Caballero B. (ed.) Encyclopaedia of Food Science, Food Technology and Nutrition. Ten-Volume Set

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that have been derivatized, the maximum being 3.) In

some reactions, the pH needs to be controlled by the

metered addition of dilute sodium hydroxide solu-

tions. Following modification to the desired level,

the starch is recovered by centrifugation or filtration,

washed, and dried.

0042 Chemical reactions currently both allowed and

used to prepare modified food starches in the USA

are as follows:

0043 Esterification with acetic anhydride, succinic an-

hydride, the mixed anhydride of acetic and adipic

acids, 1-octenylsuccinic anhydride, phosphoryl

chloride, sodium trimetaphosphate, sodium tri-

polyphosphate, or monosodium orthophosphate

0044 Etherification with propylene oxide

0045 Acid modification with hydrochloric or sulfuric

acids

0046 Bleaching with hydrogen peroxide, peracetic acid,

potassium permanganate, or sodium hypochlorite

0047 Oxidation with sodium hypochlorite

0048 Various combinations of these reactions

Other reagents may be used in other countries.

0049 Waxy maize starch modifications are especially

popular in the US food industry because the inherent

properties of waxy maize starch make them preferred

over modifications to common corn starch.

Cross-Linking

0050 Cross-linking is the most important modification of a

food starch. Cross-linking occurs when starch gran-

ules are reacted with difunctional reagents to connect

hydroxyl groups on two different molecules within

the granule. Cross-links reinforce the granule and

reduce both the rate and the degree of granule swell-

ing and subsequent disintegration, i.e., reduce sensi-

tivity to processing conditions (high temperature;

extended cooking times; low pH; high shear during

mixing, milling, homogenization, and/or pumping).

Cooked pastes of cross-linked starches are more vis-

cous, heavier-bodied, shorter-textured, and less likely

to break down with extended cooking times, greater

acidity, or severe agitation than are pastes of the

native starches from which they are prepared. Only

a small amount of cross-linking is required to produce

a noticeable effect; for example, one cross-link for

every approximately 1200 a-d glucopyranosyl units

greatly reduces both the rate and the degree of

granule swelling, greatly increases paste stability, and

changes dramatically both the viscosity profile as the

starch is cooked and the textural characteristics of its

paste. Three times that much crosslinking, for

example, produces a product in which granule swell-

ing is restricted to the point that a peak viscosity is

never reached in a slurry heated to 95



C and held at

that temperature with moderate stirring. As the

number of cross-links increases, the granules become

more and more tolerant to physical conditions and

acidity, and swell and disintegrate (solubilize) upon

cooking less and less. Energy requirements to reach

maximum swelling and viscosity are also increased.

0051By far the most common cross-links are distarch

phosphate esters. These distarch phosphates are pre-

pared with either phosphoryl chloride or sodium tri-

metaphosphate. Phosphoryl chloride is very reactive

and undoubtedly reacts near granule surfaces. To

prepare cross-linked starches with phosphoryl chlor-

ide, the reagent is added to an aqueous starch suspen-

sion of pH 8–12. To cross-link a starch with sodium

trimetaphosphate, it is slurried in a solution of the

reagent at pH 5.0–8.5; the suspension is filtered, and

the starch is dried. In this case, the cross-links are

undoubtedly more evenly distributed throughout the

granule.

0052A relatively small amount of cross-linked starch is

made by reaction of corn starch with the mixed an-

hydride of adipic and acetic acids in aqueous alkaline

suspension.

Stabilization

0053Derivatization of a starch with monofunctional re-

agents reduces the intermolecular associations which

result in gelation of its paste and/or precipitation of

the starch polymers (combined processes termed

retrogradation or setback). Pastes of unmodified

starches generally will gel, and the gels will usually

be cohesive, rubbery, long-textured, and prone to

syneresis. (Waxy maize starch pastes gel to a very

limited extent at room temperature, but will become

cloudy and chunky and exhibit syneresis when stored

under refrigerator or freezing conditions.)

0054The most common derivatives employed for starch

stabilization are the hydroxypropyl ether and acetate

and monostarch phosphate esters. Acetylation is

accomplished by treating a starch slurry with acetic

anhydride at pH 7–11, the optimum pH depending

on the reaction temperature. Acetylation of starch

lowers the gelatinization temperature, an indication

of a weakening of granules. Upon cooking, a higher

peak viscosity is obtained due to greater granule

swelling. Upon cooling of the resulting paste, the

viscosity becomes lower than that obtained from the

unmodified starch, an indication of improved stabil-

ity, i.e., less retrogradation. Acetylated starches with

an acetyl content of up to 2.5% (degree of substitu-

tion, DS, 0.09) can be used in food products (USA).

(A DS of 0.09 indicates an average of nine acetyl

groups per 100 a-d-glucopyranosyl units.)

0055Sodium phosphate monoesters are prepared by

impregnating the starch with a solution of sodium

STARCH/Modified Starches 5577

tripolyphosphate. After adjustment of the pH to

5.0–8.5, the slurry is mixed, then filtered, and the

filter cake is dried and heated. Sodium tripolypho-

sphate is used to make products of up to 0.002 DS

(one phosphate group per 500 a-d-glucopyranosyl

units), the maximum allowed in the USA. Monoso-

dium orthophosphate in the pH range 5.0–6.5 is

also used to produce monostarch phosphates in the

same way.

0056 Monostarch phosphates produce stable pastes that

are clear and have a long, cohesive texture. Paste

viscosity can be controlled by varying the concentra-

tions of phosphate salt, time of reaction, temperature,

and pH. Increasing substitution lowers the gelatin-

ization temperature; products become cold-water-

swelling at DS 0.07. Corn starch phosphates of DS

0.01–0.03 produce pastes with hot viscosity, clarity,

stability, and texture more like those of potato starch.

Starch phosphates are good emulsion stabilizers and

produce pastes with improved freeze–thaw stability.

0057 Hydroxypropyl ether derivatives of starches are

prepared by reacting an alkaline slurry with propyl-

ene oxide. To a starch slurry is added sodium sulfate

and sodium hydroxide. The reactor is charged with

propylene oxide and sealed. Reaction is continued for

about 24 h at about 49



C. The maximum allowable

moles of substitution in the USA is 0.2 (7.0% of

hydroxypropyl groups). (Moles of substitution, MS,

is essentially the same as DS but is used in place

of DS because each hydroxypropyl group contains

a hydroxyl group that can itself be etherified, so

that the maximum number of substituent groups per

glucosyl unit can be more than 3.)

0058 Low-MS hydroxylpropylstarches behave much like

low-DS starch acetates and are used because of

similar improvements in texture and appearance.

The hydroxypropyl ether linkage is, however, much

Further Reading

Eliasson A-C (ed.) (1996) Carbohydrates in Food. New

York: Marcel Dekker.

Light JM (1990) Modified food starches. Cereal Foods

World 35: 1081–1092.

Whistler RL, BeMiller JN and Paschall EF (eds) (1984)

Starch: Chemistry and Technology, chapters 10, 17, 19.

Orlando, Florida: Academic Press.

Wurzburg OB (ed.) (1986) Modified Starches: Properties

and Uses, chapters 2–4, 6, 7, 9, 12. Boca Raton, Florida:

CRC Press.

Resistant Starch

R M Faulks, Institute of Food Research, Norwich, UK

Introduction

0001The term ‘resistant starch’ was first coined by Englyst,

Wiggins, and Cummings in 1982 to describe a small

STARCH/Resistant Starch 5579

fraction of starch that was resistant to hydrolysis by

exhaustive a-amylase and pullulanase treatment in

vitro. This starch fraction was found during the

destarching of food samples for dietary fiber

analysis and consists of retrograded amylose, B-type

starch granules, starch encapsulated within plant

or tissue structures, and some modified starch

structures.

0002 Recognition that this small fraction of starch could

be a significant contaminant of the dietary fiber led to

calls to have it included in the definition of dietary

fiber. More fundamentally, it also led to a reappraisal

of the way starches behave in vivo and, in particular,

whether or not resistant starches possessed any of the

properties usually associated with dietary fiber, e.g.,

fecal bulking, laxation, shorter transit times. How-

ever, because starch reaching the large intestine may

be more or less fermented by the gut microflora, some

workers prefer to consider resistant starch as that

portion of dietary starch which escapes digestion in

the small intestine. This is an important distinction

because resistant starch measured as a dietary fiber

contaminant can only account for a fraction of the

starch lost from the terminal ileum. (See Dietary

Fiber: Physiological Effects.)

0003 Because of the possible nutritional significance of

unavailable dietary starch, it is now widely accepted

that resistant starch is best defined as that fraction

of dietary starch which escapes digestion in the

small intestine. Resistant starch is therefore a variable

fraction of starch, defined by physical, chemical, and

physiological factors, but not necessarily resistant to

hydrolysis by starch-degrading enzymes in vitro. This

definition permits the fractionation of resistant starch

into different categories according to the reason for

its nondigestion (e.g., enclosed in food matrix, slowly

digested, enzymatically resistant), and allows the de-

termination of any functional or physiological role in

the small intestine. Furthermore, this classification

may permit a clearer understanding of how much

starch is lost from the small intestine, and of its

function and fate in the large intestine.

Formation

0004 The formation of resistant starch, or, more accurately,

the change in the degree of resistance of starch to

hydrolysis by a-amylase and glucoamylases, is a func-

tion of the structure and composition of the starch,

its processing environment, and subsequent storage

conditions.

0005 The well-known granular structure of starches is

lost if starches are heated above about 70



Cinan

aqueous environment, although some starches, par-

ticularly those from some legumes or high in amylose,

may not gel unless heated to a much higher tempera-

ture, e.g., 120



C. In conditions of limited or no mois-

ture, heating may not result in significant changes in

starch granules, which may retain their native crystal-

line form. Formation of resistance normally occurs

most extensively in high-amylose starches that have

been thoroughly dispersed by cooking (100–120



in water and subsequently allowed to cool to produce

a set gel containing 10–30% starch. Prolonged

storage of gels under cool conditions, freezing and

thawing, and reheating and cooling may also increase

the degree of resistance.

0006As the dispersed gel cools, sections of the linear

amylose molecules rapidly associate by hydrogen

bonding to produce retrograded crystalline struc-

tures. Starch gels can therefore be considered as gel-

atinized granules embedded in an amylose matrix. If

the amylopectin in the gelatinized granules is not too

highly branched, parts of its structure may also crys-

tallize. This retrogradation of amylopectin is a much

slower process. The result is therefore the production

of a gel system containing two phases: a highly struc-

tured phase, containing predominantly retrograded

amylose, and a less structured amorphous phase,

containing some retrograded amylopectin.

0007Examination of retrograded starch gels by differen-

tial scanning calorimetry clearly shows two endo-

therm peaks relating to the melting of retrograded

amylopectin (c.70



C) and retrograded amylose

(c. 160



C).

0008It is the formation of physically ordered structures

within the gel which is believed to confer resistance to

amylolysis. The degree of association (retrograd-

ation) of cooked starches may be influenced by a

range of factors, including amylose-to-amylopectin

ratio, starch concentration, pH, ions, temperature,

age, and the presence of lipids.

Physical and Chemical Properties

0009Resistant starch is not a chemically defined entity.

Its properties in vivo or in vitro can therefore be

described only in general terms.

Resistant Starch Granules

0010Raw starch granules embedded in the cellular struc-

ture of food fragments may escape digestion because

they are inaccessible to a-amylase and glucoamylases

in vivo. This occurs mainly with the consumption of

‘whole-foods’, in which the particle size is large, or in

tissues that have thick cell walls, or in some processed

foods, in which the food structure restricts access by

amylolytic enzymes. In these cases, both the chemical

and physical properties of the starch are normal for

the type of starch.

5580 STARCH/Resistant Starch

0011 Although most native starches undergo quite rapid

hydrolysis in vivo, in that they are found only in trace

amounts in the terminal ileum, some native starches

are very resistant to digestion. The main dietary

starches to show marked resistance to amylolysis

in vivo are raw banana and potato. It has been sug-

gested that resistance to digestion is caused by the B

form of crystallinity of the granule, unlike the A

forms (wheat, maize) and C forms (sweet potato,

tapioca) more commonly found. Retrograded

starches are also known to develop the B-form struc-

ture. (See Bananas and Plantains; Potatoes and Re-

lated Crops: Processing Potato Tubers.)

Resistant Cooked Starch

0012 The amorphous phase of retrograded starch gels is

rapidly hydrolyzed (in minutes) by a-amylase, leaving

the structured phase which is only slowly attacked

(days). The hydrolysis curve of cooked retrograded

starch gels takes the general form shown in Figure 1.

The initial rate and extent of hydrolysis are dependent

on the botanical origin of the starch and factors influ-

encing the degree of retrogradation and the a-amylase

activity. Prolonged treatment with a-amylase may

lead to almost 100% hydrolysis of the starch, except

in the case of pure amylose, when the degree of

hydrolysis is much lower. In vivo, therefore, resistant

starch from cooked foods is probably composed of

a variable proportion of highly a-amylase-resistant

starch (depending on the efficiency of digestion) and

any residual starch that is truly a-amylase-resistant

(retrograded amylose).

0013Enzymatic treatment of starch gels may be used to

isolate the more resistant fractions for further study.

0014Isolated starch with a high degree of resistance to

a-amylase is a water-insoluble white solid which

has none of the functional characteristics normally

associated with water-soluble polysaccharides (e.g.,

viscosity) or with the heterogeneous mixture of poly-

saccharides that make up the cell wall of plants.

0015Retrograded amylose isolated by a-amylase treat-

ment has been found to consist mainly of linear glu-

cose polymers with a degree of polymerization of 60.

Careful acid hydrolysis of retrograded amylopectin

yields the associated regions which have been shown

to contain branched fragments in which the individ-

ual chains have a degree of polymerization of 15.

0016Resistance to a-amylase can be effectively abol-

ished by melting the retrograded structures (c.70



for amylopectin and c. 160



C for amylose).

0017Disruption of the H bonds that hold the crystalline

structures together may be achieved chemically

by treatment with cold dilute alkali (potassium

hydroxide, 2 mol l

1

) or with hot (100



C) anhydrous

dimethyl sulfoxide (DMSO).

0018Some of the resistant structures from high-amylose

starch gels (retrograded amylose) may also show con-

siderable resistance to hydrolysis in dilute mineral

acid (sulfuric acid, 1 mol l

1

) and may need a Seaman-

type hydrolysis (cold sulfuric acid, 12 mol l

1

followed by 1 mol l

1

at 100



C) to achieve 100%

yield of glucose.

Physiological Effects

0019When resistant starch residues were first described in

‘destarched’ dietary fiber residues, it was considered

appropriate to allocate them to the dietary fiber com-

plex, despite the fact that they are predominantly

a-1,4-linked glucans and potentially digestible in the

small intestine. It was later realized that the amount

of resistant starch could be manipulated by process-

ing and would therefore be variable as well as chem-

ically distinct from the nonstarch polysaccharides

(NSP). More recently, it has been demonstrated that

highly a-amylase-resistant starch fractions that have

been isolated from different sources differ with re-

spect to the extent of digestion in the small intestine

and fermentation in the large intestine. Resistant

starch is therefore a variable fraction of the starch,

and the fact that it escapes digestion either in vivo or

in vitro is insufficient reason to allocate it as dietary

fiber. All the current main methods for the measure-

ment of dietary fiber (Englyst, Uppsala, Association

of Official Analytical Chemists) have modifications

that allow the inclusion or exclusion of residual

starch from the dietary fiber value. (See Dietary

100

01h2 3 4 5 6 7

Time (da

Extent of hydrolysis (%)

fig0001 Figure 1 Typical hydrolysis curve of retrograded starch gel

treated with a-amylase in vitro. Reproduced from Starch, Encyclo-

paedia of Food Science, Food Technology and Nutrition, Macrae R,

Robinson RK and Sadler MJ (eds), 1993, Academic Press.

STARCH/Resistant Starch 5581

Fiber: Properties and Sources; Dietary Fiber: Deter-

mination.)

0020 There is little experimental evidence for the func-

tional properties or physiological effects of resistant

starch in vivo. This is because (1) it is difficult to

define resistant starch other than in physiological

terms; (2) there are inadequate in vitro methods for

resistant starch measurement; and (3) feeding trials

have tended to use foods in which there are overlying

effects of the food matrix and a great excess of readily

digestible starch.

0021 It is known that when isolated, highly a-amylase-

resistant starch is fed to rats, there is some (20–60%)

digestion of the carbohydrate in the small intestine,

the remainder being lost to the cecum. This starch is

digested and absorbed more distally in the ileum than

rapidly digestible starch. More distal absorption of

glucose might be expected to have some effects on gut

physiology and gut hormones. However, no differ-

ences were found in glucagon, enteroglucagon, and

cholesterol between rats fed isolated resistant starch

from high-amylose maize starch as the only complex

carbohydrate, and those fed corn starch. However, in

the resistant-starch-fed rats, the crypt cell production

rate in the ileum was significantly greater.

0022 The presence of resistant starch in the diets of

unadapted rats appears to cause a slowing in both

the rate and velocity of ileal digesta transport. It may

be that this constitutes some evidence for an ‘ileal

brake’ mechanism which slows digesta movement as

a result of presence of absorbable carbohydrate in the

distal ileum. This effect is lost during adaptation.

0023 Starch that is not digested in the small intestine is

lost to the cecum or large intestine, where it may

undergo fermentation by the microflora. Evidence

from rat-feeding experiments has shown that the

amount of fermentation is dependent on the source

of the resistant starch and the extent of adaptation of

the animal.

0024 The ceca of rats fed resistant starch are generally

much enlarged, and the pH of the cecal contents is

lower than normal, indicating fermentative produc-

tion of fatty acids. By analogy with fermentable NSP,

the main fatty acids produced from the fermentation

of resistant starch are believed to be acetate, propion-

ate, and butyrate.

0025 Initially, a large proportion of resistant starch is

voided in the feces but this falls quite quickly as the

cecal microflora adapts. The rate of adaptation also

depends on the origin of the resistant starch. Rats fed

resistant starch from normal maize adapted quickly

(5 days), losing only a few percent of the ingested

resistant starch whereas those fed resistant starch

from wrinkled peas or high-amylose maize adapted

more slowly (28 days) and were still losing a high

percentage of the ingested resistant starch. Resistant

starch may therefore contribute significantly to fecal

bulk but this is variable depending on the state of

adaptation of the animal. (See Microflora of the

Intestine: Role and Effects.)

Significance

0026At this stage it is difficult to estimate the significance

of resistant starch. It is not well defined, and pro-

posed methods of analysis may determine fractions

that possess different structures and functions or that

may be physiologically irrelevant.

0027What can be said relates mainly to the properties of

resistant starch by analogy to dietary fiber and, by

extension, its deliberate production, either within a

food or as a food additive. Where resistant starch is

present in a food it will help to flatten the glycemic

response curve by reducing the amount of rapidly

digestible starch and by moderating the rate of diges-

tion. The resistant starch that reaches the more distal

part of the small intestine may exert an influence on

the rate of transport along the ileum, and help the

more uniform absorption of carbohydrate. This type

of ‘slow-release’ carbohydrate may find application

in improving the carbohydrate tolerance of diabetics,

and it has implications for the utilization of energy.

(See Glucose: Glucose Tolerance and the Glycemic(-

Glycaemic) Index.)

0028Any starch that is lost from the small to the large

intestine represents a loss of dietary energy since, even

if it is fully fermented, the energy salvaged is only

about 50% of that in the original carbohydrate.

Some additional losses from the small intestine of

other energy sources may occur as a result of any

‘carry-over’ effect.

0029The energy value of resistant starch is therefore

somewhat less than 16.8 kJ g

1

; its actual value

depends upon how much can be salvaged by fermen-

tation in the large intestine, the pattern of short-chain

fatty acids produced, and any energy lost to hydrogen

or methane.

0030The fermentation of dietary fiber in the large intes-

tine is believed to be a protective factor against large-

bowel cancer because the short-chain fatty acids

produced lower the pH and inhibit 7a-dehydroxylase.

This prevents the formation of secondary bile acids

that are suspected of a promotional role in colonic

carcinogenesis. In addition, butyrate, produced by

fermentation, is known to be a major energy source

for the colonic tissue and is believed to be a protective

factor in the development of colon cancer.

0031Carbohydrate fermentation in the large intestine

may therefore be a major protective factor against

the development of colon cancer. Because resistant

5582 STARCH/Resistant Starch

starch is at least partially fermentable, it may play a

crucial role in these mechanisms. (See Colon: Cancer

of the Colon.)

0032 Resistant starch has also been demonstrated to

have fecal-bulking properties and this could also

play a role in the dilution of fecal carcinogens by

reducing the time and level of exposure of the colonic

mucosa.

0033 From the energy point of view, there may be some

health benefits from consuming foods high in resist-

ant starch. Foods in which resistant starch may be

created in situ would have a lower energy density but

otherwise maintain their nutrient density. Starches

high in resistant starch or isolated resistant starch

fractions may be created with suitable functional

characteristics to find application in the food industry

as an ingredient or simply as a lower-energy bulking

agent.

See also: Bananas and Plantains; Colon: Cancer of the

Colon; Dietary Fiber: Properties and Sources;

Determination; Physiological Effects; Glucose: Glucose

Tolerance and the Glycemic (Glycaemic) Index;

Microflora of the Intestine: Role and Effects; Potatoes

and Related Crops: Processing Potato Tubers

Further Reading

Asp N-G, van Amelsvoorth and Hautvast JGAJ (1996)

Nutritional implications of resistant starch. Nutrition

Research Reviews 9: 1–31.

Englyst HN and Kingman SM (1990) Dietary fibre and

resistant starch. A nutritional classification of plant

polysaccharides. In: Kritchevsky D, Bonfield C and

Anderson JW (eds) Dietary Fibre, pp. 49–65. New

York: Plenum Press.

Englyst HN, Wiggins HS and Cummings JH (1982) Deter-

mination of the non-starch polysaccharides in plant

foods by gas–liquid chromatography of constituent

sugars as alditol acetates. Analyst 107: 307–318.

Faulks RM, Southon S and Livesey G (1989) Utilisation of

a-amylase (EC.3.2.1.1) resistant maize and pea (Pisum

sativum) starch in the rat. British Journal of Nutrition

61: 291–300.

Gee JM, Faulks RM and Johnson IT (1991) Physiological

effects of retrograded a-amylase resistant cornstarch in

rats. Journal of Nutrition 121: 44–49.

Livesey G, Davies IR, Brown JL, Faulks RM and Southon S

(1990) Energy balance and energy value of a-amylase

resistant maize and pea (Pisum sativum) starches in the

rat. British Journal of Nutrition 63: 467–480.

Miles MJ, Morris VJ, Orford PD and Ring SG (1985a) The

roles of amylose and amylopectin in the gelation and

retrogradation of starch. Carbohydrate Research 135:

271–281.

Miles MJ, Morris VJ and Ring SG (1985b) Gelation of

amylose. Carbohydrate Research 135: 257–269.

Orford PD, Ring SG, Carrol V, Miles MJ and Morris VJ

(1987) The effect of concentration and botanical source

on the gelation and retrogradation of starch. Journal of

the Science of Food and Agriculture 39: 169–177.

Ring SG, Colonna P, l’Anson KJ et al. (1987) The gelation

and crystallisation of amylopectin. Carbohydrate Re-

search 162: 277–293.

Ring SG, Gee JM, Whittam M, Orford P and Johnson IT

(1988) Resistant starch: its chemical form in foodstuffs

and effect on digestibility in-vitro. Food Chemistry 28:

97–109.

Zobel HF (1988) Molecules to granules: a comprehensive

starch review. Starch 40(2): 44–50.

STARTER CULTURES

L Durso and R Hutkins, University of Nebraska,

Lincoln, NE, USA

Introduction and History

0001 Starter cultures are an essential component of nearly

all commercially produced fermented foods. Simply

defined, starter cultures consist of microorganisms

that are inoculated directly into food materials in

order to bring about desired and predictable changes

in the finished product. These changes may include

enhanced preservation, improved nutritional value,

modified sensory qualities, and increased economic

value. Although many fermented foods can be made

without a starter culture, the addition of concentrated

microorganisms, in the form of a starter culture, pro-

vides a basis for insuring that products are manu-

factured on a consistent schedule, with consistent

product qualities.

0002Fermented foods and beverages have long been

manufactured without the use of commercial starter

cultures. Traditional methods of production include

backslopping, or using a small amount of the finished

specifically preserved product to inoculate a new

batch, the use of microorganisms found naturally on

the product, and the use of special containers that

allow for the survival of the starter culture microor-

ganisms within cracks and pores. These traditional

STARTER CULTURES 5583

methods allow for the development of individual var-

ieties of fermented foods and beverages, and they are

still practiced today for small- to mid-scale produc-

tion facilities, as well as in less developed countries

and in homemade-type products. Traditional

methods, however, are prone to slow or failed fermen-

tations, contamination, and inconsistent quality. In

contrast, modern large-scale industrial production of

fermented foods and beverages demands consistent

product quality and predictable production sched-

ules, as well as stringent quality control to insure

food safety.

0003 Given that pure culture techniques in microbiology

were not developed until Pasteur in the 1860s and that

lactic acid bacteria (LAB), in particular, were identi-

fied by Lister in the 1870s, it is noteworthy that an

industry devoted to producing pure cultures had its

beginnings only a short time later. In the late 1880s,

Storch in Denmark, Weigman in Germany, and Conn

in the USA showed that pure cultures could be used to

ripen cream, and soon the role of flavor-producing

bacteria (i.e., citrate-fermenting diacetyl-producers)

was established. By 1878 Christian Hansen began a

culture business that continues even today to be a

major supplier of starter cultures for the dairy, meat,

brewing, baking, and wine industries.

0004 Initially, starter strains were prepared by the manu-

facturer by growing pure strains in heat-sterilized

milk. Calcium carbonate was often added as a buffer

in order to maintain a neutral pH. These liquid cul-

tures remained popular until relatively recently, even

though they had a relatively short shelf-life due to the

loss of cell viability and fermentative activity. Eventu-

ally, rather crude dry culture preparations were pro-

duced which required several transfers in milk to

revive the culture to an active state. Freeze-dried cul-

tures also became available, but the early product also

required growth in intermediate or mother cultures.

Frozen cultures, now the most common form for

dairy cultures, were not introduced until the 1960s.

Significant improvements in freezing and freeze-

drying technologies have led these types of cultures

to dominate the starter culture market.

0005 The modern starter culture industry provides cul-

tures for nearly every type of fermented food and

beverage. Most culture houses also produce and sell

the media used to propagate starter cultures, the

enzymes used to coagulate milk for cheese produc-

tion, and other ancillary products. In addition, the

starter culture companies maintain a staff of highly

trained microbiologists who provide expert technical

service and support when issues or problems related

to culture performance arise. And although there are

many small culture manufacturers throughout the

world, specializing in cultures for specific products

or applications, the industry is dominated by a small

number of large companies.

0006This article will review the types of microorgan-

isms used as starter cultures, how starter cultures are

produced and preserved, criteria used by manufactur-

ers to assess culture performance, and ultimately

how these cultures are used by the fermented foods

industry. Issues facing the starter culture industry and

the new technologies being developed to address

these issues will also be examined. Throughout this

chapter, dairy starter cultures will be emphasized,

since the cheese and cultured dairy products industry

represents the largest user of commercial starter

cultures.

Starter Culture Microorganisms

0007Given that fermented foods have historically been

made using various species of bacteria, yeasts, and

molds, it follows that modern starter cultures also

contain these organisms. In general, bacteria are

used in the production of cheese, cultured dairy prod-

ucts, fermented sausages, and fermented vegetables.

Yeasts are used in the manufacture of bread and

alcoholic beverages, and molds are used for cheeses

and sausages as well as for soy-derived products, such

as soy sauce and tempeh. In most cases, the organisms

contained within a starter culture preparation are

well-defined, often to the species or even strain

level, and are carefully selected based on criteria rele-

vant for a particular product. Some starter culture

organisms, however, are not well defined, but instead

are selected based on a history of successful use.

Bacterial Starter Cultures

0008LAB are clearly the most important group of bacteria

used as starter organisms (See Lactic Acid Bacteria).

The LAB consist of a cluster of low G þC Gram-

positive cocci and rods from at least 11 genera; how-

ever, only a few of these are used as starter cultures

(Table 1). Lactococcus lactis subsp. lactis has long

been considered the most important LAB, due to its

widespread use in cheese manufacture, but

other species, such as Streptococcus thermophilus,

which has gained in importance due to its use in

yogurt and Mozzarella cheese, are also widely used

(Figure 1).

0009In general, the LAB are catalase-negative hetero-

trophs having complex nutritional requirements.

They have a fermentative metabolism, deriving

energy, in the form of adenosine triphosphate (ATP),

via substrate-level phosphorylation. Sugar metabol-

ism is either homofermentative or heterofermenta-

tive. In homofermentation, nearly all (> 90%) of the

5584 STARTER CULTURES

sugar substrate is converted directly to lactic acid,

whereas in heterofermentation, lactic acid, acetic

acid, CO

, and ethanol are produced. Although

most genera are either one or the other, some species

of Lactobacillus have the biochemical capacity for

both pathways. Both hetero- and homofermentative

LAB are used as starter cultures. Their optimum

temperature for growth varies; however, most LAB

used as dairy starter cultures are either mesophilic

(Lactococcus lactis) or are moderate thermophiles

(S. thermophilus and Lactobacillus spp.).

0010Since the primary function of starter culture bac-

teria is to ferment sugars and to produce acids, the

ability of LAB to metabolize carbohydrates is of crit-

ical importance. Indeed, the specific substrate range

and rate at which metabolism occurs may be used as

criteria for strain selection. In dairy fermentations,

for example, rapid lactose fermentation is required,

whereas in sourdough fermentation, maltose and

glucose metabolism are relevant and in sausage

fermentation glucose or sucrose metabolism is most

important. In addition, these bacteria are often

selected based on their ability to perform other meta-

bolic functions. In cheese, degradation of proteins via

proteolytic and peptidolytic enzymes is essential to

develop proper aged cheese flavor and texture. In

sour cream and cultured buttermilk manufacture,

the ability of the starter culture bacteria to utilize

citrate and to form the aroma compound diacetyl is

required. Production of exopolysaccharides by strains

of S. thermophilus is desirable in yogurt in order to

impart greater viscosity. In wine-making, the conver-

sion of malic acid to lactic acid by starter malolactic

tbl0001 Table 1 Lactic acid bacteria used as starter cultures

Organism Application

Lactococcus lactis

subsp. lactis

Cheese, cultured dairy products

Lactococcus lactis

subsp. cremoris

Cheese, cultured dairy products

Lactococcus lactis

subsp. lactis biovar.

diacetylous

Cheese, cultured dairy products

Lactobacillus

helveticus

Cheese, cultured dairy products

Lactobacillus delbrueckii

subsp. bulgaricus

Cheese, yogurt

Lactobacillus

sanfranciscensis

Sourdough bread

Lactobacillus casei Cheese, cultured dairy products

Lactobacillus sakei Sausage

Lactobacillus plantarum Sausage, fermented vegetables

Lactobacillus curvatus Sausage

Lactobacillus sanfrancisco Sourdough bread

Streptococcus thermophilus Cheese, yogurt

Pediococcus acidilactici Sausage, fermented vegetables

Pediococcus pentosaceus Sausage

Pediococcus halophilus Soy sauce

Oenococcus oeni Wine

Leuconsotoc lactis Cheese, cultured dairy products

Leuconsotoc mesenteroided

subsp. cremoris

Cheese, cultured dairy products,

fermented vegetables

fig0001 Figure 1 Electron micrograph of Streptococcus thermophilus.

STARTER CULTURES 5585

bacteria is necessary to reduce the acidity of certain

grapes.

0011 Although LAB are clearly the most widely used and

most important group of bacteria used as starter cul-

tures in fermented foods, other bacteria are also used

(Table 2). In the cheese industry, Propionibacterium

shermanni and Brevibacterium linens are used in

the Manufacture of Swiss (and related varieties) and

Limburger, Muenster, and Brick cheeses, respectively

(See Cheeses: Types of Cheese; Starter Cultures

Employed in Cheese-making; Chemistry of Gel For-

mation; Chemistry and Microbiology of Maturation;

Manufacture of Extra-hard Cheeses; Manufacture of

Hard and Semi-hard Varieties of Cheese; Cheeses with

‘Eyes’; Soft and Special Varieties; White Brined Var-

ieties; Quarg and Fromage Frais; Processed Cheese;

Dietary Importance; Mold-ripened Cheeses: Stilton

and Related Varieties; Surface Mold-ripened Cheese

Varieties). Some fermented sausage manufacturers

add Micrococcus spp. to meat batters in order to

develop flavor and color attributes (See Fermented

Foods: Fermented Meat Products). The manufacture

of vinegar requires Acetobacter aceti, which oxidizes

the ethanol generated from alcohol fermentations.

(See Vinegar.)

Yeast Starter Cultures

0012 Nearly all alcoholic beverages are made using one or

more strains of yeast (Table 2). Although many of the

wines produced around the world are made via nat-

ural fermentations, relying on yeasts ordinarily pre-

sent on the processing equipment or on the grape

surface, the trend for most wine manufacturers has

been to use starter culture yeasts, mainly strains of

Saccharomyces cerevisiae (See Wines: Types of Table

Wine; Production of Table Wines; Production of

Sparkling Wines; Dietary Importance; Wine Tasting).

Like LAB, strain selection is based primarily on the

desired attributes of the final product. In addition,

traits such as the ability to flocculate (and, hence, be

easily separated from the wine), to grow at high sugar

concentrations, and to produce adequate ethanol

levels, are all considered. Yeast starter cultures are

also used by the brewing industry, and although

many large breweries maintain their own proprietary

cultures, commercial ale (S. cerevisiae) and lager

(S. carlsbergensis) cultures are widely used (See

Beers: History and Types; Wort Production; Raw

Materials; Chemistry of Brewing; Biochemistry of

Fermentation; Microbreweries). Again, strain selec-

tion is based on characteristics relevant to the needs

of the particular brewer, and these include floccula-

tion, flavor development, and ethanol production

rates. Bread manufacturers represent the other large

user of yeast starter cultures (See Bread: Dough

Mixing and Testing Operations; Breadmaking

Processes; Chemistry of Baking; Sourdough Bread;

Dietary Importance; Dough Fermentation). Yeast

starter cultures used for bread-making are optimized

for the production of CO

from sugar or starch,

whereas brewing strains used in the production of

alcohol are generally optimized for the production

of ethanol.

Mold Starter Cultures

0013Mold cultures are used for several types of cheeses,

including all varieties of blue cheese, as well as the

white surface mold-ripened cheeses typified by Brie

and Camembert (Table 2). Blue mold cheeses are

made using spore suspensions of Penicillium roque-

forti and white mold cheeses are made using

P. camemberti. Mold cultures are also used in the

production of so-called Asian or soy-derived fer-

mented foods. Examples include tempeh, made

using Rhizopus microsporus subsp. oligosporus, and

miso and soy sauce, made using Aspergillus oryzae.

Fungal starter cultures are also occasionally used for

the production of European-style sausages and hams.

Types of Starter Cultures

0014One of the first requirements of a starter culture is that

it rapidly initiates a fermentation. Thus, the initial

inoculum must contain large numbers of microorgan-

isms. As the volume of the food (or liquid) increases,

then either larger starter culture volumes or greater

tbl0002 Table 2 Other organisms used as starter cultures

Organism Application

Bacteria

Brevibacterium linens Cheese: pigment, surface

Propionibacterium freudenreichii

subsp. shermanii

Cheese: eyes in Swiss

Staphylococcus carnosus

subsp. carnosus

Meat: acid, flavor, color

Mold

Penicillium camemberti Cheese: surface ripens white

Penicillium roqueforti Cheese: blue veins,

protease, lipase

Penicillium chrysogenum Sausage

Aspergillus oryzae Soy sauce, miso

Rhizopus microsporus

subsp. oligosporus

Tempeh

Yeast

Saccharomyces cerevisiae Bread: carbon dioxide

production

Saccharomyces cerevisiae Ale beers

Saccharomyces carlsbergensis Lager beers

Saccharomyces cerevisiae Wine

5586 STARTER CULTURES