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Undistilled Beverages
Beer Fermentation
Beer is an undistilled product of grain-mash fermentation brought about by yeast. Beer
fermentation involves the conversion of starchy raw materials into sugars and then into
alcohol. It is generally prepared from malted barley but other starchy grains, e.g., maize, rice
are also used as raw materials. Yeasts especially Saccharomyces cerevisiae or S.
carisbergensis are used in the fermentation process.
Steps in Commerical Production of Beer
Five major steps are involved in the manufacture of beer They are:
(a) Malting. The starchy grains are first malted. The malt is prepared by first soaking the
grains in water and then allowing it to germinate at 170C. During germination, large amount
of amalyses, an enzyme, are produced which are subsequently involved in hydrolysing the
starch into fermentable sugars. After the germination is completed, the grains are dired at
650C.
(b) Mashing. After malting, the pure barley malt is invariably mixed with other grains such as
corn, rye, sorghum, wheat, etc. It is called ‘ground malt’. The latter is first mixed with warm
water at about 700C and 5.0 pH. Mashing brings about partial hydrolysis resulting in the
digestion of starch and protein; the partially hydrolysed solution of mash is filtered and this
filtrate is called ‘beer wort’. Beer wort serves as a rich nutrient medium for the
microorganisms. Beer wort is now boiled with hops.
Commercial Production of Beer
which are the papery scales of the female flowers of the hop vine, Humulus lupulus. The hops
are added for flavour, aroma and mild antibacterial activity to prevent the growth of spoilage.
(c) Fermenting. The beer wort is now inoculated with a pure culture of Saccharomyces
cerevisiae of S. carisbergensis and allowed to ferment at low temperature (5°C-14° C) for
longer period (5-10 days).
(d) Maturing. The fermented beet wort is refrigerated at 0°C for several months (usually 6-8)
to remove the harsh flavour and other undesirable characteristics.
(e) Finishing. This is the final step in which the matured beer is carbonated, filtered and
finally bottled, canned or barreled. Bottled or canned beer is usually pasteurized at 60°C for
about 20 minutes to prevent microbial spoilage.
Types of Beer
(a) Lager beer. It means literally, the stored beer. Lager beer is produced by ‘bottom’
fermentation and is rather high in alcohol (3.93%) and extract with a relatively low
proportion of hops.
(b) Rock beer. It is a heavy beer, dark in colour and high in alcohol (4.69%) and contains
more consumption in early spring.
(c) Ale beer. Ale beer is pale in colour, tart in taste, high in alcohol (4.75%) and contains
more hops. This beer is produced by ‘top’ fermentation.
(d) Porter beer. It is a dark ale but sweeter than the usual ale in taste. It is brewed from dark
or black malt to produce a wort of high extract. The flavour of hops is less distinct than that
of normal.
(e) Weiss beer. This beer is produced mainly from wheat as a result of ‘top’ fermentation. It
is rather light (2.75% alcohol), possesses a distinct flavour of malt and hop, is tart and
contains a large quantity of natural fermentation gas. Weiss beer is somewhat turbid in
appearance.
(f) Stout beer. This beer is stronger porter beer possessing high alcohol concentration. It is
dark in colour and possesses a sweet taste and strong flavour of malt. The flavour of hops is
more pronounced than that of the porter beer.
(g) Cereal beverage. This beer contains less than 0.5% alcohol. It is sometimes referred to as
“near beer”.
Wine Fermentation
Wine is an undistilled product of fruit juice fermentation brought about by yeast. Wine is
produced by the normal alcoholic fermentation of fruit juices, especially the grape juice. The
microorganisms used in the wine fermentation are the various strains of Saccharomyces
cerevisiae such a S. cerevisiae var. ellipsoidens.
Commercial Production
Five major steps are involved in commercial production of wines. They are :
(a) Crushing. Grapes are harvested and ripened to a stage they contain highest sugar
percentage. These fruits are crushed in a wine press and the crushed fruits with juice are
called “must”. The must is generally treated with SO2 to prevent microbial spoilage.
(b) Fermenting. The “must” is now inoculated with the starter culture of selected strain of
the yeast and is aerated slightly to promote vigorous yeast growth. Once the fermentation
starts, the rapid production of CO2 maintains anaerobic condition. The temperature is kept
usually at 25-30°C during fermentation period ranging from 5-11 days in order to inhibit
multiplication of wild yeast and undesirable bacteria that live high temperature.
(c) Tanking.
When most of the sugar is fermented the juice is separated from solid parts of fruits by
allowing it to pass into tanks. These tanks, provided with valves to let the CO2 escape, are
completely filled with juice. The anaerobic condition for alcoholic fermentation is allowed to
continue for about 12 days to increase the percentage of alcoholic concentration.
(d) Maturing.
The wine is then allowed to mature in wooden tanks for 2 to 5 or more years. During
maturing period the wine clears and develops the desired flavour of volatile ester.
(e) Finishing.
The wine may be finally cleared with the addition of gelation, casein or Spanish clay. The
cleared wine is filtered, bottled and pasteurized to prevent microbial spoilage.
Types of Wine
Variety of grape, strain of yeast, nature of fermentation, etc. result in various varieties of
wines. The latter differ from each other in so many attributes that it becomes difficult to
classify them properly. However, some important varieties of wine are as follows:
(a) Red wines.
The wines red in colour are put under this category. In preparing them the grapes are crushed
and stemmed but their skins and seeds are left in the ‘must’. The alcoholic content percentage
of these wines ranges from 11 to 12. Examples – Rose wines, Burgundy wines, Claret wines,
Vinorosso wines, etc.
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Method of Commercial Production
Vinegar is commercially produced by various methods such as ‘Orleans method’. ‘Acetator
method’ and ‘Generator method’. However, it is the ‘Generator method’ which is quicker one
and is generally used in commercial vinegar production (Fig. 17.2). This method is discussed.
Commercial Production of Vinegar by Generator Method
Generators of various sizes (15 feet in diameter and 20 feet in length) are used. The
generator consists of a cylindrical tank with a perforated false bottom supporting beechwood shavings or similar material that will help increase the of air from this bottom that has
an exit at the top. A mix is prepared which consist of an adjusted solution of alcohol
acidified with acetic acid and special nutrients for the growth of acetic acid bacteria. The
latter, spies of the genus Acetobacter, are inoculated into the beech-wood shavings. The mix
is applied in a trough at the top of the chamber and allowed to trickle down over the
shavings. The mix is collected at the bottom of the generator sand is recirculated over the
shavings resulting in more oxidation of alcohol until vinegar of the desired strength is
obtained. Oxidation of alcohol by bacteria may result in the development of temperatures
high enough top kill them. In order to keep the temperature down to 25-300C. cooling coils
need to be provided. The generator method is quicker in comparison to other methods and
the vinegar may be produced within a period of 10 days.
Precautions. Sometimes, undesirable microorganisms spoil the vinegar and the vinegar
eelworm (Turbatrix aceti) causes deterioration of flavour and appearance.
Therefore, care should be taken to check contamination by undesirable bacteria ands moulds
and establishment of eelworm. The generator may be periodically disinfected for the purpose.
Uses
(i) The vinegar is used in pickling, in preserving meat and vegetables.
(ii) It is also used in the manufacture of saladdressing and ketchup.
Antibiotic Production
‘Antibiotics’ are biochemicals secreted by microorganisms which, in low concentration,
inhibit the growth or kill other microorganisms, i.e., the antibiotics are ‘antimicrobial agents
of microbial origin’.
Although some 6,000 antibiotics are known to exist, only about 100 are marketed. About 200
new antibiotics are discovered every year. Microorganisms belonging to three orders viz.,
Eubacteriales, Actinomycetales and Moniliales have contributed immensely to the rise and
development of the antibiotic industry. However, out of 6,000 antibiotics, about 1,000 have
been obtained from just six genera of filamentous fungi (e.g., Penicillium, Cephalosporium)
and about 2,000 from just three genera of bacteria (e.g., Streptomyces, Bacillus).
Sites of Antibiotic Action on Microbial Cells
Properties of useful antibiotic
To be truly useful, an antibiotic must possess following qualities in it :
(i) Antibiotic should be of “broad spectrum”, i.e., it should have the ability to inhibit/kill a
number of different types of pathogenic microorganisms.
(ii) Is should check the development of resistant forms of pathogenic microorganisms.
(iii) It should not result in undesirable side effects in the host.
(iv) It should not disturb the “normal” microbial flora of the host. Such a disturbance may
upset the balance of nature.
Antibiotics of Fungal Origin
Penicillin
Alexander Flemming discovered penicillin secretion by the mould Penicillium notatum in
1929. He reported that a contaminating colony of the fungus lysed adjacent colonies of
staphylococci; but the lytic agent seemed too unstable to be useful. However, when Chain
(1939) purified the active material, called penicillin, it proved remarkably effective in certain
infections.
Penicillin is not a single chemical compound but a group of substances of related structure
and activity. There are six penicillins : Penicillin G (benzyl penicillin V (phenoxymethyl
penicillin), Penicillin F (A2 – pentenyl penicillin), penicillin X (p-hydroxybenzyl penicillin),
penicillin K(n-heptyl penicillin) and penicillion O (allyl-mercaptomethyl penicillin).
Penicillin is selective for Gram-positive bacteria, some spirochaetes and the Gram-negative
diplococci (Neisseria).
Commercial Production
It was World War II that provided stimulus for commercial production (Fig. 17.9) of
penicillin; various strains of Penicillium chrysogenum are used for the purpose. At present,
almost 50 years later, such strains of the fungus are available that yield 104 times more than
the original isolate. Production has also been dramatically increased by improvements in
media and to fermenter design. Phenyl ethanoic acid (phenyl acetic acid) is now routinely
added to the media since this induces the synthesis of a metabolic precursor of penicillin G
(the most active form of penicillin).
To produce penicillin commercially*, deep fermentation tanks with the capacity of several
thousand gallons are filled with fermentation medium consisting of corn steep liquor, nutrient
agar salts, lactose, glucose and phenyl ethanoic acid (phenyl acetic acid). This medium is
inoculated with conidia-suspension of the fungus and is constantly aerated as pellets. The
fermentation is completed in 20 hours. When the fermentation is complete, the masses of
fungal growth are separated leaving a clear fluid (the broth). The broth is passed over a filter
and washed. The filtrate is when mixed with potassium ions, filtered again and dried. The
result is a crystalline potassium salt of penicillin G. Penicillin is assayed to determine its
potency before being bottled and sold.
Vitamins Production
Out of all vitamins now available commercially, vitamin B12 and vitamin B2 (riboflavin) are
the main that are produced by microbial fermentations.
Vitamin B12 (Cyanocobalamine; Cobalbumin)
This vitamin is recovered as a by-product of streptomycin and aureomycin antibiotic
fermentations. A soluble cobalt salt is added to the fermentation reaction as a precursor to
vitamin B12. Relatively high amounts of this vitamin accumulate in the fermentation medium
at concentrations that are not toxic to Streptomyces species.
Vitamin B12 (Cyanocobalamine) is also produced on large scale by direct fermentation.
Propionibacterium shermanii or Pseudomonas denitrificans are the bacteria which are used
now-a-days for fermentation processes. P. shermanii is grown in anaerobic culture for 3 days
at 30°C and in aerobic culture of 4 days; the fermentation medium (growth medium) contains
glucose, corn steep liquor (a waste product of starch manufacture), ammonia and cobalt
chloride. Ammonia is used in the form of ammonia hydroxide that maintains the pH of the
medium at 7. Pseudomonas denitrificans is grown for 2 days in aerobic culture medium
containing sucrose, betaine, glutamic acid, cobalt chloride, 5, 6 dimethylbenzimidazol, and
salts.
The vitamin resulted in the fermentation are retained within the cells. The cells are, therefore,
collected by high-speed centrifugation and the vitamin B12 is recovered by releasing the
vitamin from the cells by treating with acid, heating, cyanide, etc. The vitamin thus released
from the cell is absorbed on ion-exchange resin IRC-50 or charcoal. It is then purified further
by phenol and water. The vitamin is finally crystallized from aqueous-acetone solutions. The
normal yield is 23 mg/litre when Propionibacterium is used.