Tolerance of cultivated and wild plants of different taxonomy to soil contamination by kerosene

Science of the Total Environment 424 (2012) 121–129

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Tolerance of cultivated and wild plants of different taxonomy to soil contamination by kerosene Natalia Sharonova, Irina Breus ⁎ Kazan Federal University, Division of Environmental Chemistry, 18 Kremlevskaja Str., Kazan 420008, Russian Federation

a r t i c l e

i n f o

Article history: Received 12 August 2011 Received in revised form 4 February 2012 Accepted 5 February 2012 Available online 21 March 2012 Keywords: Kerosene Soil contamination Plants Seed germination Soil phytotoxicity Tolerance

a b s t r a c t In laboratory experiments on leached chernozem contaminated by kerosene (1–15 wt.%), germination of 50 plants from 21 families (cultivated and wild, annual and perennial, mono- and dicotyledonous) as affected by kerosene type and concentration and plant features was determined. Tested plants formed three groups: more tolerant, less tolerant, and intolerant, in which relative germination was more than 70%, 30–70% and less than 30%, respectively. As parameters of soil phytotoxicity, effective kerosene concentrations (EC) causing germination depression of 10%, 25% and 50% were determined. EC values depended on the plant species and varied in a wide range of kerosene concentrations: 0.02–7.3% (EC10), 0.05–8.1% (EC25), and 0.2–12.7% (EC50). The reported data on germination in soils contaminated by oil and petrochemicals were generalized. The comparison showed that at very high contamination levels (10 and 15%) kerosene was 1.3–1.6 times more phytotoxic than diesel fuel and 1.3–1.4 times more toxic than crude oil, and at low (1 and 2%) and medium (3 and 5%) levels the toxicity of these contaminants was close differing by a factor of 1.1–1.2. Tolerance of plants to soil contamination had a species-specific nature and, on the average, decreased in the following range of families: Fabaceae (germination decrease of 10–60% as compared to an uncontaminated control) > Brassicaceae (5–70%) > Asteraceae (25–95%) > Poaceae (10–100%). The monocotyledonous species tested were characterized as medium- and low-stable to contamination, whereas representatives of dicotyledonous plants were met in all groups of tolerance. Tested wild plants, contrary to reference data on oil toxicity, were more sensitive to kerosene than cultivated. No correlation was observed between degree of plant tolerance to kerosene and mass of seeds. The evidence indicates factors as structure and properties of testa, structure of germ, type of storage compounds, and type of seed germination (underground or aboveground) are more important. © 2012 Elsevier B.V. All rights reserved.

1. Introduction Petroleum hydrocarbons (PHCs) are constituents of engine fuels, industrial solvents and many other products and are the most widespread among the organic contaminants due to extensive current use of oil and petroleum products throughout the world. They are found in surface soils due to human activities, including spills during extraction, refining and transportation of oil and petroleum products, accidents at chemical and petrochemical enterprises (20–30 million t/year), presence in the atmosphere (50–90 million t/year) as a result of burning petroleum products, etc. (Panin, 2002). After reaching the soil surface and then the aerated soil environment, PHCs degrade very slowly. PHCs accumulated in the upper soil layers significantly transform their physical–chemical and microbiological characteristics and usually depress germination and growth of plants, thus making soils phytotoxic (Salanitro, 2001). They also

⁎ Corresponding author. Tel.: + 7 843 233 73 19; fax: + 7 843 233 74 16. E-mail address: [email protected] (I. Breus). 0048-9697/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.scitotenv.2012.02.009

have direct toxic influence on plants when they contact plant tissues. However, plants respond to PHCs differently. Some of them may resist PHC contamination and some tolerant plants may be useful for cleaning up contaminated soils. Data on the influence of PHCs on plant tolerance are scarce. Furthermore, the results are often inconsistent and poorly systematized (Breus and Larionova, 2006; Newman and Reynolds, 2004) because of differences in experimental conditions, i.e. greenhouse or field studies, different soil type and water regime, plant species and sorts, PHC classes and concentrations, duration of contamination, etc. Germination is the most widespread parameter used for estimating toxicity of PHC contaminated soils (Baud-Grasset et al., 1993; Chaîneau et al., 2003; Gong et al., 2001; Kireeva, 2003; Ogbo et al., 2010; Peng et al., 2009; Petukphov et al., 2000; Wang et al., 2001), plant tolerance to contamination (Gong et al., 2001; Kireeva, 2003; Tesar et al., 2002), and even plant phytoremediation activity (Dorn and Salanitro, 2000; Gaskin et al., 2008; Henner et al., 1999; Kummerova et al., 2008; Maila and Cloete, 2002; Mang et al., 2010). In some cases, germination dynamics is also considered as a characteristic of soil phytotoxicity (Gilyazov and Gaisin, 2003; Kireeva, 2003).

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Around 90 plant species have been investigated to assess how PHC soil contamination affects them, and about 30% of these plants are from the Poaceae family. However, as yet there are no clear data about the differences in plant sensitivity to PHCs at both species and family levels. The effect of soil contamination on germination has been investigated to various degrees for different PHC types. Crude oil, diesel fuel and mineral oil are most frequently studied. Other fuels and individual hydrocarbons have received little attention. Likewise the germination data vary significantly for species and even sorts of plants. At present one of the most typical representatives of petroleum products used quite extensively is kerosene. However, there are only a few available experimental data regarding plant tolerance to soil contamination by kerosene. The problem of soil contamination by PHCs is very acute in the Republic of Tatarstan which is located in the Middle-Volga region of Russia and is one of the largest oil recovery and refinery regions in the country. According to the tentative calculations made during the period from 1943 to 1995, the recovery of 1 t of oil was accompanied by destruction and contamination of 1–1.3 m 3 of soil (Gilyazov and Gaisin, 2003). It is very important to have a clear view of plant resistance to contamination of Tatarstan's soils by PHCs. However, the existing experimental studies are insufficient to draw any meaningful conclusions regarding this issue. Gilyazov and Gaisin (2003) pointed out the inhibitory influence of contamination of leached chernozem by oil on the survivability and productivity of agricultural plants. According to the scale developed by these authors, the following levels of soil contamination can be used when considering the effects of hydrocarbon contamination of leached chernozem on plants: low (less than 3 wt.% PHC), medium (3–6 wt.% PHC), high (6–12 wt.% PHC) and very high (above 12 wt.% PHC). The aims of the present work were: experimental estimation of the tolerance of different species of cultivated and wild plants based on their germination in soil over a wide range of kerosene concentrations, establishment of the phytotoxicity parameters of soil contaminated by kerosene, and comparative analysis of seed germination data for different types of PHC contaminants including kerosene and reference data on crude oil, diesel fuel and individual hydrocarbons.

2.3. Plant species Fifty plants of different taxonomy units were screened. The seeds of plants cultivated in central Russia included 12 families (28 species and 22 sorts) and were obtained from the Scientific and Production Association «Niva Tatarstana», the Research Science Institute TatNIISHOZ, and the Botanical Garden of Kazan State University (Kazan, Tatarstan, Russia). In addition, wild plants from 9 families (16 species) were studied. All plants tested in this study are enumerated and characterized in Table 1. There were 21 kinds of annual and 23 kinds of perennial (including biannual) plants among those investigated. Among the investigated plants, the plants from Poaceae family (Grass family, 8 species), Fabaceae family (Pea family, 6 species) and Asteraceae family (Aster family, 10 species) were best represented. In other families, individual representatives had been investigated. 2.4. Testing facilities Germination was determined according to (Adam and Duncan, 1999, 2002; ISO 11269-2:1995; Kireeva, 2003; The state branch standard, 12038-84, 1984). Experiments were performed in Petri dishes. Undamaged, husked seeds were used in all experiments. Uncontaminated, leached chernozem was used as a control. To obtain an even distribution of a PHC in soil, kerosene was added to the air-dried soil, mixed thoroughly with the soil and then kept in hermeticallysealed vessels for two weeks. Samples of contaminated and uncontaminated soil (50 g each) were placed in the Petri dishes (diameter 10 cm) and moistened to 60% of total moisture capacity. The airdried seeds (batches of 10 seeds in case of maize, pea and sunflower and of 25 seeds in case of all other plants) were then planted. The filled Petri dishes were incubated at 26 °C in the dark. Germination was examined on the 3d, 7th, 10th and 14th day following the start of incubation (Adam and Duncan, 1999, 2002; Kireeva, 2003). When analyzing the results obtained under contaminated soil conditions germination is discussed on a relative basis, as a percentage of the germination in the uncontaminated control. 2.5. Calculation of soil phytotoxicity parameters

2. Materials and methods 2.1. Soil Leached chernozem was the soil used in this study as it is typical for oil contaminated areas in central Russia. Chernozems are the most widespread soil types in Tatarstan. They form 88% of the overall soil fund, and the leached and podzolic chernozems account for 40% of these. Soil samples were taken in the Alexeevsky District of Tatarstan and had 5.9 pH (KCl); humus contents 5.0%; texture: sand 22%, silt 40%, clay 38%; N (alkaly hydrolyzed) 122 mg/kg; P2O5 and K2O (available) 189 mg/kg and 251 mg/kg respectively.

2.2. PHC contaminant Kerosene is a complex mixture of aliphatic and aromatic hydrocarbons (C6-C16). It is one of the main types of fuels used in industry and for transportation purposes in Russia. In our experiments with the contaminated leached chernozem, the kerosene concentrations used were: 1, 2, 3, 5, 10 and 15 wt.% of air-dried soil (10,400, 20,800, 31,200, 52,100, 104,200, and 156,200 mg/kg of dry soil). In accordance with the scale developed by Gilyazov and Gaisin (2003), the kerosene concentrations corresponded to the four levels of soil contamination: low (1 and 2 wt.%), medium (3 and 5 wt.%), high (10 wt.%) and very high (15 wt.%).

From the obtained experimental data, soil phytotoxicity parameters for all tested plants were determined, i.e. the efficient concentrations EC10, EC25, EC50 that cause various levels of plant inhibition — of 10, 25 and 50% respectively (Dorn and Salanitro, 2000; ISO 11269-2:1995; Salanitro, 2001). These parameters were calculated from our experimental data using the approximation equation proposed previously (Larionova et al., 2008). 3. Results 3.1. Influence of different soil kerosene concentrations on seed germination The experimental data obtained on germination for tested plants with different kerosene concentrations are presented in Appendices A–D. The effects for representatives from individual families will be discussed in the context of the contamination level (Gilyazov and Gaisin, 2003). The results show that the toxic effect of kerosene on germination depended on the PHC concentration in the soil. However, for different plant species the magnitude of this effect was different and varied from no change to complete suppression of germination. Soil kerosene concentrations of 1 and 2% (low level contamination) were practically non-toxic for most annual cultivated plants from the Poaceae (Appendix A), Asteraceae (Appendix C), and Brassicaceae (Appendix D) families, as well as annual and perennial cultivated

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Table 1 List of plant names, seed germination in uncontaminated soil, and effective kerosene concentrations (EC) as parameters of soil phytotoxicity. Name of plant species

Poaceae/Grass family (Monocotyledons) Corn (Zea mays L.)a b a b

Common oat (Avena sativa L.) Cereal rye (Secale cereale L.)a b Broomcorn millet (Panicum miliaceum L.)a

b

Timothy (Phleum pratense L.)c b Smooth brome (Bromus inermis Leyss. ssp. inermis var. inermis)c Meadow fescue (Schedonorus pratensis (Huds.) P. Beauv.)c b Sudangrass (Sorghum bicolor (L.) Moench ssp. drummondii (Nees ex Steud.) de Wet & Harlan)a b

b

Name of plant sort (plant number)

Seed germination in uncontaminated soil, %

Mass of 1000 seeds, g

EC, % on dry basis EC10

EC25

EC50

Katerina (1a) Ross-151MV (1b) Los 3 (2) Radon (3) Luchistoe (4a) Kazanskoe kormovoe (4b) Kazanskaya (5) Kazanskyi (6) Kazanskaya (7) Gerkules (8)

100 ± 0 88 ± 6 94 ± 5 90 ± 2 94 ± 6 82 ± 6 82 ± 4 81 ± 3 82 ± 6 54 ± 6

290 210 40 50 7 7 0.5 4 2 33

4.4 2.3 2.7 2.2 2.2 3.3 1.8 1.2 1.2 1.9

4.8 3.2 3.1 3.0 2.6 3.6 2.6 1.7 1.9 2.3

5.4 4.5 3.6 4.0 3.0 3.9 3.8 2.5 3.2 2.9

99 ± 2

7

3.3

3.6

3.9

90 ± 7 74 ± 5 90 ± 6 66 ± 5 76 ± 5 98 ± 3 90 ± 0 75 ± 6 56 ± 5

2 2 2 2 60 270 270 20 6

2.8 3.8 0.4 7.3 4.9 3.3 2.7 3.4 0.6

6.6 8.1 3.5 – 6.0 4.3 3.2 5.9 2.3

–d – – – 7.5 5.6 3.8 10.7 -

Cyperaceae/Sedge family (Monocotyledons) Blister sedge (Carex vesicaria L.)c bW (9) Fabaceae/Pea family (Dicotyledons) Red clover (Trifolium pratense L.)c b Alfalfa (Medicago sativa L. ssp. sativa)c

Rannii-2 (10a) Trio (10b) Muslima (11a) Aisilu (11b)

b

Garden vetch (Vicia sativa L.)a (12)b Garden pea (Pisum sativum L.)a b

Kazanec (13a) Venec (13b) Petushok (14) Gale (15)

c e

Sainfoin (Onobrychis arenaria (Kit.) DC.) Eastern galega (Galega orientalis Lam.)c e

Brassicaceae/Mustard family (Dicotyledons) White mustard (Sinapis alba L.)a b (16) Rape (Brassica napus ssp. oleifera annua Metzg.)a,c b (17) Shepherd's purse (Capsella bursa-pastoris (L.) Medik.)a b W (18)

94 ± 7 52 ± 4 7±2

7 5 0.1

1.0 1.1 1.3

2.4 2.6 1.9

7.3 8.1 2.8

Asteraceae/Aster family (Dicotyledons) Common sunflower (Helianthus annuus L.)a b (19) Golden tickseed (Coreopsis tinctoria Nutt.)a,c b (20) French marigold (Tagetes patula L.)a b (21) Dahlia (Dahlia x cultonim Thorsr. et Reis.)c f Blessed milkthistle (Silybum Marianum (L.) Gaertn.)a,c b W (23) Chicory (Cichorium intybus L.)c b W (24) Common dandelion (Taraxacum officinale F.H. Wigg.)c b W (25) Pot marigold (Calendula officinalis L.)a b W (26) Greater burdock (Arctium lappa L.)c b W (27) Coltsfoot (Tussilago farfara L.)c b W (28)

78 ± 3 68 ± 4 55 ± 4 54 ± 5 94 ± 5 36 ± 6 50 ± 4 88 ± 6 6±3 64 ± 4

80 0.5 3 10 30 1 0.3 10 10 0.2

5.3 2.7 1.6 1.8 1.9 0.3 0.2 1.8 0.02 0.4

7.2 3.9 3.0 2.5 2.5 0.5 0.4 2.3 – 0.7

9.8 6.0 6.0 3.6 3.3 0.7 0.8 2.8 – 1.4

56 ± 5

10

1.1

1.3

1.6

90 ± 3 94 ± 3

30 30

1.9 2.3

2.6 3.2

3.5 4.4

Veselie rebyata (22)

Solanaceae/Potato family (Dicotyledons) Jimsonweed (Datura stramonium L.)a b W (29) Polygonaceae/Buckwheat family (Dicotyledons) Buckwheat (Fagopyrum esculentum Moench)a b

Kamaz (30a) Saulic (30б)

Caryophyllaceae/Pink family (Dicotyledons) Maiden pink (Dianthus deltoides L.)c b W (31) Feathered pink (Dianthus plumarius L.)c b W (32) Siberian pink (Dianthus amurensis Jacq.)c f W (33)

16 ± 5 66 ± 4 74 ± 6

0.2 0.6 0.6

3.9 0.1 0.02

4.2 0.9 0.05

4.3 – 0.2

Apiaceae/Carrot family (Dicotyledons) Dill (Anethum graveolens L.)a b (34)

62 ± 4

2

0.5

1.4

4.8

95 ± 6

2

2.4

7.2

–

0.4

0.7

1.5

Hydrophyllaceae/Waterleaf family (Dicotyledons) Lacy phacelia (Phacelia tanacetifolia Benth.)a b

Ryazanskaya (35)

Rosaceae/Rose family (Dicotyledons) Small burnet (Sanguisorba minor Scop.)c bW (36)

22 ± 3

Linaceae/Flax family (Dicotyledons) Common flax (Linum usitatissimum L.)a

83 ± 6

5

2.2

7.7

–

76 ± 0

5

3.5

5.0

7.3

44 ± 4

0.2

0.2

0.4

1.2

b

(37)

Malvaceae/Mallow family (Dicotyledons) Shoeblackplant (Hibiscus rosa-sinensis L.)c

b

(38)

Plantaginaceae/Plantain family (Dicotyledons) Sand plantain (Plantago psyllium L.)a b W (39)

(continued on next page)

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Table 1 (continued) Name of plant species

Name of plant sort (plant number)

Seed germination in uncontaminated soil, %

Mass of 1000 seeds, g

EC, % on dry basis EC10

EC25

EC50

Chenopodiaceae/Goosefoot family (Dicotyledons) Common beet (Beta vulgaris L.)a,c b (40)

66 ± 3

10

0.9

1.6

2.8

Lamiaceae/Mint family (Dicotyledons) Catnip (Nepeta cataria L.)c b W (41)

34 ± 3

1

1.2

1.8

2.9

Balsaminaceae/Touch-me-not family (Dicotyledons) Spotted snapweed (Impatiens balsamina L.)a b (42)

80 ± 0

Amaranthaceae/Amaranth family (Dicotyledons) Red amaranth (Amaranthus cruentus L.)a b (43) Redroot amaranth (Amaranthus retroflexus L.)a b W (44)

76 ± 5 82 ± 5

0.5 0.5

0.8 0.02

2.5 0.1

12.7 4.1

W — Wild plants. a Annuals. b Plant taxonomy corresponds to http://plants.usda.gov (05.08.2011). c Biennials and perennials. d All studied kerosene concentrations decreased seed germination less than on 50%. e Plant taxonomy corresponds to http://www.fao.org/ag/AGP/AGPC/doc/Gbase (05.08.2011). f Plant taxonomy corresponds to http://flower.onego.ru/lukov/dahlia.html (05.08.2011), http://irapl.altervista.org/botany/main.php (05.08.2011).

plants from the Fabaceae family (Appendix B). The germination of these plants in the contaminated soil decreased by less than 10%. The same effect was observed for some species and sorts of other families: for annual and perennial cultivated plants (buckwheat «Saulic», spotted snapweed, lacy phacelia, common flax, shoeblack plant), as well as perennial wild plants (maiden pink, blister sedge). The most affected group at these kerosene concentrations consisted of plants from 13 families, whose germination was reduced by 10–50% and included sudangrass, eastern galega, alfalfa «Muslima», blessed milkthistle, pot marigold, shepherd's purse, dill, red amaranth, redroot amaranth, buckwheat «Kamaz», common beet, jimsonweed, sand plantain, small burnet, catnip, and feathered pink. For perennial wild plants from the Asteraceae (chicory, common dandelion, greater burdock and coltsfoot) and Caryophyllaceae (Siberian pink) families, the toxic effect of low level contamination was the highest: germination was reduced by 50–100% even at soil kerosene concentrations of 1%. Soil kerosene concentrations of 3 and 5% (medium level contamination) were toxic to all tested plants; on average germination was reduced by 20% and 80%, respectively. The lowest toxic effect was observed for most of the plants from the Fabaceae family, cultivated

species of the Asteraceae family and for individual plants from other families (lacy phacelia, common flax and shoeblack plant). The greatest toxicity was observed for perennial wild plants from the Asteraceae family, as well as for jimsonweed. Soil kerosene concentrations of 10 and 15% (high and very high contamination levels) were highly toxic to most plant species. In this case germination of cultivated plants (Poaceae family; buckwheat «Saulic» and «Kamaz», common beet, garden pea), as well as wild plants (Asteraceae family; jimsonweed, small burnet, catnip) was completely inhibited. The cultivated annual plants from the Fabaceae (garden vetch), Asteraceae (common sunflower, golden tickseed, French marigold) and Brassicaceae (white mustard, rape) families had reductions in germination of between 50 and 95%. The lowest toxic effect at these kerosene concentrations was observed for perennial Fabaceae plants, which retained 45–90% of germination when contaminated. Tested plants were ranked in accordance with their degree of tolerance (Table 2). For each kerosene concentration three groups of plants were defined: highly tolerant (relative germination of 70% and more); mid- and low tolerant (germination of 30–70%), and intolerant (germination less than 30%). The table shows that the

Table 2 Plant grouping according to their tolerance to kerosene in contaminated leached chernozem. Kerosene concentration, % on dry basis (soil contamination rate according to Gilyazov and Gaisin (2003))

Seed germination, % relative to uncontaminated control >70 (more tolerant plantsa)

30–70 (less tolerant plantsa)

b30 (intolerant plantsa)

1% (low)

1aa, 1b, 2, 3, 4a, 4b, 5, 6, 7, 8, 9, 10a, 10b, 11a, 11b, 12, 13a, 13b, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 26, 29, 30a, 30b, 31, 33, 34, 35, 37, 38, 40, 41, 42, 43 1a, 1b, 2, 3, 4a, 4b, 5, 8, 9, 10a, 10b, 11a, 11b, 12, 13a, 13b, 14, 15, 16, 17, 19, 20, 21, 22, 23, 26, 30a, 30b, 31, 33, 34, 35, 37, 38, 40, 41, 42, 43 1a, 1b, 2, 3, 4b, 5, 9, 10a, 10b, 11a, 11b, 12, 13a, 13b, 14, 15, 16, 19, 20, 21, 30a, 30b, 31, 35, 37, 38, 42, 43 10a, 10b, 11a, 11b, 12, 13a, 14, 19, 21, 35, 37, 38, 42

25, 28, 32, 36, 39, 44

24, 27

6, 7, 18, 25, 28, 36, 39, 44

24, 27, 29, 32

4a, 6, 7, 8, 17, 18, 22, 23, 26, 33, 34, 39, 40, 41, 44 1a, 1b, 3, 5, 7, 15, 16, 17, 20, 22, 30b, 33, 34, 39, 40, 43, 44 10a, 10b, 11a, 14, 15, 16, 17, 19, 21, 32, 33, 34, 43, 44

24, 25, 27, 28, 29, 32, 36

2% (low) 3% (average) 5% (average) 10% (high) 15% (very high) a

11b, 35, 37

11a, 11b

Plant numbering corresponds to Table 1.

10a, 10b, 14, 15, 16, 17, 33, 34, 35, 37, 43, 44

2, 4a, 4b, 6, 8, 9, 13b, 18, 23, 24, 25, 26, 27, 28, 29, 30a, 31, 32, 36, 41 1a, 1b, 2, 3, 4a, 4b, 5, 6, 7, 8, 9, 12, 13a, 13b, 18, 20, 22, 23, 24, 25, 26, 27, 28, 29, 30a, 30b, 31, 36, 38, 39, 40, 41 1a, 1b, 2, 3, 4a, 4b, 5, 6, 7, 8, 9, 12, 13a, 13b, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30a, 30b, 31, 32, 36, 38, 39, 40, 41

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The most rapidly sprouting plants were cultivated annual dicotyledonous plants from the Asteraceae (common sunflower, French marigold), Polygonaceae (buckwheat «Saulic») and Balsaminaceae (spotted snapweed) families, a perennial shoeblack plant of the Malvaceae family, as well as monocotyledonous plants from Poaceae (corn «Katerina», an annual, cultivated plant) and Cyperaceae (blister sedge, a wild perennial plant) families. At soil kerosene concentrations of 1–5%, the rate of germination for these plants on the third day of the experiment fell by less than 10% compared to the uncontaminated control. The slowest sprouting seeds were those of red amaranth, smooth brome, meadow fescue, rape, dahlia, chicory, jimsonweed and Siberian pink. Even at 1 and 2% kerosene concentrations, the rate of germination for these plants was reduced by 40– 100%. At 10 and 15% soil kerosene concentrations, the rate of germination for all plants significantly decreased (by 60–100%). The one exception was the sunflower, which was only reduced by 12% at 10% soil kerosene concentration. Generalization of the data for dynamics of germination showed that the best results were demonstrated by cultivated annual plants, mostly dicotyledons, particularly members of the Asteraceae family.

100

3.2. Parameters of soil phytotoxicity

number of plants in each group decreased with increasing soil contamination level. The number of resistant plants was the highest (84% of the total number of plants) at the 1% soil kerosene concentration, decreased to 76% at 2% kerosene, 56% at 3% kerosene, 26% at 5% kerosene, 6% at 10% kerosene, and only 4% at 15% kerosene content. The most tolerant throughout the range of kerosene concentrations were alfalfa, phacelia and common flax and the least tolerant were wild plants, including perennial chicory, greater burdock and feathered pink, and annual jimsonweed. The dynamics of germination are also important for assessing the impacts of PHC contamination, and is considered an additional criterion of plant tolerance during their germination in contaminated soils (Nicolaeva et al., 1999). However, it has been poorly studied under conditions of soil contamination by PHCs. For the majority of plants tested in our experiments, soil contamination by kerosene led to an increase in germination time. Fig. 1 shows three examples for plants with extremely different dynamics.

A

80

60

Calculated values of EC10, EC25 and EC50 depended on the plant species and varied over a wide concentration range: EC10 — in the range of 0.02–7.3%, EC25 — 0.05–8.1%, EC50 — 0.2–12.7% (Table 1). The lowest EC10 values (0.02%) were found for dicotyledonous wild plants (redroot amaranth, Siberian pink and greater burdock). These values were significantly lower than the maximum allowable concentrations of oil and petroleum products in soils (0.1–0.2%) (McGill and Rowell, 1980; Methodological recommendations about revealing of the degraded and polluted soils, 1995). The lowest EC25 values (less than 0.1%) were also noted for redroot amaranth and Siberian pink, and the lowest EC50 values (0.2%) — for Siberian pink. The highest EC50 values (more than 10%) were characteristic for dicotyledonous cultivated plants (sainfoin and common sunflower). For five species of dicotyledonous cultivated plants, EC50 values were not observed in the entire range of investigated kerosene concentrations, even at 15%.

40

4. Discussion

20

It is known that plants are subjected to soil contamination (including soil contamination by PHCs), especially at infancy stages of their lifecycle (Kummerova et al., 2008; Nicolaeva et al., 1999). During germination, PHCs have a direct toxic effect on plants through contact and interaction with seeds, as well as indirect effects by disturbing physical, chemical and microbiological soil properties (Adam and Duncan, 2002; Anoliefo and Vwioko, 1995; Kireeva, 2003; Kummerova et al., 2008). The type and concentration of the PHC and biological features of the particular plant are the baseline factors determining tolerance. The influence of these factors in our experiments on the toxicity of kerosene will be discussed below together with reported data regarding the effects of soil contamination by different PHCs (oil, diesel fuel, aromatic and polyaromatic PHC, kerosene) on germination. It should be noted that the reported data are rather scattered and have been obtained for various PHCs and petrochemicals of different composition, under different soil-climatic conditions (not specified in some studies) and by applying different testing methods. Most studies have focused on soil contamination by aromatic PHCs (primarily polyaromatic), oil and diesel fuel.

60 40 20 0

Seed germination, % relative to uncontaminated soil

125

0

2

4

6

8

10

Time after seeding, days

B 120 100 80

0 0

2

4

6

8

10

8

10

Time after seeding, days

C 120 100 80 60 40 20 0 0

2

4

6

Time after seeding, days 0% KS 5% KS

1% KS 10% KS

2% KS 15% KS

3% KS

Fig. 1. Dynamics of seed germination in leached chernozem contaminated by kerosene.Plants: A — corn, sort “Katerina”, B — lacy phacelia, sort “Ryazanskaya”, C — alfalfa, sort “Aisilu”.Kerosene concentrations: 0, 1, 2, 3, 5, 10, and 15% on dry basis.

4.1. Germination affected by the type of the contaminant It is known that the level of PHC toxicity is determined by the physico-chemical properties of the compounds present. Havis et al.

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(1952) showed that PHC toxicity in relation to germination decreases as follows: aromatic PHCs > naphthenic PHCs > olefins > noncyclic paraffins. Aromatic and polyaromatic PHCs are most toxic and even low concentrations in soil (0.005–0.12%) can inhibit germination by 20–100% (Henner et al., 1999; Kummerova et al., 2008; Maila and Cloete, 2002; Salanitro, 2001; Smith et al., 2006). Oil toxicity has been studied in relation to plants from the Poaceae (27 species and kinds), Fabaceae (eight species and kinds), Brassicaceae (four species and kinds), Pinaceae (two species), Caryophyllaceae (one species) and Salicaceae (one species) families (Amakiri and Onofeghara, 1984; Anoliefo and Vwioko, 1995; Chaîneau et al., 2003; Dorn and Salanitro, 2000; Gilyazov and Gaisin, 2003; Gong et al., 2001; Issoufi et al., 2006; Kireeva, 2003; Nicolotti and Egli, 1998; Petukphov et al., 2000; Salanitro, 2001; Schwendinger, 1968; Shilova, 1988; Wang et al., 2001). It has been shown that the toxicity varies depending on oil composition and concentration in soil. The seed germination at oil concentrations between 0.04 to 20 wt.% is reduced by 30–75% on the average. However, Gilyazov and Gaisin (2003) showed that soil oil concentrations of 0.25 and 0.5% stimulated germination of common oat and common wheat. Regarding oil fractions, the lowest toxic effect, as compared with polyaromatic PHCs and oil, was determined for heavy oil fractions (b.p. 350 °C and above) and petroleum products (fuel oil, distillate and residual oil). These fractions caused a 10–60% decrease in germination (Kireeva, 2003; Morel et al., 1999; Petukphov et al., 2000). Some authors noted very low phytotoxic effects for oil fractions even at high concentrations. Thus, the reduction of germination of cultivated radish (Brassicaceae family) in soil contaminated by tar, cracking residues, distillate of carbonization of heavy gas–oil fraction of cracked residue, and asphaltite in concentrations of 0.5–8% made up only 1.4–1.8% on the third day after contamination (Kireeva, 2003). Light oil fractions (b.p. 40–180 °C) and light petrochemicals (petroleum benzene, gasoline) are highly toxic. It is assumed that their acute toxicity is caused by PHCs, particularly by low-molecular weight PHCs. The latter are relatively water-soluble and can easily penetrate through membranes into plant cells (Gunter et al., 1996; Schwendinger, 1968). They also chemically react with components of plant cells disturbing vital functions. In most cases, this leads to plant death. Therefore contamination of soil with gasoline at concentrations of 0.5% and higher can lead to total inhibition of germination of cultivated radish (Kireeva, 2003), and concentrations of 2% for fuels with high light fraction contents can fully inhibit germination of corn seeds (Salanitro, 2001). However, the toxic effect of light oil fractions is not long-lasting due to their volatility (Panin, 2002). The toxicity of medium oil fractions (b.p. 140–350°C) and of petrochemicals like diesel fuel, naphtha, kerosene and gas–oil is intermediate. These fractions include more than 95% of high-molecular aliphatic PHCs that are essentially insoluble in water and form a mechanical barrier impeding water–air uptake and mineral nutrition for plants in contaminated soils (Kireeva, 2003). The reported data show that at low and average (up to 5%) concentrations of these fractions, germination of different plants decreased from 0 to 50%, and in the case of lettuce and common barley — up to 100% (Denisova et al., 2010, 2011; Kireeva, 2003; Mang et al., 2010; Morel et al., 1999; Ogbo et al., 2010; Salanitro, 2001; Shen and Bartha, 1994; Serrano et al., 2009; Tang et al., 2011; Tesar et al., 2002; Wang et al., 2001). In contrast, soil contamination by diesel fuel (0.5–5%) had the effect of stimulating germination, with increases of 2–27% as compared to an uncontaminated control (Adam and Duncan, 1999, 2002). Concentrations of diesel fuel of 5% or more were more toxic, reducing the germination of Poaceae plants by 36–100% and Fabaceae plants by 11–82%. The most resistant were Brassicaceae plants which retained 95% of their germination in uncontaminated soil (Adam and Duncan, 1999, 2002). Diesel fuel containing 40% aromatic PHCs was 20 times more toxic (Morel et al., 1999).

Reference information related to kerosene is extremely limited. With the exception of our own data, we were only able to find two other experimental studies. It was shown that mixtures of kerosene and water in concentrations of 2.5, 5.0 and 10% totally inhibited germination of cultivated radish, and concentrations of 0.3, 0.625 and 1.25% caused decreases in germination of 70–90% (Noto, 1992). According to Dibble and Bartha (1979), 0.34% kerosene had no influence on germination, but caused a delay in germination of corn seeds. In our previous studies, we investigated the effects of soil contamination by different kerosene concentrations on germination of 15 fodder and wild plants and observed decreased germination between 0 and 95% (Breus and Larionova, 2003; Larionova et al., 2008). In this work, the average reduction of germination for 50 plants of different taxonomy in the range of kerosene concentrations from 1 to 15% varied from 19 to 82%, respectively. 4.2. Germination affected by the concentration of the contaminant It is known that with increasing soil contamination by different types of PHCs, plant germination is usually reduced. Based on the obtained results (Appendices A–D), we calculated average values of relative reduction in germination at different kerosene concentrations for all investigated plants. For kerosene concentrations of 1 and 2%, the average reduction in germination was 19%; at 3 and 5% kerosene it was 48%; and at 10% and 15% kerosene the reductions were 79% and 82%, respectively. These average reductions in germination due to kerosene contamination were compared to corresponding values for diesel fuel and crude oil, calculated by the analogy with kerosene on the basis of reported data for various plants (Adam and Duncan, 1999, 2002; Amakiri and Onofeghara, 1984; Anoliefo and Vwioko, 1995; BaudGrasset et al., 1993; Breus and Larionova, 2006; Chaîneau et al., 2003; Denisova et al., 2010, 2011; Dorn and Salanitro, 2000; Gaskin et al., 2008; Gilyazov and Gaisin, 2003; Gong et al., 2001; Issoufi, 2006; Kireeva, 2003; Mang et al., 2010; Morel et al., 1999; Nicolotti and Egli, 1998; Ogbo et al., 2010; Petukphov et al., 2000; Salanitro, 2001; Schwendinger, 1968; Serrano et al., 2009; Shen and Bartha, 1994; Shilova, 1988; Wang et al., 2001). At low soil contamination levels (less than 3% PHC), average values of germination in soils contaminated by kerosene and diesel fuel were very similar and were more than 80%, and in the case of oil the corresponding value was lower (less than 70%). At medium soil contamination level (3–6% of PHC), the differences in the average germination for kerosene (52% reduction in germination) and diesel fuel (44%) did not exceed 10%, and the toxicity of oil (57% reduction in germination) was slightly lower. The maximum toxicity was observed at high (6–12%) and very high (more than 12%) PHC contamination levels. Under these conditions the values of average germination were 22 and 19%, 30 and 24%, and 35 and 25% for kerosene, oil and diesel fuel, respectively. It is apparent from the data reported here that the decrease in germination due to soil contamination by kerosene was similar to that of diesel fuel and crude oil at similar concentrations. At low and medium contamination levels for these PHCs it differed by the factor of 1.1–1.2. At high contamination levels this difference was slightly larger: kerosene was 1.3–1.6 times more toxic than diesel fuel, and 1.3–1.4 times more toxic than crude oil. The similarity is probably due to the similar chemical composition of these PHCs. It is known that kerosene, diesel fuel and crude oil contain between 40 and 95% normal alkanes, from n-nonane to hexadecane. 4.3. Germination affected by plant characteristics According to some references, plant sensitivity to soil contamination by PHCs has a species-specific nature (Adam and Duncan, 1999, 2002; Peng et al., 2009; Robson et al., 2004). However, the existing

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Seed germination, % relative to uncontaminated soil

120 100 80 60 40 20 0 1

2

3

5

10

15

Kerosene concentration, % on dry basis Poaceae

Fabaceae

Brassicaceae

Asteraceae

Fig. 2. Average seed germination of plants from Poaceae, Fabaceae, Asteraceae and Brassicaceae families in leached chernozem contaminated by kerosene.

A 120 100 80 60 40 20

Seed germination, % relative to uncontaminated soil

data are insufficient to determine this with certainty. Generally, the experimenters investigate the same plant species (corn, common oat, common barley, beans, common wheat, rye, ryegrass and others), the majority of which belong to three or four families. Based on our experimental data, a link between the systematic affiliation of plants and their germination in soil contaminated by kerosene has been established. Fig. 2 presents the generalized data on germination of plants from the Poaceae, Fabaceae, Asteraceae and Brassicaceae families, which comprise 64% from the total number of tested plant species. It is evident that at low kerosene concentrations, average germinations of plants from all these families were similar, and with increased contamination level the germination of Fabaceae plants was least reduced among those plants examined. On the whole, the tolerance of the tested families to soil contamination decreased in the following order: Fabaceae (germination decrease of 10–60%) > Brassicaceae (5–70%) > Asteraceae (25–95%) > Poaceae (10–100%). Next, we identified the link between plant taxonomy and germination in the soil contaminated by various PHC contaminants by comparing data obtained for kerosene with reference data for crude oil and diesel fuel. We performed comparative analysis for Poaceae and Fabaceae plants as most experimental data were obtained for these families. Averaged values of relative germination of representatives of these families at soil contamination by three PHC types are presented in Fig. 3. Despite wide scatter in the data for each family (5–50%) due to differences in experimental conditions and specific plant features, the comparison indicated the existence of a common factor for kerosene, diesel and oil. At low contaminant concentrations, relative germination of plants from both families did not differ greatly (77% and 73% for kerosene, 86% and 97% for diesel fuel, and 63% and 73% for crude oil; for Poaceae and Fabaceae plants, respectively). In contrast, at higher PHC concentrations, the average germination of Poaceae plants was significantly lower than that of Fabaceae plants. For medium contamination level germination occurred in 41% and 63% for kerosene, 26% and 68% for diesel fuel, 61% and 77% for crude oil; at high concentrations: 0% and 34% for kerosene, 32% and 68% for crude oil for Poaceae and Fabaceae families, respectively (for diesel fuel no data are known). Very high concentrations yielded the lowest levels of plant germination (0% and 32% for kerosene; 29% and 65% for crude oil; for diesel fuel no data are known). Using the data acquired in this study, we have attempted to establish relationships between plant tolerance to PHC contamination and the structure and properties of their seeds. Seed mass and the morphological and anatomical structure of the seeds have been reviewed in the literature as factors that affect seed

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0

B 120 100 80 60 40 20 0

C 120 100 80 60 40 20 0

Kerosene concentration, % on dry basis Poaceae

Fabaceae

Fig. 3. Average seed germination of plants from Poaceae and Fabaceae families in soils contaminated by crude oil (A, various soils, reported data), diesel fuel (B, various soils and composts, reported data), and kerosene (C, leached chernozem, own data).

tolerance to high levels of contamination (Anoliefo and Vwioko, 1995; Robson, 2004). Some researchers indicated that small-seeded plants are more sensitive to soil contamination by heavy metals and pesticides as compared to large-seeded (Clark et al., 2004; Robson et al., 2004). However, in our study the values of EC10, EC25 and EC50 (parameters of phytotoxicity of the contaminated soil) did not correlate with the seed mass (Table 1). The effect of testa structure and thickness on toxicity may be one reason for this. The testas of small seeds are usually several times thicker than those of large seeds and may prevent PHC penetration (Lotova, 2000). In this study, germination of monocotyledonous (two families, 11 species and sorts) and dicotyledonous (16 families, 39 species and sorts) plants were compared and indicated that, while the former fell into the group of medium- and low-tolerance to kerosene contamination, representatives of the latter were found in all tolerance groups (Table 2). At low contamination levels relative germination of monocotyledonous plants made up 91% and 99% respectively on the average; at medium levels — 49% and 51%; at high and very high kerosene concentrations no germination of plants was observed

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in either family (Appendix A). In comparison, germination of dicotyledonous plants varied significantly at all levels of soil contamination. Other characteristics of seeds are well described in the literature for the Poaceae and Fabaceae families. Analysis of differences between some species may provide explanations for higher tolerance of Fabaceae plants to soil contamination by kerosene. Fabaceae seeds have better protection against environmental factors. Their testa consists of five cell layers which reflects their hard-seedness and, correspondingly, reduces permeability of water-dissolved substances (Lotova, 2000; Nicolaeva et al., 1999). In contrast, seeds of Poaceae have two-to-three layers of cover cells with hollow cells (Nicolaeva et al., 1999), where PHCs can accumulate. Fabaceae seed germ is rather large and contains two cotyledons, whereas Poaceae germ is small and immature, with only one cotyledon. In addition, Poaceae endosperm adjoins the germ only at one side making it more susceptible to PHC toxicity. Fabaceae seed storage compounds are concentrated in cotyledons and are represented primarily by proteins (up to 50%), which apparently protect cells against PHC toxicity. Poaceae seeds have well matured endosperm where carbohydrates (starch) are accumulated. Underground germination is typical for many Poaceae seeds, whereas aboveground germination is typical for Fabaceae seeds (Nicolaeva et al., 1999) and this shortens the contact time of their germ with contaminated soil and porewater considerably. The experimental data presented here does not reveal clear connections between the degree of plant tolerance and a plant's life expectancy. Among the Fabaceae plants, perennial species were more stable, and among representatives of Poaceae, Brassicaceae, Asteraceae, Hydrophyllaceae and Linaceae families, annual species were more stable. There are very few data comparing the tolerance of cultivated and wild plants to PHC contamination. According to Shilova (1988) and Kireeva (2003), cultivated plants, crops in particular, are more sensitive to contamination by oil than are wild plants. However, our data show that for soil contaminated by kerosene, wild plants were only somewhat tolerant (mid- and low, with germination of 30–70% relative to the control) or intolerant (b30%) (Tables 1, 2). 5. Conclusions The tolerance of plants to soil contamination by kerosene has a species-specific nature and, on average, decreases in families by the following pattern: Fabaceae > Brassicaceae > Asteraceae > Poaceae. The monocotyledonous species tested here were characterized as having medium- and low-tolerance to soil contamination by kerosene, whereas representatives of dicotyledonous plants were observed in all tolerance groups. Comparison of the tolerance of annual and perennial plants did not indicate any definitive difference. Wild plants were more sensitive to kerosene toxicity than cultivated plants, which is contrary to reference data on oil toxicity. No correlation was observed between degree of plant tolerance to kerosene and mass of plant seeds. Structure and properties of testa, structure of germ, type of storage compounds, and type of seed germination are more important. Plant species exhibiting the highest sensitivity to kerosene (redroot amaranth, Siberian pink, greater burdock, jimsonweed) are recommended for biotesting of PHC-contaminated soils. The most tolerant plants tested, including representatives of the Fabaceae and Asteraceae families as well as corn (Poaceae family), lacy phacelia (Hydrophyllaceae family) and common flax (Linaceae family), may be useful at sites contaminated by petrochemicals. The latter should also be considered for further testing as potential phytoremediators of PHC contaminated soils. Supplementary materials related to this article can be found online at doi:10.1016/j.scitotenv.2012.02.009.

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