Abbreviation:

IAA; Indole acetic acid; KIN: Kinetin; Chl: Chlorophyll

1. Introduction

Pigeon pea (Cajanus caja. L) is a perennial legume from the family Fabaceae. It is common food in Asia, Africa and Latin America. It is also regarded as fast growing, hardy, widely adaptable, and drought resistant [1]. Cajanus caja either originate from North-Eastern Africa or India and its cultivation dates back at least 3000 years. It is regarded as a pantropical and subtropical species particularly appropriate for rainfed agriculture in semi-arid areas due to its deep taproot, heat tolerance and fast growing habit (and Mallikarjuna et al., 2011). It form an important food staple in many traditional diets that can improve public health and nutrition (Serrano et al., 2012). The seeds are valued worldwide as an inexpensive meat alternative and are considered the second most important food source after cereals (Kouris and Belska, 2016). The consumption rate surpasses production rate for some common legumes such as cowpea and soybean, so there is need to direct attention to the growth and development of various underutilized legumes, to increase the yield, utilization and production. However, several factors contribute to the limited use of these legumes. These include the low yield, poor seed availability and lack of marketingin addition to presence of anti-nutrients, bloating and flatulence which are particular to legumes as well as hard-to-cook phenomenon. Therefore there is need to improve upon these limitations to increase the production both at local and commercial farming hence this study on the effects of Indole acetic acid and Kinetin on physiological and biochemical properties of igeon pea. Indole acetic acid and Kinetin belong to classes of plant growth hormone that have been shown to regulate and promote plant growth and development. Indole acetic acid is a natural plant growth hormone belong to classes called auxin while kinetin is a type of synthetic plant growth hormone belong to class cytokinnin.

2. Material and Method

2.1 Seed materials

Two cultivars of Pigeon pea seed (CAJ324-15 & CYJ728-42) were obtained from Institute of Agricultural Research and Training Ibadan and were sorted out.

2.2 Preparation of IAA and Kinetin

10mm and 20mm of each of IAA and Kinetin was prepared according to the methods of R. Coolbear. And W. Heydecker. (1997) with slight modification.

2.3 Seed /Plant Treatment

For pre-treatment of the seed, each of the two cultivars of the seed was treated with 10mm & 20mm of IAA and Kinetin respectively in a clean container. Distilled water was used instead of hormones for other part of the seeds which represent control while for post-treatment, foliar method was done 4weeks after planting and this was repeated twice until maturity.

2.4 Estimation of Photosynthetic Pigments

The photosynthetic pigments estimated (chlorophyll a, chlorophyll b and carotenoids) were estimated spectrophotometric according to the method of (Ricardson et al., 2002). The concentrations of chlorophyll a, chlorophyll b, and total Carotenoids were determined using Arnon's equation (1949).

Chlorophyll a = 10.3 E₆₆₃-0.918E₆₄₄ = mg/ml

Chlorophyll b = 19.7E₆₄₄-3.870E₆₆₃ = mg/ml

Carotenoids = 4.2 E₄₅₂-(0.0264 chlorophyll a + 0.426 chlorophyll b) = mg/ml

2.5 Estimation of Physiological Parameters

Fresh weight, Dry weight, Number of seed/Pod, Percentage germination and Plant height was determined according to modified methods of Singh and Gosh (2010).

3. Estimation of Phytohormones

3.1 Extraction of phytohormones

 IAA and Kinetin was extracted from the fresh plant according to the method of Marrillia et al., 2014 using 1mlof methanol and water (8:2).

3.2 Determination of phytohormone concentration

Concentration of phytohormones was determined according to the modified method of Parth et al., 2014. Standard each of IAA and Kinetin was prepared in 0.01ppm, 0.1ppm, 1ppm, 10ppm and 100ppm. The concentration of IAA and Kinetin was measured using UV spectrophotometer at absorbance of 254nm and 535nm respectively and result was calculated from calibration curve.

3.3 Statistical Analysis

Data were analysed using statistical software (SAS). Means were separated using Duncan Multiple Range Test at P<0.05 level of significance.

4. Results

Table 1: Effect of IAA and Kinetin on Photosynthetic Pigments of CAJ324-15

Treatment mg/L	Chl a mg/L	Chl b mg/L	Car mg/L
Control	1.88 ± 0.003b	0.12 ± 0.002gh	0.30 ± 0.001e
IAA 10mm Post	0.82 ± 0.002g	0.20±0.002f	0.11 ± 0.002h
IAA 10mm Pre	1.15 ± 0.002e	0.69±0.011a	0.48 ± 0.003c
IAA 20mm Pre	0.77 ± 0.001h	0.26±0.006e	0.13 ± 0.002g
IAA 20mm Post	0.66 ± 0.001i	0.14±0.002g	0.09 ± 0.002i
KIN 10mm Post	1.63 ±0.003c	0.42±0.001c	0.59 ± 0.003b
KIN 10mm Pre	2.57 ±0.003a	0.30±0.005d	0.99 ±0.002a
KIN 20mm Post	1.37 ±0.002d	0.11±0.003h	0.17 ±0.001f
KIN 20mm Pre	0.90 ±0.001f	0.63±0.002b	0.39 ±0.002d

Values are Mean±standard error, Means with the same letter are not significantly different from each other

Table 2: Effect of IAA and Kinetin on Growth Parameters of CAJ324-15

Treatment mg/L	Number of seed/Pod	Plant height(cm)	Fresh weight(g)	Dry weight(g)
Control	101.00±0.577b	96.67 ± 1.202b	0.67±0.012c	0.26 ± 0.006bc
IAA 10mm Post	50.67±0.667e	56.00 ± 0.577e	0.43 ±0.006f	0.18 ± 0.006e
IAA 10mm Pre	84.00±0.577c	77.00 ± 0.577c	0.56 ±0.003d	0.15 ± 0.006f
IAA 20mm Pre	20.00±1.000h	31.50 ± 0.500f	0.33± 0.010h	0.23 ± 0.015d
IAA 20mm Post	46.50±0.500f	57.50 ± 0.500e	0.37±0.010g	0.24 ± 0.010cd
KIN 10mm Post	65.00±1.000d	121.00 ±1.000a	0.72 ± 0.010b	0.36 ± 0.015a
KIN 10mm Pre	161.00±1.000a	120.50 ± 0.500a	0.89 ± 0.005a	0.28 ± 0.005b
KIN 20mm Post	52.33±1.453e	66.00 ± 0.577d	0.53 ± 0.009e	0.25 ± 0.003bc
KIN 20mm Pre	44.33±0.667g	64.00 ±0.577d	0.52± 0.006e	0.22 ± 0.009d

Values are Mean±standard error, Means with the same letter are not significantly different from each other

Table 3: Effect of IAA and Kinetin on Photosynthetic Pigments of CYJ728-42

Treatment mg/L	Chl a (mg/L)	Chl b (mg/L)	Car (mg/L)
Control	1.12 ± 0.002e	0.01 ± 0.002k	0.07 ± 0.001i
IAA 10mm Post	0.92 ± 0.002h	0.12 ± 0.001j	0.14 ± 0.002h
IAA 10mm Pre	0.92 ± 0.001g	0.76 ± 0.002d	0.56 ± 0.001b
IAA 20mm Pre	0.95 ± 0.001f	0.42 ± 0.001e	0.26 ± 0.001e
IAA 20mm Post	1.27 ± 0.002c	0.16 ± 0.001i	0.18 ± 0.002g
KIN 10mm Post	1.64 ± 0.002a	0.21± 0.002f	0.27 ± 0.001d
KIN 10mm Pre	1.23 ± 0.002d	0.92 ± 0.003b	0.71 ± 0.001a
KIN 20mm Post	1.45 ± 0.002b	0.17 ± 0.002h	0.21± 0.002f
KIN 20mm Pre	0.89 ± 0.002i	0.85 ± 0.001c	0.53 ± 0.006c

Values are Mean±standard error, Means with the same letter are not significantly different from each other

Table 4: Effect of IAA and Kinetin on Growth Parameters of CYJ728-42

Treatment mg/L	Number of seed/pod	Plantheight(cm)	Fresh Weight(g)	Dry Weight(g)
Control	42.00 ±0.580f	68.00 ± 0.58d	0.71 ± 0.012b	0.17 ± 0.009c
IAA 10mm Post	40.33±0.880fg	51.33 ± 0.33 g	0.48 ± 0.006 e	0.13 ± 0.006de
IAA 10mm Pre	78.33±0.333b	87.67 ± 0.33b	0.52 ± 0.006d	0.18 ± 0.006c
IAA 20mm Pre	86.50±0.500a	68.50 ± 0.50d	0.23 ± 0.010h	0.13 ± 0.015e
IAA 20mm Post	48.50±0.500cd	62.50 ± 0.50e	0.27 ± 0.010g	0.19 ± 0.005c
KIN 10mm Post	46.50± 0.500de	57.50 ± 0.50f	0.77 ± 0.015a	0.37 ± 0.010a
KIN 10mm Pre	38.50 ± 0.500g	75.50 ± 0.50c	0.79 ± 0.005a	0.18 ± 0.005c
KIN 20mm Post	40.00 ± 0.577fg	47.00 ± 0.58h	0.61 ± 0.012 c	0.22 ± 0.007b
KIN 20mm Pre	46.00±0.577e	96.00 ± 0.58a	0.42 ± 0.006f	0.22 ± 0.009b

Values are Mean±standard error, Means with the same letter are not significantly different from each other

4. Discussion

Table 1 and 2 shows that there is significant difference in the level of Chl a, Chl b and carotenoid, Number of seed/pod, Plant height, Fresh weight and Dry weight for all the treatment applied as compared with control except treatment with KIN 20mm Post for CAJ324-15 which show no significant differences as compared with the control. Also, among the IAA and KIN foliar method and pre-treatment, IAA 10mm Pre and KIN 10mm Pre show highest mean value for chl a. b and carotenoid respectively. Also for CYJ728-42, Table 3 and Table 4 show that only IAA 20mm Post, KIN 10mm Post, KIN 10mm Pre and KIN 20mm Post show significance difference with respect to control for Chl a, while IAA, KIN for both 10mm and 20mm Pre & Post only show significance difference with respect to control for Chl b and carotenoids while only IAA 10mm & 20mm Pre, 20mmPost, KIN 10mm Post, KIN 20mmPre is significant different with respect to control for Number of seed/pod, only IAA 10mm Pre, KIN10mm Pre & KIN 20mm Pre show significance difference with respect to control for plant height, KIN 10mm Pre & Post only show significance difference with respect to control for Fresh weight, KIN 10mm Post, KIN 20mm Pre & Post only show significance difference with respect to control for dry weight. This means these concentration is most suitable for higher growth and production hence accelerating the nutrients uptake that is responsible chlorophyll formation. In addition, Figure 1 and 2 shows that there is significant difference for both IAA and KIN concentration in all the treatment applied as compared to the control except for IAA 10mm Pre which show no significant difference as compared with the control. The highest mean value for IAA and KIN concentration wasfound in treatment IAA 10mm Post and KIN 20mm Pre respectively. Moreover, Figure 3 and 4 show that only IAA 10mm and 20mm for both Pre and Post show significance difference with respect to control for IAA treated group while for KIN treated group, only KIN 10mm and 20mm Pre & Post show significance difference with respect to control. This was as a result of their inductive effects which therefore upregulate the level of each endogenous phytohormone respectively. Also the implication of the upregulation was observed in their physiological and biochemical composition which is an indication of positive response to both pre and post-treatments this was also in accordance with the work of Mukhtar 2008 on which reported that 100ppm IAA and 100ppm GA₃ improve growth and nutritional value of Hibiscus sabdariffa L. (Red sorrel). Mona et al., 2013 also reported the promoting effect of foliar application of IAA and NAA on physiological and biochemical parameters of barley.

However, despite these plant growth hormones (indole acetic acid and kinetin) induce and accelerate growth in plant via different mechanism for instance IAA which is an endogenous or natural hormone serve to promote root formation and kinetin, an example of synthetic hormones serve to promote shoot elongation, it must be emphasized they conforms to general mechanisms of bioregulators in which there is promoting effects at low concentration but inhibitory effects at high concentration (El-al and Faten, 2009).

5. Conclusion

The present work show that the co application and pre-treatment of Pigeon pea with indole acetic acid and kinetin could promote both physiological and biochemical properties hence resulting to enhanced growth and productivity. However, this study show that response of pigeon pea to either pre- and post-treatment of indole acetic acid and kinetin solely depend on concentration applied and cultivars of the seed.

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