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Emulsifying Activity Of Pectin Extracted From Apple Pomace Biology Essay

The apple is the pomaceous fruit of the apple tree, species Malus domestica in the rose family (Rosaceae). It is one of the most widely cultivated tree fruits, and the most widely known of the many members of genus Malus that are used by humans. Apples grow on small, deciduous trees. The tree originated in Western Asia, where its wild ancestor, Malus sieversii, is still found today. Apples have been grown for thousands of years in Asia and Europe, and were brought to North America by European colonists. Apples have been present in the mythology and religions of many cultures, including Norse, Greek and Christian traditions. In 2010, the fruit's genome was decoded, leading to new understandings of disease control and selective breeding in apple production.

There are more than 7,500 known cultivars of apples, resulting in a range of desired characteristics. Different cultivars are bred for various tastes and uses, including in cooking, fresh eating and cider production. Domestic apples are generally propagated by grafting, although wild apples grow readily from seed. Trees are prone to a number of fungal, bacterial and pest problems, which can be controlled by a number of organic and non-organic means.

About 69 million tonnes of apples were grown worldwide in 2010, and China produced almost half of this total. The United States is the second-leading producer, with more than 6% of world production. Turkey is third, followed by Italy, India and Poland. Apples are often eaten raw, but can also be found in many foods (especially desserts) and drinks. Many beneficial health effects have been found from eating apples; however, the seeds are slightly poisonous and two forms of allergies are seen to various proteins found in the fruit.

http://en.wikipedia.org/wiki/Apple

2.2 Nutrition.

Apples may reduce the risk of colon cancer, prostate cancer and lung cancer.[60] Compared to many other fruits and vegetables, apples contain relatively low amounts of vitamin C, but are a rich source of other antioxidant compounds.[55] Apple's antioxidant property prevents the damage to cells and tissues. The fiber content, while less than in most other fruits, helps regulate bowel movements and may thus reduce the risk of colon cancer. They may also help with heart disease,[65] weight loss,[65] and controlling cholesterol. The fiber contained in apples reduces cholesterol by preventing reabsorption, and (like most fruits and vegetables) they are bulky for their caloric content.[62][65]

http://en.wikipedia.org/wiki/Apple

2.3 Pectin background

Pectin is defined as complex mixtures of polysaccharides that make up approximately one third of the cell-wall dry substance of most types of plants (Van Buren, 1991). The function of pectin in plants is to contribute structural integrity to the 11 cell wall and adhesion between cells. Pectin is primarily made up of D-galacturonic acid joined by K-(1-4) glycosidic linkages (Van Buren, 1991). As a part of the plant structure, pectin is a complex mixture of blocks of homogalacturonic acid called ‘smooth regions’ mixed with blocks of homogalaturonic acid containing many neutral sugars including rhamnose, galactose, arabinose, xylose, and glucose called ‘hairy regions’ (IPPA,2001). A percentage of the galacturonic acid residues are generally esterified with methanol. The pectin’s of a plant can be water-soluble, chelator soluble, or protopectins (VanBuren, 1991). The methods of extraction will vary based on the actual makeup for each particular plant type. The general makeup of the pectin content varies with ripening of the plant and it is fairly easily brought into solution depending on the plant type (Van Buren, 1991). After extraction pectin consists of smooth galacturonic acid regions with a few neutral sugars still attached (IPPA, 2001). Commercial pectin extraction is mainly from citrus peel and apple pomace, but several other sources exist such as sugar beets and sunflower heads. Pectin is the most important gelling agent used for jam jelly and marmalade. Because of its gelling, stabilizing, thickening and emulsifying properties, pectin is widely used in food industry as a food additives (IDOSI, 2010). Today, however, the largest use of pectin has changed to be a stabilizer for acidified milk drinks. Pectin has also been investigated for its usefulness in the pharmaceutical industry. Among other uses it has been considered in the class of dietary fibers known to have a positive effect on digestive processes and to help lower cholesterol(Braddock, 1999). It also is utilized to stabilize liquid pharmaceutical emulsions and suspensions.

2.3 Pectin extraction method

A few generations ago, powdered pectin wasn't readily available, and the skill of making pectin at home was common knowledge for the family cook - yet today it is a rare individual who knows how to do this (Sam Thayer,2001). Pectin is produced commercially from citrus peel and apple pomace. The extraction conditions vary from facility to facility and are dependent on the pectin source. Extraction most commonly occurs using a dilute mineral acid, usually hydrochloric, sulfuric, or nitric acids. Laboratory scale extractions have been conducted to determine optimal pectin extraction conditions and the feasibility of pectin extraction from a number of different plant materials. The effects of temperature, time, and pH on pectin yield for orange pectin using nitric acid extraction was investigated by Aravantinos-Zafiris et al. (1991).Optimal extraction conditions of pH 1.6, 84CC, and 64 min resulted in yields up to nearly 26% of the dried peel weight. Optimal extraction conditions found through varying extraction time, pH, and temperature for pectin extraction from sugar beet pulp were reported as the use of hydrochloric acid to adjust pH to 1.5 extracted for 4hours at 80CC (Phatak et al., 1988). The resulting pectin yield was 19.53% dry basis at these extraction conditions. Extraction pH, time, and liquid to solid ratio were optimized in a study on pectin extraction from tropical fruits (Simpson et al., 1984).

3.0 Objective

To optimize the best conditions to produce pectin from apple pomace.

To optimize the best solvent to produce pectin from apple pomace.

To determine the yield and the characteristics of extracted pectin.

To study the emulsifying activity and emulsion stability of the pectin.

4.0 Problem Statement

4.1 Aim

Because of the success found in the orange industry for extraction of pectin from peel. The extraction of orange juice yields 55% juice with 45% wet mass residue left over, resulting in a large amount of waste material for disposal (Braddock, 2004). It is of interest to explore pectin extraction from apple pomace. The pectin by itself or by its gelling properties, was employed in pharmaceutical industry, health promotion and treatment. It has been used potentially as a carrier for drug delivery to the gastrointestinal tract, such as matrix tablets, gel beads, film coated dose form (Pornsak Sriamornsak). It also is utilized to stabilize liquid pharmaceutical emulsions and suspensions. Pectin is the most important gelling agent used for jam jelly and marmalade. Because of its gelling, stabilizing, thickening and emulsifying properties, pectin is widely used in food industry as a food additives (IDOSI, 2010). The aim of this project is to study the yield and the characteristics of extracted pectin by using different types of solvents (hydrochloric acid, nitric acid). To study the emulsifying activity and emulsion stability of the pectin. In addition, this also contribute to optimize the best conditions or methods to produce pectin from apple pomace.

5.0 Methodology

5.1 Part 1: Extraction of pectin from apple pomace.

5.1.1 Apple pomace preparation.

Materials/Equipments : Green apple, knife, grinder, fruit press.

Chemicals : Hydrochloric acid, citric acid.

Method :

Green apples were washed, cut and minced in grinder. Crushed pulp was then pressed and the mash was dried. The dry apple pomace pool was crushed and mixed to produce apple flour. Pectin was extracted under reflux in a condensation system at 97oC for 30 minutes (solute/solvent ; 1kg:50L) using known weight of the apple flour as raw material and the dilute hydrochloric acid (pH = 2.5) as the solvent. The step were repeated using dilute citric acid at pH = 2.5. De – ionized water was used to dilute both acids.

5.1.2 Isolation of pectin.

Materials/Equipments : Cheesecloth bag, dessicator.

Chemicals : Ethanol, distilled acetone.

Method :

Hot acid extract was pressed and the concentrated ‘juice’ was cooled at 40oC. The apple pectin was precipitated with alcohol and was stirred for 10 minutes. And then left undisturb for 60 and 120 minutes to allow the pectin flotation. The pectin will remained at the surface of the water/alcohol mixture and then was collected. The collected pectin was dried, cooled in dessicator and the yield was calculated. The hardened pectin cake was broken up, grounded and sieved to obtain powdered pectin. Distilled acetone was used to ensure complete removal of any acid and impurities in the isolated pectin.

5.1.3 Characterization and analysis of extracted pectin by physico – chemical parameter.

Pectin extracted was characterized by FT – NMR spectrometer. Equivalent weight, methoxyl content and anhydrogalacturonic acid (AGA) were determined by the standard methods of Owen et al. (1952).

Equivalent weight.

Materials/Equipments : Conical flasks, burette, dropper, stirrer, pH indicator.

Chemicals : Ethanol, sodium chloride, phenol red indicator.

0.5g of pectin (moisture free) was weight in a 250ml conical flask, moistened with 5ml ethanol and 1g of sodium chloride was added.

Methoxyl content.

Materials/Equipments : Stopper flask, burette, pH indicator.

Chemicals : 0.25 N of NaOH, 25ml of HCl, 0.1 N of NaOH.

Method :

Solution from the equivalent weight was added with 100 ml of distilled water and phenol red indicator. The solution were stirred rapidly and then titrated with 0.1 N NaOH until the colour of the indicator changed. 25ml of 0.25 N NaOH was added to the natural solution, mixing thoroughly in a stopper flask. 25 ml of 0.25 N HCL was then added and titrated with 0.1 N NaOH.

C:\Users\muhamad\Desktop\meo.PNG

Anhydrogalacturonic acid (AGA).

The AGA content can be calculated from equivalent weight and methoxyl content of pectin.

C:\Users\muhamad\Desktop\aga.PNG

Degree of esterification (DE).

The DE of pectin can be determine by methoxyl and AGA content.

C:\Users\muhamad\Desktop\deg of ester.PNG

Moisture and ash content.

Moisture and ash content was determined according to the Association of Analytical Chemists (AOAC) methods (AOAC, 1990).

Moisture content.

Material : Aluminium weighing dish, oven, dessicator.

Method :

The moisture content was determined by oven drying method. The aluminium weighing dish was initially weighed. Then, 1g of pectin plus that aluminium weighing dish was also weighed. Next, the sample was placed into an oven. Lastly, the sample was cooled down in desiccator before weighed again.

Ash content

Materials/Equipments : Porcelain crucibles, muffle furnace, desiccator.

Method : for the ash content determination, porcelain crucibles was initially dried in muffle furnace for 3 hours and then was put in desiccator before weighed. 1 g of sample then was put into the porcelain crucibles and put into the muffle furnace at 600oC for 3 hours. Then it was allowed to cool down in desiccators before it was weighed.

5.2 Part B : Determination of the emulsifying activity and emulsion stability of extracted pectin.

For this experiment, other extraction method were used in order to obtain pectin from apple pomace.

5.2.1 Apple pectin preparation.

Materials/Equipments : Air circulated oven, polyethylene bags, whatman no 4 filter, vacuum dryer.

Chemicals : Hydrochloric acid (0.5 N), citric acid (0.5 N), aluminium salts (sulfate and chloride).

Method :

Apple pomace were dried in air circulated oven then packed in polyethylene bags for further usage. Extraction of pectin from apple pomace was carried out by hydrolysis in acid aqueous solution with HCl adjusted to pH 1.5 at 8oC for I hour. After extraction, the pectin solution was filtered with a whatman no4 filter paper. Concentrated pectin solution was precipitated with aluminium salts (sulfate and chloride). Pectin precipitate was then filtered and dried in vacuum dryer. This step was repeated by using diluted nitric acid with pH 1.5.

5.2.2 Determination of emulsifying activity is based on the ratio of the emulsified layer volume and the whole volume of the solution (Dalev and Simeonova [13] procedure).

Materials/Equipments : Graduated tubes, vortex mixer, centrifuge machine.

Chemicals : Vegetables oil, pectin solution, 0.02% sodium azide (bacteriocide).

Method :

In graduated tubes, emulsions were prepared by adding 3ml of vegetables oil to 3ml of pectin solution containing 0.02% sodium azide. Vortex mixer was used to homogenized the mixture. Sample were then centrifuged at 23oC for 3 minutes. After centrifugation, the whole volume of the solution (Wr) and the emulsified layer volume (ELV) were determined by formula below ;

C:\Users\muhamad\Desktop\emulsifying activity formula.PNG

5.2.3 Determine the emulsion stability of the extracted pectin.

Method :

Similar emulsion sample from determination of emulsifying activity experiment were prepared after 1 and 7 days of storage at 4oC and 23oC. Sample were centrifuged for 5 minutes at 4oC and 23oC. The initial emulsified layer volume (VEi) were measured. After each storage period, sample were centrifuged and the remaining emulsified layer volume (VEr) were measured. Emulsion stability (ES) was calculated as ;

C:\Users\muhamad\Desktop\emulsion stability formula.PNG

6.0 Data collection.

6.1 Part A experiment ;

Table 1 : Yield of pectin obtained by using different solvent at a different extraction time.

PHCl 1 (60min)

PHCl 2 (120min)

PCA 1 (60min)

PCA 2 (120min)

pH of solution

Types of solvent

Initial weight of sample (g)

Final weight of sample (g)

Extraction time (min)

Volume of sample (ml)

Volume of ethanol (ml)

Yield of pectin obtained (%)

Table 2 : physico – chemical characteristics of pectin.

Pectin

Yield (%)

Equivalent weight

Methoxyl content (%)

Anhydrogalacturonic acid (%)

Degree of esterification (%)

PHCl 1 (60min)

PHCl 2 (120min)

PCA 1 (60min)

PCA 2 (120min)

6.2 Part B experiment ;

Table 3 : Emulsion stability of pectin solutions.

Pectin types

P.H.C

P.H.S

P.C.C

P.C.S

Stability (1 day, 4oC )

 

 

 

 

Stability (7 days, 4oC )

 

 

 

 

Stability (1 day, 23oC )

 

 

 

 

Stability (7 days, 23oC )

 

 

 

 

P.H.C ; Pectin extracted with Hydrochloric acid precipitated with aluminium chloride, P.H.S ; Pectin extracted with Hydrochloric acid precipitated with aluminium sulfate, P.C.C ; Pectin extracted with Citric acid precipitated with aluminium chloride, P.C.S ; Pectin extracted with Citric acid precipitated with aluminium sulfate.

7.0 Expected results.

7.1 Part A experiment ;

Figure 1 : Yield of extracted pectins.

Table 2 : physico – chemical characteristics of pectin.

Pectin

Yield (%)

Equivalent weight

Methoxyl content (%)

Anhydrogalacturonic acid (%)

Degree of esterification (%)

PHCl 1 (60min)

14.55

833.33

4.82

59.52

45.98

PHCl 2 (120min)

PCA 1 (60min)

16.65

1666.30

6.21

67.14

52.51

PCA 2 (120min)

7.2 Part B experiment ;

Figure 2 : Emulsifying activity of extracted pectins.

Table 3 : Emulsion stability of pectin solutions.

Pectin types

P.H.C

P.H.S

P.C.C

P.C.S

Stability (1 day, 4oC )

88

91

92

80

Stability (7 days, 4oC )

86.2

79

83

80

Stability (1 day, 23oC )

80

88

85.1

76.5

Stability (7 days, 23oC )

76.7

78

85.2

80

P.H.C ; Pectin extracted with Hydrochloric acid precipitated with aluminium chloride, P.H.S ; Pectin extracted with Hydrochloric acid precipitated with aluminium sulfate, P.C.C ; Pectin extracted with Citric acid precipitated with aluminium chloride, P.C.S ; Pectin extracted with Citric acid precipitated with aluminium sulfate.

8.0 Research plan.

Progression

Jul

Aug

Sep

Oct

Nov

Week

1

2

3

4

1

2

3

4

1

2

3

4

1

2

3

4

1

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3

4

Title published to students

 

 

 

 

 

 

 

 

R

A

Y

A

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Fill in form 1 & submit to committee

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Proposal workshop

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Discussion of project with supervisor

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Proposal preparation

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Proposal submission

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Proposal presentation

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

FYP execution (start)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Fiber treatment

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Thesis writing

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Samples preparation

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Samples testing

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

1st draft submission (chapter 1,2,3)

Progress report presentation

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Thesis writing (chapter 4,5)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Final draft submission

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

FYP execution (end)

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Thesis presentation

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Submission of hard copy

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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