By: Mochamad Nurcholis Food Science Department Brawijaya University 2013

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1 PHYSIOLOGY & METABOLISMS of Microorganisms By: Mochamad Nurcholis Food Science Department Brawijaya University 2013

2 What is metabolisms? Can you explain it?

3 Overall biochemical reaction within cells of living organisms which require or produce energy, then used for cellular activities.

4 What is the role of Metabolisms? Coordinating, controlling of cellular activities Maintain homeostatic of living organisms

5 METABOLISMS PATHWAY Role : Synthesize macro-molecules or breakdown macro-melecules. Metabolisms pathway can be divided into three types : linear, branched or cyclic.

6 ENZYMe in metabolisms Enzyme as biocatalyst Protein that influence the rate of chemical reaction Protein that decrease energy of activation Have a unique structure and active site or specific site Lock and key (complex of enzyme-substrate),

7 ENZyMe Enzyme will be released from the substrate without change its structure. Enzyme can be used for several times and capable in catalyze next reaction. Optimally active when : 1. co-enzyme (non protein organic molecules, such as : vitamin) 2. co-factor (mineral ion, such as : Zn, Cu, Fe

8 Complex of Enzyme-Substrate

9 METABOLISMS

10 CATABOLISMS Breakdown/ hydrolysis/ of macro-molecules into simple molecules through biochemical pathway. Release more energy than energy required (exergonic).

11 CATABOLISMS Cell Respiration Respiration is an example of catabolisms. Respiration process produce energy (ATP, NADH, FADH). Catabolisms Type : 1. Glycolysis 2. Lipolysis 3. Proteolysis

12 Important Aspect of Respiration Provide energy for growth and homeostatic Respiration rate of each organisms is various The highest respiration rate occur at cell differentiation and reproduction Respiration rate is influenced by internal and external factors

Respiration Overall reaction : C 6 H 12 O 6 + 6 O 2 6 CO 2 + 6 H 2 O + Energy Cellular respiration is oxidation

13 Respiration Overall reaction : C 6 H 12 O O 2 6 CO H 2 O + Energy Cellular respiration is oxidation process of macro-molecules (ex : foods) Energy is the product of cellular respiration Energy provide cellular activities

14 Respiration 3 Stages : 1. Breakdown large macromolecules to simple unit 2. Breakdown of simple unit to acetyl coa, limited amounts of ATP & NADH 3. Complete oxidation of acetyl coa, large amounts of ATP & NADH

15 Cellular Respiration

Cellular Respiration Glycolysis (Embden-Meyerhof-Parnas) Glucose is oxidized to pyruvic acid with ATP and energy-containing NADH produced Pyruvic acid is converted acetyl CoA with NADH produced TCA

16 Cellular Respiration Glycolysis (Embden-Meyerhof-Parnas) Glucose is oxidized to pyruvic acid with ATP and energy-containing NADH produced Pyruvic acid is converted acetyl CoA with NADH produced TCA Cycle (Kreb s cycle) Acetyl CoA is oxidized to CO 2 with ATP, NADH and FADH 2 is produced Electron Transport Chain NADH and FADH 2 are oxidized through a series of redox reactions and a considerable amount of ATP is produced

17 Kreb s Cycle/TCA cycle/citric Acid Cycle

18 Electron Transport

19 SUMMARY OF AEROBIC RESPIRATION

20 Glycolysis Acetyl coa Formation Kreb s Cycle Electron transport

21 Aerobic & anaerobic respiration

23 Fermentation Pathway Glucose ATP Acetaldehyde Ethanol + CO2 Pyruvate (C3) Acetate (C-2) NADH ATP NADH Acid + gas Kreb Cycle CO2 + H2O

24 Alternative Product End in Fermentation

25 Fermentation versus Respiration The Difference Fermentation Respiration Final e- acceptor Electron donors Organic compounds Organic compounds Process Glucose is coverted to 1 or 3 Carbon compound Oxygen Glucose is oxidized to CO2 Product Step Organic acid, alcohol & 1-2 ATP Glycolysis acid or alcohol fermentation 6 CO2, 6 H2O & energy (38 ATP) Glycolysis TCA cycle electron transport

26 ANABOLISMS Biosynthesis pathway Assimilation process / synthesize complex or macro-molecules from smallmolecules Involve dehydration reaction (releasing water) and require more energy input (endergonic)

27 Plant synthesize sugar and polysaccharide through photosynthesis Microorganisms synthesize cellulose through fermentation (ex: nata) ANABOLISMS

30 The Study of Microbial Growth Microbial growth occurs at two levels: growth at a cellular level with increase in size, and increase in population Division of bacterial cells occurs mainly through binary fission (transverse) parent cell enlarges, duplicates its chromosome, and forms a central transverse septum dividing the cell into two daughter cells

31 BINARY FISSION OF BACTERIA

32 Rate of Population Growth Time required for a complete fission cycle is called the generation, or doubling time Each new fission cycle increases the population by a factor of 2 exponential or logarithmic growth. Generation times vary from minutes to days.

33 Rate of Population Growth

34 Rate of Population Growth Equation for calculating population size over time: N ƒ = (N i )2 n N ƒ is total number of cells in the population. N i is starting number of cells. Exponent n denotes generation time. 2 n number of cells in that generation

35 The Population Growth Curve 1. Lag phase flat period of adjustment, enlargement; little growth 2. Exponential growth phase a period of maximum growth will continue as long as cells have adequate nutrients and a favorable environment 3. Stationary phase rate of cell growth equals rate of cell death caused by depleted nutrients and O 2, excretion of organic acids and pollutants 4. Death phase as limiting factors intensify, cells die exponentially in their own wastes

36 The Population Growth Curve

37 Conditions that Impact Microbial Growth Food Acid/pH Tempera ture Time Oxygen Moisture Salinity Pressure Radiation

38 Microbial Nutrition Nutrition process by which chemical substances (nutrients) are acquired from the environment and used for cellular activities Essential nutrients - must be provided to an organism Two categories of essential nutrients: macronutrients required in large quantities; play principal roles in cell structure and metabolism proteins, carbohydrates, lipids micronutrients or trace elements required in small amounts; involved in enzyme function and maintenance of protein structure manganese, zinc, nickel, calcium, cuprum, natrium, vitamins, growth hormones and metabolic precursors

39 Nutrients Inorganic nutrients atom or molecule that contains a combination of atoms other than carbon and hydrogen metals and their salts (magnesium sulfate, ferric nitrate, sodium phosphate), gases (oxygen, carbon dioxide) and water Organic nutrients- contain carbon and hydrogen atoms and are usually the products of living things methane (CH 4 ), carbohydrates, lipids, proteins, and nucleic acids

40 Chemical Analysis of Microbial Cytoplasm 70% water Proteins 96% of cell is composed of 6 elements: carbon hydrogen oxygen phosphorous sulfur nitrogen

41 Sources of Essential Nutrients Carbon sources Heterotroph must obtain carbon in an organic form such as proteins, carbohydrates, lipids and nucleic acids, made by other living organisms Autotroph - an organism that uses CO 2, an inorganic gas as its carbon source not nutritionally dependent on other living things

42 Carbon as the Major Sources of Cellular Carbon and Energy. Aerobic fermentations About 50% of substrate carbon is incorporated into cell mass and about 50% of it is used as energy sources. Anaerobic fermentation A large fraction of substrate carbon is converted to products and a smaller fraction is converted to cell mass (less than 30%).

43 Sources of Essential Nutrients Nitrogen Sources Main reservoir is nitrogen gas (N 2 ); 79% of earth s atmosphere is N 2. Nitrogen is part of the structure of proteins, DNA, RNA & ATP these are the primary source of N for heterotrophs. Some bacteria & algae use inorganic N nutrients (NO 3 -, NO 2 -, or NH 3 ). Some bacteria can fix N 2. Regardless of how N enters the cell, it must be converted to NH 3, the only form that can be combined with carbon to synthesis amino acids, etc.

44 Sources of Essential Nutrients Oxygen Sources Major component of carbohydrates, lipids, nucleic acids, and proteins Plays an important role in structural and enzymatic functions of cell Component of inorganic salts (sulfates, phosphates, nitrates) and water O 2 makes up 20% of atmosphere Essential to metabolism of many organisms

45 Sources of Essential Nutrients Hydrogen Sources Major element in all organic compounds and several inorganic ones (water, salts and gases) Gases are produced and used by microbes. Roles of hydrogen: maintaining ph forming H bonds between molecules serving as the source of free energy in oxidationreduction reactions of respiration

46 Sources of Essential Nutrients Phosphorous (Phosphate Sources) Main inorganic source is phosphate (PO 4-3 ) derived from phosphoric acid (H 3 PO 4 ) found in rocks and oceanic mineral deposits Key component of nucleic acids, essential to genetics Serves in energy transfers (ATP)

47 Sources of Essential Nutrients Sulfur Sources Widely distributed in environment, rocks; sediments contain sulfate, sulfides, hydrogen sulfide gas and sulfur Essential component of some vitamins and the amino acids: methionine and cysteine Contributes to stability of proteins by forming disulfide bonds

48 Other Nutrients Important in Microbial Metabolism Potassium essential to protein synthesis and membrane function Sodium important to some types of cell transport Calcium cell wall and endospore stabilizer Magnesium component of chlorophyll; membrane and ribosome stabilizer Iron component of proteins of cell respiration Zinc, copper, nickel, manganese, etc.

49 Micronutrients Micronutrients could be classified into the following categories (required less than 10-4 M) : - Most widely needed elements Fe, Zn and Mn. Such elements are cofactors for some enzyme and regulate the metabolism. - Trace elements needed under specific growth conditions Cu, Co, Mo, Ca, Na, Cl, Ni, and Se. For example, copper is present in certain respiratory-chain components and enzymes.

50 Micronutrients - Trace elements rarely required B, Al, Si, Cr, V, Sn, Be, F, Ti, Ga, Ge, Br, Zr, W, Li and I. These elements are required in concentrations of less than 10-6 M and are toxic at high concentration. - Growth factor is also micronutrient. Growth factor stimulates the growth and synthesis of some metabolites. e.g. vitamin, hormones and amino acids. They are required less than 10-6 M.

51 Growth Factors: Essential Organic Nutrients Organic compounds that cannot be synthesized by an organism because they lack the genetic and metabolic mechanisms to synthesize them Must be provided as a nutrient essential amino acids, vitamins

52 Nutritional Types Main determinants of nutritional type are: carbon source heterotroph, autotroph energy source chemotroph gain energy from chemical compounds phototrophs gain energy through photosynthesis

53 Classification of microbes according to their oxygen responses. a. Aerobic b. Anaerobic c. Facultative d. Microaerobic e. Aerotolerant

54 Methods of Analyzing Population Growth Turbidometry most simple Degree of cloudiness, turbidity, reflects the relative population size Enumeration of bacteria: viable colony count direct cell count count all cells present; automated or manual

55 Methods of Analyzing Population Growth

56 REFERENCES Lehninger AL Principles of Biochemistry. Worth Publisher, Inc. Maryland Talaro KP Foundation in Microbiology 6th Edition. The McGraw Hill Companies. Ray B Fundamental Food Microbiology. CRC Press. Boca Raton. Pelczar and Chan Elements of Microbiology. McGraw Hill Book Company. Scientific articles from internet/website

57 Te Ri Ma Ka Sih

Foundations in Microbiology Seventh Edition

Lecture PowerPoint to accompany Foundations in Microbiology Seventh Edition Talaro Chapter 7 Elements of Microbial Nutrition, Ecology, and Growth Copyright The McGraw-Hill Companies, Inc. Permission required