TOPIC 3: TRADITIONAL AND FOOD FERMENTATION I

Question 1

TEMPEH

Answer 1

• Tempeh is a soyfood made by controlled fermentation of cooked soybeans with a Rhizopus molds (tempeh starter).

• This fermentation binds the soybeans into a compact white cake.

• But it is now rapidly becoming popular all over the world as people look for ways to increase their intake of soy, known for its health benefits.

• Normally, tempeh is sliced or cut in cubes and fried until the surface is crisp and golden brown.

• Tempeh can be used as ingredient in soups, spreads, salads, and sandwiches.

Question 2

Why ferment soybeans?

Answer 2

Soybeans are difficult to digest, owing to the presence of oligosaccharides and trypsin inhibitors, and are also associated with flatulence and indigestion.

They also contain very high levels of phytic acid, a
mineral chelator, which binds magnesium, calcium, and especially zinc, making them difficult for the body to assimilate

Question 3

Types of tempeh

Answer 3

• Tempe bongkrek –made from or with coconut press cake

• Tempeh bosok (busuk) rotten tempeh – used in small amounts as a flavouring

• Tempeh gembus – made form okara

• Tempeh godhong – tempeh made in banana leaves

• Tempeh goring – deep-fried tempeh

• Tempeh mendoan –raw-fried tempeh

• Tempeh kedelai – simply tempeh, made form soybeans

• Tempeh oncom also onchom – made from peanut press cake; orange color

Question 4

Characteristics of tempeh

Answer 4

• Mycelia of the fungus Rhizopus oligosporus mass holds the soybeans together (cake)

• Inexpensive source for dietary protein (19%)

• 15 g/day/person in Indonesia

• Contain vitamin B12

• “Vegetable meat”

• Bland, mushroom-like before cooking à nutty flavorful

Question 5

Rhizopus mould

Answer 5

• Industrial tempeh is made with certain edible strains of Rhizopus oryzae or Rhizopus oligosporus.

• In Indonesia, where tempeh originated, homemade tempeh normally contains a mixtures of both Rhizopus oryzae and Rhizopus oligosporus strains, together with other bacteria.

• They make tempeh starter by incubating boiled soybeans in hibiscus leaves, incubate 2-3 days.

• Inoculation level: 10^7-10^8 spores (~1g/kg of beans)

Question 6

Rhizopus mould (Oligosporus)

Answer 6

• Rhizopus oligosporus is a fungus that belongs to the class of zygomycetes

• Rhizopus oligosporus has large, subglobose to globose spores, and high proportion irregular spores

• Rhizopus oligosporus also has spores with nonparallel valleys and ridges, and plateaus that (sometimes are granular)

Question 7

Rhizopus oligosporus role in tempeh fermentation

Answer 7

• Spores with a tendency for fast germinability are needed, as well.

• In order for the tempeh to attain its characteristic compact ‘cake’ form after fermentation, the soybeans become compressed due to the mycelia of Rhizopus oligosporus.

• Rapidly growing mycelia helps to (increase growth of this fungus).

• Because mycelia are quite sensitive to dehydration and adverse temperatures, preserving tempeh for extended periods of time can be challenging.

• When the soybeans are bound together by the white mycelium, the fungus releases enzymes that make this heavily protein-rich product more digestible for humans.

Question 8

Effects of Rhizopus oligosporus

Answer 8

It is consumed, Rhizopus oligosporous produces an antibiotic limits gram-positive like Staphylococcus aureus (potentially harmful) and Bacillus subtilis (beneficial).

This fungus can also treat waste and wastewater, produce industrial enzymes, and ferment other substrates like other legumes and cereals.

Question 9

Nutritive value of tempeh

Answer 9

• Tempeh is very nutritive and contains many health promoting phytochemicals such as isoflavones and saponins.

• Tempeh fermentation produces natural antibiotic

• Tempeh is a complete protein food that contains all the essential amino acids.

• Protein - High quality

• Nutritional loss -Very low during process

• Rich in Phosphorus (P), Potassium (K), Ferum (Fe)

• Good source of vitamin B (thiamine) and vitamin K

Question 10

Tempeh spoilage and defects

Answer 10

• OK if eaten within a day or two of manufacture

• Otherwise pH high, other bacteria can grow

• Shelf-life short at room temperature (before sporulation)

• Vacuum packing in oxygen impermeable plastic

• Freezing

• Dehydrated or cooked or processed prior to packaging

Question 11

CHEESE

Answer 11

•Cheese is the curd of milk separated from whey and pressed into solid mass

•It is a product made from milk by coagulation of casein with the help of enzyme rennet in the presence of lactic acid produce by Lactobacillus and Streptococcus

•Then moisture is removed and pressed, and then
ripened for some time at a suitable temperature

Question 12

Cheese production

Answer 12

1) Standarization of milk
2) Inoculation of starter culture of lactic acid bacteria
3) Addition of rennet for coagulum formation
4) Shrinkage of curd
5) Salting of curd and pressing into shapes
6) Cheese ripening

Question 13

1) Standarization of milk (Cheese)

Answer 13

• Quality of milk decide the nature of cheese

• Skimmed milk cheese-hard and leathery

• More fat a cheese contain smoother it feels

• If starter culture is slow acting or souring is delayed then Steptococci could produce toxin over cheese

• Pasteurization gives better control over cheese production

• Milk may be homogenized to reduce milk fat globules to produce soft cheese

Question 14

2) Inoculation of starter culture of lactic acid bacteria (Cheese)

Answer 14

• Starter culture for the cheese prepared at less than 40οC –Lactococcus lactis
• Cheese prepared at high temperature - Steptococcus thermophiles, Lactobacillus bulgaricus

Question 15

3) Addition of rennet for coagulum formation

Answer 15

• Casein is converted into fibrous form by rennin

• 1st pH is brought down from 6.8-7 to pH 5.5 by the action of lactic acid bacteria and increasing temperature up to 45 C

• On the addition of rennet active component renin hydrolyse k-casein to para k-casein and k-casein macropeptide

• Para k-casein remain part of casein micelles and bind together to form curd following by removal of carbohydrates with k-casein macropeptide

• Fat and other particulate matter entrapped in curd

• When casein removed remaining liquid protein, lactalbumin, globulin, riboflavin known as whey use for whey cheese by the precipitation of whey
protein by heat

Question 16

Sources of rennet

Answer 16

• The material used in the formation of coagulum is rennet obtained from the stomach of freshly slaughtered calves

• Young calves contain 94% renin and 6% pepsin

• Due to high cost of animal renin, other sources mainly of microbial origin have been discovered.

• For example: Mikrozyme from Bacillus subtilis

Question 17

4) Shrinkage of curd (Cheese)

Answer 17

• The shrinkage of curd is facilitated by heating
it, cut into smaller pieces, applying some pressure on it and lowering the pH

• Lactic acid produced by starter culture
introduce elasticity desirable qualities in cheese

Question 18

5) Salting of curd and pressing into shapes (Cheese)

Answer 18

• Salt is important for the taste of cheese

• Salt contribute to moisture and acidity control

• Helps in limit the growth of proteolytic bacteria
which is undesirable

• Curd is pressed into shape before allowed to
matured

Question 19

6) Cheese ripening

Answer 19

• Except from fresh cheese, the curd is ripened
or matured at various temperature and time
until the characteristics flavor, body, and
texture achieved

• During ripening, there is degradation of lactose
by ripening agent like;

oBacteria and enzyme of milk
oRennet
olipases

Question 20

IDLI

Answer 20

• Indian idli is a small, white, acidic, leavened, steam-cooked cake made by lactic fermentation of a thick batter from polished rice and dehulled black
gram dhal, a pulse (Phaseolus mungo)

• The cakes are soft, moist, and spongy and have a pleasant sour flavor

Question 21

DOSA

Answer 21

• Dosa, a closely related product, is made from the same ingredients, both finely ground

• The batter is generally thinner, and dosa is fried like a pancake

Question 22

IDLI & DOSA Production

Answer 22

L. mesenteroides and S.faecalis develop during soaking, then continue to multiply following grinding.

Each eventually reaches more than 1 x 10^9 cells per gram, 11 to 13 hours after formation of the batter.

These two species predominate until 23 hours
following batter formation

Question 23

WINE MAKING

Answer 23

• Wine making is full of tradition. Each vineyard has its own history and ways of making wine.

• Famous vineyards are in France, Spain, California, New South Wales (Australia)

• The history of wine making started 6000 years ago

• It is thought that wine making was first started in Sourthern Caucasia (Turkey, Northern Iraq and Azerbaijan) and later expanded to the European continents

Question 24

Types of wine

Answer 24

• Red Wines.-Red wines are made from black grapes fermented with the grape skins (which is where the red colour of the wine comes from), seeds, and stems.e.g. Cabernet Sauvignon (dry), Cabernet Merlot (sweet)

• White Wines- e.g. Chardonnay

• Rosé Wines-e.g. Cabernet Sauvignon Rosé, Pinot Noir Rosé

• Sparkling Wines-e.g Champagne

• Dessert Wines-e.g. Port, Madeira

• Fortified Wines-e.g. Sherry, Vermouth

Question 25

What do people look for when drinking wine?

Answer 25

• Flavour-distinct from different districts (warm wine under tongue to get the flavour out)

• Astringency-dry pucker taste

• Bitterness-red wine is more bitter than white wine

• Body-alcohol and sugar content of the wine

• Burning sensation at the back of the throat

• Bubbles bursting in the mouth produce prickling, tingling sensation (sparkling wine)

Question 26

Grape juice (Wine)

Answer 26

• Wine production because it contains high levels of fermentable sugars (mainly glucose and fructose; 160–240 g/L)

• The natural acidity (pH 2.8–3.8) inhibits potential spoilage organisms

• Grapes also have very pleasant flavour and aroma components.

• Most wines are table wines that normally contain up to 14% (v/v) ethanol.

• However, some regions produce wines fortified with ethanol

Question 27

Unique features of grapes

Answer 27

• Grapes store carbohydrates as soluble sugar (glucose, fructose, arabinose, rhamnose), and this is readily metabolised by yeast to ethanol.

• Most other fruits store carbohydrates as starch and pectin, which can not be metabolised by yeast

• The major acid found in mature grapes are tartaric acid. This acid is seldom found in fruits but is commonly found in leaves

Question 28

Wine making process

Answer 28

1) Harvest
2) Destemming and sorting
3) Fermentation
4) Press
5) Aging
6) Bottling

Question 29

1) Harvest (Wine)

Answer 29

• Harvesting is done by hand or mechanically. Once picked, the bunches are taken to the winery and sorted

Question 30

2) Destemming and sorting

Answer 30

• Once sorted, grape bunches need to be destemmed and crushed.

• Mechanical destemming, crushing, and pressing to form the must

• Sophisticated machinery can remove stems without damaging the fruit, and the grapes can then be crushed.

• In white wines, grapes move from crushing to pressing immediately, with little or no contact between the grape skins and juice (earlier removal of skin and seeds)

• For red wine, grape pressing usually doesn’t take place until after fermentation.

• Red wine must consists of the macerated fruit, whereas for white wines, it is only the clarified juice that is fermented.

Question 31

Pectin (Grapes)

Answer 31

• Pectin found in the grape cell wall is responsible for the firmness of grape. The more pectin there is, the less juice is extracted.

• Thus, pectinase are added to improve juice separation from grape fruit.

• When grapes are crushed, polyphenol oxidase (PPO) will be released and SO2 is added immediately to control the polyphenol oxidase (PPO) oxidation which can cause “browning” of wine (Maillard reaction on sugar in grape juice)

Question 32

3) Fermentation

Answer 32

• Before the fermentation step, the juice is aerated, to promote yeast growth and then the process is made anaerobic for CO2 and ethanol production.

• With the absence of oxygen, yeast — wild occurring or inoculated — will convert the sugars of a wine grape to alcohol and carbon dioxide. (The more sugar in the grape, the higher the potential alcohol).

• Alcoholic fermentation of wine is carried out by indigenous yeast or added starter yeast.

Question 33

The fermentation rate is influenced by: (WINE)

Answer 33

-The starter yeast strain used or yeast strains that
make up the natural microflora

-temperature

-pH

-initial sugar concentration

-nutritional components of the batch of grape or supplement.

Question 34

Traditional spontaneous wine fermentations

Answer 34

• Traditional fermentations are comparatively slow. They can also be prone to some flavor and aroma defects due to the over activity of certain yeasts.

• To ensure complete fermentation, and avoid production of unwanted flavourand aroma compounds, winemakers encourage the growth and dominance of S. cerevisiae.

• This may be achieved by adding SO2 to the grape must at a concentration of 30–50 mg/L, which inhibits non- Saccharomyces yeast and spoilage bacteria.

Question 35

Advantages of starter culture (Wine)

Answer 35

• It ensures completion of the fermentation and allows the fermentation rate to be controlled
• To give a shorter lag period, and more even and rapid fermentation rates.
• Also, there is less opportunity for the production of off-flavours by wild yeasts and bacteria, giving more consistent flavor characteristics and quality.

Question 36

Genetically modified yeast (Wine)

Answer 36

The yeast (Saccharomyces ellipsoideus) used are genetically modified to be:

• Alcohol-tolerant

• Easier to sediment for easier separation of yeast from wine

• Able to carry out malolactic fermentation (secondary fermentation) by cloning malolactic genes into yeast

• Resistant to SO2

• When yeast cells die, they sink to the bottom of the fermentation tank and rest there with the skins, seeds and pulp fragments of the grape, creating the lees.

• Resting wine on the lees, or stirring the lees in the fermentation process, adds stability, body and flavor to the wine.

• For red wine, the skins are left for several weeks before being removed.

Question 37

Secondary wine fermentation

Answer 37

• In red wine, secondary fermentation converts malic acid to lactic acid (malolactic fermentation)

• The produced lactic acid will improve the taste of red wine and gives it a special flavor.

• White wine generally do not undergo 2◦ fermentation.

• A bacterial culture such as Leuconostoc oenosis added to induce 2◦fermentation (especially in US where the native flora do not produce 2◦ fermentation)

Question 38

4) Press (Wine)

Answer 38

• Pressing is the process of extracting the grape’s juice.

• White wine uses only the juice of white grapes. After pressing, the juice is clarified to remove the suspended solid (skin)

• For red wine, the skin is kept in the juice to transmit the red color and to enhance the taste of the wine.

• Basket presses are one of the earliest forms of wine presses, and look like a basket or wine barrel with a disc at the top that presses down on the
contents.

• Other press styles include a bladder press, membrane press, moving-head press and continuous

Question 39

5) Aging (Wine)

Answer 39

• Chemical processes take place during aging and react over time, creating different flavors as a wine matures.

• Aging allow tannin compounds to combine with protein and form precipitates, allowing further clarification.

• Pectinase and bentonite can be added to help remove the precipitates (cloud).

• During aging of wine, the color change from red to light brown, and the flavor becomes more complex with the development of bouquet (flavor).

Question 40

Oak barrels

Answer 40

Oak barrels were originally chosen for convenience — oak was prevalent, easy to shape, and had tight
grain for watertight storage.

But it turned out that oak aging made the wine
smooth and added pleasant flavors.

Today, wines are typically aged in French
and American oak.

American white oak is considered more “oaky” and brings a vanilla aroma, where French oak contributions are more subtle and spicy.

Question 41

6) Bottling

Answer 41

• When wine is clear, stable, and “finished,” according to the winemaker, it’s bottled. This can involve state-of-the-art machinery or simple methods.

• Once bottled, the wine is kept for several years to develop rounder and fuller flavor and texture

• Bottling protects the wine — tinted bottles help keep out the light — and then time does the rest.

Question 42

How does one get “sparkling wine”?

Answer 42

• Inject CO2
• Add sugar and yeast to produce CO2 (2◦ fermentation)

Question 43

BEER BREWING

Answer 43

• Beer brewing is the largest biotechnology industry in the world.

• Beer brewing is as old as baking bread. The Sumerians produced an alcoholic beverage from fermented grain

• The Assyrians are known to have brewed beer about 4000 B.C.

• Beer comes from the Latin word “biber” which means “beverage” or “drink”.

Question 44

What is beer?

Answer 44

It is a malt beverage resulting from the alcoholic
fermentation of the aqueous extract of malted barley flavored with hops with or without other cereal grains (adjunct)

Question 45

What are the ingredients of beer?

Answer 45

1) Hop
2) Malt
3) Yeast
4) Water
5) Malt Adjunct

Question 46

What is malt

Answer 46

Malt is a grain that has been moistened and allowed to germinate (bercambah) under controlled
conditions and then dried to a stable form. Malt
contains a lot of enzymes

Question 47

What is the most common malted grain used?

Answer 47

• Barley
• Barley malt is also sometimes called the green malt

Question 48

What are adjuncts?

Answer 48

Adjuncts are (non-malted carbohydrate) containing materials that supplements malt barley such as corn, rice, wheat which are cheaper.

Question 49

Beer brewing process

Answer 49

1) Malting
2) Mashing
3) Production of wort
4) Fermentation
5) Post Fermentation

Question 50

1) Malting (Beer)

Answer 50

• Partial germination of barley grains -allow the formation of highly active α-amylase, β-amylase and proteases enzymes as well as various flavor and color components

• Modify hard vitreous grains into friable with more degradable starch and hydrolytic enzymes

• Soak barley in water for 2 days at 10-16°C

• Increase moisture 45%

• Water drained out

• After steeping barley partially germinated for 3-5 days at 16-19°C

• Spread grains on malting floor

• Aerated with moist cool air

Question 51

2) Mashing (Beer)

Answer 51

• Hot water is added to malt to germinate the malt. This is known as mashing.

• When subjected to hot water at a very specific temperature, those enzyme become active.

• When active, they convert the readily available starch in the malt into sugars (maltose) and dextrins (think mouthfeel).

• The process of mashing will release the enzyme systems in barley such as:

i) Hemicellulase and β-glucanase-degrade cell wall of the endosperm to release the starch
ii) Glucanase-digest hemicellulose

• Dry adjuncts are also added. The starch from dry adjunct will not undergo enzymatic hydrolysis during mashing, so the adjuncts need to be boiled to solubilise and gelatinise the starch.

• Nowadays, liquid adjuncts, which consist of fermentable sugars and dextrins are used so that boiling is not necessary

• The germination of malt is stopped by heat, which also stops the enzymatic process and produce a certain color and flavor to beer

Question 52

3) Production of wort (Beer)

Answer 52

• Production of aqueous fermentation media.

• Reasons for boiling of wort:
• For extraction of hop flavor from hop flower

• Boiling coagulate remaining protein and partially hydrolyze protein and help in removal of protein (precipitate protein)

• Boiling inactivates enzymes that were active during mashing, otherwise causes caramelization of sugar

• Sterilize the liquid

• Concentrate the wort

• Caramelise the sugar

Question 53

4) Fermentation (Beer)

Answer 53

• Wort is then aerated to provide oxygen for the growth of yeast

• Beer production utilize strain of Saccharomyces carlsbergensand S. varum which are bottom yeast and S. cerevisiae which is a top yeast.

• Fermentation is usually carried out at 3-4 °C but it may range from 3- 14°Fermentation usually completes in 14 days.

• Undergoes alcoholic fermentation

• The yeast metabolises the sugar and amino acid to produce CO2, ethanol and other products like diacetyl and diketone that give beer flavor.

• CO2 evolution is maximum by fifth day of fermentation. Last for 2-7 days because yeast cells become inactive and flocculate.

Question 54

5) Post Fermentation

Answer 54

• Maturation of beer (at 0°C for several weeks to several month)-precipitation of protein, yeast, resin and other undesirable substances take place and beer become clear.

• Ester and other compounds are also produced during ageing which gives taste and aroma.

• Condition for consumption

• Cask and bottle conditioning of ales

• Krauening

• Lagering

• Storage ageing

• Prevention of oxidation