HYDROLYSIS OF

NUCLEIC ACIDS
Group 8
Objectives
• To be able to prepare a hydrolyzed and unhydrolyzed DNA and RNA
• To be able to perform qualitative tests to determine the characteristics of nucleic
acid present
THEORIES
Hydrolysis
• Breaking of a bond in a molecule using water.
• Site of cleavage: sugar, phosphate backbone, or in the base
• Result: purine and pyrimidine bases, oligonucleotide, nucleosides, ribose or
deoxyribose, and phosphate.
Base-catalyzed hydrolysis
• Occurs in RNA
• Hydroxyl ion assist a
nucleophilic attack on the 2’
hydroxyl group on the
phosphorus
• yields intermediate cyclic-
2’,3’-phosphonucleosides
that hydrolyze to nucleoside-
2’-phosphates and
nucleoside-3’- phosphates
• UNSTABLE
• Use of water to stabilize
forming 2’ or 3’ nucleoside

• Stable in DNA
Acid-catalysed Hydrolysis
• Causes depurination

• Cleaves the purine N-

glycosyl bonds

• Adenine and guanine are

liberated and apurinic site
remains.
• Some phosphodiester bond
are cleaved

• Much harsher and acidic is

required to remove
pyrimidine N-glycosides
QUALITATIVE TEST
DATA
DNA
Test Control DNA Acid Hydrolysate Base Hydrolysate
Benedict’s Test Sky blue Sky blue Sky blue Sky blue
Orcinol Test Rusty brown Light brown Yellow brown
Deoxyribose White sol’n with Red sol’n with White sol’n with
brown ppt brown ppt brown ppt.
Purine bases Cloudy lavender to Clear solution with Clear sol’n with
clear solution with White precipitate brown ppt
white ppt
Uracil and clear solution More cloudy Bright yellow to
Cytosine clear
solution(Ba(OH)2
Biuret Test Light blue Light blue Light blue
Xanthoproteic Test Light yellow Clear solution Light yellow
Inorganic
Phosphate
 RNA
Test RNA Acid Hydrolysate Base Hydrolysate
Benedict’s Test Sky blue Apple green Sky blue
Orcinol Test Dark bue-green Dark green with black Dark blue-green
particles solution
Deoxyribose White cloudy with brown Red solution with brown Layer of orange, white
ppt ppt and brown solutionwith
brown ppt.
Purine bases Cloudy dirty white sol’n Light pink sol’n with Clear sol’n with black ppt
brown ppt
Uracil and Cytosine More cloudy Cloudy white cloudy white
Biuret Test Light purple Light purple Light blue
Xanthoproteic Test Bright yellow Light yellow Bright yellow
Inorganic Phosphate
sample
Benedict’sTest
Orcinol test
Test for deoxyribose
Test for Purine bases
Test for uracil and cytosine
Biuret test
Xanthoproteic test
ANSWERS TO
QUESTIONS
How can intact DNA be obtained from a
solution of DNA and RNA
• Dna could be obtained from a solution of DNA and RNA by using
a base-catalyzed hydrolysis, since in alkaline solution RNA is
destabilized forming nucleosides while the DNA remains stable
and intact.
Based on the qualitative test, How can you
differentiate hydrolyzed from unhydrolyzed RNA
• Hydrolized Rna is positive for orcinol test, test for purine bases, biuret test,
xanthoproteic test.
-acid-catalalyzed hydrolysis: presence of pentoses, pyrimidines, and
phosphoric acid
- base-catalysed hydrolysis: only for pentoses and pyrimidines.

• Unhydrolized Rna is only positive for phosphate test.

Based on this experimental result, do you think you
obtained pure RNA from yeast? Why or Why not?
• We were not able to obtain pure Rna from yeast because of the positive result
from our xanthoproteic and biuret tests
Based on these experimental result, do you think you
obtained pure DNA from onion? Why or why not?
• We were not able to obtain pure DNA because of the positive test of
xanthoproteic on our DNA samples.
CONCLUSION
• Base-catalysed hydrolysis occurs only in RNA
• Acid hydrolysis of nucleic acids yeilds purines, pyrimidine bases and phosphoric
acid
• Despite the errors in the hydrolysis and qualitative test experiment we have
understood the nucleic acid hydrolysis
Reference:
• http://www.scribd.com/doc/38672208/Hydrolysis-of-Nucleic-Acids#scribd
• http://
www.chem.uky.edu/courses/che554/5_MolBio/NucleicAcidTechnologyIntroChpt.p
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