RATIFICATION PAGE

The complete report of Biotechnology experiment with the title “Making Nata

De Coco” created by:

Name : Nur Fahmi Utami

Reg. num : 101 404 155

Class : ICP

Group : One (I)

After checked by Assistant and Coordinator Assistant, so this report has

accepted.

Makassar, December 6th 2013

Assistant Coordinator Assistant

Rachmayani Ardiansyah, S.Pd Muhammad Nur Arsyad

Known byResponsibility of Lecturer

Prof . Ir. Hj. Yusminah Hala, M.S NIP: 1961 1212 1986 01 1 002

CHAPTER IINTRODUCTION

A. Background

Experts in United States anticipate the world’s population in 2050 to be

approximately 8.7 billion persons. The world’s population is growing, but its

surface area is not. Compounding the effects of population growth is the fact that

most of the earth’s ideal farming land is already being utilized. To avoid

damaging environmentally sensitive areas, such as rain forests, we need to

increase crop yields for land currently in use. By increasing crop yields, through

the use of biotechnology the constant need to clear more land for growing food is

reduced

Biotechnology holds the key to increasing the yield of staple crops by

allowing farmers to reap bigger harvests from currently cultivated land, while

preserving the land’s ability to support continued farming. Malnutrition in

underdeveloped countries is also being combated with biotechnology. Survey

research over the past decade shows that biotechnology is not likely to become an

important issue for most American consumers. Consumers find biotechnology

acceptable whenthey believe it offers benefits and it is safe. Surveys have

consistently found that a majority of American consumers are willing to buy

insect-protected food crops developed through biotechnology that use fewer

chemical pesticides, as well as more nutritious foods. American consumers also

appreciate the role that biotechnology can play in feeding the world. Research

shows that European consumers are much less supportive of all biotechnology

applications.

Research shows that consumers will accept biotech foods if they see a

benefit to themselves or society and if the price is right. Their responses to foods

developed through biotechnology are basically the same as for any other food -

taste, nutrition, price, safety and convenience are the major factors that influence

our decisions about which foods to eat.

Therefore one of the products by using biotechnology in particular the

principles of conventional biotechnology is nata de coco fermentation using

Acetobacter xylinum starter

B. PurposeThe purpose this experiment is to know the steps are making Nata de Coco

and fermentation process in making Nata de Coco

C. Benefit

The benefit this experiment is the student university can be know the steps

are making Nata de Coco and fermentation process in making Nata de Coco

CHAPTER IIPREVIEW OF LITHERATURE

Plants coconut (Cocos nucifera) has a huge benefit in everyday human life .

This commodity is a food ingredient important and a major contribution to the

economy of the community and the state. Coconut water in a large amount of

industrial byproduct of making copra and desiccated coconut are dumped into the

ground to form acid which lowers the pH of the soil , which eventually interfere with

the growth of surrounding plants and cause odor . In quite a lot of coconut water

contains nutrients that can be utilized by living things . The composition of coconut

water include carbohydrates (sucrose , glucose , fructose and sorbitol) minerals (K,

Na, Mg, P, Cl, Fe and Cu), protein (essencial amino acids) and vitamins B and C

(Wahyudi, 2003).

In fact, nata is cellulose bacterial or synthesis bacterial, synthesis result from

sugar by bacterial that form nata namely Acetobacter xylinum. This bacterial is

acetate acid bacterial, aerobic, gram-negative, and short stalk form. In liquid medium,

Acetobacter xylinumforms a layer that can reach thick until some centimeters. That

bacteria is trapped by its layer that has made by itself. In producing strong, elastic,

thick, white, and transparent, we need to attend the incubation temperature,

composition, and pH (Rochintaniawati, 2013).

Acetobbakter xylinum is acetic acid bacteria that are gram-negative , aerobic ,

rod-shaped , nonmotil , the optimum growth temperature of 25-30 0C , and is able to

oxidize ethanol to aetat acid at pH 4.5 ( Madigan et al : Anam,2010) . Nata -making

process by the bacterium A. xylinum cellulose synthesis activity is catalyzed by the

enzyme cellulose synthesis bound to the bacterial cell membrane. Decomposition /

fermentation of sugar is done through the hexose monophosphate and the citric acid

cycle ( Susilawati and Mubarik : Anam, 2010).

When viewed from the properties of coconut water helps in the treatment of

kidney inflammation, treatment of stomach worms are exposed to the disease , the

treatment of digestive disorders and as a result is very useful for handling cases of

cholera . presence and the salts it contains albumin can prevent vomiting and can

eliminate blotches as chicken pox and measles (Wahyudi, 2003).

The above considerations, utilization of waste coconut water are an effort to

utilize waste into value-added products. Nata de Coco Fermentation is done through

the following stages: preparation of materials and tools, a pure culture of acetobacter

xylinum, maintenance making a starter , fermentation , harvesting , processing and

packaging (Misgiyarta, 2007)

Adding the Ammonium sulfate (NH4) 2SO4 or ZA (Zinc ammonium sulfate)

as a source of nitrogen (N) will help stimulate the growth of bacteria and the

formation of a thick compact structure Nata de Coco. Addition KH2O4 (Potassium

dihidropospat) serves as a buffer in the medium, so that will be a constant pH of

around 3-4 . The optimum pH stabilization is essential for the growth of Acetobacter

xylinum in order to obtain a good nata. Acetobacter xylinum can grow well in aerobic

conditions, which is necessary to free oxygen from the air and under acidic conditions

( pH 3-4 ) . To create an atmosphere for aerobic fermentation container usually has a

large surface and closing with a cover that can still be penetrated by air , for example

with porous paper . The cover can be used newsprint, because the price is relatively

cheaper and easier to use. Meanwhile, to make an appropriate acidic conditions

necessary for the growth of bacteria added organic acids , such as acetic acid .

Desired optimum temperature of around 28 º C - 32 º C, whereas at low temperatures

slow growth activity. Inoculation temperature should not be too high or over 40 º C,

because it can inactivate the bacteria (Wahyudi, 2003).

Nata de coco production is began with the mixing of various concentrations of

sugar as a carbon source, urea as a nitrogen source, and glacial acetic acid as a pH

regulator in coconut water medium, then the nourished medium of these nutrients is

heated for 5 minutes. This heating serves as a sterilizer so that the medium does not

contaminated by undesired bacteria. After it do cooling process at room temperature

for 24 hours, then add starter (Acetobacter xylinum) into the coconut water medium.

After nine days of fermentation process, it will form a white gel on the surface of the

medium. In the early stages of nata de coco making process, Acetobacter xylinum

that have been incorporated into the coconut water medium will get a rapid increase

in the number of colonies, then the bacteria that exist in this medium produce large

amounts of cellulose fibers with the help of isomerase and polymerase enzymes

which are also produced by this bacteria, so that the surface of coconut water medium

will look turbid or gel is formed with a higher viscosity than the fluid beneath. As

longer gel layer as thicker it and very clearly visible, whereas the amount of fluid in

the medium is getting decrease (Saputra and Darmansyah, 2010).

According to research done by [Bambang Piluharto et al., 2003 and Kongruang,

S et al., 2008 : Saputra and Darmansyah, 2010], glucose that play a role in the

formation of cellulose is glucose in β form so that all glucose in α form will be

changed into β form through isomerase enzyme in Acetobacter xylinum. The next

stage is glucose binding to other glucose through 1,4 β-glycoside bond. The last stage

is polymerization, the formation of cellulose. Polymerization occurs through

polymerization enzymes in Acetobacter xylinum. The reason why some

microorganisms (Acetobacter xylinum) make cellulose in a large amount biologically

is to maintain its existence in the surface of growth medium, so that the bacteria are

still able to obtain sufficient oxygen to do activity and produce cellulose, and also

defend themself from unwanted substances such as: dirt, other bacteria, and

ultraviolet light [Brigid, A et al., 2009: Saputra and Darmansyah, 2010].

Success in making nata de coco influenced by viability (viability ) of bacteria ,

nutrient content of coconut water and environmental media. Viability of the good

bacteria will produce nata good and fast. The content of nutrients, especially sugars as

a carbon source for the formation of raw materials indispensable nata. Similarly, the

availability of nitrogen and mineral resources, although not used directly forming

nata , is indispensable for growth of Acetobacter xylinum (Wahyudi,2003).

CHAPTER IIIOBSERVATION METHOD

A.Time and Date

Day/Date : Wednesday/November 13rd 2013

Time : at 11.00 a.m. – 02.00 p.m.

Place : Biology Laboratory second floor to the west of FMIPA UNM

B. Tool and Material

1. Tools

a. A stove

b. 2 pieces of containers

c. A pan

d. Some newspaper

e. Raffia fibre

f. Wood spatula of stirring spoon

g. Storage chamber

2. Materials

a. 1 liter of coconut water

b. 12 mL of acetate acid 100%

c. 8 g of sugar

d. Starter (Acetobacterxylinum bacteria)

e. 4 tablespoon of ZA fertilizer

f. pH meters

C.Work Procedure

1. Coconut water is filtered and boiled, after boiled leave for 10 minutes.

2. Added the other materials and simmer 10 minutes.

3. Checked the pH up to 4.

4. Poured the coconut water into two containers that have been sterilized. Then

cover with newspaper that sprayed alcohol first. Stored in safe place, let it for

24 hours.

5. Measured the weight of coconut water before inoculation.

6. Poured the starter into coconut water in container, stored in safe place.

7. Let if for 7 days.

8. Measured the weight of coconut water after inoculation.

CHAPTER IVOBSERVATION RESULT

A. Observation Result

No ZA (Gram)

Weight Before (Gram) Weight After (Gram)

Container

1

Container

2

Container

1

Container

2

1 4 500 650 425 550

2 8 430 530 375 425

3 16 970 825

Group 1 (8 g)

Before

530 430 g

B. Discussion

From observation obtained that coconut water from all groups not

become nata de coco because there found sedimen mey be factorcoconut to

young or making proces nata de coco is not good.so coconut water still in

liquid.There are many factor is that influence the unsucces ful. Carbon source is

an important factor in the fermentation process. Bacteria needs it to make nata

for metabolism process, glucose to enter the cells and used for energy supply that

needed for breeding, fructose that there will be synthesized into cellulose. The

amount of sugars adding should be noted that sufficient for metabolism and

pellicle formation nata. Although there is sugar in the coconut water but the

sugar is not sufficient to form pellicle, so that need to add from the outside.

According observation result by Anon (2010) first step in making nata de

coco were adding sugar, urea ZA fertilizer, and acetate acid.Acetobacter xylinum

is cellulose bacterial or synthesis bacterial, synthesis result from sugar by

bacterial that form nata. This bacterial is acetate acid bacterial, aerobic, gram-

negative, and short stalk form. In liquid medium, Acetobacter xylinumforms a

layer that can reach thick until some centimeters. That bacteria is trapped by its

layer that has made by itself.Nata de Coco is fermented coconut water with the

help of microbe Acetobacter xylinum, which is produces solid, white,

transparent, sweet taste, and chewy texture.

Carbon source is an important factor in the fermentation process. Bacteria

needs it to make nata for metabolism process, glucose to enter the cells and used

for energy supply that needed for breeding, fructose that there will be

synthesized into cellulose. The amount of sugars adding should be noted that

sufficient for metabolism and pellicle formation nata. Although there is sugar in

the coconut water but the sugar is not sufficient to form pellicle, so that need to

add from the outside.

Nitrogen is used for making nata. ZA generally is the relatively cheapand

tends to sour. Nitrogen is required in the formation of proteins that are important

in cell growth and enzyme formation. Nitrogen deficiency causes cells to grow

less well and inhibit the formation of enzymes necessary so that the fermentation

process may fail or not perfect.

Warming has function to sterilize the media from other bacteria

contaminate.In theearly stages ofthe process of makingnatadecoco,

Acetobacterxylinumwhichhas been incorporatedinto thecoconutwatermediumwill

experiencerapidincrease inthe number ofcolonies, thenthe bacteriathat exist inthe

media. Thecellulosefibersproducedin large quantitieswith thehelp

ofenzymesisomeraseandpolymeraseenzymesarealsoproduced

bythebacteria,soonthe surfacelooksturbidmediumcoconutwaterorformeda gelwitha

higherviscositythan theliquidthat is inbelow it. The longerthegellayeris

gettingthickerandveryclearvisible, whereasthe amount offluidinthemediais

increasinglybit.

The reason why a microorganism (Acetobacterxylinum) makecellulose in

large numbers in biology is to maintain its existencein order to remain at the top

surface of the growth medium, so that the bacteriastill be able to obtain sufficient

amounts of oxygen for activity andproduce cellulose, and also can defend

itselffrom foreign substancessuch as: dirt, bacteria and other ultraviolet light.

CHAPTER VCLOSING

A. Conclusion

Nata de Coco is fermented coconut water with the help of microbe

Acetobacter xylinum, which is produces solid, white, transparent, sweet taste, and

chewy texture.

B. Suggestion

1. Laboratory

Prepare the fulfill tools and materials, so the experiment is success.

2. Assistant

Assistants have to guide the apprentice when doing experiment until it

finish, so the apprentice not confused or panic when something happens in

experiment.

3. Apprentice

Apprentices have to serious when doing experiment and notice the

experiment procedures also every apprentice has to make a temporary report.

BIBLIOGRAPHY

Anam, 2010. Pengantar Bioteknologi. Makassar: Jurusan Biologi FMIPA UNM.

Misgiyarta, 2007. Nata de Coco. http://tulisanpuji.blogspot.com/2011/01/nata-de-

coco.html. Accessed on December 4th 2013 in Makassar.

Rochintaniawati, Diana. 2013. Pembuatan Nata de Coco. Jakarta: Universitas

Pendidikan Indonesia.

Saputra and Darmansyah, 2010. Bioremediasi Limbah Cair Nanas Sebagai Bahan

Baku Pembuatan Nata De Pina. Malang: FMIPA UM.

Wahyudi, 2003. Bioteknologi Pertanian. Jogyakatra: Gadjah Mada University Perss.