Reactions of the Nitrogen Cycle Lecture PDF

Microbiology and Environmental Biotechnology Reactions of the Nitrogen cycle Eva Gonzalez Flo [email protected] Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 1 https://www.youtube.com/watch?v=vDrCZCAIz-A Eva Gonzale...

Microbiology and Environmental Biotechnology Reactions of the Nitrogen cycle Eva Gonzalez Flo [email protected] Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 1 https://www.youtube.com/watch?v=vDrCZCAIz-A Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 2 Content 1. Introduction 2. Nitrogen cycle 3. Ammonification 4. Nitrification 5. Denitrification 6. Nitrogen fixation 7. Anammox process 8. Exercices Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 3 Where N is found in the Environment? The largest source of N is the Atmosphere Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 4 Why Nitrogen is important? Nitrogen (N) is required to make PROTEINS Amino acids are the building blocks of all proteins N is a component of amino acids - Amino froup - Peptide linkage Proteins comprise not only structural components such as muscle, tissue and organs, but also enzymes and hormones essential for the functioning of all living things Most living organisms are unable to use N as exists in the atmosphere as N2 To enter the food chain N must be converted into Nitrates (NO3) which can easily be metbolised to make DNA and Proteins Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 5 Where does Nitrogen come from? 1 2 3 light, UV N + O2 Precipitation NO3- Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 6 Why is it necessary to treat Nitrogen? N is highly toxic to aquatic life Eutrophication happens when too much N enriches the water causing excessive growth of plants and algae Methemoglobinemia or “blue baby syndrome” is a serious health condition called caused by drinking water with high levels of nitrate Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 7 Introduction to the N cycle Nitrogen cycle is a biogeochemical process trought which N is converted into many forms as: !# N gas Denitrification (0) Nitrogen fixation (anaerobic) organic C !)* & !) * nitrite # !"# − % nitrate organic N (+ V) (+ III) (- III) Anammox Ammonification Nitrification (aerobic) !"& − !"'( Ammonia, ammonium (- III) O, Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 8 Nitrogen chemical species An oxidizing agent, or an oxidant, is a chemical species that accepts electrons given up by another, called a reducing agent. The Oxidizing Agent accepts the electrons from another chemical species, thus increasing its negative charge and therefore being reduced. REDUCTION The Reducing Agent, on the other hand, gives up its electrons and therefore oxidizes. OXIDATION reduction Oxidation Reduction oxidation e- donnor e- acceptor Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 9 Ammonification Organically bound nitrogen of microbial, plant and animal biomass and the excreted nitrogenate compounds are degraded/recylced after their death. !N #gas Denitrification (0) Nitrogen fixation (anaerobic) organic C * * !) & nitrate !) # nitrite !"# − % organic N (+ V) (+ III) Anammox (- III) Ammonification Nitrification (aerobic) !"& − !"'( Ammonia, ammonium (- III) O, Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 10 Ammonification Fungi and prokaryotes then decompose the tissue and release organic nitrogen back into the ecosystem as ammonia in the process known as ammonification. Ammonia is found in large quantities in wastewater due to urea excreted by humans. Urea is rapdly hydrolised into ammonium. Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 11 Nitrification The biochemical oxidation reaction that converts ammonia (NH3) into nitrate (NO3-) is called nitrification. Nitrification is a two-step reaction: !N #gas Denitrification (0) Nitrogen fixation (anaerobic) organic C * * !) & nitrate !) # nitrite Anammox !"# − % organic N (+ V) (+ III) (- III) Ammonification Nitrification (aerobic) !"& − !"'( Ammonia, ammonium (- III) O, Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 12 Nitrification The biochemical oxidation reaction that converts ammonia (NH3) into nitrate (NO3-) is called nitrification. Nitrification is a two-step reaction Chemical Species Oxidation stage Name NH2-R -3 Organic Nitrogen NH3, NH4+ -3 Ammonia, ammonium N2 0 Nitrogen gas Step 1 NO2- +3 Nitrite Step 2 NO3- +5 Nitrate Living beings oxidize ammonia present in the soil into useful forms of Nitrogen that can then be consumed by various organisms Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 13 Nitrification Living beings oxidize ammonia present in the soil into useful forms of Nitrogen that can then be consumed by various organisms The bacteria that carry it out are called nitrifiers and are AEROBIC CHEMOAUTOTROPHS OBLIGATE AEROBERS CHEMOSYNTEHTIC Energy source is inorganic matter O2 is the electron acceptor Obtains energy by the oxidation of e- donors Nitrogen e- donors release less energy per e- eq. AUTOTROPHS C source is Inorganic Carbon Fix and reduce inorganic carbon Is an energy-expensive process Small fso and Y Nitrifiers are slow growers Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 14 Nitrification Nitrification is a two-step reaction 1. Ammonia-oxidizing bacteria (AOB) of the genus Nitrosomonas oxidize ammonia/ ammonium (NH3 / NH4+ ) to nitrite (NO2-): 2 NH4+ + 3 O2 ® 2 NO2- + 4 H+ + 2 H2O (-3) (0) (+3) (-2) oxidation reduction 2. Nitrite-oxidizing bacteria (NOB) of the genus Nitrobacter oxidize nitrite (NO2-) to nitrate (NO3-): 2 NO2- + O2 ® 2 NO3- (+3) (0) (+5) (-2) The second step of nitrification is faster than the first, so the concentration of NO2- generally observed in water is usually lower than that of NO3-. Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 15 How to adjust a reaction REACTION = fe · energy reaction + fs · synthesis reaction fe + fs = 1 fs = YX/S Identify which element acts as an electron donor and electron acceptor Write for each condition the semi-reactions Oxidation Reduction Electron (e-) donor Electron (e-) acceptor e- donnor e- acceptor N. Write two minimal semi-reactions O. Adjust number of principal element O. Adjust number of Oxygen with H2O O H. Adjust number of H with H+ H e-. Balance q with electrons e- x multiply each semi-reaction to equal e- or adjust by Equivalent Electrons xn + Sum up the two semi-reactions an cancel terms in both sides of the reaction IMPORTANT e- must be cancelled, all elements must be adjusted OBTAIN THE FINAL EQUATION OF THE PROCESS Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 16 Nitrification energy reaction Step 1: NH4+ ® NO2- Electron (e-) donor Electron (e-) acceptor # N NH4+ ® NO2- O O ® H2O " 2 # O NH4+ + 2 H2O ® NO2- O O ® H2O " 2 # H NH4+ + 2 H2O ® NO2- + 8 H+ H O + 2 H+ ® H2O " 2 # e- NH4+ + 2 H2O ® NO2- + 8 H+ + 6 e- e- O + 2 H+ + 2 e- ® H2O " 2 x1 NH4+ + 2 H2O ® NO2- + 8 H+ + 6 e- x3 + # O + 2 H+ + 2 e- ® H2O " 2 ! NH4+ + 2 H2O + " O2 + 6 H+ + 6 e- ® NO2- + 8 H+ + 6 e- + 3 H2O $ NH4+ + O2 ® NO2- + 2 H+ + H2O % Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 17 Nitrification energy reaction Step 2 : NO2- ® NO3- Electron (e-) donor Electron (e-) acceptor ! N NO2- ® NO3- O O ® H2O " 2 ! O NO2- + H2O ® NO3- O O ® H2O " 2 ! H NO2- + H2O ® NO3- + 2 H+ + H O + 2 H+ " 2 ® H2O ! e- NO2- + H2O ® NO3- + 2 H+ + 2 e- e- O + 2 H+ + 2 e- ® H2O " 2 x1 NO2- + H2O ® NO3- + 2 H+ + 2 e- + ! O + 2 H+ + 2 e- ® H2O x1 " 2 ! NO2- + H2O + O2 + 2 H+ + 2 e- ® NO3- + 2 H+ + 2 e- + H2O " # NO2- + O2 ® NO3- $ Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 18 Nitrification energy reaction To obtan the energy reaction we sum up the reactions from each step ! Step 1: NH4+ + O2 ® NO2- + 2 H+ + H2O " + # Step 2 : NO2- + O2 ® NO3- " NH4+ + 2 O2 ® NO3- + H2O + 2 H+ Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 19 Nitrification synthesis reaction Electron (e-) donor Electron (e-) acceptor N NH4+ ® NO3- C 5CO2 + NH4+ + ® C5H7O2N O NH4+ + 3 H2O ® NO3- O 5 CO2 + NH4+ + ® C5H7O2N + 8 H2O H NH4+ + 3 H2O ® NO3- + 10 H+ H 5 CO2 + NH4+ + 19H+ ® C5H7O2N + 8 H2O e- NH4+ + 3 H2O ® NO3- + 10 H+ + 8 e- e- 5 CO2 + NH4+ + 19 H+ + 20 e- ® C5H7O2N + 8 H2O x2.5 NH4+ + 3 H2O ® NO3- + 10 H+ + 8 e- + 5 CO2 + NH4+ + 19 H+ + 20 e- ® C5H7O2N + 8 H2O 2.5 NH4+ + 7.5 H2O + 5 CO2 + NH4+ + 19 H+ + 20 e- ® 2.5 NO3- + 25 H+ + 20 e- + C5H7O2N + 8 H2O 3.5 NH4+ + 5 CO2 ® 2.5 NO3- + C5H7O2N + 0.5 H2O + 6 H+ R à P q= 3.5 3.5 N= 3.5 3.5 C= 5 5 H= 14 14 O= 10 10 Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 20 Nitrification global reaction To obtain the global reaction we sum up the energy reaction and the synthesis reaction Energy reaction: NH4+ + 2 O2 ® NO3- + H2O + 2 H+ + Synthesis reaction: 3.5 NH4+ + 5 CO2 ® 2.5 NO3- + C5H7O2N + 0.5 H2O + 6 H+ 4.5 NH4+ + 5 CO2 + 2 O2 ® 3.5 NO3- + C5H7O2N + 1.5 H2O + 8 H+ with !s = 0.067 1.1675 NH4+ +1.866 O2 + 0.335 CO2 ® 1.1 NO3- + 0.067 C5H7O2N + 0.96 H2O + 2.27 H+ High oxigen consumption Low biomass productivity Acidification Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 21 Nitrification key concepts Nitrification is important in many organisms as it is the only process for obtaining N These organisms convert ammonia into nitrates which is more soluble than ammonia and thus can be taken into the system more conveniently In agricultural systems ammonia is used as a fertilizer. The ammonia is then converted into nitrate which facilitates nitrogen leaching into the plants Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 22 Denitrification In anoxic environments nitrate (NO3-) or nitrite (NO2-) can be used as an electron acceptor in the respiration process and be converted to N2 (gas) Nitrogen is released back into the atmosphere from the ground. !N #gas Denitrification (0) Nitrogen fixation (anaerobic) organic C Anammox !)* & !)* # !"# − % nitrate nitrite organic N (+ V) (+ III) (- III) Ammonification Nitrification (aerobic) !"& − !"'( Ammonia, ammonium (- III) O, Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 23 Denitrification The bacteria that carry it out are denitrifiers and are ANAEROBIC CHEMOHETEROTROPHS FACULTATIVE AEROBERS CHEMOSYNTEHTIC O2 respiration Energy source is inorganic matter NO3 or NO2 respiration when O2 is limted Obtains energy by the oxidation of e- donors HETEROTROPHS C source is Organic Carbon Methanol Glucose Acetic acid Gram - bacteria Proteobacteria: Pseudomonas, Alcaligenes, Paracoccus, and Thiobacillus. Gram + bacteria Bacillus Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 24 Denitrification energy reaction Metanol (CH₃OH) as e- source and Nitrate (NO3-) as e- acceptor Electron (e-) donor Electron (e-) acceptor C CH₃OH ® CO2 N 2 NO3- ® N2 O CH₃OH + H2O ® CO2 O 2 NO3- ® N2 + 6 H2O H CH₃OH + H2O ® CO2 + 6 H+ H 2 NO3- + 12 H+ ® N2 + 6 H2O e. CH₃OH + H2O ® CO2 + 6H+ + 6 e- e- 2 NO3- + 12 H+ + 10 e- ® N2 + 6 H2O x 10 CH₃OH + H2O ® CO2 + 6H+ + 6 e- + x6 2 NO3- + 12 H+ + 10 e- ® N2 + 6 H2O 5 CH₃OH + 6 NO3- + 6 H+ ® 5 CO2 + 3 N2 + 13 H2O 5 CH₃OH + 6 NO3- ® 5 CO2 + 3 N2 + 7 H2O + 6 OH- Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 25 Denitrification energy reaction The general formula for denitrification is NO3- + organic matter® N2 + CO2 + H2O + OH- The reaction energy for metanol, acetate, etanol and glucose are: metanol (CH₃OH): 5 CH₃OH + 6 NO3- ® 5 CO2 + 3 N2 + 7 H2O + 6 OH- acetate (CH₃COOH): 5 CH₃COOH + 8 NO3- ® 10 CO2 + 4 N2 + 6 H2O + 8 OH- ethanol (CH₃CH2OH): 5 CH₃CH2OH + 12 NO3- ® 10 CO2 + 6 N2 + 9 H2O + 12 OH- glucose (C5H12O6): C5H12O6 + 4 NO3- ® 5 CO2 + 2 N2 + 4 H2O + 4 OH- Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 26 Denitrification synthesis reaction metanol (CH₃OH) Electron (e-) donor Electron (e-) acceptor C CH₃OH ® CO2 C NO3- + 5 CO2 ® C5H7O2N O CH₃OH + H2O ® CO2 O NO3- + 5 CO2 ® C5H7O2N + 11 H2O H CH₃OH + H2O ® CO2 + 6 H+ H NO3- + 5 CO2 + 29 H+ ® C5H7O2N + 11 H2O e. CH₃OH + H2O ® CO2 + 6H+ + 6 e- e- NO3- + 5 CO2 + 29 H+ + 28 e- ® C5H7O2N + 11 H2O x 28 28 (CH₃OH + H2O ® CO2 + 6H+ + 6 e- ) + 6 (NO3- + 5 CO2 + 29 H+ + 28 e- ® C5H7O2N + 11 H2O ) x6 28 CH₃OH + 6 NO3- + 2 CO2 + 6 H+ ® 38 H2O + 6 C5H7O2N Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 27 Denitrification Energy reaction: 5 CH₃OH + 6 NO3- ® 5 CO2 + 3 N2 + 7 H2O + 6 OH- + Synthesis reaction: 28 CH₃OH + 6 NO3- + 2 CO2 + 6 H+ ® 38 H2O + 6 C5H7O2N With !s = 0.267 11.141 CH₃OH + 6 NO3- + 1.6 H+ ® 3.131 CO2 + 2.199 N2 + 1.60 C5H7O2N + 15.27 H2O + 4.39 OH- Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 28 Denitrification Denitrification is applied when the complete removal of N is required Advanced treatment of wastewater discharged to watersheds to protect against eutrophication Treatment of drinking waters containing elevated NO3- + NO2- levels Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 29 Nitrification-denitrification key concepts The processes of nitrification - denitrification is used to remove N from urban wastewater The N in fresh wastewater is usually in the form of ammoniacal N or organic N, both in dissolved and particulate form. 1. Organic N is transformed into ammoniacal N which is then nitrified until it becomes NO3-. 2. The water is then subjected to anoxia conditions, in the presence of oxidizable organic matter, so that the NO3- is converted by denitrification to N2 (gas) which escapes into the air. The overall result of these processes is a net loss of the N initially contained in the water. Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 30 Anammox process ANAMMOX stands for ANaerobic AMMonium OXidation Microorganisms performing the Anammox process are bacterium from the Planctomycetes phylum !# Denitrification N gas Nitrogen fixation (0) (anaerobic) organic C !)* & !)* # !"# − % nitrate nitrite Anammox organic N (+ V) (+ III) (- III) Ammonification Nitrification (aerobic) !"& − !"'( Ammonia, ammonium (- III) O, Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 31 EXERCICES 1. Nitrification is a component of the nitrification/denitrification process used in wastewater treatment plants for Nitrogen removal. Write a balanced equation for the aerobic, autotrophic oxidation of ammonium (NH4+) to nitrate (NO3-). Use a value of fs = 0.1. 2. One way to remove nitrogen from wastewater is by biological nitrification/denitrification. In the Denitrification step, an organic such acetic acid (CH₃COOH) is used as an electron donor under anaerobic conditions and nitrate is used as an electron acceptor. During denitrification nitrate is reduced to nitrogen gas which can escape to the atmosphere. (1) Write the energy equation for the reaction. (2) Write the synthesis equation for the reaction (use the biomass formula). (3) Write a balanced equation for the overall reaction considering fs = 0.4. (4) Normalise to one mole of NO3-. Eva Gonzalez Flo Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 32 Microbiology and Environmental Biotechnology Reactions of the Nitrogen cycle Eva Gonzalez Flo [email protected] Microbiology and Environmental Biotechnology: Reactions of the Nitrogen cycle 33