Polyalcohols from periodate oxidized seeds polysaccharide of

Int J Ayu Pharm Chem
RESEARCH ARTICLE
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Polyalcohols from periodate oxidized seeds polysaccharide of
Wrightia tinctoria R.Br. (Roxb.) plant by Smith degradation method
R. B. Singh 1
*
*
Department of Zoology, School of Life Sciences, Dr. B. R. Ambedkar University,
Khandari Campus, Agra-282002 U.P., (India)
Received: 7th September 2014/ Accepted: 31st October 2014 / Published: 10th November 2014
Greentree Group
©International Journal of Ayurveda and Pharmaceutical Chemistry, 2014
R B Singh
[email protected]
Int J Ayu Pharm Chem Vol. 1, Issue 2, 2014
Int J Ayu Pharm Chem
Abstract
Water soluble polysaccharide was extracted from Wrightia tinctoria R.Br. (Roxb.) seeds, on acid
hydrolysis with sulphuric acid and obtained hydrolysate on paper chromatography led to
separation of D-galactose and D-mannose in the molar ratio of 1:3. Periodate oxidized
polysaccharide on reduction with sodium borohydride by Smith degradation method followed by
acid hydrolysis (H2SO4), yielded polyalcohols as glycerol, erythritol and thritol in the molar ratio
of 1.20:4.95:0.009 on paper chromatogram. Derivatives of polyalcohols were produced from
seeds polysaccharide as glycerol tri-O-p-nitrobenzoate and tetra-O-tosyl-erythritol. The
absorbance of polyalcohols were recorded in photoelectrocolorimeter at 540 mµ for glycerol and
erythritol.
Keywords
Polyalcohols, Wrightia tinctoria seeds polysaccharide
INTRODUCTION
Wrightia tinctoria R.Br. (Roxb.) plant[1]
soluble polysaccharide contains D-galactose
belongs to the family-Apocynacceae and
and D-mannose in 1:3 molar ratio. In our
called as Indrajau. It occurs in Northern
earlier communications the nature of sugar
India, Eastern & Western Coast, Myanmar,
are obtained from the water soluble seeds
Nepal, Sri Lanka, Thailand and Australia.
polysaccharide[2], methylation studies[3] to
Medically the bark is used in the treatment
obtained
of nervous system disorder, Paralysis, fever,
determination of proposed polysaccharide
snakebites, dysentery, diarrhoea, piles and
structure and periodate oxidation studies[4]
leucoderma.
for
Powdered
seeds
are
the
the
methyl
sugars
confirmation
the
of
proposed
of
Wrightia
carminative tonic and useful in chest
polysaccharide
infection, asthma, leprosy and burning
tinctoria R.Br. (Roxb.) seeds. The present
sensation. Flowers and fruits are a source of
manuscript highlights the identification of
blue dye, fresh fruit
polyalcohols by Smith degradation[5] studies
juice used for
coagulating milk, and leaves are used as
of
wrapper for Bidis. Seeds yielded water
polysaccharide for the confirmation of
Greentree Group
125
the
structure
for
periodate
oxidized
seeds
Int J Ayu Pharm Chem
proposed seed polysaccharide structure of
hydrolysed with H2SO4 (1 N,
100 ml)
Wrightia tinctoria R.Br. (Roxb.) plant.
at 1000C for 12 hrs. Hydrolysate was
neutralized with barium carbonate slurry,
MATERIALS AND METHODS
filtered and the filtrate was deionised by
Separation of products
Amberlite Ion-exchange resins[12], IR-120
Polyalcoholic sugars were separated from
(H+) and IR-45 (OH-) then concentrated to a
periodate oxidized hydrolysed compounds
thin syrup.
by paper chromatographic analysis
[6]
on
RESULTS AND DISCUSSION
Whatman No. 3 MM filter paper sheet. The
following upper phase of solvent mixture
Characterization of polyalcohols
(v/v) were used as : (A) n-butanol, ethanol,
Hydrolysed syrup of periodate oxidized
water (4:1:5)[7], (B) ethyl acetate, pyridine,
seeds polysaccharide was resolved into its
[8]
water (2:1:2)
and used spray reagent as
component
by
paper
chromatographic
(R) acetonical silver nitrate, alcoholic
separation method on Whatman No. 3 MM
sodium hydroxide[9] was applied for the
filter paper sheet in solvent mixture (A) and
detection of polyalcohols.
The
used (R) as spray reagent to revealed the
evaporation was carried out under reduced
presence of three spots of polyalcohols
pressure at 45-500 C.
corresponding to the glycerol, erythritol and
thritol. The component sugar strips were cut
Identification of polyalcohols by Smith
out with the help of guide spots and eluted
degradation
with
water
according
to
the
Dent’s
Purified seeds polysaccharide (1.5 gm) were
method[13], which on evaporation gave
oxidized[10]
glycerol,
with
sodium
metaperiodate
(0.125 M, 30 ml) for 72 hrs at 4-80C in
refrigerator. It was further reduced
[11]
and
thritol
were
characterized and identified as follows:
with
Fraction-I: Glycerol
sodium borohydride (1 gm) for 24 hrs at
Sugar syrup (380 mg) was dissolved in
R.T. and excess periodate was removed by
ethanol (50 ml) and decolourized with
ethylene glycol (5 ml) to decompose the
aqueous solution of animal charcoal and
excess of periodate ions and reaction
filtered off. The filtrate was concentrated to
mixture was dialysed against running water
syrup and it moved a single spot on paper
for 48 hrs. It was concentrated to syrup and
Greentree Group
erythritol
126
Int J Ayu Pharm Chem
chromatogram
corresponding
to
the
chloride (1.5 gm), then the reaction mixture
authentic sample of glycerol. The residue
was left for 24 hrs at R.T. Content was
(350 mg) was dissolved in pyridine (5 ml)
poured into ice cold water (50 ml) to
and p-nitrobenzoyl chloride (2.4 gm) then
crystallized
the content was heated for 1 hr at 70-750C.
derivatives of erythritol, which were washed
The reaction mixture was poured into ice-
with water followed by ethanol were dried
cold solution of sodium bicarbonate to
in air. On recrystallisation with acetone and
obtain a precipitate which was filtered off.
ethanol
The filtrate gave the crystals of glycerol-tri-
erythritol, had m.p. & mixed m.p. 165-
O-p-nitrobenzoate derivative were obtained
1670C, Lit. m.p. 166-1680C[15].
on cooling the reaction mixture, which were
out
the
mixture
needle
gave
shaped
tetra-O-tosyl-
Fraction-III: Thritol
separated by filtration. It on recrystallisation
with acetone, had m.p. & mixed m.p. 188-
Sugar syrup (50 mg) moved as a single spot
1900C, Lit. m.p. 186-1880C[14].
on paper chromatogram parallel to thritol.
It was obtained in traces, which have Rf
Fraction-II: Erythritol
values more than the D-galactose and D-
Syrup (850 mg) was treated with aqueous
mannose was identified as thritol. The spot
solution of animal charcoal (50 ml), filtered
of thritol is visible only in ultraviolet light.
and the filtrate concentrated to a syrup. It
moved a single spot on paper chromatogram
Quantitative estimation of polyalcohols
corresponding to the erythritol. It was again
Polyalcohols were quantitatively estimated
dissolved in ethanol in ethanol (5 ml), on
by
cooling the crystal of erythritol was obtained
Respective polyalcohols were separated by
after recrystallisation with ethanol then
descending
filtered. It had m.p. & mixed m.p. 118-
chromatographic examined on Whatman
1200C, Lit. m.p. 117-1180C[14] and 120-
No. 3 MM filter paper sheets in upper phase
1220C[15].
of solvent mixture (B) and used (R) as spray
by
pyridine
(5
dissolving
ml),
Greentree Group
in
anhydrous
p-toluene
sulphonyl
acid
techniques
method[16].
of
paper
reagent. Polyalcohol components were cut
Derivative of erythritol syrup (200 mg) was
prepared
chromatographic
out with the help of guide spots and eluted
with
water
according
to
the
Dent’s
method[13], producing glycerol, erythritol
127
Int J Ayu Pharm Chem
and thritol in the molar ratio of 1.20 : 4.95 :
polyalcohols as glycerol, erythritol and
0.009. The colour intensity and absorbance
thritol in 1.20 : 4.95 : 0.009 molar ratio by
were
read
at
photoelectrocolorimeter
540
mµ
in
paper chromatographic analysis. The large
and
results
are
proportion of erythritol released with acid
given in Table-1.
hydrolysis of polyalcohols produced by
sodium borohydride serves as evidences that
Table-1 Absorbance of polyalcohols from Wrightia
tinctoria
R.Br.
(Roxb.)
seeds
polysaccharide
S.
No
.
Amount in
micrograms
the main polymer linkages are of (1→4)-βtype
Klett reading
(absorbance) at 540
mµ
Glycero Erythrito
l
l
with
D-galactopyranose
and
D-
mannopyranose units. The ratio of erythritol
to the amount of glycerol was obtained due
Glycero
l
Erythrito
l
1.
2.0
2.0
28
18
reducing end with (1→6)-α-type linkages in
2.
4.0
4.0
54
37
the main chain of the polysaccharide
3.
6.0
6.0
77
55
4.
8.0
8.0
100
73
5.
10.0
10.0
126
89
to the presence of D-galactose at the non-
structure. It indicated one branch point on
the average of
the
main
8 hexoses units in
chain
and
side
chain
in
polysaccharide structure. The derivative of
glycerol was obtained by usual manner as
glycerol-tri-O-p-nitrobenzoate
while
erythritol as tetra-O-tosyl-erythritol. The
absorbance of polyalcohols was recorded in
photoelectrocolorimeter on 540 mµ for
glycerol and erythritol. It indicated one
CONCLUSION
branch point on the average of seven
Wrightia tinctoria R.Br. (Roxb.) seeds
hexoses units are in the backbone and one
yielded a water soluble polysaccharide by
hexose unit in the non-reducing end for the
usual manner as D-galactose and D-
support
mannose in the molar ratio of 1 : 3. Purified
Greentree Group
method.
It
earlier
proposed
Wrightia tinctoria R.Br. (Roxb.) seeds
sodium borohydride and sulphuric acid by
degradation
the
polysaccharide structure of water soluble
oxidized polysaccharide was reduced with
Smith
of
polysaccharide as shown in Figure-1.
yielded
128
Int J Ayu Pharm Chem
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at Meerut College, Meerut (U.P.) on 4th-6th September, 2010, Abst. No.: OP-8, (2010),
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