studies on acacia exudate gums. part v. structural features of acacia seyal

Studies on acacia exudate gums. Part V. Structural

features of Acacia seyal

C. Flindta,b, S. Al-Assaf a, G.O. Phillipsc,*, P.A. Williamsd

aThe Glyn O. Phillips Hydrocolloids Research Centre, North East Wales Institute, Mold Road, Wrexham LL11 2AW, UKbWolff and Olson (GmbH&Co), P.O. Box 10 66 20, D-20044 Hamburg, GermanycPhillips Hydrocolloid Research Ltd, 45 Old Bond Street, London W1S 4AQ, UK

dCentre for Water Soluble Polymers, North East Wales Institute, Mold Road, Wrexham LL11 2AW, UK

Received 28 November 2003; accepted 16 September 2004

Abstract

An investigation of the molecular structure of Acacia seyal relative to Acacia senegal provides an indication of the components which

influence emulsification effectiveness. Samples of A. seyal var. seyal and A. seyal var. fistula were fractionated using gel permeation

chromatography. The fractions and the whole gum were analysed to determine the molecular weight, sugar content, amino acid, protein

content, nitrogen, and intrinsic viscosity. The results confirm earlier findings that samples of A. seyal, as a broad grouping, have weight

average molecular weights several times greater than A. senegal, due to the greater proportion of the high molecular weight component.

Although the molecular weight of A. seyal is considerably greater than A. senegal, the intrinsic viscosity is less. The structure is, therefore,

more compact, than the structure of A. senegal. The sugar composition and amino acids in each of the gums are identical but are present in

different proportions, which is the main reason why A. seyal is dextrorotatory and A. senegal is laevorotatory.

The distribution of the protein is different between the various components which constitute the gums. In A. senegal the protein is mainly

located in association with the high molecular weight component (AGP-peak 1). Enzyme hydrolysis points to two components being

associated with the high molecular weight material in A. seyal, only one of which is degraded by the enzyme pronase. In hydrophobic

fractionation studies, a protein rich component of extremely high molecular weight (Fraction 3) was found in A. seyal but not in A. senegal. In

size fractionation this would co-elute with the main component, which is further evidence for the presence of the two different high molecular

weight components in A. seyal, unlike A. senegal. The adsorption of these high molecular weight protein fractions of A. seyal onto an oil

droplet was tested and found not to be a highly efficient emulsifying component. Structurally, while A. seyal may have the same core

structural linkages as A. senegal, the degree of branching is greater, with the protein distributed differently.

q 2004 Elsevier Ltd. All rights reserved.

Keywords: Emulsification; Molecular weight; Molecular structure

1. Introduction

It was demonstrated in previous parts of this Series that

A. seyal gum has a higher weight average molecular weight

than that of A. senegal, and that this is mainly due to

greater proportions of the high molecular weight com-

ponents, as identified by various fractionation methods.

Unusually, this higher molecular weight is not reflected in a

higher intrinsic viscosity for A. seyal, relative to A. senegal.

0268-005X/$ - see front matter q 2004 Elsevier Ltd. All rights reserved.

doi:10.1016/j.foodhyd.2004.09.006

* Corresponding author. Address: 2 Plymouth Drive, Radyr, Cardiff

CF15 8BL, UK.

E-mail address: [email protected] (G.O. Phillips).

This would indicate that the molecular structure is more

compact in A. seyal than in A. senegal. Hydrophobic

interactive chromatography (HIC) identified a high mol-

ecular weight protein rich component (AGP), a lower

molecular weight (AG) component with low protein, and a

third component in A. seyal of high protein content and

high molecular weight. It is probable, therefore that the

high molecular weight material identified by gel per-

meation chromatography is made up of at least two

separate components.

Our objective now is to further examine the molecular

structure of A. seyal relative to A. senegal with a view

to establishing which of the components influence

Food Hydrocolloids 19 (2005) 687–701

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C. Flindt et al. / Food Hydrocolloids 19 (2005) 687–701688

the emulsification functionality. We have also undertaken

a comparison of the two varieties of A. seyal, namely var.

seyal and var. fistula, which were shown in Part III to have

somewhat different characteristics.

2. Methods

2.1. Materials

Three samples of A. senegal, four samples of A. seyal

var. fistula and four samples of A. seyal var. seyal were used

in this investigation and were labelled as AS, ASF and ASS,

respectively. All reagents were of analytical grade. The

origin of gum arabic samples is given in Table 1. Pronase

Type XXV from Streptomyces griseus was obtained from

Sigma, Aldrich, UK.

2.2. Amino acid analysis

The amino acid analysis was carried out by Chembiotech

Laboratories, University of Birmingham, UK. The protein

was hydrolysed to its constituent amino acids using

hydrochloric acid (5.8 M), at 110 8C for 24 h. After cooling,

the hydrolysates were buffered to pH 2 prior to analysis by

cation exchange HPLC. For the cation exchange HPLC an

Aminex A8 column (300!3.5 mm ID) with 0.2 M sodium

citrate buffer as eluent was used. Elution of the amino acids

was obtained by stepwise increase in eluent pH. A post-

column ninhydrin assay detection system was used with the

reagent added at a flow rate of 0.2 ml/min and reaction time

of 3 min at 120 8C. For the quantification of hydroxyproline

a second analysis was performed.

2.3. Sugar analysis

The HPLC system used had a LC-9A Shimadzu pump

(Shimadzu, Japan) equipped with a Rheodyne valve, with a

50 ml sample loop linked with a Supelcosil NH2 column

(4.6!250 cm) (Supelco, USA) and a IOTA differential

Refractometer (Instrument S.A., France). Acetonitrile/water

Table 1

Origin of the gums and their sugar composition

Sample Origin Rha (%) Ara (%) Gal (%) GlcA (%)

AS1 Sudan 11.5 26.5 34.9 11.6

AS2 Sudana 11.6 30.4 37.3 11.3

AS3 Sudana 12.5 30.2 36.8 10.8

ASF1 Nigeria 3.5 32.9 27.6 8.6

ASF2 Chad 1.8 42.3 31.6 10.3

ASF3 Chad 1.9 43.2 33.0 11.2

ASF4 Chad 4.6 36.0 35.0 10.3

ASS1 Ethiopia 3.7 30.5 28.6 15.1

ASS2 Sudana 3.3 40.1 28.5 8.6

ASS3 Sudana 6.3 33.1 36.1 7.8

ASS4 Sudana 3.1 42.4 31.1 10.7

a Supplied by the Botanical Institute of Khartoum, Sudan, and the

remainder via Wolff and Olsen, Hamburg, Germany.

(80/20, v/v) was used as the mobile phase. Data evaluation

was performed using the peak areas and an external

standard.

The respective sample of about 0.05 g was weighed out

accurately into stoppered Pyrex tubes and 5 ml of sulphuric

acid (4%, w/w) added to each The tubes were heated for 4 h

in a water bath at 100 8C, re-weighed and made up to the

original weight by adding distilled water. The solutions

were neutralised by adding 1 g barium carbonate and left to

mix overnight at room temperature. The hydrolysates were

filtered (0.45 mm) and analysed by HPLC.

The glucuronic acid was determined using the method of

Blumenkrantz and Asboe-Hansen (1973).

2.4. GPC fractionation

The samples of about 10 g, dissolved in distilled water to

a total weight of 50 g, was hydrated for 24 h at 4 8C. The

sample solution was filtered through a blue-ribbon filter and

a 0.45 mm membrane filter. The solution was stored at 4 8C.

Fractionation was undertaken using a GPC-system with a

6-way-valve (Merck, Darmstadt, Germany) equipped with a

1 ml sample loop, with a 2.5!90 cm column packed with

Sephacryl S-500 HR (Pharmacia Biotech, Sweden and fitted

with a UV detector), UV-Monitor Model 1306 (BioRad,

USA) operated at 220 nm. The eluent in 0.5 M NaCl, was

degassed prior to use. The fractions were collected with a

RediFrac collector (Pharmacia Biotech, Sweden) with a

fraction size of 7 ml. The fractions were pooled from several

GPC runs and dialysed against distilled water and

lyophilised.

2.5. Nitrogen content

The nitrogen content was determined according to the

Kjeldahl method. The sample of about 1.5 g weighed and

25 ml of concentrated sulphuric acid added. The sample was

hydrolysed and solubilised by heating the mixture to 100 8C

until the solution was clear and greenish. The digest was

placed in the Kjeldahl distillation apparatus and the

distillate collected into 3% boric acid solution. The obtained

boric acid solution was titrated with 0.01 M chloric acid

using Tashiro as indicator. The amount of nitrogen was

calculated according to the expression

N½%� ZV !0:1401!100

1000!E

where V is the amount of 0.01 M HCl (ml) and E is the

amount of sample in grams.

2.6. Intrinsic viscosity

Solutions were prepared by dissolving 0.25 g of gum (dry

weight basis) in 5 ml of 1 M NaCl. The gum solution was

filtered through a 1.0 mm filter and 2 ml of the filtrate was

transferred into a Cannon Ubbelohde Capillary Viscometer

Table 2

Chemical properties of samples used in this investigation

Sample Minerals

(%)

Nitrogen

(%)

Optical

rotation (8)

Intrinsic

viscosity

(cm3/g)

AS1 3.4 0.38 K31 17.7

AS2 3.5 0.38 K26 16.6

AS3 3.6 0.32 K27 16.2

ASF1 3.2 0.18 29 14.7

ASF2 2.2 0.09 56 11.6

ASF3 2.7 0.08 59 14.8

ASF4 3.1 0.16 40 12.6

ASS1 3.3 0.26 47 14.8

ASS2 3.0 0.12 52 12.5

ASS3 2.6 0.20 18 12.4

ASS4 2.6 0.13 50 12.4

All results are on a dry weight basis.

C. Flindt et al. / Food Hydrocolloids 19 (2005) 687–701 689

(No. 75) in a water bath adjusted to 25G0.1 8C and the flow

time in seconds was measured three times. At least four

dilutions of the gum solution were made in situ and the flow

time was measured for each. The reduced and inherent

viscosities were plotted as a function of sample concen-

tration and the intrinsic viscosity [h] was obtained by

extrapolating to zero concentration.

2.7. Molecular weight distribution

A GPC-MALLS system was used for the determination

of the molecular weight and molecular weight distribution

as in Part I of this Series. The only difference in this part is

the processing of the data and peak definition. Here, we

used the UV profile to define the three peaks. The first peak

is for the AGP and usually elutes w11–15 min. The second

peak appears as a shoulder immediately after the AGP and

corresponds to the AG (w15–27 min). Finally, the third

peak elutes before the total volume and it corresponds to the

GP (w28–35 min). The expressions Mw and Mn are used for

the weight and number average molecular weights and

Mw/Mn for the polydispersity index (Mw/Mn). Rg is the root

mean square radius of gyration.

2.8. Enzyme degradation

The sample (0.09 mg) was dissolved in 9 ml 1 M NaCl at

pH 7.8 and protease solution

One millitre was added and the pH adjusted to 7.8. The

solutions were incubated at 37 8C in stoppered test tubes.

Aliquots (1 ml) were taken after 24, 48 and 72 h, diluted,

filtered through a 1 mm cellulose nitrate membrane filter and

analysed with the GPC-MALLS system.

2.9. Ash content

Ash content was determined by weighing the sample

(1 g) in a pre-weighed ashing dish after heating at 550 8C

for 3 h. The dish was cooled to room temperature in a

desiccator and was weighed again. The weight loss was

calculated as a percentage of the initial weight taken.

2.10. Emulsification properties

The emulsification properties were determined according

to the following method. D-limonene stock solution was

prepared by mixing 5 g of D-limonene and 2.5 g of

saccharose acetate isobutyrate (SAIB, Eastman, UK). The

respective gum samples were dissolved in 0.13% benzoic

acid solution. Gum solution of about 1.5 ml was homogen-

ised for 2 min with 0.5 ml of D-limonene stock solution in a

15 ml Pyrex tube (13 mm internal diameter) using Ultra-

Turax T25 (IKA, Stafen, Germany) equipped with S25

N-8G dispersion tool at 24,000 U/min. The emulsions

were diluted about 2000 to 3000 fold, depending on the

actual droplet size and measured with a Mastersizer 2000

(Malvern Instrument, UK), equipped with a syringe for the

injection of small sample volumes. To find the amount of

proteinaceous material adsorbed onto the oil droplets the

concentration of A. senegal, A. seyal of both variants was

determined before and after emulsification using nitrogen

content. ASF2 and ASS4 of about 0.45 g and 0.3 g of AS1

(Table 2) were used to make the emulsions. The emulsion

was centrifuged for 1 h at 13,000 rpm. The aqueous

phase was diluted to a concentration of 4!10K4 g/ml and

100 ml was injected into the GPC-MALLS for the

determination of molecular size and distribution.

3. Results and discussion

The origin and carbohydrate composition of the gum

samples used for this study are shown in Table 1. The ash

content, nitrogen and optical rotation are given in Table 2.

The results given in Tables 1 and 2 are typical of gums from

the botanical source of the samples (Jurasek, Kosik, &

Phillips, 1993; Phillips & Williams, 1993). For A. seyal

samples the optical rotation is positive, the amount of

rhamnose and the amount of nitrogen is relatively low

compared with the commercial A. senegal sample (AS1).

AS2 and AS3 have negative optical rotation and high

content of rhamnose and nitrogen. Also the arabinose and

galactose ratio is less than 1 which is typical of A. senegal

(Anderson & McDougall, 1987).

Typical GPC chromatograms obtained with a Superose

6HR column are shown in Figs. 1 and 2 for ASF2 and AS3,

respectively. The quantitative results are shown in Table 3

for the various samples. Due to the polydispersity of the

gums, and the particular column packing, the separation of

peaks 1, 2 and 3 (as defined in previous papers in this Series)

is not sharp and there is some overlap of individual

components.

The GPC-MALLS molecular weight measurements

again confirm that A. seyal has a higher weight average

molecular weight than A. senegal. Samples AS2 and AS3

Fig. 1. Chromatogram of ASF2 obtained with Superose 6 HR (&: light scattering detector at 908, C: RI detector, *: UV detector at 214 nm); the y-axis scaling

is for the light scattering detector, the chromatograms for the other two detectors have been set to 100%. The peak height, therefore, does not give the

true value.


also fit into the A. senegal norm, identified in Part I of this

Series, from the point of view of molecular dimensions,

intrinsic viscosity, optical rotation and nitrogen content

(Al-Assaf et al., Part I). AS1 has a higher molecular weight

than the norm but not unusually so. It is otherwise typical of

A. senegal. Despite their higher average molecular weight,

the A. seyal samples show a lower intrinsic viscosity

(Table 3). This further indicates that the overall dimensions

of the macromolecule are more compact for the A. seyal

than A. senegal, but with higher overall weight average

molecular weight. The average Mw values for peak 1

are 2.87, 3.92 and 4.04!106 for AS, ASS and ASF,

Fig. 2. Chromatogram of AS3 obtained with Superose 6 HR (&: light scattering de

for the light scattering detector, the chromatograms for the other two detectors hav

respectively. We find higher weight average molecular

weight for A. seyal var. fistula compared to A. seyal var.

seyal within the small selection used here (Table 3).

Apart from the molecular weight differences, the main

distinction between A. senegal and A. seyal is the protein

distribution as also identified by the differences in the RI and

UV elution profiles (Randall, Phillips, & Williams, 1988).

The majority of the protein is near the high molecular weight

fraction in A. senegal samples (Fig. 2), but is mainly

associated with peak 2 for the A. seyal samples (Fig. 1). Fig. 3

shows the direct comparison of the UV profiles in A. senegal

sample (AS3) and two A. seyal samples (ASF2 and ASS4).

tector at 908, C: RI detector, *: UV detector at 214 nm); the y-axis scaling is

e been set to 100%. The peak height, therefore, does not give the true value.

Table 3

Molecular weights of A. senegal and A. seyal var. fistula samples

Sample [h] (cm3/g) Processing Mw (!105) Mass

recovery (%)

A. senegal

AS1 17.7 Average 7.467 94.5

Peak 1 39.36 8.1

Peak 2 5.134 70.3

Peak 3 1.456 16.4

AS2 16.6 Average 5.686 95.6

Peak 1 22.13 12.3

Peak 2 3.579 72.3

Peak 3 1.085 11.6

AS3 16.2 Average 5.665 95.8

Peak 1 24.76 10.1

Peak 2 3.401 85.4

Peak 3

A.seyal var. fistula

ASF1 14.7 Average 8.207 86.3

Peak 1 35.74 6.1

Peak 2 7.100 66.0

Peak 3 1.535 14.5

ASF2 11.6 Average 10.71 93.2

Peak 1 44.19 8.9

Peak 2 7.354 81.9

Peak 3 2.194 3.2

ASF3 14.8 Average 10.36 91.3

Peak 1 38.84 9.3

Peak 2 7.480 77.2

Peak 3 1.479 5.8

ASF4 12.6 Average 9.433 89.5

Peak 1 48.13 5.6

Peak 2 7.285 77.7

Peak 3 1.575 6.4

A. seyal var. seyal

ASS1 14.8 Average 6.119 83.4

Peak 1 19.90 13.9

Peak 2 4.085 54.6

Peak 3 0.7047 14.7

ASS2 12.5 Average 9.624 96.1

Peak 1 40.74 9.0

Peak 2 6.744 81.4

Peak 3 1.665 5.8

ASS3 12.4 Average 5.186 89.3

Peak 1 53.53 2.3

Peak 2 4.608 70.7

Peak 3 0.8486 16.4

ASS4 12.4 Average 7.745 90.9

Peak 1 47.61 5.6

Peak 2 5.320 81.3

Peak 3 0.8929 4.7


The overall protein concentration is significantly less for the

A. seyal samples, which is consistent with the total nitrogen

content of the samples. There is also an obvious shift of the

main peak towards a higher retention time for the A. seyal

samples (Table 4).

The retention time of the UV peak maximum is shifted

from approximately 16.5 min for A. senegal to 19.5 min

for A. seyal var. fistula and A. seyal var. seyal. Since

the separation is according to size, it supports the view that

the structure of the high molecular weight component of

A. seyal gum is more branched than that of A. senegal gum.

This is shown in Fig. 4 where the molar mass of an A.

senegal (AS1) and two A. seyal (ASF2 and ASS4) samples

are plotted against the retention time. The line for AS1 is

especially in the high molecular weight region below the

lines for ASF2 and ASS4, showing that the Mw of AS1 is

lower at the same retention time than for ASF2 and ASS4.

3.1. Molecular size fractionation

Two well-characterised samples ASF2 and ASS4 were

chosen as representative for fractionation by GPC. These

were fractionated several times and six fractions were

pooled from the different runs as shown in Figs. 5 and 6.

The percentage of each of the fractions recovered are given

in Table 5.

The recovery in this investigation was 95% for ASF2 and

85% for ASS4, respectively. The fractionation of A. senegal

in a comparable investigation had recoveries of 94 and 68%,

respectively (Randall et al., 1988; Ray, Bird, Iacobucci, &

Clark, 1995). The quality of the fractionation was checked

by rechromatography of the fractions by GPC using

Superosew (6HR) as shown in Fig. 7 for A. seyal var.

fistula (ASF2) and Fig. 8 for A. seyal var. seyal (ASS4).

Fraction 2 of ASS4 also includes material belonging to

Fraction 3 due to the failure to get sharp fractionation with

the Sephacryl 500 column. Fraction 3 of ASS4 does not

have as broader peak as Fraction 3 of ASF2. This could also

explain the differences in the quantities recovered for

Fraction 2 of ASF2 and ASS4 (Table 5).

A. senegal was fractionated by Ray et al. (1995) on a

preparative scale using the same column material (Ray

et al., 1995) as in the present study (column size: 5!90 cm;

eluent: 0.25 M NaCl; flow: 140 ml/min). They separated

five fractions and from their elution profile, their fractions

can be related to the fractions in this study (Table 6).

Ray et al. (1995) reported that their Fractions 1 and 4

gave cloudy solutions, with particulates present, which was

also be observed with Fractions 1, 5 and 6 in the present

study. This effect could be due to agglomeration of the

molecules during the freeze drying process. For both

A. seyal varieties a similar GPC (Sephacryl 500) elution

pattern was found as for A. senegal (Ray et al., 1995). This is

in agreement with our previous observations (Part III) and

the findings of Underwood and Cheetham (1994). Using

hydrophobic interaction chromatography, they found the

same elution pattern for A. seyal as for A. senegal by

Randall, Phillips, and Williams (1989) and Osman,

Menzies, Williams, and Phillips (1994). However, the

fraction quantities differ somewhat with A. seyal (Tables 7

and 8) compared with A. senegal (Table 9). For A. senegal

one major fraction was found to contain 72% of the total

gum (Ray et al., 1995), whereas two almost equal fractions

Fig. 3. Comparison of the chromatograms obtained with GPC using UV detection at 214 nm for AS1, ASF2 and ASS4.


were found for A. seyal (containing 41 and 49% of the total

gum for ASF2 and 32 and 56% for ASS4; Table 5).

Table 4

Comparison of the elution time and molecular weight at the UV peak

maximum

Sample Elution time (min) Mw!105 (g/mol)

at UV peak maximum

A. senegal

AS1 17.0 12.8

AS2 16.1 9.20

AS3 16.4 11.0

A. seyal var. fistula

ASF1 20.0 7.12

ASF2 19.2 9.21

ASF3 19.2 8.81

ASF4 20.0 8.06

A. seyal var. seyal

ASS1 16.2 8.64

ASS2 20.0 7.42

ASS3 20.9 5.67

ASS4 21.2 5.49

3.2. Chemical characterisation

All the analytical data for the fractions are given in

Tables 7 and 8 when sufficient material was available. There

is no significant difference in the sugar composition of the

different fractions. In comparison to the amount of total

sugars, the amount of uronic acid is rather high in Fraction 5

of both samples and rather low in Fraction 6 of ASF2 and

Fraction 4 of ASS4 (Table 7). The arabinose/galactose ratio

does not differ significantly in the various fractions; only

Fraction 6 of ASF2 shows a significantly higher amount of

arabinose compared to galactose. With the exception of

Fractions 1 and 6, the fractions of the two varieties do not

differ in the amount of protein present (Table 8). Fraction 1

of ASF2 with 1.42% contains a higher amount of protein

than Fraction 1 of ASS4 (0.83%). The opposite was found

for Fraction 6 (1.14% for ASS4 and 76% for ASF2). In

comparison with A. senegal Fraction 1 of both A. seyal

varieties contains much less protein (Table 9).

Ray et al. (1995) found that Fractions 1 and 2 of

A. senegal contain 57% of the total protein, but only 23% of

the total gum. The high molecular weight material is

associated with AGP, which is of functional interest. In

comparison, for both A. seyal varieties a different distri-

bution of the protein is found. The two major Fractions 3

and 4 contribute 86.7 and 81.6 for ASF2 and ASS4,

respectively, to the overall protein content. Fractions 1 and 2

of A. senegal contain 15.4 and 6.0% protein, respectively

(Table 9), whereas Fractions 1 and 2 of ASF2 contain 1.42

and 1.28 protein, respectively, and of ASS4 0.83 and 1.43

(Table 8). These results are in accord with previous

observations that the nitrogen content of the A. seyal

samples, hence protein content, is lower than for A. senegal.

However, it is not only that A. seyal contains less protein,

but also that the distribution of the protein within

the molecular size range is different in comparison to

A. senegal.

Fraction 2 of the two varieties differ significantly in their

weight average molecular weight. ASF2 and ASS4 have Mw

of 1.01!107 and 2.93!106, respectively (Table 8). This

could be due to the imperfect separation of Fractions 2 and 3

for ASS4. Since Fraction 2 of ASS4 contains a proportion of

Fraction 3, this would decrease the overall Mw of Fraction 2.

Overall Fractions 3, 4 and 5 of ASS4 have a lower Mw at the

same Rh than of ASF2. It is possible, therefore, that the

separation of ASS4 is not only according to size, but is

influenced by other interactions. Therefore, the retention

times could decrease.

3.3. Amino acid distribution

The amino acid composition has been determined for all

fractions and the whole gum. The results are given in

Tables 10 and 11. From the amino acid composition, the

protein content of the fractions can be calculated. The amino

Fig. 4. Molar mass plotted as function of elution time combined with the GPC using refractive index detector for AS1, ASF2, ASS4.


acid composition of the whole gum is in agreement with that

previously reported for A. seyal gum (Osman, Williams,

Menzies, & Phillips, 1993). A. seyal contains less protein

than A. senegal, but the overall distribution of the amino

acids is not significantly different. It has already been

reported by Osman et al. (1993) and Biswas, Biswas, and

Phillips (1995). The amino acid composition of the two

A. seyal varieties was similar.

Fig. 6. GPC fractionation of A. seyal var seyal (ASS4) using Sephacryl 500

column.

Fig. 5. GPC fractionation of A. seyal var fistula (ASF2) using Sephacryl 500

column.

Based on the amino acid composition of the fractions of

ASF2, three groups can be identified. Fractions 1 and 6

contain a low amount of hydroxyproline and a high amount

of glycine. Fractions 2 and 5 contain a medium amount of

hydroxyproline and glycine and Fractions 3 and 4 a high

amount of hydroxyproline and a low amount of glycine. The

hydroxyproline content is of interest since the binding of

the peptide chain to the polysaccharide is considered to be

via hydroxyproline and possibly serine (Fincher, Stone, &

Clarke, 1983). Fractions 1 and 6, 2 and 5 and 3 and 4 differ

in the amounts of aspartic acid, serine and glutamatic acid,

respectively. Fractions 3 and 4 contain more histidine than

all other fractions.

Two groups can be detected for the fractions of ASS4.

Fractions 1, 5 and 6 contain a significantly lower amount

of hydroxyproline than Fractions 2, 3 and 4. The amount of

glycine is highest for Fractions 1 and 6 and lowest for

Fraction 3. Fraction 5 differs from Fractions 1 and 6 in the

amount of aspartic acid, serine, glycine and alanine.

Fractions 2, 3 and 4 vary in the amount of aspartic acid,

which is the highest for Fraction 4 and serine, which is

highest for Fraction 3.

The overall composition of the fractions is comparable

for the two varieties ASF and ASS. For Fraction 2 the

amount of glycine is higher in ASF2 and the amount of

hydroxyproline is higher in ASS4. Fraction 5 of ASF2

Table 5

Fractions after GPC separation using Sephacryl 500

ASF2 (%) recovery ASS4 (%) recovery

Fraction 1 0.1 0.2

Fraction 2 1.5 3.8

Fraction 3 41.1 31.6

Fraction 4 48.8 56.3

Fraction 5 5.4 6.5

Fraction 6 3.0 1.7

% recovery 95 85

Fig. 7. Comparison of the GPC (Superose 6HR) RI profiles of the fractions collected from A. seyal var. fistula (ASF2).

Fig. 8. Comparison of the GPC fractions collected from A. seyal var. seyal (ASS4) using a Superose (6HR) and detection by refractive index.

Table 7

Chemical analysis of the fractions (on dry basis)

Recovery

(%)

Rha

(%)

Ara

(%)

Gal

(%)

GlaA

(%)

Ara/gal Total

sugar

ASF2

F1 0.1 nd nd nd nd nd nd


F3 41.1 !1.0 42.1 31.9 9.9 1.32 85.2

F4 48.8 !1.0 40.2 34.6 10.4 1.16 86.4


the amount of hydroxyproline is significantly higher

compared with Fraction 5 of ASS4.

For the GPC fractions of A. senegal (data not shown) the

overall amino acid composition of the fractions does not

differ significantly. For all fractions higher hydroxyproline

content was found. Fraction 1 of A. senegal shows a

significantly higher content of leucine in comparison with

the A. seyal fractions. The threonine content shows

variations in the different fractions of A. senegal, whereas

it is almost constant in all A. seyal fractions. The amount of

aspartic acid is higher in all A. seyal fractions, except

Fraction 1 (Ray, Bird, Lacobucci, & Clark, 1995). Lysine is

one of the main parameters which influences most

the chemometric characterisation of gum arabic (Biswas

Table 6

Comparison of fractions obtained by Ray et al. (1995) and the present study

Ray et al. (1995) Present study

Fraction 1 Fraction 1

Fractions 2-1 and 2-2 Fractions 2 and 3

Fraction 3 Fraction 4

Fraction 4 Fractions 5 and 6

et al., 1995), but the content of this amino acid does not

differ between the various fractions of the two species.

When calculating the hydroxyproline content as percen-

tage of the total protein, no significant differences between

the two varieties were found (Tables 10 and 11). Fractions 3

F5 5.4 !1.0 20.7 18.4 6.0 1.13 46.2

F6 3.0 !1.0 30.1 10.8 3.8 2.79 47.5

ASS4



F3 1.6 1.6 44.7 36.7 11.4 1.22 95.6

F4 1.3 1.3 54.9 41.8 10.4 1.31 109.7

F5 1.0 1.0 36.4 29.2 9.8 1.25 77.7


Table 8

Analysis and molecular weight parameters of fractions (on dry basis)

Recovery

(%)

Protein (%)a Mw!105 Rg (nm)

ASF2

F1 0.1 1.42 nd nd

F2 1.5 1.28 100.6 53

F3 41.1 0.78 17.41 20

F4 48.8 0.48 6.14 13

F5 5.4 0.77 4.44 12

F6 3.0 0.76 9.12 19

ASS4

F1 0.2 0.83 nd nd

F2 3.8 1.43 29.3 32

F3 1.6 0.95 10.26 16

F4 1.3 0.56 3.67 11

F5 1.0 0.99 2.20 10

F6 1.7 1.14 3.42 30

a From amino acid analysis.

Table 10

Amino acid composition of A. seyal var. fistula (ASF2)

Whole

gum

F1 F2 F3 F4 F5 F6

Asp 79 80 106 80 159 165 115

Thr 89 62 73 66 43 62 53

Ser 197 152 156 220 145 114 112

Glu 46 116 71 36 60 70 140

Pro 87 57 76 74 58 63 55

Gly 41 130 90 25 45 74 112

Ala 35 71 58 28 34 45 65

Val 48 51 52 32 43 63 52

Met 0 6 5 0 0 1 4

Ile 14 27 25 10 16 21 27

Leu 70 57 63 65 67 69 65

Tyr 6 18 16 14 11 10 14

Phe 25 28 32 17 29 34 35

His 58 43 46 70 65 43 45

Lys 19 24 18 8 13 22 28

Arg 8 27 23 10 9 14 35

H-Pro 178 52 90 248 204 129 43

N-conver-

sion factor

6.62 6.17 6.38 6.71 6.56 6.45 6.14

Protein (%) 0.51 1.42 1.28 0.78 0.48 0.77 0.76

% of total

protein

0.3 3.0 50.1 36.6 6.5 3.6

H-Pro (%)

protein

19.0 5.7 9.7 26.2 21.5 13.8 4.6

Table 11

Amino acid composition of A. seyal var. seyal (ASS4) (mol/1000 mol)

Whole

gum

F1 F2 F3 F4 F5 F6

Asp 87 67 119 93 150 183 108

Thr 80 86 68 69 58 68 68

Ser 181 161 149 192 142 108 126

Glu 39 94 64 41 52 83 99

Pro 78 60 68 80 65 60 65

Gly 48 111 68 31 50 82 126

Ala 38 75 51 31 35 47 79

Val 54 53 52 37 56 73 58

Met 2 0 4 0 0 1 4

Ile 15 26 22 11 17 24 23

Leu 72 60 65 68 69 76 65

Tyr 9 20 19 16 12 12 7

Phe 30 30 33 20 33 41 39


and 4, which contain most of the gum have the lowest

protein contents, but also the highest hydroxyproline

contents. The hydroxyproline could be involved in the

linkage to the polysaccharide.

3.4. Enzyme degradation

Five representative samples were arbitrarily selected for

enzyme degradation with the proteolytic enzyme pronase, as

described by Connolly, Fenyo, and Vandevelde (1988) and

Osman et al. (1993). The changes in the Mw and mass

distribution is reflected by the elution profile as shown in

Table 12 and Fig. 9 (monitored by the refractive index

detector) and in Fig. 10 (monitored by the ultraviolet

detector). Generally for both A. seyal varieties peak 1 is

degraded by approximately 50–60%, but not completely as

A. senegal (AS1). For all A. seyal samples protein can be

detected in peak 1, even after 48 h of incubation with

pronase (Fig. 10). This is again a clear indication of the

denser structure of A. seyal in comparison with A. senegal.

For A. senegal, a ‘wattle blossom’ type structure has been

proposed, typical of arabinogalactan protein complexes

generally where blocks of carbohydrate are covalently

linked to a common peptide chain (Fincher et al., 1983;

Randal et al., 1989). It appears that the peptide chain of the

high Mw fraction of A. seyal is not as accessible as in

A. senegal. Since approximately 50–60% of peak 1 is

degraded of both A. seyal varieties, it is a further indication

Table 9

Analytical data for A. senegal fractions (Ray et al., 1995)

Recovery

(%)

Mw!105 Nitrogen

(%)

Protein

(%)

Rg (nm)

1 1.0 O100 2.33 15.4 nd

2-1 11.4 65 0.91 6.0 82

2-2 10.4 0.38 2.5 44

3 71.6 1.40 0.15 1.0 24

4 5.5 0.34 0.32 2.1 129

that peak 1 contains two different types of components,

which both contain a protein component. The peptide

chain is easily accessible for one component, but lays in

the interior of the molecule for other component, and is

His 56 51 45 66 56 41 45

Lys 20 22 22 11 15 24 33

Arg 9 26 19 9 10 14 14

H-Pro 181 58 134 226 181 63 41

N-conver-

sion factor

6.64 6.24 6.52 6.70 6.59 6.36 6.16

Protein (%) 0.83 1.43 0.95 0.56 0.99 1.14

% of total

protein

0.70 0.2 7.2 39.8 41.8 8.5 2.6

H-Pro (%)

of protein

19.4 6.4 14.3 23.8 18.9 6.7 4.6

Table 12

Comparison of Mw and mass distribution in enzyme degraded and whole gum

Sample Whole gum 24 h 48 h

Mw!105 % Mw!105 % Mw!105 %

AS1 Average 7.46 4.31 4.26

Peak 1 39.36 8.6 38.47 0.8 !0.2

Peak 2 5.13 74.4 4.46 72.3 4.51 72.5

Peak 3 1.45 17.4 2.89 27.1 2.98 27.6

ASF1 Average 8.20 8.02 7.34

Peak 1 35.74 7.1 35.90 3.2 33.23 2.6

Peak 2 7.1 76.5 7.43 81.5 6.88 82.3

Peak 3 1.53 16.8 5.23 15.4 5.27 15.3

ASF2 Average 10.71 9.15 7.40

Peak 1 44.19 9.5 34.79 4.6 34.06 3.5

Peak 2 7.34 87.9 8.26 80.4 6.85 79.7

Peak 3 2.19 3.4 6.11 14.9 4.36 16.8

ASS1 Average 6.12 10.51 8.11

Peak 1 19.90 16.7 31.54 8.3 28.69 7.0

Peak 2 4.08 65.5 9.71 70.1 7.74 67.1

Peak 3 0.70 17.6 5.08 21.8 3.44 24.5

ASS4 Average 7.74 9.81 7.20

Peak 1 47.61 6.2 57.59 3.4 47.15 3.4

Peak 2 5.32 89.4 8.62 79.6 6.25 80.9

Peak 3 0.89 5.2 5.76 17.1 3.47 15.9


inaccessible to the enzyme. This is typical of AGP’s

generally (Fincher et al., 1983). These observations further

confirm the fractionation studies in Part IV of this Series.

3.5. Enzyme degradation of the vulgares and gummiferae

series

There is need to consider the generality of enzyme action

in relation to structure for a wider selection of Acacia gums.

A. senegal and A. seyal are the prototype members of these

series, and show molecular weight characteristics which are

typical of the series as a whole (Part II of this Series). Our

studies have demonstrated the greater variability of the

Gummiferae series, reflected also in the A. seyal samples

we have investigated. Here we subject samples, studied

Fig. 9. Comparison of the GPC—refractive index profiles of AS1, AS

structurally in Parts II and III of this Series, to the same

enzyme degradation studies as already described to estab-

lish the generality of the behaviour, in view of the very large

variation which seems inherent in A. seyal samples collected

from various geographical regions. Table 13 gives the

changes in average molecular weight and in the high

molecular weight component (peak 1), after enzymatic

treatment for 24 and 72 h. Detailed references to their origin

and authentication can be found in Parts II and III of this

Series.

It is evident that there is a clear distinction between

A. senegal and A. seyal, and that these are typical of the

series they represent. Table 14 gives the percentage changes

of the average molecular weight and the percentage of

the high molecular weight peak 1 after enzyme treatment to

F1, ASF2, ASS1 and ASS4 after 48 h incubation with pronase.

Fig. 10. Comparison of the GPC fractionation profiles of AS1, ASF1, ASF2, ASS1 and ASS4 after 48 h incubation with pronase, monitored by UV detector.

Table 13

Effect of pronase digestion on Gummiferae and Vulgares gums

Digestion

time (h)

Mw/1 peak Mw/Mn % Mass Mw/2 peaks Mw/Mn % Mass % As of original

Av. Mw Peak 1

(A) A. senegal var. senegal. Origin: Sudan; unprocessed (Hashab)

0 4.92!105G0.25 1.74 100 1.64!106G0.08, 2.90!105G0.15 1.35, 1.29 15.2, 86.1

24 3.37G0.13!105 1.46 98 1.76G0.10!106, 2.91G0.10!105 1.20, 1.29 3, 95 68 20

72 3.13G0.14!105 1.49 100 1.89G0.12!106, 2.76G0.12!105 1.27, 1.35 2.28, 98 62 15

(B) A. seyal (Talha) (Gummiferae). Origin: Sudan

0 1.15!106G0.33 2.34 103 2.98!106G0.05, 5.43!105G0.10 1.37, 1.28 26.2, 82.3

24 1.07G0.04!106 1.74 97 2.62G0.11!106, 6.03G0.10!105 1.27, 1.18 22.5, 74.4 93 88

72 1.06G0.03!106 1.76 98 2.94G0.10!106, 6.32G0.17!105 1.27, 1.22 18.3, 80.3 92 98

(C) A. karoo (Gummiferae). Origin Zimbabwe; GA 10 as in Part II

0 2.99G0.08!106 1.84 100 4.26G0.12!106, 1.00G0.01!106 1.56, 1.03 59.9, 37.2

24 2.49G0.09!106 1.73 93 3.88G0.15!106, 1.00G0.03!106 1.39, 1.05 48.2, 45.0 83 81

72 2.36G0.05!106 1.72 96 3.71G0.08!106, 9.63G0.18!105 1.41, 1.04 48.9, 47.5 75 80

(D) A. seyal var fistula (Gummiferae). Origin Sudan; GA 20 as in Part II

0 1.34G0.03!106 2.27 104 3.57G0.08!106, 5.54G0.09!105 1.57, 1.19 27.1, 76.2

24 1.12G0.03!106 2.04 92 3.20G0.09!106, 5.74G0.16!105 1.33, 1.26 19.3, 73.2 80 70

72 1.09G0.04!106 2.09 100 3.25G0.14!106, 5.70G0.21!105 1.32, 1.28 19.6, 80.0 80 71

(E) A. seyal var seyal (Gummiferae). Origin Mali; GA 28 as in Part II

0 1.11G0.02!106 2.24 106 3.19G0.07!106, 4.76G0.05!105 1.66, 1.26 21.0, 85.0

24 8.79G0.38!105 1.92 92 2.60G0.10!106, 4.89G0.19!105 1.40, 1.25 17.0, 75.2 80 80

72 8.94G0.34!105 2.04 98 2.93G0.13!106, 4.93G0.19!105 1.47, 1.29 16.0, 81.7 80 90

(F) A. seyal var. seyal (Gummiferae). Origin Chad; GA 35 as in Part II

0 1.06G0.09!106 1.78 103 2.55G0.05!106, 5.60G0.07!105 1.34, 1.16 26.0, 76.9

24 9.82G0.39!105 1.67 96 2.44G0.09!106, 5.92G0.23!105 1.23, 1.19 20.2, 76.3 93 80

72 9.58G0.37!105 1.68 100 2.58G0.10!106, 6.04G0.22!105 1.21, 1.22 18.2, 83.0 90 70

(G) A. seyal var. seyal (Gummiferae). Origin: Mali; GA 80 as in Part II

0 1.04G0.01!106 2.08 106 2.78G0.05!106, 5.42G0.16!105 1.38, 1.28 23.9, 81.5

24 9.34G0.31!105 1.88 94.5 2.53G0.09!106, 5.42G0.16!105 1.26, 1.29 18.4, 75.5 90 80

72 8.89G0.21!105 1.84 97.9 2.55G0.08!106, 5.56G0.13!105 1.23, 1.33 16.5, 81.7 85 70

(H) A. laeta (Vulgares). Origin: Chad, GA 39 as in Part II

0 6.38G0.15!105 2.05 102 2.71G0.08!106, 3.43G0.05!105 1.54, 1.24 12.7, 89.4

24 3.92G0.13!105 1.48 96.6 1.76G0.06!106, 3.25G0.11!105 1.21, 1.27 4.5, 92.2 62 36

72 3.60G0.16!105 1.47 100 1.74G0.10!106, 3.16G0.13!105 1.18, 1.32 3.2, 96.6 59 29

Weight average molecular weight (Mw) for the whole gum processed as 1 peak is given in column 2. % mass is the recovered mass. Mw/Mn is the polydispersity

index. Mw processed as two peaks means that the high molecular weight peak was processed separately and the remainder of the gum processed as the

second peak.


Table 14

Enzymic degradation of Gummiferae and Vulgares gums (data from Table 13)

% initial average Mw % initial, % peak 1

Gummiferae

(B) A. seyal 93 88

(C) A. karoo 84 80

(D) A. seyal

var. fistula

80 70

(E) A. seyal 85 80

(F) A. seyal 92 75

(G) A. seyal 91 75

Vulgares

(A) A. senegal 60 18

(H) A. laeta 60 33


both Gummiferae and Vulgares Gums. Generally, the

enzymic hydrolysis is completed after 24 h.

These enzyme experiments again confirm our previous

conclusion that the high molecular weight component near

peak 1 for A. seyal is made up of two components, one of

which can be enzymatically degraded, and the other which

is resistant. This component can probably be identified

with the high molecular weight component (Fraction 3) in

the hydrophobic fractionation studies in Part IV of this

Series.

3.6. Differences in adsorption onto oil droplets

The adsorption properties of A. senegal have been

studied by Snowden, Phillips, and Williams (1987a,b) and

Randall et al. (1988). It was found that mainly the

proteinaceous high molecular weight component of the

gum adsorbs onto model polystyrene latex and orange oil,

respectively. The adsorption onto latex is completed after

4 days and 100% of the proteinaceous material is adsorbed

(Snowden et al., 1987a,b), whereas only 25% of the

proteinaceous material is adsorbed to orange oil droplets

(Randall et al., 1988).

Fig. 11. Comparison of GPC profiles monitored by refractive

The emulsification properties of AS1 and ASS4 were

investigated. The emulsions prepared as already described,

were centrifuged and the water-phase analysed using GPC-

MALLS.

For all samples only small amounts of gum were

adsorbed onto the oil droplets as can be shown by the

comparison of the GPC/RI profiles (Fig. 11). This can

be calculated from the total amount of sample detected by

the chromatographic system recovery. It is difficult to

compare the percentage adsorbed because it depends on the

initial gum concentration. What we should be comparing is

the amount adsorbed (mg/m2). A surface coverage with of

4 mg/m2 can be calculated for the A. senegal sample (AS1),

which is in good agreement with the value of also 4 mg/m2

calculated by Randall et al. (1988) and 5 mg/m2 latex

surface determined by Snowden et al. (1987a,b). For the A.

seyal sample (ASS4), a value of each 5 mg/m2 was

determined, revealing no significant difference of the

surface coverage in comparison with AS1.

From the calculated weight percentage distribution of the

peaks, it is not possible to determine which of the fractions

has been preferably adsorbed onto oil droplets, since those

distributions show only slight differences in comparison

with the data for the whole gum (Fig. 11). However on

comparing the UV profiles of the samples before and after

emulsifying, the differences become clear. For AS1 it is

mainly the proteinaceous material of peaks 1 (11–15 min)

and 2 (15–26 min) which is adsorbed onto the oil droplet

(Fig. 12), which is in agreement with the findings of Randall

et al. (1988). On the other hand, ASS4 shows a different

behaviour and it is mainly the proteinaceous material

belonging to peak 2 (15–26 min) and 3 (26–35 min) which

is adsorbed (Fig. 13).

There are, therefore, again distinct differences between

A. senegal and A. seyal. Whereas, the AGP (peak 1), which

is readily all degraded by pronase is adsorbed onto the oil

droplets, it is protein of different molecular weight which

adsorbs for A. seyal.

index changes for ASF2 before and after emulsification.

Fig. 12. Comparison of GPC profiles monitored by UV detection for AS1 before and after emulsification.


3.7. Structural differences between A. senegal and A. seyal

Throughout our investigations, we are mindful that we

are dealing with a natural product of wide geographical

origin. Differences between and within sample groupings

might therefore, be expected and are observed. Here,

however, we point to gross differences which distinguish

between A. senegal and A. seyal, indicating also comparable

differences between the taxonomic series of which these

widely used food additives are prototypes.

1.
The A. seyal series, as a broad grouping, have an average
molecular weight 2–4 times greater than A. senegal. This

is due to a greater proportion of the high molecular

weight component.

2.
The sugar composition and amino acids making up the
protein moiety in each of the gums are identical, but are

present in different proportions This is the main reason

why A. seyal is dextrarotary and A. senegal is laevorotary

(Biswas et al., 1995).

Fig. 13. Comparison of GPC/UV profiles of

3.

ASS

Although the molecular weight is considerably greater,

the intrinsic viscosity of A. seyal is considerably less

than A. senegal. The structure is, therefore, even more

compact than for A. senegal.

4.
Enzyme hydrolysis also points to two components being
associated with the high molecular weight material in A.

seyal, one only which is degraded by pronase. In

hydrophobic fractionation studies, a protein rich com-

ponent of extremely high molecular weight (Fraction 3)

was found in A. seyal but not in A. senegal. In GPC

fractionation this would co-elute with the main com-

ponent, which is further evidence for the presence of the

two different high molecular weight components in A.

seyal, unlike A. senegal.

5.
These high molecular weight fractions adsorb onto oil
droplets during emulsification experiments. The mol-

ecular weight of the AGP, established as the active

moiety for A. senegal is greater than the fraction which

binds to the oil in A. seyal, and in itself is not a highly

efficient emulsifying component.

4 before and after emulsification.


6.
There are small indications of differences between
varieties, but overall these cannot be regarded as

significant. Essentially, the greatest differences is in

molecular weight. A. senegal var. karensis has a higher

weight average molecular weight than A. senegal var.

senegal due to a greater proportion of Peak 1 (as

described in Part 1 of this Series). Similarly, A. seyal var.

fistula has a higher molecular weight than A. seyal var.

seyal for the same reason.

Fig. 14. Type 1 repeating unit. GZD-galactopyranose, R1Zarabinose
7.
chain of varying length.

The differences identified here for A. seyal and A. senegal

represent also the same differences which extend to their

taxonomic species Gummiferae and Vulgares.

8.

Fig. 15. Type 2 repeating unit. R2 may be H, D-glucuronic acid; D-4-O-

methylglucuronic acid, or a-(1/4)-L-rhamnopyranosyl-glucuronic acid;

glycosidic bonds:—Zb-1,3;/Zb-1,6.

Structural studies carried out some time ago support

these observations, allowing us now to visualise the

gums structures of the two gums. Emphasis should be

made of the diversity which seems to be associated with

A. seyal. There does not seem to be a typical A. seyal.

The diversity is evident from the plot of log intrinsic

viscosity against log Mw. Whereas, it is linear for

A. senegal. The plot is widely scattered for A. seyal and a

linear plot cannot be achieved. The finding is in keeping

with the chemometric observation, with A. seyal of the

Gummiferae series forming much broader clusters than

A. senegal and the Vulgares series.

During Smith degradation, the enrichment of pepti-

des/amino acids within the interior of the gum

macromolecules is considerably greater than the corre-

sponding enrichments reported for the gum from

A. senegal (Anderson & McDougall, 1987; Anderson

& Yin, 1987). The molar ratio from polysaccharide to

protein is 113:1, 58:1, 27:1, 5,5:1 and 4:1 for the whole

gum and the sequential Smith degradations SD-I, SD-II,

SD-III and SD-IV products, respectively, for A. seyal

compared to 31:1, 18:1, 11:1, 11:1, 11:1 for A. senegal.

There is, therefore, evidence that only few of the amino

acids in A. seyal gum are attached to periodate-

vulnerable sugars in peripheral positions and that the

majority of the amino acids are located at deep-seated

locations within the complex gum macromolecules

(Anderson & Yim, 1987).

The early work of Anderson’s group (Anderson, Dea,

& Hirst, 1968) could not distinguish any different

fundamental linkages between A. seyal and A. senegal.

However, they noted that long chains of periodate-resistant

b 1–3 linked galactose residues are not such a dominant

structural feature of A. seyal as for A. senegal. It

was suggested that A. seyal has a galactan framework

which is more highly branched than A. senegal. In these

studies also, Anderson noted a considerable variation in

composition between different nodules of well-authenti-

cated A. seyal.

Street and Anderson (1983), in a reinterpreation of the

work of Churms, Stephen, and Siddiqui (1981) suggested

different structures of the polysaccharide for A. senegal and

A. seyal gum. A. senegal is suggested to contain entirely of

Type 1 repeat units (Fig. 14), whereas A. seyal consists of

small blocks of two or three modified Type 1 repeat units

separated by significant blocks of Type 2 repeat units

(Fig. 15). A. seyal is proposed, therefore, to be more highly

branched than A. senegal.

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