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III. Studies on Soil Polysaccharides
G. D. SWINCER, J. M. OADES, A N D D. J . GREENLAND
When preceded by complete hydrolysis into monosaccharides and removal of interfering compounds from the hydrolyzate, colorimetric
methods can give useful estimates of the carbohydrate content of the soil,
either before or after extraction, and of crude extracts. The colorimetric
methods are normally applicable only to single classes of monosaccharides, such as the hexoses, pentoses, uronic acids, or hexosamines, and
even within a particular class none of the methods gives the same color intensity for equimolar concentrations of the different individual monosaccharides. Moreover, optimum hydrolysis conditions vary at least from
one class of sugars to another, and probably also from one individual
sugar to another. Clearly, therefore, the most precise determinations of
the proportion of the total carbohydrates extracted would require the
measurement of the amount of each individual sugar in both the soil and
Individual sugars, or the different classes of sugars, have been measured in soils and purified soil extracts (Graveland and Lynch, 1961;
Thomas and Lynch, 196 1; Ivarson and Sowden, 1962; Sowden and Ivarson, 1962; Gupta et al., 1963; Gupta and Sowden, 1965; Cheshire and
Mundie, 1966). Only Parsons and Tinsley (1961), Lynch et al. (1957,
1958), and Swincer et al. (1 968a,b) have attempted to relate the amounts
extracted with the amounts originally present in the soil or left in the
soil residue after extraction.
The results of Parsons and Tinsley are probably not accurate, particularly with respect to uronic acids, as the authors themselves admit. Although Lynch et al. (1 957, 1958) claimed to have measured the recovery
of the original carbohydrates in various extracts by separating and estimating seven different sugars, the yields reported were undoubtedly too
high as a result of the low values for “total soil carbohydrate” that must
have accompanied the very mild hydrolysis conditions used.
b. Polymer Degradation. The detection and evaluation of damage to
the polysaccharide molecules during the extraction process is by no
means easy. Positive and conclusive evidence against changes in the
carbohydrate polymers during extraction is virtually unobtainable because it is not yet possible to know the properties of these molecules before isolating them from the soil. Any information that can be obtained
must be either indirect or of a negative kind.
Whitehead and Tinsley ( 1964) made a useful indirect assessment of the
likely degradative effect of their extraction procedure on soil polysaccharides by subjecting several other natural polymers (starch, alginic
acid, chitin, cellulose, gluten) to the same treatment.
Bernier (1 958a) compared the viscosities of polysaccharides extracted
from soils by different reagents, and considered that the preparations with
higher viscosity were those that had suffered least degradation during
extraction. This is a worthwhile approach provided samples are free from
salt and ash.
Swincer et a f . (1968a) preferred to use the results from analysis of
extracts on Sephadex (3-25 as an indication of degradative treatments.
When reagents caused extensive fragmentation of polysaccharides by
hydrolysis or other means, a large proportion of the extracted materials
were “low molecular weight” according to gel filtration on Sephadex
(3-25. Thus, it was found that all methods involving high temperatures,
even methods based on water, led to extensive polysaccharide degradation, and 0.5 N NaOH at 20°C. was the most efficient, single, nondegradative extractant. Again it should be emphasized that such experiments
cannot provide conclusive evidence against degradation (e.g., by a relatively large average polymer size or the absence of specific degradation
products), even though they can give indications of whether damage has
occurred and thus help to rule out unsuitable extraction treatments.
3 . Extractants of Soil Polysaccharides
Extractants that have been used for obtaining polysaccharides from
soils include water, aqueous buffers and complexing reagents, dilute
mineral acids, organic reagents, and alkalis.
The yields of soil polysaccharides that have been obtained by various
workers using a number of different procedures are nearly all based on
gravimetric determinations and, therefore, provide only an approximate
guide to the efficiency of each extractant.
a. Water. Water has been a popular extractant for soil polysaccharides because of the simplicity of extraction, the relatively low
simultaneous extraction of humic materials, and the ease of subsequent
purfication. Most of the procedures involve extraction at temperatures of
at least 70”C., either with continuous shaking or by use of Soxhlet’s
apparatus (Mortensen, 1961; Keefer and Mortensen, 1963; Mortensen
and Schwendinger, 1963; Thomas et al., 1967). Forsyth ( I 950) found that
autohydrolysis of an extracted polysaccharide occurred in distilled water
at 1 OO’C., 90 percent of the arabinose and considerable amounts of ribose
being released in 24 hours. Furthermore, Swincer et al. ( 1968a) considered hot water extraction to be degradative on the basis of analysis of the
extract on Sephadex (3-25.
What quantitative information there is shows that hot water extracts
up to about 2 percent of the soil organic matter, which could represent
up to one-quarter of the total carbohydrates.
G. D. SWINCER, J. M. OADES, A N D D. J. GREENLAND
b. Aqueous Buffers and Complexing Reagents. Buffer solutions
(Bernier, 1958a) and complexing agents, such as sodium phyrophosphate
(Bernier, 1958a; Lynch et al., 1958; Mortensen and Schwendinger,
1963) and disodium EDTA (Barker et af., 1965, 1967), have been used
in attempts to obtain polysaccharides with a minimum of alteration from
their natural state. In general, yields were low (less than 5 percent of the
soil carbohydrate), and it is impossible to know whether the material
extracted was representative of the total. For this reason such mild reagents are probably of limited value for investigating the whole soil
polysaccharide fraction, in spite of their ideality in terms of the risk of
hydrolysis. A possible exception is the use of metal complex reagents
such as Schweitzer's reagent (cuprammoniurn hydroxide) for extracting
Schweitzer's reagent was first applied to soils by Daji (1932), and its
usefulness has been confirmed by Gupta and Sowden ( 1964). However,
it might well be replaced to advantage by one of the more recently developed metal complex reagents with high cellulose dissolving power
(Jayme and Lange, 1963).
c . Dilute Mineral Acids. Although inorganic acids are relatively inefficient reagents for extracting humic materials from soil (Mortensen and
Himes, 1964), they are more effective for the polysaccharide fraction.
Barker et al. ( 1 965, 1967) chose 0.6 N H2SO4as the extractant for routine
isolation of larger quantities of polysaccharide materials from an English
muck soil. Other reagents were superior in terms of yield, but the acid
extracts were more easily purified. Furthermore, treatment with acid
apparently hydrolyzed bonds between polysaccharides and humic materials, thus simplifying the separation of polysaccharides from these
materials. Acid hydrolysis of the polysaccharides themselves was prevented by carrying out the extraction at 3°C.
Black et af.( 1955) also used dilute mineral acid (0.09 N HCl) to extract
a polysaccharide complex from peat. The yield was low compared with
those obtained by other workers with different reagents: furthermore, it
is quite likely that some degradation occurred under the conditions used
( 1 hour at 60" to 70°C.).
H F at 20°C. was the most efficient acid extractant used by Swincer et
af.( 1 968a), but the extracts are not easily analyzed; thus, HCI was preferred and released 10 to 20 percent of the carbohydrate from a range of
great soil groups, being most efficient in soils rich in calcium carbonate
or free iron and aluminum oxides.
Another acid extractant that has been tried is a suspension in water of
the H+ form of a cation exchange resin (Amberlite 1 R-120) (Barker et al.,
1965, 1967). This treatment extracted slightly less carbohydrate material
than 0.6 N H&O, under the same conditions, and the mineral acid was
preferred for this and other reasons.
d. Organic Reagents. Parsons and Tinsley ( 1961) extracted polysaccharides from a variety of soils by double reflux with 0.2 N lithium bromide in anhydrous formic acid. The proportions of soil organic matter
(3.5 to 17.5 percent) extracted as polysaccharides are some of the highest
on record. However, after extraction, a considerable amount of carbohydrate still remained with each soil residue, and the authors themselves
admitted that the yields quoted were probably overestimates because
some of the C 0 2 evolved in the decarboxylation method for determining
uronic acids almost certainly arose from nonuronide materials. Deuel
et al. (1960) have established that ready decarboxylation is a general
property of the colored humic substances. I t is possible that the reducing
sugar content was also overestimated due to the presence of other reducing substances in hydrolyzates of the isolated polysaccharide materials.
Whitehead and Tinsley ( 1964) extracted high proportions of the organic
matter from several soils by refluxing with a mixed reagent consisting of
0.4 M boric acid, 0.4 M oxalic acid, and 0.2 M lithium chloride in dimethyl formamide (DMF). The polysaccharide contents of the preparations were not determined and attempts to separate the dissolved polysaccharides from the nonpolysaccharide components were not successful.
Although exposure to high temperatures was not prolonged, hydrolysis
of soil polymers almost certainly occurred as appreciable degradation
was observed when the procedure was applied to such natural polymeric
material as starch, alginic acid, chitin, and gluten. Swincer et al. ( I968a)
also considered this procedure to be degradative, based on analysis of
the extract by Sephadex (3-25.
DMF (without dissolved acids or salts) was also among the extractants
examined by Barker et al. (1965, 1967). Extraction was for 24 hours at
room temperature, and the carbohydrate yield was very low. However,
the yield was increased almost 30-fold if the soil was hydrogen ion saturated before extraction with DMF. Of the other organic extractants examined, N-methyl-2-pyrrolidone was only slightly more efficient than
DMF (without the pretreatment), but 8 M urea was quite superior. Not
enough data were given to allow adequate assessment of the potential
of the different reagents as extractants for soil polysaccharides.
The only soil examined was a muck, and relative yields could be quite
different in mineral soils where many of the polysaccharides are likely
to be strongly associated with inorganic colloids.
e. Alkalis. I n spite of some objections and criticisms, dilute NaOH,
G . D. SWINCER, J. M . OADES, A N D D. J. GREENLAND
the classical extractant for soil organic matter, has been used frequently
for isolating soil polysaccharides (Forsyth, 1947, 1950; Rennie et al.,
1954; Dubach et af., 1955; Chesters et al., 1957; Muller et af., 1960;
Graveland and Lynch, 196 1 ; Acton et al., 1963a; Dormaar, 1967; Oades
and Swincer, 1968; Swincer et al., 1968a,b). This extractant is more
efficient than most others, an observation which has been made repeatedly in studies of the extraction of other soil organic materials (e.g., Bremner
and Lees, 1949; Choudri and Stevenson, 1957; Evans, 1959). In contrast
to most other workers, Whistler and Kirby ( I 956) reported that hot water
and cold 0.5 N NaOH extracted comparable amounts of “purified” polysaccharide from an American soil. However, the amount of polysaccharide in their initial crude 0.5 N NaOH extract may have been much
higher than they reported, as considerable losses could have occurred
during the purification treatment.
Large amounts of humic materials are always extracted with the polysaccharides. The separation of these materials from the polysaccharides
has proved to be particularly difficult, and is a significant objection to
the use of this extractant.
Another objection to the use of NaOH has been the possibility of
damage either by hydrolysis of the polymers or by autoxidation (Bremner,
1954; Tinsley and Salam, 1961 ; Dubach and Mehta, 1963). There is good
evidence that some organic materials are, indeed, altered under alkaline
conditions (Bremner, 1950; Choudri and Stevenson, 1957; Evans, 1959)
although Kononova (196 1 ) discusses other evidence which, she says,
shows that extraction with alkaline solutions does not change the nature
of humic substances essentially. As these experiments were concerned
only with overall effects, the conclusions reached do not necessarily apply
directly to the polysaccharide materials, which constitute only a rather
small proportion of the extracted organic matter. Bremner (1 950) found
that the oxygen taken up during alkaline extraction of soils went into the
acid-insoluble (humic acid) fraction rather than into the fulvic acid fraction which contains most of the carbohydrates (Acton et af., 1963a;
Dormaar, 1967; Swincer et al., 1968a).
Some alkaline degradation of the polysaccharides might be expected.
Neuberger and Marshall ( 1966b) and Horton and Wolfrom (1 963) state
that glycosidic linkages between monosaccharides are ordinarily stable
to alkali, but that in certain circumstances a slow stepwise degradation
may take place from the reducing end of a polysaccharide molecule.
Saccharinic acids are formed and eliminated and new reducing groups are
exposed until this “peeling” reaction is intercepted by a “stopping” reaction in which release of the saccharinic acid is inhibited by an unfavor-
able glycosidic linkage. The rate of the “peeling” reaction, which can
occur in the absence of oxygen, is influenced by the position at which the
penultimate sugar residue is attached (Kenner and Richards, 1957;
Whistler and BeMiller, 1958). If this linkage is (1 +. 3) the terminal
residue is readily detached, but ( I +. 4) and ( 1 + 6) links are much more
stable, and ( 1 -+2 ) links are resistant to the action of alkali even under
very vigorous conditions.
Thus under mild alkaline conditions, some shortening of polysaccharide chains is possible even though there is no random fragmentation
such as occurs in hot mineral acids. The presence of oxygen in an alkaline solution of a polysaccharide can lead to additional damage. I n this
case the individual sugars may be oxidized without changes in polymer
size (by hydrolysis of glycosidic bonds). In order to reduce this risk,
Choudri and Stevenson (1957) advocated the addition of stannous
chloride to the soil sample before extraction with NaOH; other workers
(e.g., Bremner, 1950) have carried out alkaline extraction of soil organic
matter under an atmosphere of nitrogen.
Tinsley and Salam ( 196 1) mentioned two other undesirable features of
NaOH as an extractant of soil organic matter. First, silica is dissolved
from the mineral matter in the soil and contaminates the organic fractions
separated. This can be overcome by concentration of a solution of the
partly purified polysaccharide; this results in polymerization and precipitation of the silica (Swincer et al., 1968a).
Secondly, NaOH is known to dissolve some protoplasmic and structural components of fresh organic tissues. The problem is eliminated if,
before extraction, the free organic residues are first removed from the
soil by flotation sieving (Roulet et al., 1963a) or densimetric fractionation
with heavy liquids (Ford et al., 1969).
$ Sequential Extraction Treatments. No single reagent has given
complete extraction of soil polysaccharides, and for maximum extraction multiple treatments are necessary. This is so with other organic
materials, e.g., humic materials (Choudri and Stevenson, 1957; Bremner
and Harada, 1959) and organophosphates (J. K. Martin, 1964). Studies
on ten great soil groups showed that a double treatment, using I N HCI
followed by 0.5 N NaOH, removed more than SO percent of the soil carbohydrate in all except two cases (Swincer er d.,1968a). A third treatment using acetic anhydride containing 2.5 percent concentrated H & 0 4
at 60°C. for 2 hours brought the total yield up to 80 percent or more except in an ando soil and a red earth. Extraction with NaOH can be increased by ultrasonic dispersion. Thus the sequential procedure should
give almost complete extraction in many cases and although certain soils
G. D. SWINCER, J . M . OADES, AND D. J . GREENLAND
may require special approaches this procedure would appear to have
general applicability (Fig. 1). These results were obtained on soils after
the removal of partly decomposed plant remains by a densimetric procedure (Ford et d . , 1969).
Ultrasonic dispersion in a
LIGHT FRACTION - partly
decomposed plant and
I N HCI 20"
1 N HCI EXTRACT - add EDTA,
neutralize with NaHCO,.
Concentrate for gel filtration on
Sephadex and Biogel
Ultrasonic dispersion in
Acetic anhydride + 2.5%
coric. H2SO4 60"
0.5 N NaOH extract - pass
through H+ Dowex 50 and
Polyclar AT columns.
Concentrate for gel filtration
on Sephadex and Biogel
REACTION MIXTURE -dilute
tenfold with water and extract
acetylated polysaccharides in
CHCI,. Concentrate the extract
for gel filtration on Sephadex
FIG.1. Procedure for isolation of polysaccharides from other soil materials. The light
fraction usually contains from 10 to 50 percent of the total soil carbohydrate. Of the remainder, 10 to 20 percent occurs in the acid extract, 30 to 50 percent in the NaOH extract,
20 to 30 percent in the acetic anhydride extract, and 10 to 20 percent may be left in the final
residue. Details of this procedure are described by Swincer et af. (1968a,b).
C . PURIFICATION
I . Introduction
After removal of solids by centrifugation, the first step in the recovery
of polysaccharides from an extract depends on the particular extractant
used. The specific primary treatments that have been applied will be discussed very briefly before consideration is given to the more general
procedures used for subsequent purification.
SOIL POLY SACCHARIDES
2 . Primary Purijication Treatments
With water extracts, the initial purification treatment is simplified because the problem of removing solutes contributed by the extractant does
not arise. In fact some workers (Duff, 1952a; Whistler and Kirby, 1956)
have omitted the “primary treatment” and have applied the more general
procedures for purifying polysaccharides directly to the concentrated
extract. However, Mortensen and co-workers (Mortensen, 196 1 ; Keefer
and Mortensen, 1963; Thomas et al., 1967) favored an initial acidification step (adjustment to pH 2 with N HCI) to precipitate humic materials.
To neutralize acid extracts, Black et al. ( 1 955) used 40 percent NaOH
while Barker et al. (1 965, 1967) used NaHC03. The latter authors examined polysaccharides both in the resulting precipitate and in the
supernatant. The precipitated polysaccharides were brought into solution with 0.3 N HCI before the application of further purification treatments.
To avoid precipitation of polysaccharides during the neutralization of
acid extracts Swincer et al. (1968a) added EDTA before NaHCOa. The
EDTA prevented precipitation of di- and trivalent cations in the acid
extract and consequently hydroxides of metals did not occur as precipitates associated with polysaccharides.
Parsons and Tinsley (1 96 I ) used diisopropyl ether to precipitate
organic materials from anhydrous formic acid extracts of soil and Whitehead and Tinsley ( 1 964) treated DMF extracts with diethyl ether in the
presence of acetic acid. The precipitates were dissolved in water and
formic acid, respectively, in readiness for further purification treatments.
Where alkaline solutions have been used for polysaccharide extraction,
the first purification step has been acidification to pH 2 to 3 to precipitate
humic acids. The acidification treatment removes a large proportion of
the organic components of the extract, as well as some of the inorganic
materials, but the precipitate may also contain significant amounts of
carbohydrate material (Lynch et al., 1957; Acton ef al., 1963a; Dormaar,
1967). Dormaar ( 1967) reported that some humic acid precipitates contained up to 9 percent of the total soil carbohydrates, and Acton et al.
obtained values as high as 12 percent.
This association of carbohydrates with the humic acid fraction is
probably due to coprecipitation involving metal cations. Swincer et al.
(1968a) found that 8 percent of the soil carbohydrate was precipitated
with the humic acid fraction obtained by direct addition of acid to the
NaOH extract, but only 1 percent of the soil carbohydrate was present
in the humic acid fraction obtained by passage of the NaOH extract
through a column of coarse H+ Dowex 50. The humic acid fraction was
G . D. SWINCER, J. M. OADES, AND D. J. GREENLAND
trapped in the resin column, and the fulvic acid solution emerging from
the column was free of metal cations and was easily concentrated without the formation of precipitates.
Cellulose extracted from soils with Schweitzer’s reagent was recovered by precipitation with 5 volumes of 80 percent ethanol (Gupta
and Sowden, 1964). The precipitate was washed successively with water,
N HCl, and water before being dried. This product was reported to be
reasonably pure without further treatment.
3 . Secondary Purification Treatments
The purification procedures commonly applied to soil polysaccharide
extracts after the appropriate initial treatment include: (a) dialysis to
remove salts, solvents, and other low molecular weight materials; (b)
charcoal filtration to remove colored compounds; (c) precipitation of the
polysaccharides from aqueous solution by the addition of acetone,
alcohol, or other reagents; and (d) various deproteinization procedures.
Dialysis in Visking tubing against distilled water has been the most
commonly used purification treatment, having been applied in all cases
of extraction with hot water (Duff, 1952a; Whistler and Kirby, 1956;
Clapp, 1957; Mortensen, I96 1 ; Keefer and Mortensen, 1963; Mortensen
and Schwendinger, 1963; Thomas et al., 1967) and acid (Black et al.,
1955; Barker et al., 1965, 1967) as well as to phosphate buffer extracts
(Bernier, 1958a), a sodium pyrophosphate extract (Mortensen and
Schwendinger, 1963), and an acidified NaOH extract (Muller et al.,
1960). Losses of carbohydrate during dialysis of various extracts against
distilled water are considerable (Clapp, 1957; Muller et al., 1960).
Swincer et al. (1968a) showed that much of this material could be recovered from the dialysis water as low molecular weight fragments, but
not as monosaccharides. However, significant amounts were sorbed on
dialysis tubing and could not be removed by washing with water.
Forsyth (1 947) introduced the use of acid-washed animal charcoal for
the sorption of organic materials from classical fulvic acid extracts.
Charcoal will remove most of the organic materials and all the color from
such extracts and has been used to purify polysaccharides not only from
the fulvic acid fraction of NaOH extracts (Dubach et al., 1955; Muller
et al., 1960; Swincer et al., 1968a), but also from a hot water extract
(Whistler and Kirby, 1956) and phosphate buffer extracts (Bernier,
1958a). However, recoveries of polysaccharides from charcoal are poor.
Whistler and Kirby (1 956) measured a recovery of 14 percent and
Swincer et al. (1968a) reported a 12 percent recovery. Thus while relatively pure polysaccharides can be obtained by this procedure, losses
are very high. Swincer et a f . (1968a) increased recoveries to 30 percent
by using 8.5 N acetic acid as eluant, and even up to 65 percent by pretreating the charcoal with stearic acid as described by Dalgleish (1955),
but results obtained were difficult to reproduce.
Polyclar AT (cross-linked polyvinyl pyrrolidone) which is a strong
adsorbent of polyphenolic compounds (Sanderson and Perera, 1966)
proved to be most useful for separating colored materials from polysaccharides in “fulvic acid” solutions. Recoveries of carbohydrates from
columns of Polyclar AT were greater than 95 percent, and almost complete removal of colored compounds occurred when the pH was lowered
to 2 (Swincer et al., 1968a).
Very often polysaccharide materials have been recovered from
aqueous solution by precipitation with excess ethanol or acetone (Forsyth,
1947, 1950; Duff, 1952a; Rennie et al., 1954; Black et al., 1955; Chesters
et al., 1957; Bernier, 1958a; Mortensen, 1961; Salomon, 1962; Acton
et al., 1963a; Dormaar, 1967). Polysaccharides have been purified by
precipitation from 0.5 N lithium chloride solutions at pH 7 by the addition of cetyl trimethylammonium bromide (Parsons and Tinsley, 196 I ) ,
from formic acid solution with diisopropyl ether containing 1 percent
acetyl chloride (Whitehead and Tinsley, 1964),and from alkaline solution
with copper sulfate (Forsyth, 1950). The purity of some preparations was
increased by redissolving the precipitate in water and repeating the step,
sometimes with an alternative precipitant such as ether (Black et al.,
1955) or a mixture of alcohol and acetic acid (Duff, 1952a).
Although they have been widely used, these precipitation methods are
far from satisfactory, particularly when not preceded by a desalting step.
They are not specific for polysaccharides, as some noncarbohydrate
materials are also precipitated, and they leave polysaccharides in the
supernatant (Acton et al., 1963a; Dormaar, 1967). In fact, Acton et al.
( 1963a) found that the precipitate obtained by one purification procedure
(Rennie et al., 1954; Chesters et al., 1957) had an ash content of more
than 60 percent and a carbohydrate content of less than 20 percent, measured by an anthrone method (Brink et al., 1960). They also found that
there was much more carbohydrate in the supernatant than in the precipitate.
Much of the protein material that invariably remains with soil polysaccharide preparations, even after several different purification treatments, has been removed either by the emulsification procedure of Sevag
et al. (1938) (Bernier, 1958a; Mortensen, 1961; and Thomas et al.
1967) or by calcium sulfate precipitation and sorption on Fuller’s earth
in N acetic acid (Bernier, 1958a). However, amino acids were always
detected in hydrolyzates of the “deproteinized” polysaccharides.
Roulet et al. (1963b) used gel filtration and ion exchange chroma-
G . D . SWINCER, J . M . OADES, AND D . J. GREENLAND
tography with the primary purpose of purifying the humic substances in
acid extracts. The best separation of high molecular weight carbohydrates (measured as uronic acids) from “nitrogenous compounds” (as
well as from humic substances) occurred with Sephadex G-75, but no
pure fractions were recovered. The “nitrogenous substances” were
mainly of low molecular weight, but they occurred in all fractions. Oades
and Swincer (1968) and Swincer et al. (1968a,b) have confirmed the
partial separation of nitrogenous materials from carbohydrates by means
of gel filtration. This separation occurred because materials of low
molecular weight obtained by gel filtration invariably contained higher
proportions of amino acids than larger polymers. Thus materials separated
as the high molecular weight fraction on Biogel P- 100 contained the least
proportions of amino acids.
A more direct application of gel filtration to the purification of soil
polysaccharides has been made by Barker et af. (1 965, I967), who used
Sephadex G- 100 to remove humic materials from the high molecular
weight polysaccharides. Limitations of the technique were that good
separations could be achieved only with acidic extracts, and the problem
of separating humic materials from the smaller polysaccharides remained
Other purification procedures that have been used to advantage with
soil polysaccharides are desalting by ion exchange (Mortensen, 196I),
and removal of clay minerals by treatment with a mixture of 0.3 N with
respect to both H F and HCI for 10 minutes at 40°C. (Mortensen and
In summary, it can be said that in spite of numerous intensive attempts
to purify the polysaccharides extracted from soils, it is doubtful if any
of the resulting preparations have been completely free from noncarbohydrate materials. The possibility of carbohydrate-humic and carbohydrate-protein links will be considered later.
1 . Introduction
The large number of different sugars in the hydrolyzates of “purified”
polysaccharides and viscosity and ultracentrifuge studies (Bernier,
I958a; Ogston, 1958) which indicate a wide range of molecular size and
shape show that soil polysaccharides are complex mixtures. Their fractionation has thus proved to be even more difficult than fractionation of
other polysaccharide mixtures. Most of the well-established methods of
polysaccharide and polymer chemistry have been used, but in spite of a