To extract pectin from fruit and study various factors that affects the amount of pectin of the fruit. Abstract: Pectin is a family of complex polysaccharide. The quantity, structure and chemical composition of pectin varies between plants to plants, within a plant over time and in different parts of a plant. DuringÂ ripening, pectin is broken down by the enzymesÂ pectinaseÂ andÂ pectinesterase; in this process the fruit becomes supple as the middle lamella breaks down and cells become split from each other.
Pectin is a structuralÂ heteropolysaccharideÂ enclosed in the primaryÂ cell wallsÂ ofÂ terrestrial plants. Part of the carboxyl groups of the anhydro-galacturonic acid is esterifies with methanol. Pectin constitutes a compound collection of polysaccharides in plant cells and is found in the majority of major cell walls and especially in profusion in all non-woody terrestrial plants. Pectin is found all through the primary cell walls and also in the middle lamella (a term for a plate like structure and appearing in multiples) between the plant cells where its primary function is to bind these cells together.280405.jpg
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The aim of the present work is to identify a specific trend in the availability of pectin in fruits and factors that affect the content of pectin in a fruit.
Pectin is found in fruits such as apples, plums, lemons, cranberries, grapefruit, and oranges in plenty. Some vegetables such as carrots, lettuce, and spinach also contain pectin. The amount, structure and chemical composition of pectin differs between plants, within a plant over time and in different parts of a plant. DuringÂ ripening, pectin is broken down by the enzymes pectinase and pectin-esterase; in this process the fruit becomes softer as the middle lamella breaks down and cells become separated from each other.
This is the first factor affecting the availability (amount) of pectin in the fruits. As the above paragraph recommend that the pectin is broken down in the plant over time, "I suggest that the amount of pectin should also decrease overtime as the fruit ripens."
To find a trend in the fruit and what are the effects of the ripening of the fruit on its pectin content I have undertaken three different fruit for experimentation. As mentioned earlier fruits like Apple and Citrus fruit contain good amount of pectin and hence, I have chosen Apple, Orange and Pear among the fruit for extraction of pectin and to study the effect on the amount of pectin due to the ripening of these fruit. I have undertaken one variable as time (the stage of ripening). For this, I took three sets of unripe fruit of same size and same weight (almost). The day on which the first experiment was performed on the first fruit is regarded as to be the initial stage or the unripe stage. The duration of ripening of seven days was kept constant for all the fruit. This implies that the next experiment conducted to obtain the amount of pectin at the next stage of ripening was conducted after seven days and so on.
In the process of extraction of pectin I first extracted the liquid pectin by the method of milking lotion in the form of a thick saucy liquid. By adding appropriate amount of alcohol i.e. ethanol and isopropyl (rubbing alcohol) to this sauce I obtained loose solid mass like jelly, this is pectin. (The concentration of the alcohol was kept constant)
Another factor that accounts for the formation of pectin is the presence of Ca2+ ions or a solute at low pH. Although the exact mechanism of gel formation is not clear, significant progress has been made in this direction. Depending on the pectin, coordinate bonding with Ca2+ ions or hydrogen bonding and hydrophobic interactions are involved in gel formation. In low-methoxyl pectin, gelation results from ionic linkage via calcium bridges between two carboxyl groups belonging to two different chains in close contact with each other. In high-methoxyl pectin, the cross-linking of pectin molecules involves a combination of hydrogen bonds and hydrophobic interactions between the molecules. \
Another factor is the pH content
Structure and Properties of Pectin:
Pectin has a complex structure with a Î±-(1->4)-linked D-galacturonic acid polysaccharide backbone. Preparations consist of sub-structural entities that depend on their source and extraction methodology.
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Pectin gelation characteristics can be divided into two main types: high methoxy gelation and low methoxy gelation.
Gelation of high methoxy pectin usually takes place at a pH of below 3.5 and total solids content of above 55%. This is the typical gel formed during jam making. High methoxy pectins are characterised by their setting time and the gel strength. Setting time is usually categorised as rapid set, medium set and slow set. High methoxy pectins gel slower as more of the methoxy groups are removed during processing. The gel strength is measured SAG units and the pectin garde is often expressed as the number of units of sugar that a unit of pectin can gel.
Low methoxy pectin is gelled with calcium ions and hence is not dependant on the presence of acid or high solids content. The less ester groups present the more sensitive the pectin becomes to pectin and hence a rapid set, low methoxy pectin has the lowest level of esterification. Amidation can interfere with the gelation causing the gelation to be delayed. Another useful property of amidated pectins is the ability of the gel to re-heal after shearing. http://www.cybercolloids.net/library/pectin/introduction-pectin-properties
Approach to research: (279)
Pectin is widely used as a functional ingredient in the food industry due to its ability to form aqueous gels and has been used in jams and jellies, fruit preparations, fruit drink, deserts etc. Commercial pectin is currently classified according to the degree of esterification. It is produced commercially as a white to light brown powder, mainly extracted fromÂ citrus fruits, and is used in food as aÂ gelling agent particularly inÂ jamsÂ and jellies. It is also used in fillings, sweets, as a stabilizer in fruit juices and milk drinks and as a source ofÂ dietary fibre. Due to the wide distribution of pectin in nature, their availability and their relatively low cost of production gives them great latent importance for creating agent in the medical domain.
Pectin is also anÂ antioxidant.Â Antioxidant foodsÂ appear to confer a number ofÂ healthÂ benefits, including lowering cholesterol, helping peopleÂ manageÂ diabetes, and potentially reducing the risks of certainÂ cancers. People who consume apple pectin can alsoÂ eatÂ otherÂ antioxidant foodsÂ to keep their diets varied. Pectin is found in fresh and dried apples along with supplements, making it easy to access and integrate into the diet.
In addition to this what interests me is the oldÂ aged proverbÂ "an apple a day keeps the doctor away". As a child I always used to get baffled and did not really understand the meaning of this saying. But now to my knowledge its nothing but the Pectin; present in Apples which is a naturally available medicine that can cure number of ailments and illnesses within our body. Pectin reduces cholesterol level in the body which in turn reduces the risk of heart attacks etc. and perhaps this saying holds true.
However, its increasing demand and prosperity in the food industrial sector interest me and made me curious to know more about its availability, strengths and limitations.
Background information: (156)
Pectin was first isolated and described in 1825 byÂ Henri Braconnot, though the action of pectin to make jams and marmalades was known long before.Vauquelin stated its chemical nature in 1790 and Braconnot showed the characteristic of jellification and gave it the name pectin. To obtain well set jams from fruits that had little or only poor quality pectin, pectin-rich fruits or their extracts were mixed into the recipe.
During industrialization, the makers of fruit preserves soon turned to producers of apple juice to obtain dried appleÂ pomaceÂ that was cooked to extract pectin.
Later, in the 1920s and 1930s, factories were built that commercially extracted pectin from dried apple pomace and later citrus-peel in regions that produced apple juice in both the USA and in Europe.
At first pectin was sold as a liquid extract, but nowadays pectin is often used as dried powder that is easier to store and handle than a liquid
Aim: To extract liquid pectin from oranges:
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I cleaned the oranges by rinsing them under cool water.
By using a vegetable peeler I removed the peel from the oranges and cut them into small slices.
Put the orange peelings into a medium sized beaker.
Cutting the orange (slices) in half using a sharp knife I removed the seeds and Placed the seeds aside in a Petri dish.
I squeezed the juice of the oranges into the same beaker that contains the orange peelings.
And kept the remaining part of the orange, the membrane, in the Petri dish in which I had kept the seeds.18102010363.jpg
I placed the orange seeds and membranes into a muslin cloth and tied it with a thread.
Then I placed this muslin bag containing the seeds and membrane into the beaker that contained orange peels and juice.
Added enough water to this beaker so as all the substances along with the muslin bag were immersed into the water.
With the help of a burner and tri-pod stand I heated this beaker for exactly 30 minutes.
In the middle of the process I stirred the content of the beaker to heat them uniformly.
Then I removed the muslin bag from the beaker and let it cool until it was slightly warm to touch.
I obtained my liquid orange pectin by squeezing the muslin bag into another Petri dish.
Aim: To extract liquid pectin from apple and pear:
I chopped the fruit (Apple and Pear) into small chunks using a sharp knife, and filled it in a beaker.
Remember except for the stem of these fruit everything else was put into the beaker. (the seed was also put)
Added just enough water to almost cover the fruit chunks.
I covered the pot using a watch glass and placed it on low heat for a long time, until the fruits were fully cooked. ( in my case it took approximately 2 hours)19102010387.jpg
Now what I had looked like a runny fruit sauce with skins and seeds in it.
I stirred the fruit every twenty minutes or so while they were been cooked.
Â To obtain the liquid pectin from this sauce I should have arranged a strainer by placing filtering paper inside a funnel and set it aside above a beaker.
Then I poured the sauce into the funnel and allowed it to drip.
But since I had to do this experiment within the school period I directly poured it into muslin cloth, held the muslin cloth as a bag and squeezed it gently to remove the liquid pectin.
What dripped out from the bottom was a clear thick liquid that is little bit slimy to the touch.
Aim: to extract solid pectin from the liquid
To further extract pectin from this liquid pectin sauce,
I poured a little bit of ethanol (15ml) and rubbing alcohol (5ml) into Petri dish and then add the extracted liquid fruit pectin. The pectin coagulates into a jelly-like mass.
What did not coagulate into a jelly like-mass is the water and the juice of the fruit.
Hence, I separated the Solid gel of pectin from the liquid.
This was also used to compare the strength of pectin of different fruits. If this mass can be pulled out with a fork and it forms a heaping gob on the tines, it is concentrated enough to jell perfectly. 18102010377.jpg
If it can be picked up by the fork, but mostly hangs from it, then it will jell loosely.
If it cannot be picked up by the fork in mostly one mass, then the concentration is too weak for it to jell.
(Note: the alcohol test doesn't work right if the pectin is hot.)
Aim: To determine the acid content of the fruits
I chopped the fruit into halves and squeeze the juice into a beaker.
(For conducting experiment with apple and pear I had to use a juicer to extract the juice)
Then I filtered all of the juices through the funnel covered with a muslin cloth into another beaker.
I diluted all fruit juices in a volumetric flask, by factor of ten, made standard solutions of all.(0.1m)
Using 0.1m hydrochloric acid (HCl) and sodium hydroxide (NaOH) I performed around three to four titrations to accurately determine the concentration of the sodium hydroxide.
I slowly added the NaOH into the flask containing the fruit juice (with a 25ml burette).
And when the point of neutralisation is reached i.e. when the indicator shows a colour change, the amount of used NaOH is noted (titre).
I used the same sodium hydroxide to titrate against all of the diluted fruit juices.
Phenolphthalein was used as indicator.
Care must be taken that the NaOH is dropped directly into the solution and does not adhere to the glass; otherwise the reading may be false.
While titrating care must be taken to continually swirl the solution in the beaker to keep it thoroughly mixed.
Aim: To Calculate the Sugar/Acid Ratio in the fruits
The calculations for determining the sugar/acid ratios of all produce are the same, but as some products contain different acids the appropriate multiplication factor must be applied to each calculation. A list of these acids and multiplication factors of the used fruits are given below:
Factor for: - citric acid: 0.0064 (Orange)
- Malic acid: 0.0067 (Apples)
- Tartaric acid: 0.0075 (Grapes)
Results expressed as percentage acid:
Percentage acid =
The sugar acid ratio =
Amount of pectin(ml)
Amount of pectin(gms)
(Note: the colour of the cells indicate the quality of the obtained pectin; Yellow indicates that the jelly was loose when picked up by the fork; green indicates that the jelly was good enough to be picked up by the fork; red indicates that the jelly was not of good quality and could not be picked up by the fork)
Here we can see that the trend in the content of pectin is ambiguous. The amount of pectin increases as the fruit proceeds from un-ripe stage to the ripe stage. And then again decrease as the fruit forwards from the ripe stage to the over-ripe stage. However the orange shows a gradual increase in the content of pectin.
(I have to work on the pictures yet....)
Many factors including the ripeness of the fruit, pH, presence of other solute, and degree of methoxylation influence the formation of the gel.
Apple provided me with the most pectin and pear did the least. But what is strange is there I could not propose a specific trend in the pectin content in these fruits. At the unripe stage the fruit contained less pectin but of a poor quality. In middle ripening stage the fruit contained more pectin than the earlier stage and also of a better quality except of oranges where the pectin was not of a good quality. And finally in the later stage i.e. Over-ripe stage the amount of pectin was found to be low than the middle stage in apple and pear but it was more in the oranges. Moreover the quality of pectin of this stage was deteriorating quality but surprisingly in oranges the quality was remarkable.
In the first stage (under-ripe) pectin is of a poor quality because did not gel firmly and I could not pick it up in one mass. This is possible because the fruit were still not well ripened and hence the content of pectin was less and of an immature quality.
In the second stage (ripe) the pectin is of a better quality because they were firmly gelled and could be picked up in one piece by a spatula. This is possible because the fruit is now suitably ripened and as a result the amount of pectin is more and of a better quality.
In the last stage (over-ripe) the pectin content and quality both are declining. This is possible because the pectin is been broken is broken down by the enzymes within the fruit over time.
Another concern over here is the diverging trend in orange, to my understanding, it due to the difference in ripening period. Perhaps Orange must be a slowly ripening fruit or demands certain environment for ripening. Perhaps orange takes more than seven days time to ripe or show some change. As we can see in the last stage were other fruit have revealed a declining characteristic in quality orange has revealed just the opposite, because it must have just entered in its ripe stage and not the over-ripe one.
Secondly I admire that the formation of gel is not analogous in all the fruit. This is because the gel sets with activation of pectin. When ethanol is added to the solution (liquid pectin) it starts cooling and once the temperature falls below the gelling temperature, a gel starts to coagulate. What happens is in high-ester pectins with a pH-value between 2.8 and 3.6, hydrogen bonds and hydrophobic interactions take place. This binds the individual pectin chains together forming a 3-dimensional molecular net that creates the macromolecular gel. This gelling mechanism is known as the sugar-acid-pectin gelation.
Whereas, what happens in low-ester pectins is that ionic rifts are formed between calcium ions (Ca2+) and the ionised carboxyl groups of the galacturonic acid. But low-ester pectins need calcium to form a gel, however, they can form gel at a higher pH-values than high-ester pectins.
This proposes that high-ester pectin set at higher temperature and low-ester pectins set at a relatively low temperature
High-ester pectins set at higher temperatures than low-ester pectins. However, gelling reactions with calcium increase as the degree of esterification falls. Similarly, lower pH-values or higher soluble solids (normally sugars) increase gelling speed.
If gel formation is too strong, syneresis or a granular texture are the result, whilst weak gelling leads to excessively soft gels.
Pectin which is found between the cell wall and the middle lamella did show a gradual difference in its content. It can be concluded that as the fruit ripens from the under-ripe stage to ripe stage the amount of pectin increases and the quality of the pectin improves. And as it further ripe from the ripe stage to over-ripe stage the pectin content of the fruit decreases and possessing a poor quality of pectin.
Moreover, the quality of the pectin delineates the outer texture of the fruit. When the fruit is ripe fairly denotes that it comprises of good quality pectin. Due to the good quality of pectin the fruit is hard and lustrous. If the fruit is over-ripe then it possesses a poor quality of the pectin and as a result the fruit is soft and dull.
Firstly, the Liquid pectin that is been extracted should be considered as the real pectin (liquid) but since I had to do it within the school time I had to squeeze the muslin bag in order to extort the pectin. One can get more pectin by pressing it, but then it comes out a little cloudy and carries more of the fruity flavour. As a result there is a great chance of the liquid pectin to contain water and the fruit squash along with the pectin. To avoid this I could have let it strain overnight but again the school time restrictions follow. And hence I have taken the solid jelly like mass as pectin (which is obtained by adding appropriate amount of alcohol to the liquid pectin) for comparison and contrast.
Secondly I assume that the constant period(of a week) for the fruit to ripe should have been taken more than a week so as to give the fruit enough time to ripe and differ it by the other stages as a result of which the analysis and conclusion would have been more precise.
The apparatus which were used to extract pectin from a fruit were kept unvarying as I used the same set of apparatus for all sets of fruit to keep the systematic error as low as possible.
The major concern out here is that the stage of the three fruits. I have tried my best to choose three sets of each fruit which were very green (green here refers to the un-ripeness of the fruit) and weighed almost the same. But I have no
To generalize and boil down to a precise trend of the amount of pectin in the fruit I should have taken more than three fruits under consideration. As a result the experimental errors would have identified and addressed and I would have been able to draw a precise conclusion.
Health and Safety Guidelines:
Apron and goggles should be worn when dealing with acids and fire.
Sodium Hydroxide in its undiluted form is extremely corrosive to body tissue. Skin contact causes irritation almost immediately and continued contact causes burns. The 0.1 Molar solution used in this test is much safer. However, it is recommended that protective coats are worn when using, and that it is used only in a well ventilated room.
Phenolphthalein is highly flammable and should be used with care. It should be stored and used away from naked flames or other sources of ignition. It is toxic if ingested.
Fruits should be free from defects such as sun scorch and pest or disease damage, which may have affected the normal ripening process.
It is the sugar/acid ratio which contributes towards giving many fruits their characteristic flavour and so is an indicator of commercial and organoleptic ripeness. At the beginning of the ripening process the sugar/acid ratio is low, because of low sugar content and high fruit acid content, this makes the fruit taste sour. During the ripening process the fruit acids are degraded, the sugar content increases and the sugar/acid ratio achieves a higher value. Overripe fruits have very low levels of fruit acid and therefore lack characteristic flavour. Titration is a chemical process used in ascertaining the amount of constituent substance in a sample, e.g. acids, by using a standard counter-active reagent, e.g. an alkali (NaOH). Once the acid level in a sample has been determined it can be used to find the ratio of sugar to acid