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4: ISOLATION
AND CHARACTERIZATION OF CARBOHYDRATES
ABSTRACT
Carbohydrates are the most abundant class of organic compounds found in living organisms. It is defined as any of a
group of organic compounds that includes sugars, starches, celluloses, and gums and serves as a major energy source
in the diet. The objective of this experiment is to isolate the polysaccharide glycogen from chicken liver and explain
the principle involved in it and in the general tests done to determine the polysaccharide content of the sample. Some
of the other goals of the experiment are to prepare a dialyzing bag, to perform TLC properly, to microscopically
examine the different osazone and mucic acid crystals, and to classify unknown carbohydrates. Initially, the glycogen
from chicken liver is isolated by heating and adding 0.1% acetic acid and then adding 5-10 drops of ethanol. It then
undergoes the general tests for polysaccharides, including Molischs Test which uses 5% naphthol in 95% ethanol (
+ : blue-violet colored ring) and I2 Reaction involving 0.01 M I2 ( + : bluish purple). The sample is also hydrolysed via
acidic and enzymatic hydrolysis, after which it undergoes the qualitative tests for carbohydrates, namely Benedicts
Test ( + : brick-red precipitate), Barfoeds Test ( + : brick-red precipitate), SeliwanoffS Test ( + : yellow to faint pink
solution), Bials-Orcinol Test ( + : blue-green solution), Muric Acid Test, and Phenylhydrazone Test. TLC or thin layer
chromatography is then performed, followed by a quantitative analysis of the samples using Nelsons method of
measurement.
Seliwanoffs
flame to the volume of 10 mL. Test for the presence of
Benedicts
Barfoeds
reducing sugar in the hydrolysate by performing Carbohydrat
Bials
Benedicts Test. e Solution
The glycogen elicited a positive result upon the The purple interface is shown as the product of
addition of Molischs Reagent and conc. sulfuric acid the Molisch Test, a test for carbohydrates. A deep red
because the Molischs Test is a test for carbohydrates. It colour, indicating a positive reaction, is meanwhile seen
also produced a positive result for I 2 Reaction because it as the product of the Iodide test which detects the
will react with glycogen found in a polysaccharide or in a presence of glycogen.
solution.
Figure 2. Positive Result for Benedicts,
Table 2. Results for Qualitative Tests for Barfoeds, Seliwanoffs, and Bials Tests
Carbohydrates Respectively
http://www2.chemistry.msu.edu/faculty/reusch/VirtT
xtJml/carbhyd.htm
http://www.answers.com/topic/carbohydrate
http://chemistry2.csudh.edu/rpendarvis/monosacch
.html
http://www.chacha.com/question/what-is-the-
formula-for-glycosidic-linkage-formation
http://themedicalbiochemistrypage.org/carbohydrat
es.php
http://answers.yahoo.com/question/index?
qid=20080911213302AACHBrn
http://www.harpercollege.edu/tm-
ps/chm/100/dgodambe/thedisk/carbo/molisch/molis
ch.htm
Figure 3. Microscopic Observations of the
Osazones of Sucrose, Xylose, Fructose, http://www.biosci.ohiou.edu/introbioslab/Bios170/17
Lactose, Starch, and Glucose Respectively 0_2/benedict.htm
http://www.harpercollege.edu/tm-
ps/chm/100/dgodambe/thedisk/carbo/barf/barfoed.
htm
http://www.harpercollege.edu/tm-
ps/chm/100/dgodambe/thedisk/carbo/seli/seli.htm
http://en.wikipedia.org/wiki/Ketose
http://www.harpercollege.edu/tm-
ps/chm/100/dgodambe/thedisk/carbo/bial/bials.htm
http://www.slideshare.net/katealyssacaton/mucic-
and-barfoeds-test
http://autumnmarcita.wordpress.com/2010/10/11/c
arbohydrates/
http://himedialabs.com/TD/HTBC002.pdf
References