Group No.

3 MT205 M,TH; 7:30 12:00

LACUANAN, Kim Basil March 5, 2017

LAGUA, JasminTherizza March 7, 2017
MANGAYA, Princess Kaye
MINANO, Czefiah Jade

Proteins:

Abstract

Proteins is found throughout the body, they do most of the work in cells and are required for
the structure, function, and regulation of the body's tissues and organs. It is made up of amino acids
building block of proteins, which are attached to one another in long chains. There are 20 standard
amino acids which play critical roles in the body. The sequence of amino acids determines each
proteins unique structure and its specific function. The amino acids found in proteins are always -
amino acid which the amino group and the carboxyl group are attached to the -carbon atom. This
certain experiment emphasizes on how to distinguish and determine the different result of proteins
conducted from different tests Biuret test, Xanthoproteic test, Ninhydrin test and Reaction with
NaOH and Lead Acetate, also to know the different preparation in making solution like Albumin,
Casein and Nucleoprotein. The group successfully identified the different result from conducted tests
in Properties of proteins, also the group victoriously made solutions in the Preparation of proteins.

Introduction distinguish the positive result as guide for

discovering effect or product. To gain
The experiment was conducted in
knowledge about how hydrolysis of RNA
order to understand the Ribonucleic acid
may results to polynucleotides,
(RNA), its different result when conducted
mononucleotides, nucleotide, purine and
in different tests and the factors that are
pyrimidine bases, sugar and phosphates.
affecting RNA in making negative results.
The main component in this experiment
Methodology
was by the use of RNA combined with
Separation of Ribonucleic Acid (RNA)
yeast and with other chemicals to
from Yeast
formulate various outcomes in different
In 50 mL water, 1% NaOH were
tests. To have knowledge about effects of
diluted for about 10 mL. After diluting, dry
RNA whether it is positive or negative
yeast about 18 g were added and heated
showed in different tests. To point out the
for 30 minutes, After heating, it was
factors that affect the negativity of the
filtered and centrifuge for 10 minutes. The
tests in determining RNA. Also to
Group No. 3 MT205 M,TH; 7:30 12:00

solution
were
decanted
into a
beaker
and cool
the
solution. After cooling, add glacial acetic
until became acidic. After determining the
acidity, evaporate the supernatant liquid,
filter if needed. If the supernatant
evaporated successfully, cool it below 40*
C in 40 ml ethyl alcohol with 2 drops of
HCL. Decant and wash the solution with
95% alcohol and dry if the RNA settled.

Hydrolysis of RNA

Small portion of RNA were added in

10 mL of 10% sulfuric acid, it was boiled
for 30 minutes. After boiling, wait for it to
dry and weigh it. Divide it by two naming
RNA test solution A (hydrolyzed)and B
(unhydrolyzed).With these two division,
different test were conducted: Orcinol test
- 1mL of test sol'n (A & b) and 1mL orcinol
rgt. and heat for 10 minutes; For
Phosphate - 2ml NH3were added in 1mL
test sol'n, add 2mL HNO3 and 1mL
ammonium
molybdate; For
Purine - 1mL
2N HCl were
added in 1mL
test sol'n and
heated for
about 20 mins.,
1ml 2N NaOH
and 2mL
acetate buffer were added for about 5
min.of heating, after heating 1mL CuSO4
and NaHSO3 were lastly added and
heated for about 5 mins.; Lastly,
Benedict's test - 2mL of Na2CO3 were
added to 2mL test sol'n and 1mL of
Benedict's sol'n were added.
Group No. 3 MT205 M,TH; 7:30 12:00

Results and Discussion

The
Orcinol test for Ribose
The test was used to identify the
presence of pentoses. Our group didnt
able to see the pentoses which should have
been present on the solution because both
Separation of the RNA the solution turned pale orange. The
(Ribonucleic Acid) from the yeast requires positive result would show a blue color.
theFigur
disruption of Fig
the cell membrane.
Figu That The pentose
is the reason behind heating it with NaoH. will be dehydrated to form furfural which
The Sodium then
Hydroxide is a reacts with the orcinol
strong base, the that will generate a
addition of this colored substance. The
to the mixture solution will turn bluish and a
of Yeast and precipitate may form.
Distilled water resulted a neutralization,
Figure 4the pH level. Resulting in
thus increasing
the denaturation of contaminant proteins,
inactivates nucleases which can degrade
RNA. Heating helped loosen the
cell membrane by increasing the
kinetic energy of the lipid
molecules, making it release
more RNA. After filtering the
solution, the color became
beige(Figure 1.)

The centrifugation of the mixture

was used to remove the lysed lipids,
denatured
Test proteins and other contaminants
(Figure
tube 2.) Glacial acetic was added
1:
dropwise
Orcin
to lower the pH and help
denature
ol more proteins, prevented alkali
RNA hydrolysis, ensuring that the desired
RNA was not degraded. (Figure 3.) The
mixture was decanted and centrifuged
repeatedly to eliminate the precipitated
proteins. The filtrate was cooled below
40C, after the addition of ethyl alcohol
with concentrated HCL, allowing the RNA
to settle down (Figure 4.)
Group No. 3 MT205 M,TH; 7:30 12:00

This test is used to identify the

Test for inorganic phosphates presence of reducing
This test is used for the identification sugars. The result turned
of Phosphate Ions. The Ammonium out to be negative, because
molybdate will react to the acidified the solution turned blue.
solution. The positive result would
The
Testaddition
tube 5: of the mixture show Brick red color, that
of NH3, 6N HNO3 and 1mL of
Benedict's would identify that the solution is
ammonium molybdate to both
hydrolyzed containing more than 2% of reducing
hydrolyzed and unhydrolyzed sugars. It can also be Green, but with
Test tube 2:
RNA solution showed different smaller amout of reducing sugars which is
Phosphates
results. The unhydrolyzed aboutboth0.1-0.5% only. This will make
solution turned pale yellow. The Copper (II) Ion ( Cu2+) to be reduced to
hydrolyzed solution remained colorless. Copper (I) (Cu+).
Our result turned out both negative. The
positive result would show a Yellow
solution,
indicating the
presence of
phosphate ions
on the test tube.

Purine bases
To determine the presence of purine
bases. The presence of purine bases didnt
appear on our experiment because the
solution turned dark blue for unhydrolyzed Conclusion
and light blue for hydrolyzed, instead of a
foamy white gelatinous substance. The
principle behind this is that the B b
Hydrolysis of N-B-Glucosidic bonds
between purine bases and ribose or
deoxyribose results in a release of bases
caused by NH4OH.

Test Test
tube 3:
tube 4:

Benedicts Test
Group No. 3 MT205 M,TH; 7:30 12:00

The experiment was done to perform 2. Pearson - The Biology Place. (n.d.).
different tests to identify the products of Retrieved February 06, 2017, from
RNA hydrolysis. After performing the
Orcinol test, we conclude that the RNA http://www.phschool.com/science/biology_
has a ribose, which is an Aldopentose. place/biocoach/biomembrane1/hypertonic.
Our test for Inorganic Phosphates turned html
out negative because we didnt get enough
RNA from the yeast on the Separation
process, thats why we didnt saw the 3. Steele, E. (n.d.). Osmotic Pressure:
positive color Yellow, which will prove the
Definition & Formula. Retrieved February
presence of phosphates ions on the RNA.
On the Purine Bases and Benedicts, our 06, 2017, from
experiment on both of the results turned http://study.com/academy/lesson/osmotic-
out negative also, the white foamy
substance didnt appear on the Purine base pressure-definition-formula-quiz.html
and the Brick Red color didnt show up on
the Benedicts, we assume that the reason 4. Cell Membrane. (n.d.). Retrieved
behind this is the same with the test for
Inorganic phosphates, which is the lack of February 06, 2017, from
RNA of the hydrolyzed and unhydrolyzed
solotuion. The positive result on these two
can prove that the RNA is a sugar and
contains purine bases.
We highly recommend that during
the extraction process of the RNA, make
sure to use as many as you can or if
possible, all of the filtered mixture of the
Yeast and distilled water, for this will be
the key in order to properly conduct the
mentioned experiments. The lack of RNA
on the experiments will give a hard time
getting a positive result and make sure to
properly follow all of the procedures
especially the allotted time of heating of
every solutions.

References

1. Bailey, R. (2016, April 25). Cell

Membranes Function, Structure and
Composition. Retrieved February 06,
2017, from
http://biology.about.com/od/cellanatomy/ss
/cell-membrane.htm
Group No. 3 MT205 M,TH; 7:30 12:00

Appendix

SEPERATION OF BENEDICTS TEST

RNA FROM YEAST

10mL of 1% NaOH 2mL test solution

with 50mL water
18g Yeast 1mL of benedict reaagent

40 mL of 95% ethyl
alc.

PURINE BASES

Pinch of RNA in 1mL water

& another 1mL hydrolysate
1mL of 2N HCl

1mL 2N NaOH& 2mL acetate

buffer
0.5mL of 10% CuSO4

10 drops of NaHSO3

INORGANIC
PHOSPHATE
NH3 to 1mL test
solution
6N HNO3

1mL of ammonium
molybdate reagent