PHYSIOLOGICAL BASIS OF SEED HARDENING IN CHICKPEA

PHYSIOLOGICAL BASIS OF SEED HARDENING

IN CHICKPEA (Cicer arietinum L.)

Thesis submitted to the University of Agricultural Sciences, Dharwad in partial fulfilment of the requirements for the

Degree of

MASTER OF SCIENCE (AGRICULTURE)

IN

CROP PHYSIOLOGY

By

MANJUNATHA B. L.

DEPARTMENT OF CROP PHYSIOLOGY COLLEGE OF AGRICULTURE, DHARWAD

UNIVERSITY OF AGRICULTURAL SCIENCES, DHARWAD-580 005

JULY, 2007

ADVISORY COMMITTEE

DHARWAD (M.M. DHANOJI) JULY, 2007 MAJOR ADVISOR

Approved by :

Chairman :

__________________________

(M.M. DHANOJI)

Members : 1. _________________________

(M.B. CHETTI)

2. _________________________

(S.M. HIREMATH)

3. _________________________

(B.S. VYAKARANHAL)

CONTENTS

Sl. No.

Chapter Particulars

CERTIFICATE

ACKNOWLEDGEMENT

LIST OF TABLES

LIST OF FIGURES

1. INTRODUCTION

2. REVIEW OF LITERATURE

2.1 Effect of Seed hardening on morpho-physiological parameters

2.2 Effect of Seed hardening on growth parameters

2.3 Effect of Seed hardening on biochemical parameters

2.4 Effect of Seed hardening on yield and yield components

3. MATERIAL AND METHODS

3.1 Experimental site

3.2 Climatic conditions

3.3 Soil and its characteristics

3.4 Experimental details

3.5 Cultural operations

3.6 Collection of experimental data

3.7 Biochemical parameters

3.8 Statistical analysis

4. EXPERIMENTAL RESULTS

4.1 Morphological characters

4.2 Growth Parameters


4.4 Yield parameters

5. DISCUSSION

5.1 Morpho-physiological parameters

5.2 Dry matter production and its distribution

5.3 Growth parameters


5.5 Yield and yield components

5.6 Future line of work

6. SUMMARY AND CONCLUSIONS

7. REFERENCES

LIST OF TABLES

Table No.

Title

1. Monthly metrological data for 2007 and average 55 years (1950-2004) at Main Research Station, University of Agricultural Sciences Dharwad.

2. Physical and Chemical properties of the soil from experimental site

3. Influence of seed hardening chemicals on plant height (cm) at different stages in chickpea

4. Influence of seed hardening chemicals on number of leaflets at different stages in chickpea

5. Influence of seed hardening chemicals on leaf dry weight (g plant-1

) at different stages in chickpea

6. Influence of seed hardening chemicals on stem dry weight (g plant-1


7. Influence of seed hardening chemicals on reproductive dry weight (g plant-1


8. Influence of seed hardening chemicals on total dry matter (g plant-1


9. Influence of seed hardening chemicals on leaf area (cm-2


10. Influence of seed hardening chemicals on leaf area index at different stages in chickpea

11. Influence of seed hardening chemicals on specific leaf weight(mg cm-2


12. Influence of seed hardening chemicals on leaf area duration (days) at different stages in chickpea

Contd…..

Table No.

Title

13. Influence of seed hardening chemicals on absolute growth rate(g plant-1

day-1


14. Influence of seed hardening chemicals on crop growth rate (g dm-2

days-1


15. Influence of seed hardening chemicals on relative growth rate (g g-1

day-1

) and net assimilation rate (g dm

-2 day

-1) at different stages in chickpea

16. Influence of seed hardening chemicals on biomass duration (g-days) at different stages in chickpea

17. Influence of seed hardening chemicals on specific leaf area (cm2) at different

stages in chickpea

18. Influence of seed hardening chemicals on leaf area ratio(cm2 g

-1) at different

stages in chickpea

19. Influence of seed hardening chemicals on relative water content (%) at different stages in chickpea

20. Influence of seed hardening chemicals on chlorophyll ‘a’ and ‘b’ content (mg g

-1 fr.wt.) at different stages in chickpea

21. Influence of seed hardening chemicals on total chlorophyll content ( mg g-1

fr.wt.) at different stages in chickpea

22. Influence of seed hardening chemicals on chlorophyll stability index (%) at different stages in chickpea

23. Influence of seed hardening chemicals on proline content (µg g-1


24 Influence of seed hardening chemicals on epicuticular wax content (mg dm-2


25. Influence of seed hardening chemicals on yield and yield components at different stages in chickpea

LIST OF FIGURES

Figure No.

Title

1.

Plan of layout of the experiment

2.

Influence of seed hardening chemicals on leaf dry weight (g plant-1) at different stages in chickpea

3.

Influence of seed hardening chemicals on stem dry weight (g plant-1) at different stages in chickpea

4.

Influence of seed hardening chemicals on reproductive dry weight (g

plant-1) at different stages in chickpea

5.

Influence of seed hardening chemicals on total dry matter (g plant-1) at different stages in chickpea

1. INTRODUCTION

India is the major pulse growing country in the world accounting 68.32 m ha area (35.2%) and 57.51 m tons (26.65%) of world production. Pulses play an important role in Indian agriculture, because of their inherent ability to fix atmospheric nitrogen through biological nitrogen fixation (BNF), which is economically sound and environmentally acceptable (Anon., 2003). The pulses are known to improve the physical characteristics of soil by virtue their deep penetrating tap root system, which contribute substantially to the loosening up of soil and also enable them to utilize available moisture efficiently. Pulses have been traditionally recognized as indispensable basic ingredient in the diets of vast majority of vegetarian Indians as they provide a perfect mix of high biological value in cereal based diet.

The ever increasing population of the country coupled with inadequate progress in the pulse production is posing serious challenges on the food front. The progressive decline in per capita availability of pulses (69 g in 1961 to 30 g in 2002) in India is a matter of great concern. This is attributed to steady marginalization of their cultivation in the wake of “Green Revolution” and burgeoning population with assured supply of cereals at an affordable price. To alleviate protein-energy malnutrition, a minimum of 50 g pulses per capita per day is required, in addition to other sources of protein such as cereals, milk, meat, eggs, etc. To make up this shortfall in supply, besides further demand from burgeoning population, about 20 m tons of pulses are required by 2007, which is expected to touch 28 m tons by 2020. This can be realized only by adopting increasingly more productive technologies along with aggressive developmental efforts and favourable Government policies (Bangal et. al., 1984).

Per cent contribution of pulses in total food grain production in India has declined during the last three decades. Pulses are grown on 22.24 m ha area, producing 13-15 m tons. The production of pulses reached to its highest (14.91 m tons) during 1999. There was a considerable decline in pulses production during 2000-01 (11.06 m tons). This sudden decline was mainly due to reduction in area under rabi pulses, because of early cessation of monsoon and failure of winter rains.

In order to get rid the people from the shocks of protein malnutrition, the per capita availability of pulses has to be stepped up. Based on the projections made elsewhere, 20 m tons of pulses are required by 2007, which is expected to touch 28 m tons by 2020. This can be realised by adopting more productive technologies along with aggressive developmental efforts.

Among the pulses, chickpea (Cicer arietinum L.) is the third most important food legume grown in over 45 countries in all continents of the world. It provides a high quality protein to the people in developing countries. It is one of the important crops grown during rabi season. On an average, it produces 126 kg protein per hectare and it is probably the highest protein yielding legume next to groundnut and soybean. In India, chickpea ranks first among the legumes in area occupying of 4.8 m ha (27.0 %) and with a production of 3.5 m tons (38.0 %) and productivity level of 720 kg/ha. Similarly in Karnataka it is grown in an area of 3.7 lakh ha with an annual production of 2.39 lakh m tons with an average productivity of 650 kg/ha (Anon., 2003).

The average yield of chickpea in India is very low, which might be due to the cultivation of this crop on residual soil moisture in cool dry season. It is generally grown on conserved soil moisture and the moisture in the profile gradually receeds as the crop grows. As a consequence, plant experience progressively increasing degree of terminal moisture stress. Thus, soil moisture stress assumes a major limiting factor determining the growth and yield of chickpea in peninsular India. Such situation particularly affects the pod formation, which is critical for determining the yield potential (Verma and Pramilakumari, 1978). Added to this, some of the important physiological constrains in productivity are intensive flower dropping, poor pod filling, low dry matter accumulation and pod shedding (Singh et al.,1991).

Moisture stress is one of the abiotic stresses, which affect the productivity. Research management practices aimed at overcoming abiotic stress limitations to increase yield have demonstrated that significant progress can be made. Most of the research work done so far has been on understanding the mechanism underlying productivity, but very little work has

been done with respect to the possibilities of overcoming stresses or limitations imposed by environmental factors.

There is an urgent need to identify suitable ameliorative measures to overcome the effect of moisture stress. There are certain avenues to induce drought tolerance under receeding soil moisture condition viz., seed hardening technique by using different chemicals. Pre-sowing seed hardening of seeds is one of the best methods that results in modifying the physiological and biochemical nature of seed so as to get the characters that are favourable for drought resistance. Pre-sowing seed hardening is the result of extensive physiological recognisation induced by dehydration process. But, the study on these aspects in chickpea is very meager and therefore there is an urgent need to improve the productivity potential of chickpea under receeding soil moisture conditions.

Keeping these views, the present investigation was undertaken with the following objectives:

i. To study the effect of seed hardening on morpho-physiological traits in chickpea.

ii. To study the effect of seed hardening on biophysical and biochemical parameters in chickpea.

iii. To study the effect of seed hardening on yield and yield parameters in chickpea.

2. REVIEW OF LITERATURE

Pulses, besides being an indispensable component of vegetarian diet, play a vital role in sustaining long term productivity of soil through biological nitrogen fixation. Chickpea (Cicer arietinum L.) is considered as one of the important grain legume crops because of its high nutritional value. In India, it is extensively cultivated as a winter crop, since it thrives well under receeding soil moisture condition. However, the productivity depends largely on the efficient utilization of available soil moisture.

The study on seed hardening in chickpea is very meager. Hence, an attempt has been made to survey the available literature pertaining to the increase in productivity potential in receeding soil moisture by using seed hardening chemicals and reviewed hereunder in this chapter.

2.1 Effect of Seed hardening on morpho-physiological parameters

Seed hardening has been reported to induce drought resistance in the plants, such seeds as indicated by its capacity to withstand dehydration and overheating. Other beneficial effects of hardening are inducing better root growth, higher rate of photosynthesis and larger dry matter accumulation (Henckel, 1964).

In wheat pre sowing seed treatment with 5% cycocel, resulted in lower height and parts were markedly dark green. The length of coleoptile, lengths of first leaf, total shoot length and dry weights of shoots were all reduced (Appleby et al., 1966).

Abdel Hafeez and Hudson (1967) reported that in the moist soil, plants from hardened seeds grow better and produced significantly more dry weight than unhardened plants. Irrespective of soil fertility status the seed hardened plants had significantly lower leaf area than unhardened plants. Salim and Todd (1968) adopted pre sowing seed treatment in wheat and barley and found that seeds are capable of germinating in higher concentration of mannitol. Austin et al., (1969) recorded 51% increase in length of the carrot embryos in hardened seeds, which were hardened compare to the unhardened seeds. Studies conducted with finger millet using CaCl2, ascorbic acid, kinetin and benzyl adenine shown to have a greater beneficial effects interms of germinability and seedling growth under simulated water conditions or salinity (Krishna sastry et.al., 1969). Similarly, Woodruff (1969) adopted the technique of seed hardening in wheat and found that the plants maintained higher leaf relative water content (RWC) under moisture stress condition as compared to unhardened plants.

Mehrotra et.al., (1970) observed reduction in the height of the okra plants by application of cycocel either to soil at three leaf stage or by soaking the seeds in 100 ppm for 24 hours. They found that the flowering was delayed by seed hardening treatment with cycocel as compared to soil application. Similarly in tomato soil application of cycocel did not affect the number of leaves, while soaking seeds in cycocel (250 mg/l) significantly increased the number of leaves (Linkens et.al., 1964). Similar results were obtained by Arora et.al., (1990) with cycocel (100 ppm) both as seed and foliar treatments.

Rajasheker et al. (1970) reported that in ragi pre sowing seed hardening increased the root growth by 27 per cent than unhardened seeds. Where as, Sadonzev et al. (1970)found that seed soaking with 10 per cent cycocel for 4 hours delayed emergence by 1 to 3 days in winter wheat.

Singh et al. (1975) indicated that, pre sowing seed hardening in barley helped the seeds to germinate a day earlier than the unsoaked seeds resulting in establishment of the crop. The grain and straw yield was also significantly increased with soaked seeds than with the unsoaked seeds. Similarly,

Pre soaking seed hardening with water increased green matter production, number of green leaves per plant, plant height, root length and seed production in grain sorghum (Corleto et al. 1977). Kamala Thirumalaiswamy and Sakharam Rao (1977) reported that, seed treatment with distilled water, 5 ppm CCC, 10 ppm Resistine and 5 ppm Kinetin slightly

increased NAR , RGR and leaf area in pearl millet. Irrespective of moisture level the size of the leaf was greatly influenced by cycocel followed by Kinetin, Resistine.

Singh and Thakur (1979) reported that in soybean the improvement in seed germination per cent to the extent of 49, 43, 71, 44 and 47 per cent by pre soaking with water, cobalt (0.1 and 1.0 ppm), molybdenum (1.0 and 2.0 ppm) respectively, over unsoaked seeds and germination relative index (GRI) to the extent of 108, 67, 26, 92 and 102 respectively, over control.

Misra and Dwivedi (1980) found that ragi seeds (Eleusine coracana) treated with potassium and distilled water recorded significantly more plant height , enhanced tiller production , green leaf number , leaf area and dry weight of shoot as compared with the control. Channakhesava (1982) reported that the seed treatment of sorghum (cv CSH 5) with distilled water, glucose and gibberellic acid has shown early growth, increased plant height and dry matter production.

De et al. (1982) noticed that in wheat seed soaking with cycocel decreases the shoot growth and leaf area per plant progressively with increase in concentration of the chemical by 10, 40 and 80 mg/l. while the radical growth was increased considerably. Dighe et al. (1983) indicated that , cycocel (500 ppm) applied to wheat cultivars by either seed treatment and foliar spray or soil drenching (1000 ppm) decreased plant height and delayed flowering and maturation by 4 to 5 days as compared to control.

Singh et al. (1984) stated that soaking of sorghum seeds in water for 12 hours over night hastened the flowering period by four days as compared to control. These four days of early in flowering can be utilized for solving synchronization problem in CSH-5 seed production.

Karivaratharaju and Ramakrishnan (1985) reported that pre-sowing seed hardening of ragi seeds in different chemical solutions significantly increased the germination, tillering, plant height, root growth and dry matter production.

Bhatia and Rathore (1986) studied the effect of seed soaking with distilled water, 5% KH2PO4, 0.25% CaCl2, 0.1% cycocel, 2.5% NaCl and saturated solutions Ca (OH) 2 on germination and seedling attributes in wheat. They found that treatment with KH2PO4 enhanced the germination. Increased the number of seedlings per meter row length, Dry matter accumulation and seedling height over control. Similarly seed hardening with 2% CaCl2 in the ratio 1:1 (seed:solution) for four hours increased drought resistance in sorghum. There was an improvement in the germination, seedling growth and development, higher relative water content (RWC) and root to shoot ratio (Patil, 1987).

Mandal and Basu (1987) indicated that pre-sowing seed hardening with water increased the germination by 10 %, shoot growth by 8% and root growth by 17 % in wheat over the control. Arjunan and Srinivasan (1989) indicated that seed hardening with CaCl2 (1 %) in groundnut has given significantly higher pod yield through increased germination per cent and higher dry matter accumulation. Eshanna and kulkarni (1990) revealed that seed treatment with CaCl2 in the ratio of 1:3 (seed: solution) recorded significantly higher plant height, LAI ,CGR,NAR and total dry matter at different growth stages in maize as compared to control. Seed hardening with 10

-5 M GA3, kinetin or IAA or 10

-4 cycocel generally increased

number of leaves per plant, root, stem and total dry weight, RGR, NAR, leaf weight ratio in wheat as compared to control (Gurudev Singh et al., 1991). Similarly, foliar spray of CCC at 1000 ppm increased the LAI, LAD, NAR, CGR and total dry matter production followed by cytozyme and CCC at 1000 and 500 ppm in ground nut (Nawalagatti et al., 1991)

Rabai et al. (1991) observed that seeds soaked in 100, 250 or 500 ppm cycocel lowered the abscission of buds, flowers and pods by 3.6-6.4 and 0.8-3.2 per cent in vicia faba cv. Giza-1 and Giza-402 respectively as compared to control. Singh and Kakralya (1992) reported that 50 mg/l of ethrel or GA or benzyl adenine may be applied as pre sowing seed soaking treatments for 12 hours to improve germination per cent, crop stand density, seedling growth and yield of pigeonpea. Rangaswamy et al. (1993) found that seed hardening with CaCl2 at 0.4% and CCC at 0.2% increased the germination per cent, vigour index and root to shoot ratio in sorghum, pigeonpea, groundnut and cowpea. In another study by Jayarami Reddy et al. (1996) reported that foliar spry of NAA (10 ppm) + KNO3 (0.5%) and NAA

(20ppm) + KNO3 (0.5%) recorded the maximum dry matter production in pigeon pea as compared to control.

Upadhyay, (1994) observed increase in the plant height, number of branches, number of flower bud, number of flowers, vegetative growth and yield in chick pea due to foliar application of NAA (10, 20 and 30 ppm), kinetin (10, 20 and 30 ppm) and KNO3 (100, 200 and 300 ppm)over control. Maitra et al. (1998) studied the effect of seed soaking treatments with distil water and agrochemicals (0.25% CaCl2, 100 ppm KH2PO4 and 100 ppm Na2HPO4) on growth and productivity in fingermillet. Treatment with 100 ppm Na2HPO4 caused remarkable improvements in growth attributes such as plant height and dry matter accumulation at different growth stages. Where as Govindan and Thirumurugan (2000) revealed that the growth parameters like plant height, LAI and dry matter production in greengram were significantly high with treatments received foliar spray of KNO3 (1%) or KCl (1%) and their combination.

Punithavathi and Palaniswamy (2001) revealed that ragi seeds soaked in 1% concentration of KCl, CaCl2, prosopis as well as pungam for 12 hours recorded higher germination per cent as well as other seed quality parameters like root length, shoot length, vigour index and dry matter production. Similarly, Pawar et al. (2003) revealed that seed hardening with 2% CaCl2 recorded significantly higher plant height, total dry matter and grain yield in sunflower as compared to control.

2.2 Effect of Seed hardening on growth parameters

The technique of growth analysis has been extensively used for better understanding of the physiological basis of yield variation in crop plants. Growth analysis is a physiological probe on the development of the crop in a chronological sequence to elucidate and account the causes for difference in yield through the events that have occurred at different stages of growth. (Krishnamurthy et al., 1997).

Extensive studies have been made on the physiological analysis of growth parameters in cereals, pulses and oilseeds emphasizing their importance in yield analysis (Chhonkar and Singh, 1959). Kamal Thirumalaiswamy and Sakharam Rao (1977) reported that seed treatment with distilled water, 5 ppm CCC and 5 ppm kinetin and moisture levels slightly increased net assimilation rate (NAR), relative growth rate (RGR) and leaf area in pearlmillet and irrespective of moisture level the size of the leaf was greatly influenced by cycocel followed by kinetin.

Specific leaf weight (SLW) is a stable character and was initially low and improved subsequently and reached the maximum value at 42 days in greengram genotypes (Kalubarme and Pandey, 1979). Crop productivity is mainly determined by crop growth rate (CGR) which depends on leaf area index (LAI) and then rate of photosynthesis. The CGR was found to be low during early vegetative stage but increased with the advancement of growth in greengram (Kalubarme and Pandey, 1979). They further reported that NAR and RGR decreased with the advancement of crop growth. In another study Misra and Dwivedi (1980) found that ragi seeds treated with potassium and distilled water produced distinctly more leaf area and shoot dry weight compared with control.

De et al. (1982) noticed that in wheat, seed soaking with cycocel decreases the shoot growth and leaf area per plant progressively with increase in concentrations of the chemical by 10, 40 and 80 mg/l, the radicle growth was increased considerably. Pre sowing seed hardening of ragi seeds in different chemical solutions increased significantly the germination, root growth and dry matter production (Karivaratharaju and Ramakrishna, 1985).

Bhatia and Rathore (1986) studied the effect of seed soaking treatments with distilled water, 5% KH2PO4, 0.25% CaCl2, 0.1% cycocel, 2.5% NaCl and saturated solutions Ca(OH)2 on germination and seedling attributes in wheat. They found that treatment with KH2PO4 enhanced the germination and increased the number of seedlings per meter row length. Dry matter accumulation and seedling height was increased by KH2PO4 followed by distilled water and both were significantly superior over control. Similarly seed treatment with 2% CaCl2 in the ratio 1:1 (seed: solution) for four hours increased drought resistance in sorghum. There

was an improvement in the germination, seedling growth and development, higher relative water content (RWC) and root to shoot ratio (Patil, 1987).

Sumabai et al. (1987) indicated that foliar spray of ascorbic acid, NAA and GA produced increased assimilate production favoured longer reproductive growth and increase in LAI, LAD and NAR in greengram. Arjunan and Srinivasan (1989) showed that seed hardening in groundnut with CaCl2 (1.0%) has given significantly higher pod yield through increased germination per cent and higher dry matter accumulation.

Eshanna and Kulkarni (1990) observed that seed treatment with CaCl2 in the ratio of 1:1 (seed : solution) recorded significantly higher plant height, LAI, CGR, NAR and total dry matter at different growth stages in maize as compared to control. Similarly, seed hardening with cycocel generally increased root, stem and total dry weight, RGR, NAR and leaf weight ratios in wheat as compared to control (Gurudev Singh et al., 1991).

Nawalagatti et al. (1991) found that foliar spray of CCC at 1000 ppm increased the LAI, LAD, NAR, CGR and total dry matter production followed by cytozyme and CCC at 1000 and 500 ppm in groundnut. Similarly Jayarami Reddy et al. (1996) reported that foliar spray of NAA (10 ppm) + KNO3 (0.5%) and NAA (20 ppm) + KNO3 (0.5%) recorded the maximum dry matter production in pigeonpea as compared to control.

Maitra et al. (1998) studied the effect of seed soaking treatments with agrochemicals (distilled water, 0.25% CaCl2, 100 ppm KH2PO4 and 100 ppm Na2HPO4) on growth in fingermillet. Treatment with 100 ppm Na2HPO4 caused remarkable improvements in growth attributes such as LAI, LAD, CGR and dry matter accumulation at different growth stages. Similarly Thirumurugan (2000) revealed that the growth parameters LAI and dry matter production in greengram were significantly high with treatments received foliar spray of KNO3 (1%) or KCl (1%) and their combination.

Punithavathi and Palaniswamy (2001) indicated that ragi seeds soaked in 1% concentration of KCl, CaCl2, for 12 hours recorded higher germination per cent as well as root length, shoot length, vigour index and dry matter production. In another study Sinha et al. (2002) found that ascorbic acid treated moong and pea plants showed an increase in the nodule number and dry weight of the plants as compared to the control.

2.3 Effect of Seed hardening on biochemical parameters

Cheema et al. (1975) revealed that seed hardening of the barley with 0.5 per cent cycocel improved the chlorophyll content to the extent of 5 to 17 mg/g of fresh leaf tissue , protein content in grains and also the relative leaf water content of the leaf tissue. Das and Shekar (1981) obtained significant increase in the chlorophyll content in rice and wheat with the application of 0.5 per cent KNO3 . Shashidhar et al. (1981) reported that ground nut cv RS-218 treated with 1% CaCl2 proline content 17 times more than the control . Pre-sowing seed treatment with 0.25% CaCl2 and soil application of 40 kg P2O5/ha recorded higher grain protein percentage in wheat (Avijit Sen and Misra, 1987). Similarly, the seed treatment with 2% CaCl2 recorded higher RWC and proline content in sorghum (Patil, 1987).

Amaregowda et al., (1994) revealed that seed treatment with CaCl2 (2%) in wheat increased free proline content and RWC at both 60 and 80 DAS. this treatment also increase chlorophyll b-content, accumulated maximum k-content , total sugar content in leaves.

Pre-sowing seed hardening with water NaH2PO4, triazol, KCl and cowdung increased chlorophyll content, RWC and proline in rice (Sheela and Thomos Alexander, 1995 ).Ghosh and Srivastava (1995) revealed that the foliar spray of Quercus serrata seedlings with 5 mM KCl resulted in higher level of total chlorophyll , total sugars, soluble protein and nitrate reductase activity in the leaves the pre- sowing treatments with 100 ppm of CaCl2, KNO3, ascorbic acid, pyridoxine hydrochloride , IAA and GA increasesd chlorophyll content in Pennisetum americanum and Sorghum bicolor (Kadiri and Hussaini,1999).

Ramesh et al.,(2001)noticed during 1998 and 1999 that the foliar spray of 25 ppm benzaldenine along with 2%DAP and 2% KCl enhanced leaf chlorophyll-a contents (1.10 and 1.14 mg/g fresh weight) chlorophyll-b contents (0.43 and 0.49 mg/g fresh weight) respectively in sorghum.

2.4 Effect of Seed hardening on yield and yield components

Grain yield is ultimate economic product of the crop, which is determined mainly by grain weight and number of grains per unit land area. Most of the yield components show direct influence on grain yield. Investigations made by Appleby et al. (1966) revealed that increase in test weight and grain yield of wheat with 5 per cent cycocel seed treatment.

Austin et al. (1969) noticed that plants from hardened seeds of carrot gave the mean yields of roots 64.0 tons compared to 59.2 tons per hectare from untreated seeds. Woodruff (1969) indicated that pre sowing seed hardening increased the grain yield in wheat by 20 per cent. Rajashekhar et al. (1970) reported that, with the use of hardened ragi seed, it was found to increase the yield on an average 12 per cent more than the unhardened seeds. Hardened and better responded to fertilizers. Application of CCC through soil or by soaking the seeds at 50 mg/l water resulted in a significantly higher yield per plant in okra (Mehrotra, 1970). Seed soaking with water increased grain yield by 21 and 12 per cent in two varieties of sorghum i.e. 5-4-1-9 and M-35-1, respectively as compared to the control (Parvatikar et al., 1975)

Filatov and Frolova (1975) found that hardening induced heat tolerance in sunflower and the hardened plants produced more seed yield from 150 to 300 kg/ha compared to unhardened plants. The increase in yield was due to high photosynthetic rate. Pre sowing seed hardening treatment increased early emergence by 13 per cent and yields by 60 per cent in carrot. Whereas in lettuce emergence increased by 9 to 11per cent and yields by 10 to 12 per cent (Pantielev et al., 1976).

Karnail Singh (1996) reported that, seed cotton yield was increased significantly with application of cycocel @ 80g ai/ha over control at 40 DAS. Nayeem and Bapat (1976) coducted experiment on sorghum cv R-16,36A x PD-3-1-11 and M-35-1. The seeds of sorghum cv R-16,36A x PD-3-1-11 and M-35-1were soaked in CaCl2 solution and distilled water for 24 hours and then the seeds were dried for few days in air and they found that among the treatments , the water soaked seeds increased a yield by 1.0,44.6and3.05 per cent respectively over control.

Misra and Dwivedi (1980) reported that, seed treatment with potassium and distilled water distinctly produced more grain and straw yields as compared to control in wheat. Sidhu et al. (1980) concluded that the foliar spray of ZnSO4 (0.5%) recorded maximum fruit weight and yield over other treatments in peech. Das and Sarkar (1981) revealed that post flowering foliar spray with 0.5 % KNO3 solution has given higher yield of both grain and straw in rice and wheat. Seed soaking of wheat in 0.5 % cycocel prior to sowing increased the grain yield (De et al., 1982).

The experiment conducted by Dighe et al. (1983) opined that among the different methods of cycocel treatments, the seed treatment and foliar spray of cycocel in wheat produced higher grain yield than the application of cycocel through soil drenching. Seed treatment with 0.25 % CaCl2 prolonged the grain filling period in wheat by way of early ear head emergence and consequently increased the grain yield (Avijit Sen and Misra 1984).

In another study pre soaking of bajra seeds in 4 % manganese solution recorded highest grain yield compare to pre soaking of potassium dihydrogen phosphate, diammonium phosphate, potassium chloride, ferrous sulphate and zinc sulphate (Kannadasan et al., 1985).

Masood Ali (1985) indicated that foliar spray of 2 % KCl solution significantly increased grain yield over water spray in chickpea. Similarly Goswami and Garg (1985) observed that application of ascorbic acid increased fodder yield by 42 to 50 per cent in Lucerne. Foliar spray of potassium, K2SO4 (1 %) and KCl (1 %) increase the seed yield by 21.2 and 33.4 per cent respectively over control in blackgram (Chandrababu et al. 1985).

Avijit Sen and Misra (1987) reported that treating wheat seeds 0.25 % CaCl2 or 2.5 % KCl increased the grain yield compared to control. Similarly Patil (1987) opined that seed treatment with 2 % CaCl2 for four hours increased drought resistance in sorghum and also increased grain yield by 10 % over control under dry land conditions.

Arjunan and Srinivasan (1989) noticed that the seed hardening of ground nut with CaCl2 (1 %) significantly increased the mean pod yield by 20 % over untreated control. Foliar

spray of green gram with K2SO4 (1%) and KCl (1%) significantly increased the seed yield by 10.1 and 7.7 per cent respectively over control (Sadasivam et al. 1990).

The application of 100 ppm cycocel in okra has increased the yield (17.69 t/ha) as compared to control (8.45t/ha) in dry land situation (Arora et al., 1990). In the same way Patel and Singh (1991) also reported the increase in pod weight of okra with the application of 200 ppm of cycocel. Sarkar and Mukhopadhyay (1990) reported that foliar spray of 0.5 per cent KNO3 solution at 50 per cent flowering stage significantly increased the grain yield of high yielding and traditional cultivars by 49.1 and 19.3 per cent respectively over control in rice. Khalate et al. (1990) reported an increase in the yield of onion due to the spray of ZnSO4 (0.5 and 1%), CuSO4 (1%) and MnSO4 (0.6%).

Singh et al., (1991) found that foliar spray at pre flowering stage with cycocel (15 mg/l), mixtalol (6 mg/l) and triacontanol (4 mg/l) effectively enhanced the seed yield and protein content by their ameliorative effects on flower retention, pod formation, seed setting and seed weight in chickpea. Nawalagatti et al. (1991) reported that foliar spray of CCC at 1000 ppm increased total dry matter production and pod yield followed by cytozyme and CCC at 1000 and 500 ppm respectively in groundnut.

Shanmugasundaram and Nanjan (1992) indicated that soil application of magnesium (60 kg/ha) and foliar spray of magnesium sulphate (1.0%) recorded maximum potato tuber yield compared to control.

Upadhyay, (1994) recorded foliar application both NAA, kinetin (10, 20,30 ppm) and KNO3 (100,200,300ppm) at both bud initiation and pod formation stage significantly increased the yield of chickpea. Similarly Amaregouda et al. (1994) observed seed treatment with CaCl2 (2%) had given higher yield by 19% in wheat as compared to control. Seed treatment with cycocel (10ppm) increased graine yield of bajra by 2.0tonnes/ha as compared to control (Bishnoi et al., 1995).

Lourduraj et al. (1996) reported that among the different seed hardening chemicals, CaCl2 recorded higher dry pod yield, shelling per cent, 100 kernal weight and finally yield in groundnut. In another study by Jayarami Reddy et al. (1996) reported that foliar spry of NAA (20ppm) + KNO3 (0.5%) recorded the maximum seed yield (8.57q/ha) in pigeon pea over control (7.35q/ha). Govindan and Thirumurugan (2000) revealed that foliar spray of KNO3 (1%) + KCl (1%) increased the grain yield in greengram by 21.8 per cent over control. Rao et al., (2000) conducted the experiment on sorghum during 1996 and 1997 and observed that, the application KCl (1%) at terminal drought stress increased the grain yield of 32 and 27 per cent respectively over control.

Karaani et al. (2001) reported that the seed treatment of ragi cultivars viz., CO-13,PR 202and Indaf 9 with combination of 1 per cent KCl and CaCl2 had registered maximum yield of 19.6,17.5and13.9 per cent respectively over control. Similarly pre-sowing seed hardening of wheat with CaCl2 (2.5%) produced significantly higher grain yield (4014 kg/ha) over control (Ugale and Mungse, 2001).The higher fodder productivity was observed in seed hardening with K2HPO4 (2%) followed by KCl (2%) and KH2PO4 (2%) in forage grasses (Swaminathan and Sujata, 2001).

Garai and Datta (2003) found that nodules per plant and yield increased due to foliar application of cycocel in greengram. Combined foliar application of planofix @ 30 ppm on 30 and 45 DAS and chamatkar @ 120 ppm at 60 DAS recorded increased number of pods per plant, number of seeds per plant, 100 seed weight and yield in blackgram (Prakash et al., 1997).

Ramesh (2004) studied that seed hardening with CaCl2 (2%) increased number of pods per plant, pod yield per plant, seed yield per plant 100 seed weight and HI in chickpea. Foliar application ZnSO4 (0.5%) increased seed yield and its attributes significantly over the other treatments in pigeonpea (Varma et al., 2004).

3. MATERIAL AND METHODS

A field experiment was conducted during rabi 2005-06 to study the effect of seed hardening treatments with water, CaCl2, KH2PO4, KNO3, KCl, Sodium molybdate, Zinc sulphate, Cycocel, Succinic acid and Ascorbic acid on morpho-physiological, biochemical, yield and yield components in chickpea (Cicer arietinum (L.)) at College of Agriculture Farm, University of Agricultural Sciences, Dharwad. The details of materials used and techniques adopted during the course of investigation are described in this chapter.

3.1 Experimental site

The experiment was carried out on medium black clay soil (Plot no. 126A of ‘E’ block) at Main Agricultural Research Station, University of Agricultural Sciences, Dharwad, which is situated at 15

0 12' N latitude and 75

0 07' E longitude with an altitude of 678 m above the

mean sea level.

3.2 Climatic conditions

The meteorological data for 2005-06 and the mean of previous 55 years were collected from the Meteorological Observatory, College of Agriculture, University of Agricultural Sciences, Dharwad and presented in Table 1.

3.3 Soil and its characteristics

The experimental site consisted of medium black clay loam soil. The composite soil samples were analysed for their physical and chemical properties as per the procedure of Piper (1966) and Jackson (1967), and the details are furnished in Table 2.

3.4 Experimental details

3.4.1 Treatment details

The experiment consisted of eleven seed hardening treatments viz., water, CaCl2 (1% and 2%), KH2PO4, KNO3, KCl, Sodium molybdate, Zinc sulphate, Cycocel, Succinic acid and Ascorbic acid with a control.

Genotype : ICCV-2

Treatments : Twelve

Replications: Three

T1 : Control

T2 : Water soaking

T3 : Seed hardening with CaCl2 (1%)


T5 : Seed hardening with KH2PO4 (1%)

T6 : Seed hardening with KNO3 (100 ppm)

T7 : Seed hardening with KCl (100 ppm)

T8 : Seed hardening with sodium molybdate (100 ppm)

T9 : Seed hardening with zinc sulphate (100 ppm)

T10 : Seed hardening with cycocel (1000 ppm)

T11 : Seed hardening with succinic acid (20 ppm)

T12 : Seed hardening with ascorbic acid (20 ppm)

Table 1. Monthly meteorological data during crop growth period (2005-06 and 2006-07) and the average of 55 years (1950-2004) at Main Agricultural

Research Station, University of Agricultural Sciences, Dharwad

2005-06 2006-07 1950-2004

Temperature (0C) Temperature (

0C)

Temperature (0C)

Months

Max. Min.

Mean relative humidity

(%)

Rainfall (mm)

Max. Min.


(%)

Rainfall (mm)

Max. Min.


(%)

Rainfall (mm)

June 30.9 21.5 76 151.0 21.5 20.6 78 212.4 22.1 21.0 87.5 145.8

July 27.4 21.0 83 290.2 26.6 20.4 87 176.1 22.0 20.6 86.5 95.3

August 27.1 20.4 81 138.8 26.3 19.6 86 115.2 28.8 20.2 82.4 100.5

September 27.5 20.3 85 194.5 29.2 19.2 77 91.4 30.1 19.3 76.4 131.0

October 29.6 19.1 70 89.4 30.0 19.1 67 38.6 29.5 15.5 68.1 32.0

November 29.4 14.9 51 38.0 29.2 18.0 70 55.4 29.2 13.4 63.8 54.5

December 28.9 13.1 53 0.0 29.1 12.8 61 0 29.2 19.2 63.3 0.09

January 29.9 12.9 52 0.0 30.4 14.0 72 0 34.2 16.0 51.2 1.2

February 32.4 14.8 39 0.0 31.9 15.7 67 0 35.7 18.8 56.5 0.1

March 34.1 18.1 45 5.2 35.3 19.7 49 12.8 37.0 21.3 77.0 48.5

April 37.1 20.3 49 1.5 36.7 21.4 55 86.4 36.5 21.5 66.7 81.4

May 35.3 20.9 61 166.8 - - - - 29.5 21.2 81.7 109.1

Total 1075.4 788.3 799.5

Table 2. Physical and chemical properties of soil from the experimental site

Sl. No.

Properties Value

obtained Method employed

I. Physical properties

1. Coarse sand (%) 6.28 International pipette method (Piper, 1966)

2. Fine sand (%) 14.27 International pipette method (Piper, 1966)

3. Silt (%) 27.52 International pipette method (Piper, 1966)

4. Clay (%) 51.99 International pipette method (Piper, 1966)

5. Bulk density (g/cc) 1.33 Core sample method (Dastane, 1967)

II. Chemical properties

1. Soil pH (1:2.5 soil:water) 7.60 pH meter (Piper, 1966)

2. Electrical conductivity (dS/m) 0.28 Conductivity bridge (Jackson, 1967)

3. Organic carbon (%) 0.52 Walkley and Black wet oxidation method (Jackson, 1967)

4. Available nitrogen (kg/ha) 221.0 Modified Kjeldahl method (Jackson, 1967)

5. Available phosphorus (kg/ha) 32.4 Olsen’s method (Jackson, 1967)

6. Available potassium (kg/ha) 318.7 Flame photometer (Jackson, 1967)

LEGEND

T1 : Control

T2 : Water soaking



T5 : Seed hardening with KH2PO4 (1%)

T6 : Seed hardening with KNO3 (100 ppm)

T7 : Seed hardening with KCl (100 ppm)

T8 : Seed hardening with sodium molybdate (100 ppm)

T9 : Seed hardening with zinc sulphate (100 ppm)

T10 : Seed hardening with cycocel (1000 ppm)

T11 : Seed hardening with succinic acid (20 ppm)

T12 : Seed hardening with ascorbic acid (20 ppm)

N

R-I R-II R-III

0.5m

2.4m

Figure 1. Plan of layout of the experiment

T1

T8

T9

T7

T2

T12

T4

T11

T3

T6

T5

T10

T7

T3

T1

T12

T4

T9

T5

T10

T8

T11

T6

T2

T12

T6

T9

T2

T7

T11

T8

T3

T4

T1

T10

T5

2.7m

3.4.2 Design and layout

The experiment was laid out in randomized block design with three replications. The plan of layout of experiment is given in Figure.1.

Plot size: Gross plot size : 2.7 m x 2.4 m

Net plot size : 1.5 m x 2.0 m

3.5 Cultural operations

3. 5.1 Land preparation

After the harvest of the previous crop, the land was ploughed with a disc plough, subsequently the land was harrowed twice, followed by clod crushing and repeated harrowings. The land was smoothened with plank to bring the soil to a fine tilth suitable for sowing. Small bunds were erected around each plot to avoid movement of fertilizer and surface flow of rain water from one plot to another. The plots were laid out according to the plan (Figure.1).

3. 5.2 Seed treatment and sowing

A day before sowing chickpea seeds were soaked separately in water, CaCl2 (1% and 2%), KH2PO4, KNO3, KCl, sodium molybdate, zinc sulphate, cycocel, succinic acid and ascorbic acid for three hours and seeds were dried under shade and used for sowing.

3. 5. 3 Plant protection chemicals

To control the pest and diseases, necessary plant protection measures were taken up as per the recommended package of practice for field crops (Anon., 1978).

3. 5. 4 Fertilizer application

The crop was fertilized with nitrogen and phosphorus at the rate of 25:50 kg ha-1

in the form of urea and super phosphate, respectively. At sowing, entire quantity of nitrogen and phosphorus was applied as a basal dose and mixed thoroughly into the soil.

3.5.5 Spacing

Inter-row spacing : 30 cm

Intra-row spacing : 10 cm

3.5.6 Seed source and sowing

Seeds were obtained from Main Research Station, Dharwad. Healthy and bold seeds were dibbled with a spacing of 30 cm x 10 cm to a depth of 5 cm. The crop was given protective irrigations as and when required.

3.5.7 Thinning operation

The seedlings were thinned out by maintaining one plant per hill after 15 days after sowing.

3.5.8 Intercultural operations

Intercultural operation was carried out at three weeks after sowing immediately after hand weeding.

3.5.9 Harvesting

The crop was harvested when plants were at 80% pod maturity, leaf lets started shedding and pods turning to pale yellow colour. The border row plants were first uprooted manually from all sides of each plot and then the net plots were harvested excluding five randomly selected and tagged plants. The harvested plants were dried in the shade for seven days. The seeds were separated manually by gently beating the dried plants with the wooden

stick. The seeds were cleaned dried in the shade and were collected for each treatment. Thereafter seed yield g.plant

-1 as well as q ha

-1 was computed.

3.6 Collection of experimental data

Five plants from each plot were selected randomly and tagged for the purpose of recording morphological, physiological, biochemical and yield parameters at different stages of growth period.

3.6.1 Morphological characters

3.6.1.1Plant height (cm)

Plant height was recorded from the ground level to the growing tip of the main shoot. Measurements were taken from five tagged plants in each treatment and the average height was calculated and expressed in centimeters.

3.6.1.2 Number of leaflets

Number of leaflets from five tagged plants was counted and the average was worked out.

3.6.1.3Total dry matter production and its partioning

Randomly selected five plant samples were separated into leaf, stem and reproductive parts and dried in oven at 80

oC until constant weight was obtained. Total dry

matter was calculated by adding dry weight of different plant parts and expressed as grams per plant at different intervals of crop growth period.

3.6.2 Growth analysis

3.6.2.1Leaf area (dm2 plant-1)

The leaves from plants selected for growth analysis from each treatment were used for the estimation of leaf area. Leaf area was computed by graphic method and expressed as

dm2 plant-1.

3.6.2.2Leaf area index (LAI)

The leaf area index was calculated by dividing the leaf area per plant by land area occupied by the plant (Sestak et al., 1971),

Leaf area / plant (cm2)

LAI = --------------------------------- Land area / plant (cm

2)

3.6.2.3Crop Growth Rate (CGR)

Crop growth rate is the rate of dry matter production per unit ground area per unit time (Watson, 1952). It was calculated by using the following formula and expressed as g m

-

2 day

-1.

(W2 – W1) 1 CGR = ------------------- x ----- (t2 – t1) A

where, W1 = Dry weight of the plant (gm-2

) at time t1

W2 = Dry weight of the plant (gm-2

) at time t2

t1-t2 = Time interval in days

A = Unit land area (m2)

3.6.2.4Absolute Growth Rate (AGR)

It expresses the dry weight increase per unit time and was calculated by using the following formula,

W2 – W1

AGR = ---------------- g day-1

t2 – t1

where, W2 and W1 are the total dry weights per plant at time t2 and t1 respectively.

3.6.2.5Relative Growth Rate (RGR)

It is rate of increase in dry weight per unit dry weight already present and is expressed in g g

-1 day

-1. Relative growth rate at various stages was calculated as suggested

by Radford (1967).

Loge W2 – Loge W1 RGR = ------------------------

t2 – t1

where, W1 = Dry weight of plants (g) at time t1

W2 = Dry weight of plants (g) at time t2

3.6.2.6Net Assimilation Rate (g dm-2 day-1)

Net assimilation rate (NAR) is the rate of dry weight increased per unit leaf area per unit time. It was calculated by using the following formula (Watson, 1952)

(W2 – W1) Loge W2 – LogeW1 NAR = --------------- x ------------------------ g dm

-2 day

-1

(t2 – t1) (L2 – L1)

where,

L1 & W1 = Leaf area (dm2) and dry weight of the plant (g), respectively at time t1.

L2 & W2 = Leaf area (dm2) and dry weight of the plant (g), respectively at time t2.

3.6.2.7 Leaf Area Duration (LAD)

Leaf area duration is the integral of leaf area index over a growth period (Watson, 1952). LAD for different growth periods was worked out as per the formula of Power et al.

(1967) and expressed in days.

Li + (L (i+1)) LAD = --------------- x (t2 – t1) 2

where,

Li = LAI at ith stage

L(i+1) = LAI at (i+1)th stage

t 2 – t1 = Time interval between ith stage and (i+1)

th stage

(days)

3.6.2.8Leaf Area Ratio (LAR)

Leaf area ratio was worked out by Radford (1967) expressed in cm2 g

-1.

Leaf area (cm2 plant

-1)

LAR = ----------------------------------- Total dry matter (g plant

-1)

3.6.2.9Biomass Duration (BMD)

The BMD was calculated by using the following formula and expressed in gram days.

TDMi + TDM (i+1) BMD = -------------------------- x (t2 – t1 2

where,

TDMi = TDM at ith stage

TDM (i+1) = TDM at (a+1)th stage

(t2 – t1) = Time interval between ith stage and (i+1)

th stage (days)

3.6.2.10 Specific Leaf Weight (SLW)

The specific leaf weight indicates the leaf thickness and was determined by the method of Radford (1967). It was expressed as mg cm

-2.

Leaf dry weight (mg) SLW = ------------------------------- Leaf area (cm

2)

3.6.2.11 Specific leaf area (SLA)

The inverse of the Specific Leaf Weight is the Specific leaf area and was calculated

as follows:

Leaf area (cm2)

SLA = ------------------------------ Leaf dry weight (mg)

3.6.3 Yield and yield parameters

Tagged plants used for recording morphological observations were harvested at physiological maturity and were used for recording the following yield and yield components.

3.6.3.1 Seed yield per plant (grams)

The five tagged plants were uprooted at harvest maturity and processed for seed yield, from which the average was calculated and expressed as gram seed yield per plant.

3.6.3.2 Seed yield per hectare (quintal)

On the basis of seed yield per plant, the seed yield per ha was computed and expressed as quintal seed yield per hectare.

3.6.3.3 100 seed weight (grams)

Hundred seeds were counted from the sample drawn from seed yield of each net plot and the weight of 100 seeds was recorded and expressed in grams

3.6.3.4 Harvest index (HI)

It was calculated by using the following formula.

Seed yield per plant (g) HI = -------------------------------------- x 100 Total dry weight per plant (g)


3.7.1 Estimation of total chlorophyll content

Chlorophyll pigments viz., chlorophyll ‘a’, chlorophyll ‘b’ and total chlorophyll in leaf were determined by dimethyl sulfoxide method (DMSO) of Hiscox and Israelstam (1979). Top fully expanded leaf was brought from the field in polythene bags, kept in ice box and it was cut into small pieces. Known weight of leaves (100 mg) was kept in test tubes containing 7 ml of dimethyl sulfoxide (DMSO). The test tubes were incubated at 65

0C for 30 minutes. Leaf

residue was removed by decanting the solution and final volume was made to 10 ml with DMSO. The absorbance of the extract was measured at 645 and 663 nm in a UV-Vis spectrophotometer (Elico, CL-54) and a blank was run using DMSO.

The total chlorophyll content was calculated by using the following formula and expressed in mg g fresh weight

-1;

V Chlorophyll ‘a’= 12.7 x (A663) - 2.69 x (A645) x ---------------

(mg g-1

fr.wt.) 1000 x w x a V

Chlorophyll ‘b’= 22.9 x (A645) – 4.68 x (A663) x --------------- (mg g

-1 fr.wt.) 1000 x w x a

V

Total chlorophyll = 20.2 x (A645) – 8.02 x (A663) x --------------- (mg g

-1 fr.wt.) 1000 x w x a

where, A645 = Absorbance of the extract at 645 nm

A663 = Absorbance of the extract at 663 nm

a = Path length of cuvette (1 cm)

V = final volume of the chlorophyll extract (10 ml)

W = Fresh weight of the sample (0.10 g)

3.7.2 Estimation of free proline

Free proline content was estimated by following the method of Bates et al. (1973). A known weight (0.5 g) of fresh leaf sample was macerated in a mortar using 10 ml of 3 per cent sulphosalicylic acid. The extract was filtered and 2.0 ml of the filtrate was used for proline estimation. To this 2.0 ml of filtrate, 2.0 ml of acid ninhydrin reagent (2.5 g of ninhydrin dissolved in 40 ml of 6.0 M orthophosphoric acid and 60 ml of glacial acetic acid), 2.0 ml of glacial acetic acid were added and placed in boiling water bath for one hour. Following this, test tubes containing the samples were transferred to an ice bath for cooling. The contents of each test tube were transferred to a separatory funnel and 6.0 ml of toulene was added, shaken thoroughly and allowed for few minutes for separation of two layers. The lower layer was discarded and the upper toulene layer containing the colour complex was taken into a test tube. The optical density was read at 520 nm using spectrophotometer (Elico, UV-vis spectrophotometer) and the proline content was calculated as follows.

36.2311 x OD x V x d Proline content (µg / g dry wt) =------------------------------

2 x f

where, OD = Optical density at 520 nm

V = Volume of aliquot made (ml)

D = Fresh weight / dry weight ratio

f = Fresh weight taken for proline estimation (mg)

2 = volume of the aliquot

3.7.3 Relative water content

The relative content was estimated by the method of Barrs was Weatherly (1962). Ten leaf discs were collected randomly in each treatment and weighed accurately upto third decimal on a single pan analytical balance. This was considered as fresh weight. The weighed leaf discs were allowed to float on distilled water in a petridish and allowed to absorb water for four hours. After four hours, the leaf discs were taken out and their surface was blotted gently and weighed. This was referred to as turgid weight. After drying in hot air oven at 72

0 C for 48 hours, the dry weight was recorded and RWC was calculated by using the

following formula,

Fresh weight – Dry weight RWC (%) = ----------------------------------- x 100

Turgid weight – Dry weight

3.7.4 Chlorophyll stability index (CSI)

Green plants pigments are thermo-sensitive and degradation occurs when they are subjected to higher temperature. This method is based on pigment changes induced by heating. Chlorophyll stability is the function of temperature and this property of chlorophyll stability was found to have good correlation with drought resistance.

Representative leaf sample was placed in two clean tubes with 50 ml of distilled water. One tube was then subjected to heat on water bath at 56

oC ± 1

oC for exactly 30

minutes. The chlorophyll in both the samples was extracted by placing the sample in 7 ml of DMSO at 65

oC for 30 minutes. The supernatant was decanted and the tissue was discarded,

then volume was made to 10 ml by DMSO. Finally, the absorbance of the extract was read at 645, 652 and 663 nm using DMSO as blank (Hiscox and Isrealstam , 1979).

V Total chlorophyll = 20.2 x (A645) – 8.02 x (A663) x -----------------

(mg g-1

fr.wt.) 1000 x w x a

where, A645 = Absorbance of the extract at 645 nm

A663 = Absorbance of the extract at 663 nm


V = final volume of the chlorophyll extract (10 ml)

W = Fresh weight of the sample (0.10 g)

Cs CSI (%) = ---------x 100

Cc

where,

CSI = chlorophyll stability index

Cs = chlorophyll content of stressed plant

Cc = chlorophyll content of control plant


3.7.5 Estimation of epicuticular wax content

Epicuticular wax content of leaf sample was determined by the rapid colorimetric method of Ebercon et al., (1977). The individual sample consisting of 10 leaf discs of known area (both surfaces) was filtered and evaporated to dryness on boiling water bath until the

smell of chloroform goes. Five ml of acidic K2Cr2O7 was added to samples and placed in boiling water for 30 minutes, after cooling, 12 ml of deionised water was added. The contents were allowed to stand for 15-20 minutes for colour development, and the optical density of the sample was read at 590 nm.

Wax was quantified by using the standard curve prepared by using carbowax 3000 (polyethylene glycol 3000) and expressed in mg per dm

2 area (both surfaces – since wax is

present on both the surfaces)

3.8 Statistical analysis

The data were subjected to the analysis of variance by following the method of Panse and Sukhatme (1967). The level of significance used in 'F' and‘t’ tests was P = 0.05. Critical differences were calculated wherever 'F' test was found significant.

4. EXPERIMENTAL RESULTS

A field experiment was conducted during rabi 2006 at Main Agricultural Research Station, University of Agricultural Sciences, Dharwad, to study the influence of seed hardening techniques on various morpho-physiological, growth, biochemical and yield parameters in chickpea (var, ICCV-2). The results obtained from the investigation are presented in this chapter.

4.1 Morphological characters

4.1.1 Plant height (cm)

The data on plant height as influenced by seed hardening chemicals showed significant differences among the treatments at all the stages except at 40 DAS. In general, plant height increased from 40 DAS to harvest as indicated in Table 3. Among the treatments, seed hardening with 2% CaCl2 recorded significantly higher values over all other treatments. However, it was on par with the treatments KH2PO4 (1%), KNO3 (100 ppm), KCl2 (100 ppm), zinc sulphate (100 ppm) and ascorbic acid (20 ppm) at 60 DAS. The plant height in the treatments viz, KNO3 (100 ppm), sodium molybdate (100 ppm) and zinc sulphate (100 ppm)were on par with each other at 80 DAS. While, KNO3 (100 ppm) and ascorbic acid (20 ppm) at harvest were on par with each other. Significantly lower plant height was observed in the treatment seed hardening with cycocel (1000 ppm). However, it did not differ significantly on seed hardening treatments with succinic acid (20 ppm) and control. Maximum plant height (48.05 cm) was recorded in seed hardening with 2% CaCl2 at harvest.

4.1.2 Number of leaflets

The data pertaining to number of leaflets presented in Table 4. indicated significant difference among the treatments at all the stages. In general the number of leaflets was increased from 40 to 80 DAS. Among the treatments, seed hardening with 2% CaCl2 recorded significantly more number of leaflets followed by seed hardening with KNO3 (100 ppm) and cycocel (1000 ppm). Significantly less number of leaflets was recorded in control at all the stages. However, seed hardening with water soaking and CaCl2 (1%) were on par with each other at 40 and 60DAS. While, water soaking seed hardening with CaCl2 (1%), KCl2 (100 ppm) did not differ significantly with each other at 80 DAS. Maximum number of leaflets (298) was recorded in seed hardening with 2% CaCl2 at harvest.

4.1.4 Leaf dry weight (g plant-1)

The Leaf Dry weight (LDW) as influenced by the seed hardening chemicals presented in Table 5. indicated significant difference among the treatments at all the stages. In general, the leaf dry weight increased from 40 to 60 DAS and decreased thereafter. Among the treatments, seed hardening with 2% CaCl2 recorded significantly higher leaf dry mater followed by seed hardening with cycocel (1000 ppm) and succinic acid (20 ppm) at 40 DAS. Significantly lower leaf dry matter was recorded in control at all the stages. However at 60 DAS seed hardening with CaCl2 (2%), KH2PO4 (1%), KNO3 (100 ppm) and cycocel (1000 ppm) were on par with each other. While, seed hardening with CaCl2 (2%), KNO3 (100 ppm), cycocel (1000 ppm) and succinic acid (20 ppm) at 80DAS did not differ significantly. At harvest KNO3(100 ppm), cycocel (1000 ppm), succinic acid (20 ppm) and zinc sulphate (100 ppm) were on par with each other. Maximum leaf dry matter (2.16 g plant

-1) was recorded in

seed hardening with 2% CaCl2 at harvest (Figure 2).

4.1.5 Stem dry weight (g plant-1)

The stem dry weight increased from 40 to 80 DAS and decreased thereafter as shown in Table 6. treatments differed significantly at all the stages. Among the treatments, seed hardening with 2% CaCl2 recorded significantly higher stem dry weight followed by seed hardening with cycocel (1000 ppm) and ) succinic acid (20 ppm). However, it was on par with sodium molybdate (100 ppm), zinc sulphate (100 ppm) and cycocel (1000 ppm) at 60 DAS. While, at 80 DAS KNO3 (100 ppm) and cycocel (1000 ppm) were not differ significantly with each other. Significantly lower stem dry weight was recorded significantly in control. But it did

Table 3. Influence of seed hardening chemicals on plant height (cm) at different stages in chickpea

Plant height Treatments 40DAS 60DAS 80DAS At harvest

T1 : Control 25.4 31.0 36.7 40.6

T2 : Water soaking 26.1 35.0 41.3 43.5

T3 : CaCl2 (1%) 24.4 34.8 41.0 43.7

T4 : CaCl2 (2%) 27.4 38.9 44.1 48.1

T5 : KH2PO4 (1%) 24.7 35.6 41.8 44.7

T6 : KNO3 (100 ppm) 25.4 36.8 43.6 46.0

T7 : KCl (100 ppm) 25.7 35.8 41.1 45.0

T8 : Sodium molybdate (100 ppm) 24.9 34.5 43.2 43.8

T9 : Zinc sulphate (100 ppm) 25.1 35.3 42.1 44.7

T10 : Cycocel (1000 ppm) 24.1 28.8 35.6 39.6

T11 : Succinic acid (20 ppm) 25.0 29.3 36.7 40.3

T12 :Ascorbic acid (20 ppm) 25.6 36.2 41.9 45.6

Mean 25.3 34.3 40.7 43.8

S.Em ± 1.17 1.28 0.73 0.91

CD (5%) NS 3.67 2.11 2.63

NS : Non-signfiicant DAS : Days after sowing

not differ significantly with CaCl2 (1%) at 80 DAS and water soaking and seed hardening with KCl (100 ppm) at harvest. Maximum stem dry weight was recorded in seed hardening with 2% CaCl2 (8.14 g plant

-1) at harvest (Figure 3).

4.1.6 Dry weight of reproductive parts (g plant-1)

Significant differences in the Table 7. were due to various treatments with respect to dry weight of reproductive parts at all the stages except 40 DAS. The dry weight of reproductive parts was increased from 40DAS to harvest in all the treatments. Among the treatments seed hardening with 2% CaCl2 as recorded significantly higher reproductive dry weight followed by seed hardening with cycocel (1000 ppm) and succinic acid (20 ppm) as compared to control at 60 and 80 DAS. Significantly lower dry weight of reproductive parts was recorded in control. But it did not differ significantly with water soaking, and CaCl2 (1%) at 80DAS. Maximum reproductive parts dry weight was recorded in seed hardening with 2% CaCl2 (25.79 g plant

-1) at harvest (Figure 4).

4.1.7 Total dry matter (g plant-1)

The results pertaining to total dry matter (TDM) in Table 8. indicated significant difference among the treatments at all the stages. The total dry matter increased from 40 DAS to harvest in all the treatments. Among the treatments, pre-sowing seed hardening with 2% CaCl2 has recorded significant increase in total dry matter content followed by seed hardening

Table 4. Influence of seed hardening chemicals on number of leaflets at different stages in chickpea

Number of leaflets Treatments 40DAS 60DAS 80DAS

T1 : Control 81.3 161.3 201.7

T2 : Water soaking 94.0 175.7 216.0

T3 : CaCl2 (1%) 87.7 168.3 210.3

T4 : CaCl2 (2%) 170.7 265.7 298.0

T5 : KH2PO4 (1%) 115.0 209.3 247.7

T6 : KNO3 (100 ppm) 167.0 255.0 297.0

T7 : KCl (100 ppm) 98.3 187.0 222.0

T8 : Sodium molybdate (100 ppm) 102.0 191.0 231.7

T9 : Zinc sulphate (100 ppm) 124.7 216.3 256.0

T10 : Cycocel (1000 ppm) 159.0 247.3 285.0

T11 : Succinic acid (20 ppm) 139.3 225.7 267.0

T12 :Ascorbic acid (20 ppm) 132.7 221.7 259.3

Mean 122.6 210.4 249.3

S.Em ± 4.4 7.7 8.9

CD (5%) 12.6 22.2 25.6

DAS : Days after sowing

with cycocel (1000 ppm), and KNO3 (100 ppm) as compared to control. Significantly lower TDM was observed in control at all the stages. But, it did not differ significantly CaCl2 (1%) at 80 DAS. Maximum total dry weight (36.1 g plant

-1) was recorded in pre sowing seed

hardening with 2% CaCl2 at harvest (Figure 5).

4.2 Growth Parameters

4.2.1 Leaf area (cm2 plant-1)

The leaf area per pant indicated significant difference among the treatments at all the stages except 40 DAS. Leaf area increased from 40 DAS to 60 DAS and thereafter decreased, Table 9. Among the treatments, seed hardening with 2% CaCl2 showed significantly higher values followed by KNO3 (100 ppm) compare to other treatments at all the stages. However, significantly lower leaf area was observed in control, but it did not differ significantly with cycocel (1000 ppm), succinic acid (20 ppm) and Water soaking at 60 DAS. Similarly, it was on par with Water soaking and seed hardening with CaCl2 (1%), sodium molybdate (100 ppm), cycocel (1000 ppm) and succinic acid (20 ppm) at 80 DAS. Maximum leaf area (492 cm

2 plant

-1) was recorded in seed hardening with 2% CaCl2 at harvest.

Table 5.Influence of seed hardening chemicals on leaf dry weight (g plant-1


Leaf dry weight

Treatments 40DAS 60DAS 80DAS At

harvest

T1 : Control 0.96 2.83 2.05 1.48

T2 : Water soaking 1.33 3.39 2.26 1.85

T3 : CaCl2 (1%) 1.05 3.46 2.00 1.35

T4 : CaCl2 (2%) 1.53 4.88 3.70 2.16

T5 : KH2PO4 (1%) 1.11 4.37 2.74 1.96

T6 : KNO3 (100 ppm) 1.17 4.67 3.25 2.09

T7 : KCl (100 ppm) 1.12 4.35 2.44 1.48



T10 : Cycocel (1000 ppm) 1.52 4.48 3.51 2.08



Mean 1.21 4.09 2.73 1.86

S.Em ± 0.07 0.21 0.17 0.06

CD(5%) 0.19 0.60 0.49 0.17


Fig.2: Influence of seed hardening chemicals on leaf dry weight (g plant-1


Table 6.Influence of seed hardening chemicals on stem dry weight (g plant-1


Stem dry weight Treatments

40DAS 60DAS 80DAS At harvest

T1 : Control 0.64 2.73 4.55 4.05

T2 : Water soaking 0.95 3.58 5.88 4.91

T3 : CaCl2 (1%) 0.70 3.49 4.67 5.11

T4 : CaCl2 (2%) 1.29 4.35 8.28 8.14

T5 : KH2PO4 (1%) 0.81 3.83 5.70 5.68

T6 : KNO3 (100 ppm) 0.71 3.66 7.70 6.16

T7 : KCl (100 ppm) 0.66 3.39 6.05 4.26



T10 : Cycocel (1000 ppm) 1.09 4.27 7.61 7.43



Mean 0.84 3.72 6.40 5.73

S.Em ± 0.05 0.23 0.29 0.33

CD(5%) 0.15 0.67 0.84 0.95


Fig.3: Influence of seed hardening chemicals on stem dry weight (g plant-1


Table 7.Influence of seed hardening chemicals on reproductive dry weight (g plant-1


Reproductive dry weight Treatments

40DAS 60DAS 80DAS At harvest

T1 : Control 0.11 0.55 7.61 16.70

T2 : Water soaking 0.12 1.39 8.84 19.73

T3 : CaCl2 (1%) 0.12 1.15 8.99 18.10

T4 : CaCl2 (2%) 0.14 2.25 13.78 25.79

T5 : KH2PO4 (1%) 0.11 1.41 9.08 22.98

T6 : KNO3 (100 ppm) 0.13 1.44 10.73 23.32

T7 : KCl (100 ppm) 0.12 1.34 9.35 22.06



T10 : Cycocel (1000 ppm) 0.14 2.23 13.38 23.21



Mean 0.12 1.50 10.39 21.06

S.Em ± 0.009 0.21 0.59 0.99

CD(5%) NS 0.60 1.70 2.86


Fig.4: Influence of seed hardening chemicals on reproductive dry weight (g plant-1

) at stages in chickpea

4.2.2 Leaf area index

The data on leaf area index in Table 10. increased from 40 to 60 DAS and decreased there after at 80 DAS. All the treatments differ significantly at all the stages and among the treatments seed hardening with 2% CaCl2 recorded significantly higher values followed by KNO3 (100 ppm) compare to other treatments at all the stages, however significantly lower leaf area index (LAI) was observed in control, but it did not differ significantly with seed hardening with Water soaking, cycocel (1000 ppm) and succinic acid (20 ppm) at 60 and 80 DAS. Maximum leaf area index (1.64) was recorded in seed hardening with 2% CaCl2 at harvest.

4.2.3 Specific leaf weight (mg cm -2)

The results pertaining to SLW in Table 11. indicated significant difference among the treatments at all the stages. SLW increase from 40 to 60 DAS and decreased thereafter at 80 DAS in all the treatments. Among the treatments, the maximum SLW was obtained significantly in seed hardening with cycocel (1000 ppm) followed by seed hardening with succinic acid (20 ppm) and 2% CaCl2.. Significantly lower SLW was observed in control. But it did not differ significantly with water soaking and seed hardening with CaCl2 (1%), KCl (100 ppm) and sodium molybdate (100 ppm) at all the stages. Maximum SLW (8.80 mg cm

-2) was

recorded in seed hardening with cycocel (1000 ppm) at harvest.

4.2.4 Leaf area duration (days)

The data on leaf area duration (LAD) presented in the table 12 indicated significant differences due to treatments at all the stages. It increased from 40 -60 DAS to 60-80DAS. Among the treatments seed hardening with 2% CaCl2 indicated significantly increased LAD followed by KNO3 (100 ppm) and ascorbic acid (20 ppm) as compared to control at all the stages. However significantly lower LAD was observed in control at all the stages but it did not differ significantly with water soaking, CaCl2(1%), sodium molybdate (100 ppm), cycocel (1000 ppm) and succinic acid (20 ppm) at 60-80DAS. Maximum LAD (36 days) was noticed in seed hardening with 2% CaCl2 at 60-80 DAS.

4.2.5 Absolute growth rate (g plant-1days-1)

The data on absolute growth rate (AGR) indicated significant differences among the treatments at all the stages. It is evident from the Table 13. that AGR increased from 40-60 DAS to 60-80 DAS. Among the treatments seed hardening with 2% CaCl2 has recorded significantly higher values followed by cycocel (1000 ppm), KNO3 (100 ppm), sodium molybdate (100 ppm) and ascorbic acid (20 ppm), at 40-60 DAS as compared to control. However, significantly lower AGR values were observed in control at all the stages. But this treatment did not differ significantly with Water soaking, CaCl2 (1%), KH2PO4 (1%), KCl (100 ppm), sodium molybdate (100 ppm) and Zinc sulphate (100 ppm) at 60-80 DAS. Maximum AGR (0.714 g plant

-1days

-1) was recorded seed hardening with 2% CaCl2 at 60-80 DAS.

4.2.6 Crop growth rate (g dm-2days-1)

The results pertaining to crop growth rate (CGR) shown in Table 14. were indicated significant differences among the treatments at all the stages CGR increased continuously upto 80 DAS in all the treatments. Among the treatments, the maximum CGR was obtained significantly in seed hardening with 2% CaCl2 followed by cycocel (1000 ppm) and KNO3 (100 ppm) at 40-60 DAS as compare to control. However, control recorded significantly lower CGR values at all the stages but it was on par with water soaking, CaCl2(1%), KH2PO4(1%), KCl (100 ppm) and Sodium molybdate (100 ppm)at 60-80 DAS. Maximum CGR (23.8 g dm

-

2days

-1) was recorded seed hardening with 2% CaCl2 at 60-80 DAS.

4.2.7 Relative growth rate (g g-1day-1)

The results pertaining to RGR indicated significant differences due to various treatments at all stages it is evident from Table15. that RGR increased from 40-60 DAS and declined there after. However, at 40-60DAS significantly higher RGR values were recorded in ascorbic acid (20ppm) followed by sodium molybdate (100ppm), KH2PO4 (1%), KNO3

Table 8.Influence of seed hardening chemicals on total dry matter (g plant-1


TDM Treatments 40DAS 60DAS 80DAS At harvest

T1 : Control 1.7 6.1 14.2 22.2

T2 : Water soaking 2.4 8.4 17.0 26.5

T3 : CaCl2 (1%) 1.9 8.1 15.7 24.6

T4 : CaCl2 (2%) 3.0 11.5 25.8 36.1

T5 : KH2PO4 (1%) 2.0 9.6 17.5 30.6

T6 : KNO3 (100 ppm) 2.0 9.8 21.7 31.6

T7 : KCl (100 ppm) 1.9 9.1 17.8 27.8



T10 : Cycocel (1000 ppm) 2.8 11.0 24.5 32.7



Mean 2.2 9.3 19.5 28.7

S.Em ± 0.09 0.59 0.89 1.57

CD(5%) 0.28 1.72 2.56 4.53


Fig.5: Influence of seed hardening chemicals on total dry matter (g plant-1


Table 9.Influence of seed hardening chemicals on leaf area (cm-2


Leaf area

Treatments 40DAS 60DAS 80DAS

T1 : Control 189 456 393

T2 : Water soaking 213 504 408

T3 : CaCl2 (1%) 192 525 408

T4 : CaCl2 (2%) 216 588 492

T5 : KH2PO4 (1%) 195 546 438

T6 : KNO3 (100 ppm) 190 558 456

T7 : KCl (100 ppm) 191 528 432

T8 : Sodium molybdate (100 ppm) 194 534 405

T9 : Zinc sulphate (100 ppm) 193 537 438

T10 : Cycocel (1000 ppm) 195 459 399

T11 : Succinic acid (20 ppm) 191 498 399

T12 :Ascorbic acid (20 ppm) 190 552 444

Mean 196 524 426

S.Em ± 9.5 19.4 12.5

CD(5%) NS 55.6 36.2


Table 10.Influence of seed hardening chemicals on leaf area index at different stages in chickpea

LAI Treatments 40DAS 60DAS 80DAS

T1 : Control 0.63 1.52 1.31

T2 : Water soaking 0.71 1.68 1.36

T3 : CaCl2 (1%) 0.64 1.75 1.36

T4 : CaCl2 (2%) 0.72 1.96 1.64

T5 : KH2PO4 (1%) 0.65 1.82 1.46

T6 : KNO3 (100 ppm) 0.63 1.86 1.52

T7 : KCl (100 ppm) 0.64 1.76 1.44



T10 : Cycocel (1000 ppm) 0.65 1.53 1.33



Mean 0.65 1.75 1.42

S.Em ± 0.031 0.065 0.046

CD(5%) NS 0.187 0.132


Table 11. Influence of seed hardening chemicals on specific leaf weight(mg cm-2


SLW Treatments

40DAS 60DAS 80DAS

T1 : Control 5.08 6.21 5.09

T2 : Water soaking 6.24 6.73 5.54

T3 : CaCl2 (1%) 5.47 6.59 5.02

T4 : CaCl2 (2%) 7.08 8.30 7.52

T5 : KH2PO4 (1%) 5.69 8.00 6.26

T6 : KNO3 (100 ppm) 6.16 8.37 7.13

T7 : KCl (100 ppm) 5.86 8.24 5.65



T10 : Cycocel (1000 ppm) 7.79 9.76 8.80



Mean 6.16 7.81 6.40

S.Em ± 0.39 0.53 0.50

CD(5%) 1.11 1.53 1.44


Table 12.Influence of seed hardening chemicals on leaf area duration (days) at different stages in chickpea

Leaf area duration Treatments 40-60 60-80

T1 : Control 21.5 28.3

T2 : Water soaking 23.9 30.4

T3 : CaCl2 (1%) 23.9 31.1

T4 : CaCl2 (2%) 26.8 36.0

T5 : KH2PO4 (1%) 24.7 32.8

T6 : KNO3 (100 ppm) 24.9 33.8

T7 : KCl (100 ppm) 24.0 32.0

T8 : Sodium molybdate (100 ppm) 24.3 31.3

T9 : Zinc sulphate (100 ppm) 24.3 32.5

T10 : Cycocel (1000 ppm) 21.8 28.6

T11 : Succinic acid (20 ppm) 23.0 29.9

T12 :Ascorbic acid (20 ppm) 24.7 33.2

Mean 24.0 31.7

S.Em ± 0.83 1.19

CD(5%) 2.40 3.43


Table 13.Influence of seed hardening chemicals on absolute growth rate(g plant-1

day-1


AGR

Treatments 40-60 60-80

T1 : Control 0.220 0.403


T3 : CaCl2 (1%) 0.311 0.381

T4 : CaCl2 (2%) 0.426 0.714

T5 : KH2PO4 (1%) 0.379 0.396

T6 : KNO3 (100 ppm) 0.388 0.596

T7 : KCl (100 ppm) 0.359 0.438



T10 : Cycocel (1000 ppm) 0.412 0.676



Mean 0.357 0.510

S.Em ± 0.014 0.013

CD(5%) 0.042 0.039


Table 14. Influence of seed hardening chemicals on crop growth rate (g dm-2

days-1


CGR Treatments

40-60 60-80

T1 : Control 7.3 13.4


T3 : CaCl2 (1%) 10.4 12.7

T4 : CaCl2 (2%) 14.2 23.8

T5 : KH2PO4 (1%) 12.6 13.2

T6 : KNO3 (100 ppm) 13.0 19.9

T7 : KCl (100 ppm) 12.0 14.6



T10 : Cycocel (1000 ppm) 13.7 22.5



Mean 11.9 17.0

S.Em ± 0.48 0.53

CD(5%) 1.40 1.53

(100ppm), KCl (100ppm) and Zinc sulphate (100 ppm). At 60-80 DAS significantly higher RGR was recorded in succinic acid (20 ppm) followed by control and seed hardening with CaCl2 (1%). However, significantly lower RGR was observed in water soaking. Maximum RGR (0.0197 g g

-1day

-1) was noticed in seed hardening with succinic acid (20 ppm) at 60-80

DAS.

4.2.8 Nitrate assimilation rate (g g-1day-1)

The observations on net assimilation rate (NAR) indicated Table 15. were shown significant differences due to treatments at the all the stages. The NAR was maximum at 40-60 DAS and decreased thereafter. At 40-60 DAS, significantly higher NAR was found in seed hardening withCaCl2 (2%) followed by KNO3 (100 ppm), ascorbic acid (20 ppm), KH2PO4

(1%), sodium molybdate (100 ppm) and zinc sulphate (100 ppm). Significantly lower NAR was recorded in control at all the stages, which was on par with water soaking and seed hardening with CaCl2 (1%). Maximum NAR (0.339 g g

-1day

-1) was noticed in seed hardening with

succinic acid (20 ppm) at 60-80 DAS.

4.2.9 Biomass duration (g days)

The results on biomass duration (BMD) indicated significant differences among all the treatments as represented in Table 16.The BMD in all the treatments increased from 40-60 DAS to 80-harvest. Among the treatments seed hardening with 2% CaCl2 has recorded

Table 15.Influence of seed hardening chemicals on relative growth rate (g g-1

day-1

) and net assimilation rate(g dm

-2 day

-1) at different stages in chickpea

RGR NAR Treatments 40-60 60-80 40-60 60-80

T1 : Control 0.0276 0.0183 0.420 0.130

T2 : Water soaking 0.0271 0.0154 0.557 0.198

T3 : CaCl2 (1%) 0.0318 0.0144 0.680 0.208

T4 : CaCl2 (2%) 0.0295 0.0176 0.927 0.276

T5 : KH2PO4 (1%) 0.0338 0.0130 0.848 0.189

T6 : KNO3 (100 ppm) 0.0344 0.0173 0.909 0.261

T7 : KCl (100 ppm) 0.0339 0.0147 0.792 0.191

T8 : Sodium molybdate (100 ppm)

0.0348 0.0140 0.844 0.261


T10 : Cycocel (1000 ppm) 0.0301 0.0174 0.765 0.206



Mean 0.0317 0.0160 0.766 0.225

S.Em ± 0.0005 0.0007 0.048 0.025

CD(5%) 0.0015 0.0022 0.138 0.073

Table 16.Influence of seed hardening chemicals on biomass duration (g-days) at different stages in chickpea

BMD Treatments 40-60 DAS 60-80DAS 80-harvest

T1 : Control 78.3 202.7 362.6

T2 : Water soaking 107.6 253.4 434.7

T3 : CaCl2 (1%) 99.7 238.1 404.0

T4 : CaCl2 (2%) 144.4 372.4 618.5

T5 : KH2PO4 (1%) 116.3 271.3 481.4

T6 : KNO3 (100 ppm) 117.7 314.5 532.5

T7 : KCl (100 ppm) 109.9 269.2 456.4



T10 : Cycocel (1000 ppm) 137.3 354.8 572.2



Mean 114.9 288.4 481.8

S.Em ± 8.35 14.57 26.34

CD(5%) 24.12 42.12 76.15


significantly higher values followed by seed hardening with cycocel (1000 ppm) and succinic acid (20 ppm) as compared to control at all the stages. However, the control recorded the lower BMD at all the stages which was on par with water soaking and CaCl2 (1%) at 80-harvest. Maximum BMD (618.5 g days) was recorded in seed hardening with 2% CaCl2 at 80-harvest.

4.2.10 Specific leaf area (cm2)

The results pertaining to specific leaf area (SLA) indicated significant differences among the treatments at all the stages as indicated in Table 17.The SLA decreased upto 60 DAS and increased thereafter. Among the treatments, maximum SLA was obtained in control followed by CaCl2 (1%) and sodium molybdate (100 ppm) at 40 DAS. Significantly lower SLA was recorded in cycocel (1000 ppm) compare to all treatments at all the stages which was on par with 2% CaCl2 at 40 and 60 DAS. Maximum SLA (199 cm

2) was recorded in seed

hardening with 1% CaCl2 at 80 DAS.

Table 17.Influence of seed hardening chemicals on specific leaf area(cm2) at different stages

in chickpea

SLA Treatments 40DAS 60DAS 80DAS

T1 : Control 0.197 0.161 0.197

T2 : Water soaking 0.160 0.149 0.181

T3 : CaCl2 (1%) 0.183 0.152 0.199

T4 : CaCl2 (2%) 0.141 0.120 0.133

T5 : KH2PO4 (1%) 0.176 0.125 0.160

T6 : KNO3 (100 ppm) 0.162 0.119 0.140

T7 : KCl (100 ppm) 0.171 0.121 0.177



T10 : Cycocel (1000 ppm) 0.128 0.102 0.114



Mean 0.165 0.130 0.162

S.Em ± 0.006 0.006 0.005

CD(5%) 0.019 0.019 0.014


4.2.11 Leaf area ratio (cm2g-1)

The observations on leaf area ratio (LAR) indicated Table 18. were shown significant differences due to treatments at the all the stages. The LAR was maximum at 40 DAS and decreased thereafter. At 40 DAS, significantly higher LAR was found in control followed by seed hardening with sodium molybdate (100 ppm), KCl (100 ppm), CaCl2 (1%) and ascorbic acid (20 ppm). Significantly lower LAR was recorded in seed hardening with cycocel (1000 ppm) at all the stages, which was on par with seed hardening with 2% CaCl2 and succinic acid (20 ppm). Maximum LAR (27.7 cm

2g

-1) was noticed in control compared to all other

treatments at all the stages.

4.2.12 Relative water content (%)

The data on relative water content (RWC) presented in Table 19. indicated significant difference due to various treatments at all the stages except 40 DAS at it decreased from 40 to 80 DAS. Among the treatments, pre- sowing seed hardening with 2% CaCl2 significantly increased the RWC followed by seed hardening with cycocel (1000 ppm), succinic acid (20 ppm), KNO3 (100 ppm), ascorbic acid (20 ppm) and KH2PO4(1%) as compared to control. However, control recorded significantly lower RWC at all the stages but it was on par with water soaking and seed hardening with water soaking and seed hardening with CaCl2(1%) and KCl (100 ppm). Maximum RWC (59.0%) was noticed in seed hardening with 2% CaCl2 at 80 DAS.

Table 18.Influence of seed hardening chemicals on leaf area ratio (cm2 g) at different stages

in chickpea

LAR Treatments 40DAS 60DAS 80DAS

T1 : Control 111.2 74.8 27.7

T2 : Water soaking 88.8 60.0 24.0

T3 : CaCl2 (1%) 101.1 64.8 26.0

T4 : CaCl2 (2%) 72.0 51.1 19.1

T5 : KH2PO4 (1%) 97.5 56.9 25.0

T6 : KNO3 (100 ppm) 95.0 56.9 21.0

T7 : KCl (100 ppm) 100.5 58.0 24.3



T10 : Cycocel (1000 ppm) 69.6 41.7 16.3



Mean 92.3 57.1 22.4

S.Em ± 4.41 3.77 1.08

CD(5%) 12.75 10.89 3.12


Table 19.Influence of seed hardening chemicals on relative water content (%) at different stages in chickpea

RWC Treatments 40DAS 60DAS 80DAS

T1 : Control 80.3 53.0 46.9

T2 : Water soaking 81.6 59.9 50.6

T3 : CaCl2 (1%) 79.6 59.8 50.8

T4 : CaCl2 (2%) 84.2 68.8 59.0

T5 : KH2PO4 (1%) 79.4 62.5 53.4

T6 : KNO3 (100 ppm) 80.6 62.9 54.7

T7 : KCl (100 ppm) 80.9 59.2 51.6



T10 : Cycocel (1000 ppm) 83.7 65.1 56.9



Mean 81.1 61.5 53.3

S.Em ± 3.72 2.41 1.95

CD(5%) NS 6.97 5.59



4.3.1 Chlorophyll ‘a’ (mg g-1 fr.wt)

Significant differences observed in Table 20. due to various seed hardening treatments at all the stages with respect chlorophyll ‘a’ content It was observed that the chlorophyll ‘a’ content decreased from 60-80 DAS. Among the treatments, seed hardening with cycocel (1000 ppm) recorded significantly higher value followed by seed hardening with succinic acid (20 ppm) and 2% CaCl2 at all the stages. While, lower value was observed in control but, it did not differ significantly with water soaking, KCl (100 ppm) at 80 DAS. Maximum chlorophyll ‘a’ content (1.07 mg g

-1 fr.wt) was observed in seed hardening with

cycocel (1000 ppm) at 80 DAS.

4.3.2 Chlorophyll ‘b’ (mg g-1 fr.wt)

The data on chlorophyll ‘b’content indicated significant difference due to various treatments at both the stages in Table 20. The chlorophyll ‘b’ content increased at 60 DAS and decreased thereafter. Among the treatments, pre-sowing seed hardening with cycocel (1000 ppm) recorded significantly higher values which is on par with seed hardening with CaCl2 (2%), KNO3 (100 ppm), zinc sulphate (100 ppm) and ascorbic acid (20 ppm) at 60 DAS and succinic acid (20 ppm), CaCl2 (2%) and KNO3(100 ppm) at 80 DAS as compared to other treatments. While, control was recorded significantly low chlorophyll ‘b’ content. But, it did not differ significantly with water soaking and seed hardening with CaCl2 (1%), KCl2 (100 ppm) and KH2PO4 (1%). Maximum chlorophyll ‘b’ content (0.21 mg g

-1 fr.wt) was noticed in seed

hardening with cycocel (1000 ppm) at 80 DAS.

Table 20.Influence of seed hardening chemicals on chlorophyll ‘a’ and ‘b’ content ( mg g-1


Chlorophyll ‘a’ Chlorophyll ‘b’ Treatments 60DAS 80DAS 60DAS 80DAS

T1 : Control 1.34 0.47 0.27 0.13

T2 : Water soaking 1.51 0.59 0.28 0.15

T3 : CaCl2 (1%) 1.02 0.52 0.22 0.14

T4 : CaCl2 (2%) 1.72 0.86 0.45 0.18

T5 : KH2PO4 (1%) 1.55 0.80 0.29 0.17

T6 : KNO3 (100 ppm) 1.65 0.85 0.45 0.18

T7 : KCl (100 ppm) 1.22 0.54 0.30 0.13

T8 : Sodium molybdate (100 ppm)

1.37 0.75 0.34 0.16


T10 : Cycocel (1000 ppm) 1.86 1.07 0.50 0.21



Mean 1.51 0.75 0.36 0.16

S.Em ± 0.07 0.08 0.03 0.012

CD(5%) 0.20 0.23 0.07 0.034


4.3.3 Total chlorophyll (mg g-1 fr.wt.)

It was evident from the Table 21.that significant differences were observed due to various treatments at all the stages of investigation with respect to total chlorophyll content. The total chlorophyll content decreased from 60-80DAS. Among the treatments, seed hardening with cycocel (1000 ppm) followed by 2% CaCl2 recorded significantly higher values as compare to control at 60 DAS. However, significantly lower total chlorophyll content was observed in control at 80 DAS and it was found on par with water soaking and seed hardening with CaCl2 (1%) and KCl2 (100 ppm) at 80DAS. Maximum total chlorophyll content (1.28 mg g

-1 fr.wt.) was noticed in seed hardening with cycocel (1000ppm) at 80 DAS.

4.3.4 Chlorophyll stability index (%)

The data on Chlorophyll stability index (CSI) presented in Table 22. indicated significant differences due to various treatments and it increased from 60-80 DAS. Significantly higher CSI was found in seed hardening with 2% CaCl2 followed by cycocel (1000 ppm), succinic acid (20 ppm) and KNO3 (100 ppm) as compared to control at all the stages. However, significantly lower CSI was observed in control at all the stages and it was

Table 21.Influence of seed hardening chemicals on total chlorophyll content ( mg g-1


Total chlorophyll Treatments 60DAS 80DAS

T1 : Control 1.61 0.60


T3 : CaCl2 (1%) 1.24 0.66

T4 : CaCl2 (2%) 2.17 1.04

T5 : KH2PO4 (1%) 1.84 0.97

T6 : KNO3 (100 ppm) 2.10 1.03

T7 : KCl (100 ppm) 1.52 0.68



T10 : Cycocel (1000 ppm) 2.36 1.28



Mean 1.87 0.91

S.Em ± 0.069 0.086

CD(5%) 0.198 0.250


on par with water soaking, CaCl2 (1%), KCl (100 ppm) and sodium molybdate (100 ppm) at all the stages. Maximum CSI (79 %) was found in seed hardening with 2% CaCl2 at 80 DAS.

4.3.5 Proline content (µg g-1 fr.wt)

The results pertaining to proline content showed in Table 23. indicated significant differences due various treatments at all the stages. It increased from 60 to 80 DAS in all the treatments. Among the treatments, seed hardening with 2% CaCl2 recorded significantly higher values followed by cycocel (1000 ppm), KNO3 (100 ppm) and KH2PO4 (1%), as compared to control at all the stages. However, significantly lower proline was observed in control at all the stages. Maximum proline content (321 µg g

-1 fr.wt.) was recorded in seed

hardening with 2% CaCl2 at 80 DAS.

4.3.6 Epicuticular wax content (mg dm-2)

Significant differences observed due to various seed hardening treatments at all the stages with respect epicuticular wax content Table 24. It was observed that the wax content increased from 60-80 DAS. Among the treatments, seed hardening with 2% CaCl2 recorded significantly higher value followed by cycocel (1000 ppm) and succinic acid (20 ppm) at all the stages. While lower value was observed in control but it did not differ significantly with water soaking and seed hardening with KNO3 100 ppm), KCl2(100 ppm) , sodium molybdate(100 ppm) and zinc sulphate (100 ppm)at 80DAS. Maximum epicuticular wax content (0.673 mg dm

-2) was observed in seed hardening with 2% CaCl2 at 80 DAS.

Table 22. Influence of seed hardening chemicals on chlorophyll stability index (%) at different stages in chickpea

CSI Treatments 60DAS 80DAS

T1 : Control 48.0 51.8


T3 : CaCl2 (1%) 53.5 55.2

T4 : CaCl2 (2%) 75.8 79.2

T5 : KH2PO4 (1%) 64.4 67.5

T6 : KNO3 (100 ppm) 67.5 70.5

T7 : KCl (100 ppm) 54.6 57.6



T10 : Cycocel (1000 ppm) 72.2 76.3



Mean 60.9 63.8

S.Em ± 2.9 2.6

CD(5%) 8.3 7.5


4.4 Yield parameters

4.4.1 Seed yield (g plant-1)

The data on seed yield presented in Table 25. indicated that significant differences due to various treatments. Among the treatments, pre-sowing seed hardening with 2% CaCl2 recorded significantly higher seed yield (19.15 g plant

-1) followed by seed treatment with

cycocel 1000 ppm (16.10 g plant-1

) and succinic acid 20 ppm (14.67 g plant-1

) over control. Significantly lower seed yield (9.12 g plant

-1) notified in control. However, it did not differ

significantly with water soaking and CaCl2 (1%). Maximum seed yield of 19.15 g plant-1

was observed in seed treatment with 2% CaCl2.

4.4.2 Seed yield (q ha-1)

The observations on seed yield on hectare basis also indicated significant differences due to various treatments as shown in the Table 24. Among the treatments, pre-sowing seed hardening with 2% CaCl2 showed significantly higher values (26.52 g/ha) followed by seed treatment with cycocel 1000ppm (23.54 g/ha), succinic acid (20 ppm) and KNO3 100ppm (23.73 g/ha) as compare to control. Significantly lower seed yield (19.04g/ha) was noticed in control which was on par withWater soaking, CaCl2 (1%) and KH2PO4 (1%). Maximum seed yield (26.32 g/ha) was recorded in seed treatment with 2% CaCl2.

Table 23.Influence of seed hardening chemicals on proline content (µg g-1


Proline content Treatments 60DAS 80DAS

T1 : Control 164 217

T2 : Water soaking 168 250

T3 : CaCl2 (1%) 189 258

T4 : CaCl2 (2%) 220 321

T5 : KH2PO4 (1%) 207 283

T6 : KNO3 (100 ppm) 213 303

T7 : KCl (100 ppm) 200 276

T8 : Sodium molybdate (100 ppm) 197 264

T9 : Zinc sulphate (100 ppm) 200 284

T10 : Cycocel (1000 ppm) 218 312

T11 : Succinic acid (20 ppm) 191 275

T12 :Ascorbic acid (20 ppm) 207 287

Mean 198 277

S.Em ± 7.3 12.2

CD (5%) 21.0 35.3


Table 24.Influence of seed hardening chemicals on epicuticular wax content (mg dm-2


Epicuticular wax content Treatments 60 DAS 80DAS

T1 : Control 0.273 0.423


T3 : CaCl2 (1%) 0.357 0.500

T4 : CaCl2 (2%) 0.460 0.673

T5 : KH2PO4 (1%) 0.376 0.535

T6 : KNO3 (100 ppm) 0.331 0.490

T7 : KCl (100 ppm) 0.325 0.473



T10 : Cycocel (1000 ppm) 0.427 0.627



Mean 0.363 0.519

S.Em ± 0.021 0.027

CD (5%) 0.063 0.079


Table 25.Influence of seed hardening chemicals on yield and yield components at different stages in chickpea

Treatments Seed yield

(g plant-1

)

100seed waight

(g)

Seed yield

(q ha-1

)

Harvest index (%)

T1 : Control 9.12 26.64 19.04 41.1

T2 : Water soaking 11.11 28.69 19.41 42.0

T3 : CaCl2 (1%) 11.58 29.22 19.12 47.1

T4 : CaCl2 (2%) 19.15 30.77 26.32 53.1

T5 : KH2PO4 (1%) 13.48 26.66 22.02 44.0

T6 : KNO3 (100 ppm) 13.77 30.33 23.73 43.6

T7 : KCl (100 ppm) 12.85 24.89 20.65 46.2



T10 : Cycocel (1000 ppm) 16.10 27.70 23.54 49.2



Mean 13.38 27.47 21.98 46.5

S.Em ± 0.62 0.73 1.16 2.05

CD (5%) 1.81 2.11 3.38 5.95

4.4.3 Hundred seed weight (g)

It is evident from the Table 24. that there were significant differences in 100 seed weight due various seed hardening treatments. Among the treatments pre- owing seed hardening with 2% CaCl2 (30.77 g) recorded significantly higher test weight followed by seed treatment with KNO3 (100 ppm), CaCl2 (1%) and water soaking as compared to control (26.64 g). Significantly lower was noticed in control which was on par with KCl (100 ppm), zinc sulphate (100 ppm) and ascorbic acid (20 ppm).

4.4.4 Harvest index (%)

The data on harvest index indicated significant differences due to various treatments Table 24. Among the treatments seed hardening with 2% CaCl2 recorded significantly higher values (53.1 %) followed by cycocel 1000 ppm (49.2 %), zinc sulphate 100ppm ( 48.71%) and Sodium molybdate 100 ppm (48.58%) as compared to the control (41.1%).

5. DISCUSSION

Chickpea is essentially a rainfed (or) post monsoon winter crop. The average yield in India is very low compared to other countries. The main challenges in the future scenario will revolve around attaining self-sufficiency in chickpea production, to meet the increasing demand of protein and ensuring the environmental security. The area under chickpea is predominantly during rabi season and is characterized not only by the limited availability of soil moisture but also by high air temperature, particularly at pod formation stage. These conditions would invariably limit the productivity.

Depending on the extent of residual soil moisture and the scarce winter rains, chickpea suffers to varying degree of mounting moisture stress and consequently, productivity declines. It has been envisaged that better technology, particularly use of seed hardening techniques are known to cause the physiological changes in seeds which bring the seeds to resist drought conditions

To combat adverse moisture scarcity conditions, matching agro techniques needs to be evolved for various agro-ecological regions. Pre-sowing seed treatment to induce hardening (Misra and Dwivedi, 1980; Henckel, 1964 and Singh and Chatterjee, 1980), foliar spray of fertilizer solution during ontogenesis (Frewal and Mittal, 1982 and Dev and Dev, 1980) and mulching (Ali and Prasad, 1974 and Umrani et al., 1973) have been reported to increase productivity of pulses in rainfed areas. Most of the research work done so for has been on understanding the mechanism underlying productivity, but very little has been done with respect to the possibilities of overcoming stress or limitations imposed by environmental factors. It is important at this juncture to see and understand how best the stress effect can be minimized by adopting different strategies and to elucidate the impact of such strategies in enhancing productivity potential under water limited conditions.

In this direction, investigations were carried out to know the influence of seed hardening techniques on various morphological, physiological, biophysical and biochemical parameters, yield and yield components in chickpea (var. ICCV-2). The results obtained from this investigation are discussed in this chapter.

5.1 Morpho-physiological parameters

The seed hardening treatments significantly influenced the morphological characters viz., plant height, number of leaves and total dry matter production and it’s partitioning. Basically, plant height is a genetically controlled character, but several studies have indicated that the plant height can be increased or decreased by the seed hardening treatments.

However, in the present investigation, significant differences were noticed in the plant height due to seed hardening treatments at 60, 80 and at harvest. It was interesting to note that there was increase in the plant height over control except in seed hardening with cycocel (1000 ppm), where there was a decrease in plant height compared to control. Further, the plant height was significantly higher in pre-sowing seed hardening with CaCl2 (2%) followed by KH2PO4 (1%), KNO3 (100 ppm), KCl2 (100 ppm), zinc sulphate (100 ppm) and ascorbic acid (20 ppm). This clearly indicates that the mode of action is quite different for different chemicals studied. Similarly in finger millet, seed hardening with 2% CaCl2 was more effective and increased the plant height and such effect was due to redistribution of nutrient reserves leading to cell enlargement and increase in normal cell division (Karivartharaju and Ramakrishnan, 1985). Studies conducted with finger millet using CaCl2, ascorbic acid kinetin and benzyladenine showed greater beneficial effect in terms of germinability and seedling growth under simulated water stress condition or salinity (Krishna Sastry et al., 1969). Increase in the plant height was due to the seed hardening with KNO3, ascorbic acid and zinc sulphate by maintaining the turgidity of guard cells , which would further help for better plant growth through stomatal regulation .

The decrease in plant height in cycocel (1000 ppm) may be attributed to anti-auxin activity of CCC. It blocks the synthesis of IAA in the plant system and thereby reduces the plant height. The mechanism of reduction of plant height due to the seed hardening with cycocel appears to be due to reduced cell size and cell wall thickening or reduction in the cell

division activity (Ginzo, 1977). It has been suggested that the embryonic tissue provided a factor the gibberllin which is upon depression of repressed genes in the endosperm thereby activating the amylase, which further induced the breakdown of starch. Since cycocel is a retardant, the absorption of the chemical by the seed is likely to cause anti-gibberllin effects. The lesser availability of metabolic energy thus, may be responsible for the dwarfism exhibited by the cycocel treated seedlings (Paleg, 1969). Similarly, Dighe et al. (1983) indicated that cycocel (500 ppm) significantly reduced the plant height in wheat. It was observed that number of leaves due to various seed hardening treatments were differed significantly at all the stages. Significantly higher number of leaflets were noticed in seed hardening with 2% CaCl2 followed by seed hardening with KNO3 (100 ppm) and cycocel (1000 ppm) over control.

5.2 Dry matter production and its distribution

The dry matter production is of greater significance in the determination of yield and that of reproductive parts, particularly is an important yield contributing character. Although, the dry matter production in general is an indicative of efficiency of the genotypes and the pattern in which it is distributed in different plant parts would give a better understanding of genotypes potential. It is well documented that seed hardening treatments will have their influence on the production of dry matter and the way in which it is partitioned between different parts of the plant. In the present investigation, it is found that there was a significant increase in dry matter production of leaf, stem and reproductive parts due to seed hardening treatments at different concentrations. Among the treatments studied, seed hardening with 2% CaCl2 followed by cycocel (1000 ppm) and succinic acid (20 ppm) recorded significantly higher leaf, stem and reproductive parts dry weight. Further, it was observed that the extent of increase in dry matter production was more in the reproductive parts as compared to leaf and stem. Thus, it indicates the partitioning of dry matter in reproductive parts. The decline in the leaf dry weight at later stages could be due to translocation of stored assimilates towards the development of reproductive parts.

The amount of total dry matter (TDM) produced is an indication of the overall efficiency of the utilization of resources and better light interception. The data pertaining to total dry weight indicated that it increased continuously from 40 DAS to harvest. At later stage of the growth, dry matter accumulated at a decreasing rate, which could be attributed to reduced source activity leading to lesser dry matter accumulation in leaf and stem. These results are in concurrence with Arjunan and Srinivasan (1989), whose results also revealed that seed hardening with 2% CaCl2 significantly increased total dry matter production in groundnut. Singh et al. (1999) also revealed that seed treatment with cycocel (10

-4M)

significantly increased the root, stem and total dry weight as compared to control in wheat.

The increase in TDM towards maturity may be due to indeterminate growth pattern, higher rate of CO2 fixation and RuBP carboxylase activity during crop growth. The association of TDM with grain yield was more significant at all the stages of crop growth. Thus, TDM is an important parameter in boosting the source-sink relationship and yield potential. Similar observations were made by Nam et al., (1998) and Katti et al., (1999) in pigeonpea.

5.3 Growth parameters

The assessment of yield variation in terms of growth and development is very complex, since it involves the effect of external factors on all the physiological processes in plants.

It is well established fact that the infrastructure of the plant is decided by the growth parameters such as leaf area index (LAI), leaf area duration (LAD), crop growth rate (CGR), relative growth rate (RGR), net assimilation rate (NAR), specific leaf weight (SLW) and biomass duration (BMD). Growth analysis technique has made substantial contribution to the current understanding of the physiological basis of yield variation in different crops, but the information on chickpea is very meager. Leaf area fairly gives a good idea of the photosynthetic capacity of the plant.

In the present study the leaf area and leaf area index (LAI) increased drastically upto 60 DAS and decreased thereafter towards maturity. In general, the seed hardening chemicals

showed profound significant effect over these parameters. However, seed hardening with 2% CaCl2 recorded significantly higher LAI followed by KNO3 (100 ppm) as compared to other treatments. Maitra et al (1998) also noticed that seed hardening with 2 per cent CaCl2 recorded significantly higher LAI as compared to control in finger millet. Similarly, Govindan and Thirumurugan (2000) revealed that foliar spray of KCl (1%) or KNO3 (1%) or in combination, increased the LAI over control in green gram.

Leaf area duration (LAD) is another parameter that is determined by the LAI of two consecutive growth stages. This was a useful concept, not only for predicting the efficiency of photosynthetic system but also for dry matter production (Chetti and Shirohi, 1995). It indicated the maintenance of assimilatory surface area over a period of time, which is a pre-requisite for the prolonged photosynthetic activity and the ultimate productivity in crop plants. The increase in LAI and LAD could be mainly due to the maintenance of more green leaf area. In the present investigation, it increased upto 80 DAS. The seed hardening with 2% CaCl2 recorded significantly higher values followed by KNO3 (100 ppm) and ascorbic acid (20 ppm) as compared to other treatments. These results are in conformity with Wright et al., (1983) who observed higher LAI and LAD due to higher grain yield in sorghum. Maitra et al., (1998) reported that seed hardening with 2% CaCl2 recorded significantly maximum LAI and LAD over control in finger millet.

Relative growth rate (RGR) represents the increase in dry matter per unit of dry matter already present per unit time. It was found in the present study that RGR declined with advancement of the crop growth. The decline in RGR with the advancement in crop growth could be due to decline on the rate of dry matter production. The increase in the RGR due to seed hardening treatments could be due to the effectiveness of chemicals in increasing not only dry matter production but also the rate of increment in total dry matter. This also could be attributed to increase in photosynthetic efficiency by increasing leaf thickness and retaining more chlorophyll content. Efficient translocation of photosynthates. Similarly, Kamala Thirumalaiswamy and Sakharam rao, (1977) opined that, seed treatment with cycocel significantly increased RGR under moisture stress condition and also the size of the leaves was greatly influenced by cycocel.

According to Watson (1952), crop growth rate (CGR) is useful growth parameter for estimating production efficiency of crop stand. The computation of CGR at different growth stages indicated that CGR was maximum at 60-80 DAS. In the present study seed hardening treatments significantly increased CGR over control. Among the treatments seed hardening with 2% CaCl2 recorded significantly higher values followed by cycocel (1000 ppm) and KNO3 (100 ppm) as compared to control. These results are in accordance with Maitra et al., (1998) who revealed that seed hardening with 2% CaCl2 significantly increased the CGR over control in finger millet.

In the present study, seed hardening treatments, use of hardening chemicals increased absolute growth rate (AGR) over control at all the stages. The seed hardening with 2% CaCl2 significantly increased the AGR followed by cycocel (1000 ppm), KNO3 (100 ppm), sodium molybdate (100 ppm) and ascorbic acid (20 ppm), as compared to control.

Net assimilation rate (NAR), synonymously called as “unit leaf rate”, express the rate of dry weight increase at any instant on a leaf area basis with leaf representing an estimate of the size of the assimilatory area (Gregory, 1926). The decline in NAR with advancement of the crop growth could be attributed to a decline in the rate of dry matter production coupled with leaf area though the leaf area increased upto 60 DAS, the NAR declined from 60-80 DAS. This is not only due to reduce rate of leaf area but also due to reproductive dry matter which is evidenced from RGR. The highest values for the NAR were recorded in seed hardening with 2% CaCl2 followed by cycocel (1000 ppm), succinic acid (20 ppm), KNO3 (100 ppm), Ascorbic acid (20 ppm) and KH2PO4 (1%) as compared to control. Similarly, Black (1957) also reported that higher NAR coupled with higher RGR in most of the stages resulted in higher dry matter accumulation in barley. Eshanna and Kulkarni (1990) who revealed that pre-sowing treatment with CaCl2 (1:3 proportion) recorded maximum NAR over control in maize.

The specific leaf weight (SLW) indicates leaf thickness and which is due to compactness and stacking of mesophyll cells. Since chickpea is a C3 plant, the

photosynthetic efficiency per unit leaf area is low and the increase in leaf thickness could probably be due to enhanced photosynthetic efficiency and more stacking of mesophyll cells and bundle sheath cells thereby recapturing the CO2 released in photo-respiratory process and leading to an increased total dry matter accumulation. In the present study, the SLW increased upto 60 DAS and declined thereafter .The specific leaf weight was more in seed treatment with cycocel (1000 ppm) followed by succinic acid (20 ppm) and 2% CaCl2 as compared to control.

Biomass duration (BMD) indicates the maintenance of dry matter over a period of time and is essential for prolonged supply of photosynthates to the developing sink. The biomass duration significantly increased due to the seed hardening with 2% CaCl2 followed by cycocel (1000 ppm) and succinic acid (20 ppm) as compared to control which could be attributed to increased dry matter production and its maintenance. Similarly, Koti (1997) showed positive association between BMD and seed yield in soybean. Chougale (1997) also indicated that significant differences in biomass duration with the application of growth regulators. Similarly, Padmavathi (1998) in onion, Prabhu (2000) in blackgram and Sai Sankar (2002) in greengram showed positive association between BMD and yield and also reported that higher BMD was recorded with plant growth regulators. The increase in BMD with age due to growth regulator treatments and could be mainly attributed to increase in total dry matter, which again depends on the development of leaf area and which is evident from the LAI and LAD.

Leaf area ratio (LAR) expresses the ratio between the leaf lamina or photosynthesising tissue and the total respiring plant tissue or a total biomass. It reflects the leafiness of the plant. In our study LAR differ significantly ay all the stages. Lower LAR was recorded in seed hardening with cycocel 1000 ppm (16.3 cm

2g

-1) compared to all other

treatments. Lower LAR was may be due to growth retardant property of cycocel. Since cycocel is a retardant, the absorption of the chemical by the seed is likely to cause anti-gibberllin effects. The lesser availability of metabolic energy thus, may be responsible for the dwarfism exhibited by the cycocel treated seedlings (Paleg, 1969)

A better understanding of different biophysical characters and their relationship with yield and other physiological process is essential. In this direction, influence of seed hardening treatments was studied. Relative water content (RWC) is a measure of the amount of water present in the leaf tissue in relation to turgid condition and the treatments having higher RWC under drought condition would be preferable to maintain higher water balance. In the present study, the seed hardening with 2%CaCl2 recorded significantly higher RWC values followed by cycocel (1000 ppm), succinic acid (20 ppm), KNO3 (100 ppm) , ascorbic acid (20 ppm) and KH2PO4 (1%) compared to control. These results are in accordance with Patil (1987) who revealed that pre-sowing seed treatment with 2%CaCl2 recorded higher RWC in sorghum. Similar results were reported in wheat by Amaregouda et al., (1994)


Identification of substances which play an important role in osmoregulation is another approach of screening for drought tolerance in crop plants. It is well documented that under drought conditions many osmotically active solutes accumulate in leaves of higher plants which lower the osmotic potential and maintain the turgor of both shoot and roots (Munns et

al., 1979 and Jones et al., 1980). In analogy to this, attempts have been made to find out the influence of exogenously applied synthetic plant growth regulators and also seed hardening chemicals on various plant growth and developmental process in several crops. Studies indicate that this minute molecules trigger various physiological processes such as photosynthesis, photorespiration, partitioning of photoassimilates, mineral ion uptake and nitrogen metabolism, which in turn is linked by numerous interactions leading to productivity.

Significant increase in the chlorophyll ‘a’, chlorophyll ‘b’ and total chlorophyll contents was observed due to seed hardening treatments at all the changes. The chlorophyll content increased upto 60 DAS and decreased thereafter at all the stages. Among the treatments, the maximum chlorophyll ‘a’, chlorophyll ‘b’ and total chlorophyll contents were observed in seed hardening with cycocel (1000 ppm). The delay in leaf senescence, increased SLW, reduction in cell size with denser cytoplasm and inhibition of chlorophyll breakdown could be attributed to higher chlorophyll content in cycocel treated plants. These results are in accordance with

Cheema (1975) who reported that seed treatment with cycocel increased the chlorophyll content in barley. Similarly Jayakumar and Thangaraj (1998), who explained that the application of cycocel to groundnut resulted in higher chlorophyll content without the modification of leaf anatomy and decreased chlorophyll degradation.

Chlorophyll stability index (CSI) was increased from 60 to 80 DAS. Significantly higher CSI was recorded in seed hardening with 2%CaCl2 over control. Here the destruction of chlorophyll pigment changes due to terminal stress, because chlorophyll stability is a function of temperature .This property of chlorophyll stability was found to correlate well with drought resistance. High CSI was corresponded with more drought tolerance (Hiscox and isrealstam ,1979). This is evendent from the specific leaf weight.

Free proline content has been shown to accumulate upon desiccation in leaves of many plant species. It has been suggested by Jones et al., (1980) that proline accumulation could make useful contribution to the osmotic adjustment. Blum and Ebercon (1976) indicated that proline plays an important role as storage compound for carbon and nitrogen, detoxification of NH3, preserving the hydration of proteins in dehydrated tissues thereby contributing to the survival of cellular functions. In the present study, it is clear from the data that, seed hardening with 2% CaCl2 recorded significantly higher proline content followed by cycocel (1000 ppm), KNO3 (100 ppm) and KH2PO4 (1%) as compared to control at all the stages. These results are in accordance with Patil (1987) who reported that seed treatment with2% CaCl2 significantly increased proline content in sorghum and similar results were reported by Amaregouda et al. (1994) in wheat. Epicuticular wax content was increased from 60-80DAS. Here there was gradually increase in the wax content on the cuticular surface of leaves. This indicated that epicuticular wax content (0.673 mg dm

-2) was observed in seed

hardening with 2% CaCl2 at 80 DAS.This was because to decrease the transpiration losses in terminal drought (Ebercon et al., 1977).

5.5 Yield and yield components

Grain yield is the manifestation of morphological, physiological, biochemical, biophysical and growth parameters. Improvement in yield according to Humphries (1979) could happen in two ways i.e., adopting the existing varieties to grow better in their environment or by altering the relative proportion of different plant parts so as to increase the yield of economically important parts. The influence of seed hardening chemicals significantly increased the seed yield. The increased seed yield may be attributed to higher dry matter production and its partition in reproductive parts, higher NAR, CGR, BMD, LAD, LAI, enhanced chlorophyll content and proline content.

In the present investigation, it is observed that the seed yield, 100 seed weight both on plant and area basis increased due to different seed hardening chemicals. The increase in seed yield could be attributed to significant enhancement in the growth parameter viz., CGR, SLA, LAD, and BMD in addition to harvest index.

The present study also revealed that the increase in seed yield was significantly more in seed hardening with 2% CaCl2 followed by cycocel. Data clearly indicated that seed hardening treatments are very effective in increasing yield land yield attributes compared to in control. The increase in the vegetative characters that intern enhanced cell division and quick cell multiplication, while the higher yield may be due to better carbon assimilation, better accumulation of carbohydrates and reduced respiration in plants. The results are in agreement with the findings of Mahabir singh and Rajodia (1989), Singh and Dohare (1964) and Das and Prusty (1982).

In addition, the present study also indicated that seed hardening with CaCl2, ascorbic acid and CCC significantly increased seed yield per plant, 100 seed weight and harvest index, which are most important yield determining components in chickpea. The increase in seed yield with respect to seed hardening treatments was probably due to maximum water absorbing capacity of seeds, more intense photosynthetic activity and more tissue hydration thereby, enabling the plant to resist soil moisture stress more efficiently (Henckel, 1964). This is in conformity with the findings of earlier research workers that pre-sowing seed hardening with cycocel (50 mg/1) recorded significantly higher yield per plant in okra (Mehrotra et al.,

1970). Seed hardening with CaCl2 (1%) recorded significantly higher number of pods per plant and pod yield in groundnut (Arjunan and Srinivasan 1989).

Several research workers have indicated increase in the yield of chickpea due to different seed hardening chemicals. Sen and Misra (1987) reported that treating wheat seeds with 0.25 % CaCl2 or 2.5 % KCl increased the grain yield compared to control. Similarly Patil (1987) opined that seed treatment with 2% CaCl2 for four hours increased drought resistance in sorghum and also increased grain yield by 10 per cent over control under dryland condition. Shindhe et al. (1991) opined that foliar spray of NAA and KNO3 increased pod yield in cowpea. Singh et al., (1991) found that foliar spray of cycocel, mixtalol and triacontanol effectively enhanced seed yield in chickpea. Masood Ali, (1985) indicated that foliar spray of 2% KCl solution significantly increased grain yield in chickpea. Amaregouda et al. (1994) noticed that treatment with CaCl2 (2 %) had given higher yield in wheat.

5.6 Future line of work

1. It is necessary to understand the changes in the genetic make up of the plant due to seed hardening chemicals at molecular level.

2. Little is known about the influence of seed hardening chemicals at metabolic level on enzyme activity etc. and synthesis of proline under drought and there is ample scope for such studies.

3. It is also important to study seed hardening chemicals in the manipulation of source-sink relationship.

4. It is required to study seed hardening chemicals with different concentrations, to determine optimum concentrations which have profound influence on yield and yield components in chickpea.

5. It is worthwhile to study the anatomical changes brought by different seed hardening techniques under soil moisture stress conditions

6. SUMMARY AND CONCLUSIONS

A field experiment was undertaken to find out the effect of different seed hardening techniques on morphological, biochemical, yield and yield attributing characters under rainfed condition in chickpea during rabi 2006 at College of Agriculture Farm, University of Agricultural Sciences Dharwad. The experiment consisted of 12 treatments with various seed hardening chemicals at different concentrations. The experiment was laid out in randomized block design with three replications. The results obtained from the present investigations are summerised in this chapter.

1. The plant height increased significantly due to the seed hardening treatment with 2% CaCl2, KNO3 (100 ppm) and ascorbic acid (20 ppm) as compared to control.

2. Number of leaflets increased significantly due to seed hardening treatment with 2% CaCl2,

KNO3 (100 ppm) and cycocel (1000 ppm) as compared to control.

3. Dry weight of leaf, stem and reproductive parts increased significantly due to seed hardening with 2% CaCl2 and cycocel (1000 ppm) and succinic acid (20 ppm) as compared to control. It was found that leaf dry weight increased upto 60 DAS and decreased thereafter but the dry weight of reproductive parts increased upto harvest.

4. The leaf area index increased upto 60 DAS and declined thereafter. The treatment of seed hardening with 2% CaCl2 and KNO3 (100 ppm) increased the leaf area index as compared to control.

5. The pre-sowing seed hardening treatments significantly increased CGR, AGR from 40 to 80 DAS and RGR and NAR significantly increased upto 40 to 60 DAS and declined thereafter.

6. Leaf area duration (LAD) increased from 40-60 DAS to 60-80 DAS due to seed hardening chemicals. Biomass duration (BMD) also followed the similar trend.

7. Specific leaf weight (SLW) increased significantly upto 60 DAS and declined thereafter due to seed hardening chemicals. However, higher SLW was found with seed hardening with cycocel(1000 ppm) , succinic acid (20 ppm) and 2% CaCl2.

8. Maximum leaf area ratio (LAR) was recorded in 40 DAS and decreased thereafter due to seed hardening chemicals. However, lower LAR was recorded with seed hardening with cycocel 1000 ppm.

9. There was a significant increase in chlorophyll ‘a’, chlorophyll ‘b’ and total chlorophyll contents due to various seed hardening treatments at all the stages. However, maximum chlorophyll content was observed in seed hardening with cycocel (1000 ppm) and 2% CaCl2.

10. There was a significant increase in chlorophyll stability index from 60-80DAS due to various seed hardening treatments. However, maximum CSI was observed in seed hardening with 2% CaCl2 ,cycocel (1000 ppm) and succinic acid (20 ppm) over control..

11. The relative water content (RWC) was significantly higher in seed hardening with CaCl2 (2 %) followed by cycocel (1000 ppm) and seed hardening with succinic acid (20 ppm).

12. There was a significant increase in proline content due to various treatments at all the stages. However, the maximum proline content was observed in seed hardening with 2% CaCl2 at all the stages.

13. The epicuticular wax content was recorded significantly higher in seed hardening with 2% CaCl2 , cycocel (1000 ppm) and succinic acid (20 ppm).

14. The seed yield and yield components viz., seed yield per plant, seed yield per hectare, test weight and harvest index were significantly higher due seed hardening with 2% CaCl2 followed cycocel (1000 ppm) and succinic acid (20 ppm).

15. Based on the above results, it is concluded that the seed hardening with 2% CaCl2 and cycocel (1000 ppm) is more effective in increasing the yield of chickpea.

Conclusions

Improvement of seed quality by seed hardening is a simple, easy and inexpensive approach to enhance the seed performance and agricultural productivity especially in the drylands and marginal lands of resource poor farmers. This will maintain higher water balance in the tissue and enhance the photosynthetic activity which maight have ultimately contributed to an increse in the drought tolerance capacity and incresed palnt vigour. As per the results obtained due to various seed hardening treatments, it is concluded that seed hardening with 2% CaCl2 and cycocel (1000 ppm) is more effective in incresing the yeild of chickpea.

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PHYSIOLOGICAL BASIS OF SEED HARDENING IN CHICKPEA (Cicer arietinum L.)

MANJUNATHA B. L. 2007 Dr. M. M. DHANOJI MAJOR ADVISOR

ABSTRACT

A Field experiment was conducted at College of Agriculture Farm, University of Agricultural Sciences, Dharwad during rabi 2006 to study the influence of various seed hardening chemicals on various on morpho-physiological, biochemical, yield and yield components in Chickpea (var ICCV-2). The experiment was laid out in randomized block design with three replications. The treatments consists of seed hardening treatments with water, CaCl2, KH2PO4, KNO3, KCl, sodium molybdate, zinc sulphate, Cycocel, Succinic acid and Ascorbic acid.

Significant differences were observed for various morpho-physiological biochemical, yield and yield components due to various seed hardening chemicals. Significant increse in plant height, dry matter in leaf, stem and reproductive parts and total dry matter content was due to treatments as compare to control. The growth parameters viz., LAI, AGR, CGR, RGR, NAR, LAD, LAR, SLW and BMD increase significantly due to seed hardening with calcium chloride (2 %), cycocel (1000 ppm) and succinic acid (20 ppm). The biochemical parameters viz., total chlorophyll content, prolin content and epicuticular wax content incresaed significantly due to seed hardening with calcium chloride (2 %), cycocel (1000 ppm) and succinic acid (20 ppm).

Seed hardening with CaCl2 recorded significantly higher seed yield followed by cycocel (1000 ppm) and succinic acid (20 ppm) and the incresed yield was due to higher test weight and harvest index. As per the results obtained seed hardening with 2% CaCl2 and cycocel (1000 ppm) is more effective in increasing the yield of chickpea.

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PHYSIOLOGICAL BASIS OF SEED HARDENING IN CHICKPEA