

�4�0

5.2 Recommendation

The following suggestions are recommended in order to monitor and protect the Quality juice

due to the environmentalpollution increasesthrough the time.

ðØ This study focused on only four brands of mango juice. Hence, it is recommended to

perform research works on other juice that have the ability to understand the physico

chemical characteristics of juices.

ðØ Due to the wide consumption of fruit juice drinks,it contributesa large Quantity of heavy

metals intake and therefore all people„sstrict control of these elements is advisable.

ðØ Government authorized department should take initiatives for providing training to

manufacturersto awarenesson maintaining the rules and regulation of Quality

assessmentfor juices production.

�4�1

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�5�1

APPENDIX

Table13: Optimizationof Parameters for Wet Digestion of Juice Sample by using HNO3

HNO3 (mL) Temp.(oC) Time (min) Observation(Result)

1 105 45 Yellow

2 105 45 Yellow

4 105 45 Yellow

6 105 45 Yellow

8 105 45 Yellow

10 105 45 Yellow

�5�2

Table 14: Optimization of Parameters for Wet Digestion of Juice Sampleby using HNO3 and

HCl

Reagent volumes (mL) Temp.(oC) Time

(min)

Observation(Result)

HNO3 HCl Total

0.5 0.5 1 105 60 Yellow

1 1 2 105 60 Yellow

1 2 3 105 60 Yellow

2 1 3 105 60 Yellow

1.5 1.5 3 105 60 Light Yellow

2 2 4 105 60 Very light Yellow

1 3 4 105 60 Light yellow

3 1 4 105 60 Yellow

2.5 1.5 5 105 60 Light yellow

1.5 2 5 105 60 Light yellow

2 3 5 105 60 Yellow

3 2 5 105 60 Light Yellow

4 1 5 105 60 Yellow

1 4 5 105 60 Very light Yellow

2.5 2.5 5 105 60 Almost color less

�5�3

Statically for one way ANOVA test value.

Table15: One way ANOVA test value forPhysical parameterof four mangojuice brand

ANOVApH value

Sum of

Squares

Df Mean

Square

F Sig.

Between

Groups

.738 3 .246 140.603 .000

Within Groups .014 8 .002

Total .752 11

Electrical conductivity

Sum of

Squares

Df Mean SquareF Sig.

Between Groups2908569.667 3 969523.222 2302.906 .000

Within Groups 3368.000 8 421.000

Total 2911937.667 11

TDS value

Sum of

Squares

Df Mean SquareF Sig.

Between Groups1268760.333 3 422920.111 3033.498 .000

Within Groups 1115.333 8 139.417

Total 1269875.667 11

�5�4

Table16: Post Hoc Tests for multiple comparison of mango juice

Dependent Variable: pH value

Tukey HSD

(I)

brand

(J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat .69000* .03416 .000 .5806 .7994

Rani .38667* .03416 .000 .2773 .4960

Texas .26333* .03416 .000 .1540 .3727

Prigat

3D -.69000* .03416 .000 -.7994 -.5806

Rani -.30333* .03416 .000 -.4127 -.1940

Texas -.42667* .03416 .000 -.5360 -.3173

Rani

3D -.38667* .03416 .000 -.4960 -.2773

Prigat .30333* .03416 .000 .1940 .4127

Texas -.12333* .03416 .028 -.2327 -.0140

Texas

3D -.26333* .03416 .000 -.3727 -.1540

Prigat .42667* .03416 .000 .3173 .5360

Rani .12333* .03416 .028 .0140 .2327

*. Themean difference is significant at the 0.05 level.

�5�5

Dependent Variable:TDS value

Tukey HSD

(I) brand (J) brand Mean

Difference (I-

J)

Std. ErrorSig. 95% Confidence Interval

Lower BoundUpper Bound

3D

Prigat 802.33333* 9.64077 .000 771.4602 833.2065

Rani 773.33333* 9.64077 .000 742.4602 804.2065

Texas 633.00000* 9.64077 .000 602.1269 663.8731

Prigat

3D -802.33333* 9.64077 .000 -833.2065 -771.4602

Rani -29.00000 9.64077 .066 -59.8731 1.8731

Texas -169.33333* 9.64077 .000 -200.2065 -138.4602

Rani

3D -773.33333* 9.64077 .000 -804.2065 -742.4602

Prigat 29.00000 9.64077 .066 -1.8731 59.8731

Texas -140.33333* 9.64077 .000 -171.2065 -109.4602

Texas

3D -633.00000* 9.64077 .000 -663.8731 -602.1269

Prigat 169.33333* 9.64077 .000 138.4602 200.2065

Rani 140.33333* 9.64077 .000 109.4602 171.2065

*. The mean difference is significant at the 0.05 level.

�5�6

Dependent Variable: E.C

Tukey HSD

(I)

BRAND

(J) BRANDMean

Difference (I-

J)

Std. Error Sig. 95% ConfidenceInterval

Lower BoundUpper Bound

3D

PRIGAT 1221.000000* 16.753109 .000 1182.36726 1259.63274

RANI 1159.333333* 16.753109 .000 1120.70059 1197.96607

TEXAS 969.666667* 16.753109 .000 931.03393 1008.29941

PRIGAT

3D -1221.000000* 16.753109 .000 -1259.63274 -1182.36726

RANI -61.666667* 16.753109 .006 -100.29941 -23.03393

TEXAS -251.333333* 16.753109 .000 -289.96607 -212.70059

RANI

3D -1159.333333* 16.753109 .000 -1197.96607 -1120.70059

PRIGAT 61.666667* 16.753109 .006 23.03393 100.29941

TEXAS -189.666667* 16.753109 .000 -228.29941 -151.03393

TEXAS

3D -969.666667* 16.753109 .000 -1008.29941 -931.03393

PRIGAT 251.333333* 16.753109 .000 212.70059 289.96607

RANI 189.666667* 16.753109 .000 151.03393 228.29941

*. The mean difference issignificant at the 0.05 level.

�5�7

Table 17: One way ANOVA test value for metal analysis of Ca, Cr, Cu, Fe, Mg, Mn, and Zn

calciumSum of

Squares

df Mean

Square

F Sig.

Between Groups643.377 3 214.459 627.914 .000

Within Groups 6.831 20 .342

Total 650.208 23

Chromium

Sum ofSquares

df MeanSquare

F Sig.

BetweenGroups

.000 3 .000 9.496 .000

Within Groups .000 20 .000Total .000 23

Copper

Sum ofSquares

df MeanSquare

F Sig.

BetweenGroups

.011 3 .004 166.208 .000

Within Groups .000 20 .000Total .011 23

�5�8

Iron

Sum ofSquares

df MeanSquare

F Sig.

BetweenGroups

10.762 3 3.587 12209.851 .000

Within Groups .006 20 .000Total 10.768 23

Magnesium

Sum ofSquares

df MeanSquare

F Sig.

BetweenGroups

154.594 3 51.531 3191.982 .000

Within Groups .323 20 .016Total 154.916 23Manganese

Sum ofSquares

Df MeanSquare

F Sig.

BetweenGroups

.070 3 .023 6201.674 .000

Within Groups .000 20 .000Total .070 23Zinc

Sum ofSquares

Df MeanSquare

F Sig.

BetweenGroups

15.307 3 5.102 5884.975 .000

Within Groups .017 20 .001Total 15.325 23

�5�9

Table18: Tukey HSDmultiple comparisons test formetal analysis of Ca, Cr, Cu, Fe, Mg, Mn,and Zn.

Calcium

Tukey HSD

(I)

brand

(J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat 3.047167* .367617 .000 2.01823 4.07610

Rani 5.081000* .367617 .000 4.05206 6.10994

Texas 13.885250* .367617 .000 12.85631 14.91419

Prigat

3D -3.047167* .367617 .000 -4.07610 -2.01823

Rani 2.033833* .367617 .000 1.00490 3.06277

Texas 10.838083* .367617 .000 9.80915 11.86702

Rani

3D -5.081000* .367617 .000 -6.10994 -4.05206

Prigat -2.033833* .367617 .000 -3.06277 -1.00490

Texas 8.804250* .367617 .000 7.77531 9.83319

Texas

3D -13.885250* .367617 .000 -14.91419 -12.85631

Prigat -10.838083* .367617 .000 -11.86702 -9.80915

Rani -8.804250* .367617 .000 -9.83319 -7.77531

*. The mean difference issignificant at the 0.05 level.

�6�0

Chromium

Tukey HSD

(I)

brand

(J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat .000333 .000808 .976 -.00193 .00259

Rani .003000* .000808 .007 .00074 .00526

Texas -.001167 .000808 .488 -.00343 .00109

Prigat

3D -.000333 .000808 .976 -.00259 .00193

Rani .002667* .000808 .017 .00041 .00493

Texas -.001500 .000808 .278 -.00376 .00076

Rani

3D -.003000* .000808 .007 -.00526 -.00074

Prigat -.002667* .000808 .017 -.00493 -.00041

Texas -.004167* .000808 .000 -.00643 -.00191

Texas

3D .001167 .000808 .488 -.00109 .00343

Prigat .001500 .000808 .278 -.00076 .00376

Rani .004167* .000808 .000 .00191 .00643

*. The mean difference is significant at the0.05 level.

�6�1

Copper

Tukey HSD

(I)

brand

(J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat .045000* .002698 .000 .03745 .05255

Rani .043167* .002698 .000 .03561 .05072

Texas .055667* .002698 .000 .04811 .06322

Prigat

3D -.045000* .002698 .000 -.05255 -.03745

Rani -.001833 .002698 .904 -.00939 .00572

Texas .010667* .002698 .004 .00311 .01822

Rani

3D -.043167* .002698 .000 -.05072 -.03561

Prigat .001833 .002698 .904 -.00572 .00939

Texas .012500* .002698 .001 .00495 .02005

Texas

3D -.055667* .002698 .000 -.06322 -.04811

Prigat -.010667* .002698 .004 -.01822 -.00311

Rani -.012500* .002698 .001 -.02005 -.00495

*. The mean difference is significant at the 0.05level.

�6�2

Iron

Tukey HSD

(I) brand (J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat .042000* .009896 .002 .01430 .06970

Rani .132167* .009896 .000 .10447 .15987

Texas -1.484500* .009896 .000 -1.51220 -1.45680

Prigat

3D -.042000* .009896 .002 -.06970 -.01430

Rani .090167* .009896 .000 .06247 .11787

Texas -1.526500* .009896 .000 -1.55420 -1.49880

Rani

3D -.132167* .009896 .000 -.15987 -.10447

Prigat -.090167* .009896 .000 -.11787 -.06247

Texas -1.616667* .009896 .000 -1.64437 -1.58897

Texas

3D 1.484500* .009896 .000 1.45680 1.51220

Prigat 1.526500* .009896 .000 1.49880 1.55420

Rani 1.616667* .009896 .000 1.58897 1.64437

*. The mean difference is significant at the0.05 level.

�6�3

Manganese

Tukey HSD

(I) brand (J) brand Mean

Difference (I-

J)

Std. ErrorSig. 95% Confidence Interval

Lower BoundUpper Bound

3D

Prigat -.006167* .001118 .000 -.00930 -.00304

Rani .001667 .001118 .461 -.00146 .00480

Texas -.125833* .001118 .000 -.12896 -.12270

Prigat

3D .006167* .001118 .000 .00304 .00930

Rani .007833* .001118 .000 .00470 .01096

Texas -.119667* .001118 .000 -.12280 -.11654

Rani

3D -.001667 .001118 .461 -.00480 .00146

Prigat -.007833* .001118 .000 -.01096 -.00470

Texas -.127500* .001118 .000 -.13063 -.12437

Texas

3D .125833* .001118 .000 .12270 .12896

Prigat .119667* .001118 .000 .11654 .12280

Rani .127500* .001118 .000 .12437 .13063

*. The mean difference is significant at the 0.05 level.

�6�4

Zinc

Tukey HSD

(I)

brand

(J) brand Mean

Difference

(I-J)

Std.

Error

Sig. 95% Confidence Interval

Lower

Bound

Upper

Bound

3D

Prigat .078833* .017000 .001 .03125 .12642

Rani .094167* .017000 .000 .04658 .14175

Texas -1.784833* .017000 .000 -1.83242 -1.73725

Prigat

3D -.078833* .017000 .001 -.12642 -.03125

Rani .015333 .017000 .804 -.03225 .06292

Texas -1.863667* .017000 .000 -1.91125 -1.81608

Rani

3D -.094167* .017000 .000 -.14175 -.04658

Prigat -.015333 .017000 .804 -.06292 .03225

Texas -1.879000* .017000 .000 -1.92658 -1.83142

Texas

3D 1.784833* .017000 .000 1.73725 1.83242

Prigat 1.863667* .017000 .000 1.81608 1.91125

Rani 1.879000* .017000 .000 1.83142 1.92658

*. The mean difference is significant at the 0.05 level.