Xanthan gum–gelatin complexes

C.-y. Lii a,*, S.C. Liaw a, V.M.-F. Lai b, P. Tomasik a,c

a Institute of Chemistry, Academia Sinica, Nankang, 11529 Taipei, Taiwan, ROCb Department of Food and Nutrition, Providence University, Shalu, Taichung, Taiwan, ROC

c Department of Chemistry, University of Agriculture, Mickiewicz Avenue, 21, 31120 Cracow, Poland

Received 30 May 2001; received in revised form 25 October 2001; accepted 15 November 2001

Abstract

Gelatin (G) and xanthan gum (X) formed complexes either on bringing their blends to pH 2.3 or carrying out the

electrode process in aqueous blends of X and G at pHs from 9 to 11. X carboxylic groups and G peptide moieties were

involved in the interactions between the partners. Also non-coulombic interactions with involvement of NH and OH

groups, as well as hydrophobic interactions were involved, as proved by FTIR and thermal analyses, respectively. The

reaction yield declined with increase of concentration of G in the blends. Simultaneously, the thermal stability of the

complexes slightly increased with increase in G content in the blend. The latter increased with its concentration in

the reaction mixture. Slow formation by electrosynthesis provided more an organised matrix. � 2002 Elsevier Science

Ltd. All rights reserved.

Keywords: Coacervation; Electrosynthesis; Polysaccharide–protein complexes; Plant gums

1. Introduction

For over two decades polysaccharide–protein com-

plexes have received considerable attention for both

theoretical and practical reasons [1–5]. Gelatin (G) is

often used as protein component for complexes. It

can complex to carboxymethyl cellulose (CMC) with

involvement of electrostatic interactions [6]. Non-

coulombic interactions were essential in the formation

of complexes of G with pectins and alginates [7]. The

character of the interactions was dependent on the re-

action conditions, mainly on pH and ionic strength of

the reaction mixture, as proved in complexes of G with

CMC [8] and with gum Arabic [9].

Complexation of G with CMC was utilised in puri-

fication of these molecules [6] and complex formation

with acacia gum was patented as a fat replacer in

foodstuffs [10]. Complexes of G with acacia gum [11–16],

pectin [17], gellan gum [18], alginate [19], and CMC [19]

were considered as prospective sources for microcap-

sules.

Xanthan gum (X) as a polysaccharide component of

complexes with proteins was also subjected to numerous

studies. In a complex with milk protein prepared in the

pH range between 3.7 and 6.3, the components bound

via hydrophobic interactions [20]. Complexes of X with

milk proteins appeared to be interesting as potential

shortening agents for bakery, a fat replacer [21], and an

emulsifier for food industry [22,23]. Complexes of X

with soy protein [24,25], casein [24], ovalbumin [20,24],

and whey protein [26] may be used as meat and fat re-

placers. Purification of whey proteins by complexation

to X could be achieved [27].

In this paper, the synthesis and properties of the

complexes formed from G and X are described. Simul-

taneously, the electrode process formerly applied for the

synthesis of polysaccharide–protein complexes [28–32]

with covalent bonds between the partners is shown to be

useful for slow coacervation, providing a more organ-

ised matrix of the resulting complexes.

Although research on protein–polysaccharide com-

plexes has been addressed mainly in food science and

European Polymer Journal 38 (2002) 1377–1381

www.elsevier.com/locate/europolj

* Corresponding author. Tel./fax: +886-2-2783-1237.

E-mail address: cylii@chem.sinica.edu.tw (C.-y. Lii).

0014-3057/02/$ - see front matter � 2002 Elsevier Science Ltd. All rights reserved.

PII: S0014-3057 (02 )00008-3

technology, the results of such studies might be poten-

tially utilised for non-nutritional purposes. Such mate-

rials could serve as sizes for textiles, fillers and thickeners

for pulp and paper, coatings, microcapsules, and bio-

degradable constructing materials.

2. Results and discussion

Electrosynthesis provided materials, which were 15–

7% richer in G than the blends from which they were

prepared. However, blends richer in G gave products

with a lower excess of G. Generally, the yield of the

products decreased with an increase in pH. High ratios

of G in initial blends resulted in a decreased yield. The

highest yield of the products separating on the anode

was achieved from X:G ¼ 1:2 blends (Table 1).

The current applied (Table 1) was automatically in-

creased with pH. In the blends of 1:1 and 1:2 X:G ratios,

the current rose with time. This might be an effect of a

decrease in solution viscosity caused by the separation of

the product. In the blends containing high ratio G, the

current either remained unchanged, or decreased with

time. The latter could result from the clathration of ions

and/or water in these G-rich products.

The solubility tests for X, G, and the products pre-

pared either by simple coacervation or electrosynthesis

(Table 2), particularly solubility in 7 M aq. urea, re-

vealed that coulombic interactions and formation of

hydrogen bonds between X and G were involved. Thus,

electrosynthesis did not promote formation of covalent

bonds between those partners and the process taking

place at the anode and in its vicinity was also a coac-

ervation process.

Coacervation of X with G took place at a pH of 2.3.

Differential FTIR spectra of 1:1 X–G complex (Figs. 1

and 2) and with the subtracted spectrum of X taken at

pH 2.3 (Fig. 2) showed that although the spectral pat-

terns resembled a simple combination of the spectra of

G and X, the essential band shifts could be observed.

Contrary to X CMC formed complexes with G in

which both partners were covalently bound [33]. X is a

polysaccharide, the anionic property of which results

from the presence of uronic acid moieties. The carb-

oxylic groups in X are directly bound to the pyranose

ring, and anionic properties of CMC result from the

presence of the carboxylic groups bound to the glucose

units through the 6CH2–O–CH2 linkage. The differences

in the bonding of the carboxylic groups in X and CMC

should not be reflected by any essential differences in

their ionisation. Therefore, availability of the carboxylic

Table 1

Composition and yields of X–G complexes

X:G ratio Intended G, % pH Formed Yielda, % Current change, A

Nitrogen content, % Gb, %

1:1 50 9 10.1 58.2 75 0:01! 0:03

10 10.0 57.8 73 0:01! 0:03

11 9.9 57.5 68 0:04! 0:051:2 67 9 12.6 72.6 77 0:01! 0:02

10 13.1 75.5 76 0:01! 0:02

11 12.8 73.9 73 0:04! 0:03

1:3 75 9 14.2 82.1 67 0:01! 0:0110 13.9 80.1 65 0:02! 0:02

11 13.1 75.9 55 0:05! 0:03

1:4 80 9 15.1 87.2 62 0:01! 0:0110 14.6 84.3 58 0:02! 0:01

11 14.7 85.0 54 0:05! 0:02

a The yields were calculated on the basis of the total parent polymer weight with standard deviations of 0.3–3.2%.bThe data were calculated from the nitrogen content in complexes and original G sample (17.29%).

Table 2

Solubility tests of G, X, and X–G complexes

Sample Solvent

H2O 5% Na2CO3 1% NaOH 7 M Urea DMSO 5% HCl

G–X complexesa – – þ þ – –

G – – þ þ þ þX þ – þ þ – þaAll complexes regardless the preparation method, initial composition of the blend and its pH.

1378 C.-y. Lii et al. / European Polymer Journal 38 (2002) 1377–1381

group for the complexing partner, which in X was much

more obstructed by steric hindrances seemed to be cru-

cial for the mode in which partners interact with one

another.

Electrosynthesis of G–X complexes proceeded in the

reaction mixtures of pH 9, 10 or 11 but the local pH

around anode could be different. Indeed, the FTIR

spectra of the 1:1 G–X complexes from both processes

were identical. Table 3 lists the wavelengths of the

spectral bands and their assignments.

The band shifts confirmed the involvement of the

carboxylic groups of X and peptide bond moieties of G

in the G–X interactions. Simultaneously, the shifts of the

vibration bands of the OH and NH groups showed that

non-electrostatic interactions might participate in coac-

ervate formation. The pH at which the syntheses were

carried out had no effect on the FTIR spectra, and

varying proportions of X and G in the initial blends

were reflected exclusively by relative peak intensities.

X was less thermally stable than G as shown by

thermogravimetric (TG) and differential thermogravi-

metric (DTG) analyses (Table 4).

Thermal stability of the products from electrosyn-

thesis slightly increased with an increase of G in prod-

uct. The effect of pH upon the thermal stability of

resulting products could be interpreted in terms of the

effect of pH upon the content of G in the product. At

higher pH, G content in the products decreased (Table

1) and, therefore, the latter slightly less thermally stable.

The processes of thermal decomposition for 1:1

products prepared by coacervation alone and by elec-

trosynthesis were practically identical (Fig. 3).

Slightly higher water content in the electrosynthes-

ised product might be a result of the slower and, per-

haps, more ordered building up of the matrix. The result

of the DTG analysis (Table 4) indicated that both

preparation methods produced products of different

macrostructure. Water in the product formed by coac-

ervation was held more strongly than in the product

prepared by electrosynthesis. It is suggested that the

latter procedure provided a more compact and, possibly,

a better organised matrix.

TG and DTG analyses showed that the X–G com-

plexes decomposed in three steps. The third step took

Fig. 1. FTIR differential spectrum of the 1:1 G–X product from

simple coacervation (below). The spectrum of X taken at pH

2.3 was subtracted from the spectrum of coacervate and spec-

trum of G taken at pH 2.3 was added for comparison (above).

Fig. 2. FTIR differential spectrum of the 1:1 G–X product from

simple coacervation (below). The spectrum of G taken at pH

2.3 was subtracted from the spectrum of complex and spectrum

of X taken at pH 2.3 was added for comparison (above).

Table 3

FTIR spectral patterns of G (at pH 2.3), X (at pH 2.3) and 1:1

G–X complex and the band assignments

m ðcm�1Þ Band assignment

1:1 G–X

complex

G X

3416 s 3416 s mOH intramolecularhydrogen bond

3368 s mNH intramolecularhydrogen bond

3081 sh 3081 w mOH polymeric association2928 w 2947 w 2918 w mC–H1719 m 1733 m mCOOH1652 s 1652 s 1642 m mC@O, dOH

1538 w mCOO�

1447 w 1447 w dCH1404 w 1404 w dCH

1380 m mCOO�

1333 w 1333 w dNH1252 m dCH

1242 m 1238 w dNH,mC�N1152 w 1152 m mC@O glucose units

1080 w dCOH1066 s 1061 s mC–O, mC–C, dCOH, C1–H1033 s 1028 s mC–O, mC–C, dOH, C4–O

C.-y. Lii et al. / European Polymer Journal 38 (2002) 1377–1381 1379

place at temperature above the decomposition point of

X and G and this thermal stabilisation might be ac-

counted for as a result of hydrophobic interactions.

Coacervation of X with G from solutions of X and G

suggested that such a process might be useful in recovery

of X from aqueous solution. Moreover, the increased

stability of the complexes under acidic conditions could

be of practical significance in formation of microcap-

sules, and in application as sizes and biodegradable

materials.

3. Experimental procedures

3.1. Materials

X (G1253) and G (type A from porcine skin, G2500)

were purchased from Sigma Chemical Co., St. Louis,

MO, USA.

3.2. Electrosynthesis

A 400-cm3 beaker equipped with two Pt electrodes

situated at a distance of 2.5 cm from one another was

used as an electrolytic cell. It was connected to a power

supply (model P-3003D, Taiwan). The beaker was filled

with 250 ml of aqueous solution containing 0.125 g of X

and 0.125, 0.250, 0.375 g of G. These concentrations

corresponded to X:G ratios of 1:1, 1:2, 1:3, 1:4 (w/w),

respectively. The polymer solution was completely dis-

solved in distilled water under conditions of mild heating

(�40 �C) and stirring, then cooled to room temperature.

The pH was adjusted to 9, 10 and 11 with 0.01 M

NaOH.

The electrosynthesis was conducted at a constant

potential difference of 12 V for 1.5 h. A layer of white,

gummy product was collected from the surface of anode,

centrifuged to remove liquid, and dried in vacuum at

room temperature.

3.3. Sample coacervation

Aqueous blends of X and G as described in Section

3.2 were adjusted to a pH 2.3 with 0.1 M hydrochloric

acid whilst being agitated. The resulting precipitates

were centrifuged to remove liquid, and dried in vacuum

at room temperature.

3.4. Combustion analysis

Combustion analysis was carried out with a Perkin

Elmer 2400 CHN elemental analyzer (Norwalk, CT,

USA). Analyses were duplicated. They varied by up to

0.25% of recorded values.

3.5. Solubility tests

The solubility of starting materials and the products

in water, 5% hydrochloric acid, 5% Na2CO3, 0.1–1 M

NaOH, DMSO, and 7 M aq. urea were examined at

25 �C.

Fig. 3. The TG curve of the 1:1 G–X product obtained by

simple coacervation (––) and by electrosynthesis (– – –).

Table 4

TG and DTG analysis of G, X as well as 1:1 X–G product

obtained by simple coacervation and by electrosynthesis

Sample Analysis TG DTGa

(�C)Temperature

of the effect

(�C)

Associated

weight lossb

(%)

G 156.4 10.2 75.7

227.3 11.4 307.7c

227.4 52.8

X 144.2 18.0 56.5

206.7 19.3 278.9c

324.5 56.7

1:1 G–X from

simple coacerva-

tion

124.3 9.2 66.7

180.0 10.2

240.0 17.5 234.9

270.0 27.5 293.8c

370.0 60.0 337.1

1:1 G–X from

electrosynthesis

124.6 10.1 60.9

180.0 11.2

240.0 18.0 237.5

270.0 28.0 296.4c

370.0 60.0 333.7

aAll effects were endothermic.b The weight loss from the origin.c The dominating effect.

1380 C.-y. Lii et al. / European Polymer Journal 38 (2002) 1377–1381

3.6. FTIR analysis

The infrared spectra of X, G, and their complexes

were run in KBr discs using a Perkin Elmer FTIR

spectrometer Paragon 1000 (Norwalk, CT, USA), in the

frequency range of 4000–500 cm�1. Differential spectra

of X–G complexes and pure X or G were also recorded.

3.7. Thermal analysis

The thermal characteristics of pure X, G, and X–G

complexes were determined under a nitrogen stream

using a Du Pont TGA 951 system (Wilmington, DE,

USA) scanned from 25 to 500 �C at 10 �C/min.

4. Conclusions

Regardless of preparation methods, X and G formed

complexes with the involvement of weak electrostatic

and non-electrostatic interactions between X and G. The

matrix of the product formed from electrosynthesis

could be more regularly in structure. The role of the

electrode process carried the formation around the an-

ode of a concentration of protons suitable for coacer-

vation of G and X.

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