Effect of soil management practices on soil enzyme activities

1. 11

2. Speaker:
Pradip Tripura
Ph.D (Agri.) student
Reg. No. 1010115012
College of Agriculture
J.A.U., Junagadh.
Major advisor:
Dr. K. B. Polara
Professor
Dept. of Agril. Chemistry and
Soil Science
College of Agriculture,
JAU., Junagadh.
Minor advisor:
Dr. B. K. Sagarka
Professor and Head
Dept. of Agronomy
College of Agriculture,
JAU., Junagadh.
SEMINAR ON
Effect of soil management practices on soil
enzyme activities
2

3. 3
Soil enzymes - function, classification, sources, importance and
application
 Soil enzyme as biological quality indicator
 Role of soil enzymes in maintaining soil health
 Different methods of enzyme activity measurement
 Effect of different management practices on enzymatic activity
 Conclusion
Contents

4. Soil
Quality
Soil enzyme activity
Nutrient transformationMetabolic capacity
Indicator
Soil Enzymes and Soil Quality
Management Practices
4

5. Soil Enzymes
 A substance produced by a living organism which acts as a
catalyst to bring about a specific biochemical reaction.
 A group of enzymes.
 Inhabitate in soil.
 Playing an important role in maintaining soil ecology, physical and
chemical properties, fertility, and soil health.
Functions of Enzymes in soil
 Play key biochemical functions in the overall process of organic
matter decomposition in the soil system (Sinsabaugh et al., 1991)
 Important in catalyzing several vital reactions necessary for,
 Life processes of micro-organisms in soils.
The stabilization of soil structure.
Organic matter formation.
Nutrient cycling.
5

6. 1. Oxidoreductases - Oxidation reduction reaction
(Dehydrogenase, Catalase, Peroxidase)
2. Tranferases - The transfer of group of atoms from
donor to an acceptor molecule.
(Aminotransferases, Rhodonese)
3. Hydrolases - Hydrolitic cleavage of bonds.
(Phosphatase, Cellulase, Urease)
4. Lyases - Cleavage of bonds other than hydrolysis
or oxidation.
(Aldolase)
5. Isomerases - Isomarization reaction
6. Ligases - Formation of bonds by the cleavage of
ATP.
(Acetyl-CoA carboxylase)
Enzymes Classification
6

7. 1 Constitutive
Always present in nearly constant amounts in a
cell (not affected by addition of any particular
substrate – genes always expressed).
(Pyrophosphatase)
2 Inducible
Present only in trace amounts or not at all, but
quickly increases in concentration when its substrate
is present.
(Amidase)
Both types of enzymes are present in the soil.
Kind of soil Enzymes
7

8. Although the general origins of soil
enzymes are:-
(a) Microorganisms-living and dead
(b) Plant roots and plant residues and
(c) Soil animals;
Origin of soil Enzymes
8

9. State-1: Role of Clays
a. Most activity associated with clays.
b. Increases resistance to proteolysis and microbial attack
c. Increases the temperature of inactivation.
State-2 :Role of Organic Matter
a. Humus material provides stability to soil nitrogen
compounds
b. Enzymes attached to insoluble organic matrices exhibit pH
and temperature changes.
c. Inability to purify soil enzymes free of soil organic matter
(bound to O.M. )
State-3: Role of O.M. - Clay Complex
a. Lignin + bentonite ( clay ) protect enzymes against
proteolitic attack, but not bentonite alone.
b. Enzymes are bound to organic matter which is then bound
to clay.
State of soil Enzymes
9

10. 1. Accumulated enzymes
2. Continuously released extracellular enzymes
Proliferating
microbes
Plant roots Soil faunaBound to
microbial cells
Not associated with
Cellular components
Non
proliferating
cells
Dead cells Cellular
fragments
Originating
from plant
roots
Originating
from
microbes
and soil
fauna
Microbes Plants
Dick and Tabatabai (1992)10

11. Importance of Soil Enzymes
 Release of nutrients into the soil by means of organic matter
decomposition.
 Identification of microbial activity.
 As sensitive indicators of ecological change.
Application of Soil Enzymes
 Correlation with soil fertility
 Correlation with microbial activity
 Correlation with biochemical cycling of various elements in soil
(C, N, S)
 Degree of pollution (heavy metals, SO4)
 To assess the successional stages of an ecosystem
 Rapid degradation of pesticides
 Disease studies
 Enzyme activity as biological indicator of soil health and quality
and monitoring soil quality improvement
11

12. Soil enzymes as biological soil quality indicator
Soil Quality:
Capacity of a specific kind of soil to function within ecosystem and landuse
boundaries, to sustain biological productivity, maintain environmental quality and
sustain plant, animal and human health (Doran and Parkin, 1994)
Soil health:
A state of dynamic equilibrium between flora and fauna and their sourrounding soil
environment in which all the metabolic activities of the former proceed optimally
without any hinderance, stress or impedence from the later (Goswami and Rattan,
1992)
The definition of soil quality encompasses physical, chemical and biological
characteristics, and it is related to fertility and soil health
Biological indicators of soil quality:
Properties associated with biological activity on organic matter, such as
A. Microbial biomass carbon
B. Soil respiration
C. Abundance, diversity, food chains and stability of microbial communities
D. Mesofauna such as earthworms, nematodes and arthropods
E. Biological activities such as enzyme activity
F. Potentially mineralized nitrogen
G. CO2 production
12

13. Cont….
Why soil enzymes are considered as useful indicators of
soil quality?
Soil enzyme activities,
(1)Closely related to soil organic matter, soil physical
properties and microbial activity or biomass
(2)Changes much sooner than other parameters, thus
providing early indications of changes in soil health
(3)Involve simple procedures for determination of activity
(Dick et al., 1996)
13

14. Soil Enzyme Enzyme reaction End Product Indicator of
microbial activity
ß glucosidase Cellobiose hydrolysis Glucose C Cycling
FDA hydrolysis Organic matter
decomposition
Different nurients C Cycling
Amidase N-mineralization Ammonium N Cycling
Urease Urea hydrolysis Ammonium N Cycling
Phosphatase Hydrolysis of organic P
compounds
Inorganic Phosphate P Cycling
Arylsulphatase Hydrolysis of organic S
compounds
Inorganic Sulphate S Cycling
Phenol oxidase Lignin hydrolysis Low molecular
weight C
compounds
C Cycling
Cellulase Cellulose hydrolysis Glucose C Cycling
Dehydrogenase Electron transport
system
Oxidized products C Cycling
Potential Roles of Soil Enzymes in Maintaining Soil Health
14

15. Approach I :
Two methods are used frequently. These are,
 Colorimetric assay – By using p-nitrophenol (pNP) linked substrates or L- 3, 4-
dihydroxyphenylalanine (L-DOPA) or 2,3,5 triphenyl tetrazolium chloride (TTC)
 Fluorometric assay – By using 4-methyl umbelliferone (MUF) or 7-amino-4-
methyl coumarin (AMC) linked-substrates.
Different methods of enzyme activity measurement
Approach I I :
In situ approach. Include, addition of varying concentration of a single substrate with
the intention of having non-limiting substrate and, thereby, measuring Vmax as a proxy
for enzyme pools from the Michaelis-Menten equation.
Approach III :
 Genomic tools - Allow us to determine the genetic potential for the production
of specific enzymes in complex microbial communities
 Transcriptomic tools - Enable us to examine controls on the expression of
enzyme coding genes
 Proteomic methods - Allow us to directly detect the presence of enzymes in
the environment as well as providing information about the organisms that produce
them
15

16. Land use changes
Soil amendments - lime and
gypsum addition
Conservation practices
Tillage and cultivation
Soil Management
Practices
Crop rotation
Irrigation practices
Soil management practices affecting enzyme activities
LTFE with INM
Pesticide
application
16

17. Effect of crop rotation on enzyme
activities
17

18. CCCC - continuous corn
CSbCSb - corn–soybean
CCOM - corn–corn–oats–
meadow (alfalfa)
SbSbSbSb - continuous
soybean
Fig-1. Impacts of crop rotations on urease activity
18Lowa state university, USA Klose andTabatabai (2000)

19. CCCC - continuous corn
CSbCSb - corn–soybean
CCOM - corn–corn–oats–meadow (alfalfa)
SbSbSbSb - continuous soybean
Klose et al. (1999)
Fig – 2. Impact of crop rotation on arylsulphatase activity
19Lowa state university, USA Klose andTabatabai (2000)

20. Fig - 3. application of organic and inorganic nutrients to a Maize–Wheat rotation
on soil dehydrogenase activity
20Uttarakhand Saha et al. (2008)

21. Fig – 4. Effects of different tillage and crop rotation treatments on enzyme activity
RT-CC ridge tillage with monoculture
corn, RT-CS ridge tillage with corn-
soybean rotation, NT-CC no-tillage with
monoculture corn, NT-CS no-tillage with
corn-soybean rotation
21China Zhang et al. (2014)

22. Effect of shifting cultivation on enzyme
activities
22

23.  Study site - Nokrek Biosphere Reserve in the western part of
Meghalaya, north-east India.
 Two zones of Jhum cultivation :
a) Buffer zone – Exposing different degrees of jhum fallows of
different ages (1, 3-4, 6-8 and 12 years)
b) Core zone – Undisturbed forest
 About 129 villages of the Garo tribe with a total population of
39,432 are located in the buffer zone.
 Shifting agriculture is extensively practiced by these people in
the entire buffer zone.
 This agricultural practice involves slashing or complete clearing
of the vegetation and burning of dried slash during dry winter,
followed by pure and mixed cropping.
Effect of shifting cultivation on enzyme activities
23Meghalaya Ralte et al. (2005)

24. Fig – 5. Impacts of shifting cultivation on dehydrogenase activity
TPFreleased(µg/g/24hr)
0-10 cm
10-20 cm
Ralte et al. (2005) 24
Meghalaya Ralte et al. (2005)
C - Primary forest,J12, J6, J3 and J1 = 10-12, 6-8, 3-4 and 1 year
jhum fallows

25. Fig – 6. Impacts of shifting cultivation on urease activity
NH4released(µg/1oog/6hr)
10-20 cm
Ralte et al. (2005)
 The increase in dehydrogenase and urease activities from the young to the
old jhum fallows may be attributed to the build up of organic matter and
nutrients in the soil.
25Meghalaya Ralte et al. (2005)

26. Effect of soil amendment addition on
enzyme activities
26

27.  Experimental Site : Northeast
Research Center in Nashua,
Iowa.
 Soil samples collected from
0- 15 cm surface soil after 17
years of lime application in the
form of CaCO3
Fig – 7. Effect of lime application on soil phosphatases activity
27USA Acosta-Martinez and Tabatabai (2000)

28. control
N
Lime
N+Lime
Fig – 8. Effect of liming and nitrogen fertilization on soil phosphomonoesterase
activity
Johnson et al. (2005)
28University of Sheffield, UK Johnson et al. (2005)

29. Fig – 9. Enzyme activity assay in different soil amendments
X axis - 1.rhizosphere soil, 2.fungicide treatment, 3.insecticide treatment, 4.biofertilizer
treatment, 5.bulk soil; y axis optical density
29Kerala Aswathy and Jose (2013)

30. Effect of land use changes on enzyme
activities
30

31. 31
Fig – 10. Effect of land use types on soil enzymatic activities
Bamako (Mali) Gonnety et al. (2012)

32. Fig – 11. Phytase activity of arid crops was grown under tillage and
no-tillage condition
32Rajasthan Yadav and Tarafdar (2004)

33.  Arylsulfatse activities of fodder, organic farming and soybean-wheat
systems were found to be at par.
 If season and depths are compared, the arylsulfatase activity in the 0-15
cm soil depth can be arranged as: forest >soybean-wheat ≈ fodder ≈
organic farming > barren land
Fig – 12. Effect of land use changes on enzyme activities in the
himalayan region
33
IARI, New Delhi Justin et al. (2013)

34. Enzyme activities as
affected by soil order.
Enzyme activities as
affected by land use.
Fig – 13. Enzymatic activity affected by land order and land use
34
Akron, United state Acosta-Martinezn et al. (2007)

35. Table -1. Impact of land use system on enzymatic activity in central
Himalayan region
35IARI, New Delhi Singh et al. (2014)

36. Effect of LTFE with INM on enzyme
activities
36

37. 37
Treatment
Dehydrogenase Activity
(μg TPF-1 24 hr-1 g-1 soil)
1st Year 12th Year
T1 (50 % NPK of recommended dose of Groundnut-Wheat sequence) 36.5 26.5
T2 (100% NPK of recommended dose of Groundnut-Wheat sequence.) 42.5 32.5
T3(150% NPK of recommended dose of Groundnut-Wheat sequence) 30.0 30.5
T4(100% NPK+ ZnSO4 @ 50 kg/ha once in three year to groundnut only.) 46.5 31.5
T5(NPK as per soil test) 33.5 22.5
T6(100% NP of recommended dose of Groundnut-Wheat sequence) 38.3 33.5
T7(100% N of recommended dose of Groundnut-Wheat sequence) 46.5 36.5
T8(50% NPK + FYM @ 10 t/ha to Groundnut and 100% NPK to wheat) 52.5 42.5
T9(Only FYM @ 25 t/ha to Groundnut only) 49.5 35.5
T10(50% NPK + Rhizobium + PSM to groundnut and 100 % NPK to wheat) 31.5 21.5
T11(100% NPK of recommended dose of Groundnut-Wheat sequence. (P as
SSP)
40.5 28.5
T12 (Control) 29.5 19.5
MEAN 39.7 30.0
S.Em.± 2.3 1.5
C.D. at 5 % 6.6 4.4
C.V. % 11.5 10.3
Table 2. Effect of different treatment on Dehydrogenase Activity in 1st and 12th
Year of LTFE Soils.
JAU, Junagadh Karad et al. (2016)

38. Fig – 14. Impact of long-term organic and inorganic nutrient Managements on
enzyme activities
 Long-term nutrient experiment in semi-arid Alfisol at Coimbatore.
 investigated in two successive years, 2009 and 2010 to assess the effects of
organic manure (OM) and inorganic chemical fertilizers (IC) on phosphatase
activities.
38Tamilnadu Chinnadurai et al. (2014)

39.  For improving N supply and
management in Rice Wheat
Cropping system Sesbania
sesban and the nitrification
inhibitor encapsulated calcium
carbide (ECC) is used widely.
Fig – 15. Effect of green manure and nitrification inhibitor on DHA and Nrse activity
Patra et al. (2006) 39IARI, New Delhi Patra et al. (2006)

40. Fig – 16. Enzymatic activity in soil with different treatment in LTFE soil under a
rainfed soybean–wheat system in N-W Himalaya.
Saha et al. (2008)
40Uttarakhand Saha et al. (2008)

41. Fig – 17. Effect of LTFE on enzymatic activity under a rainfed soybean–wheat
system in N-W Himalaya
Saha et al. (2008)
41Uttarakhand Saha et al. (2008)

42. 42
Effect of management practices on different
kinetic parameters of enzymes

43. What is enzyme kinetics ?
Enzyme kinetics refers in terms of rate at which substrate is
converting into product.
What is utility of studying enzyme kinetics ?
To know the efficiency of enzymes kinetic study is needed.
How to study enzyme kinetics ?
 To know the efficiency of enzymes we should know various
parameters. These parameters comes from the well known
Michaelis-Menten equation :
V= (Vmax × S) / (Km + S)
Where, V=Enzyme activity, Vmax = Maximum velocity of enzyme
reaction, S=Substrate concentration (p-NPP), Km = Michaelis-
Menten constant
 Vmax and Km are called kinetic parameters.
 Km is the substrate conc at which Vmax decreases to its half, which
tells us the affinity of enzyme for substrate and is inversely
proportional to the substrate affinity. i.e lesser the value of the Km,
more will be the affinity of enzyme for substrate.
43

44. Fig – 18. Kinetic parameters as affected by Tillage and no tillage practices
Surface (0-20 cm) meadow brown
soil samples collected from the
plots under no tillage and
conventional tillage in a 7-year
field experiment under maize
cropping.
Higher Vmax value for Pase, Pdse
and Arase in NT system than CT
system.
Lower Km value for Pase, Pdse
and Arase in NT system than CT
system.
For urease reverse is true.
44China Zhang et al. (2010)

45. Fig 19. Kinetic parameters as affected by vermicompost and fertilizers addition
0
1
2
3
4
5
6
7
8
9
10
Control VC VC + Fert Fert
Km (mM)
Vmax
(nmol/g/hr)
b
c
b
a
b b
b
a
a
c c
a
 Vmax increases markedly in organic and organomineral
treatments.
 Km was similar in vericompost treated soil and control soil
while doubled in organomineral and mineral treatments.
45Pisa, Italy Masciandaro et al. (2000)

46. Effect of pesticide addition on activitiy of
enzymes
46

47. Effect of pesticides on soil enzymes
 Pesticides reaching the soil may disturb local metabolism or
enzymatic activities.
 Negative impact of pesticides on soil enzymes like hydrolases,
oxidoreductases, and dehydrogenase activities has been widely
reported in the literature. (Malkomes, 1989; Menon et al., 2005)
 There is also evidence that soil enzymatic activities increased
by some pesticides (Megharaj et al., 1999)
 The effect of pesticides on soil enzymes particularly
extracellular enzymes are not clear due to their
multidimensional behaviour in complex soil medium and the
greater complexity of soil microbial and biochemical
interactions
47

48. A. Dehydrogenase activity
B. Phosphatase activity
Fig – 20. Effect of different concentration of butachlor on DHA and Ptse activities
Un-flooded
Un-flooded Flooded
Flooded
N (23 mg kg-1), N10 (230 mg kg-1) and N100 (23 g kg-1) at -10 cm depth of soil
48
Srinagar, J & K Rasool et al. (2014)

49. Protease :- *μg glucose per gram soil formed after 24 hrs with 1% casein.
Urease :- *μg glucose per gram soil formed after 24 and 72 hrs incubation with 2% starch.
Figures in parentheses indicate relative production percentages.
Table – 3. Effect of different concentrations of selected insecticides on Protease
and Urease activity
49
Kurnool district, AP
Protease Urease
Nasreen et al. (2012)

50. Fig – 21. Effect of pesticide on enzymatic activity under biochar - amended soil
S- control,
SBC30- biochar amended soil – 30kg/ha,
SBC40 -biochar amended soil – 45kg/ha,
50Poland Olesczuk et al. (2014)

51.  Dehydrogenase activity was highest in the non-Bt treatment
followed by Bt-cotton and least in the no-crop treatment.
 It also varied significantly between the four sampling times.
51
Table - 4. Dehydrogenase activity in the rhizosphere soils under Bt(BT) and
non-Bt(NBT) cotton crops.
IARI, New Delhi Sarkar et al. (2008)
NC indicate no crop.

52. Effect of conservation practices on
activity of enzymes
52

53. Treatments
Tillage
No of Irrigation
Conservation Conventional
2
(sub opt)
3
(optimal)
5
(supra opt)
2
(sub opt)
3
(optimal)
5
(supra opt)
T0 139.1b 142.3a 91.3a 73.4a 70.3a 65.1
T1 158.3d 231.5c 172.6b 84.4b 115.6c 87.2b
T2 262.9e 292.5c 166.5d 112.3c 93.7d 82.5c
T3 178.4c 162.6b 112.3b 93.6b 102.4a 106.3a
T4 277.7d 233.2b 211.4a 131.3c 98.9b 152.4c
T5 251.3c 158.7b 131.6e 121.7b 95.6b 121.6b
T6 310.3f 351.1d 236.7a 152.4d 127.4d 163.7d
T7 111.4a 114.4a 152.3a 62.2d 68.7b 75.4a
Urease activities in terms of µg NH4 released /g soil/hr
T0 = No external Fertilizer
T1 = Recommended dose of N (RDN) through urea
T2 = 75% RDN urea + 25% RDN FYM
T3 = 75% RDN urea + 25% RDN GM
T4 = 75% RDN urea + 25% RDN BF
T5 = 75% RDN urea + 25% RDN SS
T6 = 50% RDN FYM + 25% RDN BF + 25% RDN GM
T7 = Blank Plot
Table – 5. Effect of Conservation tillage, optimal Water supply and INM
on urease activity
53IARI, New Delhi Sharma et al. (2013)

54. Fig – 22. Effect of cover crops on arylamidase activity at different depths in
conventional tillage and no-till systems.
54Tuskegee University, United state Said et al. (2009)
arylamidase

55. Fig – 23. Effect of zero tilled and tilled on soil enzyme activities at different depth
surface (0 - 10 cm) and subsurface (10 - 20 cm) layers
55East Midlands, UK Mangalaserry et al. (2015)

56. Fig – 24. Soil phosphatase activities under no-tillage and tillage at different
residue input amounts
Wang et al. (2011)
TR0, TR50, and TR100 are conventional tillage with 0, 50, and 100% residue input
amounts. NTR0, NTR50, and NTR100 are no-till with 0, 50, and 100% residue input
amounts, respectively
56China Wang et al. (2011)

57. Effect of irrigation practices on activity
of enzymes
57

58. 58
Fig – 25. Effects of irrigation induced salinity and sodicity on phosphatase
activities
Zimbabwean Rietz and Haynes (2003)

59. 59
Fig – 26. Effects of irrigation induced salinity and sodicity on arylsulphatase
activities
Zimbabwean Rietz and Haynes (2003)

60. Groundwater (GW), and River water (RW), Sensitive Index (SI) Catalase activity (CAT),
dehydrogenase (DEH), alkaline phosphatase (ALP), β-glucosidase (GLU), arylsulphatase (ARY)
and urease (URE) activities
The sensitive index (SI) related to irrigation treatments for labile organic Carbon fractions
and enzyme activity was calculated using the following equation:
mean of RW irrigated value – mean of GW irrigated value
mean of GW irrigated value
Table – 6. Effect of Different sources of irrigation on enzyme activities of
soil
60China Gao et al. (2015)
GLU URE ALP ARY DEH CAT
Sep 2012
GW 71.8 153.1 399.2 34.5 142.7 0.58
RW 116.4 213.0 496.6 52.1 202.1 0.67
SI 62.1 39.1 24.4 51.0 41.6 14.9
May 2013
GW 74.2 161.1 418.3 35.7 157.4 0.65
RW 134.6 235.9 532.6 69.5 223.4 0.71
SI 81.4 46.4 27.3 95.0 41.9 8.8
Mean
GW 73.0 157.1 408.8 35.1 150.0 0.62
RW 125.5 224.4 514.6 60.8 212.7 0.69
SI 71.9 42.9 25.9 73.3 41.8 11.7
SI = ×100

61. Fig – 27. Effect of long term irrigation on the profile distribution of
enzyme activity of typical chernozem soil
61Republic of Moldova Senikovskaya and Filipchiuk (2011)

62. Conclusions
crop rotation greatly influences the enzyme activities such as
urease, arylsulphatase, dehydrogenase etc. in soil.
Long term application of compost and/or FYM along with NPK
resulted in stimulation of all types of enzyme activities.
soil mulching enhances the phosphatase and aryasulphatase
activity.
lime application in acid soil has favorable effect on the activity
of alkaline phosphatase.
Organic farming system has improved kinetic properties (in
terms of Vmax) than conventional one but Km remain
unchanged.
62

63. 63
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