TRANSFORMASI NITROGEN II

(Fiksasi N2 Simbiotik dan Non Simbiotik, Pengaturan

Aktivitas N2)

Oleh:
Monica Lucky
Rizcha Prawestriningrum

Bioteknologi Tanah dan Pupuk

Nitrogen Cycle
 NITROGEN FIXATION

Most of living The nitrogen

organism (N2) is quite There are three
need inert process for
nitrogen molecule nitrogen fixation

Biologic Atmosph
Industrial
al eric
Fixation
Fixation Fixation
BIOLOGICAL FIXATION

Nitrogenase is composed
of of two subunits: an
iron-sulfur protein (Fe
Protein) and a
molybdenum-iron-sulfur
protein (MoFe Protein)

The presenece of oxygen

caused Fe-proteins
oxydized, so nitrogenase
is inactived.
N2+ 8H+ + 8e-+ 16 ATP = 2NH3+ H2+ 16ADP +
16 Pi
 BIOLOGICAL FIXATION

Only a few species of microorganisms

can fix nitrogen.
Type Microorganism
Symbiotic:
Bacteria Rhizobium-Legume
Blue Green Anabaena azollae-Azolla
Algae
Asymbiotic:
Aerobic Azotobacter, Beijerinckia
Azospirillum, Thiobacillus, Spirillum
Microaerophilic Bacillus, Escherichia, Pseudomonas
Facultative Clostridium, Desulfovibrio
Anaerobic Nostoc, Anabaena, Rhodospirillum
Blue Green
BIOLOGICAL FIXATION
SYMBIOTIC
(Rhizobium-Legume) Rhizobium
Gram -, rod
Most studied symbiotic N2-fixing
bacteria
Now subdivided into several gene
Many genes known that are invol
in nodulation

Legume
The subfamilies of
legumes
(Caesalpinioideae,
Mimosoideae,
Papilionoideae), 700
genera, and 19,700
species of legumes.
Only about 15% of the
species have been
evaluated for
 BIOLOGICAL FIXATION
SYMBIOTIC (Actinorhizal
Association)

Figure A (below)
shows a young
alder tree (Alnus
glutinosa) growing
in a plant pot

Figure B shows part

of the root system
of this tree, bearing
the orange-yellow
coloured nodules
(arrowheads)
containingFrankia.
 BIOLOGICAL FIXATION
SYMBIOTIC (Actinorhizal Plant
Hosts)
Family Genera
Betulaceae Alnus
Casuarinaceae Allocasuarina, Casuarina, Ceuthostoma,
Gymnostoma
Myricaceae Comptonia, Myrica
Elaeagnaceae Elaeagnus, Hippophae, Shepherdia
Rhamnaceae Ceanothus, Colletia, Discaria, Kentrothamnus,
Retanilla, Talguenea, Trevoa
Rosaceae Cercocarpus, Chamaebatia, Cowania, Dryas,
Purshia
Coriariaceae Coriaria
Datiscaceae Datisca
 BIOLOGICAL
` FIXATION
SYMBIOTIC (Anabaena Azolla)

Heterocysts are modified cells

found in certain filamentous
cyanobacteria. They lack
photosystem II (and thus do
not produce O2), but possess
nitrogenase and can conduct
 Major metabolic reactions and nutrient exchanges
occurring in the bacteroid

bacteroid: rhizobial
cells transformed into
swollen, misshapen,
branched shapes within
the plant cell, possess
nitrogenase activity,
symbiosome:
incapable of proliferation
bacteroids (singly or
small groups)
surrounded by portions
of plant cell membrane;
N2-fixation begins O
leghemoglobin: after
2-
symbiosome
binding formation
protein; supplies
O2 to the bacteroid and
protects nitrogenase from
O2-inactivation
synthesis requires input
 Biological Fixation

Phyllosphere or rhizosphere (tropical grasses)

Azosprillum, Acetobacter
1 to 10% of rhizosphere population
Some establish within root
Same energy and oxygen limitations as
free-living
Acetobacter diazotrophicus lives in internal
tissue of sugar cane, grows in 30% sucrose,
can reach populations of 106 to 107 cells g-1
tissue, and fix 100 to 150 kg N ha-1 y-1
 BIOLOGICAL FIXATION

Factors affecting N2 Fixation

by free living
1. pH
Azotobacter sensitive to acidity
Beijerinkia & Derxia lived at pH 3-9
2. Phosporus and organic matter (nutrient status)
3. Available source energy
4. Efficiency
5. Secretion (exudate) Azospirilium lipoferum
6. Direct inoculation with non-symbiotic fixing
nitrogen bacteria
Estimated Average Rates of
Biological N2 Fixation

Nitrogen Fixation
N2 fixing system
(kg N/ha/year)
Rhizobium-legume 200-300

Cyanobacteria- moss 30-40

Rhizosphere
2-25
associations
Free- living 1-2
Atmospheric Fixation
Industrial Fixation
Pengukuran Laju Fiksasi
N

TUJUAN?

Mengetahui jumlah gas nitrogen (N2), yang diikat

oleh suatu mikroorganisme atau tumbuhan yang
bersimbiosis dengan mikroorganisme penambat
nitrogen
Pengukuran Laju Fiksasi
N

Metode Kjeldahl

Metode Pengukuran
Isotop 15N

Metode Analisis Reduksi

Asetilen
 Metode Kjeldahl
Prinsip kerja: mengukur kandungan
amonium dalam sampel dengan
distilasi alkali

3 tahapan:
Destruksi
Distilasi
Titrasi
Destruksi
Sample + H2SO4 (NH4)2SO4(aq) + CO2(g) + SO2(g) +
H2O(g)
Distilasi
(NH4)2SO4(aq) + 2NaOH Na2SO4(aq) + 2H2O(l) + 2NH3(g)
Titrasi
B(OH)3+ H2O + NH3 NH4++ B(OH)4
B(OH)3+ H2O + Na2CO3 NaHCO3(aq) + NaB(OH)4(aq) +
CO2(g) + H2O
Metode Pengukuran Isotop 15 N

3 tahapan :
Konversi isotop (sampel)
amonium
Konversi amonium + NaOBr
nitrogen
Perhitungan komposisi
isotop 15N dengan
spektrofotometer
 Metode Analisis Reduksi
Asetilen
Prinsip kerja : memisahkan gas etilen
dengan kromatografi, kemudian
mengukur konsentrasi gas etillen
tersebut dengan kolorimetri.
Metode Analisis Reduksi
Asetilen

N2 + 8H+ +8e
2NH3+H2
C2 H2+2H+2e C2
H4
Reduksi empat molekul asetilena sama dengan
reduksi satu molekul N2