Product Profile 1

Xanthan gum: Production,properties and applications

Dr. Anil Lachke & Dr. P.K. IngleNational Chemical Laboratory

Pune 411008

WE are quite familier with gum sinceour childhood. We use gum Arabic forsealing envelopes and for stickingpostal stamps. The better consistencyto ice-cream is often provided by gela-tin or dextran. The delicious jellies andjams can be moulded into beautifulshapes using China grass (sea weed,agar). Gum tragacanth gives desir-able flow property to the tooth-paste.Natural gums occur in all life forms.l\/lany types of gums can be obtainedfrom plant sources. Their collection iscostly and requires skilled labourers.Moreover, seasonal variations affectthe quality and quantity of plant gums.The polysaccharides for scientific andindustrial applications are obtainedmore conveniently from microbialsources due to several factors. Theycan be produced undercontrolted con-ditions from selected species usingrenewable sources and arebiocompatible and biodegradable.These factors have accelerated theuse of microbial gums such as pullulan,curdlan, scleroglucan, dextran andxanthan. Chemically gums are carbo-hydrate polymers or polysaccharides.(However, gelatin is a protein).Polysaccharides are present every-where. They have a number of uniquechemical and physical properties. Theyserve as structural material to the plant

kingdom, as energy reserves, adhe-sives and also information-transferagents. Microbial polysaccharides arecomposed of regular repeating unitsof simple sugars like glucose, man-nose, fructose etc. These polysaccha-rides are sometimes termed as slimeor exopolysaccharides.

Why do microorganismsproduce gums?

Most phytopathogenic bacteria donot form spores. Many of them areresistant to desiccation and surviveunder dry conditions for more than 50years at normal surrounding tempera-ture. This is due to the protective layerof the "ooze" or exudates produced bythe bacteria. The layer is nothing but acoating of specific gum that is chemi-cally a polysaccharide. This coatingmay act as a barrier against attackfrom bacteriophage, and also helpsidentification of appropriate sites onthe host plant for colonization of thebacteria. Xanthomonas campesthssynthesizes the xanthan gum.Xanthomonas is a genus of thePseudomonaceae family. All organ-isms in this genus are plant patho-gens. It is a Gram negative, yellow-pigmented bacterium. The structureof the ceJI envelope is similar to that of

the other Gram-negative cells. Thecapsular polysaccharide is the xanthangum. Staining with India ink showsthat many isplates of XanthomonasSp. have capsules with the capsularpolysaccharides often quite looselyassociated with the cells.Xanthomonas campestris is the mostcommonly employed microorganismfor industrial production of xanthan.Xanthomonas cells occur as singlestraight rods, 0.4-0.7 ^m wide and0.7-1.8 i m long. The microorganismis chemiorganotropic and an obligateaerobe with a strictly respiratory typeof metabolism that requires oxygen asthe terminal electron acceptor. Thebacterium cannot denitrify, and it iscatalase-positive and oxidase-nega-tive. The colonies are usually yellow,smooth, and viscid. Xanthomonas Sp.are able to oxidize glucose and theEntener-Doudoroff pathway is pre-dominantly used for glucose catabo-lism. The pentose phosphate pathwayalso occurs but uses only 8-16% of thetotal glucose consumed. Both the tri-carboxylic and gtyoxylate cycles arepresent. Several species ofXanthomonas pathogenize specificplant hosts. For example, cabbage isattacked by X. campestris ,sugar caneby X. vasculorum, Strawberry by X.fragaria and walnut by X. juglandis.

CHEMICAL BUSINESS -•• JULY 2006 43

Xanthan gum is a microbial polysac-charide of great commercial signifi-cance, It is a cream coloured powderthat is soluble in hot or cold water witha high viscosity even at low concen-trations. The molecular weight ofxanthan is determined by light scatter-ing, quasi-elastic light scattering andband sedimentation analysis. Thesemethods however revealed a widevariation between 2 million to 62 mil-lion in the molecular weight of xanthan.The explanations for these variationsin the reported valueswere provided by thequasi-elastic light scatter-ing techniques. Hydrogenbonding appears to beimportant in stabilizing theaggregates of xanthan inwater. In 4 molar urea so-lution, a lower molecularweight of 2 million wasobtained.

H.C C

Backbone ofXanthan

Xanthan gum is a"heteropolysaccharide. The main chainof xanlhan is built up of D-glucoseunits linked through the fi-i position ofone unit with 4th position of the nextunit, a linear backbone identical to thechemical structure of cellulose. Theprimary structure of xanthan consistsof a pentasaccharide repeating units(Figure 1). The presently acceptedstructure of xanthan consists of ( 1 ^4)-(5-D-glucopyranose units. Trisac-charide side-chains are attached toalternate sugar residues on the mainchain at the C-3 position. The side-chain consists of two mannose resi-dues and a glucuronic acid residue.The terminal p-D-mannopyranose resi-due is (1 ^ 4) linked to the ^-D-giucu-ronic acid residue, that in turn is (1->2) linked to non-terminal a-D-mannopyranose residue. The 6-OHgroup of the non-terminal D-

mannopyranose residue is present asacetic acid ester. Pyruvate acetalgroups are located on the D-mannopyranosyl end groups of side-chains. The repeated pentasaccharideunits formed by two glucose units, twomannose units, and one glucuronicacid unit, in the molar ratio 2.8:2.0:2.0.The influence of different glycosidic(or other) linkages in the backbone ofany polysaccharide is an importantfeature in modifying polysaccharidechain conformation and its character-

by phosphorylation of the substratewith a hexoklnase enzyme that uti-lizes adenosine 5"-trlphosphate. Thebiosynthesis involves conversion ofthe phosphorylated substrate to thevarious sugar nucleotides required forassembly of the polysaccharide-re-peating unit via enzyme such as UDP-Glc pyrophosphoryiase. UDP-glucose,GDP-mannose and UDP-glucuronicacid are necessary for the synthesis ofxanthan with the appropriate repeat-ing unit.

OH OH CHjOH

H H

In the biosynthesis ofxanthan on the cabbageplant by X. campestris thecabbage provides the car-bohydrate substrates, pro-teins, minerals for cellgrowth. In the laboratoryconditions or commercialfermentation, carbonsources, nitrogen sources,trace minerals and pH con-ditions are provided in away that simulates naturalconditions.

istics. It is not surprising to thatxanthans of different pyruvate levels(that is 1 to 6 %) display differentrheological (flow) properties. Pyruvicacid attached to the terminal carbohy-drate of the side chains adds anothercarboxylate group. The percent com-position of xanthan proposed for in-dustrial use is as follows: Glucose 37,mannose 43.4, glucuronic acid 19.5,acetate 4.5 and pyruvate 4.4%.

Production of Xanthan

The biosynthesis of microbialheteropolysaccharides such asxanthan is a complicated process, in-volving a multienzyme system. Theinitial step in the biosynthesis ofxanthan is the uptake of carbohydrate,which may occur by active transport orfacilitated diffusion. This is followed

Industrial production

The selected microbial strain is pre-served for possible long-term storageby proven methods to maintain thedesired properties. A small amount ofthe preserved culture is expanded bygrowth on solid surfaces or in liquidmedia to obtain the inoculum for largebioreactors. The industrial scale pro-duction of xanthan \e carried out usinginexpensive substrates and nutrients.Carbohydrate sources such as su-crose, sugarcane molasses, and wheyhave been successfully used in theproduction medium. The whey alsoprovides adequate nitrogen and somegrowth factors. Efficient conversion ofcarbon source to desired polysaccha-ride production requires a high carbonto nitrogen ratio. Inorganic nitrogensources like ammonium or nitrate salts

44 CHEMICAL BUSINESS -f JULY 2006

are suitable, and a wide variety ofcomplex nitrogen sources like yeastextract, soy-meal peptone soybeanwhey are also useful for xanthan pro-duction. The nutritional requirementsof X. campestris necessitate the pres-ence of phosphorus and this is usuallyadded in the form of a phosphatebuffer. Beneficial production isachieved using cereal grains. Gener-ally batch cultivation with complexmedia is favoured. However, simplesynthetic media can be used. Thebatch growth cultivation requires twodays. Although most often a stirred-tank reactor is used with batch fer-mentation operation, there is no uni-formity with regard to the operationalconditions employed such as tempera-ture, oxygen mass transfer, pH, andthe composition of the culture me-dium. The production is generally car-ried out using a temperature range of25to 35" C. Several investigators havestudied the influence of temperatureon growth in the temperature range of22-35^; 28°C is the optimal growthtemperature in the media used. Ahigher media temperature increasesxanthan production but lowers its pyru-vate content (Cadmus et al.,Biotechnol. Bioeng. 20:1003-1014,1978). Oxygen mass transfer is influ-enced by both agitation speed andairflow rate.

Proper maintenance of X.Campestris stock culture is importantfor consistency in the production ofxanthan, as variation is a recognizedcharacteristic in Xanthomonas spe-cies. During the initial growth phasepolysaccharide accumulation startsand continues after growth. NeutralpH is the optimum value for growth ofX. campestris. The pH decreases dur-ing the fermentation due to the forma-tion of organic acids. If pH falls below6.0, the formation of xanthan drasti-cally reduces. Thus, it is necessary tocontrol the fermentation medium at

the optimum pH of 7.0 using a bufferor addition of base during process.Adequately designed agitation is nec-essary to disperse the introduced airevenly throughout the medium. Agita-tion of the medium is useful for en-hancing the rate of transport of nutri-ents across the cell membrane, whichin turn supports the growth rate of themicroorganism.

Isolation of the gum

The fermentation broth of thexanthan gum is apparently very vis-cous. The crude broth contains bacte-rial cells, and after solidifying the gum,their weight will be about 15 g per 100g of xanthan gum. The weight of thecells is 1-10 g/L. Recovery of thexanthan from the fermentation broth isgenerally difficult and expensive. Ahigh viscosity complicates biomassremoval from the broth. The removalof the cells is essential if the gum is tobe used in optically clear food prod-ucts and cosmetics. Products specifi-cations are less restrictive for usessuch as in textile processing. In the'Enhanced Oil Recovery ' application,the presence of the cells in the poly-mer is undesirable because such poly-mer fluid causes the plugging of thepores in the oil-bearing rocks. Theseparation of cells from viscous brothby centrifugation, filtration or by floc-culations is very expensive. The mainsteps of the recovery process are de-activation and removal (or lysis) of themicrobial cells, precipitation of thebiopolymer, dewatering, drying andmilling. Processing must be done with-out degrading the biopolymer. A fewmethods have been developed to de-activate, lyse, or remove cells from thebroth. Treatment with chemicals likealkali, hypochlorite or enzymes, bymechanical means, and thermal treat-ment are used. Chemical treatment athigh pH can cause depyruvylation ofthe product. When enzymes are used,

they must be removed from the me-dium and this adds to costs. Gener-ally, the fermentation broth is pasteur-ized or sterilized to kilt the cells. Pas-teurization of the fermentation broth ata high temperature often causes ther-mal degradation of the microbialexopolysaccharides. When the brothis treated under proper conditions (80-ISCC, 10-20 min, pH 6.3-6.9) en-hanced xanthan dissolution occurswithout thermal degradation and dis-ruption of cells is observed.

In the laboratory method, the cul-ture broth is diluted with 33% ethanolto 100 cP (Centipoise, a unit of viscos-ity). The cells are removed by centrifu-gation and 1% KCI is added to thebroth. Precipitation of the gum isachieved by the addition of 3 volumesof chilled 95% ethanol. The loweralcohols (methanol, ethanoi, isopro-panol) and acetone, which are non-solvents for the polysaccharide, canbe added to the fermentation broth notonly to decrease the solubility untilphase separation occurs, but also towash out impurities such as colouredcomponents, salts, and cells. Solventprecipitation, precipitation using diva-lent metal ion salts of the polymer atalkaline pH, precipitation as aquarternary ammonium complex atacidpH, or precipitation as fatty aminecomplex at acid pH have been suc-cessfully attempted. Whole broth con-taining xanthan can be spray or drumdried. However, the final product ishighly coloured because the bacteriapossess a yellow pigment. The gumitself is colourless. The bacteria havea yellow pigment in the wall, but it isextractable only with organic solvents.

Unique Properties

Xanthan exhibits pseudoplasticity(shear-reversible property) in aque-ous solutions. This characteristic so-lution property of the xanthan gum can

CHEMICAL BUSINESS -f JULY 2006 45

be explained on the basis of its helicalstructure. The viscosity of xanthan insolution instantaneously and revers-ibly decreases as the shear rate in-creases. Most aqueous solutions ofother polysaccharides show a sharpdecrease in viscosity as the tempera-ture is increased. Xanthan exists insolution at moderate temperature in anative, ordered conformation. On in-creasing the temperature of an aque-ous solution of xanthan, the viscosityincreases suddenly, suggesting acon-formational change. A xanthan solu-tion in presence of salts like sodium orpotassium chloride can maintain itsordered structure and viscosity up to100°C. Generally, at a low ionicstrength, polyelectrolytes adopt ahighly expanded conformation. On theaddition of salt, the conformation col-lapses-to form a more compact coildue to charge screening. Xanthansolutions (0.2-0.5%) show an increasein viscosity with an addition of 0.1%sodium chloride solution due to stabi-lization of the xanthan structure in theordered form, giving rise to intermo-lecular association.

The measurement of optical rota-tion of a salt free xanthan gum solu-tion has shown that the viscosity in-crease is exactly parallel to a decreasein the optical rotation. This is consis-tent with the unwinding of the orderedconformation such as helix into a ran-dom coil with a consequent increasein resultant shape and size of themolecules. This alters the viscosity ofthe solution.

At lowtemperatures, xanthan has acompact conformation that undergoesa transition to an expanded form at aspecific temperature, reported to be55°C. By NMR spectroscopy it is dem-onstrated that there is an ordered con-formation for xanthan below 55°C anda disordered random coil at highertemperatures. These changes have

also been examined by circular dichro-ism spectroscopy. Fourier transformIR spectroscopic investigation ofxanthan solution showed that an or-der-disorder transition occurs, as thetemperature is increased. This is cor-related with the sharp increase inviscosity in the same temperaturerange.

A tight relationship

There is a special relationship be-tween plant polysaccharides andxanthan gum. A synergistic viscosityrise is observed when the xanthan isadded to the solutions of plant polysac-charides such as locust bean gum andguar gum. The galactomannans ofthese gums link with helices of xanthanto form a more rigid molecular struc-

Table 1

Application

Food Industry

Juice drinks, chocolatespickles, fruit pulp.

Canned food, jams, jellies,milk products

Fro2en foods, sauces, gravies

Bakery products

Cheese, creams, meat productsmargarine

Industrial chemicals

Agriculture

Paints and inks

Ceramics

Paper manufacturing

Textile

Abrasives, adhesives, polish,toothpaste's, cosmetics, gelledexplosives

Enhanced oil recovery

Applications of xanthan

Properties

Suspending and thickening agentparticulate suspension {for chocolate)

Shear thinning properties ensuresfavourable viscosity under processingconditions. Good gelling agent, easypouring due to high pseudoplasticity.

Provides favourable emulsion, suspension,stability and viscosity, improves freeze-thaw

stability of starch thickened products.

For better texture to bread, mouth feel andrelease. Xanthan can replace gluten.

Combination of xanthan with plant gumslike locust bean gum. guar gum issuggested

Longer coniact with crops, clings duringspraying and controls drift of fungicide,herbicide, pesticide and fertilizers.

Stabilizer, emutsifier for thixotropic paints.compatible with other thickeners and waterbased emulsion Inks, prevent sagging andallows easy and even application.

Suspending agent for heavy particles inceramics glazes. Maintains proper viscosity

and lubricating power of suspension.

Clay coating for paper finish. As a rheologymodifier for high size press, roll coaling.

Suspending agent for dyes, pigments,controlling agent for printing-

Viscosity control, flow modifier.

Flocculent stable to pH and temperaturechanges and to high salt concentrations,effective lubricant, high pseudoplasticityallows easy injectability.

46 CHEMICAL BUSINESS -f JULY 2006

ture of a gel. It is this particular prop-

erty that facilitates use of xanthan gum

in many daily applications such as ice

cream, pasteurized process cheese

dips and spreads as well as a variety

of frozen desserts. Locust bean gum

is preferred over guar gum as it has

fewer galactose side chains. A greater

proportion of guar gum (80:20) is re-

quired for optimal synergy compared

to locust bean gum (50:50). On heat-

ing up to 55°C and cooling, such solu-

tions form thermally reversible stable

gels with a three dimensional network.

These are true gels, which do not flow

and do not recover from mechanical

damage.

Many Applications...!

The major applications of xanthan

gum are in food industry as a sus-

pending and thickening agent for fruit

pulp and chocolates. United States

Food and Drug Administration have

approved Xanthan on the basis of toxi-

cology tests for use in human food.

Many of today's foods require the

unique texturization, viscosity, flavour

release, appearance and water-con-

trol properties. Xanthan gum improves

all these properties and additionally

controls the rheology of the final food

product. It exhibits pseudoplastic prop-

erties in solutions, and has less

'gummy' mouth-feel than gums with

more Newtonian characteristics.

Xanthan gum is used in textile print-

ing pastes, ceramic glazes, slurry ex-

plosive formulations, and rust remov-

ers. Xanthan has wide applications in

the chemical industry (Table 1). A mix-

ture of xanthan and locust bean gum

can be used in deodorant gels. A spe-

cial gel of xanthan can form in pres-

ence of borate. This combination has

been used for preparation of explo-

sives. The ability of xanthan to impart

the desired dispersion stability at rest,

and low viscosity under application is

used to obtain a right consistency to

the toothpaste. Ceramic glazes con-

tain suspension of heavy particles.

Xanthan gum solution maintains

proper viscosity and lubrication power

of the suspension.

The special rheological properties

of xanthan are technologically suit-

able for the 'Enhanced Oil Recovery'

(EOR) application. At low concentra-

tion, the gum forms high viscosity so-

lution that exhibits pseudoplasticity.

The crude oil (petroleum) is held in the

tiny pores of the small sand stone

rocks. For the efficient displacement

of oil, the pumping of xanthan gum

solution in the rocks is necessary. As

a result, oil held in the pores of the

sand stone rocks is displaced.

The microbia! polysaccharides have

gained a significant commercial im-

portance during the last 10 years.

Considerable scope exists for further

improving the production and recov-

ery of xanthan, particularly through

modeling of the ferentation design.

The applications of xanthan are based

on its ability to alter the rheological

properties of aqueous solutions. How-

ever, carboxymethyl cellulose and

polyacrylamides are also competitive

to xanthan because of their lower price.

The commercial production of xanthan

requires a very high level of engineer-

ing and technical expertise as well as

marketing skill. Only those firms en-

gaged on developing a whole range of

microbial products, for example Tate

and Lyie or Keico Company, presently

produce xanthan. •

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