the role of fat in the stabilisation of gas cells in bread dough

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Journal of Cereal Science 24 (1996) 187–198 The Role of Fat in the Stabilisation of Gas Cells in Bread Dough B. E. Brooker Institute of Food Research, Earley Gate, Whiteknights Road, Reading, Berkshire RG6 6BZ, U.K. Received 7 August 1995 ABSTRACT Electron microscopy has shown that during dough mixing by the Chorleywood Bread Process, fat crystals develop a crystal–water interface as they emerge from droplets of shortening and that they then adsorb to the gas–liquid interface of bubbles. In this process, the interface surrounding each crystal coalesces with the gas–liquid interface of the bubble. This adsorption process was not observed when triglyceride was added to doughs in the form of oil. The expansion of bubbles during proofing leads to the adsorption of many more fat crystals as they are encountered in the aqueous phase. During baking, fat crystals melt and thereby make it possible for the crystal-liquid interface to be incorporated into the surface of the bubble as it expands. This transfer of interfacial material from crystals to bubble surface explains how the addition of shortening to dough allows bubbles to expand during baking without rupturing, thus producing high volume bread with fine crumb structure. It follows that the amount of interface transferred to bubble surfaces for any given weight of fat is inversely proportional to the size of the crystals in the shortening and that therefore shortenings containing small crystals are more eective in producing high quality bread than those containing large crystals. A mechanism is now proposed which, for the first time, explains the precise role of fat crystals in this process and why the addition of oil is not eective. 1996 Academic Press Limited Keywords: bread, dough, fat, gas cell, stabilisation. the significance of this observation in terms of a INTRODUCTION general mechanism for air bubble stabilisation has It has been known for some time that the addition not been elucidated. of small amounts of shortening to bread dough None of the numerous hypotheses discussed in leads to improved loaf volume and finer, more the literature provides an acceptable explanation uniform crumb structure with thin cell walls. A for the mechanism by which solid fat improves direct relationship has been shown between the loaf volume during baking although even in some solid/liquid ratio of the shortening and the per- of the earliest publications it was accepted that formance of that fat in the baked product 1,2 ; as fat somehow acts by allowing bubbles to expand the proportion of solid (crystalline) fat increased during baking without rupturing 1,6 . Solid fat has to an optimum level, baking performance also been hypothesised as providing a reservoir of crys- improved. Interestingly, the most eective crys- tals that melt in the dough as the temperature talline fats were those with high melting points rises, thus providing a continuous supply of melted that remained solid at the end of proofing 2–5 , but fat to seal oleaks in the expanding dough film and thus improve gas retention 2,6 . However, the observation that bread dough continued to expand long after the fat crystals in the shortening had : CBP=Chorleywood Bread Pro- melted 4 threw doubt on this idea. cess; CLSM=Confocal Laser scanning microscopy; Work with short-time Chorleywood Bread Pro- FU=functional units; NMR=nuclear magnetic reson- ance; TEM=Transmission electron microscopy. cess doughs indicated that shortening delays the 0733–5210/96/060187+12 $18.00/0 1996 Academic Press Limited 187

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Page 1: The Role of Fat in the Stabilisation of Gas Cells in Bread Dough

Journal of Cereal Science 24 (1996) 187–198

The Role of Fat in the Stabilisation of Gas Cells inBread Dough

B. E. Brooker

Institute of Food Research, Earley Gate, Whiteknights Road, Reading, Berkshire RG6 6BZ, U.K.

Received 7 August 1995

ABSTRACTElectron microscopy has shown that during dough mixing by the Chorleywood Bread Process, fatcrystals develop a crystal–water interface as they emerge from droplets of shortening and that theythen adsorb to the gas–liquid interface of bubbles. In this process, the interface surrounding eachcrystal coalesces with the gas–liquid interface of the bubble. This adsorption process was not observedwhen triglyceride was added to doughs in the form of oil. The expansion of bubbles during proofingleads to the adsorption of many more fat crystals as they are encountered in the aqueous phase.During baking, fat crystals melt and thereby make it possible for the crystal-liquid interface to beincorporated into the surface of the bubble as it expands. This transfer of interfacial material fromcrystals to bubble surface explains how the addition of shortening to dough allows bubbles to expandduring baking without rupturing, thus producing high volume bread with fine crumb structure. Itfollows that the amount of interface transferred to bubble surfaces for any given weight of fat isinversely proportional to the size of the crystals in the shortening and that therefore shorteningscontaining small crystals are more effective in producing high quality bread than those containinglarge crystals. A mechanism is now proposed which, for the first time, explains the precise role offat crystals in this process and why the addition of oil is not effective. 1996 Academic Press Limited

Keywords: bread, dough, fat, gas cell, stabilisation.

the significance of this observation in terms of aINTRODUCTIONgeneral mechanism for air bubble stabilisation has

It has been known for some time that the addition not been elucidated.of small amounts of shortening to bread dough None of the numerous hypotheses discussed inleads to improved loaf volume and finer, more the literature provides an acceptable explanationuniform crumb structure with thin cell walls. A for the mechanism by which solid fat improvesdirect relationship has been shown between the loaf volume during baking although even in somesolid/liquid ratio of the shortening and the per- of the earliest publications it was accepted thatformance of that fat in the baked product1,2; as fat somehow acts by allowing bubbles to expandthe proportion of solid (crystalline) fat increased during baking without rupturing1,6. Solid fat hasto an optimum level, baking performance also been hypothesised as providing a reservoir of crys-improved. Interestingly, the most effective crys- tals that melt in the dough as the temperaturetalline fats were those with high melting points rises, thus providing a continuous supply of meltedthat remained solid at the end of proofing2–5, but fat to seal off leaks in the expanding dough film

and thus improve gas retention2,6. However, theobservation that bread dough continued to expandlong after the fat crystals in the shortening had : CBP=Chorleywood Bread Pro-melted4 threw doubt on this idea.cess; CLSM=Confocal Laser scanning microscopy;

Work with short-time Chorleywood Bread Pro-FU=functional units; NMR=nuclear magnetic reson-ance; TEM=Transmission electron microscopy. cess doughs indicated that shortening delays the

0733–5210/96/060187+12 $18.00/0 1996 Academic Press Limited187

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B. E. Brooker188

release of carbon dioxide during baking7. How- phenomena being studied were independent of thesource of the ingredients.ever, this could not be confirmed in work with

conventionally mixed and fermented doughs A white flour (10·9% protein, flour weight basis,14·4% moisture and 0·6% ash) produced with awhich showed that carbon dioxide was released

at the same time and rate from doughs with and laboratory Buhler mill from a commercial grist,was stored at −25 °C before being used. Twowithout shortening8. In this work it was also found

that dough containing shortening continues to commercial white bread flours (11·1 and 11·8%protein, 14·6 and 14·8% moisture and 0·7 andexpand at a higher temperature than does no-

shortening dough8, indicating, perhaps, that short- 0·6% ash, respectively) and were used to makebread doughs immediately after purchase.ening delays gelatinisation, a possibility suggested

previously9 but refuted in later work10,11. More A commercial bakery shortening (Ambrex) wasobtained from Peerless Foods (slip point=45 °C);recently, a mechanism for the involvement of hard

fat in bubble stabilisation was proposed12 in which in addition, an experimental shortening consistingof a mixture of hardened palm oil and rapeseedfat crystals disrupt the natural bilayer structure of

polar lipids and enable them to be incorporated oil in a ratio of 1:4 (slip point=41 °C) was preparedon a laboratory scale by melting and mixing to-into the gas–water interface.

In several respects, the role of crystalline fat in gether the oils followed by re-crystallisation in aSchroder 3-stage scraped surface heat exchanger.bread dough closely resembles that already established

in cake batters where it provides good crumb structure, Pulsed NMR (Minispec, Bruker) showed that at theproof temperature of 40 °C, the solid fat content oflarge cake volume and expansion without collapse

during baking. For both cake batters and doughs Ambrex was 7·6% and that of the experimentalshortening, was 5·9%. Shortenings were stored atalike, oils used in place of shortening do not impart

any of these physical properties2. 4 °C prior to use.Recent work on the role of fat in the stabilisation

of air in ‘all in one’ cake batters13 has shown that fatcrystals are ejected from shortenings during mixing, Methodsbecome enveloped by a fat(crystal)–water interface

Analysis of floursand are able to stabilise large numbers of small airFlours were analysed using the following methods:bubbles by adsorbing to their surface. During baking,moisture (102 °C, 18 h), protein (Kjeldahl, N×5·7)air bubles can expand without rupturing because ofand ash (500 °C, 16 h).extra interfacial material provided by adsorbed fat

crystals when they melt. Macroscopically, these eventsproduce a batter that can expand during cooking Bread-making processwithout collapsing to produce a high volume cake of A medium-scale short-time, high work inputfine crumb structure. method based on the CBP was used to produce

It seems likely that elements of this basic mechanism two types of dough. The standard dough consistedfor water continuous batters could be equally well of flour (100 parts), compressed yeast (2·05 parts),applied to explain the functional properties of solid salt (2·8 parts), ascorbic acid (0·01 parts) and waterfats in bread doughs. The aim of the present study (60 parts). The alpha-amylase activity of the flourwas therefore to investigate (a) the structural basis of was adjusted to 80 FU by the addition of fungalthe improved performance conferred by fat; (b) the alpha-amylase. Another simplified formulationmechanism by which fat crystals allow bubbles to contained only flour, water and yeast in the sameexpand without rupturing; and (c) why fat is more proportions as above.effective than oil. The experimental approach was to To follow the fate of fat crystals when addeduse freeze fracturing in conjunction with electron to dough, experimental doughs were preparedmicroscopy to follow the fate of fat during dough containing 1% (with respect to flour weight) short-development and after baking. ening. Other experimental doughs containing 1%

(with respect to flour weight) soya oil were alsoEXPERIMENTAL prepared. Control doughs contained no added

shortening or oil.Materials Ingredients were mixed in a Morton mixer(200 rev/min) to a total work input of 40 kJ/kg ofA number of different flours and shortenings were

selected for this study to ensure that the interfacial dough before proofing to a constant height of

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The role of fat in bread 189

Figure 1 Confocal laser scanning micrograph of commercial bread shortening showing numerous (black) fat crystals dispersedin a continuous (white) oil phase. Note that most of the crystals are in the form of clumps or aggregates. Stained with NileRed; excited at 488 nm.

12 cm in 400 g bread tins at 40 °C in a chamber holder or by microtoming the specimen on thewith high relative humidity to prevent skin for- brass table. The fractured surface was then uni-mation. Loaves were baked at 250 °C for 30 min directionally coated with 2 nm of platinum at anin a gas-fired oven and allowed to cool for 6 h and angle of 45° and 20 nm of carbon at an angle of24 h before being examined further by electron 90°. The platinum/carbon replicas were floatedmicroscopy. from the surface of the dough by soaking in distilled

water for 24 h, cleaned in 70% sulphuric acid orchromic acid for 24 h, washed in distilled waterTransmission electron microscopy (TEM)for 24 h, transferred to acetone for 1 h andSamples of dough (approximate volume 6–8 mm3)mounted on 300 mesh copper grids prior to ex-following mixing and proofing were removed fromamination in a Hitachi H-600 TEM operating ata centre portion using a pair of scissors. They werea voltage of 75–100 kV.then either (a) mounted in a Balzer complementary

To examine the structure of the internal surfacereplica holder or (b) attached to a brass specimenof bubbles in bread, a thin slice, 2 mm thick andtable with Tissue-Tek and then rapidly frozen byof total volume 16–20 mm2, was cut using a double-plunging into a well, 70 mm deep, containingedged razor blade to expose the inside of many ofliquid propane (−189·6 °C). They were thenthe air bubbles. The slice was attached to a brasstransferred to the pre-cooled (−135 °C) stage oftable with Tissue-Tek, frozen, microtomed anda Balzer BAF 400T freeze etching unit fitted withcoated with layers of platinum and carbon followedelectron beam guns for the evaporation of carbonby processing to clean the replica as describedand platinum and then fractured. This was done

either by opening the complementary replica above.

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B. E. Brooker190

Figure 2 Dispersed fat crystals (C) in bread dough at the end of mixing prepared using a commercial shortening. Y=yeastcell. Freeze fracture, TEM.

Freeze fracturing produced fractures through 8 °C. Samples were examined using a Zeiss CLSMII confocal microscope and images obtained usingbubbles so that only a segment of each was re-

vealed. For each specimen, the average number of the 488 nm laser line.fat crystals adsorbed to the surface of 50 fracturedbubbles was determined by direct examination of

Resultstransmission electron micrographs. To obtain anestimate of the total surface area of the segments, When Nile Red-stained bread shortenings werestereo pairs of electron micrographs were digitised examined by CLSM, the fat crystals could beand stored in a LINK AN10000 spectrometer seen by negative contrast against a background of(Oxford Instruments) and stereological meas- intensely fluorescing oil phase. Fat crystals do noturements taken using the ‘stereo’ image processing stain with Nile Red14. In Ambrex, crystals weresoftware. up to 8 lm long and 1 lm wide and were not

uniformly dispersed in the oil phase but insteadwere predominantly arranged in clumps of 10–20Confocal laser scanning microscopy (CLSM)

A CLSM was used to examine the crystal structure crystals (Fig. 1). In the experimental shortening,crystals were smaller (up to 3 lm long and 0·5 lmof the shortenings. Large samples (20 mm×20 mm

×3 mm thick) of the Ambrex and experimental wide) but were uniformly distributed throughoutthe continuous oil phase.shortening stored at 8 °C were applied to glass

microscope slides and a light dusting of Nile Red When doughs containing shortening were ex-amined by TEM ex-mixer, single fat crystals (re-was flicked onto their surface using a small brush.

A coverslip was then applied and the dye allowed cognisable from their characteristic lamellarstructure) or small aggregates of crystals wereto diffuse into the specimen for several hours at

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The role of fat in bread 191

Figure 3 A fractured small air bubble (B) (>5 lm diameter) from dough after mixing showing adsorbed fat crystals (arrows).Commercial shortening. Freeze fracture, TEM.

found dispersed between yeast cells, starch gran- of less than 20 lm (Fig. 3). This important processof crystal adsorption involved the coalescence ofules, gluten and air bubbles, as were occasional

droplets of shortening from which the fat crystals crystal–liquid interface with the gas–liquid in-terface of the bubble and resulted in the directhad originated (Fig. 2). This agreed with previous

lipid binding studies15, in which it was found that exposure of naked triglyceride to the gas phase.As the bubbles expanded during proofing, theat the end of mixing 86% of the solid fat remained

free and 14% was bound. However, because of number of adsorbed fat crystals increased pro-gressively until, at the end of the process, anthe small amount of shortening contained in the

doughs, it was rare in the present study to find average of 0·2 crystals/lm2 of bubble surface werefound in the case of the experimental shortening;crystals emerging from a droplet of shortening

in the manner described previously from cake for Ambrex, the corresponding figure was 0·06crystals/lm2 although it should be noted that theybatters13. By the end of mixing, fat crystals had

adsorbed in small numbers to the gas–liquid in- tended to adsorb as widely spaced clumps ratherthan as individual crystals (Fig. 4). At the endterface of some air bubbles (<0·001 crystals/lm2),

including the very small bubbles with a diameter of this process, adsorbed crystals were typically

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B. E. Brooker192

Figure 4 Part of the inside of a large gas bubble in proofed bread dough (containing experimental shortening) showing theadsorption of clusters of needle-like fat crystals. Freeze fracture, TEM.

oriented with their long axes parallel to the plane bubbles but there was no interfacial adsorptioncomparable to that seen with fat crystals. Similarly,of the gas–liquid interface (Fig. 5). In proofed

doughs, the gas–liquid interface pressed hard in control doughs containing no added triglyceride,droplets of endogenous flour lipid were never seenagainst starch granules in the continuous phase,

resulting in indentations in the bubble surface (Fig. adsorbed to the gas–liquid interface.When doughs containing shortening had been5). These interfacial phenomena were similar in

doughs with and without salt, ascorbic acid and baked and the bread allowed to cool, examinationof the internal surface of bubbles showed that thealpha-amylase additives and were independent of

type of fat or flour used. Neither fat crystals nor interface was covered with a discontinuous layerof oil (Fig. 6) in which there was only a limitedfat crystal adsorption to the surface of bubbles

were seen in control doughs to which no shortening indication of fat re-crystallisation (Fig. 7), evenafter some days of storage. All doughs containinghad been added.

In doughs to which soya oil had been added, fat had a similar interfacial morphology. In ex-perimental doughs containing soya oil and in con-oil droplets were found close to the surface of

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The role of fat in bread 193

Figure 5 The inside surface of a gas bubble in proofed dough containing an experimental shortening showing the adsorptionof a number of fat crystals (C). Most crystals lie with their long axis in the plane of the interface. Large starch grains (arrows)bulge into the bubble but do not penetrate the gas–liquid interface. Freeze fracture, TEM.

trols with no added triglyceride, the inside surface become enveloped in a crystal–liquid interfacialof the bubbles was smooth in appearance with no layer which coalesces and becomes continuousindication of fat crystals or oil. with the gas–liquid interface of bubbles when they

collide. As a result of this adsorption process,crystals are brought into direct contact with the

Discussion gas in the bubble.The observations made in the present studyPrevious work on the stabilisation of air in cake

are entirely consistent with there being a similarbatters led Brooker13 to propose a mechanism inmechanism operating in bread doughs and thewhich fat crystals effectively act as vehicles for theessential elements of this are represented in Figuretransfer of additional interfacial material to the8. This not only satisfactorily explains many ofsurface of expanding bubbles during baking andthe questions that have been posed in the pastthereby prevent their rupture. In this mechanism,about the role of fats in bread doughs but alsofat crystals emerge from droplets of shortening in

large numbers during the mixing process and embodies the principles by which the functionality

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B. E. Brooker194

Figure 6 The inside surface of a bubble from bread 6 h after baking showing areas of air-water interface (∗) and streams oflipid representing the pooled oil from the many fat crystals that melted during baking (arrows). Individual droplets of oilproduced when single crystals or clusters of crystals melted can also be seen (open arrows). Very little of the oil has re-crystallised (C). Freeze fracture, TEM.

of bread shortenings might be predictably op- due to differences in surface area. The superiorperformance of shortenings containing small fattimised in the future. In particular, it can be seen

that for any given mass of solid fat, a large number crystals has been suspected for some time5,6.When crystals melt during baking, the fat–liquidof very small crystals would be expected to convey

more interface to the surface of expanding bubbles, interface of the many adsorbed crystals providesa source of extra interfacial material for the bubbleand as a consequence, produce greater im-

provement to crumb and loaf volume, than the surface which allows expansion without rupturing.The consequence is that large numbers of smallsame mass of much larger, but fewer, crystals.

Similarly, it would be expected that aggregates of bubbles may survive baking and thereby contributeto crumb quality, whereas in the absence of fatfat crystals, of the type found in the commercial

bread shortening which adsorb to the air–water many bubbles rupture or are able to decreasetheir surface to volume ratio by coalescing withinterface as aggregates would be less effective in

transferring interfacial material to the surface of neighbouring bubbles, resulting in coarser crumbstructure. The improvement in crumb colour pro-bubbles than the same number of dispersed crystals

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The role of fat in bread 195

Figure 7 The inside surface of a bubble from bread 12 hours after baking showing the air–water interface (∗) with a streamof lipid (arrows), only some of which shows signs of re-crystallisation (C). Note isolated droplet of oil from the melting of smallcrystal clusters (open arrow). Freeze fracture, TEM.

duced by fat5 is probably the result of increased crystal adsorption then takes place. When lowermelting point fats are used, all the crystals soonreflection of light from the increased number of

small bubbles that survive baking. melt to form small oil droplets and there are nocrystals available for adsorption to bubbles, withThe proposed mechanism in Figure 8 also ex-

plains why it is important to use bread shortenings all the disadvantages this has for the expansion ofbubbles during baking.in which the melting point or slip point of the fat

is higher than that of the proofing temperature, The finding in the present study that dropletsof added oil are unable to adsorb to the surfacei.e. some fat crystals persist to the end of proofing.

At the end of mixing, the bubbles are relatively of bubbles probably explains why oil does notproduce the same improvements in bread qualitysmall with only few fat crystals adsorbed to their

surface. However, when proofing commences, the that have been attributed to solid fats1,2. Theadverse effect of oil compared with no-fat controls2expanding bubble surfaces are brought into con-

tact with many more of the free fat crystals dis- implies that there is a destabilising interactionbetween oil and the surface of bubbles but thepersed in the continuous phase and most of the

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B. E. Brooker196

Fat crystalsFat crystals

Aqueous phase

Mixing

Air

Aqueous phase

Final product

Air

Proofing

Air

Baking

OilOil

OilOil

Proteininterface Oil

Air

(a)

(b)

(c)

(d)

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The role of fat in bread 197

structural basis of this phenomenon was not ob- suggests that oil-soluble components in the gas–liquid interface (such as polar lipids) may dissolveserved in the present study. However, the puzzling

effectiveness of castor oil as a dough improver16 in it during baking and later inhibit re-crys-tallisation, as described by Garti19 for some sur-may be due to the presence of the hydroxyl group

in ricinoleic acid and its presumed surfactive prop- factants.The presence of this thin layer of triglycerideerty that would be important in the stabilisation

of bubbles during mixing, proofing and baking15. in close contact with the air–water interface mayhave interfered with some previous attempts toThe apparent paradox that only solid fat confers

improved baking performance but that the be- locate flour proteins in the gas–liquid interface bytreatment with antibody prior to embedding andneficial effects continue during baking long after

all fat has melted4,17 can now be reconciled by sectioning. For example, in a study in which alabelled anti-gliadin monoclonal antibody wasFigure 8. Once, during mixing and proofing, the

critical transfer of extra interface by crystal ad- used to locate storage proteins in bread, it wasfound that although the antibody bound stronglysorption to bubbles has taken place, subsequent

melting of the fat is irrelevant and serves only to to proteins in the matrix, there was little or nolabelling of the gas cell surface20. This suggestsmake an interface available for bubble expansion.

Whereas in cake batters the escape of fat crystals either that gliadins are not present at the interfaceor that future attempts to locate interfacial proteinsfrom droplets of shortening into the aqueous phase

is mediated by added emulsifiers that lower the in this way should use bread containing no addedshortening.surface tension of the oil–water interface, the

agents responsible for this same process in doughs(in the absence of any added emulsifier) have not

Acknowledgementsbeen identified. However, the flour polar lipids inthe aqueous phase are likely candidates18 especially This work was made possible by funding from theas their concentration is higher in bread dough BBSRC. Grateful thanks are due to John Price for

technical assistance.than in batters.One feature of note in Figure 5 is that during

proofing of bread doughs, most crystals come toREFERENCESlie with their longest surface in the plane of the

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2. Baker, J.C. and Mize, M.D. The relation of fats toever, this happens to only some of the crystals13

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Figure 8 Diagrammatic representation of the interfacial behaviour of fat during the mixing, proofing and baking of breaddough: (a) dispersion of fat crystals and their occasional adsorption to the gas–liquid interface during mixing; (b) increasingnumbers of crystals adsorb during proofing as the surface of bubbles expands; (c) during baking, crystals melt and the fat–liquidinterface is incorporated into the bubble surface; (d) in the baked bread, the fat forms a discontinuous layer on the insidesurface of bubbles.

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