Review

The coming
Recent advances in probiotic research have finally confirmed
the health benef~s of some probiotic bacterial strains. In ad-
dition to their application to dairy products, prabiotic bacteria
of age of probiotics
are now being applied to new food products including fer-
mented cereals, infant formulae and therapeutic foods. It is
very important to be able to enhance the stability d probiodc Yuan-KunLeeandSe@oSalminen
bacteria both in old and new applications in order to expand
their new uses and also to guarantee minimum effective doses
for particular products. recently been reviewed~. It was concluded that the
frequeney of occurrence of lactic acid becteda as ep-
po~unistic hacteria is exUemaly ram Also, the necur-
rence of apy vindence ~ in lactic acid bacteria
Almost a hundred years have passed since the introduc- remains to be shown. The International ~ of
tion of the theories on the prolongation of life by the Microbiological Societies reviewed the literatme, and
modulation of the intestinal ecosystem. However, only concluded that the clinicaldata do not ~ that
recently has the scientific basis of probiotic studies been l a ~ and related organisms z ~ such risks.
firmly established and sound clinical studies of soma However, as new pmbiotic preparations also im:lude
strains published. The physiological and nutritional bilidobacteria, enterococ~ Wopionlbtgte~ and even
properties of some strains of probiotic bacteria are now Saccharomyces strains, it is important to verify the
well understood; furthermore, it is possible to verify that safety of new probiotic strains. Although this is an im-
soma strains are probiotic with documented examples of portant aspect, no firm guidelines exist for safety test-
their ability to maintain and promote the health of the ing. An example of a suitable safety assessment pro-
host. cedure is given in Ref. 2. Safety assessment should
While clinical research has progressed, developmants include studies on: the risk of microbial invasina (infec-
have also occurred in the assessmant and improvement tivity), basic toxicology, including the development of
of the stability of probiofic bacteria in dairy products deleterious metabolites in the intestine; the degradation
and functional foods. Pmbiotic bacteria are often un- of the intestinal mneosa; as well as provide epidemio-
stable in retail products. With their stability improved logical data and evidence on safety. Further guidance
and probiotic effects confirmed, we are now approach- may be found in the European 'novel food' guidelines.
ing the general acceptance of probiotics as documented Further evidence of safety can be a s s m ~ by selecting
functional ingredients of foods and beverages. strains of human origin, and strains that m'e relatively
common in the human intestinal U'ecP. Such strains
Requirements for probiotic bacteria have been isolated and ro-adminigtexed gO ~
The basic requiremants for strain selection are given jects and include Lactobac//~ ~ m m u m ,
in Box 1. It is important that probiotic strains are of svuteri and Lactobacillus casei anbsp, rlmmr.osus.
human origin because some health-promodng effects
may be species dependent. Other required propezties
include acid and bile resistance and the ability to colo-
nize the human intestinal tract, at least temporarily. It is
likely that some mechanisms for adhering and/or bind-
ing to the intestinal cells are also required. Adherent
strains of probiotic bacteria are favoured because they
are more likely to becoma established in the intestinal
tract, and thus have greater probiotic effects. It is also
important for probiotic strains to be antagonistic against
cariogenic (i.e. those that cause dental caries) and patho-
genic bacteria either by antimicrobial substance produc-
tion or competitive exclusion. Finally, the strains should
be safe and tested for human use.
The use of lactic acid bacteria in foods has a long
history, and lactobaciIli have a 'generally recognized
as safe' status. The safety of lactic acid bacteria has

Yuan.KunLee is at the Departmentof Microbiology,Facultyo~ Medicine,

NationalUnive~P/of Singapore,KentRidge,Sinsapore0511 (fax:+65-776-
6877; e-mail:micleey~leonis.nus.ss).SeppoSalmlnenis at the Department
o~ BiachemisUyand Food ChemisUy,Unive~ of Turku, 20500 Turku,
Finland.

Trendsin Food Science & Technology July 1995 [Vol. 6]

 T~de 1. Fml~o~c bacterid ~mdye~t ~ m~t thair reposed effects

~aJn e~l~eea effe~ in d i n i ~ ~ i e ~ ~~ec~_..e~~,ferer~

Lactobacillusacidophilus LCI
Lactobacillusacidophilus
NFCO1748
LactobacillusGG
LactobacilluscaseiShirota
Lactobacillusgassed(ADH)
Bifidobac~um bifidum
I.actobacillusbulgaricus
Saccharomy~ boulardii
Probiotic mixture (seetext)
Immune enhancing; vaccine adjuvant; adherenceto human intestinal cells; 5-7
balancing of internal microflora
Lowering of faecal enzymes,prevent~n of radiotherapy-relateddiarrhoea; 2, 27
Ueatmentof constipatino
Preven~onof antibiotic-amsuciateddian'hoea;treal~nentand prevention of 2,10-14
rotavinJsdiarrhoea; beatment of relapsingClostddiumdi~Mle diarrhoea;
p~eventionof acute diarrhoea; alleviation of Crohn's disease;antagonistic
againstanticariogenic bacteria
Preventionof intestinal disturbances;balancing of intestinal bacteria; lowering 15,16
of faecal enzyme activities; inhibition of superficial bladder cancer
Faecalenzyme reduction; survived in inteslinal tract 17
Treatmentof rotavims diarrhoea; balancing of intestinal microflora; treatment 18,19
of viral dim'noea
No effect on rotavirus dian'neea;no immune-enhancingeffect during rotavims 4,12
dianhoea; no effect on faecal enzymes
Preven~ of uavelleds diarrhoea; prevention and ueatment of C difficile diananoea 20-22
Preventionof Vavelle~sdiarrhoea 23

Bacteria commonly found in yoghurt are rarely isolated
from the lower parts of the intestinal tract4.
Present slate of tim a r t - m c a ~ d u i strains
There ate only a few well-documented probiofic
strains available at present that are applied to dairy
products; the most important ones are listed in Table 1.
However, it should be noted that some pmbiotic strains
such as ,~,ccharomyces boulordii are not used in dairy
products or other foods at present.Documenled examples
of the beneficial effects of probiofics include their use
in the treatment of vosious types of dian4mea, the allevi-
ation of the g a s t r o i n t e s ~ sidv effects of antibiotic
treaunent, the alleviation of lactose intolerance, the relief
of comfipafion, and the general balancing and stabiliz-
ation of intesl~d integrity. More recent claims for some
strains include immune-enhancing and vaccine-adjuwmt
effects as well as an ability to lower serum cholesterol
levels to affectcancer-related parameters. The immune-
enhancing effects have been reported in several studies
for two l, actobacillus strains, l~tobacillus acidophilus
LC1 and Laclobacillus GG. l, actobacillus sb'afin GG
(ATCC 53103) has just recently been identified as Lacto-
baci//us rkomnosus, but most published studies wm the
name l, actobacillus GG. Both strains appear to have
that may enhance natural immunity and also
appear to act ~ immurm-~v~mrs.
Studies on the treatment of acute infant diarrhoea
have been successfully canied out with l, actob~illus
C-G, and verified in ~veral publicationsm..3.-~.Similarly,
a good study on the treatment of mtavims dimhoea
using B/fu/obacter/um b/fu/urn preparations has been
published recentlf 9.
Traveller's diarrhoea has been successfully prevented
in a study using a probiotic mixtme (L ocidophilus, B.
bi.fufum, Streptococcus thermophilus and Loctobacillus
bulgaricus)"~. $. boulordii p ~ o n s have also been
successfully used for the prophylaxis of Waveller's
dian'hoea by Auslfiem tourists. A recent review on trav-
eller's diantmea covem most probiodc, as well as other,
studies~.
It has been shown that isolated strains of Lac-
tobacillus and Bifutobacwrium influence the metabolic
activity of the resident micmflora. The consumption
of L. oc/doph/lus of human origin was originally shown
to C~L.~¢a significant decline in the faecal activflies of
b a c t e ~ ~-~ucoronidase, azorcd_,,ms¢ and niuoreduc-
tase~. Later, Ma~.au et a/.'s fed human volunteers a
fermented dairy product containing human strains of
both L. oc/doph/lus and B. b/fu/um, and demonslrated
the survival of both slrains, and a significant zeducfion
in faecal niu'o~luctme activity but no change in either
p - g l u c o ~ d a s e or ~mxluctmm activity, l~lrosa et
a/.t~ found a significant reduction in all three enzyme
activities in elderly subjects after supplementation with
Lactobacillus gassed for 12 days, but not after sup-
plementafion with S. thermophilus and L bulgaricus,
conflmfmg the findings reported earner for these two
yoghurt strain~. Infozmafion on faecal enzyme activities
is increasing constantly, and several probiofic prep-
arafions are known to decrease such activities. Re-
po~dly, some p~bimic preparmions also decrease the
mutagenicity of the intestinal contents, thus altering
intestinal metabolism in a beneficial rammer~. Further
studies are needed to verify the health effects of the
altermion of intestinal metabolism.

242 Trends in Food Science & Technology July 1995 [Vol. 6]

 Similarly, effects on cholesterol levels remain to be at acidic pH values~ s . Av,e n g ~ to isolate 'stable' hac-
verified by further studies, as many confounding factors ~ial s~ains by the seqeeae~ ~ of eeUs e m
have contributed to the effects observed in different remained viable a0x'r ~ incubetion were unsuc-
population groups. In most studies, lactobac~ and cessfuP2. Almost all of the acid4ekram 'aged" mains
bifidobacteria have been repofted to deconjugate bile isolated were non-lactic-acid-geducing bacteria. The
salts, producing free bile acids, which are more inhibi- loss of the acid-producing ability of aged cells may have
tory to susceptible bacteria than the deconjugated form2s. involved genotypic changes, as the aged cells could not
It has also been reported that Lactobacillus acidophilus readily revert to the acid-godecing yonng phenot3q~.
NCFM has an appreciable ability to assimilate choles- To obtain acid-producing and acid-t~nmt lactic acid
terol. Few studies are available on the effects of pro- bacteria likely to survive in femlented foods, selectioa
biotic bacteria on faecal bile acids or neutral sterols. procedures should be performed in growing culea~.
More well-defined clinical work is needed before any Such a selection system has been described n~:eatly~.
firm claims, especially in case of the cholesterol-lower- The selection process involves the addition of fresh mb-
ing effects, can be made. strates into a growing lactic acid bacteria culture when
the pH of the culture drops below a Ineset value owing
s u b i ~ of prot~ic ~ c t n in ~t~t products to fermentation resulting in the production of lactic
Only decades ago, cultured milk was consumed acids from lactose.
mainly for its taste, texture and improved shelf life
compared with milk. In recent years, the probiotic Substrateenhancementof
properties of cultured milk have become desirable by Tha prima~ mechanismof pH maintemncein lactic
consumers. Because the probiotic properties of desirable acid bacteria at med~mely low pH values (-pH5.0) is
bacteria are largely dependent on their ability to remain dependent upon the expulsion of protons from the cyto-
viable and to colonize the surface of human intestinal plasm by acid-dependent adenosine ~ at
cells, sufficient numbers of viable bacteria must be the expense of ATP (adenosine triph~,phate)~ . Thus,
present at the time of consumption. A concentration the inclusion of a slow-metabofizing energy source,
of I × 10s colony-forming units per g or ml of the final such as argiulne and fructose, in non-growing cultmes
product has been suggested as the 'therapeutic mini- provides energy (ATP), which enables cells to remain
mum'29. The consumption of a viable cell number of viable for longer in culture at low pH vahte~. At lower
1 × 10~-1 × 109 cells per day is necessary in order pH values (<4.0), a different mechanism is involved,
for any beneficial effects in humans to develop~3°~L which involves the consmnptinn of intraceilul~ H ÷ ions
Most pmbiotic bacteria, for example L acidophilus and through the degration of L-malic a c i ~ ~ , and the diver-
B. b/fu/um, show a short stationary growth phase, which sion of pyruvate to other ucumfl products, such as ace-
is followed by rapid losses in cell viability, even in cold toin~. Thus, the inclusion of malic acid and the pymvate
storages2. This short shelf life represents a logistic prob- somr~, for example citrate~ , contribute to pH homeo-
lem for both manufacturers and retailers, and a technical stasis and the stability of lactic acid bacteria. Malic
challenge for researchers. and citric acids are already included in some probiotic
Of the various probiotic bacteria, L casei and L products to enhance their fruity taste, and ate generally
plantarum appear to have longer shelf fifes than accepted by consumers.
L acidophilus, L reuteri and Bifutobacterium spp. in Other researchers found that the inclusion of reducing
cultured milk. However, the reason for the difference in agents, such as thioglycollate, cysteine of dithiothteitol,
their stability in cultured milk is not entirely clear. The conferred greater stability on the glycolytic activity of
ability to tolerate lower internal pH values is a mech- non-growing lactobacilli~. However, some of the afore-
anism that allows L. plantarum to survive in culture at mentioned chemicals have yet to be approved as food-
lower pH values33. Further investigations are necessary grade antioxidants. Because of their oxygen-excluding
to elucidate the mechanisms that impart various degrees protective effect, the use of aPlgopriate packaging ma-
of acidostability to the lactic acid bacteria. Such teriais becomes a practical alternative for maintaining
research should lead to target-specific genetic improve- the stability of fermented products. The desirable pack-
ments to the probiotic bacteria. aging materials need to be non-permeable to oxygen and
preferably non-penetrable by fight.
sek~0n of~ u e gob~,Ucbact~ Some carbohydrates that cannot be digested by
The viability of f~eeze-dried bacteria and bacteria humans were found to promote the growth of selected
in non-fermented products, such as sweet acidophllus indigenous prohiotic bacteria, especially bifidobucl~a,
milk, could ha maintained for many months3°~u. How- in the intestinal trace ~. Of these indigestible of slow-
ever, the preferred form of prohiotic bacteria for con- absorbing carbohydrates, most polyols me considered
sumers is in fermented foods and beverages, such as 'safe' of 'no health hazards ate fo~oseen' for them, for
cultured milk, because they have both a better taste and example sofbitol, mannitol~ , lactitol and xyfitol~, and
texture. A typical cultured milk is acidic (pH 3.5-4.5) they are commercially available. Oligosacch~-ides and
and contains 0.5-1.5% (w/v) lactic acid. Lactate at these lactulose, however, have not been evaluated in extensive
c o n c e n ~ i o n s is not inhibitory to the microbial culture, toxicological studies, but it is assumed that they do not
but the 14" ions cause a decrease in the viability of cells pose great health hazards as they are either naturally

Trendsin FoodScience& TechnolosyJuly 1995 [Vol. 6] 243

 occurring or made from natural components. Lactolose
has been used as a therapeutic agent for many years,
and the oligosuccheuride galactosyl lactose is presently
being added to connnercial infant formulae in Japan, to
promote the growth of bilidobacteria in the guts of
infants.
The fulure 12
Future research on probiotic bacteria will probably 13
still centre on the selection of new and more-specific
slrains of lactic acid and other bacteria to promote the
wellbeing of the host. It may well be that different
regions of the gastrointestinal tract require different pro-
biotic bacteria; furthermore, it may be necessary to se-
lect disease-specific mains. This is ~ y true with
diseases such as rotavims diarrhoea and gastritis caused
by Helicobacter pylori. Today, most probiotic strains
are used in dairy products. Present and furore studies
indicate that new probiotic functional foods will include
infant formulae, baby foods, fermented fruit juices, fer-
mented soy products and cereal-based woducts (such as
fermented oatmeal) without milk and also disease-
specific clinical foods containivg viable pmbiotlc bacteria.
All these woducts will be requiged to have good stabil-
ity and in many cases an extensive shelf life. For such
development to occur, it is important that the require-
ments for good clinical studies are followed even more
carefully. Such requirements, listed in Box 2, are s'umlm"
to those t~lulred for any pharmaceutical studies. With
carefully controlled studies on clearly defined selected
strains the future will provide targeted probiotic bacteria
for specific diseases and their pzevantlon. In addition,
the effects of mixtures of different p ~ o i o t l c slrains
should be carefully studied.
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Conference Report
The 4th Food Choice Conference was the fourth in a se-
ries of conferences, all devoted to food choice. The 1st
Food Choice Conference was held in Brussels, in 1992,
as a satellite meeting of a psychology congress; the
2rid and 3rd Food Choice Conferences were held in
Australia and the USA, and were attached to nutrition
and ingestive behavior congresses, respectively. Thus
this was the first of the food choice conferences to be
held on its own, without any other congress being held
at the same time.
The founding father of these conferences, which appear
to be a great success, is David Booth from the School
of Psychology of the University of Birmingham, UK.
Booth, who is one of the editors of the food science
journal Appetite, noticed that food choice issues were
discussed at a number of congresses, but never in their
own fight. The food choice conferences now attract
scientists from a wide variety of disciplines, such as
physiology, nutrition, marketing, psychology and soci-
ology. Although most of the 200 attendants were from
Europe, a substantial proportion were from the USA.
The field of food choice was revealed as an exciting
area with many challenging issues for research. Several
symposia were held at the conference; however, owing
to space limitations, this report deals only with a limited
array of the subject matters that were raised at the
meeting.
Comumer concert, about food safety
It is believed that consumer concerns about food
safety do ~ t edways concur with the ideas that scientists
have about hazards related to foods. Whereas consumers
may think of artificial food additives and chemical
residues as being significant hazards, scientists are
mainly concerned about the generalcomposition of the
diet, such as the high fat content. In a nationwide survey
carried out in the UK, C. Fife-Schaw and G. Rowe,
*Heldin Birminsham,UK, 24-26Ap~11995
Ceet de Ca'ad is at the I ~ of Human Nutr~on, Wageningen
Ag~cultural University, PO Box 8129, 6700 EV Wagen/nsen, The
(fax:+31-8370-83347;e-maihkees.~.veed.wau.n|).
Trends in Food Science & Technology July 1995 INol. 6]
47 Salminen,$., I~mos,p. andFonden,R.0SO3)m ta:~c Ac/d
B,~m~/af , a ~ - n , S.endyenV,~ht, &, ed~, m 2~-306,
MarcelOet~
WHO 0 982) Tox/co~ f - n / m ~ of'Ca~ F ~ M d ~ tWHO
FoodAdd~veSeries),Wodd Hee~ OqlamzaemGemn,
49 FASEB/~GOS(1973)Eva/uat/onofSod~/and~as
Food~n~d/e~ (Re~ 9-10, NTB Pe7.21.-~'1and~3),
~ of ~n~'ican Sodetim~m"E ~ :.,,e,~ m o ~ , Bee~esda,
MD, USA
4th FoodChoice
Conference*
Ceesde Graaf
Univershy of Surrey, UK, investigated whether percep-
tions nf food-related risks ceuld be slmcam~ according to
some underlying dimensions. The results nf their study
showed that perceptions of food-related hazards could
be intexwated aecording to fla'ee dimensiom. One im-
portant dimension reflects the comrollabi~ of the risk.
People thought that they have little coatroi over things
such as food irradiation or the use of genetk engineer-
ing in the production of foods, whereas batter-known
risks such as relatively high fat and sugar intakes we,re
thonght to be under their own control. A second import-
ant dimension concerning perceptions in food-related
risks is their perceived seriousness. Food-related risks
such as bovine spongifmm ¢mcephalolmthy (BSE; mad-
cow disease) are viewed as very serious by the con-
sumer. The third dimension in the perception of food-
related risks deals with the naturalness versus the
artificiality of the risk. R is obvious that hazards of
microbiological origin rate low on this dimemion,
whereas risks related to artificial additives rate highly.
These results are congruent with those found earlier by
P. Sparks and R. Sbepeni, Institute of Food R ~ h ,
Reading, UK.
In relation to this issue, it was noted that the results of
these studies may depend on whether the researchers
generated the risk characteristics or whether the charac-
teristics were elicited from the consumers themselves.
Sparks and M. Raats. Inslimte of Food Research,
Reading, UK, demonstrat~ thet the dimensions in food-
related risks were similar in both cases. In addition, they
showed that attitudes towards hazards varied with dif-
ferences in social and environmental attitudes.
Although moiw people have the ~ that they
have good control over their fat intake, and think that
m~s, LkevterSciem:eLid0924.2244~,~$09.S0 245