National seminar: Environmental Challenges: Sustainable development Proceeding: Environmental Challenges: Sustainable development; March 29-30, (2010), M.D.

University, Rohtak, pp: 326-331

Effect of sewage effluent on protein content during seed germination in Pigeon pea (Cajanus cajan) plant
Ajit Singh, J.S. Laura, Anju Rana, Jyoti Rana Department of Environmental Sciences, M.D. University Rohtak (Haryana) Corresponding authors: jslmdu@gmail.com Abstract Seed germination represents a crucial phase in the life cycle of angiosperms. With seed independence of next generation of plants begins. Sewage is used for the irrigation of crops in field around most cities .Analysis of the sewage revealed presence of heavy metals which are known to suppress seed germination. Hence in the present investigation, the effect of sewage on protein content was found to be significantly affected by higher concentration of sewage effluent due to unacceptable range of heavy metals in sewage effluent. It was found that protein content in the cotyledons decrease day by day after imbibitions. But high decrease was observed in 100% sewage application as compare to control and 50% sewage treatment. On the other hand in embryonic stage increase was observed in protein content in control and 50% conditions but decrease was found in 100% condition. Protein was considerably suppressed particularly during initial stages of seed germination, interestingly as compare to control higher protein content got depleted form cotyledons germinated in sewage. Key words: Protein, germination, sewage effluent, cotyledons and embryonic axis. Introduction Over the last few decades due to rapid urbanization and industrialization coupled with an ever increasing Sewage serves as cheap source of plant nutrients. But Sewerage water contains a large variety of wastes ranging from domestic to industrial; therefore, quality of such water is not suitable to irrigate any crop because of presence of many toxic chemicals (Furedy et al., 1999; Zarsky & Hunter, 1999; Murtaza et al., 2003; Ghafoor et al., 2004). Sewage application 326 although increase crop

population as resulted in production of huge amount of sewage. Treatment of sewage requiring a large sum of money hence sewage is generally discharged without any treatment directly onto the river, canal and agricultural field.

National seminar: Environmental Challenges: Sustainable development production because it contains has nutritive value and a cheaper source of protein. Several enzyme activities and other metabolic processes are known to be disturbed in plant by the presence of high concentration of toxic contaminants present in sewage (Hemalatha et al., 1996, Neelam and Jaganmohase, 2003).

agronomically important plant nutrients such as N, P, K, Zn, Mn, Cu, Fe. But with these it also contains hazardous heavy metals such as Cd2+, Pb2+, Al2+, Ni2+ etc ( Gupta et al., 1996, Sancherz et al., 1999). Thus the uncontrolled use of sewage water for irrigation can result in accumulation of some potentially toxic metals in soil and plants and cause adverse effects (Gupta et al., 1999). In India total sewage produced annually from cities is about 2600 mm3 (Paroda, 1999). Out of which 70% untreated are applied on agricultural land and 30% discharge after treatment. Haryana produces raw sewage 485 million liter/day. It is estimated that it has the potential to contribute about 56.4 tones/day and 20.872 tones/day

Material and Methods In the present study the seeds were collected form Hisar Agricultural University Krishi Vigyan Kendra,

Rohtak. Sewage water samples were collected from different main sewage discharge site of Rohtak in clean plastic bottles. Concentrated Nitric acid @ 5 ml was added at the time of sample collection to avoid adsorption of heavy metals on wall of bottles and to preserve it. Electric conductivity, pH were

cumulatively of all nutrients. Seed is the beginning point of the next generation. Seed contains the food reserve in the form of carbohydrates, proteins and fats in more concentration than in plants. In present investigation the effect of sewage application on protein content in Pigeon Pea plant has been studied. Pigeon Pea is the most important pulse crop grown in India both for vegetable and dry seed purpose. It 327

estimated in unacidified sample. All other parameters were analyzed by Standard methods given by APHA (1979). Heavy metals were estimated as such after filtering by digestion 100 ml of sample with diacid digestion and then determined by AAS.

Protein content The study was conducted for the observation of protein content in the

National seminar: Environmental Challenges: Sustainable development form of total protein, soluble protein and insoluble protein in cotyledons and in embryonic axis. The experiment was conducted in Petri plates. Marbach and Mayer (1976) was adapted for extraction of soluble protein and insoluble protein from finely ground samples. Soluble protein was extracted by thoroughly macerating 20 mg of oven dried sample in mortar and pestle in 0.1 M phosphate buffer (pH 7.3). The homogenate was decanted the supernatant and pellet was washed with same buffer and Result and Discussion The characteristics of physicochemical sewage water in alkaline supernatant were

quantitatively estimated. Both soluble and insoluble proteins extracted as descried above were determined by Lowry et al., (1951). Then protein content was determined by a standard curve prepared using graded

concentration (0.200 g) of Bovine serum albumin.

recentrifuged. Supernatant obtained were pulled and used for estimation of soluble protein. The extracted protein in

presented in Table1 indicate that pH, EC and BOD were within permissible limit. Heavy metals in sewage are given in Table 1. Except Zinc all heavy metals were found out of permissible range when compared with drinking water standards given by ICMR.

supernatant is soluble proteins. The extracted protein in supernatant fraction was precipitate by addition of TCA to a final concentration 7.5% (W/v) and then collected by centrifugation of 3,000 g for 10 min. The precipitates were dissolved in 5 ml of 0.1M NaOH for quantitative estimation of soluble

Protein content During the study the protein content was found to be dependant on the concentration of sewage. In early stages of seed germination protein content decrease from day 4-8 and highest decrease was found in seed with 100% application. As on day 4 the total protein content was 10.78, 18.19, 16.23 mg protein/pair cotyledons respectively 328

proteins. To the pellets which were left over after extraction of soluble protein with buffer, 5 ml of 0.1M NaOH was added and then the tubes were kept in boiling water both for 15 min. After centrifugation of 1,000Xg for 10 min the insoluble proteins which were recovered

National seminar: Environmental Challenges: Sustainable development in control, 50%, 100% sewage diminished capacity of embryonic axis to synthesize proteins.

application. But decrease on day 8 i.e. 7.25, 10.89, 8.41 mg protein/pair

cotyledons as shown in Table 2. On embryonic stage total protein content increase from day 4-8 in control and 50% sewage application but decrease in 100% sewage treatment. As on day 4 the total protein content was 1.70, 1.52 and 1.25 mg protein/embryonic axis

Conclusion The present investigation showed that the sewage concentration inhibit the protein synthesis. It was conclude that the protein synthesize in seed in present study is dose dependent. In 100% sewage treatment the protein content was found to be lower than control and 50% sewage treatment in both the early and later stages of seed germination. At later stages (4-8 days) of germination of rate of disappearance of proteins from cotyledons of seeds in sewage exceeded than in control. of seed An impeded was

respectively in control, 50% and 100% sewage application and on day 8 was 2.58, 1.82 and 1.10 mg

protein/embryonic axis as shown in Table 2. Same trend was found in case of soluble and insoluble protein as shown in Table 3 and Table 4

respectively. So the protein content varies due to sewage concentration. The sewage effluent due to high

mobilization

protein

accompanied by a reduction in free amino acids which in turn likely to affect denovo synthesis and of proteins axis in of

concentration of heavy metals also affect utilization of cotyledonary proteins

cotyledons germinating

embryonic During

particularly during initial stages of germination and this was accompanied by lower amount of free amino acids in cotyledons and embryonic axis.

seeds.

study

germination in seeds was found to be less in effluents and waste waters when compared with control.

Embryonic axis of these seeds with sewage treatment contained much lower quantities of total, soluble and insoluble proteins than controls. This indicates References APHA 1979. Standards methods for the examination of water and waste

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National seminar: Environmental Challenges: Sustainable development water. 14th Ed. American Public Health Association, New York. Chugh, L.K. and Sawhney, 1995. Hemalatha, S., A. Anburoy and K. Francis 1997. Effect of heavy metals on certain biochemical constituents and nitrate reductase activity in Oryza sativa seed. J. Environ. Biol. 18(3): 313-319. Murtaza, G., A. Ghafoor, M. Qadir and M.K. Rashid, 2003. and

Environmental pollution: Effect of Cadmium on germination, amylase and roll of respiration of germinating seeds, 92(1): 1-5. Furedy, C.V., Maclaren and J. Whitney, 1999. Reuse of waste for food production Health Perspectives. in and In: Asian Cities:

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Bioavailability of Cd, Co and Mn in Soils and Vegetables Irrigated with City Effluent. Pakistan J. Agric. Sci., 40: 1824. Neelam, P. and K. Jaganmohase 2003. Differential effect of Cd and Hg on growth and metabolism of S. melongena. J. Environ. Biol. 24(4): 453-460. Paroda, R.S. and Mruthyunjaya (1999). NARS in the Asia Pacific Region: A perspective, of Asia Pacific

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Table 1. Average physico-chemical characteristics of sewage effluent Parameters Odour pH Conductance (ds/m) BOD Cadmium Chromium Nickel Lead Copper Zink (mg/l) (mg/liter) (mg/liter) (mg/liter) (mg/liter) (mg/liter) (mg/liter) Sewage Effluent Odour 7.4 1.7 250 0.065 0.183 0.195 0.595 0.0674 0.238

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National seminar: Environmental Challenges: Sustainable development Table 2. Total Protein Content of Pigeon Pea Seeds Germinated Under Sewage Irrigation
Cotyledons (mg protein/pair cotyledons) Concentration of Sewage 4 Control 50% 100% 10.780.23 18.191.03 16.230.94 No. of Days 8 7.250.14 10.891.23 8.410.54 4 1.700.08 1.520.62 1.250.51 Embryonic axis (mg protein/ pair embryonic axis) No. of Days 8 2.580.87 1.820.71 1.100.92

Table 3. Soluble Protein Content of Pigeon Pea Seeds Germinated Under Sewage Irrigation
Concentration of Sewage Cotyledons (mg protein/ pair cotyledons) No. of Days 4 Control 50% 100% 5.67 10.5 8.90 8 3.30 6.80 4.96 Embryonic axis (mg protein/ pair embryonic axis) No. of Days 4 0.95 0.71 0.56 8 1.20 0.84 0.49

Table 4. Insoluble Protein Content of Pigeon Pea Seeds Germinated Under Sewage Irrigation
Concentration of Sewage Cotyledons (mg protein/ pair cotyledons) No. of Days 4 Control 50% 100% 5.10 7.69 7.13 8 3.95 4.10 3.49 Embryonic axis (mg protein/ pair embryonic axis) No. of Days 4 0.77 0.82 0.69 8 1.35 0.98 0.63

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