NAMA(NO BP)

BIOCHEMICAL PRACTICUM

REPORT

(ENZYMATIC BROWNING)

BY:

FITRA AINI FIRMAN

(2210422019)

GROUP 1 KBI

ASISSTANT RESPONSIBILITY:

ALDILA DIFA FEBRI

GRESCIA EKA PUTRI

EDUCATIONAL LABORATORY III

DEPARTMENT BIOLOGY

FACULTY OF MATHEMATICS AND NATURAL SCIENCES

ANDALAS UNIVERSITY

PADANG, 2023
 FITRA AINI FIRMAN (2210422019)

ENZYMATIC BROWNING

I. Working Principle

. Enzymatic browning occurs due to oxidation events by the enzyme

II. Practicum Method

2.1 Time and Place

This practicum was held on Thursday, March 29 2023 at 08.00 until finished
at the Teaching III Laboratory, Department of Biology, Faculty of Mathematics and
Natural Sciences, Andalas University, Padang.

2.2 Tools and Materials

The tools used in this practicum are a cutter knife, HVS paper, and four
containers to put the solution at the treatment stage on the fruit. While the ingredients
used are green apple (Malus domesticus), avocado (Persea americana), starfruit
(Averrhoa carambola), sweet potato (Ipomea batatas), lime (Citrus aurantifolia),
vinegar (CH3COOH), water, and vitamin C.
2.3 Ways to Work
All ingredients are cut into 6 equal parts and then thinly skinned. After that,
each fruit was given 6 treatments. The first treatment, the pieces of fruit were left
without treatment (control). The second treatment, the fruit pieces were dipped in a
solution of vitamin C. The third treatment, the fruit pieces were dipped in lime juice.
The fourth treatment, the fruit pieces were dipped in a solution of vinegar
(CH3COOH). The fifth treatment, pieces of fruit dipped in water. The fifth
treatment, pieces of fruit crushed mechanically like pounded - mashed. Then observe
the changes that occur every 15 minutes.

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III. Result and Discussion
3.1 Result
No Material Time Perlakuan
(minute) Control Vitamin Lime Vinegar water Mechanical
C bruise
Solution
1 Persea americana 15 ++++ ++ + ++ ++ +++
(alvocado) 30 ++++ +++ + ++ ++ +++
45 ++++ +++ +++ ++ ++ +++
60 ++++ +++ +++ ++++ +++ ++++
2 Malus domestica 15 ++ + ++ +++ ++ +++
(green apple) 30 ++ ++ ++ +++ +++ ++++
45 +++ ++ ++ +++ +++ ++++
60 +++ ++ +++ ++++ ++++ ++++
3 Ipomea batatas 15 + + + + + ++
(sweet potato) 30 ++ + + + + ++
45 +++ + ++ ++ + +++
60 +++ ++ ++ +++ + +++
4 Overrhoa 15 +++ ++ ++ + + +++
carambola 30 +++ ++ ++ + ++ ++++
(Startfruit) 45 +++ +++ +++ ++ +++ ++++
60 +++ +++ +++ +++ +++ ++++

Note book
+ : very slowly
++ : slowly
+++ : fast
++++ : very fast

3.2 Discussion
Based on the practicum that has been carried out, it was found that fruits that
were cut and given several treatments experienced a browning process or what is
known as "Browning". Browning is divided into two, namely enzymatic and non-
enzymatic browning. Enzymatic browning is caused due to the activity of enzymes in
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fresh foodstuffs, such as fresh milk, fruits and vegetables. Meanwhile, non-enzymatic
browning is caused by a browning reaction without the influence of enzymes, which
usually occurs during processing. Enzymatic browning occurs in fruits that contain a
lot of phenolic substrates. Phenolic compounds with adjacent orthodihydroxy or
trihydroxy types are good substrates for the Browning process.
According to Wiley Blackwell (2012), this browning event can occur due to
a reaction. This reaction occurs when plant tissue is cut, peeled off, and due to
mechanical damage which can cause damage to the integrity of plant tissue. This
causes the enzyme to come into contact with the substrate which is usually the amino
acid tyrosine and phenolic components such as catechins, caffeic acid, and
chlorogenic acid so that the phenolic substrates in plants will be hydroxylated to 3,4-
dihydroxyphenylalanine (dopa) and oxidized to quinone by the phenolase enzyme. In
this event, you can see the color change on the surface of the fruit influenced by
external factors that trigger the change in color to brown, and this does not affect the
ripeness of a fruit in the short term but will accelerate fruit decay in the long term.
According to Winarno FG (2008) The inhibiting factors for browning are
temperature, sugar concentration, amino concentration, pH, and type of sugar. On fruit
that is mechanically treated, bruises will brown due to the oxidation and
dehydrogenation of polyphenols found in plants. The reaction is catalyzed by
polyphenol oxidase so that a reddish brown o-quinone is formed. This reaction is very
reactive and will then undergo a non-enzymatic browning reaction which forms brown
to black melanin pigments (Busch, 2000).
However, in other treatments on the fruit, the browning process became
slower. Fruits that are dipped in lime juice and vitamin C solution browning process
occurs more slowly. This is because the citric acid contained in lime and vitamin C
can inhibit the browning process. Citric acid can inhibit the browning process because
it can complex copper ions which in this case act as a catalyst in the browning
reaction. As an antioxidant compound, vitamin C also inhibits oxidation on the surface
of the fruit.
Meanwhile, at a pH in an acidic solution, this antioxidant treatment can
inhibit enzymatic browning, such as ascorbic acid and citric acid (Ceroli et al., 2018).
Citric acid has an inhibitory effect on PPO through two mechanisms, namely by
binding copper ions which are important for enzymes and lowering PH (Jiang et al.,
1999; in Ceroli et al., 2018). Meanwhile, ascorbic acid inhibits the enzymatic
browning process by reducing the quinone component (Ceroli et al., 2018). It also
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affects the activity of polyphenol oxidase (PPO) enzymes found in plant tissues.
Where this enzyme will be very active in the normal to acidic pH range. In very acidic
conditions, namely at a pH below 3.5, the activity of this enzyme will be very small.
Generally, acidifying agents such as citric acid and malic acid are widely used to
control pH in order to inhibit the browning process.
In fruits that are immersed in water, inhibition occurs in the browning
process due to reduced oxygen so that contact with the substrate is reduced. Due to the
lack of contact with the substrate containing phenolic compounds, the enzymatic
oxidation will stop or at least be reduced. In fruits dipped in vinegar, the activity of the
polyphenol oxidase (PPO) enzyme was reduced as well as those dipped in acid
because vinegar itself has a pH of less than 3.3. The browning
process on fruits that were soaked in water occurred faster than the immersion
treatment in lime juice and vinegar solution but slower when compared to those that
were not treated (control). Although water can prevent oxidation of fruits treated with
immersion in water, at the most influential level, pH appears to be more significant in
inhibiting enzymatic browning compared to efforts to reduce oxygen.
From the results of the practicum observations above, the practicum can
determine which treatment is more influential in controlling enzymatic browning. In
the first order it can be observed that the control treatment has a faster browning
process than the others, because this treatment does not have a browning inhibiting
factor but leaving it at room temperature makes enzymatic browning occur faster. At
the second order or at the same level as the first, the browning process will take place
faster than the treatment other than control. In the third place, the fruits were treated
with immersion in a solution of vitamin C browning, then followed by the fruits
treated with dipping in a vinegar solution, followed by water and finally lime.

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IV. Closing

4.1 Conclusion

From the practical results above, it can be concluded that to trigger the browning
reaction of fruits/vegetables, there are three components that must act as
conditions for the reaction to work, namely polyphenol oxidase (PPO) enzymes,
phenolic substrates and oxygen. In plant tissues, this enzyme is located separately
from the phenolic substrate so that oxygen is needed to oxidize it with the
polyphenol oxidase enzyme,

To control enzymatic browning, inactivation with heat, chemical inhibition of

4.2 Message

From the practicum that has been carried out, it can be suggested that the
practitioner can be more careful in observing changes in the practicum object
carefully then in the selection of materials it is hoped that the materials used must be
representative so that they can achieve maximum and best results.

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BIBLIOGRAPHY

Aftab, U., M. Ashraf and Z. Jiang. 1999. Low Diet For Broilers. World Poult. Sci. 62(4):688-701
Blackweel, Wiley, 2012. Food Biochemistry and Food Processing, 2nd (ed). New York
Busch, J. 2000. Enzymic Browning in Potato: a Simple Assay for a Polyphenol. Biochemical
Education. 27:171-173
Ceroli, P., Proccacini, L., Corbino, G., Monti, M & Huarte, M. 2018. Evaluation of Food
Conservation Technologies for Potato Cubes. Journal Of The European Association For
Potato Research. Vol : 61, No 4: 67-82.
Winarno, F.G. 2008. Kimia Pangan dan Gizi. Bogor : Mbrio Press.
Hartoyo, T. 2010. Teknologi Pangan Susu Kedelai dan Aplikasi Olahannya. Jakarta: Trubus Agri
Sarana.

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ATTACHMENT

Picture 1. Lime solution Picture 2. vinegar

Picture 3. Vitamin C solution Picture 4. Water

Picture 5. Change at 45 minutes

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enit 45
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Picture 6. Change at 60 minutes