Changes in the chemical composition of sugar cane (Saccharum officinarum) during storage.
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Date
1973
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Abstract
An outline is given of the South African sugar industry,
with particular emphasis on the unit operations which make up the
industrial process for manufacturing sugar from cane.
Current knowledge of the chemistry of soluble polysaccharides
is reviewed and the structures of several polysaccharides,
including starch, dextran, and pullulan, are discussed.
It has been found that changes take place in the chemical
composition of the juice in sugar cane (Saccharum officinarum) during
post-harvest storage. With increasing storage time, there is a proportional
decrease in the starch content of the juice, and a considerably
larger proportional increase in the soluble polysaccharide
content. The increased polysaccharide content was found to be due to
a single glucan which, contrary to most previous publications on this
subject, is definitely not a dextran. Following structural analysis,
it has been established that the polysaccharide formed in stored cane
had not been described before and the name "sarkaran" , derived from
the Sanskrit word "Sarkara", meaning "sugar" is proposed for it.
The polysaccharide was isolated from cane juice by precipitation
with ethanol after the starch in the juice had been
removed by centrifugation. The polysaccharide was purified by
repeated dissolution in water and reprecipitation with ethanol.
Analysis by gel chromatography resulted in a single
symmetrical peak, indicating that the isolated polysaccharide is
homogeneous. This was confirmed by hydrolysing fractions representing
a section of the ascending and a section of the descending
part of the peak of the chromatogram, using the enzyme pullulanase.
Chromatographic separation and quantitative analysis of the isolated
oligosaccharides showed that the compositions of the two enzymes
digests were identical.
Acid hydrolysis of the polysaccharide resulted in a
single hexose. This was identified as glucose by paper chromatography,
comparing the Rf value with that of pure glucose. Confirmation
was obtained by comparing the osazone with that of
glucose, using microscopic examination and determination of the
melting points.
Paper electrophoresis showed the molecule to be
uncharged.
Several techniques, both absolute and non absolute,
were used to determine the molecular weight of the polysaccharide.
A method involving viscosity determination indicated a molecular
weight of 34 000 while a figure of 50 000 was obtained by gel
chromatography on a Sephadex column, comparing the peak elution
volume of the polysaccharide with that of dextrans of a defined
molecular weight. Both these techniques are non absolute and
yield rough estimates of the molecular weight. Osmometric
measurement, an absolute method, showed the number average
molecular weight to be 51 500. An absolute value for the weight
average molecular weight of 250 000 was obtained by light
scattering techniques. Data from the light scattering experiments
were also used to determine a value of 200 - 250 A for the radius
of gyration RG of the polysaccharide. End group analysis after
exhaustive methylation resulted in a value of 24 000 for the
number average molecular weight Mn. This indicates either that
some degradation of the polysaccharide molecule occurs , during the
methylation procedures or that there is a certain degree of
association between individual molecules.
Periodate oxidation showed that 32 percent of the
glucosidic linkages are in ( 1 + 6 ) position.
The polysaccharide was exhaustively methylated by
several Haworth methylations followed by a number of Kuhn methylations.
The fully methylated product was methanolysed and the
methyl glucopyranosides analysed by gas liquid chromatography. The
results were compared with those obtained from fully methylated
starch and dextran. From the absence of disubstituted methyl
derivatives in the methanolysate it was concluded that the
polysaccharide is an unbranched glucan.
From the quantities of Methyl 2,3,4,6 tetramethyl-O-Dglucopyranoside,
Methyl 2,3,6, trimethyl-O-D-glucopyranoside and
Methyl 2,3,4? trimethyl-0-D-glucopyranoside, it was concluded that
the only linkages in the glucan are ( 1 + 4 ) and ( 1 + 6 ) and
that these are present in the ratio 68:32.
Enzymic hydrolysis, using pullulanase, was followed by
paper chromatographic separation. Quantitative determination of
the oligo-saccharides present in the enzyme digest resulted mainly
in two oligosaccharides, maltotriose and maltotetraose, in nearly
equal proportions. For this reason it was postulated that the
polysaccharide is a maltotriose-maltotetraose polymer, and that
the individual units are linked in ( I + 6 ) position, a linkage
for which pullulanase is specific in certain configurations.
The sequence of the maltotriose and maltotetraose units
in the polymer has not been investigated further, although this
could be carried out by partial acid hydrolysis, followed by
isolation and identification of the various oligosaccharides
formed. An alternate method for the determination of the sequence
of the monomers is discussed.
It was subsequently shown that the linkages in the
polysaccharide are in the a configuration. The polysaccharide
is highly dextra rotary and the magnitude of the rotation is comparable
to that of other polysaccharides linked in a position, .
such as starch and dextran.
Infrared spectroscopy was used to confirm the configuration.
The spectrogram of the polysaccharide contained an absorption
peak at 840 cm-1 , which is typical of the a-anomeric absorption
occurring, for example, in the IR spectrum of starch. The spectrogram
exhibited no absorption peak at 891 cm-1 , the wavelength
typical of the B-anomeric absorption in the IR spectrum of cellulose.
In addition, it was found that all polysaccharides containing
a ( 1 + 4 ) linkages show an absorption peak at 700 cm 1. This
absorption peak was absent in all IR spectra obtained from
various dextrans. This phenomenon has not been reported previously
and it is suggested that the presence of this absorption peak in
the IR spectrum of a glucan can be used to support the evidence of
the presence of a( 1 + 4 ) linkages.
It was not possible to correlate the formation of the
polysaccharide with the occurrence of a specific micro organism.
It is suggested that the formation of the polysaccharide is the
result of enzymic reactions in the sugar cane after harvesting.
The investigation of the composition of juices from
deteriorated cane has not been confined to polysaccharides.
Ethanol has been isolated from the juice of some samples of stored
cane which had been burnt before harvesting. The ethanol was
isolated by fractional distillation and identified by measurement
of the boiling point. It was confirmed, by the formation of the
molybdate-xanthate complex, that the product isolated was an
alcohol. The identification was further confirmed by oxidising
the ethanol to acetic acid and proving the identity of the acids
by paper chromatography.
It has been shown that, with the exception of two acids,
the carboxylic acid composition of cane juice remains unaltered
during post-harvest storage of the cane.
The two exceptions , succinic and aconitic acids, were
identified from their melting points and by specific spot tests.
Ion exchange was used to isolate the acids from the juice. The
eluate from the ion exchange column was concentrated and the acids
separated by liquid-liquid chromatography, using a silica gel column.
The levels of both aconitic and succinic acids were found to increase
during the early period of storage but decreased again slowly thereafter.
The percentage change was greater in the case of succinic
acid, although aconitic acid was the most abundant carboxylic acid
in the juice.
Lactic acid was absent from the cane juices analysed.
This is surprising, as lactic acid is a common product of the metabolism
of carbohydrates by micro organisms. It is suggested that
the changes in acid composition during the storage of harvested
cane are caused by deactivation of enzymes of the Krebs cycle.
Post-harvest deterioration of sugar cane can have serious
consequences which can affect the whole Sugar Industry. Not only is
crystallisable sugar lost but the products of the deterioration have
adverse effects on factory processing and laboratory analysis. The
problem, which will become more acute with the introduction of
mechanical cane harvesting, can only be resolved through the cooperative
efforts of all the parties concerned.
Description
Thesis (Ph. D.)-University of Natal, Durban, 1973.
Keywords
Sugarcane--Analysis., Theses--Chemical engineering.