Effect of different water regimes and organic manures on indole acetic acid ( IAA ) , oxidase activity , leaf area , light transmission ratio , chlorophyll stability index , relative water content and yield attributes of noni ( Morinda citrifolia L . )

An experiment was conducted at Horticultural College and Research Institute, Tamil Nadu Agricultural University, Periyakulam in split plot design with irrigation regimes on main plot (four levels) and organic manures on sub plot (eight levels) with two replications. Among the different treatment combinations, M2S4 (100% WRc through drip irrigation + 50% farmyard manure + 50% vermicompost) exhibited superior performance for leaf area, light transmission ratio, chlorophyll stability index, relative water content and fruit yield as against the poor performance by M4S8 (check basin method of irrigation + no manures and no fertilizers) at different growth phases. Indole acetic acid (IAA) oxidase activity was found to be the lowest in M2S4 (100% WRc through drip irrigation + 50% farmyard manure + 50% vermicompost) as against the highest in M4S8 (check basin method of irrigation + no manures and no fertilizers).


INTRODUCTION
Rise in population, inadequate supply of life saving essential drugs in certain parts of the world, prohibitive cost of treatments for even some common ailments, side effects of several allopathic drugs in current usage and development of resistance to these drugs for infectious disease have lead to increased emphasis on the use of herbal plant materials as source of medicines for the range of human ailments (Meena et al., 2009).Cultivation of medicinal and herbal plants has assumed greater significance in recent days due to the tremendous potential, they offer in formulating newer drug formulations against many diseases.Chemical residue free plant based drugs are much valued than that obtained from plants cultivated inorganically (Padmanabhan, 2003).
Valid information related to organic production techniques in medicinal plants, especially for noni is scanty.Hence, the field level study on the "Development of organic production techniques for noni (Morinda citrifolia) under varying water regimes through drip irrigation for higher yield" was conducted at Horticultural College and Research Institute, TNAU, Periyakulam.
Morinda citrifolia L., popularly known as Indian mulberry or Indian noni belonging to the family Rubiaceae.It is grown in tropical regions of the world.It is one of the most significant sources of traditional medicines among Pacific Island societies (Mathivanan et al., 2005).The roots, stems, bark, leaves, flowers and fruits of the noni plant are all involved in various combinations in almost 40 known and recorded herbal remedies.Noni fruit has excellent levels of carbohydrates and dietary fiber and is a good source of protein.Noni pulp is low in total fats.About 160 phytochemical compounds have been identified in the noni plant (Rethinam and Sivaraman, 2007).Noni fruit contains a number of enzymes and alkaloids that are believed to play a pivotal role in maintaining a good health.The fruit juice is in high demand in alternative medicine for different kinds of illnesses such as arthritis, diabetes, high blood pressure, muscle aches and pains, menstrual difficulties, headaches, heart disease, AIDS, cancers, gastric ulcers, sprains, mental depression, senility, poor digestion, atherosclerosis, blood vessel problems and drug addiction.Current trends reflect that people have a more inclination towards herbal medicines for its healing properties and no side effects.Noni best suits the current proposition where it has proved that it has got tremendous medicinal properties and is being used even in the treatment of cancer.Looking at the wide range of prospects that noni can offer in terms of medicinal properties, it is quite clear that the plant can be exploited for the benefit of human race.Given the conditions that are required for the noni to grow, it would be a good idea if its cultivation is encouraged in the state.If successful, it may give a boost to the local economy through commercialization.
Any crop management practice should aim in keeping the physiological process of the plants as an active stage and plant enzymatic activities in favourable condition so that the plants can produce biomass inturn yield with the least destructive processes.Indole acetic acid (IAA), a premier bioregulator, regulates the apical dominance and initiation of vegetative and flower buds in various crop plants.The amino acid tryptophan and zinc level in the leaves influences the IAA.IAA oxidase is the enzyme responsible for destruction of auxin through the process of oxidation.Therefore, the enzymatic activity causes reduction in auxin content and thereby decreases the normal growth of the plant.Total leaf area at any stage of the crop growth is an important aspect of noni as it has a close association with photosynthetic efficiency, reflecting on biomass production.Greater leaf area aids the plant to synthesize more metabolites exhibiting high photosynthetic Kumar and Ponnuswami 551 rate during the period of growth and development (Mahadevan, 1988).A higher photosynthetic activity is a good indication of physiological efficiency in plants.This primarily depends upon the chlorophyll content in the leaves.This content in leaves indicates the efficiency of photosynthesis where the solar energy is converted into chemical energy.A slight fluctuation in chlorophyll content is enough to trigger changes in physiological processes of the plants particularly photosynthesis.
Relative water content represents the ability of the plants to maintain its tissue water status even under stress situations and the plants retaining more tissue water are expected to perform better through maintaining proper hydration of plant protoplasm and turgidity of the assimilatory cells.Yield is a complex trait influenced by many factors.Apart from modern day cultivars, nutrient management system plays a crucial role on yield.Yield per plant is the culmination of the interplay of several factors like morphological traits, physiological parameters, soil biological properties, fruit characters and yield parameters.The purpose of all improved cultural operations is to manipulate these parameters thereby to attain increased yield level in crops.

MATERIALS AND METHODS
This study was conducted at Horticultural College and Research Institute, TNAU, Periyakulam, Tamil Nadu, India which is situated at 77°E longitude, 10°N latitude and at an altitude of 300 m above mean sea level.The nature of soil of the experimental plot is sandy loam.The details of the initial soil chemical and physico-chemical characteristics of the experimental field were furnished in Table 1.

Computed water requirement
Computed water requirement of noni was calculated by using the following formula The quantity of water applied during the study period is presented in Table 3.

IAA oxidase
The IAA oxidase enzyme activity in the leaf sample was determined colorimetrically at 540 nm as per the method of Parthasarathy et al. (1970).The OD values were referred to a standard curve using auxin (IAA -10 to 100 µg/l) and expressed as µg un-oxidized auxin g -1 h -1 of the fresh leaf sample.

Leaf area per plant
The leaf area was calculated by using graph sheet and expressed in cm 2 .

Light transmission ratio (LTR)
LTR measurements were made by placing the Lux meter above the canopy and also on the ground surface (Salki, 1963) and the ratio is expressed in per cent.
LTR (%) = Where: I0, Light intensity above the canopy; I1, Light intensity at the ground surface 5 -10 measurements were made in one canopy and average values were obtained.

Chlorophyll stability index (CSI)
CSI was assessed following the method of Murty and Majumder (1962).Two clean test tubes were taken (control and treatment).250 mg of fresh leaf samples were weighed and cut into 8 to 10 leaf bits and transferred to test tubes.20 ml of distilled water was added to the control test tube and 20 ml of hot water was added to the treatment test tube.The treatment test tube was kept in hot water bath for exactly 30 min.The other tube was kept as control.After completion of the reaction time, the leaf bits were taken from test tubes and macerated with 10 ml of 80% acetone.The contents were centrifuged at 3000 rpm for 10 min.The supernatant was collected and volume was made to 25 ml by using 80% acetone.The optical density was measured at 652 nm in a spectrophotometer.
Total chlorophyll content (treatment) Total chlorophyll content (control)

Relative water content (RWC)
The relative water content of leaves was calculated as per the method suggested by Barrs and Weatherly (1962) to find out the percentage of water held by leaves relative to fully turgid tissue.
Leaves were punched uniformly and the fresh weight of 20 leaf discs was recorded.Then the leaf discs were made to float in water for 2 h, after which the turgid weight was recorded after removing the excess water by blotting them thoroughly.The dry weight was recorded after drying in hot air oven at 70°C.
Fresh weight -dry weight Turgid weightdry weight Fruit yield per hectare The yield was recorded after weighing fully matured fruits at each harvest, summed and expressed in tones.

Statistical analysis
The statistical analysis of data collected was done by adopting the standard procedures of Panse and Sukhatme (1985).The AGRES software (Version 3.01) was used for analysis of data.

Leaf area
There was a progressive increase in leaf area per plant in all the treatments as the age of the crop advanced (Table 5 and Table 6).The treatment M 2 (100 per cent WRc through drip irrigation) produced the highest leaf area per plant (23670.19,69669.69 and 174016.14cm 2 ) in vegetative, flowering and harvesting stages respectively.Among the main plots, M 4 (check basin method of irrigation) exhibited the lowest leaf area per plant (8561.51,23072.83and 67017.29 cm 2 ) in all stages of crop growth.Regarding the various manure treatments, combined application of 50% FYM + 50% VC (S 4 ) registered more leaf area per plant in vegetative (23027.16cm 2 ), flowering (68171.99cm 2 ) and harvesting (179726.75 cm 2 ) stages.While S 8 (no manure and no fertilizers) showed the lowest leaf area per plant of 5916.31,14687.42 and 41619.91 cm 2 in these three stages respectively.
Among the interactions, the treatment combination of M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC) produced the highest leaf area per plant (33655.16,100507.68 and 271346.10cm 2 ) in vegetative, flowering and harvesting stages respectively.Leaf area per plant was found to be the lowest (4531.74,11692.80 and 33166.92cm 2 ) in the treatment combination M 4 S 8 (check basin method of irrigation + no manure and no fertilizers).

Fruit yield
When main plot treatments were rated based on their performance (Table 10), it was known that application of 100% WRc through drip irrigation (M 2 ) had resulted in the highest fruit yield per hectare (19.35 t) while the lowest (10.91 t) was found to be with check basin method of irrigation (M 4 ).Regarding the sub plots, application of 50% FYM + 50 % VC (S 4 ) exhibited the superior scores for fruit yield per hectare (19.96 t).While the treatment S 8 (no manure and no fertilizers) showed very poor performance for fruit yield per hectare with 5.66 t.In the combined effect of treatments, the treatment combination comprising 100% WRc through drip irrigation + 50% FYM + 50% VC (M 2 S 4 ) recorded the highest scores for fruit yield per hectare with 25.68 t.Fruit yield per hectare was found to be the lowest in M 4 S 8 (check basin method of irrigation + no manure and no fertilizers) with 4.87 t.

DISCUSSION
IAA oxidase estimated at various crop growth stages revealed differences among water regimes and organic manures.Among the various treatment combinations, M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC) exhibited the lowest IAA oxidase activity.IAA oxidase is an enzyme very sensitive to both biotic and abiotic stresses.
It is responsible for inactivation of auxin in plant system.Galston and Dalberg (1954), measured IAA oxidase activity and growth response of 7 to 8 days old etiolated pea seedlings, and they revealed that IAA oxidase activity was low in regions of high auxin content.IAA stimulates the cellulose synthetase and regulates cell wall synthesis (Aref and Ray, 1967).It revealed that the high yielding plants had favorable auxin balance through the destruction of the IAA oxidative degradation.In the plants with lesser levels of available moisture and nutrients in soils the IAA synthesis might have been insufficient for encouraging IAA oxidative metabolism.Combined application of 100 per cent WRc through drip  irrigation + 50% FYM + 50%VC (M 2 S 4 ) has led to higher leaf area.This may due to continuous and uninterrupted judicious supply of water and nutrients.This finding was strengthened by previous research of Umesha et al. (2011).The plants maintain a turgid condition during the day time under drip irrigation as compared to check basin method of irrigation.There is a possibility of wide opening of stomata for longer period which might have resulted in high exchange of gases.Similarly, noni leaves might have remained turgid and produced more leaf surface.Thus, in turgor state helps in absorption of more sun light and solar radiation.It could have resulted in higher rate of photosynthesis and increased the photosynthetic capacity, which ultimately might have resulted production of leaf area and in turn the LTR (Kadam, 1990).The crops experienced period of water stress before each irrigation under check basin method irrigation due to the limited availability of moisture and nutrients for the roots to absorb.As results of that, there is reduction in leaf area and LTR.The higher leaf area is as a result of maintenance of favorable soil moisture in the rhizosphere and effective absorption by plants.The optimum P uptake and greater P mobility through frequent or continuous low volume irrigation can maintain three dimensional distribution patterns of water and nutrients and provide improved conditions for growth, water and nutrient uptake (Gal and Dudley, 2003).Similarly the higher leaf area and LTR may be due to the optimum uptake of nutrients especially nitrogen, iron and magnesium from the soil resulted in higher chlorophyll content which might have enhanced the photosynthetic rate and production of more leaf area.Similarly, in ginger, farmyard manure influenced the length and breadth of leaf and it is in accordance with the present investigation (Khandkar and Nigam, 1996).
The inoculation of farmyard manure in the treatment would have increased the friability, promoted aggregation of soil and increased the level of humus, thereby increasing the microbial activity.This in turn would have enhanced the production of photosynthates due to efficient photosynthesis leading to accumulation of more carbohydrates.This may be responsible for increased leaf area and LTR.Furthermore, inoculation of the biofertilizer, Azospirillum would have increased the activity of root exudates which in turn might have accelerated the activity of beneficial microbes by higher nitrogen fixation and secretion of growth promoting substances as reported by Okon and Kapulnik (1986) which owed to the luxuriant vegetative growth that was reflected on improved leaf area.
Two specific physiological attributes indicating abiotic stress tolerance viz., CSI and RWC of leaves was found to be the highest in the treatment combination M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC).Koleyreas (1958),  Murty and Majumder (1962) and Kilen and Andrew (1969) while working with pine, rice and corn respectively found significant correlations between CSI and drought tolerance.Sanandachari (1978) reported that, CSI appeared to be more reliable index to assess the drought resistance or tolerance capacity in sugarcane varieties.Todd and Webster (1965) and Alizada and Sultanov (1970) interpreted the RWC of leaves as an evidence of higher drought tolerance in crops.The leaves of drought tolerant cultivars show more turgidity than that of the susceptible, which was evident in grapevine (Barkousi et al., 1979) and brinjal (Panchalingam, 1983).RWC is considered as an important measure of plant water status since, it reflects on the metabolic activity of the tissues and lethal leaf water stress, particularly at hot summer (Flower and Ludlow, 1986).Sunflower cv.Ida when grown under water deficit conditions showed a reduction of 12% in relative water content (Izzo et al., 1992).Reddy and Krishnasastry (1984) while analyzing the RWC of safflower under moisture stress, observed a drastic reduction in RWC of crops under stress.The increased RWC of M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC) may also be due to reduced water loss, continued water availability and continued water uptake under trickle irrigation system.The treatment combination comprising 100% WRc through drip (irrigation + 50% FYM + 50% VC (M 2 S 4 ) recorded the highest fruit yield.Better and utilization of nutrients and water may be the possible reason for the promotary effects.Roots can easily translocate absorbed water from the soil where available soil moisture content was optimum at 100% WRc through drip irrigation.
Required energy for water absorption was very less under this treatment and ultimately led to easy energy translocation to the reproductive parts.Application of farmyard manure and vermicompost had the soil organic matter content and improved the soil structure and biological activity.
This would have reduced the loss of nitrogen by increased cation and anion exchange capacities in soil thereby enhancing the fruit development and yield.Further, by improving the structure of the soil by more aggregation, water holding capacity and air permeability were increased.
These comprehensive changes in soil might have improved the fruit development and number in turn fruit yield.The neutral to mildly acidic pH will be of highly useful in potential utilization of available macro and micro nutrients, which chiefly interrupt the major plant physiological processes viz., photosynthesis and partitioning of photosynthates from source to sink.This might have ultimately increased the yield of fruits.Further, more humic substances present in farmyard manure could have mobilized the reserve food materials to the sink through increased activity of hydrolysing and oxidising enzymes.This combined application of farmyard manure and vermicompost would have helped in better availability and utilization of nutrients.All these scavenging effects might have made quick mobilization and availability of nutrients, which would have aided better plant growth and development.This in turn could have assisted for higher fruit yield of noni.Presence of humic substances in farmyard manure improves the soil tilth as a result of favourable physical conditions of soil viz., water holding capacity and aeration.This might have enhanced the yield of fruits.
Farmyard manure contains humic acid, fulvic acid and humin.These organic acids would have stimulated photosynthetic organelles namely chloroplast more particularly on number of grana.The higher photosynthetic activity would have enhanced the yield.Another reason which can be attributed may be the increase in cell wall permeability resulting in more entry of essential elements into plants, thereby activating photosynthetic apparatus giving rise to enhanced formation of carbohydrates and proteins.This inturn would have increased the photosynthetic rate.The presence of humic substances in farmyard manure was the additional source of polyphenols that might have acted as respiratory catalysts, which inturn increased the rate of respiration and metabolic activity of the noni plants.This might have triggered higher photosynthetic rate and inturn fruit yield.Farmyard manure may form chelates with organic compounds during decomposition that might have increased the availability of micronutrients to plants.These micronutrients may involve electron transport system in photosynthesis and respiration process, thereby resulting in enhanced photosynthetic rate (Srivastava, 1996).
As in any other plant, leaves of noni are chief functional photosynthetic units.For normal production, a noni crop needs sufficient number of leaves at different growth phases.In that situation sustaining leaf production in any crop depends again on water and nutrient availability.The results of the present study also revealed a positive association between total number of leaves produced and the yield of the crop which is maximum in the treatment M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC).Several studies indicated the positive influence on production of more number of leaves from planting to various crop growth phases due to the incorporation of organic manures along with biofertilizers (Vidhya, 2004).Greater accumulation of dry matter conferred greater ability to give higher yield.This was obviously due to vigorous growth of plant.The results of the present study indicated more number of leaves, increased the photosynthetic activity resulting in higher accumulation of carbohydrates.Relatively higher level of carbohydrates could have promoted the growth rate and inturn increased the fruit yield.This was in accordance with the result of Padmapriya (2004) andVanilarasu (2011).
The increase in yield was due to the improved performance of all crop growth and yield attributing characters due to better availability of soil moisture and plant nutrients throughout the crop growth period under drip combined organic farming system.Drip irrigation maintains the soil moisture around the field capacity between two irrigation intervals.On the other hand, check basin method has high fluctuation of moisture between field capacity and permanent wilting point.This might have resulted in lower fruit yield in noni under conventional check basin method of irrigation.These results are in confirmation with the findings of Aladakatti et al. (2012) and Behera et al. (2012).
In this study, the low yield was obtained in the treatment M 4 S 8 (check basin method of irrigation + no manure and no fertilizers) this might be probably due to the low availability moisture and nutrients resulting in production of less number of leaves ultimately less photosynthates which inturn reflects low yield.The check basin method of irrigation had high fluctuation of moisture between field capacity and permanent wilting point.Thus, this result to lower fruit yield.These results corroborate with the findings of Behera et al. (2012).The reduced moisture availability in M 4 S 8 (check basin method of irrigation + no manure and no fertilizers) creates water stress condition.As water stress increases, CO 2 assimilation per unit leaf area decreases (Acevedo et al., 1971).When soil moisture stress intensifies, photosynthesis gets restricted to few hours and peak rate reduces.As a result biomass accumulations become slower (Suanez et al. 1989).This inhibition of CO 2 assimilation by water stress is closely linked to the extent of closure of stomata in the leaf.In addition to that, the metabolic capacity of the cells for photosynthesis is also reduced in general (Hsiao, 1993).Studies have indicated that electron transport and photophosphorylation may be inhibited as a result of reduction in cell water content (Leebot et al. 1998).

Conclusion
Water stress generate active oxygen species which are extremely reactive and cytotoxic and it can affect the respiratory activity in mitochondria, which can cause pigment break down and thereby less of carbon fixing capacity of chloroplasts (Scandalios, 1993).As result, there is reduction in fruit yield.
The treatment combination of M 2 S 4 (100% WRc through drip irrigation + 50% FYM + 50% VC) exhibited the highest yield through improved leaf area, light transmission ratio, chlorophyll stability index and relative water content.
a. Statistical design: Split plot design b.Factors: 2 c.Replications: 2 d.Spacing: 3.6 m x 3.6 m e. Number of plants per replication: 5 WRc = CPE x Kp x Kc x A x Wp lit plant -1 Where WRc = Computed water requirement (lit plant -1 ) CPE = Cumulative Pan Evaporation for two days (mm) Kp = Pan Co-efficient (0.75) Kc = Crop factor (0.90 for vegetative stage, 0.95 for flowering and harvesting stage) (Allen et al., 1998) A = Area occupied by the noni tree (3.6 m x 3.6 m) Wp = Wetting percentage (40)

Table 1 .
Initial soil chemical and physico-chemical characteristics of the experimental field.

Table 2 .
Nutrient content of organic manures.

Table 3 .
Quantity of water used during the study period.

Table 5 .
Effect of different water regimes and organic manures on leaf area per plant (cm 2 ) at vegetative and flowering stage.

Table 6 .
Effect of different water regimes and organic manures on leaf area per plant (cm 2 ) at harvesting stage.

Table 7 .
Effect of different water regimes and organic manures on light transmission ratio of noni leaves.

Table 8 .
Effect of different water regimes and organic manures on chlorophyll stability index (per cent) of noni leaves.

Table 9 .
Effect of different water regimes and organic manures on relative water content (per cent) of noni leaves.

Table 10 .
Effect of different water regimes and organic manures on noni fruit yield per hectare (tones).