Inclass Assignment

Question 1

What is the iodine test used for?

Answer 1

Distinguish starch and glycogen from other polysacchrides

Question 2

How do you interpret the iodine test?

Answer 2

Lugol’s iodine reagent (I2/KI) forms a dark blue complex in the presence of starch and blue brown for glycogen
Others produce no colour change

Question 3

How does the iodine test work?

Answer 3

Starch and glycogen form helical coils
Iodine atoms fit in hélices to form complex
Starch has less branches and therefore binds more iodine producing a more intense colour

Question 4

What are reducing agents?

Answer 4

Sugars that have free or potentially free aldehyde groups

Question 5

What does the benedicts test determine?

Answer 5

If it is a reducing sugar
If non reducing = sucrose or trehalose (no colour change)
If red precipitate = reducing sugar

Question 6

How does the benedicts test work?

Answer 6

Reducing sugars are oxidized by copper ion in solution to form a carboxylic acid and a reddish precipitate of copper oxide

Question 7

What is in benedicts reagent?

Answer 7

Copper sulfate (CuSO4) 
Sodium carbonate (Na2CO3)
Sodium citrate

Question 8

What does the barfoeds test do?

Answer 8

Determines if it is a monosaccharide or disaccharides

Question 9

How do you read the barfoeds test?

Answer 9

If red précipité forms in 2-3 minutes = monosaccharide

10 min = disaccharides

Question 10

Describe the reaction of the barfoeds test

Answer 10

Also uses the reduction of Cu2O

Less reactive and the rate at which red precipitate forms can be used to distinguished

Question 11

What is the Seliwanoff test used for?

Answer 11

Differentiates between ketohexoses and aldohexoses

Question 12

Describe the reaction for seliwanoff test

Answer 12

When a ketose is heater with strong mineral acid (HCl), 5-hydroxymethylfulfural is formed, creating a red complex with resorcinol in 2 minutes
Aldoses will give red colour more slowly

Question 13

What is HPLC?

Answer 13

High Preformance Liquid Cromatography

Method used to identify constituents monosaccharides of oligosacchrides

Question 14

How does HPLC work?

Answer 14

Based on differential partitioning of each sugar between a liquid and solid phase
Mobile phase (solution) passes though column of stationary phase and each sure is partitioned between the phases
Less soluabke sure in mobile phase, the more it is partitioned into the solid phase and longer it is retained in the column
If stationary phase is more polar than mobile = normal phase
Reverse phase is opposite

Question 15

Are sugars polar or non polar? Why does this matter for HPLC?

Answer 15

Sugars are moderately polar and dissolve in polar media
If solution is chromatographed in normal phase, the retention time is long
If done in reverse phase, retention time is short
In practice, polarity of mobile phase is reduced to the point where the unknown sugar are well separated but elites in a conveniently short time

Question 16

Describe the mechanism associated with anthrone assay

Answer 16

Treat oligosacchrides with concentrated sulphuric acid hydroxylase, all inter-ring links, and oxidizes each of the m monomers to produce hydroxymethyl furfural
M oxidized monomers condense with anthrone reagent to form m moles of coloured products
If treated with sodium borohydride, the aldehyde group on terminal aldose ring is reduced to a hydroxyl group
Since hydroxyl group is inert, only produces (m-1) oxidized monomers of coloured product

Question 17

How do you determine the degree of polymerization?

Answer 17

m = A / A-B

Question 18

How do you determine absorbance for anthrone assay?

Answer 18

If treated with sulphuric acid:
A = m (Ecl)mon
Where A=absorbance, c=original concentration of oligosacchrides, and m=moles of coloured product

Question 19

How do you determine the specific rotation of an unknown disaccharide?

Answer 19

Specific rotation of a compound, [alpha] at t degrees Celsius with polarized light is defined as:
[alpha]^t = 100alpha^obs / Cxl
Where alpha obs=observed rotation, C=concentration of sample in g/100mL, and l=pathlength of light in decimetres

Question 20

How do you interpret optical rotation?

Answer 20

For a pair of glucoses or glycosides, alpha anomer has a higher [alpha]^t than bêta anomer under identical conditions

Question 21

What are some specific rotations of some common disaccharides?

Answer 21

D-Glucopyranose = +112.2 (alpha) or +18.7 (beta)
D-Galactipyranose = +150.7 (alpha) or +43.7 (beta)
D-Mannopyranose = +29.3 (alpha) or -17 (beta)

Question 22

How do you interpret an IR spectrum of an unknown disaccharide?

Answer 22

Present several characteristic features from which structural assignments can be made
In region of 750-950 cm^-1 there are three principal tests of absorption bands which are particularly useful for the assignment of anomeric configuration of carbohydrates