APPROVAL SHEET

The complete report of experiment Basic Microbiology with title “The

Influence of Environmental Factor to the Microbial Growth” created by
Name : Mutmainna Ekawati
Reg. No : 071404189
Group : IV (Forth)
Have been checked and consulted by assistant and assistant coordinator and this
report is accepted.
Makassar, January 2010
Assistant Coordinator, Assistant,

Rezky Amelia Waji, S. Si. Sulfianto

061 414 025

Known by
Lecturer of responsibility

Prof. Dr. Ir. Yusminah Hala

NIP: 1961 12 12 1986 01 20 02
 CHAPTER I
INTRODUTION

A. Background
A simple and specific medium consisting of chitosan, trypticase, Tween-80
and agar is devised to isolate the organisms directly from the clinical specimens and
to produce germ tubes. The isolation of independent bacterial species from various
environmental sources is important in all branches of microbiology since bacteria are
ubiquitous and live in microbial communities of mixed populations. Populations in
microbial communities or ecosystems may interact and cooperate in their efforts to
obtain nutrients from the environment with the waste products from one group of
microorganisms serving as nutrients for another. Alternatively, the metabolic wastes
from fermentation processes of some bacteria in a population (for example acid
production) may provide a favorable ecological niche for bacteria that prefer to grow
at low pH.
New appreciation of the importance of cellular heterogeneity, coupled
with recent advances in technology, has driven the development of new tools and
techniques for the study of individual microbial cells. Because observations made at
the single-cell level are not subject to the "averaging" effects characteristic of bulk-
phase, population-level methods, they offer the unique capacity to observe discrete
microbiological phenomena unavailable using traditional approaches. As a result,
scientists have been able to characterize microorganisms, their activities, and their
interactions at unprecedented levels of detail.
All living things need nutrient to their growth and development. Nutrient
is main material that is used to build new cellular components and to produce energy
that is needed in the cell life process. To stimulate the microbe’s growth the media
that is used must contain components that are needed by those microbes. Growth and
death rates of microorganisms are greatly influenced by number of environmental
factors such as solutes and water acidity, temperature, oxygen requirement, pH,
pressure and radiation. To prove that statement we must to do this experiment.
B. Purpose
This experiment was done to know the influence of environmental factors
to the microbial growth.
C. Benefit
The benefits of this experiment are:
1. As literature reference to the next observation.
2. As materials to the next scientist.
 CHAPTER II
PREVIEW OF LITERATURE

Bacteria live within the widest temperature range in all organisms. Some
species live at ocean depths of 27m3 meters where the temperature is close to 350ºC.
They have been found growing in Artic ice in the atmosphere. Bacteria are present in
the nose, intestines, skin, and in the air you breathe. One reason bacteria can survive
in so many environments is that some bacteria do not require oxygen. Most species of
bacteria need oxygen to live. They are called aerobic bacteria. Bacteria that do not
need oxygen to live are called anaerobic (Daniel, 1987: 94).
Microbial growth is greatly affected by chemical and physical nature of
their surroundings instead of variations in nutrient levels and particularly the nutrient
limitation. For successful cultivation of microorganisms it is not only essential to
supply proper and balanced nutrients but also it is necessary to maintain proper
environmental conditions. Thus, understanding of environmental influences on the
growth of microorganisms becomes mandatory. As bacteria shows divers food habits,
it also exhibits diverse response to the environmental conditions. Growth and death
rates of microorganisms are greatly influenced by number of environmental factors
such as solutes and water acidity, temperature, oxygen requirement, pH, pressure and
radiation (Anonima, 2010).
Bacterial growth usually refers to an increase in the number of bacteria,
rather than to an increase in the size of an individual cell. A colony is a large group of
bacteria, such as that grown on a nutrient plate in a laboratory. All the members of a
colony are descendants of a single bacterium. All bacteria need food and water and
many need oxygen. Other factors that influence growth are temperature, sunlight, and
chemicals. Limiting these growth factors helps prevent food spoilage due to bacterial
action (Graham, 1986: 302-303).
According to Anonimb (2010), knowledge of the factors that favor or
inhibit the growth of the microorganisms is important in understanding the principles
of food six major factors: 1. Moisture 2. Oxygen¬spoilage and preservation. 3.
Temperature 4. Acidity, pH 5. Nutrients 6. Growth inhibitors
1. Effect of moisture on microbial growth, water is essential for the growth of all
living organisms. Has many important functions in the growth of microoranisms
and in enzyme activity. Preserving techniques such as dehydration,
concentration, freezing¬ Mainly¬are based on making water unavailable for the
microorganisms. 3 methods for making water unavailable: 1. Increasing the
solute concentration; removing water, adding of solutes (salt, sugar) 2. Addition
of hydrophilic (water-binding) colloids (gels, pectins, gums) 3. Bringing water to
a solid phase (freezing)
2. Osmotic Effect, water is transported into and out of the¬ microbial ¬cell
through the semi-permeable cell membrane (wall) by osmosis. If solute¬Water
passes from the less dense medium to the more dense. Concentration of the food
(growth medium of microorganisms) is increased water flows from the microbial
cell to its surrounding. Microbial cell shrivels (shrinks) when osmoregulatory
capacity of the cell is exceeded (plasmolysis). Shrinking of cells retards the
metabolic activity.
3. Existance of water in foods, bound water: An integral part of the living tissue,
vital to all Free water: Exists in and around¬processes associated with the cell.
The tissue or cell. Important for metabolism and survival of Amount of water
available for microorganims is¬microorganisms. Expressed in terms of water
activity. aw = Po/Pow (ERH) at a given T, 0.0 < aw < 1.0
Temperature, especially on land, is an important ecological factor.
Bacteria can live within the greatest temperature range. Some bacteria form
endospores and can withstand temperatures above the boiling point and far below the
freezing point of water. For this reason, bacteria are found almost everywhere. Some
sores and seeds can survive temperatures that would kill a mature plant. In this way,
the species survives extreme temperatures (Oram: 1986: 648).
 CHAPTER III
OBSERVATION METHOD
 CHAPTER IV
OBSERVATION RESULT AND DISCUSSION
B. Discussion
In our experiment we just use 3 environmental factors; they are
temperatures, acidity (pH), and toxic chemist compound.
1. The influence of temperature
Based on our group data, seemed that in the temperature 5ºC, the Nutrient
Broth that contaminated with Escherichia coli still transparent, found thin sediment
and sheath, and there is not found foam. In the temperature 30 ºC the medium change
to turbid, has no foam, has thick sediment, and still has sheath. While in the
temperature 50 ºC the medium become transparent, has no sediment and foam.
From the explanation above, we can see the turbidity of the medium is
always changing. In the 5 ºC, the medium is transparent because the microbe is in
desired condition. The enzyme is still in inactive condition, it can be seen by the
sediment that found in the top of tube. In the 30 ºC, the medium become turbid. It
signs that the enzymes start to active. In the temperature 50 ºC, the medium become
transparent. It signs that the denaturation of enzyme is has done. Most of the others
group has similar data wit us.
Environmental temperature is one of the most important factors affecting
the growth rate of microbes. There is a minimum temperature, below which growth
does not occur. As we rise above the minimum, rate of growth increases in
accordance with the laws governing the effect of temperature on the chemical
reactions that make up growth. These reactions are mostly enzyme catalysed.
However, a point is reached the optimum temperature when there is also a very rapid
increase the rate of inactivation of heat sensitive cell components, like enzymes,
ribosomes, DNA, membranes etc. Above an optimum temperature, this heat
denaturation will occur so rapidly that there is a corresponding rapid drop in the rate
of growth to give a maximum temperature for growth for that particular
microorganism. Most microbes are capable of growth in a temperature range of 20-
3O"C. Most microorganisms have a growth optimum between 20 and 40°C and are
called mesophilic. Those inhabiting cold environments such as polar areas can prow
at much lower temperatures (Anonima, 2010).
2. The Influence of Acidity (pH)
In this experiment we use pH 3, 7, and 9 to determine the microbe that can
be found. In the pH 3, the NB is transparent, has no sediment and foam. In the pH 7,
the medium become turbid, much sediment has sheath and has no foam. While in the
pH 9 the medium become rather turbid, little sediment, has sheath and has no foam.
Increasing the acidity of foods, either through fermentation or the addition
of weak acids, has been used as a preservation method since ancient times. In their
natural state, most foods such as meat, fish, and vegetables are slightly acidic while
most fruits are moderately acidic. A few foods such as egg white are alkaline. It is
well known that groups of microorganisms have pH optimum, minimum, and
maximum for growth in foods. As with other factors, pH usually interacts with other
parameters in the food to inhibit growth. The pH can interact with factors such as aw,
salt, temperature, redox potential, and preservatives to inhibit growth of pathogens
and other organisms. The pH of the food also significantly impacts the lethality of
heat treatment of the food. Less heat is needed to inactivate microbes as the pH is
reduced (Anonimc, 2010).
Same with the range of temperature, the pH also has role to the bacterial
growth. It can be seen from the sediment that found. In the neutral condition, the
enzyme of bacterial is work optimally so we can see in the pH 7 the medium become
turbid. It condition also found in the others group.
3. The influence of toxic chemist compound
In this experiment, we use many kinds of materials like soap, sulfur soap,
medicines, and tooth paste. But in this case, we just found 4 data, and the others can
be said failed.
For the first, the paper dish was submerging to the Amoxylin (material).
This material has different dilution there are 30%, 50% and 100%. After 60 minutes,
the paper dish was taken in the surface of medium. We used 2 methods (pour and
spread method). After incubated for 24 hours, we found the zone area. In the spread
method, especially in the dilution 30% the shortest zone is 2.1 cm while the longest is
3.4 cm, in the dilution 50%, the shortest zone is 2 cm while the longest zone is 2.9
cm. in the 100% solution, the shortest zone is 1.8 cm while the longest zone is 2.5
cm. while if each of them are decreased with the diameter of paper dish, so the result
in order from 30%, 50% and 100% are 2.6 cm, 2.3 cm and 2.2 cm. In this, the pour
method is failed.

The Amoxylin form zone area in the medium (pour

method) that contaminate with Stapylococcus
aureus.

The next is using Resik V and Candida

albicans. The paper dish is also submerged in the Resik V for 60 minutes. After
incubated for 24 hours, we found the zone area. In the spread method, especially in
the dilution 30% the shortest zone is 2.0 cm while the longest is 3.0 cm, in the
dilution 50%, the shortest zone is 1.5 cm while the longest zone is 2.0 cm. in the
100% solution, the shortest zone is 2.0 cm while the longest zone is 2.5 cm. while if
each of them are decreased with the diameter of paper dish, so the result in order from
30%, 50% and 100% are 2.35 cm, 1.6 cm and 2.1 cm. In the pour method, in the
dilution 30% the shortest zone is 1.7 cm while the longest is 3.2 cm, in the dilution
50%, the shortest zone is 1.0 cm while the longest zone is 2.3 cm. in the 100%
solution, the shortest zone is 2.0 cm while the longest zone is 2.5 cm. while if each of
them are decreased with the diameter of paper dish, so the result in order from 30%,
50% and 100% are 2.3 cm, 1.5 cm and 2.5 cm. this data indicate the pour and spread
method just has little different.
 In the tooth paste (pepsodent) we also found zone area. In the 50% the
shortest zone is 1.5 cm while the longest zone is 2.2 cm. in the 100% solution, the
shortest zone is 1.5 cm while the longest zone is 2.0 cm. while if each of them are
decreased with the diameter of paper dish, so the result in order from 50% and 100%
are 1.7 cm and 1.6 cm. in this the spread method is failed.

From the picture seem the zone is entered by

microbial, it can be happened because there is
another microbe except the Staphylococcus aureus
that contaminate with the medium.

Halogen is group of chemist compound that mostly used as preventive

materials for microbes. Halogen is strong oxidation, so finally this character very
effective in damage the important compound in the microbial cell. Iodine is one of
microbial preventive that nice to use (Hala, 2009: 54).
Added antimicrobial compounds can have an interactive or
synergisticeffect with other parameters of the formulation. One example is the
interaction with pH. Many preservatives have an optimum pH range for effectiveness.
Other factors include aw, presence of other preservatives, types of food constituents,
presence of certain enzymes, processing temperature, storage atmosphere, and
partition coefficients. The effective use of combinations of preservatives with other
physicochemical parameters of a food formulation can stabilize that food against
spoilage organisms or pathogens. Leistner systematically developed the “hurdle
concept” to describe these effects. The hurdle concept states that several inhibitory
factors (hurdles), while individually unable to inhibit microorganisms, will,
nevertheless, be effective in combination. A classic example of applying the hurdle
concept is the antibotulinal stability of certain shelf-stable processed cheese
formulations. Combinations of moisture, total salt, and pH have been shown to allow
for the safe storage of these products at room temperature for extended time even
though the individual factors, taken singly, would not support that practice. In
combination products, the effectiveness of an antimicrobial may be altered by other
factors including the potential for migration of the antimicrobial to other components
of the food and the different food parameters at the interface areas (Anonimc, 2010).
Many chemical compounds selectively inhibit microbial activity These
compounds act either by inactivating a necessary enzyme needed¬ for growth, by
denaturing the protein portion of a cell or by causing physical damage to certain parts
of the cell like breaking the cell A microrganism growing in food may produce
substances¬wall. inhibitory to other organisms or to destroy compounds inhibitory to
its growth. Basis for production of antibiotics. Example: Pencillin is produced by
moulds (aspergillus) and retards growth of bacteria (Anonima, 2010)
 CHAPTER V
CONCLUSION AND SUGGESTION

A. Conclusion
Based on the observations that have done, we can conclude that the
microbial growth can be influenced by the environmental factors like the
temperatures, acidity (pH) and the toxic chemist compound.
B. Suggestion
. 1. Ought to all assistant come at the practice day, so can guide the practical for
each groups.
2. Ought to the practical came in the laboratory no late, must be discipline.
3. Ought to keep the cleaning the laboratory and ought to comply the rule, when
after equable and difficult for using the tools, in order that observation, the
tools when has used is cleaned and saved for it’s place.
 BIBLIOGRAPHY

Anonima. 2010. Factors that Influence Microbial Activity.

http://www.scribd.com/doc/17275885/5268-Factors-That-Influence-
Microbial-Activity. Accessed on January 02nd 2010.

Anonimb. 2010. The Influence of Environmental Factors on Growth.

http://www.studentsguide.in/microbiology/microbial-nutrition-
growth/the-influence-of-environmental-factors-on-growth.html.
Accessed on January 02nd 2010.

Anonimc. 2010. Factors that Influence Microbial Activity.

http://www.publichealthgreybruce.on.ca/FoodSafety/Temperature_Etc/
MicrobialGrowthInfluences.html. Accessed on January 02nd 2010.
.

Daniel, Lucy. 1987. Focus on Life Science. Merrill Publishing Company. United
States of America.

Graham, Linda. 1986. Biology. Harcourt Brace Jovanovich. New York.

Hala, Yusminah. 2009. Penuntun Praktikum Mkrobiologi Dasar. Laboratorium

Mikrobiologi Jurusan Biologi UNM. Makassar

Oram. 1986. Biology Living Systems. Merrill Publishing Company. London.

Ini adalah tembolok Google' untuk http://www.scribd.com/doc/17275885/5268-Factors-That-Influence-
Microbial-Activity. Gambar ini adalah jepretan laman seperti yang ditampilkan pada tanggal 24 Des
2009 17:09:26 GMT. Sementara itu, halaman tersebut mungkin telah berubah. Pelajari Selengkapnya
Factors that influence microbial activity
¬ A knowledge of the factors that favor or inhibit the growth of the microorganisms is important in understanding
the principles of food Six major factors: 1. Moisture 2. Oxygen¬spoilage and preservation. 3. Temperature 4.
Acidity, pH 5. Nutrients 6. Growth inhibitors
Effect of moisture on microbial growth
¬ Water is essential for the growth of all living organisms. Has many important functions in the growth of
microoranisms and in enzyme activity. Preserving techniques such as dehydration, concentration, freezing¬
Mainly¬are based on making water unavailable for the microorganisms. 3 methods for making water unavailable:
1. Increasing the solute concentration; removing water, adding of solutes (salt, sugar) 2. Addition of hydrophilic
(water-binding) colloids (gels, pectins, gums) 3. Bringing water to a solid phase (freezing)
Osmotic Effect
Water is transported into and out of the¬ microbial ¬cell through the semi-permeable cell membrane (wall) by
osmosis. If solute¬Water passes from the less dense medium to the more dense. concentration of the food
(growth medium of microorganisms) is increased water flows from the microbial cell to its surrounding.
Microbial cell shrivels (shrinks) when osmoregulatory capacity of the cell is exceeded (plasmolysis). Shrinking of
cells retards the metabolic activity.
(0.85% salt)
Existance of water in foods
¬ Bound water : An integral part of the living tissue, vital to all Free water : Exists in and around¬processes
associated with the cell. the tissue or cell. Important for metabolism and survival of Amount of water available
for microorganims is¬microorganisms. expressed in terms of water activity. aw = Po/Pow (ERH) at a given T,
0.0 < aw < 1.0
Effect of oxygen on microbial growth
Essential element for the metabolic¬ activity Free atmospheric oxygen in air. Some organisms can utilize
oxygen in¬ Microorganisms are classified according to their oxygen¬C-H-O requirements: aerobic : grow in the
presence of atm. oxygen anaerobic : grow in the absence of atm. oxygen facultatively anaerobic : grow either in
absence or presence of oxygen microaerophilic : grow in the presence of limited oxygen
In the presence of atmospheric oxygen¬ microbial activity leads to a slow oxidation of foods: Carbohydrates →
CO2 and H2O Amino In the¬acids → NH3 , CO2 and H2O Fats → free fatty acids, CO2 and H2O presence of
limited oxygen oxidation results in intermediary products; alcohol, lactic acid, acetic acid, formic acid. C6H12O6
+ 6O2 → 6CO2 + 6H2O + 674 calories C6H12O6 → 2C2H5OH + 22 calories
Effect of temperature on microbial growth
Optimum, minimum and maximum temperatures for¬ microbial Microorganisms are classified w.r.t.
their¬growth. min. T. : refrigeration, freezing max. T. : heat processing temperature requirements.
Dependence of growth of microorganisms on¬ temperature Three different effects of¬is largely related to
activation and inactivation of enzyme systems in microorganisms. temperature contribute to the death of
microbial cells: 1. Denaturation of proteins (enzymes) by heat 2. Intoxication due to accelerated metabolic
reactions 3. Changes in essential lipids. Melting points of the fats found in the organisms and temperature ranges
of death are related.
Effect of acidity on microbial growth
Microorgisms¬ affected by pH of foods, since they do not have any mechanism to adjust Microorganisms have
different minimal, maximal and¬their internal pH. optimal pH-levels for growth: optimum Bacteria Molds Yeasts
6.5-7.5 4.0-6.8 4.5-6.5 maximum 9.0 8.0-11.0 8.0-8.5 minimum 4.5 1.5-3.5 1.5-3.5
Acid foods : pH¬ < Pasteurization (heat treatment below boiling¬4.5 . Pathogenic micro-organisms cannot
survive. temperature) For pH¬is sufficient for acid foods. > 4.5, sterilization (severe heat treatment above
boiling temperatures) is needed to destroy pathogenic microorganisms and their toxins.
pH ranges for major foods
Fruits ¬: 3.0 – 4.5 Vegetables : 4.6 – 6.6 Meat : 5.5 – 6.8 Milk : 6.0 – 6.8 Acetic-,¬Pickling : Production of acids
in foods by fermentation. citric-, lactic-, propionic-, benzoic-, and sorbic acids added to foods pH also affects
resistance of¬for preservation (acidulants) microorganisms to destruction during heating, drying and other
processes
Effect of nutrients on microbial growth
¬ ¬Microorganisms depend on nutrients for both energy and growth. Different microorganisms possess different
enzyme systems which are ¬specific in breakdown of certain nutrient compounds. Microbial growth can be
enhanced by enriching the growth medium with specific Creating¬nutrients and it can be retarded using growth
inhibitors. specific nutrient media is a very useful tool both in laboratory work and in industry for isolation and
growth of certain microorganisms.
Growth inhibitors
Many chemical compounds selectively inhibit¬ microbial activity These compounds act either by inactivating a
necessary enzyme needed¬ for growth, by denaturing the protein portion of a cell or by causing physical damage
to certain parts of the cell like breaking the cell A microrganism growing in food may produce substances¬wall.
inhibitory to other organisms or to destroy compounds inhibitory to its growth. Basis for production of antibiotics.
Example: Pencillin is produced by moulds (aspergillus) and retards growth of bacteria.
Antibiotics used to control microbial growth
1. 2. 3.
Cell wall inhibitors Protein synthesis inhibitors DNA replication inhibitors
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The Influence of Environmental Factors on Growth
Microbial growth is greatly affected by chemical and physical nature of their surroundings instead of
variations in nutrient levels and particularly the nutrient limitation. For successful cultivation of
microorganisms it is not only essential to supply proper and balanced nutrients but also it is necessary to
maintain proper environmental conditions. Thus, understanding of environmental influences on the
growth of microorganisms becomes mandatory. As bacteria shows divers food habits, it also exhibits
diverse response to the environmental conditions. Growth and death rates of microorganisms are greatly
influenced by number of environmental factors such as solutes and water acidity, temperature, oxygen
requirement, pH, pressure and radiation
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Chapter III
Factors that Influence Microbial Growth

Increasing the acidity of foods, either through fermentation or the addition of weak acids, has been used as a preservation
method since ancient times. In their natural state, most foods such as meat, fish, and vegetables are slightly acidic while most
fruits are moderately acidic. A few foods such as egg white are alkaline. Table
3–3 lists the pH ranges of some common foods. Added antimicrobial compounds can have an interactive or synergistic effect
with other parameters of the formulation. One example is the interaction with pH. Many preservatives have an optimum pH
range for effectiveness. Other factors include aw, presence of
other preservatives, types of food constituents, presence of certain enzymes, processing temperature, storage atmosphere, and
partition coefficients. The effective use of combinations of preservatives with other physicochemical parameters of a food
formulation can stabilize that food against spoilage organisms or pathogens. Leistner systematically developed the “hurdle
concept” to describe these effects (Leistner 1995). The hurdle concept states that several inhibitory factors (hurdles), while
individually unable to inhibit microorganisms,
will, nevertheless, be effective in combination. A classic example of applying the hurdle concept is the antibotulinal stability of
certain shelf-stable processed cheese formulations. Combinations of moisture, total salt, and pH have been shown to allow for
the safe storage of these products at room temperature for extended time even though the individual factors, taken singly,would
not support that practice (Tanaka and others 1986). In combination products, the effectiveness of an antimicrobial may be altered
by other factors including the potential for migration of the antimicrobial to other components of the food and the different food
parameters at the interface areas.There are a number of food formulations that, either by addition of preservatives or through the
application of the hurdle concept do not require refrigeration for microbiological stability or safety. However, in the absence of a
well defined and validated microbiological model, it is usually difficult to evaluate the microbiological safety of these products.
In the majority of these cases, the application of appropriate microbiological challenge testing is the most effective tool for
judging the suitability of these formulations
for nonrefrigerated storage.