Yeast and sourdough

1. Raw materials

1.3. Yeast and sourdough

1.3.1 Yeast

What is yeast ?

Yeast is a micro-organism that is grown in yeast factories. Its scientific name is Saccharomyces cerevisiae. In this word you can see the Latin word "saccharo", which means sweet or sugar and the word
"myces", which means, "mould". Baker's yeast is a unicellular mould that reproduces through a process, which is known as "budding".

We know that bakers use it to make the dough "rise"; without it, our bread would be like flat, hard cakes. In the days when people made their own bread, they would go to a brewer and get a jug of
brewer's yeast. It was fluid and yellow. Nowadays, yeast is made commercially on a large scale. The yeast you buy at your market, the yellow lumps done up in paper, has been compressed for convenient
handling.

Commercial yeast obviously is widely available. The baking characteristics of the yeast are the most significant feature of commercial yeast. Different types of yeast are available for different types of
bread-making processes (f.i. osmotolerant yeast suitable for doughs that contain a lot of sugar). Properties such as the shelf life of the yeast are controlled by various fermentation parameters. For
instance, a high percentage of budding cells at the time of the harvest will result in a product with a shorter shelf life.

Yeast is a plant, according to the biologists, and is capable of reproducing itself. A piece of yeast consists of minute cells, with walls composed of cellulose, and an interior of living matter called protoplasm.
You can feed it with a solution of sugar to make it grow, or it can be "killed" by starvation or heat. The ancients did not use yeast as we know it today; they prepared a leaven or 'barm' (which has the same
action) from ground millet kneaded with "must" out of wine-tubs.

Wheat bran was also used, kneaded with a three-days-old must, dried in the sun, and then made into little cakes. When required for making bread, the cakes were soaked in water, and then boiled with the
finest flour, after which the whole was mixed in with the meal. Another old method for making barm was to prepare cakes of barley meal and water; these were baked on a hot hearth, or else in an earthen
dish upon hot ashes and left until they turned reddish-brown. Afterwards, the cakes were kept shut up in a vessel until they turned quite sour. When wanted for leaven, they were first steeped in water.
Two hundred twenty five grams of this was enough to make a quantity of bread of about 6,5 kg to rise.

General description of yeasts

Yeasts are not, like moulds, a clearly defined taxonomic group. They belong to the same club as the moulds. Yeasts are said to be unicellular fungi in contrast to moulds, which are multi-cellular.

Yeasts can be found in nature mainly in habitats, where substrates rich in sugars are present i.e. flower nectar, on all kind of fruits etc. They can cause spoilage of certain foodstuffs such as fruit juices,
syrups, honey, meat, wine, beer, yoghurt etc. They also are used in a number of food preparation processes such as bread, beer, wine, vinegar, cheese and for the production of enzymes. Although there are
pathogenic yeasts (such as Candida albicans) they are rarely found in foodstuffs.

As with many biological materials yeast activity is temperature sensitive. Changes in dough temperature at the end of mixing of during the proof will have drastic effects on the CO2 production. The lower
the dough temperature the slower the yeast will produce gas and the longer will the proof time be in order to obtain a given dough volume before the dough can enter the oven. Higher dough temperatures
give an increased gas production but require better process control.

Morphological characteristics.

Most of the time yeasts are ovoid but they also can be round, lemon or pear shaped, cylindrical or even be triangular. The size can vary quite a bit but in general yeasts are bigger then bacteria. The
diameter varies between 1 and 5 µ, while the length might vary between 5 and 30 µ. Yeasts do not have flagellae.

Fermentation

The primary function of yeast is to produce carbon dioxide gas, which expands the dough during proof and the early stages of baking (oven spring). Yeast prefers slightly acid conditions to work best. A pH
ranging from 4,5 to 6,0 gives the best results. Bread doughs are generally in the region of pH 5,5 so in normal bread-making the effect of the pH is not a particular consideration. However' some
ingredients used in the bakery, such as mould inhibitors in some bread improvers, lower the pH of the dough and do have a retarding effect on yeast fermentation. This effect is usually taken into account
when deciding on the yeast level to use in a given recipe. There are other ingredients which can retard yeast activity i.e. spices or raisins.

Carbon dioxide cannot form a gas bubble on its own it requires a "nucleating site" (i.e. somewhere it can gather to form a bubble). In fizzy drinks microscopic projections on the side of the bottle provide
those sites, which is why when you release the pressure as you open the bottle you see "streams" of gas running from the sides. In bread dough the nucleating sites are provided by the nitrogen gas
bubbles, trapped in the dough during mixing. The yeast has used up the oxygen from the air.

During proof stages the carbon dioxide goes into solution until the solution is saturated and then any more, which is generated, makes its way into the nitrogen gas bubbles, which grow in size, and the
dough expands. The more yeast and the warmer the temperature the faster the expansion - we get oven spring because the maximum gassing rate occurs at 40 - 45°C.

In bulk fermentation stages we also get dough expansion from carbon dioxide generation but most of that is lost when the dough is knocked back and divided, so the yeast has to start over again.

Yeast also contributes to dough maturity/development. Though its role is minor compared to improvers in no-time doughs, it is more significant in bulk fermentation where the enzymes, especially the
proteolytic ones (they modify the gluten proteins), play a significant role.

To sum up, then, the dough is aerated by the action of the yeast. The little cells we mentioned ferment the dough, and produce tiny bubbles of gas inside it. As a result, the dough gets fatter and bigger,
and rises, of course. Thus when the dough is baked, you have a 'bold' loaf, light and airy; when you cut it you can see all the tiny holes formed by the gas, so that it looks like a sponge.

Yeasts constitute a group of single-celled (unicellular) fungi, a few species of which are commonly used to leaven bread and ferment alcoholic beverages. Most yeasts belong to the division Ascomycota. A
few yeasts, such as Candida albicans, can cause infection in humans. More than one thousand species of yeasts have been described. The most commonly used yeast is Saccharomyces cerevisiae, which was
domesticated for wine, bread, and beer production thousands of years ago.

Yeast physiology can be either obligate aerobic or facultative anaerobic. In the presence of oxygen, yeast will multiply through a mechanism known as budding. In the yeast factory this mechanism is used
to grow yeast into a commercially available product called compressed yeast. Compressed yeast contains about 75 % water of which about 1/5th sits between the yeast cells and not in the yeast cells.
Depending on the moisture content and the size of the individual yeast cells, 1 gram of compressed yeast contains 8 to 13 billion yeast cells. In these circumstances, the yeast does not have access to
nutrients and it must survive on the reserves stored in the cell. The yeast cell will slowly but surely use these reserves. As a result the cells grow weaker and weaker i.e. the yeast will lose its gassing power.
This process is called "autolysis". The speed at which this process progresses, depends on the proteolytic activity of the cell. Proteases are enzymes that split proteins. The proteolytic activity depends on
the temperature. The higher the temperature the faster the breakdown of proteins will take place. Therefore yeast should be stored in the fridge.

In the absence of oxygen, fermentative yeasts produce their energy by converting sugars into carbon dioxide and ethanol (alcohol). In brewing, the ethanol is bottled, while in baking the carbon dioxide
raises the bread, and the ethanol evaporates. There is no known obligate anaerobic yeast.

An example with glucose as the substrate is

C6H12O6 -> 2C2H5OH + 2CO2

Yeasts for leavening bread may be produced commercially or caught from the environment. Many types of yeast can be isolated from sugar-rich environmental samples. Some good examples include fruits
and berries (such as grapes, apples or peaches), exudates from plants (such as plant saps or cacti). Some yeasts are found in association with insects.

The use of potatoes, water from potato boiling, eggs, or sugar in bread dough accelerates the growth of yeasts. Salt and fats such as butter slow yeast growth down. A common medium used for the
cultivation of yeasts is called potato dextrose agar (PDA) or potato dextrose broth. Potato extract is made by autoclaving cut-up potatoes with water for 5 to 10 minutes and then decanting off the broth.
Dextrose (glucose) is then added (10 g/L) and the medium is sterilized by autoclaving.

Yeast fermentations comprise the oldest and largest application of microbial technology. They are used for beer and wine fermentations and bread production.

There are many different kinds of yeast, which all belong to the Saccharomyces family. They don't have all the same capacity to ferment sugars. Certain species only ferment one type of sugar, others two
different types of sugar. In the case of bread we need a yeast which can transform glucose and fructose into CO 2 and alcohol.

Yeast budding

Saccharomyces cerevisiae is also known as budding or baker's yeast. It is used as a model organism by biologists studying genetics and molecular biology (in particular the cell cycle) because it is easy to
culture but as a eukaryote, it shares the complex internal cell structure of plants and animals.

Yeasts can reproduce asexually through budding or sexually through the formation of ascospores. A new organism is formed by the protrusion of part of another organism. When yeast buds, one cell
becomes two cells. This is an example of reproduction. This is very common in plants, but may be found in animal organisms, such as the hydra, as well. Usually, the protrusion stays attached to the primary
organism for a while, before becoming free. The new organism is naturally genetically identical to the primary one (a clone).

During asexual reproduction a new bud grows out of the parent yeast when the condition is right, then, after the bud reaches an adult size, it separates from the parent yeast. Under low nutrient
conditions yeasts that are capable of sexual reproduction will form ascospores. Yeasts that are not capable of going through the full sexual cycle are classified in the genus Candida.

The cell cycle, or cell division cycle, is the cycle of events in an eukaryotic cell from one cell division to the next. It consists of interphase, mitosis, and usually cell division. The cell cycle is regulated by
cyclins and cyclin-dependent kinases. Leland H. Hartwell, R. Timothy Hunt and Paul M. Nurse won the 2001 Nobel Prize in Physiology or Medicine for their discovery of these central molecules in the
regulation of the cell cycle.

Schema of the cell cycle. I = interphase, M = mitosis.

The duration of mitosis in relation to the other phases has been exaggerated in this diagram

The phases of the cell cycle are:

The G0 phase is a period in the cell cycle where cells exist in a quiescent state.
The G1 phase is the first growth phase.
S phase, during which the DNA is replicated (S stands for the synthesis of DNA)
G2 phase is the second growth phase, also the preparation phase for the cell.
M phase or mitosis and cytokinesis, the actual division of the cell into two daughter cells.

A surveillance system, so-called "checkpoints", monitors the cell for DNA damage and failure to perform critical processes. Checkpoints can block progression through the phases of the cell cycle if certain
conditions are not met. For instance, there is a checkpoint, which monitors DNA replication and keeps cells from proceeding to mitosis before DNA replication is completed. Similarly, the spindle checkpoint
blocks the transition from metaphase to anaphase within mitosis if not all chromosomes are attached to the mitotic spindle.

Saccharomyces cerevisiae was the first eukaryotic genome that was completely sequenced. The yeast genome database is highly annotated and remains a very important tool for developing basic knowledge
about the function and organization of eukaryotic cell genetics and physiology. The Munich Information Centre maintains another important S. cerevisiae database for Protein Sequences

Physiological characteristics

Most yeasts grow will in a liquid substrate or a substrate that is rich in water. Although yeasts require more free water than moulds, some of them grown pretty well in substrates containing a lot of
dissolved substances such as sugar or salt. Where for "normal" yeasts the minimal aw-value for growth lies between 0,88 and 0,94 (for instance 0,94 for yeast used in breweries, 0,905 for baker's yeast),
osmophilic yeasts will also grow at a w values of 0,62 (for instance Zygosaccharomyces rouxii) to 0,65. This kind of values are found in syrups, jams, honey etc. Some osmophilic yeasts will not grow at a w
values higher than 0,78. Obviously the optimum a w value as well as the a w range in which yeasts will grow depend also on other factors such as pH, temperature, oxygen, presence inhibitory substances etc.

Yeasts also have an optimum temperature at which they will grow. Normal values range between 25 and 30°C. The maximum temperatures yeasts can withstand are in the 35 – 47°C bracket. Some types
grow at 0°C. Candida curiosa for instance grows between 0°C and 15°C but not any more at 25°C.

An acid environment (pH 4,5 – 6,0) will favour the growth of yeast. Growth in an alkaline environment is feeble. Exceptions are Zygosaccharomyces balii which still grows at a pH value of 1,8 while other
species such as certain Schizosaccharomyces types grow well at a pH > 7.

Types of commercial yeast

Cream yeast is liquid and contains about 16 - 20 % solids and generally speaking no additives. The shelf life is about 2 weeks at 4°C. It needs to be stirred continuously unless it has been stabilised with
hydrocolloids. This type can be pumped which is an advantage for large industrial bakeries.

The second type is compressed yeast which is the most familiar form of yeast. Sold as blocks or as crumbled yeast in 20 kg bags. The solid content is about 29 - 34 % and the shelf life is about 4 weeks at
4°C. However opened bags of crumbled yeast have to be used as soon as possible as they seem to get warmer and warmer even in the fridge. In this way the yeast loses its vitality and its fermentative
power.

Finally also dried yeast is available. Solid level about 92 - 97 % and the shelf life is easily 1 year if packed under vacuum. Dosage level is in general 1/3th of compressed yeast. It is advised to mix the yeast
with some water and leave it for a couple of minutes before adding it to the dough. This type of yeast is particularly suitable in countries with a poor distribution system and/or countries which have a high
temperature and high humidity climate (such as Brazil or the Democratic Republic of Congo)

1.3.2. Sourdough

Introduction

How is it possible that out of four relatively tasteless or even bad tasting raw materials the baker can make such a fantastic product? In a poll, where people were asked to list according to their
preference different smells, 71 % of all respondents put the smell of freshly baked bread on the first spot. The results were the following:

Smell of freshly baked bread 71 %

Freshly cut lawn 60 %

Freshly brewed coffee 51 %

Smell of air at the sea 50 %

Perfume 45 %

Smell of air after snow 42 %

Partner during love making 34 %

Air after rain 33 %

Smell of banknotes 14 %

Smell of suntan oil 12 %

New car 7%

The quality of bread is characterised by its flavour, nutritional value, texture and shelf life. In the baking industry there characteristics are improved by addition of so called "improvers" or enzymes
(which normally are incorporated in the improvers). Alternatively the addition of sourdough influences all aspects of bread quality and thus meets the consumer demand for a reduced use of these
improvers which contain all kind of additives. As sourdough is an intermediate and not an end product, its impact on the bread can only be determined on the basis of the quality of the bread. Biochemical
changes during sourdough fermentation occur in protein en carbohydrate components of the flour. The rate and extent of these changes greatly influence the properties of the sourdough and consequently
the quality of the bread. The effects are associated with the metabolites produced by the lactic acid bacteria and yeast during fermentation, including organic acids, enzymes and CO2.

Definitions

Let's start by giving some definitions because there is quite a lot of confusion when it comes to sourdough. People also use rather carelessly various expressions, saying one thing but meaning another.

Sourdough is made from flour and water, which starts to ferment spontaneously and which is allowed to ferment for a certain time at a certain temperature. Indeed flour contains naturally lactic acid
bacteria, which will develop in the mixture and which will acidify it. Sometimes the baker adds lactic acid bacteria himself. In French this is called a "levain", in Italian or Spanish "madre", in Dutch
"zuurdesem" and the German baker will talk about "Sauerteig".

A sponge is made from flour, water and commercial available yeast. As a sourdough it is kept for a certain time at a certain temperature. In French this is called a "poolish", in Dutch a "zetsel". Italians will
call it "biga" and the Germans use the word "Hefestück".

It seems appropriate to make a list of all terms used in the bakery and to give a definition of the various expressions.

Yeast Unicellular plant, which metabolises simple sugars to alcohol and CO 2. This process is called proofing or fermentation.
Different species of yeast are used in different types of fermentations i.e. baker's yeast in the bakery, beer yeast in the
brewery

Baker's yeast In 1860 Louis Pasteur discovered why bread rises and what the role of the yeast was in this process. He discovered that
a particular type of yeast – Saccharomyces cerevisiae – was particularly apt to be bred industrially.

Natural or wild yeast Numerous types of yeast can be found on plants, cereals, fruits, vegetables etc. They belong often to the family of
Saccharomyces exigus to which belong also the varieties Candida krusei and Candida milleri. Not all natural yeasts are
appropriate for use in the food industry.

Lactic acid bacteria Generic term for a whole series of gram positive bacteria (Enterococcus, Lactococcus, Leuconostoc, Lactobacillus) which
produce mainly or exclusively lactic acid. They are used abundantly in the food industry for the preparation of yoghurt,
cheese, sauerkraut, wine, beer, sourdough etc.

Anstellgut (German) A piece of dough kept from previous productions, which is refreshed at regular intervals by adding fresh flour and water.

barm All natural sourdough made from whole wheat flour or wheat kernels, which begin to ferment spontaneously thanks to the
presence of natural lactic acid bacteria and/or natural yeasts.

biga Leaven made on the basis of commercial yeast. Water, flour and a small amount of commercial yeast are mixed. The
consistency is the same as normal dough. The mixture is allowed to ferment for 24 hours and subsequently added to
(Italian) freshly made dough, to which, if necessary, more yeast is added.

chef A piece of dough kept from previous productions. It is used to make "levain". A "chef" however is normally less solid than
levain

levain Is made from the "chef" to which more flour and water is added in order to get the consistency of normal dough.
Normally it is kept at relatively low temperatures (20 – 23°C) in order to encourage the formation of lactic acid to the
(French) detriment of acetic acid.

desem Natural leaven made from whole wheat flour. Traditionally it is kept cool ( 18°C) and completely covered by flour in order
to favour the development of the lactic acid bacteria and yeasts that are naturally present in the flour.

amish Natural leaven or leaven made on the basis of commercial yeast in which, instead of water, milk is used. Sometimes a small
amount of sugar is added as well.

Anfrischsauer German expression, which indicates the first step in the production process of sourdough

Grundsauer German expression used to indicate the second step in the production of sourdough. This process is described in detail
later on.

Vollsauer German expression used to indicate the third step in the production of sourdough. This process is described in detail
later on.

madre (Spanish or Italian) Natural leaven made with wheat flour that is allowed to ferment spontaneously.

poolish Mixture of water, flour and commercial yeast. In most cases quite a bit of water is used so the final consistency
resembles more a batter than dough.

Preferment Is basically the same as a poolish although certain preferments are made with lactic acid bacteria (used in the production
of crackers for instance).

Sauerteig Leaven obtained by the natural and spontaneous fermentation of flour and water. After complete maturation a piece is
kept aside as "Anstellgut".

Sourdough Leaven obtained by the natural and spontaneous fermentation of flour and water. After complete maturation a piece is
kept aside to start the process all over again

starter (English or Dutch) Can have two meanings

pure culture of lactic acid bacteria freely available from specialised companies
piece of dough which is kept aside and used as basis in a process of continuous refreshment.

sponge Mixture of water, flour and commercial yeast. The consistency is similar to that of normal dough or slightly less solid.

Processing

Although yeast in a compressed, dried or liquid form has become standard raw material for industrial baking worldwide, wheat or rye sourdough remain popular in many countries and in fact is gaining in
popularity. We should never forget that the oldest method of making fermented bread was to use wheat or rye sourdough and the today's yeast is a rather recent invention (about 125 years or so). During
the fermentation of sourdough acetic acid is produced and this created the optimal conditions for the swelling of and baking of the flour, simultaneously preventing the growth of spoilage organisms in the
dough and imparting an excellent flavour to the bread.

Sourdough preparation can be done through many different techniques. However the main objective is to obtain a leavening agent that contains well-adapted resident micro-organisms. They have to produce
sufficient CO2 to leaven the dough, organic acids and other metabolites to provide a bread with good crumb characteristics and pleasant sensory properties. Also a shelf life life extension, in terms of
microbilogical shelf life, can be obtained by the use of sourdough. To produce a standardised acidified dough the ripening method will depend upon:

type and quality of the substrate (important is the ash content of the flour but there is no law that says we have to use wheat flour to make sourdough. As long the substrate is a source of
carbohydrates, it can be used to as substrate. I have seen sourdough made from chickpeas, sorghum or sourdoughs where a part of the flour was replaced by potatoes etc.)
type of lactic acid bacteria used
conditions of time, temperature and consistency with which the sourdough is produced

The production of a good sourdough requires an exact control of the acidity developed during the process. Temperature, time, dough yield (dough firmness) and type and species of microflora will
determine the quality of the sourdough and the bread. Even in a modern industrial bakery, control of these parameters can prove difficult and lead to quality variations in the ripened dough, which,
unfortunately, is only discovered in the baked bread.

Wheat sourdough cultures depend on various species of lactobacilli and yeasts contained in the "mother-doughs" (see definitions above). The contents of these starter doughs are carefully guarded secrets
in may bakeries. Their preparation, control and perpetuation has been carried out with great care and individualism by idealists over generations. This of course is a dangerous situation because if
something goes wrong during the process or the so-called idealist disappears for what ever reason, all the know how might be disappear as well.

The mother-dough or starter culture is made into a stiff dough containing water and flour, stored overnight at 2 - 8°C for up to 8 hours. The degree of acidulation will depend on the storage time and
temperature. Again a word of warning: in this case one allows the natural microflora present in the flour to develop. I'm not sure that is such a good thing. I prefer to add a known species or mixture of
species to add to the mother-dough (combined or not with some yeast), so at least the type and strain of micro-organism is under control.

Traditionally the typical wheat flour sourdough process involves 3 steps prior to the final mixing stage. Normally this would extend over about 18 - 24 hours fermentation to obtain the typical flavour
associated with sourdough breads. In order to produce an excellent sourdough and optimise the conditions necessary for the microorganisms to produce the type and quantity of acids, one can use the
following empirical system:

1. Stage 1: water and wheat flour are mixed together with the starter culture to give a dough yield of 200. The basic definition of dough yield is the amount of dough prepared from 100 parts of flour
(the Germans call this TA or "Teigausbeute"). So in this particular example we mix 100 parts of flour with 100 parts of water. The dough is allowed to ferment 6 hours at 26°C. During this time the
yeast will grow and some acidity will develop.
2. Stage 2: adjust the dough yield to 170 by adding more flour and ferment for 8 hours at a temperature between 24 and 28°C. During this time the dough ripens and develops a considerable amount of
acidity and aroma. To do this, one can add to the dough of stage one 100 parts of flour and 40 parts of water. In total we then have 200 parts of flour and 140 parts of flour i.e. a dough yield of 170.
3. Stage 3: adjust the dough yield to 190 and allow the sourdough to mature during 3 hours at 28 - 32°C. To obtain a dough yield of 190 at this stage, one has to add 100 parts of flour and 130 parts of
water to the dough of stage 2.

After this 17 hours of ripening the pH of the dough will be somewhere between 4,0 and 4,5 again depending on the type of microorganisms and the temperature at which the ripening took place.

Yeast fermentation contributes little to dough acidity in the form of lactic and acetic acid, but the lactic acid bacteria, depending on type and species are capable of converting citric and malic acids in
lactic and acetic acid. This induces the development of aromatic compounds.

The famous San Francisco sourdough bread has been made continuously for over 150 years but the microbiology of the starter cultures were only characterised in the seventies by Dr. Leo Kline. The yeast
identified as Saccharomyces exiguus and the lactic acid bacteria, named by Dr. Kline, Lactobacillus sanfranciscensis coexist in the pH range 3,8 - 4,5. The bacteria utilises maltose as a carbohydrate
source, whereas the yeast does not utilise maltose at all, thus avoiding any competition for the same carbohydrate in the dough. This is a typical example of microbiological cohabitation or symbiosis.

Sourdoughs can be dried and milled into powders. Many commercial products use this technique. The quality and consistency of dried powdered sours depend on the type, species and blend of the
microorganisms and the efficiency of the drying process. Freeze-drying techniques offer the best protection for the microorganisms, since heat damage is reduced to its minimum. However I would advise
bakers to prepare their own sourdough if they want to produce an unique quality. On top of that I'm convinced that the future of the industrial bakery lies in developing specific sourdoughs which impart
specific characteristics to the bread. Possible examples are elimination of emulsifiers, creating pre- or probiotics in the sourdough which will then enhance the nutritional aspects of the breads etc.

Depending on the type of bread many individual methods of sourdough processing have been developed. I always say that there are as many systems as there are bakers and that all systems are good and
acceptable. The clue of the matter is to be able to reproduce the process and that is not always an easy thing to do. By choosing appropriate process parameters the microbial metabolism of the microflora
can be controlled to give a balanced build up of aromas. There is a close relationship between the microbial composition of the sourdough, the formation of metabolic products and the process parameters.

In general a sourdough is ready to be used when the following basic characteristics have been achieved:

the prevalence of a microbial flora different from that of the flour used as a substrate
a stable ratio between lactic acid bacteria and yeasts to the order of 100:1 or more
obligatory heterofermentative lactic acid bacteria have to be predominant over other types of lactic acid bacteria.

Lactic acid bacteria

As said before, the term "lactic acid bacteria" is a collective noun and there are numerous species and subspecies of lactic acid bacteria. Lactobacillus literally means, "milk rod" because the take the shape
of a little rod. They have very complex nutritional requirements for amino acids, peptides, vitamins, salts, fatty acids and fermentable sugars. The requirements are normally specific for each different
kind of lactic acid bacteria. The temperature for optimal growth lies between 30°C and 40°C but they are active between 2°C and 53°C. Most lactic acid bacteria are micro-aerobe to anaerobe but most of
them are aerotolerant. They survive well in acid conditions and the optimal pH lies between 5,5 and 6,2. They remain active at a pH as low as 3,5.

Lactic acid bacteria can be divided into two main families

Obligatory homofermentative lactic acid bacteria : produce exclusively lactic acid out of maltose and glucose in anaerobic circumstances. They use the fructose diphosphate pathway, converting
glucose to yield over 90 % of lactic acid. As they do not produce CO2, yeast has to be added to make the dough rise. The most common homofermentative lactic acid bacteria are Lactobacillus
plantarum, Lactobacillus casei, Lactobacillus farciminis, Lactobacillus alimentarius and Lactobacillus acidophilus.

Heterofermentative lactic acid bacteria (such as Lactobacillus sanfranciscensis) produce not only lactic acid but also ethanol and CO2 using all kind of hexoses as a substrate (pentose phosphate
cycle). In the presence of certain ingredients (such as fructose, oxygen, citrates), not only ethanol will be produced but also acetic acid. As fructose is always present in dough but as it is not
metabolised by lactic acid bacteria, a certain amount of acetic acid is always formed. The most important heterofermentative lactic acid bacteria are L. brevis lindneri, L. fermentum, L.
sanfranciscensis, L. fructovorans and L. buchneri. Of all these types, L. brevis lindneri, L. fermetnum, L. plantarum and L. casei are important for flavour building during dough fermentation.

Both groups will contribute to the flavour profile of the bread. Important is to remember that lactic acid bacteria not necessarily impart a acid taste to the bread. The taste depends on the dominant
group of lactic acid bacteria present in the dough. The total count of lactic acid bacteria in a sourdough is on average between the 2.10 8 and 6.10 9 cells per gram of sourdough.

The most important strains of lactic acid bacteria are :

Lactobacillus is a strain of rod shaped bacteria. They can be found in all kinds of vegetable origin but also in the intestines and the mouth. They are important in the production of yoghurt,
sauerkraut, cottage cheese, beer, wine and obviously sourdough. In yoghurt one will find about 10 million lactobacilli in 1 gram of product.
Streptococci is a strain of sphere shaped bacteria and the name literally means "spheres which appear in strands". They can also be found in all kind of products and are amongst other things
important during the production of sausages.

Whilst the lactic acid bacteria produce acids and flavour compounds, the yeasts will produce carbon dioxide for leavening and the formation of crumb structure and volume. In the initial stage the yeast will
consume the oxygen present in the dough thus creating favourable conditions for the metabolism and reproduction of the facultative anaerobe lactic acid bacteria. Spicher has identified over forty yeast
strains which belong to the following four types: Saccharomyces cerevisiae, Pichia saitoi, Candida krussei and Torulopsis holmii.

Apart from the indigenous flour microflora, the sourdough microflora will depend on the following variables:

initial microflora of the starter culture

the processing parameters

Assuming that the process parameters and dough yield are controlled, the rate of reproduction and metabolism within the sourdough should remain constant over a given period of time. Therefore by
increasing or decreasing the amount of started used, the ripening time can be decreased or increased accordingly. The maturing time of the sourdough depends on the growth curve of the microflora.
Normally the growth curve can be divided into four phases, by plotting the logarithm of the cell count versus time, as follows:

1. Phase 1 is the lag phase or induction phase during which the microorganisms adapt themselves to the substrate and take in the nutrients for reproduction and metabolism. The duration of the
induction phase is inversely proportional to the cell count of the inoculate. The lower the amount of starter used, the longer the induction phase. The average duration of the lag phase is bout 1 hour.
2. Phase 2 is an acceleration phase in which exponential reproduction occurs. During this phase the microorganisms reach their maximum growth potential. The total duration of this phase is about 4
hours.
3. Phase 3: when approximately 4 hours have elapsed the exponential growth lapses into a period of delayed growth in order to reach the so-called stationary phase. At this point a state of equilibrium
exists between newly formed cells and dying cells.
4. Finally, during the 4th phase the lethal phase sets in whereby the availability of nutrients diminishes and the the concentration of metabolites reaches such a level that cell autolysis occurs. This, in
real life, is the moment one has to add again substrate and water to revive the process.

In a sourdough bacteria and yeasts coexist, but each group has different optima regarding temperature, time, pH and consistency of the dough. Therefore the optimal conditions for both bacteria and
yeasts cannot be simultaneously provided. Either bacteria or yeasts predominate and one group can have a negative influence on the other. The optimal temperature for bacterial metabolism and hence the
acid production is between 30 and 40°C but the optimum differs according to the strain of bacteria. The function of the bacteria is to transform the carbohydrates (mainly maltose) and the proteins (or
amino acids to be more precise) of the flour into lactic and acetic acid and other volatile aroma components.

Acid production continues down to a pH of about 4,0. Below this pH the bacteria become inhibited by their own metabolites. The nature of metabolites depend on the type of lactic acid bacteria which all
use different sugars and their metabolism uses different pathways. All sourdough bacteria ferment glucose, maltose and sucrose except L. brevis which does not ferment sucrose. Lactose is also fermented
except by L. delbrueckii and L. leichmanii. L. plantarum and L. brevis also ferment pentoses especially arabinose.

The generation time (that is the time necessary for the bacteria count to double) for lactic acid bacteria ranges from 70 to 200 minutes, depending on the species, temperature and consistency of the
dough. Every micro-organism demands certain optimal conditions for both its reproduction and its metabolism and these factors are important for all sourdough processes. Now lets have a look at these
different process variables and their importance and influence on the sourdough.

The first one is the temperature. The development of the desired micro-organism can be selective to a degree centigrade! Bakers underestimate this aspect and most of them will not believe that 1°C will
make a difference. But an expert in tasting - such as a sommelier in wine tasting - will be able to tell the taste difference. The controllable variables must be used in coordination in order to optimise the
chosen technological process. This is also the reason why different temperatures are used at different stages of the sourdough production. Optimal temperatures for growth and development are:

25 - 27°C for sourdough yeasts: production of CO2 (and thus leavening), production of alcohols, aldehydes and organic acids for flavour and aroma. At the same time they act as antibiotic to wild
gram-negative bacteria
20- 30°C for heterofermentative lactic acid bacteria. They are responsible for the development of an aromatic bread flavour. They produce lactic acid and acetic acid in a ratio of 80:20.
30 - 40°C for homofermentative lactic acid bacteria for the production of lactic acid and to obtain a milder more balanced flavour.

The second process variable is the consistency of the dough i.e. the amount of water present in the dough. Flour water ratio's varying between 1:1 to 1:2 produce a more intensive flavour then sourdoughs
containing less water. Both yeasts and lactic acid bacteria develop better in softer doughs. This is due to the nutrients' solubility and improved distribution during mixing. The homogeneous distribution of
the nutrients in a softer dough results in an acceleration of microbial activity of the homofermentative lactic acid bacteria. This gives the bread a mild harmonious flavour note which is very distinctive.

Of course also the type of substrate will influence the flavour development in the sour. As substrate one can use any carbohydrate source. What I mean is that the substrate does not have to be wheat or
rye flour. It also can be cornflour or potato. However it is common in the bakery industry to use either wheat flour or rye flour. A first point remark is that the assimilative forms of carbon and nitrogen,
trace elements and vitamins are very different in these type of cereals. This important complex of variable nutrient content will have a definite influence on the ripening of the sour. Flour extraction rate
is involved in this equation: lower extraction flours producing less acid build up than the higher ones this being due to differences in enzyme activity. Thus when processing a rye flour of lower extraction
rate a higher refreshment rate is required than in the case of higher extraction flours.

Important is of course the enzymatic activity in the substrate and in the sour. Salt and acid together reduce enzyme solubility. At comparatively low acidity, in the presence of salt, enzyme activity
becomes inhibited.

Finally there is the ripening time. Each type of four requires a definite standing time and good bread quality results from allowing the various sourdough stages, enough time to reach maturity. Standing
times can vary considerably as they depend on the other process variables (dough consistency, temperature, characteristics of the substrate). The average rage is form 2 to 5 hours. But to slow the
process down and to be able to use overnight ripening, one can add more flour, giving a firmer consistency, thus slowing down the activity of the microorganisms.

The following tables give an idea of the results in aroma development with changing process parameters.

volatile component resembles whole wheat white flour

(E,E)-2,4-nonadienal greasy, fried 512 4

phenylacetaldehyde floor wax 256 4

methional boiled potatoes 1024 16

(E,E)-2,4-decadienal greasy, fried 512 16

4-vinyl-2-methoxyphenol cloves 512 16

3-hydroxy-4,5-dimethyl-2-furanon sweat 4096 256

(E)-4,5-exposy-(E)-2-decanal metallic 4096 64

(E,Z)-2,6-nonadienol cucumber 1024 64

vanillin vanilla 1024 64

2,3-methyl-1-butanol sweat 256 16

The numbers in the table indicate how many times the extract can be diluted in order to just still smell or taste the volatile component. It can also be seen that using whole wheat as a substrate will give a
more intense sourdough with regards to flavour and aroma. It also can be seen that in both breads the same volatile compounds are present but that their concentration is totally different.

component dough yield 150 dough yield 300

ethanol 18200 8471

n-pentanol 52 49

n-hexanol 290 373

ethylacetaat 10400 1096

ethyllactaat 31 8

2,3-methyl-1-butanol 31 18

In the above table you can see the influence of the dough yield on the quantity of volatile components produced during ripening. A dough yield of 150 means 100 parts of flour + 50 parts of water while a
dough yield of 300 means 100 parts of flour and 200 parts of water. So the thinner the sour is, the less aromatic compounds will be produced.

Finally the next table gives an idea what happens if the type of lactic acid bacteria is changed.

component L. sanfrancisc L. fermentum L. alimentaris

ethanol 18200 5700 23

n-pentanol 52 89 166

n-hexanol 290 423 384

ethylacetate 10400 4960 17

ethyllactate 31 12 8

2,3-methyl-1-butanol 31 34 98

Researchers in Finland have discovered lactic bacteria that naturally produce hydrocolloids in wheat bread using sourdough, and could be used to make additive-free products that meet taste and texture
requirements.

Sourdough always contains lactic acid bacteria, which are responsible for the fermentation process. But Kati Katina, senior research scientist at VTT Technical Research Centre of Finland led a three-year
project to screen over 100 other cereal and food-based microbes to find out which ones work in a wheat matrix and can yield helpful hydrocolloids. The first phase of the project involved a modelling
system.

Once the team had narrowed down the potential candidates, it progressed to the baking stage. Katina told Food Navigator that the team was “quite lucky” to find four or five lactic bacteria that helped the
mechanical processability of the dough, improved shelf life, and increased volume. The taste was mild and lacked the pungency often associated with sourdough bread.

The effect is attributed to the production of exopolysaccharides during the fermentation process, which act as coagulants and emulsifiers.

In a report on the work published in the journal Food Microbiology, Katina and her team said that Weissella confusa was identified as a strain with particular potential. Others from the general
Leuconostoc, Lactobacillus, and Weissella were seen to produce exopolysaccharides, but with some strains the positive technical results were marred by acidification. This was not the case with Weissella
confusa.

The indication is that the addition of the lactic bacteria early on means there is no need for other additives to be used in the manufacturing process, in order to achieve the same high quality results.

The project as funded by the Finnish Funding Agency for technology and Innovation Tekes, as well as by VTT. Katina said that since it is a nationally funded project, the findings have been made public and
some bakeries are already putting them to use in their projects.

However Katina’s work is not yet over: There is potential to use the technology for producing ingredients for other cereal products and foods, such as extruded snacks, she said.

She is now investigating the nutritional aspects of the technology. Sourdough already has a reputation for having a low glycaemic index, and it is thought that the formation of the hydrocolloids could
enhance this effect.

Organic acids

The production of organic acids and consequently the pH drop in sourdough have a major effect on structure forming components such as starch, gluten and arabinoxylans. The primary effect of the
organic acids on the protein fraction is the increased swelling and solubility of gluten proteins. This results in a softer dough with less stability and shorter mixing time. Furthermore softness of the gluten
promotes swelling and increased water uptake. Partial hydrolysis of starch also exerts positive effects on the starch granules leading to an increased water binding capacity.

A secondary effect of the pH drop relates to the change in the activity of the enzymes present in the dough (either "naturally" present or enzymes added via the so called "improvers"). Flour proteases
have their pH optimum below pH 7 and increased proteolysis occurs in doughs at pH 4 compared to non-acidified systems. Of course the increased activity of cereal proteases can also be attributed to
longer fermentation times. The rheological consequence of gluten degradation is a major reduction of elasticity and firmness of the sourdough and subsequent bread dough. Whether this has positive or
negative effects on the volume of the bread and staling depends on the acidity profile and gluten network.

Physicochemical changes in the protein network resulting from sourdough fermentation enhance gas retention and allow greater expansion due to softer and more extensible doughs. It has been proved
that the gluten in dough made with sourdough has a more amorphous nature and there were greater areas of aggregated material, stronger and ticker protein strands. The presence of thicker strands
could be the reason for the increase in loaf volume. And no doubt there is a correlation between higher volume and softer crumb as well as a reduced rate of staling.

However if the acidity of the sourdough becomes too high, the bread volume will decrease. This is attributed to the hydrolysation of the gliadins and glutenins. This can clearly be observed in chemically
acidified doughs. Especially high molecular glutenins are completely degraded, which leads to strong gluten softening. Weaker gluten increases the expansion of the dough but also decreases gas retention.
Needless to say that the acidity level of the sourdough as well as the bread dough must be carefully controlled in order to avoid volume loss.

The drop in pH not only has an effect on the proteases but also on the amylases. Acid conditions partially inactivate amylases. This is an important aspect of the production of rye bread since excessive a-
amylase activity results in a sticky crumb, very open grain and a reduction of loaf volume. In wheat flour a-amylases are practically absent while ß-amylases are abundantly present. However ß-amylases have
little influence on the starch granules and are inactivated before starch gelatinisation.

Kefir sourdough

Kefir can be successfully used for sourdough-type bread making, leading to bread of good quality, increased shelf-life and better flavour, states new research from Greece. The authors, who published
their findings in Food Chemistry, said that kefir degrades at lower rates compared to conventional baker’s yeast breads and as it can ferment lactose, and consequently, cheese whey, the main liquid waste
of the dairy industry, it is a an extremely sustainable ingredient.

“The fundamental idea behind the use of kefir for food production, apart from its proven positive effect on quality and preservation, is the utilisation of a low cost, yet seriously polluting raw material,”
they argue.

The food scientists explained that kefir is a natural mixed culture in which lactic acid bacteria (LAB), yeasts and other bacteria co-exist in symbiotic associations.

The starter culture originates from the Caucasus region in Russia, is popular in Eastern and Central Europe but is also gaining awareness among West European consumers for its probiotic and nutraceutical
properties.

Staling

The losses of aroma volatile compounds during storage – staling - are part of the overall physicochemical changes that occur in bread from the moment of production and during storage.

Controlling staling and maintaining the quality and freshness of bread for longer periods, obviously lead to financial gain, and the researchers thus argue that the rate of staling, including the rate of
flavour loss, is one of the most intriguing concerns for investigation in bread manufacture research.

Studies on the changes of bread flavour during storage have indicated that within three to four days most of the volatile compounds are dramatically lost. The use of sourdough seems to be able to
efficiently control this problem, slowing down bread flavour losses, compared to yeast leavened breads, said the authors.

“However, there are other factors to be taken into account, such as the type of flour, the sourdough fermentation conditions (pH and temperature) and the selection of starter cultures with specific and
desirable metabolic properties (e.g. production of specific volatile compounds),” they added.

The scientists said that while previous studies have shown that kefir is able to improve the overall quality of bread, there is no research published on monitoring of changes in the aroma volatile compound
profiles of sourdough breads made with kefir during storage, and thus they aimed to evaluate the efficiency of the culture in this regard.

The study

The aroma volatile compositions of sourdough breads containing kefir grains were monitored by SPME GC–MS analysis during a five day ambient storage period.

The researchers explained that breads were made with 20 per cent and 10 per cent kefir sourdough (Breads A and B respectively), and were compared with breads made with commercial sourdough (Bread
C) and sourdough prepared in the laboratory without the addition of a starter culture (Bread D).

A dramatic decrease of volatiles was observed during storage for all samples, but the kefir sourdough breads (A and B) exhibited more complex profiles of volatiles with lower loss rates during storage,
noted the authors.

They also reported observed differences in the percentages of esters on total volatiles - 6.2 per cent, 5 per cent, 2.8 per cent and 2 per cent in the case of breads A, B, C, and D, respectively.

And the researchers said that the customer oriented sensory evaluation revealed significant differences among the tested samples, with best results scored in the case of bread A in all days of storage,
agreeing with the analytical data.

The authors concluded that their findings reveal the superiority of kefir sourdough over conventional yeasts in terms of bread aroma and the losses observed during storage.

Altamura bread

Altamura bread has received PDO-status i.e. protected denomination of origin. This means that Altamura bread can only be made in the Puglia region of Italy using specific cultivars of durum wheat and the
use of a 3 stage sourdough. The full sour is obtained by a 3-stage process in order to gradually increase the amount of acidified dough. At each step, water and durum flour are mixed with the previous
fermented dough, which is added at the proportion of ± 20 % based on flour weight. On the basis of the ratio between the ingredients that are indicated in the original recipe (100 kg of durum flour + 20
kg of sourdough + 60 kg of water + 2 kg of salt) and assuming a full sour with a dough yield of 160, which contains 12,50 kg of flour, the final dough should have a dough yield of 162.

Noël Haegens

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