1. OSAZONE TEST

2. Introduction The technique was developed by Emil Fischer, , a German chemist.

3. Introduction Phenylhydrazine is the chemical compound with the formula C6H5NHNH2.

4. Introduction Osazones are formed when the sugars react with a compound known as phenylhydrazine at boiling point. General Reaction:

5. Introduction These sugars are reducing ones which have either a free aldehyde or a ketone group to react with the phenylhydrazine.

6. Procedures 1. Pipette 1 ml of the phenylhydrazine mixture to a test tube.

7. Procedures 2. Add 5 ml of the test solution

8. Procedures 3. shake vigorously to insure complete solution

9. Procedures 4. Stopper the mouth of the tube loosely with cotton

10. Procedures 5. Place the tube immediately in a boiling water bath

11. Procedures 6. Record the time

12. Procedures 7. Observe the tube

13. Procedures 8. If a precipitate is formed, record the time and remove the tube from the bath

14. Procedures 9. Cool off the tube

15. Procedures 10. Examine the crystals under low power objective

16. Results and Discussions The reaction is stepwise;

17. 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. Mannose Glucose Fructose

18. Osazone crystals have a characteristic shape under the light microscope and help in the identification of the sugar type. Monosaccharides - like needle-shaped or broomstick Disaccharides - like sunflowers Lactose - like tight balls of needles

19. Making Osazone Crystals The 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.

20. Viewed under the microscope: Glucosazone

21. Viewed under the microscope: Fructosazone

22. Viewed under the microscope: Galactosazone

23. Viewed under the microscope: Maltosazone

24. Viewed under the microscope: Xylosazone

25. Viewed under the microscope: Sucrose

27. glucose, four to five minutes;

28. xylose, seven minutes;

29. galactose, 15-19 minutes;

30. maltose, osazone soluble in hot water

31. Actual Time:

32. Xylose – 20 mins

33. Galactose – 25 mins

34. Glucose – 30 mins

35. Sucrose – 47 mins

36. Fructose – 57 mins

38. References http://www.ehow.com/info_8392949_different-osazone-crystals.html#ixzz1Rx9lFGBJ http://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