• Yogurt is defined as a product resulting from milk by fermentation with a mixed starter culture consisting of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus.
• However, in some countries, including Australia, other suitable lactic acid bacteria are permitted for use as starter cultures. As a result, some yogurt manufacturers use Lactobacillus helveticus and Lactobacillus jugurti for yogurt manufacture
Milk (100 g)
Yoghurt (100 g)
Some typical water-soluble vitamin contents of skim milk and low-fat yoghurt
– On the basis of the method of production and the physical structure of the coagulum:
a) Set yogurt: product formed when the fermentation of milk is carried out in a retail container, and the yogurt produced is in a continuous semisolid mass.
b) stirred yogurt: when the coagulum is produced from milk, and the gel structure is broken before cooling and packaging.
Fluid yogurt can be considered as stirred yogurt of low viscosity.
– On the basis of flavorings, yogurts are divided into three categories.
a) Plain or natural yogurt is the traditional product, which has a typical sharp ‘nutty’ flavor.
b) Fruit yogurts are made by addition of fruits, usually in the form of fruit preserves, puree or jam.
c) Flavored yogurts are prepared from plain or natural yogurt by adding sugar and or other sweetening agents, flavorings and colorings.
Ultrafiltration and reverse osmosis:
• Milk concentrated by ultrafiltration to 18–20% total solids has been reported to produce good-quality yogurt.
• Casein is the major contributor of viscosity followed by fat and whey proteins.
• gelatin, starch, vegetable gums, carrageenan, and pectin are used most widely as stabilizers for yogurt. The best yogurt texture is achieved by using gelatin at 0.3–0.8%.
Sweeteners are added to plain yogurts, generally with fruits.
• Addition of sugar is a method of cutting the sharpness of yogurt flavor.
• Up to 10% sucrose is usually used when fruit is added to yogurt.
• A mix containing 4% or more sucrose, may reduce acid production and lower cell counts of both microbial species when incorporated in the mix prior to fermentation.
• Nonnutritive or intense sweeteners such as aspartame (NutraSweet) may find favor among diet conscious consumers. 200 times sweeter than sucrose, only a small amount is required for desirable sweetness so yogurts with sweeteners require a bulking agent such as polydextrose.
• used to effect stabilization of the lipid phase against separation by gravity.
• specific gravity of milk fat = 0.9, skim milk= 1.036.
• D of the fat globules in milk ranges from 1 to15 μm, with an average of 3–4 μm.
• Homogenization reduces the average diameter of fat globules to 1 or 2 μm.
• There is usually a four to six fold increase in the surface area.
• The fat-globule membrane protects the fat globules from lipase.
• Upon homogenization, the fat-globule membrane is destroyed, and the fat globule is vulnerable to attack by lipase, which is naturally present in milk.
• Because of this, the milk is pasteurized to inactivate lipase before homogenization.
• Since the efficiency of homogenization is much better at higher temperatures, the mix is warmed to around 65 °C to liquefy fat and then forced at high pressure (1.8 × 104 kPa) through a small orifice to reduce the size of the fat globules
• typically 85 °C for 30 min.
(1) to destroy all pathogenic bacteria,
(2) to inactivate all the enzymes that may be present in milk, including lipase,
(3) to destroy most of the spoilage-causing bacteria, including thermodurics,
(4) to denature whey proteins, β-lactoglobulin (90%) and α-lactalbumin ( 60 %).
• Increased hydrophilic properties of the casein leads reduced syneresis, forms stable coagulum.
• The hydration of the protein decreases when milk is heated over 85 C
• the heated milk forms a smooth gel-like structure at pH=4.6 (isoelectric point of casein , casein proteins carry no electrical charge).
Inoculation and Incubation
• mix is cooled to 45 °C and inoculated at a level that varies from 0.5 to 5%.
• The maximum amount recommended is 5%. This level will cause very rapid acid production, but leads to defects in aroma, and a large amount of culture must be prepared.
• The optimum level is 2%, 1% each of Lb. delbrueckii ssp. bulgaricus and Sc. thermophilus.
o Streptococcus thermophilus and Lactobacillus bulgaricus ( symbiotic growth)
mixed culture produces more L.acid than individual pure cultures
mixed culture produces more acetaldehyde than individual pure cultures
L delbrueckii ssp. bulgaricus hydrolyzes milk proteins, the caseins, thus releasing essential amino acids, including valine, which stimulate the growth of S thermophilus. Initially, S thermophilus grows rapidly, reducing the pH to around 5.4, which stimulates the growth of Lb. delbrueckii ssp. bulgaricus, which is acid-tolerant and produces large amounts of lactic acid, which reduces the pH.
S. thermophilus uses oxygen during its growth, which makes oxidation–reduction potential more favorable for L. bulgaricus; it also produces formic acid, which stimulates the growth of the lactobacillus.
• the inoculated mix stirred for 10–15 minutes after the addition of the cultures.
• This is followed by dispensing the mixes into consumer-sized containers and incubating at 42 °C for approximately 4 h until the pH decreases to 4.5.
• For the manufacture of stirred yogurt, the inoculated mix is incubated in bulk, and when the pH reaches the desired value, the yogurt is stirred, cooled, mixed with various flavorings, if required, and filled into retail containers
• the opt.temp. S thermophilus 37 °C, for Lb. delbrueckii ssp. bulgaricus is 45 °C.
• The ratio of lactobacilli and streptococci in natural yogurt for optimum flavor should be 1:1.
• When a 2% inoculum is used at an incubation temperature of 42 °C, the milk will coagulate, and a firm gel will form in 3.5–4 h, and the pH will decrease to 4.4–4.5.
• When frozen concentrated cultures are used, an incubation period of 5–8 h may be required.
• Similarly, freeze-dried cultures may require longer times than fresh and active cultures.
Biochemistry of Fermentation:
• Fermentation of carbohydrates is energy supply of lactic acid bacteria.
• Lactose is the only sugar in milk and it is used for this purpose.
• Catabolism of lactose by S.thermophilus and L.bulgaricus takes place inside the cell and hence the initial step is to transport the lactose molecule across the cell wall membrane.
Phosphotransferase (PTS) system:
lactose is phosphorylated during its dislocation (lactose-P)
• Inside the cell: lactose-P ® D- glucose +galactose-GP ® hydrolysed by beta-D- phosphogalactosidase
Glucose (by EMP pathway) ® pyruvate (by lactate dehydrogenase) ® lactic acid
• But metabolism of galactose-GP is somewhat different
(by D-tagatose-GP pathway) ®glyceraldehyde-3P +EMP ® pyruvate +LDH ® lactic acid
• Mostly glucose utilized, very little galactose utilized.
• Especially in the presence of a more readily fermentable sugar, i.e. glucose, galactose pathway is suppressed.
Production of Lactic Acid:
• Lactic acid helps to destabilise the casein micelles by progressively converting the colloidal calcium phosphate/phosphate complex (in the micella) ® soluble calcium
• So micelles being gradually depleted of calcium… leading to coagulation of casein at pH=4.6-4.7 and the formation of the yoghurt gel.
• Also, L.acid gives sharp and acidic flavor.
COOH COOH ½ ½ HO-C – H L(+) lactic acid (S. thermophilus) H- C – OH D(-) lactic acid (L.bulgaricus) ½ CH3 CH3
• During the manufacture of yoghurt, S.thermophilus grows faster than L.bulgaricus and hence L(+) lactic acid is produced first followed by D(-) lactic acid.
• Yogurt usually contains 45-60% L(+) and 40-55% D(-).
• if yogurt contains >70% of L(+) lactic acid
Probable reasons ?????
• starter culture was predominantly S.thermophilus
• fermentation temperature <40 C
• product is cooled at low acidity and cooled product has <0.8% Lactic acid
A new trend in yogurt making:
• to incubate the yogurt mix at a significantly lower temperature than normal for a longer period.
• One of the advantages is that products require less starter cultures and shorter cooling times.
• An incubation temperature of 30 °C with 0.25% starter culture may take 12–14 h.
• The final acidity of yogurt is 0.9–1%.
• The US standard requires a titratable acidity of 0.9% or higher, whereas the Australian standard requires a pH of 4.5 or lower.
• According to IDF, the minimum acidity of yogurt should be 0.7%.
• Once the desired acidity has been reached, the product is cooled to <10 °C as quickly as possible.
• stirred yogurt,
• In one-phase cooling: cool to <10 °C prior to the addition of flavoring material and filling.
• In two-phase cooling, the temperature of the product is reduced to 15–20 °C during the first phase cooling before addition of flavoring materials and filling of containers followed by the second stage cooling to <10 °C in a cold store.
• The viscosity of yogurt improves during storage for 1–2 days.
• So important to delay the sale or distribution of yogurt for 24–48 h.
• During cold storage, minimize rough mechanical handling of the packaged yogurt, and to maintain the temperature of <5 °C.
• During transport, especially in summer, shaking of yogurt can lead to a reduction in viscosity and syneresis.
Continous Stirred Yoghurt Production:
A two stage process;
• Milk ® pasteurized ® homogenized ® a pH-stat prefermentation and plug flow
• pH-stat fermentor: prefermentation of milk until pH decreases to 5.7 ( at 45 C)
• Plugflow fermentor: formation of the yoghurt texture due to further acidification to pH 4.5 (37 C)
• The specific gravity of coagulated milk is slightly more than that of milk and the plug flow was therefore directed downwards.
• Flowing through the tank a gradient of acidity will rise ® texture will build up.
Three problems to establish and maintain plug flow;
• Transferring milk without disturbing coagulating milk in the second tank
• The coagulating milk had to be prevented from adhering to the wall of the coagulation
• The coagulated milk had to be stirred and removed from the coagulation tank without disturbing the plug flow.
Solution to these problems:
• Milk is fed through a centrifugal distributor on to metal screen in to plugflow fermentor to minimize disturbance of coagulating milk
• poly tetrafluoroethylene (PTFE) or lecithin (cheaper, long life) can be used to cover stainless steel to prevent adhering of yoghurt coagulum soya lecithin and ethanol mixture sprayed on the wall before process
• Bottom portion of coagulum, where fermentation is completed has to be stirred and pumped out without disturbing the upper portion.
A horizontal plate with holes move up and down (coagulum is sheared through the holes in the plate and downward movement pushes the yoghurt out)
Advantages of continuous fermentation of yoghurt:
• space saving
• reduction in size of equipment
• reduction of yoghurt losses in fermentation tanks and pipelines
• reduction in capacity of cooling and filling sections
• greater flexibility in relation to the total amount production
• no need for all the milk to be in stock at the start of production
• uniformity of product quality and characteristics
• better control over acid development
Fed-batch prefermentation is a new method to accelerate the production of stirred yoghurt.
• The fermentation can be greatly accelerated by using a high inoculation percentage of a culture in the exponential growth phase instead of a low inoculation percentage of concentrated
• The quality of the yoghurt prepared by a fed-batch prefermentation is comparable to that of yoghurt produced in the traditional way.
• A fed-batch experiment with a duration of 51 hours shows that it is possible to perform fed-batch fermentations for a long time with the same culture without infections or loss of product quality.
SPECIAL YOGURT PRODUCTS
Low-lactose Yogurt or Lactose Hydrolysed Yoghurt (LHY):
• Only small parts of lactose is converted (fermented) to L.acid. Lactose malabsorbers do not produce sufficient lactase (β–galactosidase): cannot hydrolyze the ingested lactose completely, gastrointestinal discomfort, stomach upset, diarrhea.
• Low-lactose or lactose-free milk for lactose sensitive (intolerant) individuals can be prepared either by the physical removal of lactose by ultrafiltration or by hydrolysis of lactose into the corresponding monosaccharides, glucose, and galactose.
• Lactose can be hydrolyzed using strong mineral acid or enzymes ( β-galactosidase, lactase, catalyzes the hydrolysis of the β-1 → 4-galactosidic linkage present in lactose)
• isolated commercially from the fungi, Aspergillus niger, A. oryzae, and A. flavus, or the yeast Saccharomyces lactis or from the bacterium Escherichia coli.
• Yogurt bacteria (L delbrueckii ssp. bulgaricus and S thermophilus)possess the highest amount of β-galactosidase activity among the lactic acid bacteria.
• Low-lactose yogurt is then produced by the use of low-lactose milk during processing.
• One can increase sweetness of yoghurt without increasing calorific value by hydrolysing lactose (0.4 as sweetness) by beta-D-galactosidase
Lactose ® galactose + glucose
Sweetness 0.4 0.6 0.7 (sweetnes of sucrose is 1)
• this is desirable for manufacture of fruit/flavored yoghurt
• For sweetness it is cheaper to add sweeting agents.
Flavored with natural orange, lemon, cherry or apple flavors
• Liquid or dry form ® gradually releases the CO2 when it is reconstituted with water.
• carbonated yoghurt less acidic (because of metal carbonates)
• Ca-carbonate is more advantageous than Na-carbonate because ,it releases CO2 much slower
• Addition of Calcium compounds improves opacity
• (calcium + acids ® insoluble salt ¯)
• Since soybean is plentiful, relatively inexpensive and rich in protein some effort has been devoted to exploiting it for the manufacture of more acceptable and palatable food products.
• Main problems : beany flavor and flatulence (CO2 production, hydrogen and methane production by intenstinal flora from oligosaccharides)
• using L.bulgaricus alone, an acceptable yoghurt-like product can be produced
• Lactic acid (optimum) = nearly 1.15 %
• Flavor of fermented soy milk was directly related to the levels of n-pentonal (produced by S.thermophilus) and n-hexonal (naturally present in soy milk)
• Reduction of oligosaccharides was insignificant.
• Color problem is another problem for soy-milk yoghurt.
Heated or Pasteurized Yogurt
• at 4 °C, the titratable acidity increases due to residual activity of the starter bacteria ( ‘post-acidification,’
• Without heat treatment, the shelf-life of yogurt is 4–5 weeks at 4 °C.
• Heating destroy most of the starter bacteria and yeast&molds: the shelf-life of the product can be extended to 8 we
• No post-acidification occurs in heated yogurt.
• by heat-treating yogurt in the package at about 55 °C for 30 min, followed by cooling.
• The main problems: loss of flavor and syneresis. Stabilizers can be used to overcome the latter problem.
• yogurt contains very few or no viable yogurt bacteria after heating ( raises questions about the definition of yogurt)
Prebiotics, Probiotics, and Synbiotics
• ‘probiotic’ : from two Greek words meaning ‘for life.
• ‘they are live microbial food or feed supplements that provide a beneficial effect on hosts (human or animal) by improving the microbial balance in the intestine.’
• The intake of these bacteria is reported to help restore the balance in the intestinal microflora, which may have been lost due to stress, antibiotic use, or illness.
• The major strains of bacteria used in probiotics, L acidophilus and Bifidobacterium spp., are dominant organisms of human large intestines.
• These microorganisms are claimed to inhibit the growth of pathogenic organisms through the production of organic acids and bacteriocins.
• Other benefits include reduction in lactose malabsorption, suppression of potentially harmful enzymes, increased immune response due to increased production of secretory immunoglobulins A, reduction in serum cholesterol, and antimutagenic effects.
• The growth of bifidobacteria is dependent on the presence of complex carbohydrates such as oligosaccharides and other substrates such as N-acetyl glucosamine and lactulose.
• These carbohydrates that stimulate the growth of bifidobacteria are known as ‘bifidogenic factors.’
• Some oligosaccharides, due to their chemical structure, are resistant to digestive enzymes and therefore pass into the large intestine, where they become available for fermentation by bifidobacteria.
• Compounds that are either partially degraded or not degraded by the host and are preferentially utilized by bifidobacteria as carbon and energy sources are defined as ‘prebiotics.’
• Some of the bifidogenic factors that are of commercial significance include fructo-oligosaccharides, lactose derivatives (such as lactulose, galacto-oligosaccharides), isomalto-oligosaccharides, xylo-oligosaccharides, gluco-oligosaccharides and soybean oligosaccharides.
• Resistant starch and nonstarch polysaccharides are classified as colonic foods but not as prebiotics because they are not metabolized by certain beneficial bacteria.
• Products that contain both prebiotics and probiotics are referred to as ‘synbiotics.’
• An example of synbiotic includes SymBalance yogurt, which contains inulin as prebiotic and L reuteri, Lb. acidophilus, and L casei as probiotics.
• Inulins are a group of naturally occurring oligosaccharides (several simple sugars linked together) produced by many types of plants
• Inulins with a terminal glucose are known as alpha-D-glucopyranosyl-[beta-D-fructofuranosyl](n-1)-D-fructofuranosides, abbreviated as GpyFn. Inulins without glucose are beta-D fructopyranosyl-[D-fructofuranosyl](n-1)-D-fructofuranosides, abbreviated as FpyFn where n is the number of fructose residues and py is the abbreviation for pyranosyl.
• Inulin is indigestible by the human enzymes ptyalin and amylase, which are designed to digest starch. As a result, inulin passes through much of the digestive system intact. It is only in the colon that bacteria metabolise inulin, with the release of significant quantities of CO2 and/or methane.
• (rich in chicory(hindiba), garlic,leek( pırasa)
• It is widely accepted that because of acid production by L acidophilus and bifidobacteria, the enteropathogenic bacteria are unable to grow.
• The growth of clostridia and E. coli, when cocultured with bifidobacteria, has been found to be inhibited, even at a neutral pH, suggesting that acid production may not be solely responsible for inhibition.
• Metabolites produced by bifidobacteria may be partly responsible for the inhibition of pathogens.
Acidophilus and Bifidus Yogurt (AB Yogurt)
• Lb. acidophilus and bifidobacteria are normal inhabitants of the intestine of many animals including man (predominate the gut flora in breast-fed infants.)
• 11% of all yogurt sold in France now contains L acidophilus and Bifidobacterium spp.
• L acidophilus and Bifidobacterium spp. are difficult to propagate because of their specific nutritional requirements.
• Bifidobacteria are not as acid-tolerant as L acidophilus, and the growth of Bifidobacterium species is significantly retarded below pH 4.0.
• L acidophilus and Bifidobacterium spp. are slow acid producers; the slow growth rate of these organisms can be compensated by adding a higher level of inoculum, such as 5 or 10%
• Yogurt bacteria are usually added to carry out fermentation.
• If pure cultures of Lb. acidophilus and or Bifidobacterium spp. are used, the time required to reduce the pH of milk to 4.5 could be as long as 18–24 h at 37 °C.
Recent Advances in Probiotic Yogurt
• The most commonly used species in commercial probiotic products are Lb. acidophilus, Lb. casei, Lactobacillus GG , B. bifidum, B. longum, B. breve, and B. infantis.
• Additional blends are also being investigated, such as Lb. reuteri, Lb. plantarum, and Lb. casei.
• In Australia and Europe, yogurt containing Lb. acidophilus and Bifidobacterium spp. is referred to as AB yogurt.
• The recent trend is to incorporate Lb. casei in addition to Lb. acidophilus and Bifidobacterium spp, and such products are known as ‘ABC yogurt.’
• Because of sensitivity to acid, Lb. acidophilus and Bifidobacterium spp. in yogurt begin to die within a few days after manufacture because of acid produced during manufacture and storage.
• In order to provide health benefits, the suggested level for probiotic bacteria is 106 cfu per gram of a product.
• However, studies have shown a low viability of probiotics in market preparations.
• Many yogurt manufacturers use a starter culture devoid of L delbrueckii ssp. bulgaricus but a combination of L.acidophilus, bifidobacteria and Sc. thermophilus (known as ‘ABT’) as starter cultures to overcome the postacidification problem.
• However, the use of ABT starter culture increases incubation time significantly as S.thermophilus is the main organism responsible for fermentation in ABT starter cultures, and this organism is less proteolytic than L.delbrueckii ssp. bulgaricus.
Concentrated-Strained yogurt ( Süzme yogurt )
• Concentrated or strained yogurt is a popular product in the Middle East region.
• The product is known as labneh, labaneh, or lebneh (in most Arab countries), mast or mastou (in Iraq), leben zeer (in Egypt), tan or than (in Armenia), Greek yogurt or Greek-style yogurt (in the UK), stragisto or tzatziki (in Greece), torba or süzme (in Turkey), basa or zimne (in the Balkans), and yogurt cheese (in some parts of the world).
• The cold natural plain yogurt is stirred and emptied into cloth bags of about 25 kg.
• The bags are stacked on top of each other in a vertical press that is located in a refrigerated room.
• Pressure is applied in order to assist in whey drainage for duration of 12–18 h.
• On the following day, the concentrated product is emptied into a mixing bowl to obtain a uniform texture prior to packaging.
• Alternatively, long, horizontal filter cloths can be used; the long sides are supported on poles and gently oscillated up and down, while slight lateral pressure is applied.
• This method of processing is known as the modified ‘Berge’ system, and was developed in France in the mid-1960s for the production of fresh curd cheese.
• The application of more pressure and a longer dewheying stage will yield a product that contains ≥ 30 g 100 g−1 total solids, and is known as ‘yogurt cheese.’
• The production of strained yogurt by centrifugation of heated yogurt has been used successfully in experimental trials and commercially in different countries; concentration is achieved using a nozzle or Quarg separator.
• Skimmed milk should be used for the manufacture of the yogurt, and the fermented milk is stirred vigorously, heated to about 60 °C, cooled to about 40 °C and concentrated to 18 g 100 g−1 total solids, cooled to about 12 °C, blended with cream, and finally packaged.
• If whole milk is used instead, the nozzles of the separator will clog. ( recent developments in the design of the centrifugal separators have made it feasible to use fermented whole milk)
• After acidification (i.e., pH 4.6–4.8), the fermented milk is heated to 60 °C to inactivate the culture and control the level of acidity, and then deaerated for 15–20 min to assist the separation of whey in the separator.
• A centrifugal pump transports the fermented milk through a switchable double strainer to break up any lumps before it enters the separator.
• The concentrated product leaving the separator is blended with cream and seasoning (e.g., salt, herbs or fruit flavors – optional), cooled, and packaged.
• A typical chemical composition for strained yoghurt is 24 g 100 g−1 total solids and 9.6 g 100 g−1 fat
• Capacities of such separators are up to 6.5 tonnes h−1
Strained yogurt production-Ultrafiltration (UF)
• Two different systems of UF have been used for the production of strained yogurt.
A) the standardized milk is concentrated by UF to the desired solids content before homogenization, heat treatment, and fermentation,
B) the warm yogurt (about 45 °C) is concentrated by UF.
• The quality of strained yogurt made by UF of warm yogurt closely resembles the traditional product in terms of elasticity, firmness, and structure.
(1) production of yogurt from whole milk,
(2) UF of yogurt at 35–45 °C using batch or multistage systems,
(3) partial cooling to 20 °C, fruit blending (optional), and packaging,
(4) cooling in the retail container to < 10 °C.
Freezing (Frozen yogurt )
• Frozen yogurt resembles ice-cream, in that the fresh stirred yogurt is stabilized, fortified with fruit base (syrup or pieces), whipped, and frozen.
• In general, the product is classified into three main categories: soft, hard, or mousse.
• The outlet freezer temperature of these two products is −6 °C, and the storage temperature for soft and hard frozen yogurts is −6 °C and −25 °C, respectively.
• Although air is normally used at the whipping freezing stage, a longer shelf-life product can be achieved by using nitrogen rather than air.
• powdered yogurt, dried or instant yogurt : aimed towards the do-it-yourself consumer market, the baby food manufacturers, and the foodbaking industries.
• The concentration stage, before drying, should be carried out at a low temperature (about 50–60 °C) to minimize scorching on to the surfaces of the evaporator, or discoloration of the final powder,
• The acidified product is concentrated using an ordinary evaporator to 25–36 g 100 g−1 solids at 57–58 °C.
• The processing conditions must be moderate in order to ensure a high viable cell count of starter culture organisms in the dried product.
• The concentrated buttermilk, which is highly viscous, is pumped to the spray drier at 43 °C and dried using an inlet air drying temperature between 175 and 190 °C.
• The moisture content in the dried product is around 4 g 100 g−1, and the tapped bulk density is 0.77–0.83 g cm−3.
• It was first produced in the Caucasus Mountains, where the drink was fermented naturally in bags made of animal hides.
• Kefir is now produced commercially in large quantities in the countries of the former Soviet Union, and in appreciable quantities in Poland, Germany, Sweden, Romania and Turkey.
• It is also still produced traditionally, under a variety of names including kephir, kefer, kiaphur, knapon and kippi.
•Cow’s and goat’s milk are usually preferred for the manufacture of kefir, and either whole or skimmed milk, or a mixture, is used.
• The composition of the product is determined primarily by the raw material and the microflora of kefir grains, and is subject to regional variations.
• Kefir is a self-carbonated, fermented beverage containing about 0.8–1% lactic acid and 1–2% alcohol.
• During fermentation, lactose-forming yeasts produce alcohol and CO2 and lactic acid bacteria convert lactose to lactic acid.
• Some proteolysis occurs in the milk, and a yeasty aroma develops.
• The flavour of kefir is mildly alcoholic, yeasty and sour, with a tangy effervescence, and is related to the composition of the kefir grains.
Methods of production Traditional method:
• Pasteurized milk is inoculated with kefir grains, which are composed of lactic acid bacteria, yeasts, streptococci and acetic acid bacteria in various proportions.
• The usual initial heat treatment is at 90°C for 30 min.
• The inoculum consists of a ‘mother culture’ (2–10%, w/v).
• Fermentation is achieved at 20–25°C over about 24 h.
• The kefir grains are then removed, by straining.
• The filtrate is refrigerated overnight and the beverage, which contains live microorganisms from the grains, is ready for consumption
A two-step fermentation (known as the Russian method)
• It is applied, to stimulate the activity of the microorganisms and accelerate the changes in the milk. In the first step,
• The milk is inoculated with kefir grains (2–3%, ) to prepare a mother culture.
• Fermentation takes about 24 h at 20–25°C.
• The grains are removed and the filtrate, which is the mother culture, is added to pasteurized milk (1–3%, v/v).
• The second fermentation lasts 12–18 h at 20–25°C
Many problems are associated with traditional kefir production, and this has led to more modern methods.
• The production of CO2 by yeasts often leads to ‘blown’ containers, which are mistakenly judged by the consumer to be ‘spoiled’.
• The shelf life of traditional kefir is as short as 2–3 days.
• In order to resolve these difficulties and to facilitate large-scale production, the use of standard starter cultures, consisting mainly of streptococci, has gained popularity.
• The second starter is chosen so as to include either aroma-producing or acid-producing bacteria, depending on the required characteristics of the kefir.
• The starter for lactic cultures is a blend of Leuconostoc cremoris and Lactococcus lactis subsp. lactis biovar diacetylactis, while that for yoghurt cultures is a blend of Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus.
• producing kefir from pure, defined cultures to have better control of the microorganisms involved, greater ease of production and more consistent quality. In addition, the shelf life of the kefir can be extended to 10–15 days at 4°C and its modification and improvement, e.g. interms of health-related and nutritional aspects,are facilitated.
• The main problem when using pure cultures is finding the balance between the bacteria and yeast strains which creates a product with the characteristic properties of traditional kefir, including both the organoleptic qualities and the health benefits.
• The optimal inoculation is in the range 1 : 50 (culture : milk).
• High inoculation kefir grains (1 : 20) shorten the fermentation, but the growth rates of the yeasts and of both homo- and heterofermentative streptococci decrease compared with growth at lower inoculation rates (e.g. 1 : 50).
• The balance of microorganisms in kefir grains is also affected by agitation: frequent agitation during the fermentation may cause the numbers of bacteria and yeasts to increase.
• frequent washing of the grains with water leads to a rapid decrease in the number of microorganisms; also, the fermentation takes longer and the taste and consistency of the end product become nonrepresentative of kefir.
• Thus the microflora of kefir can be controlled by altering the conditions of fermentation.
The following procedures should ensure a product of high quality and, to some extent, standardization:
• The milk should be changed at the same time every day.
• The ratio of grains to milk should be in the range 1 : 50.
• The milk should be heat-treated (90–95°C for 5–15 min, or ultra-high temperature).
• The milk should be fermented at 18–20°C.
• During incubation, the fermenting milk should be stirred two to three times
• Another critical step in ensuring the quality of kefir is post-fermentation cooling. Cooling must take place slowly (over 10–12 h) for the pronounced taste and aroma of kefir to be acquired, so that the necessary accumulation of homofermentative lactic streptococci and yeasts can occur.
• Leakage and blowing of containers, as a result of excess CO2 production, are major problems in the kefir industry.
• To avoid them, non-lactose-fermenting yeasts should be used – in which case sucrose supplementation is recommended.
Microbiology of Kefir Grains
• Kefir grains are gelatinous granules, some 2–15 mm in diameter, consisting of a mixture of microorganisms grouped in a highly organized manner.
• Kefir grains have the chemical composition 89–90% water, 0.2% lipid, 3% protein, 6% sugar (mainly polysaccharide) and 0.7% ash.
• They should be stored wet at 4°C, or should be dried at room temperature for 36–48 h.
• Dried kefir grains retain their activity for 12–18 months, whereas wet grains retain their activity for only 8–10 days.
Storage at −20°C is another effective method of preserving kefir grains.
• The bacterial and yeast cells in the kefir grains are embedded in a slimy polysaccharide material named ‘kefiran’.
• Early studies revealed that Lactobacillus brevis was able to produce kefiran, but later a homofermentative lactobacillus, which was described as an a typical streptobacterium, was found to be responsible.
• Subsequently, a capsule-forming homofermentative bacterium, Lactobacillus kefiranofaciens, was isolated from kefir grains.
• The appearance of kefir grains is similar to that of the tiny florets of cauliflower.
• The proportions of bacteria and yeasts vary, depending on the source of the kefir grains.
Freezing Kefir Grains
• One method for storing kefir grains for periods of up to 2 months, is by freezing the grains.
• To freeze kefir grains effectively, wash the grains with pre-boiled then COOLED water, pat them dry between pre-ironed cooled white towel to remove excess moisture.
• Place the grains in a jar or in a plastic bag, with the addition of dry milk powder ( DMP) then freeze.
• DMP is added as a protective agent.
Drying Kefir Grains
• Kefir grains may be dehydrated for long term storage of up to 12 to 18 months.
• To dehydrate fresh kefir grains, rinse the grains with pre-boiled COOLED water.
• Place the grains in between two sheets of pre-ironed white cotton or linen cloth.
•Leave to dry in a well ventilated warm spot, until the grains become quite firm and yellow in colour.
• Place the dry grains in an airtight jar and store in a cool place e.g., in the refrigerator I add a little dry milk powder [DMP] with dehydrated kefir grains, adding enough DMP to completely cover the grains in a jar or in a zip lock plastic bag
• Mesophilic homofermentative lactic streptococci (Lactococcus lactis subspp. lactis and cremoris) are the most active components of kefir grains, and they cause a rapid increase in acidity during the first hours of fermentation.
• Lactobacillus brevis, which is always heterofermentative, and the facultatively heterofermentative Lactobacillus casei subsp. rhamnosus are common in kefir starters.
• Mesophilic heterofermentative lactic streptococci (Leuconostoc mesenteroides and L. mesenteroides subsp. dextranicum) are mainly responsible for the development of the characteristic taste and aroma of kefir, and they may cause gas formation in association with yeasts.
• Yeasts have a symbiotic relationship with some of the other microorganisms in kefir, and contribute to the formation of its specific taste and aroma and to the release of CO2.
• The kefir microflora is dominated by mesophilic homofermentative lactic streptococci, and so about 10 times more (+)-lactic acid is formed than (−)-lactic acid. CO2 is produced as a result of the activity of yeasts and heterofermentative lactic streptococci.
• CO2 plays an important role in the organoleptic characteristics of kefir, giving it the slight effervescence that is highly appreciated by consumers.
• Alcohol (1–2% (v/v). ), mainly ethanol, is another metabolite synthesized by the yeasts, and also by the lactic streptococci if they are not inhibited by the high acidity and the anaerobic conditions resulting from tight packing.
• Kefiran is perhaps the most important metabolite in kefir, because it acts like glue and keeps the kefir grains intact.
• Kefiran consists of glucose and galactose in equal proportions, and is present in the capsular material of some large, rod-like bacteria, especially lactobacilli.
• Some 30–35% of the wet weight of kefir grains may be attributed to kefiran.
• The accumulation of free amino acids, as a result of proteolysis, contributes to the nutritive value of kefir, as does the formation of B and P group vitamins.
• CO2 produced by the yeasts stimulates the appetite and aids digestion.
• Koumiss is a drink with ancient origins, and is common in eastern Europe and central Asia.
• It is traditionally produced from mare’s milk by a combined fermentation to lactic acid and alcohol, and its highly nutritive and curative characteristics are well known.
• the product of a previous day to seed freshly drawn mare’s milk (usually unpasteurized) in ‘saba’ or ‘turdusk’, made from smoked horsehide.
• Fermentation takes some 3–8 h.
• Production ceased at the end of lactation, in late autumn, and the koumiss starter was put into a glass bottle which was sealed tightly and stored in a cool dark place until early summer, when production began again.
• The starter microorganisms were reactivated by keeping the koumiss starter at room temperature for about 24 h and then mixing with fresh milk three or four times.
• Alternatively, koumiss was mixed with cow’s milk in the middle of winter and kept at room temperature.
• The koumiss starter can also be preserved by drying
• Before use, the dried starter (3–4 tablespoonfuls) is added to 5 lt of fresh mare’s milk and then left at room temperature for about 2–3 days.
• This fermented milk is blended with 6–7 lt of fresh milk and further fermented.
• Mare’s milk has lower levels of fat, protein, ash and total solids than cow’s, goat’s and sheep’s milk.
• Koumiss made from mare’s milk is sweeter than that made from cow’s milk.
• It is milky-green in colour, light and fizzy, and has a sharp alcoholic and acidic taste.
• The digestibility of mare’s milk is good, because it contains a high level of whey protein.
• Tarhana is a fermented food resulting from the combined fermentation of yoghurt with cracked wheat or flour.
• Tarhana is a traditional Turkish food, which is widely known and consumed in the Balkans and the Middle East under different names, including‘trahana’ (Bulgaria), ‘trahanas’ (Greece), ‘taron’ (Macedonia), ‘tarhonya’ (Hungary), ‘kisk’ (Iraq), ‘kishk’ (Egypt, Syria and Lebanon) and ‘goce’ (Turkmenistan).
• Tarhana is used for making soup, and has a considerable share of the ready-soup market in the Balkans and the Middle East.
• The method of manufacture of tarhana does not vary significantly from one region to another, although slight variations in the microbiological and compositional characteristics may occur depending on the locality and its traditions.
• Production is based on a yoghurt fermentation, but concentrated yoghurt or sour milk can also be used.
• Usually, equal quantities of yoghurt and cracked wheat or flour are used.
• Saccharomyces cerevisiae is generally added to the mixture, to give the characteristic aroma of tarhana.
• Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus, from the yoghurt, are responsible for the production of lactic acid and the yeast produces ethanol and CO2.
• Lactobacillus casei, L. plantarum and L. brevis can also be added in order to improve the aroma and flavour.
• After fermentation, the mixture is spread onto a large cloth (traditional method) or a pulsating tray (industrialized method), and is dried at 40°C for 20–30 h (traditional method) or 60°C for 5–8 h (industrialized method).
• The dried mixture is milled and stored at room temperature.
• According to Turkish standards, tarhana must contain protein (min. 14%), moisture (max. 10%) and salt (max. 5%).
• A low level of lactose and a high level of hydrolysed starch facilitate the digestion of the product.
• It is rich in protein, calcium, iron and zinc.
• Unless dried in the sun, it is an important source of group B vitamins – direct sunlight causes the loss of riboflavin, which can be avoided by drying on pulsating trays.
• wheat flour is replaced ( totally or partialy with soy flour)
• No color problem
• Better amino acid profile
• The severe heat treatment received by the process milk, together with the low pH of the final product, make yoghurt extremely safe in terms of public health because none of the recognized pathogens can survive or grow at 4.3 pH.
• the spores of Bacillus cereus will not germinate at low pH
• organisms of concern in soft cheeses, e.g. Listeria monocytogenes, will be inactivated long before the yoghurt reaches the consumer.
• In addition, there is good evidence that some metabolites of yoghurt organisms can actively depress the viability of many enteric pathogens such as Campylobacter, Escherichia and Salmonella species.
• Hydrogen peroxide is one such metabolite released by L. bulgaricus, some strains of which are reported to secrete an antibiotic called ‘bulgarican’.
• S. thermophilus may also release a compound of low molecular weight, with bactericidal properties. However, the activity of these bacteriocins is strictly limited and in practice they do little more than reinforce the effects of acidity
• In natural yoghurt, the principal sugar available is lactose and, because few yeasts can ferment it, the major concerns are species like Kluyveromyces marxianus var. lactis or K. marxianus var. marxianus.
• Both of these lactose-utilizing species grow readily on poorly cleaned surfaces, and hence high standards of hygiene are essential if contamination after heat treatment is to be avoided.
• The same figure applies for moulds – genera including Mucur, Rhizopus, Penicillium and Aspergillus can grow readily at the yoghurt/air interface of an undisturbed carton.
• In contrast to yeasts, just one spore of a fungus can spoil a carton of yoghurt by growing over the surface of the product. Hence protection of filling lines from airborne contamination is essential.
• if regulations permit, sorbic acid is added (usually as potassium sorbate) at a level of up to 300 mg kg−1. This preservative is extremely effective against yeasts, but has little effect on L. bulgaricus or S. thermophilus.
• Excessive acidity may result from continued starter activity during prolonged storage at temperature > 5°C, e.g. the acid-tolerant L. bulgaricus can generate lactic acid to levels of 1.7% or even above, depending on the strain.
• Such levels are too high for the palates of most consumers, and it is this post-production acidification that tends to determine the shelf life of commercially produced yoghurt.
• Some protection can be obtained by lowering the level of L. bulgaricus in the original inoculum, but this can reduce the degree of synergism and such products tend to lack the characteristic flavour of yoghurt.