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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5987_Библиотеки_им_академика_М_И_Перельмана.pdf
22
Pharmaceutical Chemistry of Natural Products
(a) Benedict’s, Fehling’s and Tollens’ reagents: Benedict’s reagent [an alkaline solution of copper
sulphate complexes with citrate ion (cupric citrate ion complex)], Fehling’s reagent
(mixture of copper sulphate and potassium sodium tartrate) and Tollens’ reagent
(ammoniacal silver nitrate solution) are mild oxidizing agents and oxidize the aldoses
to the corresponding aldonic acid. These reagents are, therefore, used for detecting the
presence of a reducing sugar in qualitative analysis. The sugars that are oxidized by these
reagents are known as reducing sugars.
Because α-hydroxy ketones are also oxidized to the acids, these reagents also oxidize
ketoses having α-hydroxy ketonic group. This explains why fructose reduces Fehling’s
reagent, Benedict’s reagent and Tollens’ reagent though it does not have any aldehyde
group.
(b) Bromine water: This mild oxidizing agent selectively oxidizes the –CHO group of aldose
and gives carboxylic acid called aldonic acid, which on evaporation gives a stable δ-lactone.
By using sodium amalgam in the presence of a trace of acid the lactone may be reduced to
the original aldose; the lactone may also be reduced to alcohol by adding aqueous lactone
to aqueous sodium borohydride. However, ketoses are not oxidized by bromine water.
CHOH
CH OH
2
CHO
CHOH
CHOH
CHOH
CHOH
CH OH
2
COOH
CHOH
CHOH
CHOH
CH OH
2
CO
CHOH
CHOH
HC
CHOH
CHOH
CH OH
2
CHOH
CHOH
CHOH
CH OH
2
[O] Evaporate NaBH
4
Alditol (Sorbitol)
Lactone
(Glucose- -Lactone)
-
Aldonic acid (Gluconic acid)
Aldose (Glucose)
Na/Hg; H
O
(c) Nitric acid: It is a strong oxidizing agent, and therefore, it oxidizes the aldehydic and
primary alcoholic groups of aldose to corresponding dicarboxylic acid known as aldaric
acid. Stronger oxidizing agents usually cleave the carbon chain of ketoses to form a
mixture of acids having fewer carbon atoms.
CHOH
CH OH
2
CHO
CHOH
CHOH
CHOH
CHOH
COOH
COOH
CHOH
CHOH
CHOH
HNO
3
Aldose
(Glucose)
Aldaric acid
(Saccharic acid)
CO
CH OH
2
CH OH
2
CHOH
CHOH
CHOH
Ketose
(Fructose)
HNO
3
Glycolic acid
CH OH
2
COOH
COOH
COOH
CHOH
CHOH
Tartaric acid
Carbohydrates
23
(d) Lead tetra-acetate and periodic acid: These reagents are frequently used in carbohydrate
analysis. These reagents oxidize the 1,2-glycolic portions of the monosaccharide molecules
with the cleavage of the C–C bond. Between every two 1,2-glycolic groups, one molecule
of the reagent is consumed.
R’
R
CHOH
CHOH
R’
CHO
CHO
R
For convenient handling, periodic acid is preferred over lead tetra-acetate. For determining
the size of the ring in aldoses, oxidation by periodic acid has been proved to be the best
method (refer to page 37 for detailed discussion).
4. Glycoside formation: It is one of the important properties of monosaccharide. Upon reaction
with methanolic hydrogen chloride, monosaccharide forms two isomeric compounds known
as methyl glycosides. In this reaction, the conversion of an aldehydic group to an acetal group
occurs.
Strong methylating agents (e.g. dimethyl sulphate) are required for the methylation of
hydroxyl groups of sugars. For example, the remaining four hydroxyl groups of methyl
glucosides can be methylated with dimethyl sulphate or methyl iodide to give methyl
tetramethyl glucoside. Acetals (and thus glycosides) are acid labile but stable in alkaline
medium. When treated with acid they undergo hydrolysis only at the hemiacetal linkage,
and not at other linkages to form tetramethyl glucose. This reaction is used in establishing
the structure of the glucose.
CH OH
2
CHO
(CHOH)
3
CHOH
CH
3
OH
Aldose
(Glucose)
-Methyl glycoside
(CHOH)
3
C
HC
CH OH
2
H
OCH
3
O
(CHOH)
3
HC
CH OH
2
HCO
3
H
O
-Methyl glycoside
Methylglycoside
(or )
(CHOH)
3
CHOCH
3
HC
CH OH
2
O
(CH)SO
32 4
(CHOCH )
33
CHOCH
3
HC
CH OCH
23
O
Methyl tetramethyl
glucoside
(CHOCH )
33
CHOH
HC
CH OH
2
O
Tetramethyl
glucose
H
HCl
24
Pharmaceutical Chemistry of Natural Products
5. Acetylation: Monosaccharides react with acetic anhydride and form the corresponding
polyacetate esters. The formation of sugar esters indicates the presence of alcohols in the
sugars. The total number of acyl groups that can be taken up by a molecule of sugar is the
measure of the number of hydroxyl groups. Both glucose and fructose form penta-acetyl
derivatives indicating the presence of fi ve hydroxyl groups in glucose and fructose.
OH
H
OH
OH
H
HO
H
H
CH OH
2
CHO
Glucose
OAc
OAc
OAc
H
H
AcO
H
H
CH
2
OAc
Glucose pentaacetate
CHO
5(CH CO) O
32
5CH COOH
3
H
OH
OH
HO
H
H
CH OH
2
CH OH
2
Fructose
O
O
H
OAc
OAc
AcO
H
H
CH OAc
2
CH OAc
2
Fructose pentaacetate
5(CH CO) O
32
5CH COOH
3
6. Reaction with phenylhydrazine: As the usual carbonyl group, aldoses and ketoses react with
one molecule of hydroxylamine to form oxime but they react with three molecules of
phenylhydrazine to yield osazones through phenylhydrazones.
The mechanism in this reaction has been studied by different scientists, but no defi nite
conclusions have been obtained. Therefore, it is concluded that different mechanisms may
apply under different conditions. Actually the mechanisms for osazone formation are of two
types.
(a) Fischer type mechanism: According to this mechanism, a molecule of phenylhydrazine
fi rst reacts with the aldose or ketose molecule to form phenylhydrazone. Then the
second molecule of phenylhydrazine oxidizes either the C
2
hydroxyl group (in case of
aldoses, e.g. glucose) or the C
1
hydroxyl group (in case of ketoses, e.g. fructose) of the
phenylhydrazone to a carbonyl group. The newly formed carbonyl group reacts with the
third molecule of the phenylhydrazine to yield osazone.
OH
H
OH
OH
H
HO
H
H
CH OH
2
CHO
Glucose
C H NHNH
65 2
HO
2
OH
H
OH
OH
H
HO
H
H
CH OH
2
HC
Glucose phenylhydrazone
NNHC H
65
C H NHNH
65 2
( CHNH, NH)
65 2
3
H
OH
OH
HO
H
H
CH OH
2
HC
Phenylhydrazone of
glucosone
O
NNHC H
65
Glucosazone
H
OH
OH
HO
H
H
CH OH
2
HC
NNHC H
65
NNHC H
65
C H NHNH
65 2
HO
2
C H NHNH
65 2
HO
2
Fructosazone
C
CH OH
2
(CHOH)
3
CH NNHC H
65
NNHC H
65
C
CH OH
2
(CHOH)
3
CH OH
2
O
Fructose
C H NHNH
65 2
HO
2
(CHOH)
3
C
CH OH
2
NNHC H
65
Fructose
phenylhydrazone
CH OH
2
Carbohydrates
25
This reaction reveals that even though glucose and fructose are different sugars they both
yield the same osazone.
Fischer’s mechanism of osazone formation suffers from the following objections:
(i) We know that phenylhydrazine is a powerful reducing agent, then how does it
behave as an oxidizing agent in this reaction?
(ii) It also does not explain why the reaction stops as soon as the two phenylhydrazine
residues are introduced at C
1
and C
2
of aldoses and ketoses.
(b) Weygand’s mechanism: In this mechanism, the phenylhydrazone (I) formed initially
undergoes Amadori rearrangement to give an intermediate (II), which loses aniline
through an intramolecular process to form iminoketone (III). The iminoketone (III) reacts
with two molecules of phenylhydrazine forming osazone accompanied with the loss of
ammonia.
Shortcomings of the Fischer’s mechanism are suitably explained by this mechanism. When
phenylhydrazone (prepared by the reaction of glucose with
15
N-labelled phenylhydrazine)
is treated with ordinary phenylhydrazine, unlabelled osazone is obtained with the
expulsion of labelled ammonia. This reaction supports Fischer’s mechanism. Osazone
from fructose formation can also be written in the same way.
C
C
R
H
H
O
O
H
C
C
R
H
H
O
H
C
C
R
H
O
H
II
H NNHC H
265
*
NNHC H
65
*
(N = N)
15
*
I
H
N
*
NHC H
65
CHNH
65 2
C
C
R
H
O
NH
*
H NNHC H
265
C
C
R
H NH
*
NNHC H
65
NH
3
*
H NNHC H
265
C
C
R
H
Osazone
NNHC H
65
NNHC H
65
III
But Weygand’s mechanism also fails to explain why only the fi rst two carbon atoms
are involved in osazone formation. However, in 1944, Fieser and Fieser suggested that
as soon as the osazones are formed they are stabilized by internal hydrogen bonding
(chelation) and hence no other (except C
1
and C
2
) carbon atom takes part in osazone
formation. Thus, for comparing confi gurations at asymmetric centres below C
2
in aldoses
and ketoses, osazone formation is an important tool.
26
Pharmaceutical Chemistry of Natural Products
A convenient way of identifying a pair of epimers is by the preparation of osazones. In the
formation of osazones, asymmetry at C
2
of the aldose is lost and thus epimers (e.g. glucose
and mannose) yield identical osazones.
D-Glucose
D-Mannose
D-Fructose
CHO
OH
H
HHO
OHH
OHH
CH OH
2
CHO
H
HO
H
HO
OH
H
OHH
CH OH
2
C
O
HHO
OHH
OHH
CH OH
2
CH OH
2
Identical configurations
at C ,C ,C
345
3C H NH.NH
65 2
3C H NH.NH
65 2
3C H NH.NH
65 2
CH
C
HHO
OHH
OHH
D-Glucosazone
NNHC
6
H
5
NNHC
6
H
5
CH OH
2
7. Action of alkali: Monosaccharides react with alkalis in different ways.
Upon warming with concentrated alkalis, the sugar fi rst turns yellow, then brown and fi nally
resinifi es.
In the presence of dilute alkali or amines, these sugars undergo rearrangement to form a
mixture of more than one sugar. For example, in the presence of very dilute alkali (0.02
N NaOH), glucose is converted into a mixture of glucose, mannose and fructose (Lobry-de
Bruyn–van Ekenstein rearrangement).
8. Reaction with hydrogen cyanide: Aldose reacts with hydrogen cyanide and the cyanohydrins
formed can be hydrolysed to the higher aldonic acid, which is then converted into two
isomeric sugars.
This is one of the most important synthetic reactions for the preparation of sugars, and it is
used to prepare aldoses of up to 10 carbon atoms (refer to Kiliani’s synthesis under page 18
for details).
Carbohydrates
27
CHO
OHH
HHO
OHH
OHH
D
( )-Glucose
C
C
OH
HHO
OHH
OHH
CHO
HHO
HHO
OHH
OHH
OHH
Enediol
D
( )-Mannose
0.02 N NaOH
CH OH
2
CH OH
2
CH OH
2
C
HHO
OHH
OHH
O
D
( )-Fructose
CH OH
2
CH OH
2
Ring Size of Monosaccharides
The fi ve- and six-member oxygen-containing heterocyclic rings (furan and pyran, respectively)
of monosaccharides bear a normal relationship to the heterocyclic compounds furan and pyran,
respectively.
O
Furan
O
Pyran
O
1
2
3
4
5
O
1
2
3
4
Hence, the fi ve- and six-member rings in the structures of sugars (e.g. glucose) are indicated by
α-D-glucofuranose and α-D-glucopyranose, respectively, which indicates the oxygen heterocyclic
ring (furanose = furan and pyranose = pyran).
28
Pharmaceutical Chemistry of Natural Products
Thus, the two glucopyranose and two glucofuranose may be represented as below.
O
OH
OH
H
H
OH
OH
H
HCOH
1
2
3
4
O
OH
OH
H
H
OH
H
OH
HCOH
1
2
3
4
O
H
OH
H
OH
OH
H
OH
H
H
O
H
OH
H
OH
H
HO
OH
H
H
CH OH
2
CH OH
2
CH OH
2
6
5
CH OH
2
6
5
- -GlucopyranoseD - -GlucopyranoseD - -GlucofuranoseD - -GlucofuranoseD
Structures are represented by thin lines throughout the book for the sake of simplicity.
Ring structure of aldoses
The monosaccharides, polyhydroxy aldehydes (e.g. glucose) or ketones (e.g. fructose) with open
chain formula failed to explain the following properties:
1. Glucose does not yield a bisulphite (upon reaction with NaHSO
3
/KHSO
3
) and aldehyde–
ammonia compound (upon reaction with NH
3
).
2. Glucose does not answer for aldehyde with Schiff’s reagent.
3. When glucose is dissolved in water its specifi c rotation gradually changes until it reaches
a constant value. The phenomenon known as mutarotation is exhibited by almost all the
reducing sugars (except few ketoses).
4. Two isomeric penta-acetates are obtained upon acetylation of glucose, neither of which reacts
with carbonyl reagents.
5. Glucose reacts with only one molecule of methanol in presence of hydrogen chloride, to
form two isomeric compounds, each containing one methoxy group exhibiting the properties
of acetals. They have different melting points, specifi c rotations and enzymatic hydrolyses.
Normal aldehyde usually reacts with methanolic hydrogen chloride to yield hemiacetal (I)
and acetal (II).
R
C
H
O
CH OH
3
R
C
H
OH
OCH
3
H
CH OH
3
R
C
H
OCH
3
OCH
3
Aldehyde
Hemiacetal (I) Acetal (II)
H
One of the ethers called the
α
-form undergoes ready hydrolysis by maltase, and the β-form is
hydrolysed by emulsin. Hydrolysis of both the ethers, whether carried out with enzymes or acids
(but not by alkali), always yields only D(+) glucose. These glucosides do not undergo mutarotation,
and does not react with Tollens’ or Fehling’s reagent.
D( )-Glucose
CH OH
3
HCl
Methyl- − -glucoside D
m.p.
[]
D
165°C
157°
Methyl- − -glucoside
D
m.p.
[]
D
101°C
33°
Carbohydrates
29
This behaviour can be demonstrated on the basis that the carbonyl group is not free in glucose
but bound up in a form that destroys its reducing properties. On this basis and the stability of
γ-lactones of the aldonic acid, Tollens proposed a cyclic hemiacetal structure (γ-oxide ring structure)
for D(+) glucose.
CHOH
OHH
HHO
H
OHH
O
*
Ring structure
CHO
OHH
HHO
OHH
OHH
Open-chain structure
CH OH
2
CH OH
2
D
(+)-Glucose
The objections against the open-chain structure are explained by the ring structure for glucose
in the following ways:
1. As the ring structure of glucose is not having free aldehyde group, it does not react with certain
weak carbonyl reagents (does not restore Schiff’s reagent colour, does not form a bisulphite
or an aldehyde–ammonia compound), which are characteristic for aldehydes. But it gives
certain reactions of the carbonyl group, such as the formation of cyanohydrins, oxime and
phenylhydrazone, because the strong carbonyl reagents such as HCN, NH
2
OH, C
6
H
5
NHNH
2
are able to open the ring to a free aldehydic group that reacts with these reagents.
2. One new chiral centre (C
1
, marked by asterisk) has been introduced in the ring structure and
hence gives two different isomeric glucoses and its derivatives. Practically, glucose, methyl
glucoside and glucose penta-acetate are found to exist in two forms, i.e. α and β. By studying
the various aldoses it has been found and also confi rmed by both physical and chemical
methods that in the α-form the hydroxyl group or substituted hydroxyl group is attached to
the right, and in the β-form the hydroxyl group is attached to the left.
C
OH
H
H
OH
H
OHH
OHH
O
CHO
OHH
HOH
H
OHH
OH
CH OH
2
CH OH
2
C
OHH
HOH
H
OHH
HOH
O
CH OH
2
-D-Glucose -D-Glucose
Aldehydic form
30
Pharmaceutical Chemistry of Natural Products
The two isomers differing only in the confi guration of C
1
(in aldoses) or C
2
(in ketoses) are
known as anomers, while such carbon atom is known as anomeric carbon atom.
The existence of two methyl glucosides and penta-acetates can be explained in same way.
H
RO
H
H
H
OR
OR
OR
H
O
RO
RO
H
H
H
OR
OR
H
H
CH OR
2
CH OR
2
O
Methyl glucoside when R = CH
Glucose penta-acetates when R = COCH
3
3
3. Mutarotation: The cyclic structure can open and reclose, which allows the rotation to occur
about the carbon bearing the reactive carbonyl to yield two distinct confi gurations (α and
β) of the hemiacetals and hemiketals. The carbon about which this rotation occurs is called
anomeric carbon and the two forms are called anomers. Carbohydrates can change spontaneously
between α and β confi gurations. This process is known as mutarotation.
When glucose crystallized from water below 50°C is dissolved in water, its initial specifi c
rotation of +113° falls gradually to a constant value of +52.5°; similarly, when the glucose
crystallized from water above 95°C is dissolved in water, its initial specifi c rotation of +19.5°
gradually rises to +52.5°. This phenomenon is known as mutarotation in glucose.
OHH
HHO
H
OHH
OHH
O
CHO
OHH
HHO
OHH
OHH
CH
2
OH
OHH
HHO
H
OHH
HHO
O
Aldehydic form
(36%)
(64%)
CH OH
2
CH OH
2
-Glucose
[ ] 113°
D
-Glucose
[ ] 19.5°
D
[ ] 52.5°
D
Carbohydrates
31
The ring structure for glucose explains that the change in specifi c rotation is due to
interconversion of the α-form [(α)
D
= +113°] of glucose to β-form [(α)
D
= +19.5°) and vice versa
through the aldehydic structure till an equilibrium is reached between the two structures. The
specifi c rotation value of this equilibrium mixture corresponds to +52.5°.
The question at our hand is how the open-chain form of glucose is obtained from the ring
form. It can be explained that this happens because protonation by acid HA and deprotonation by
base B takes place at the same time.
C
C
OHH
C HHO
C OHH
C OHH
H O
C
C
OHH
C HHO
CH
C OHH
O H
O
H
Aldehydic form
- D( )-Glucose -( )-GlucoseD
C
C
OHH
C HHO
CH
C OHH
H O H
:B
O
H
A
CH OH
2
CH OH
2
CH OH
2
The concentration of aldehydic form can be increased by adding methanethiol and esterifi cation,
followed by the removal of methanethiol.
CHO
OHH
HHO
OHH
OHH
Glucose
OHH
HHO
OHH
OHH
Pyridine
CH(SCH
3
)
2
OAcH
HAcO
H
H
CHO
OAcH
HAcO
OAcH
OAcH
CH
2
OH
2CH SH
3
–H O
2
CH
2
OH
CH(SCH )
32
Glucose dimethyl mercaptal
Ac O
2
CH OAc
2
HgCl
2
H O/CdCO
23
2,3,4,5,6-Penta-acetyl
glucose
CH OAc
2
OAc
OAc
Two types of ring structures: Fischer isolated a new D-glucoside, which differs from that of already
known α- and β-glucosides and called it γ-D-glucoside. But later on Haworth explained that this
γ-glucoside was found to be a mixture of two different glucosides (α and β) different from that of
previously known α and β-glucosides. These observations reveal that the sugars have two different
ring structures, namely γ- and δ-oxide ring structures.