In last 250 years, CO2 levels have increased from 280 to 390 ppm (IPCC 2007). Elevated CO2 reduces N content and alters the concentrations of secondary metabolites in plants (Sun et al. 2013). Therefore, the present studies were undertaken on the effects of increased levels of CO2 on enzymatic activity and secondary metabolites in chickpea. Helicoverpa armigera damages chickpea plants at all the stages.

1. Conclusions
Amounts of polyphenols and condensed tannins increased in plants kept at elevated
levels of CO2. Increased activities of PAL, TAL, PPO, and POD under high CO2 might
have resulted in increased production of C-based secondary metabolites, which will
increase the levels of plant resistance to insects.
References
Intergovernmental Panel on Climate Change. 2007. Climate Change 2007; the physical science basis.
Summary for policy makers. Report of Working Group I of the Intergovernmental Panel on Climate
Change. [WWW document]. Available: http://www.ipcc.ch/pub/spm18-02.pdf
Sun YC, Cao HF, Yin J, Kang L and Ge F. 2010. Elevated CO2 changes the interactions between
nematode and tomato genotypes differing in the JA pathway. Plant Cell Environ 33: 729–739.
Sharma HC. 2005. Heliothis/Helicoverpa management: Emerging Tends and Strategies for Future
Research. New Delhi, India: Oxford and IBBH Publishing Co. 469 pp.
* For more information, please write to: Dr HC Sharma, Principal Scientist – Entomology, ICRISAT. Email: h.sharma@cgiar.org
Introduction
In last 250 years, CO2 levels have increased from 280 to 390 ppm (IPCC 2007). Elevated
CO2 reduces N content and alters the concentrations of secondary metabolites in plants
(Sun et al. 2013). Therefore, the present studies were undertaken on the effects of
increased levels of CO2 on enzymatic activity and secondary metabolites in chickpea.
Helicoverpa armigera damages chickpea plants at all the stages (Plate 1).
Materials and Methods
Chickpea plants: Fifteen-day old chickpea plants of ICCL 86111 (R) and JG 11 (S),
were transferred to open top chambers (OTC) from the greenhouse (Plate 2). Three CO2
levels (350, 550, and 750 ppm) were maintained in the OTCs connected to CO2 cylinders.
There were three replications for each genotype, 10 pots in each replication, with 5 plants
in each pot. One set of chickpea genotypes were kept outside the OTCs under ambient
conditions.
Larval infestation: Neonates of H. armigera larvae were released on the chickpea
plants. Each plant was infested with 20 neonates of H. armigera. Uninfested plants were
maintained as a control. After 7 days of infestation, data were recorded on insect damage,
larval survival, and larval weights.
Estimation of secondary metabolites and the enzymatic activities: After
7 day of infestation, amounts of secondary metabolites (phenols, tannins) and hydrogen
peroxide (H2O2), and the activities of defensive enzymes [peroxidase (POD), polyphenol
oxidase (PPO), phenylalanine ammonia lyase (PAL) and tyrosine ammonia lyase (TAL)]
were estimated using standard protocols. Amounts of chlorophyll, polyphenols and
nitrogen basal index (NBI) were estimated by using Force A phenol meter.
CO2
Concentration
(ppm)
Polyphenols (μg cm-2) Chlorophyll (μg cm-2)
JG 11 ICCL 86111 JG 11 ICCL 86111
Infested Control Infested Control Infested Control Infested Control
Control 1.22±0.13 1.06±0.10 0.99±0.1 0.96±0.1 27.4±2.3 21.9±3.2 16.4±3.4 29.5±5.5
350 0.97±0.12 0.43±0.07 1.03±0.2 0.64±0.1 30.2±1.9 17.2±1.2 28.2±2.5 28.6±1.7
550 1.36±0.14 1.02±0.10 1.39±0.1 0.98±0.1 19.6±1.0 30.7±1.4 23.3±1.6 27.8±2.0
750 1.36±0.13 0.86±0.19 1.27±0.1 0.47±0.1 17.7±2.1 14.9±1.2 21.7±1.1 17.4±1.5
Fig. 5. POD (A) and PPO (B) activities of H. armigera infested and uninfested chickpea plants
under different CO2 concentrations, and under ambient conditions.
Fig. 1. Tannins (A,B) and NBI (C,D), of H. armigera infested and uninfested chickpea plants
under different CO2 concentrations, and ambient conditions. NBI = Nitrogen basal index.
Table 1. Polyphenols and chlorophyll contents of insect infested and uninfested chickpea plants
under different CO2 regimes .
Results
Amounts of condensed tannins were greater in plants raised at 550 ppm of CO2 than in
plants raised at 350 ppm (Table 1, Fig. 1A). Tannin content was more in un-infested
chickpea plants under ambient conditions than at 350 ppm (Fig. 1B). Polyphenol
content increased with elevated CO2 (Table 1). The NBI was more in plants kept at 350
ppm CO2 than in plants kept under ambient conditions (Fig. 1c,d). The H2O2 content
was more in plants kept under ambient conditions than those kept at 350 ppm CO2 (Fig.
2A, B). The PAL and TAL activities increased up to 550 ppm CO2 (Fig. 3, 4). The H.
armigera infested plants showed greater PAL and TAL activities than the control plants.
Plants at 350 ppm CO2 had higher amounts of TAL than those under ambient condition
in JG 11. The POD and PPO activities showed progressive increase with elevated levels
of CO2 (Fig. 5A, B).
Fig. 3. Phenylalanine ammonia lyase (PAL) activity of H. armigera infested and uninfested
chickpea plants under different CO2 concentrations, and ambient conditions.
A
B
0
1
2
3
4
5
6
7
350 ppm 550 ppm 750 ppm
Tannins(mgCE/gFW)
CO2 conc.
JG 11 IN JG 11 UI ICCL 86111 IN ICCL 86111 UI
0
20
40
60
80
100
120
140
160
180
350 ppm 550 ppm 750 ppm
NBI
CO2 conc.
ICCL 86111 UI ICCL 86111 IN JG 11 UI JG 11 IN
0
5
10
15
20
25
350 ppm 550 ppm 750 ppm
H2O2(μmol/gFW)
CO2 conc.
JG 11 IN JG 11 UI ICCL 86111 IN ICCL 86111 UI
Fig. 2. H2O2 (A,B) content of H. armigera infested and uninfested chickpea plants under different
levels of CO2 and ambient conditions.
0
2
4
6
8
10
12
14
16
18
350 ppm 550 ppm 750 ppm
PAL(OD/min/gFW)
CO2 conc.
JG 11 UI JG 11 IN ICCL 86111 UI ICCL 86111 IN
0
2
4
6
8
10
12
14
16
350 ppm 550 ppm 750 ppm
TAL(OD/min/gFW)
CO2 conc.
JG 11 UI JG 11 IN ICCL 86111 UI ICCL 86111 IN
0
1
2
3
4
5
350 ppm 550 ppm 750 ppm
POD(OD/min/gFW)
CO2 conc.
JG 11 IN JG 11 UI ICCL 86111 IN ICCL 86111 UI
0
0.2
0.4
0.6
0.8
1
1.2
350 ppm 550 ppm 750 ppm
PPO(OD/min/gFW)
CO2 conc.
JG 11 UI JG 11 IN ICCL 86111 UI ICCL 86111 IN
Plate 2. Open top chambersPlate 1. Helicoverpa armigera infested chickpea (a) leaves and (b) pods
a b
0
1
2
3
4
5
6
IN UI IN UI
JG 11 ICCL 86111
Tannins(mgCE/gFW)
Genotypes
Ambient conditions 350 ppm
0
10
20
30
40
50
60
IN UI IN UI
JG 11 ICCL 86111
NBI
Genotypes
Ambient conditions 350 ppm
0
5
10
15
20
25
IN UI IN UI
JG 11 ICCL 86111
H2O2(μmol/gFW)
Genotypes
Ambient conditions 350 ppm
B
C D
A B
A
Fig. 4. Tyrosine ammonia lyase (TAL) activity of H. armigera infested and uninfested chickpea
plants under different CO2 concentrations, and ambient conditions.
July 2014ICRISAT is a member of the CGIAR Consortium
Effect of CO2 on Enzymatic Activity, Secondary Metabolites and Expression of Resistance to
Helicoverpa armigera
Abdul Rashid War, S MD Akbar, RS Munghate, M Pathania, SP Sharma and Hari C Sharma*
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru 502 324, Telangana, India