Research Journal of Chemical and Environmental Sciences Volume 1 Issue 2 (June 2013): 37-41 Available Online http://www.aelsindia.com/rjces.htm ©2013 AELS, India Online ISSN 2321-1040

ORIGINAL ARTICLE

Effect of Diesel Fuel Contaminated Soil on the Germination and the Growth of Festuca arundinacea Fateme Zarinkamar, Fateme Reypour, Saiedeh Soleimanpour Department of Plant Biology, Faculty of Basic Science, Tarbiat Modares University, Tehran, Iran ABSTRACT Although the toxicity of diesel fuel has decreased in recent years due to the removal of many of the aromatic compounds present in diesel fuel, at certain concentrations, it is still toxic to plants. The effects of soil diesel fuel pollution on the germination and the longitudinal growth of Festuca arundinacea were studied in a greenhouse plot-culture experiment. For this purpose, percentage germination was recorded weekly for 6 weeks after seed cultivation. Inhibition of germination increased with increasing diesel fuel concentration. Also weekly and during 4 weeks after seed germination, length of roots and leaves were measured separately. Usually the inhibitory effect of diesel fuel on seed germination and plant growth was dependent on pollution level. Despite this the obtained results in F. arundinacea suggest its high toleranance. Since a delay in germination time and growth factorsare observed, its seeds can tolerate relatively high concentrations of diesel fuel by germinating and growing in contaminated soils. Key words: Diesel fuel, Festuca arundinacea, growth, germination Received 21/04/2013 Accepted 12/05/2013

© 2013 Academy for Environment and Life Sciences, India

INTRODUCTION Petroleum hydrocarbons are a mixture of chemical substances which are produced during human activities by using petroleum products in industrial and agricultural production. Although petroleum is one of the dominant energy sources to maintain the economic and social development of a country, during the exploration, translation and processing, petroleum has caused a huge area of contamination and relevant adverse effects, and many countries and regions face serious problems of soil contamination and ecological risk by petroleum hydrocarbons [12, 11, 20]. Changes in soil properties due to contamination with petroleum-derived substances can lead to water and oxygen deficits as well as to shortage of available forms of nitrogen and phosphorus [17]. So the presence of petroleum hydrocarbons in soils is not only an adverse factor for human health, but also a negative impetus for plant growth and development [9, 14]. Their harmful effects include inhibition of seed germination, reduction of photosynthetic pigments; slow down of nutrient assimilation and shortening of roots and aerial organs. It is also expected that some fractions of petroleum can dissolve biological membranes and as a consequence, disrupt the plant root architecture [21]. In General petroleum derivatives e.g. diesel fuels, are phytotoxic to plants at relatively low concentrations. At levels below this phytotoxic level, the development of plants grown in diesel fuel contaminated soil differs greatly from plants grown in normal soil conditions [2]. Germination and root elongation are two critical stages in plant development that are sensitive to environmental contaminants. Plants that are able to germinate successfully amidst the contaminant and show root elongation are tolerant plants [18] and can be useful for phytoremediation. Phytoremediation is the green technology that uses plants to remediate contaminated soil, sediment and surface water. It is a cost-effective, ecologically compatible tool for the environmental clean-up of a wide array of contaminants such as petroleum hydrocarbons [4, 8, 15, 22, 23]. Little is known about plants suitable for the phytoremediation of different pollutants in soils. Studies have shown that the plants that are suitable for remediation of a particular contaminant must be that can tolerate the contaminant [16]. Grasses are sometimes used to remediate chemically contaminated soils, because their root systems are extensive and fibrous [18]. The fibrous root system forms a continuous, dense rhizosphere, which provides ideal conditions for phytoremediation [1, 10]. One commonly used plant for phytoremediation is Tall fescue (Festuca arundinacea), a robust, broad-leaf grass [5, 6, 7, 13, 15, 19]. The genus Festuca is a perennial herb and belongs to Geramineae (Poaceae) family.

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MATERIALS AND METHODS Plant material and experimental treatments: Seeds of Tall fescue (Festuca arundinacea) from ‘Seed and Plant Improvement Institute’, Karaj, Iran were purchased. The greenhouse plot-culture experiment was conducted to investigate the remediation effectiveness of petroleum contaminated soil by F. arundinacea, and carried out in the greenhouse of Tarbiat Modares university of Tehran, Iran. The experiment was arranged in three different levels of contamination (10000, 20000, 30000 ppm) and uncontaminated soil (concentration of diesel fuel was zero) was used as the control sample. After achieving a relatively homogenous mixture of soil samples contaminated with different concentrations of diesel fuel, about one kilogram soil was transferred per each PVC pot. Then seeds of Festuca were scaled and about two grams of them were cultivated in each pot and covered with thin soil layers. Each treatment was replicated three times. The gaps between the pots were 10 cm or so. Lighting for plant growth was natural sunlight with a light/dark cycle of approximately 14/10 h. the temperature in the greenhouse was 12–20 ◦C with 20-45% air humidity. The plants were watered every other day (or as needed). The number of germinated seeds was recorded weekly for 6 weeks after exposure to different concentrations of pollutant. Then the percentage germination in each week was calculated and the influence of diesel fuel on this parameter was measured. Also weekly and during 4 weeks after seed germination, the plants were harvested, the roots and leaves were separated and longitudinal growth of each of them was measured. Statistical analysis: Statistical analysis in this research by usage of factorial analysis and in a CRD (completely randomized design) was performed. For each of treatments three repetitions were considered. Analysis of the data by SAS 9.1 software and comparison of means by Duncan test was performed. The significance of differences among treatment means were compared by Fisher’s least–significant difference test (LSD) at P < 0.05. RESULTS AND DISCUSSION Percentage germination: The percentage germination of the seeds were affected by the quantity of diesel fuel added to the soil and decreased slightly as the level of pollution increased, nevertheless the process of seed germination was never stopped. The obtained charts from results of all six weeks were approximately as the same shapes. Percentage germination of Fescue plants were shown in Fig. 1. In this picture for brevity, just 3 charts were shown which belong to percentage germination in first, third and sixth weeks after seed cultivation. The results showed reduced seed germination in the polluted conditions than control seeds. Although this percentage did not reach zero. In other words, despite the presence of oil pollution in soil and because of Festuca seed′s high resistance, seeds were still able to germinate in polluted conditions. But smaller number of polluted condition's seeds could enter germination stage in a specified time. For example in control samples, in the first week after seed cultivation, 40% from them entered the stage of germination. Whilst in 10000 and 20000 ppm of diesel fuel contained treatments, seed germination around 40%, with delay and three weeks after seed cultivation was seen. Even thought at the highest treatment, even after six weeks, this percentage was not obtained; so that at the end of sixth week, just 37.67 % from these seeds germinated. While in this time (sixth week), the control seeds showed 67% germination; that it was over 1.7 times more than the percentage germination in the most polluted treatment (Fig. 1). In most cases, F. arundinacea could tolerate the contaminant by germinating successfully in it, but a delay in germination time was observed. A more probable reason for the inhibitory effect of diesel fuel on germination is diesel fuel’s physical water repellent property as Adam and Duncan described in 2002. Diesel fuel on entering the terrestrial environment will not migrate far into the soil profile due to the hydrophobic properties of the fuel. This means diesel fuel will be held in the surface of soil and within the rooting zone of most seed species. Seeds planted into this contaminated soil will become coated with diesel fuel. The embryo of a seed could easily be injured or killed if it were to come in contact with diesel fuel. Seeds have a primary line of defence preventing diesel fuel penetration—their seed coat. The integrity and hardness of the seed coat affects the rate of oil penetration. The film of diesel fuel around the seeds may act as a physical barrier, preventing or reducing both water and oxygen from entering the seeds. This would inhibit the germination response [1, 3]. Morphological studies: Organ elongation of Fescue plants were shown in Fig. 2. In this picture, 4 charts were shown which contained elongation of both of plant’s root and leaf, during 4 weeks after seed germination. Generally, the data presented here indicated that addition of diesel fuel in soil decreased root and leaf elongation in Fescue plants. RJCES Volume 1 [2] June 2013

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Studying of root length changes during 4 weeks represented a reduction in root growth trend for increased soil pollution. Reduction of root length in each of the four weeks, at least in the highest infection compared to the control treatment was significant (Fig. 2). Similar to the root, studying of leaf elongation during 4 weeks indicated a reduced leaf growth trend for increased soil pollution. In each of four weeks there was a significant decline in leaf length for all pollutant-treated Fescue seedlings compared to the control treatments; Except for the plants treated with concentration of 10000 ppm diesel fuel in third week. Also in all of four weeks there was a significant difference in leaf length of plants treated with 10000 and 30000 ppm pollutant (Fig. 2). These results confirmed that presence and increasing in amount of diesel fuel in soil could decrease rate of Festuca growth. By comparing the weeks in Fig. 2, it became clear that despite the presence of contaminant (up to 30000 ppm diesel fuel) in the soil, Fescue seedlings continued to grow over time, as root and leaf significant elongation were observed in the computation. However as mentioned before, with increasing the level of contamination in the soil; organ elongation rate was reduced compared to lower levels of pollution and control treatments. In addition, a comparison on Fescue seedlings’ root and leaf elongation revealed that during two first weeks of treatment, leaves elongation were more than elongation in roots; but in third and forth weeks longitudinal ratio of roots to leaves increased (Fig. 2). This property in addition to the control samples was observed in most of the treatments. This result can be justified with the nutritional needs of plant. At the beginning of seed germination and in order to lead the seedling to become independent from the seed storage source, the plant expands its leaves and consequently its photosynthetic tissues. But when the plant grows, its nutritional needs increase. So for faster and greater absorption of minerals and raw materials needed for photosynthesis, the plant increases the rate of its root elongation.

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Fig. 2: Effect of diesel fuel on root and leaf elongation of F. arundinacea seedlings during 4 weeks after seed germination. Values are mean ± SE (n = 6) and differences between means were compared by Fisher’s least significance test at P < 0.05. Root and leaf graphs in each of the weeks are independent. RJCES Volume 1 [2] June 2013

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CONCLUSION Diesel fuel like the other petroleum products affects the growth and performance of plants. In our research the percentage germination of Festuca seeds were affected by the amount of diesel fuel added to the soil and decreased slightly as the level of pollution increased. In other words, F. arundinacea could tolerate the contaminant by germinating successfully in it, but a delay in germination time was observed. Also in this study it became clear that despite the presence of contaminant (up to 30000 ppm diesel fuel) in the soil, Fescue seedlings continued to grow over the time, but there was a reduction in root and leaf growth trend for increased soil pollution. In addition, this study proposed an environmentally friendly, inexpensive way to remove diesel fuel from polluted soils because plants would provide an efficient method for removal of petroleum contaminants such as diesel fuel from soils which must accompany field experiments. REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21.

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