PROSES PRODUKSI BIOINDUSTRI

(PROSES FERMENTASI)

Prof. Nyoman Semadi Antara, Ph.D.

The Potential of Microbial Cell

In natural environment, microbial cells will almost always be in mixed cultured. They

have to interact each other, but these interaction are limited by the potential of

each cell.Each species of the environmental microbe has specific potential and some of them are

potential to produce useful industrial products.

The Potential…

The range of Microorganisms The Potential…

The range of Microorganisms

The microorganisms encompass the three groups:

Viruses, Prokaryotes, and EukaryotesOther microbiologists claim three Kingdoms of five Kingdoms of cellular life are microorganisms, namely:Monera : bacteria (procaryotae) and blue green algaeProtista : protozoa, eukaryotic algae, slime

moulds and flagellated fungiFungi : non-flagellated fungi

Viruses

It is not having a cell structure

It is dependent on its host’s metabolic machinery

Its structure of nucleic acid, DNA or RNA, is surrounded by protein and sometimes an outer lipid-rich envelope

Prokaryotes – the bacteriaProkaryotes – the bacteria Purple bacteria and the green bacteria

Gliding bacteria Sheathed bacteria

Spirochaetes Spiral and curve bacteria

Gram-negative aerobic rods and cocci Gram-negative facultative anaerobic rods

Gram-negative anaerobic bacteria Methane producing bacteria

Gram-positive cocci Gram-positive endospore-forming rods and cocci

Group of Lactobacillaceae Actinomycetes

Rickettsias Mycoplasmas

Cyanophyceae or Cyanobacteria

Purple bacteria and the green bacteria Gliding bacteria

Sheathed bacteria Spirochaetes

Spiral and curve bacteria Gram-negative aerobic rods and cocci

Gram-negative facultative anaerobic rods Gram-negative anaerobic bacteria

Methane producing bacteria Gram-positive cocci

Gram-positive endospore-forming rods and cocci Group of Lactobacillaceae

Actinomycetes Rickettsias

Mycoplasmas Cyanophyceae or Cyanobacteria

…the bacteria

The students are divided into several groups and each group…

…discuss these grouping of prokaryote microorganisms (the bacteria) and the

beneficial of them in industrial purposes.

…the bacteria…the bacteria

Bacteria are unicellular, most ca. 0.5-1.0 x 2.0-10 µm in size.

They can be motile or non-motile Cytoplasmic materials are enclosed in a rigid wall

on the surface and a membrane beneath the wall, and they are immobile.

The membrane contains energy generating components.

The genetic materials (structural and plasmid DNA) are circular, not enclosed in nuclear membrane, and do not contain basic protein such as histones.

Cell division is by binary fission. Can also have flagella, capsules, surface layer

protein, and pili for specific function. Some also form endospores (one per cell) Gram-positive cells or Gram-negative cells

Bacteria are unicellular, most ca. 0.5-1.0 x 2.0-10 µm in size.

They can be motile or non-motile Cytoplasmic materials are enclosed in a rigid wall

on the surface and a membrane beneath the wall, and they are immobile.

The membrane contains energy generating components.

The genetic materials (structural and plasmid DNA) are circular, not enclosed in nuclear membrane, and do not contain basic protein such as histones.

Cell division is by binary fission. Can also have flagella, capsules, surface layer

protein, and pili for specific function. Some also form endospores (one per cell) Gram-positive cells or Gram-negative cells

…the bacteria

If we look at the shape and size, called morphology, it is more simple to use them for grouping the bacteria. The most common shapes are rod-like, bacillus (plural: bacilli), and sperichal, coccus (plural: cocci)The rods form vary from short rods (almost look like cocci) to very long filaments. Also form spiral and corkscrew, oval (coccoid), comma, and branch structure.

Shape of bacteriaShape of bacteria

EukaryotesEukaryotesEukaryotic cells:

are generally much larger than prokaryotic cells have rigid cell walls and thin plasma membranes

(contain sterol) the cell wall does not have mucopeptide and is

composed of carbohydrates the cytoplasm is mobile (streaming) and contain

organelles the DNA is linear (chromosomes), contains

histones, and is enclosed in a nuclear membrane.The eukaryotic microorganisms are:

FungiAlgaeProtozoa

Eukaryotic cells: are generally much larger than prokaryotic cells have rigid cell walls and thin plasma membranes

(contain sterol) the cell wall does not have mucopeptide and is

composed of carbohydrates the cytoplasm is mobile (streaming) and contain

organelles the DNA is linear (chromosomes), contains

histones, and is enclosed in a nuclear membrane.The eukaryotic microorganisms are:

FungiAlgaeProtozoa

Eukaryotes - fungiEukaryotes - fungi

Yeasts – unicellularMolds – multicellular

Molds are non-motile, filamentous and branches the cell wall is composed of cellulose, chitin, or

both are composed of hyphae (large number of

filaments), an aggregate of hyphae called mycellium.

a hyphae can be vegetative or reproductive, the reproductive hyphae usually extend and form exospores, either free (conidia) or in sack (sporangium). Shape, size and color of spores are used for taxonomic classification.

Yeasts – unicellularMolds – multicellular

Molds are non-motile, filamentous and branches the cell wall is composed of cellulose, chitin, or

both are composed of hyphae (large number of

filaments), an aggregate of hyphae called mycellium.

a hyphae can be vegetative or reproductive, the reproductive hyphae usually extend and form exospores, either free (conidia) or in sack (sporangium). Shape, size and color of spores are used for taxonomic classification.

… fungi… fungi

Yeast the cells are oval, spherical, or elongated

(5-30 x 2-10 µm), non-motile. the cell wall contains polysaccharides

(glycans), protein and lipids. the cytoplasm has a finely granular

appearance for ribosomes and organelles. the nucleus is well defined with nuclear

membrane

Yeast the cells are oval, spherical, or elongated

(5-30 x 2-10 µm), non-motile. the cell wall contains polysaccharides

(glycans), protein and lipids. the cytoplasm has a finely granular

appearance for ribosomes and organelles. the nucleus is well defined with nuclear

membrane

…fungi

Eukaryotes - algaeEukaryotes - algae

The algae are photosynthetic eukaryotic microorganisms.

The major primary producers in the sea and in lakes, but are also found in the surface layer of soil.

Many can live heterotrophically.

The algae are photosynthetic eukaryotic microorganisms.

The major primary producers in the sea and in lakes, but are also found in the surface layer of soil.

Many can live heterotrophically.

…microalgae

…algae…algae

The algae have been classified traditionally by pigmentation and life cycle:

› Rhodophyceae, the red algae: marrine, multicellular, immotile, some unicellular

› Chlorophyceae: many planktonic species, freshwater and marine, motil by flagella,multicellular.

› Prasinophyceae: unicellular planctonic flagellated organisms, marine.

› Euglenophyceae: unicells, motile by single flagellum, common in nutrition rich freshwater pools, also found in the sea and in the soil.

› Bacillarophyceae, diatoms: in mcroplankton of sea and lakes, motile.

› Dinophyceae, dinoflagellates: in microplankton of seas and lake s, motile, some are non photosynthetic.

› Crysophyceae: found in freshwater, also the marine silicoflagellates, biflagellate cells.

› Haptophyceae: biflagellate planktonic algae, in marine, blooms of this species can give the sea a milky appearance.

› Cryptophyceae: unicellular flagellates found as minor components of plankton.

The algae have been classified traditionally by pigmentation and life cycle:

› Rhodophyceae, the red algae: marrine, multicellular, immotile, some unicellular

› Chlorophyceae: many planktonic species, freshwater and marine, motil by flagella,multicellular.

› Prasinophyceae: unicellular planctonic flagellated organisms, marine.

› Euglenophyceae: unicells, motile by single flagellum, common in nutrition rich freshwater pools, also found in the sea and in the soil.

› Bacillarophyceae, diatoms: in mcroplankton of sea and lakes, motile.

› Dinophyceae, dinoflagellates: in microplankton of seas and lake s, motile, some are non photosynthetic.

› Crysophyceae: found in freshwater, also the marine silicoflagellates, biflagellate cells.

› Haptophyceae: biflagellate planktonic algae, in marine, blooms of this species can give the sea a milky appearance.

› Cryptophyceae: unicellular flagellates found as minor components of plankton.

…algae…algae

algae… classified traditionally…:–Xanthophyceae: multicellular, but few unicellular in plankton and soil.–Eustigmatophyceae: unicellular, in freshwater, sometime found in soil.

Among aquatic microorganisms microalgae are a very interesting source of a wide range of compounds. They do not only have the capacity to produce high-value compounds, but also the ability to do it using only sunlight, carbon dioxide and sea water.

algae… classified traditionally…:–Xanthophyceae: multicellular, but few unicellular in plankton and soil.–Eustigmatophyceae: unicellular, in freshwater, sometime found in soil.

Among aquatic microorganisms microalgae are a very interesting source of a wide range of compounds. They do not only have the capacity to produce high-value compounds, but also the ability to do it using only sunlight, carbon dioxide and sea water.

…many algae produced commercially as source of nutrition and others functional compounds

…many algae produced commercially as source of nutrition and others functional compounds

Peptides and protein: lectins (glycoprotein) have proved to be useful for clinical diagnosis and other health application. Red algae (Bryothamnion triquetrum, Solieria robusta, Ceratodiction spongiosum)

-carotene/canthaxanthin and astaxanthin: green-algae, Haematococcus pluvialis.

Seaweed polysaccharides Lipid – polyunsaturated fatty acid

Peptides and protein: lectins (glycoprotein) have proved to be useful for clinical diagnosis and other health application. Red algae (Bryothamnion triquetrum, Solieria robusta, Ceratodiction spongiosum)

-carotene/canthaxanthin and astaxanthin: green-algae, Haematococcus pluvialis.

Seaweed polysaccharides Lipid – polyunsaturated fatty acid

Eukaryotes - protozoaEukaryotes - protozoa Exhibit a very wide range of form and way of life.

Many are predators on bacteria, fungi, algae, yeast, or other protozoa, while some others are parasitic in animals.

Sarcomastigophora: the flagellated cells and amoeboid protozoa, some are human pathogens such as Trypanosoma brucei in the bloodstream, and Entamoeba histolytica in the gut. Some flagellates are symbionts in termite guts.

Ciliphora: includes the familiar ciliates, Tetrahymena and Paramecium. Vorticella species are very important in sewage treatment processes.

Sporozoa: totally endoparasitic in animals or man, Plasmodium cause malaria.

Exhibit a very wide range of form and way of life.

Many are predators on bacteria, fungi, algae, yeast, or other protozoa, while some others are parasitic in animals.

Sarcomastigophora: the flagellated cells and amoeboid protozoa, some are human pathogens such as Trypanosoma brucei in the bloodstream, and Entamoeba histolytica in the gut. Some flagellates are symbionts in termite guts.

Ciliphora: includes the familiar ciliates, Tetrahymena and Paramecium. Vorticella species are very important in sewage treatment processes.

Sporozoa: totally endoparasitic in animals or man, Plasmodium cause malaria.

Factors that influence microbial growth – Intrinsic FactorsFactors that influence microbial growth – Intrinsic Factors

Factors inherent to the media/substrate of the growth are considered intrinsic factors which may stimulate or retard the growth of microbial, including:› pH (acidity)› Water Activity (moisture)› Oxidation-reduction potential (oxygen

or ionic)› Nutrition (food)

Factors inherent to the media/substrate of the growth are considered intrinsic factors which may stimulate or retard the growth of microbial, including:› pH (acidity)› Water Activity (moisture)› Oxidation-reduction potential (oxygen

or ionic)› Nutrition (food)

…protozoa

…extrinsic factors…extrinsic factors

Environment factors that influence microbial growth are considered extrinsic factors, such as:› Temperature

Each species of microbe has an optimal temperature of growth

Temperature regulate the expression of gene The growth temperature can also influence a cell’s

thermal sensitivity.› Gas composition

It relates to the oxygen concentration. Many microbes are inhibited in low concentration or without of oxygen, but some may grow even though in the absent of oxygen.

It is applied in food preservation using controlled or modified atmosphere storage.

Environment factors that influence microbial growth are considered extrinsic factors, such as:› Temperature

Each species of microbe has an optimal temperature of growth

Temperature regulate the expression of gene The growth temperature can also influence a cell’s

thermal sensitivity.› Gas composition

It relates to the oxygen concentration. Many microbes are inhibited in low concentration or without of oxygen, but some may grow even though in the absent of oxygen.

It is applied in food preservation using controlled or modified atmosphere storage.

Growth kineticsGrowth kinetics

Four phase of microbial growth:

Fase lag Fase logaritmik

Fase stasioner Fase kematian

Waktu

LogTotalmikroba

…growth kinetics…growth kinetics

During the log, or exponential, growth phase microbes (bacteria) reproduce by binary fision. Thus, during this phase, first-order reaction can be used to describe the change in cell number

The number of microbes (N) at any time is directly proportional to the initial number of microbes (No)

The microbiologists frequently use td to describe growth rates of microbes and µ to describe specific growth rates

During the log, or exponential, growth phase microbes (bacteria) reproduce by binary fision. Thus, during this phase, first-order reaction can be used to describe the change in cell number

The number of microbes (N) at any time is directly proportional to the initial number of microbes (No)

The microbiologists frequently use td to describe growth rates of microbes and µ to describe specific growth rates

…death kinetics…death kinetics

The killing of microbes by energy input, acid, bacteriocin, and other lethal agents is also governed by first-order kinetics

By this kinetic reaction, it can be predicted the number of microbes (viable cells) remaining after treatment, such as in sterilization process.

In microbiology, D value (amount of time required to reduce No by 90%) is the most frequently used as kinetic constant. D values are inversely proportional to the rate constant, k, for a given temperature.

The killing of microbes by energy input, acid, bacteriocin, and other lethal agents is also governed by first-order kinetics

By this kinetic reaction, it can be predicted the number of microbes (viable cells) remaining after treatment, such as in sterilization process.

In microbiology, D value (amount of time required to reduce No by 90%) is the most frequently used as kinetic constant. D values are inversely proportional to the rate constant, k, for a given temperature.

First-order kinetics can be used to describe exponential growth and inactivationFirst-order kinetics can be used to describe exponential growth and inactivation

Growtha Thermal inactivationb Iradiationc

N = Noeµt N = Noe-kt N = Noe-D/Do

2.3log(N/No) = µ∆t 2.3log(N/No) = -(k∆t)

∆t = [2.3 log(N/No)]/µ ∆t = -[2.3 log(N/No)]/k

td = 0.693/µ D = 2.3/k

Organism µ (h-1) td (h)

Bacteria

Optimal conditions 2.3 0.3

Limited nutrients 0.20 3.46

Psychotroph, 5oC 0.023 30

Molds, optimal 0.1 – 0.3 6.9 – 20

aN, cell number (cfu/g); No, initial cell number (cfu/g); t, time (h); µ, specific growth rate (h-1); td, doubling time (h); bk, rate constant (h-1); D, decimal reduction time (h); cDo, rate constant (h-1); D, dose (Gy)

Cell NutritionCell Nutrition The availability of suitable nutrients is clearly a

major factor determining whether the microbes will grow or not in a particular environment.

There are four useful ways to classify potential nutrient mlocules:› As esential, or useful but dispensable› Used as building blocks for macromolecules, or as

energy sources, or as both› As macronutrient required in large quantities, or as

micronutrients› As macromolecules requiring breakdown before entry to

the cell, or small molecules readily entering as soluble nutrients.

The major elements of the cell are carbon, hydrogen, oxygen, nitrogen, sulphur and phosphorous.

The availability of suitable nutrients is clearly a major factor determining whether the microbes will grow or not in a particular environment.

There are four useful ways to classify potential nutrient mlocules:› As esential, or useful but dispensable› Used as building blocks for macromolecules, or as

energy sources, or as both› As macronutrient required in large quantities, or as

micronutrients› As macromolecules requiring breakdown before entry to

the cell, or small molecules readily entering as soluble nutrients.

The major elements of the cell are carbon, hydrogen, oxygen, nitrogen, sulphur and phosphorous.

Cell NutritionCell Nutrition Carbon dioxide is utilized as sole carbon source

only by autotrophs. Carbohydrates are commonly utilized as sources of carbon. Organic acids are readily used directly as sources of carbon by most microbes. Protein and their constituent organic acids are utilized as carbon sources by proteolitic microbes.

Nitrogen is abundant as gaseous dinitrogen, but only few prokaryotic organisms and some of blue-green algae can utilize it. Other sources of nitrogen, such as nitrate, amonia, amino acids, nucleotides, uric acid and urea, can also provide cells’ requirements for nitrogen.

Some organisms can utilize hydrogen sulphide as source of sulphur. Organic sulphur as amino acids cysteine and methionine can also be used.

Probably all organisms can utilize soluble inorganic phosphate.

Carbon dioxide is utilized as sole carbon source only by autotrophs. Carbohydrates are commonly utilized as sources of carbon. Organic acids are readily used directly as sources of carbon by most microbes. Protein and their constituent organic acids are utilized as carbon sources by proteolitic microbes.

Nitrogen is abundant as gaseous dinitrogen, but only few prokaryotic organisms and some of blue-green algae can utilize it. Other sources of nitrogen, such as nitrate, amonia, amino acids, nucleotides, uric acid and urea, can also provide cells’ requirements for nitrogen.

Some organisms can utilize hydrogen sulphide as source of sulphur. Organic sulphur as amino acids cysteine and methionine can also be used.

Probably all organisms can utilize soluble inorganic phosphate.