OSAZONE TEST
Introduction
The technique was developed by Emil Fischer, , a German chemist.
IntroductionPhenylhydrazine is the chemical compound
with the formula C6H5NHNH2.
IntroductionOsazones are formed when the sugars react
with a compound known as phenylhydrazine at boiling
point.General Reaction:
Introduction
These sugars are reducing ones which have either a free aldehyde or a ketone group to react with the phenylhydrazine.
Procedures
1. Pipette 1 ml of the phenylhydrazine mixture to a test tube.
Procedures
2. Add 5 ml of the test solution
Procedures
3. shake vigorously to insure complete solution
Procedures
4. Stopper the mouth of the tube loosely with cotton
Procedures
5. Place the tube immediately in a boiling water bath
Procedures
6. Record the time
Procedures
7. Observe the tube
Procedures
8. If a precipitate is formed, record the time and remove the tube from the bath
Procedures
9. Cool off the tube
Procedures
10. Examine the crystals under low power objective
Results and DiscussionsThe reaction is stepwise;
Osazone formation involves hydrazone formation at C-1 of an aldose (or C-2 of a ketose) and oxidation of C-2 (or C-1) of an alcohol group to a ketone (or an aldehyde). The new carbonyl group is also converted to a hydrazone.
Fructose
Mannose Glucose
Osazone crystals have a characteristic shape under the light microscope and help in the identification of the sugar type.
Disaccharides - like sunflowers
Lactose - like tight balls of needles
Monosaccharides - like needle-shaped or broomstick
Making Osazone CrystalsThe difference in the structure of the
monosaccharides is caused by the diverse groups attached to the first and second carbons of the sugar molecules. Their needle-shaped crystals show that the position of the first and second carbons do not matter in the crystal formation.
Viewed under the microscope: Glucosazone
Viewed under the microscope: Fructosazone
Viewed under the microscope: Galactosazone
Viewed under the microscope: Maltosazone
Viewed under the microscope: Xylosazone
Viewed under the microscope: Sucrose
Time to FormThe time needed to create osazone crystals
varies among the various sugars involved, but helps to identify the sugars being tested. For an osazone crystal to be presented from a hot solution will take as long as follows:
fructose, two minutes; glucose, four to five minutes; xylose, seven minutes; galactose, 15-19 minutes; maltose, osazone soluble in
hot water
Actual Time:Xylose – 20 minsGalactose – 25 minsGlucose – 30 minsSucrose – 47 minsFructose – 57 minsMaltose – 59 mins
ApplicationFor identifying sugars esp. Reducing sugars.Osazones are used as dyes
Referenceshttp://www.ehow.com/info_8392949_different-
osazone-crystals.html#ixzz1Rx9lFGBJhttp://www.pua.edu.eg/Version2/Courses2/
Dentistry%20Courses/Freshmen/Spring/BCM101/Practical/Week%202%20practical%20_Chemistry%20of%20carbohydrates_.pdf
http://www.chemistry.ccsu.edu/glagovich/teaching/316/qualanal/tests/osazone.html
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