Review

Received: 21 December 2010 Revised: 4 February 2011 Accepted: 4 February 2011 Published online in Wiley Online Library: 28 March 2011

(wileyonlinelibrary.com) DOI 10.1002/jsfa.4367

Probiotics and prebiotics perspectives

and challenges
Ivonne Figueroa-Gonzalez, Guillermo Quijano, Gerardo Ramrez
and Alma Cruz-Guerrero

Abstract
Owing to their health benefits, probiotics and prebiotics are nowadays widely used in yogurts and fermented milks, which
are leader products of functional foods worldwide. The world market for functional foods has grown rapidly in the last three
decades, with an estimated size in 2003 of ca US$33 billion, while the European market estimation exceeded US$2 billion in
the same year. However, the production of probiotics and prebiotics at industrial scale faces several challenges, including the
search for economical and abundant raw materials for prebiotic production, the low-cost production of probiotics and the
improvement of probiotic viability after storage or during the manufacturing process of the functional food. In this review,
functional foods based on probiotics and prebiotics are introduced as a key biotechnological field with tremendous potential
for innovation. A concise state of the art addressing the fundamentals and challenges for the development of new probiotic-
and prebiotic-based foods is presented, the niches for future research being clearly identified and discussed.

c 2011 Society of Chemical Industry

Keywords: functional food; probiotics; prebiotics; synbiotics

INTRODUCTION
Nowadays, the strong relationship between diet and health is
well accepted. Although the primary role of diet is to provide
nutrients to fulfil metabolic requirements, the use of foods to
improve health and the state of wellbeing is an idea increasingly
accepted by society in the last three decades.1 3 The change
in the way of conceiving foods has led to the introduction of
the concept of functional foods. Besides exhibiting an adequate
nutritional value, a food can be considered as functional if it
beneficially affects one or more target functions in the body
in a way that is relevant to either the state of wellbeing and
health or the reduction of the risk of a disease.4 Functional
foods as a marketing term was initiated in Japan in the late
1980s. However, the world market has grown rapidly, with an
estimated size in 2003 of ca US$33 billion, while the European
market estimation exceeded US$2 billion in the same year.5 The
functionality of functional foods is based on bioactive components,
often contained naturally in the product but usually requiring
the addition of a specific ingredient in order to optimise the
beneficial properties.1 Today, functional food ingredients include
probiotics, prebiotics, vitamins and minerals, which are used in
fermented milks and yogurts, sports drinks, baby foods, sugar-free
confectionery and chewing gum.6,7 Probiotics and prebiotics are
fundamental ingredients of fermented milks and yogurts, which
account for the most important fraction of the overall market
for functional foods. Therefore most of the available research on
functional foods has focused on probiotics and prebiotics.5,6,8,9
This article constitutes a state-of-the-art review of probiotics and
prebiotics from the fundamentals to the concept of synbiotics and
challenges for the development of new probiotic- and prebiotic-
based foods. Finally, the key research niches in the field are
PROBIOTICS
Several definitions of probiotics can be found in the literature.
Hence, a decade ago, probiotics were considered as those viable
micro-organisms that exhibit a beneficial effect on the health of
the host by improving its intestinal microbial balance.10 However,
a more recent and comprehensive definition is provided by Fric:11
probiotics are non-pathogenic micro-organisms, mostly of human
origin, which confer a health benefit on the host and enable to
prevent or improve some diseases when administered in adequate
amounts. The initial notion of probiotic micro-organisms can be
traced to a century ago when the Nobel Laureate Ilya Metchnikoff
noticed that the long healthy life of Bulgarian peasants resulted
from the consumption of fermented milk products.2 Metchnikoff
suggested that, when consumed, the fermenting bacilli positively
influenced the microbiota of the gut, since these micro-organisms
were theoretically able to be established in the intestinal tract,
decreasing the pathogenic bacterial population.12 Nevertheless,
later investigations revealed that the micro-organisms involved
in milk fermentation are not able to be established in the
gut.13,14 Therefore the original idea of Metchnikoff regarding the
mechanism by which milk-fermenting micro-organisms provide
health benefits to the host was fundamentally wrong.
Nowadays, the most widely used probiotics include lactobacilli,
bifidobacteria and some non-pathogenic strains. These probiotic
Correspondence to: Guillermo Quijano, Departamento de Biotecnologa,
Universidad Autonoma Metropolitana-Iztapalapa, Av. San Rafael Atlixco No.
186 Col. Vicentina, CP 09340, Distrito Federal, Mexico.
E-mail: ggovantes@gmail.com
Departamento de Biotecnologa, Universidad Autonoma Metropolitana-
1341

Iztapalapa, Av. San Rafael Atlixco No. 186 Col. Vicentina, CP 09340, Distrito
identified and discussed. Federal, Mexico

J Sci Food Agric 2011; 91: 13411348 www.soci.org

c 2011 Society of Chemical Industry
 www.soci.org I Figueroa-Gonzalez et al.

Table 1. Key probiotic micro-organisms reported in the literature

Micro-organism Effect on human health Reference

Lactic acid bacteria Lactobacillus rhamnosus GG Reduces the intestinal permeability defects caused by exposure 10

to cows milk or rotavirus infection. May shorten the course of

rotavirus infection causing diarrhoea, travellers diarrhoea and
antibiotic-associated diarrhoea
Lactobacillus casei Reduces the severity and duration of diarrhoea. Stimulates the 10,17,18

immune system of the gut, alleviates the symptoms of Crohns

Lactobacillus casei Shirota
Lactobacillus acidophilus
Lactobacillus johnsonii
disease and possesses strong antimicrobial properties
18,19
Prevents diarrhoea caused by viruses or bacteria. Has the
strongest human health efficacy with respect to management
of lactose malabsorption, rotaviral diarrhoea,
antibiotic-associated diarrhoea and Clostridium difficile
diarrhoea. Has a preventive effect on the recurrence rate of
superficial bladder cancer after surgery
17,18,20
Secretes lactic acid that lowers the pH of the intestinal content
and helps to inhibit the development of invasive pathogens
such as Salmonella spp. or strains of Escherichia coli. Increases
antibody responses and seroconversion rates. Lowers serum
cholesterol levels
May reduce density of Helicobacter pylori and inflammation as 18

well as gastritis activity

Lactobacillus plantarum Produces short-chain fatty acids that inhibit the generation of 18

carcinogenic products by reducing enzyme activities

10
Bifidobacteria Bifidobacterium breve Activates the humoral immune system by augmenting
anti-rotavirus IgA production or anti-influenza virus
Bifidobacterium bifidum May successfully compete for space and nutrients against 17,20

pathogenic or putrefactive bacteria. Reduces the incidence of

diarrhoea and increases antibody responses and
seroconversion rates
Bifidobacterium infantis Prevents diarrhoea and constipation 11

Bifidobacterium animalis Normalises the intestinal motility of obstipated subjects. Reduces 2,19

the risk of acute diarrhoea in children and adults

Yeasts Saccharomyces cerevisiae Boulardii Prevents travellers diarrhoea and the development of colitis and 11,21

enterocolitis of pathogenic origin. Reduces the risk and

duration of antibiotic-associated diarrhoea

cultures include Lactobacillus rhamnosus GG, Saccharomyces PREBIOTICS

cerevisiae Boulardii, Lactobacillus casei Shirota and Bifidobacterium
animalis, which are by far the most studied probiotics with
proven human health efficacy.2,7,11,15 A brief summary of key
probiotics reported in the literature and their effect on health
is provided in Table 1. Most probiotics belong to the genera
Lactobacillus and Bifidobacterium, while S. cerevisiae Boulardii is
the only yeast with proven probiotic characteristics. According
to Saier and Mansour,16 probiotics possess three mechanisms of
promoting human health: (i) providing end-products of anaerobic
fermentation of carbohydrates such as organic acids that can
be absorbed by the host, these end-products being able to
influence human mood, energy level and even cognitive abilities;
(ii) successfully competing with pathogens; (iii) stimulating host
immune responses by producing specific polysaccharides.
It is important to note that health benefits provided by probiotics
are strain-specific and not species- or genus-specific. Therefore no
probiotic strain will provide all proposed benefits, not even strains
of the same species. Likewise, not all strains of the same species
will be effective against defined health conditions.18 Nonetheless,
it is observed that a common benefit of most probiotics listed in
Table 1 is their control of diarrhoea, this being associated with the
competitive exclusion of pathogens.2 Thus the ability to survive
through the upper gastrointestinal tract and colonise the intestine
1342
The concept of prebiotics has also evolved significantly over time. A
prebiotic was first defined as a non-digestible food ingredient that
beneficially affects the host by selectively stimulating the growth
and/or activity of one or a limited number of bacteria in the colon,
thus improving host health.22 However, recent literature does not
restrict the colon as the only action site and defines a prebiotic as
a selectively fermented ingredient that allows specific changes in
the composition and/or activity of the gastrointestinal microbiota
that confer benefits upon host wellbeing and health.23 The key
concept associated with both these definitions is that prebiotics
have a selective effect on the microbiota that results in improved
health of the host.24 An ingredient must fulfil three fundamental
aspects in order to be considered as a prebiotic: (i) resistance
to digestion; (ii) fermentation by the large intestinal microbiota;
(iii) a selective effect on the microbiota that has associated health-
promoting effects.25 Therefore, in order to be effective, prebiotics
need to reach the large bowel with their chemical and structural
properties essentially unchanged to further selectively stimulate
the microbiota. Most prebiotics are short-chain carbohydrates with
a degree of polymerisation of 2 or more, which are not susceptible
to digestion by pancreatic and brush border enzymes.24 A brief
summary of the main candidates for prebiotic status is provided
in Table 2. Prebiotic oligosaccharides may be manufactured by
extraction from plant materials, microbial/enzymatic synthesis

is a key criterion to consider a micro-organism as probiotic.16,17 and enzymatic hydrolysis of polysaccharides. Various prebiotics

wileyonlinelibrary.com/jsfa

c 2011 Society of Chemical Industry J Sci Food Agric 2011; 91: 13411348
Probiotics and prebiotics perspectives and challenges www.soci.org

Table 2. Main candidates for prebiotic status reported in the literature

Carbohydrate Chemical structure Method of manufacture Degree of polymerisation Reference

Inulin (2-1)-Fructans Extraction from chicory root and Agave 265 25,27,31,32

tequilana
Fructo-oligosaccharides (2-1)-Fructans Transfructosylation from sucrose or 210 25,33

hydrolysis of chicory inulin

Galacto-oligosaccharides Galactose oligomers and Produced from lactose by 25 25,34 36

some -galactosidase
glucose/lactose/galactose
units
Soya-oligosaccharides Mixture of raffinose and Extracted from soya bean whey 34 25,33

Xylo-oligosaccharides
Isomalto-oligosaccharides
stachyose
25,37
(14)-Linked xylose Enzymatic hydrolysis of xylan. Enzyme 24
treatments of native lignocellulosic
materials. Hydrolytic degradation of
xylan by steam, water or dilute
solutions of mineral acids
(14)-Glucose and Microbial or enzymatic 28 25,38

branched (16)-glucose transgalactosylation of maltose.

Enzymatic synthesis from sucrose
Pyrodextrins Mixture of Pyrolysis of potato or maize starch Various 25

glucose-containing
oligosaccharides

are produced at industrial scale and are widely available in the
market.26 As a matter of fact, prebiotics have been reported as
cheap to manufacture.27 This perception might be inspired by
the fact that prebiotics are obtained from relatively low-cost raw
materials.28 However, most prebiotics are obtained by means of
enzymatic processes and thus synthesis and further purification is
not necessarily a cheap process. In fact, several investigations on
novel aqueous/organic media for enzymatic synthesis have been
explored in order to improve the yield of specific prebiotics.29,30
Aiming to develop a quantitative tool to compare the effect
of prebiotics on the growth of probiotic bacteria, Palframan
et al.39 proposed an equation in which a prebiotic index (PI) is
introduced. The PI score is based on the selective stimulation
of probiotic bacteria (bifidobacteria and lactobacilli) over other
micro-organisms (bacteroides and clostridia) due to addition of a
prebiotic according to the following equation:
PI = Bif/Total + Lac/Total Bac/Total Clos/Total
and in vivo experiments, it is not fair to compare their PI scores.
Recently, Roberfroid23 proposed a new PI for in vivo experiments
based on the generation of new probiotic bacteria per gram of
prebiotic ingested:
PI = [(CFU/gfaeces )final time (CFU/gfaeces )initial time ]/Prebiotic dose
(2)
where CFU denotes colony-forming units and the Prebiotic dose
refers to the number of grams of prebiotic ingested per day by the
host. The advantage of calculating the PI by means of Eqn (2) is that
the effect on probiotic growth is based on prebiotic concentration.
Therefore a fair comparative analysis can be performed even for
studies with different prebiotic doses. However, the author did
not find a correlation between the PI values and the amount of
prebiotic ingested. This might have been due to the fact that
the ingested prebiotic is not necessarily the amount of prebiotic
that reaches and is consumed in the large intestine by probiotic
(1) bacteria.

where Bif, Lac, Bac, Clos and Total represent the concentration
ratios of bifidobacteria, lactobacilli, bacteroides, clostridia and total
bacteria between sampling time and at inoculation respectively.
A negative PI score indicates that the prebiotic addition did not
selectively stimulate the growth of probiotic bacteria, while a
positive PI value indicates that the tested prebiotic was able
to specifically stimulate probiotic bacteria under the working
conditions. Unfortunately, this equation has two important
drawbacks: (i) the amount of prebiotic consumed is not considered
and thus a fair comparative analysis among several prebiotics is
restricted to studies with similar prebiotic uptake; (ii) the PI defined
in Eqn (1) is useful only for experiments in vitro where the total
concentration of probiotics and pathogens is well known. In the
case of in vivo experiments the total bacterial population as well
as the prebiotic uptake in the host is fundamentally unknown.
Hence the increase in probiotic population due to prebiotic
consumption is only inferred through the content of probiotics in
SYNBIOTICS
It is nowadays common to find probiotic foods (e.g. yogurts and
fermented milks) with added prebiotics. Such a combination, in
which the concentration of prebiotic is typically below 10 g kg1 ,
is known as a synbiotic.19 Synbiotics have been considered as
a very promising area for the development of new functional
foods. In fact, the potential synergy between probiotics and
prebiotics has been considered as obvious.40 Synbiotics have
also been defined as mixtures of probiotics and prebiotics
that beneficially affect the host by improving the survival
and implantation of live microbial dietary supplements in its
gastrointestinal tract.41 In vitro simulations of the gastrointestinal
tract constitute a common experimental framework to investigate
synbiotics (Table 3). One of the main benefits of synbiotics is the
increased persistence of probiotics in the gastrointestinal tract.42
Therefore the reported in vitro experiments are mainly focused
1343

faecal matter.23 Based on these key differences between in vitro on elucidating (i) if probiotics might metabolise the prebiotic

J Sci Food Agric 2011; 91: 13411348

c 2011 Society of Chemical Industry wileyonlinelibrary.com/jsfa
 www.soci.org
Table 3. Examples of synbiotics reported in the literature
Synbiotic
Probiotic Prebiotic
Lactobacillus casei strain Shirota Oligomate 55
Bifidobacterium longum Oligofructose
Bifidobacterium lactis Lafti B94 Resistant starch
Bifidobacterium breve strain Yakult Galacto-oligosaccharides
Lactobacillus gasseri Inulin and unspecified oligosaccharides
Lactobacillus acidophilus ATCC 4962 Manitol, fructo-oligosaccharides and inulin
Lactobacillus sakei JCM Fructo-oligosaccharides and trehalose
Lactobacillus plantarum and L. acidophilus Xylo- and fructo-oligosaccharides
component under simulated conditions of the gastrointestinal
tract, (ii) if the prebiotic component may be metabolised by
pathogenic bacteria and (iii) if the synbiotic is able to inhibit
or decrease the growth of pathogenic bacteria with respect to
a control without synbiotic. Despite the relevant information
obtained from in vitro tests, authors working with this kind of
experiment clearly recognise that further in vivo trials are necessary
to corroborate the effectiveness of synbiotics.40 In this context,
de Vrese and Schrezenmeir19 highlighted that synbiotics might
improve the survival and implantation of probiotics only in the
case of a true synergistic mutual reinforcement between the
prebiotic and probiotic components. Lately, it is not clear whether
synbiotics will result in elevated levels of probiotic bacteria
owing to the prebiotic component.42 For instance, a synbiotic
based on Bifidobacterium longum and oligofructose stimulated
the proliferation of faecal bifidobacteria by 1.4 log(CFU g1 )
in Wistar rats, while an increase in faecal bifidobacteria of 1.6
log(CFU g1 ) was obtained by administering only oligofructose.43
Moreover, Morelli et al.44 assessed the effect of a synbiotic based
on Lactobacillus gasseri (strain B21090), inulin and unspecified
oligosaccharides on the growth of probiotic bacteria. Although
these authors found that the synbiotic was effective in increasing
the population of faecal probiotic bacteria in human volunteers,
the stimulation of probiotic bacteria by synbiotic administration
was not compared with the stimulation obtained with the prebiotic
alone. Consequently, a comparative analysis of the potential
superior effectiveness of the synbiotic over the prebiotic by
itself cannot be made. Therefore convincing evidence for the
real potential of synbiotics should come from more in vivo studies
comparing the performance of synbiotics with respect to their
components.
CHALLENGES FOR NEW PROBIOTIC-
AND PREBIOTIC-BASED FOODS
Owing to their perceived health benefits, probiotics and prebiotics
are now widely added to yogurts and fermented milks.5,6,50
However, the production of probiotics and prebiotics at industrial
scale faces several challenges, including (i) the use of novel
techniques and economical sources for prebiotic production,
(ii) the low-cost production of probiotics and (iii) the improvement
of probiotic viability after storage, during the manufacturing
process of the functional food and during transit through the
1344
stomach. These key challenges are addressed below.
wileyonlinelibrary.com/jsfa

c 2011 Society of Chemical Industry
I Figueroa-Gonzalez et al.
Type of experiment Reference
45
In vitro
In vivo (rats) 43
In vitro 40
In vivo (mice) 46
In vivo (humans) 44
47
In vitro
In vitro 48
In vitro 49
Novel techniques and economical sources for prebiotic
production
Most oligosaccharides with prebiotic status are normally obtained
by enzymatic treatment of cheap raw materials such as sucrose,
lactose and plant derivatives (Table 2). The amount and nature of
the oligosaccharides formed depend upon several factors such as
the enzyme source, the concentration and nature of the substrate
and the reaction conditions.32 Nevertheless, the current processes
to obtain oligosaccharides have very low yields, thus increasing
the production cost.29,51 Panesar et al.34 noted that the yield
of oligosaccharides can be increased by decreasing the water
content in the reaction medium. In this regard, del Val and Otero29
studied the enzymatic synthesis of galacto-oligosaccharides from
lactose in aqueous/polyethylene glycol media. They found that a
decrease in water content increased not only the overall yield of
galacto-oligosaccharides but also the selective production of 6 -
galactosyl lactose. In addition, Cruz-Guerrero et al.30 investigated
the synthesis of galacto-oligosaccharides from lactose using a
hyperthermophilic -glycosidase in aqueous/acetone medium.
They observed that the oligosaccharide yields were strongly
affected by the water activity (aw ), the maximum yield being
obtained at an aw of 0.57 and an initial lactose concentration
of 8 g L1 . Therefore the enzymatic production of prebiotics in
organic media represents an interesting research field to improve
the yield of prebiotics over traditional synthesis in aqueous media.
The reviewed literature already focuses on such research to
establish optimum aqueous/organic volume ratios or aw /substrate
concentration ratios when the synthesis medium consists mainly
of an organic solvent.
The production of non-dairy oligosaccharides is another
interesting option for low-cost prebiotic production at industrial
scale. In recent years, cereals have been investigated regarding
their potential as sources of prebiotics. Charalampopoulos
et al.52 concluded that at least two types of naturally occurring
oligosaccharides can be found in cereal grains, namely galactosyl
derivatives of sucrose, stachyose and raffinose and fructosyl
derivatives of sucrose. On the other hand, cereal grains are rich
in dietary fibre, which encompasses a heterogeneous range of
complex polysaccharides that are not substantially digested in the
small intestine and pass through to the colon. Thus dietary fibre
constitutes a potential source of prebiotics from cereals.53 The
dietary fibre content in grains such as rice, rye and corn can reach
values as high as 150190 g kg1 dry matter (Table 4). -Glucans,
xylans, xylo-oligosaccharides and arabinoxylans are the main
constituents of dietary fibre. Crittenden et al.53 reported that many
Bifidobacterium species and Lactobacillus brevis were able to grow
J Sci Food Agric 2011; 91: 13411348
Probiotics and prebiotics perspectives and challenges
Table 4. Total dietary fibre content in several cereal grains
Cereal Total dietary fibre (g kg1 dry matter) Reference
Wheat 121
Rye 161
Barley 100
Sorghum 107
Rice 192
Corn 150
Oat 140
at high yields using xylo-oligosaccharides. They also observed that
B. longum strains were able to grow using arabinoxylan as the
sole carbon source. In addition, hydrolysates of -glucans have
been reported to stimulate the growth of several Bifidobacterium
strains and L. rhamnosus GG.52 Recently, Moura et al.54 selectively
produced oligosaccharides with degrees of polymerisation of 34
and 56 by means of autohydrolysis techniques using corn cobs
as the raw substrate. Moreover, the obtained oligosaccharides
promoted the growth of both Bifidobacterium and Lactobacillus
species to a similar extent to commercial xylo-oligosaccharides.
Starch is a substantial component of many cereal grains such as
corn, wheat, rice and oat (785, 650, 820 and 636 g kg1 dry matter
respectively).56 Although resistant starch is found naturally in
cereal grains, it is frequently destroyed when subjected to modern
food processes. Industrial methods for manufacturing resistant
starch include partial acid hydrolysis, hydrothermal treatment,
heating, retrogradation, extrusion cooking, chemical modification
and repolymerisation.52 As in the case of dietary fibre, studies
in animals and human volunteers have shown that a fraction
of the total starch is not digested in the small intestine and
passes through to the colon. This fraction is termed resistant
starch and has been reported as prebiotic.57 Crittenden et al.40
reported that many Bifidobacterium strains such as B. adolescentis,
B. bifidum, B. breve, B. infantis, B. lactis and B. longum were able
to hydrolyse resistant starch. However, only B. lactis (strain Lafti
B94) maintained its viability under conditions similar to those
encountered during passage through the gastrointestinal tract.
The authors finally proposed a synbiotic yogurt based on resistant
starch and B. lactis. On the other hand, Topping et al.57 concluded
that evidence regarding resistant starch as prebiotic is still limited,
since much of the experimental work has been done in animals,
the available investigations in humans being of relatively short
duration. However, the authors recognised that there is a great
deal of promise, and further research is necessary to assess the
potential of resistant starch as a prebiotic in humans.
Low-cost production of probiotics
Low-cost production of concentrated cultures of probiotics is a
key challenge to satisfy the increasing demand for probiotics in
the market.58 Probiotic micro-organisms are normally difficult to
grow as they lack biosynthetic capacity for most vitamins and
amino acids, so culture media for probiotic production must
be supplemented with both amino acids and vitamins.59,60 The
reduction of production costs at industrial scale might start with
the use of some agro-industrial residual effluents as cultivation
media for probiotics. In this regard, effluents from fruit and
vegetable processing usually contain high amounts of proteins,
carbohydrates, lipids and vitamins. In the case of effluents from
J Sci Food Agric 2011; 91: 13411348

c 2011 Society of Chemical Industry
www.soci.org
cereal processing, mono-, di- and oligosaccharides can also be
found.61 A clear example of this kind of effluent is the wastewater
from industrial corn processing. According to Gutierrez-Uribe
et al.,62 the wastewater from the manufacturing process of
55
commercial fresh masa and dry masa flour (also called nejayote)
55
contains a high load of corn solids. Approximately 50% of these
52
solids are suspended and contain about 64, 20 and 1.4% non-starch
52
polysaccharides, starch and protein respectively. The remaining
55
50% consist of proteins, sugars, vitamins and phytochemicals
52
rich in phenolics and carotenoids. Other studies revealed that
52
nejayote contains polysaccharides of arabinose, xylose, glucose,
galactose and D-glucuronic acid and approximately 23% crude
fibre (whose dietary fibre fraction may stimulate the growth
of probiotics).52,53,63 65 Therefore agro-industrial effluents such
as the wastewater from cereal processing may be an abundant
source of prebiotic compounds. Efforts must be made to determine
the feasibility of using these effluents either partially or totally in
culture media formulation.
Improving viability of probiotics
The quality of probiotic micro-organisms relies greatly on their
viability, which is a fundamental fulfilment to reach and colonise
the human large intestine.66,67 Probiotics must retain their viability
during three critical stages: (i) storage; (ii) manufacturing process
of the functional food; (iii) transit through the stomach and small
intestine. Therefore the viability of probiotics is an issue of vital
importance from both an economic and a technological viewpoint.
In this regard, Ratnakomala and Widyastuti68 observed that storage
of a probiotic by freezing at 40 C decreased its viability from
1.8 1015 to 1.6 1010 CFU mL1 , while a freeze-drying pre-
treatment followed by storage at 4 C decreased its viability from
8.9 1014 to 2.4 109 CFU mL1 . Likewise, Lahtinen et al.66 noted
that probiotics might become dormant during storage. They
observed that Bifidobacterium strains can be completely dormant
after storage at 4 C for 25 days in an oat-in-water suspension. Such
results clearly indicate that storage may significantly decrease the
viability of probiotics. Freeze-drying in food matrices has been
proposed to improve probiotic viability during storage and transit
through the stomach and small intestine. For instance, Saarela
et al.69 reported that freeze-drying in a matrix of low-fat milk or
fruit juice significantly improved the viability of a Bifidobacterium
strain. They observed that viability in the juice matrix was higher
than in low-fat milk during storage, while tolerance to acid and
bile was better in cells with the low-fat milk matrix after storage.
The authors concluded that, when choosing a food matrix as
cryoprotectant for a probiotic, viability should not be the only
parameter to be evaluated but also the ability to resist adverse
conditions after storage. Therefore research on low-cost food
matrices as cryoprotectants to improve probiotic viability after
storage is a relevant field that is worthy of further investigation.
On the other hand, microencapsulation is a promising technique
recently studied to improve the viability of probiotic micro-
organisms. Microencapsulation can be defined as a technology
of packaging solids, liquids or gases in miniature, sealed capsules
that can release their contents at controlled rates under the
influence of specific conditions.70 It has been reported that
microencapsulation protects probiotics during storage and during
the manufacturing process of a determined functional food,
improved viability against heat stress and acidic conditions
also being observed.70,71 Microencapsulation techniques used
so far to protect probiotic micro-organisms include spray-drying,
1345
spray-congealing, fluidised bed coating/air suspension, extrusion,
wileyonlinelibrary.com/jsfa
 www.soci.org I Figueroa-Gonzalez et al.

for prebiotic synthesis; (ii) the reduction of probiotic production

Table 5. Some examples of coating materials used in probiotic
microencapsulation costs; (iii) the enhancement of probiotic viability during storage,
during the manufacturing process of the functional food and dur-
Coating material Probiotic Reference ing transit through the upper gastrointestinal tract. The strategies
71 to overcome such challenges must come from a multidisciplinary
Alginate/resistant starch Lactobacillus
acidophilus and approach. The alternatives herein reviewed include the following.
Bifidobacterium Production of prebiotics in two-liquid-phase systems by adding
spp.
71
an organic solvent that increases the yield of prebiotics. Such
Whey protein Lactobacillus
rhamnosus
multiphase systems may also increase the yield of prebiotics
Whey protein Bifidobacterium 70 with specific degrees of polymerisation.
longum Production of oligosaccharides from cheap and non-dairy raw
Cellulose acetate phthalate Bifidobacterium 70 materials, where cereals seem to be the most interesting
pseudolongum option as they have naturally occurring oligosaccharides such
Alginate/sodium lauryl Lactobacillus 70 as galactosyl derivatives of sucrose, stachyose and raffinose as
sulfate delbrueckii well as fructosyl derivatives of sucrose. Moreover, cereal grains
Carrageenan/locust bean Lactobacillus casei 70
are rich in dietary fibre and starch, the latter being able to be
gum modified to yield resistant starch.
70
Alginate Lactobacillus Probiotic production from agro-industrial effluents, where
acidophilus
72
corn-processing effluents appear as a very interesting option
Canola vegetable Bifidobacterium
oil/caseinate/fructo- infantis
based on their high concentrations of non-starch polysaccha-
oligosaccharides/resistant rides, starch, protein, sugars, vitamins, phytochemicals rich in
starch (or glucose syrup) phenolics and carotenoids as well as polysaccharides of ara-
Alginate/gellan gum/fructo- Bifidobacterium 48 binose, xylose, glucose, galactose and D-glucuronic acid. Thus
oligosaccharides/peptides bifidum this kind of effluent may be totally or partially used for low-cost
culture media formulation.
The viability of probiotics may be improved by using cheap
food matrices or by means of microencapsulation techniques.
coacervation/phase separation and electrostatic methods. A
Hence research on matrices such as milk or fruit juice could
detailed description of these microencapsulation technologies
potentially improve the viability of probiotics, as they act
is given by Anal and Singh.70 The common material of all
as cryoprotectants during the freeze-drying process. On the
microencapsulation techniques is the coating material in which
other hand, microencapsulation was identified as a promising
the probiotic is encapsulated. Common coating materials include
technique that protects probiotics during storage, during the
water-soluble/insoluble polymers, waxes, fatty acids and lipids.
manufacturing process of a determined functional food and
Table 5 shows some of the most common probiotics and coating
during transit through the upper gastrointestinal tract.
materials used in probiotic microencapsulation. Prebiotics such
as fructo-oligosaccharides or resistant starch may be part of
the coating formulation. For instance, Chen et al.48 included REFERENCES
fructo-oligosaccharides in the coating formulation for B. bifidum 1 Korhonen H, Technology options for new nutritional concepts. Int J
microencapsulation. They observed that a formulation including Dairy Technol 55:7988 (2002).
2 Vasiljevic T and Shah NP, Probiotics from Metchnikoff to bioactives.
prebiotics, gellan gum and peptides significantly improved
Int Dairy J 18:714728 (2008).
probiotic viability with respect to microcapsules made from 3 Wiseman A and Woods L, Addition of designer enhancers to functional
alginate as the sole coating material. Likewise, Crittenden et al.72 foods: need also for redesigned biocatalysts in fail-clean strategies
included prebiotics in the coating formulation for B. infantis of bioprocessing? J Chem Technol Biotechnol 76:10381040 (2001).
microencapsulation. They found that a combination of canola 4 Roberfroid M, Functional food concept and its application to
prebiotics. Dig Liver Dis 34:S105S110 (2002).
vegetable oil, caseinate, fructo-oligosaccharides and resistant 5 Menrad K, Market and marketing of functional food in Europe. J Food
starch (or glucose syrup) improved the viability of the prebiotics Eng 56:181188 (2003).
by ten orders of magnitude relative to free cells after storage 6 Stanton C, Gardiner G, Meehan H, Collins K, Fitzgerald G, Lynch PB,
for 5 weeks in an open container at 25 C and 50% relative et al, Market potential for probiotics. Am J Clin Nutr 73:476S483S
(2001).
humidity. Therefore the investigation of new coating materials and 7 Heller KJ, Probiotic bacteria in fermented foods: product
microencapsulation techniques also constitutes a key research characteristics and starter organisms. Am J Clin Nutr 72:374S379S
niche. Available investigations seem to direct efforts towards (2001).
research on cheap and abundant coating materials as well as 8 Senok AC, Ismaeel AY and Botta GA, Probiotics facts and myths. Clin
Microbiol Infect 11:958966 (2005).
optimisation of the coating material formulation (e.g. optimum 9 Prado FC, Parada JL, Pandey A and Soccol CR, Trends in non-dairy
prebiotic percentages). probiotic beverages. Food Res Int 41:111123 (2008).
10 Kaur IP, Chopra K and Saini A, Probiotics: potential pharmaceutical
applications. Eur J Pharmaceut Sci 15:19 (2002).
11 Fric P, Probiotics and prebiotics renaissance of a therapeutic
CONCLUSIONS principle. Cent Eur J Med 2:237270 (2007).
In brief, functional foods based on probiotics and prebiotics consti- 12 Mercenier A, Pavan S and Pot B, Probiotics as biotherapeutic agents:
tute a very important biotechnological field with tremendous po- present knowledge and future prospects. Curr Pharmaceut Des
8:99110 (2002).
tential for innovation. We herein identified three main challenges 13 Caplice E and Fitzgerald GF, Food fermentations: role of micro-
for the industrial production of probiotic- and prebiotic-based organisms in food production and preservation. Int J Food Microbiol
1346

functional foods: (i) the improvement of production techniques 50:131149 (1999).

wileyonlinelibrary.com/jsfa

c 2011 Society of Chemical Industry J Sci Food Agric 2011; 91: 13411348
Probiotics and prebiotics perspectives and challenges
14 Fuller R and Gibson GR, Probiotics and prebiotics: microflora manage-
ment for improved gut health. Clin Micriobiol Infect 4:477480
(1998).
15 Gomes A and Malcata MX, Bifidobacterium spp. and Lactobacillus acid-
ophilus: biological, biochemical, technological and therapeu-
tical properties relevant for use as probiotics. Trends Food Sci
Technol 10:139157 (1999).
16 Saier MH and Mansour NM, Probiotics and prebiotics in human. J Mol
Microbiol Biotechnol 10:2225 (2005).
17 Itsaranuwat P, Shal-haddad K and Robinson PK, The potential
therapeutic benefits of consuming health-promoting fermented
dairy products: a brief update. Int J Dairy Technol 56:203210 (2003).
18 Shah NP, Functional cultures and health benefits. Int Dairy J
17:12621277 (2007).
19 De Vrese M and Schrezenmeir J, Probiotics, prebiotics, and synbiotics.
Adv Biochem Eng Biotechnol 111:166 (2008).
20 Gill H and Prasad J, Bioactive components of milk: probiotics, immuno-
modulation, and health benefits, in Advances in Experimental
Medicine and Biology, ed. by Bosze Z. Springer, New York, NY,
pp. 423454 (2008).
21 Priebe MG, Vonk RJ, Sun X, He T, Harmsen HJM and Welling GW, The
physiology of colonic metabolism. Possibilities for interventions
with pre- and probiotics. Eur J Nutr 41:1/21/10 (2002).
22 Gibson GR and Roberfroid MB, Dietary modulation of the human
colonic microbiota: introducing the concept of prebiotics. J Nutr
125:14011412 (1995).
23 Roberfroid M, Prebiotics: the concept revisited. J Nutr 137:830S837S
(2007).
24 Steed H and Macfarlane S, Mechanisms of prebiotic impact on
health, in Prebiotics and Probiotics Science and Technology, ed.
by Charalampopoulos D and Rastall RA. Springer, New York, NY,
pp. 135161 (2009).
25 Macfarlane S, Macfarlane GT and Cummings JH, Review article:
prebiotics in the gastrointestinal tract. Alim Pharmacol Therapeut
24:701714 (2006).
26 Grajek W, Olejnik A and Sip A, Probiotics, prebiotics and antioxidants
as functional foods. Acta Biochim Pol 52:665671 (2005).
27 Venter CS, Prebiotics: an update. J Family Ecol Conservat Sci 35:1725
(2007).
28 Rastall RA, Gibson GR, Gill HS, Guarner F, Klaenhammer TR, Pot B, et al,
Modulation of the microbial ecology of the human colon by
probiotics, prebiotics and synbiotics to enhance human health:
an overview of enabling science and potential applications. FEMS
Microbiol Ecol 52:145152 (2005).
29 Del Val MI and Otero C, Biphasic aqueous media containing
polyethylene glycol for the enzymatic synthesis of oligosaccharides
from lactose. Enzym Microb Technol 33:118126 (2003).
30 Cruz-Guerrero AE, Gomez-Ruiz L, Viniegra-Gonzalez G, Barzana E and
Garca-Garibay M, Influence of water activity in the synthesis
of galactooligosaccharides produced by a hyperthermophilic -
glycosidase in an organic medium. Biotechnol Bioeng 93:11231129
(2006).
31 Kelly G, Inulin-type prebiotics a review: part 1. Altern Med Rev
13:315329 (2008).
32 Iniguez-Covarrubias G, Diaz-Teres R, Sanjuan-Duenas R, Anzaldo-
Hernandez J and Rowell RM, Utilization of by-products from the
tequila industry. Part 2: Potential value of Agave tequilana Weber
azul leaves. Bioresour Technol 77:101108 (2001).
33 Delzenne NM, Oligosaccharides: state of the art. Proc Nutr Soc
62:177182 (2003).
34 Panesar PS, Panesar R, Singh RS, Kennedy JF and Kumar H, Microbial
production, immobilization and applications of -D-galactosidase.
J Chem Technol Biotechnol 81:530543 (2006).
35 Boon MA, vant Riet K and Janssen AEM, Enzymatic synthesis of
oligosaccharides: product removal during a kinetically controlled
reaction. Biotechnol Bioeng 70:411420 (2000).
36 Olano A and Corzo N, Lactulose as food ingredient. J Sci Food Agric
89:19871990 (2009).
37 Vazquez MJ, Alonso JL, Domnguez H and Parajo JC, Xylooligosac-
charides manufacture and applications. Trends Food Sci Technol
11:387393 (2000).
38 Goulas AK, Cooper JM, Grandison AS and Rastall RA, Synthesis of
isomaltooligosaccharides and oligodextrans in a recycle membrane
bioreactor by the combined use of dextransucrase and dextranase.
Biotechnol Bioeng 88:778787 (2004).
J Sci Food Agric 2011; 91: 13411348

c 2011 Society of Chemical Industry
www.soci.org
39 Palframan R, Gibson GR and Rastall RA, Development of a quantitative
tool for the comparison of the prebiotic effect of dietary
oligosaccharides. Lett Appl Microbiol 37:281284 (2003).
40 Crittenden RG, Morris LF, Harvey ML, Tran LR, Mirchell HL and
Playne MJ, Selection of a Bifidobacterium strain to complement
resistant starch in a synbiotic yoghurt. J Appl Microbiol 90:268278
(2001).
41 Andersson H, Asp NG, Bruce A, Roos S, Wadstrom T and Wold AE,
Health effects of probiotics and prebiotics: a literature review
on human studies. Scand J Nutr 45:5875 (2001).
42 Rastall RA and Maitin V, Prebiotics and synbiotics: towards the next
generation. Curr Opin Biotechnol 13:490496 (2002).
43 Bielecka M, Biedrzycka E and Majkowska A, Selection of probiotics
and prebiotics for synbiotics and confirmation of their in vivo
effectiveness. Food Res Int 35:125131 (2002).
44 Morelli L, Zonenschain D, Callegari ML, Grossi E, Maisano F and
Fusillo M, Assessment of a new synbiotic preparation in healthy
volunteers: survival, persistence of probiotic strains and its effect
on the indigenous flora. Nutr J 2:111 (2003).
45 Figueroa-Gonzalez I, Hernandez-Sanchez H, Rodrguez-Serrano G,
Gomez-Ruiz L, Garca-Garibay M and Cruz-Guerrero A, Antimicro-
bial effect of Lactobacillus casei strain Shirota co-cultivated with
Escherichia coli UAM0403. Rev Mex Ing Quim 9:1116 (2010).
46 Asahara T, Nomoto K, Shimizu K, Watanuki M and Tanaka R, Increased
resistance of mice to Salmonella enterica serovar Typhimurium
infection by synbiotic administration of bifidobacteria and
transgalactosylated oligosaccharides. J Appl Microbiol 91:985996
(2001).
47 Liong MT and Shah NP, Optimization of cholesterol removal, growth
and fermentation patterns of Lactobacillus acidophilus ATCC 4962
in the presence of mannitol, fructo-oligosaccharide and inulin:
a response surface methodology approach. J Appl Microbiol
98:11151126 (2005).
48 Chen MJ, Chen KN and Kuo YT, Optimal thermotolerance of
Bifodobacterium bifidum in gellan-alginate microparticles.
Biotechnol Bioeng 98:411419 (2007).
49 Pan X, Wu T, Zhang L, Cai L and Song Z, Influence of oligosaccharides
on the growth and tolerance capacity of lactobacilli to simulated
stress environment. Lett Appl Microbiol 48:362367 (2009).
50 Ozer D, Akin S and Ozer B, Effect of inulin and lactulose on survival of
Lactobacillus acidophilus LA-5 and Bifidobacterium bifidum BB-02 in
Acidophilus-Bifidus yoghurt. Food Sci Technol Int 11:1924 (2005).
51 Crittenden RG and Playne MJ, Production, properties and applications
of food-grade oligosaccharides. Trends Food Sci Technol 7:353361
(1996).
52 Charalampopoulos D, Wang R, Padiella SS and Webb C, Application of
cereals and cereal components in functional food: a review. Int J
Food Microbiol 79:131141 (2002).
53 Crittenden R, Karppinen S, Ojanen S, Tenkanen M, Fagerstrom R,
Matto J, et al, In vitro fermentation of cereal dietary fibre
carbohydrates by probiotic and intestinal bacteria. J Sci Food Agric
82:781789 (2002).
54 Moura P, Carvalheiro F, Grio F, Loureiro MC and Esteves MP,
In vitro fermentation of xylo-oligosaccharides from corn cobs
autohydrolysis by Bifidobacterium and Lactobacilllus strain.
LWT Food Sci Technol 40:963972 (2007).
55 Nyman M, Siljestrom M, Pedersen B, Bach-Knudsen KE, Asp NG,
Johansson CG, et al, Dietary fiber content and composition in six
cereals at different extraction rates. Cereal Chem 61:1419 (1984).
56 Snow P and ODea K, Factors affecting the rate of hydrolysis of starch
in food. Am J Clin Nutr 34:27212727 (1981).
57 Topping DL, Fukushima M and Bird AR, Resistant starch as a prebiotic
and synbiotic; state of the art. Proc Nutr Soc 62:171176 (2003).
58 Mattila-Sandholm T, Myllarinen P, Crittenden R, Mogensen G,
Fonden R and Saarela M, Technological challenges for future
probiotic foods. Int Dairy J 12:173182 (2002).
59 Altermann E, Russell WM, Azcarate-Peril MA, Barrangou R, Buck BL,
McAuliffe O, et al, Complete genome sequence of the probiotic
lactic acid bacterium Lactobacillus acidophilus NCFM. Proc Natl Acad
Sci USA 102:39063912 (2005).
60 Ballongue J, Bifidobacteria and probiotic action, in Lactic Acid Bacteria,
ed. by Salminen S and Wright A. Marcel Dekker, New York, NY,
pp. 519587 (1998).
61 Federici F, Fava F, Kalogerakis N and Mantzavinos D, Valorisation
1347
of agro-industrial by-products, effluents and waste: concept,
wileyonlinelibrary.com/jsfa
 www.soci.org
opportunities and the case of olive mill wastewaters. J Chem Technol
Biotechnol 84:895900 (2009).
62 Gutierrez-Uribe JA, Rojas-Garca C, Garca-Lara S and Serna-Saldivar O,
Phytochemical analysis of waste water (nejayote) obtained after
lime-cooking of different types of maize kernels processed into
masa for tortillas. J Cereal Sci 52:410416 (2010).
63 Nino-Medina G, Carvajal-Millan E, Rascon-Chu A, Marquez-Escalante J,
Santana-Rodriguez V and Salmeron-Zamora J, Maize processing
waste water arabinoxylans: gelling capability and cross-linking
content. Food Chem 115:12861290 (2009).
64 Martnez-Bustos F, Martnez-Flores HE, Sanmartn-Martnez E,
Sanchez-Sinencio F, Chang YK, Barrera-Arellano D, et al, Effect of the
components of maize on the quality of masa and tortillas during the
traditional nixtamalisation process. J Sci Food Agric 81:14451462
(2001).
65 Rosentrater KA, A review of corn masa processing residues: generation,
properties, and potential utilization. Waste Manag 26:284292
(2006).
66 Lahtinen SJ, Gueimonde M, Ouwehand AC, Reinikainen JP and
Salminen SJ, Probiotic bacteria may become dormant during
storage. Appl Environ Microbiol 71:16621663 (2005).
1348
wileyonlinelibrary.com/jsfa

c 2011 Society of Chemical Industry
I Figueroa-Gonzalez et al.
67 Tuomola E, Crittenden R, Playne M, Isolauri E and Salminen S,
Quality assurance criteria for probiotic bacteria. Am J Clin Nutr
73:393S398S (2001).
68 Ratnakomala S and Widyastuti Y, Storage of probiotic Leuconostoc
citreum TSD-10 by freezing and freeze drying. Ann Bogor 7:4348
(2000).
69 Saarela M, Virkajarvi I, Alakomi HL, Sigvart-Mattila P and Matto J,
Stability and functionality of freeze-dried probiotic Bifidobacterium
cell during storage in juice milk. Int Dairy J 16:14771482 (2006).
70 Anal AK and Singh H, Recent advances in microencapsulation of
probiotics for industrial applications and targeted delivery. Trends
Food Sci Technol 18:240251 (2007).
71 Vidhyalakshmi R, Bhakyaraj R and Subhasree RS, Encapsulation the
future of probiotics a review. Adv Biol Res 3:96103 (2009).
72 Crittenden R, Weerakkody R, Sanguansri L and Augustin MA,
Synbiotic microcapsules that enhance microbial viability during
nonrefrigerated storage and gastrointestinal transit. Appl Environ
Microbiol 72:22802282 (2006).
J Sci Food Agric 2011; 91: 13411348