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Experiment 1 Carbohydrates
Experiment 1 Carbohydrates
d sweet Classifications: a. Monosaccharides simple sugars with saccharide group, which cannot be hydrolyzed further b. Oligosaccharides 2 to 10 saccharide groups linked by acetal or ketal linkages can be hydrolyzed to their respective monosaccharides or to two molecules of disaccharides. c. Polysaccharides may be branched or long linear monosaccharides in glucosidic linkages d. homopolysaccharides yield only one kind of monosaccharide upon hydrolysis (starch and glycogen) e. heterosaccharides yield a mixture of monosaccharides and derived products Cellulose and agar-agar polysaccharides with structural function For energy storage dextrin, glycogen, insulin and starch Glycogen animal starch; absent in brain but present in liver, muscles and other tissues - abundant in oysters and other shellfish - upon hydrolysis with amylase maltose - upon hydrolysis with acid glucose - 1,4- and 1,6- alpha glycosidic bonds giving a branched structure. Reagents Na2CO3 Sodium citrate CuSO4 Sample Starch RHO + 2Cu2+ + 4OHGlycogen Solubility of Polysaccharides Schweitzers Test [Cu(NH3)4(H2O)2 ](OH)2 Cellulose Amorphous precipitate + precipitation by addition of dilute acetic acid + reprecipitation of cellulose by 95% ethanol + also soluble in NaOH and CS2 for making viscose (precursor of rayon) + Iodine + polysaccharides colored complexes + alpha-amylose deep blue color + amylopectin (other component of starch), cellulose red brown solution + mono & disaccharides too small for I2 + differentiate helical from non helical polysaccharide + hemicelluloses (GA & A) vs cellulose may be hydrolyzed by dilute mineral acid RCOOH + Cu2 O + 2H2 O Chemical Reactions Visible Results Brick-red precipitate Brick-red precipitate Principle Involved + reducing sugars
y y y
B C 1
HCl ZnCl2
Cellulose
soluble
Starch
Glycogen
Deep blue solution without NaOH Wine-red solution Or red precipitate Reddish-brown solution Colorless or yellow solution Colorless or yellow solution Colorless or yellow solution
Glucose
E I Glucose Molisch Test (Naphthol Reaction) Molisch Reagent (a-naphthol in 95% Ethanol) Conc. H2SO4 dehydrates CHO
Qualitative Tests for Carbohydrates General Color Reactions Reddish-violet ring at junction Reddish-violet ring at junction Reddish-violet ring at junction
O OH
Sucrose
Starch
+ H2SO4 dehydrates pentoses to furfural and hexoses to 5hydroxymethylfurfural + glycosidic bonds are hydrolyzed + condensation with anaphthol to produce purple product + intensity dependent on amount of CHO + same principle as above + used in the assay of carbohydrates carried under appropriate conditions + sugars react with 3 mol PhNHNH2 to form 1,2diphenylhydrazones or osazones + Ketoses with configurations identical to aldoses below C-2 give the same osazones e.g. glucose and fructose + sucrose hydrolysis glucose + fructose + lactose tufts of short fine crystals + for reducing sugars with free aldehyde or ketone group + osazones of maltose and lactose are formed upon cooling + has characteristic MP and shape + reducing sugars + free or potentially free ketone or aldehyde substituent + alkali present brings about considerable decomposition of sugar into reactive fragments which may reduce metal ions
Glucose
+
2 Anthrone Test Anthrone reagent (0.2% in H2SO4 ) Sucrose
H 2SO 4
OH O
Blue/bluegreen solution Blue/bluegreen solution Blue/bluegreen solution Yellow crystalline precipitate (5 minutes) Osazone soluble in hot water Yellow crystalline precipitate (glucosazone obtained by heating) (30 minutes) Yellow crystalline precipitate (20 minutes) Brick-red precipitate Brick-red precipitate Brick-red precipitate Brick-red precipitate Brick-red precipitate
Carbohydrate + H2SO4
OH O
Starch
O OH
Glucose
Maltose
Sucrose
The Crystals:
Galactose Glucosazone (broom-shaped) Glucose Fructose 4 Benedicts Test Xylose Arabinose Maltose maltosazone (sunflower-shaped) galactosazone (fluffy ball shaped)
Sucrose
Blue solution
Starch
Blue solution
Glucose Fructose 5 Barfoeds Test Barfoeds Reagent (Cu(CH3COO)2 Acetic Acid) Maltose
Brick-red precipitate Brick-red precipitate Brick-red precipitate (slow) Blue solution Yellow to faint pink color Cherry-red solution Green solution Cherry-red solution Presence of Pentoses Green, brown or reddish brown solution Green, brown or reddish brown solution Blue-green solution Blue-green solution
H3 C
OH
Sucrose Glucose
Seliwanoffs Test
+ has quantitative version since amount of precipitate is the basis of the test + di and polysacc. may be hydrolyzed to monosacc. + sucrose the only nonreducing sugar + carried out at pH 4.6 + differentiates di and monosaccharides + monosaccharides are more readily oxidized + reducing disaccharides (slow) lactose but not sucrose + 2-3 mins. for monosaccharides + differentiating for aldohexoses and ketohexoses + pentoses green (-) + keto cherry-red + aldo - yellow to pink + ketohexoses are dehydrated faster + boil for NMT 20-30s to avoid false + results
F Glucose Orcinol (m-methylresorcinol) Condenses furfural to colored compounds FeCl3 in HCl decomposition to furfural
Fructose
Xylose
+ condensation reaction
OH HO
HO OH
CHO (
HO
CH 3
HO
+ condensation reaction
Glucose
HO
Lactose 3 Mucic Acid Test Conc. HNO3 Galactose
OH H H H O H OH OH
HNO 3 (
HO H HO H
COOH
OH
H HO
H
COOH
OH
Bakers yeast
Effervescence
Zymase + H3PO4 + coenzyme phosphorylation of sugar forming hexose PO4 CO2 + ethanol hexose hexose diphosphoric acid phosphor-glyceric acid pyruvic acid acetaldehyde ethanol
+ Mucic Acid = galactaric acid + differentiating test from fructose and glucose + galactose is [O] by HNO3 to mucic acid (sandy crystals) + lactose reducing disacc. gluc + galac + yeast ferments only the naturally occurring dsugars. Some yeasts do not ferment lactose or dgalactose at all. + Einhorn saccharometer determine amount of CO2 evolved + zymase enzyme in yeast which produces fragmentation of sugar molecule + 10-15% ethanol because it inhibits action of yeast + invertase sucrose to glucose and fructose + addition of PO43- salts catalyzes by buffering action + CHI3 hexagonal crystalline shape under the microscope
2NAOH + I2 NaIO + NaI + H2O C2H5OH + NaIO CH3CHO + NaI + H2O CH3CHO + 3NaIO I3CCHO + 3NaOH I3CCHO + NaOH CHI3 + HCOONa
CaCl2 TS
CaCO3 + HCl
White precipitate
Classification Procedure:
Haworth projection
OH H HO H
H H H HO OH OH H
OH O OH H OH H H OH H H
Classification
Simpler fragments
Glucose
H OH H
O OH H
Mono
Hexose
Red
OH
O OH
Fructose
HO H H
O H OH OH OH O
OH
Mono
Hexose
Red
Galactose
H HO HO H
OH H H OH OH
HO
Mono
Hexose
Red
Sucrose
Di
Non-red
Glucose, Fructose
Maltose
Di
Red
Glucose
Lactose
Di
Red
Galactose, Glucose
Arabinose
H HO H
OH H OH OH
O
Mono
Pentose
Red
Xylose
H HO H
OH H OH OH
Mono
H
Pentose
Red