Current research on the volatile emissions emanating from recently-mowed turf-grass indicates that these gases may have a significant impact on the environment. The purpose of this study was to observe the effect of gaseous emissions from different species on the rate of seed germination. In the first study, dandelion (Taraxacum officinale Weber) and white snakeroot (Ageratina altissima (L.) King & Robinson [Eupatorium rugosum Houtt.]) seeds were placed into resealable, plastic bags containing the freshly-harvested foliage (cut into approximately 4 cm lengths) from a species of plant that commonly grows in turf. Seeds were treated with the emissions from one of seven plant species: Cynodon dactylon (L.) Pers., Lolium perenne L., Leptochloa fascicularis (Lam.) Gray, Taraxacum officinale Weber, Trifolium repens L., Glechoma hederacea L., or Plantago lanceolata L. Germination rates were examined over a 18-day period, and the foliage-gas treatments that incurred the greatest effects were examined in more detail in a second experiment. In that study, A. altissima seeds were germinated in a replicated trial in the presence of cut foliage from one of four different species (C. dactylon (L.) Pers., T. officinale, T. repens, P. lanceolata) or an experimental control. No statistically significant differences were observed for rate of germination in any of the foliage treatments (including the control). A third experiment was conducted in which the cut foliage was placed in sealed flasks in order to allow the build up of gases from the wounded plants. The gases were then analyzed using gas chromatography. In addition, the effect of these gases on crickets, a common grass-dwelling insect, will also be presented.

 Introduction

The scent of freshly mowed lawns is well known to many, but one could ask, "What effect does this "smell" have on ecosystems?" Recently published research suggests that gas emissions from cut grass are volatile organic compounds (VOC) that play an important role in the pollution of the troposphere (de Gouw et al., 1999). Some of the gases emitted from cut turf have been identified as oxygenated VOC’s such as methanol, acetaldehyde, acetone, and butanone, and C₆ oxygenates such as (Z)-3-hexanal, (Z)-3-hexanol, and hexenyl acetate (Kirstine et al., 1998; de Gouw et al., 1999). The purpose of our experiments were to explore the following questions:

• If these gases are pollutants, what effect do they have on the germination of seeds, especially of weeds?
• What effect do these gases have on insects?
• Will the gases stimulate or inhibit growth?
• Will the gases repel insects, attract them, or have no effect?

Material and Methods

Experiment 1:
The seeds (achenes) of two species were used: Dandelion (Taraxacum officinale Weber) and White Snakeroot (Ageratina altissima (L.) King & Robinson [Eupatorium rugosum Houtt.]). The achenes were treated with the gases from one of 6 turf species from Bellarmine College's campus or a control (cheesecloth only). The turf species used were:

• Perennial Ryegrass (Lolium perenne (L.) Pers.)
• Bearded Sprangletop (Leptochloa fascicularis (Lam.) Gray)
• Dandelion (Taraxacum officinale Weber)
• White Clover (Trifolium repens L.)
• Ground Ivy (Glechoma hederacea L.)
• Narrow-Leaf Plantain (Plantago lanceolata L.)

There were 14 petri dishes used with 100 seeds in each. The achenes were planted onto filter paper with 3 ml of distilled water. Seven of the dishes contained Dandelion seeds and the others held White Snakeroot. All of the leaves (4.5g/treatment) were collected, cut 2 or 3 times and wrapped in cheesecloth. The stapled cheesecloth containers (and a control with no leaves) were placed individually into resealable plastic bags. Each plate, with seeds, was placed into a plastic bag with one of the leaf-treatments and sealed. Germination was assessed 6 days and 18 days after planting. Since this was a preliminary study, no replications were used.

Experiment 2:
White Snakeroot achene germination was examined in this experiment. Four sources of turfgrass leaf material and one control (cheesecloth only) were used:

• Bermuda Grass (C. dactylon)
• Dandelion (T. officinale)
• White Clover (T. repens)
• Plantain (P. lanceolata)

For each leaf species 4 petri dishes were used (as replicates) giving a total of 20 dishes (including 4 dishes for the control). The leaves were collected at the same time as the achenes were planted. Dishes were lined with filter paper and 3 ml of deionized water was added to each. The tissue (4.5g) was cut several times and placed (along with a dish of achenes) into a resealable plastic bag. Achenes were allowed to germinate and counted at 5, 10, 15, and 22 days. Statistical comparison was by analysis of variance (SigmaStat v2.0).

Experiment 3:
In this experiment, the gases emitted from the cut grasses were identified using gas chromatography/mass spectroscopy (GC/MS). The GC/MS used was Hewlett Packard 5890 HP-5ms. The column was 5%(crosslinked)-MeSILOXANE 30 meters, .25mm I.D. and 300⁰C max. Helium was used as the carrier flow at 1 cc/min. The oven was set at room temp (25°C), and the injection port was set at 100°C. Three species of plants were used:

• Bermuda Grass
• Dandelion
• Plantain

Four grams of leaves from each species were cut 2 or 3 times and placed into 250ml Erlenmeyer flasks. One flask, the control, contained only air and a few drops of water. Each of the other 6 flasks contained the sample of leaves from one species. Each flask was sealed with a rubber stopper and 3 were placed under a light, while the other 3 flasks were placed in the dark. The flasks were allowed to sit for 5 days. Known gases were injected into the GC-MS as standards. They were:

• Acetone
• Methanol
• Acetaldehyde
• Ethanol
• Butanone

The experimental samples were then collected (by inserting a syringe through the rubber stopper) and injected the gas into the GC-MS.

Experiment 4:
The effect of vegetative gas emissions on crickets (Order Orthoptera) was also examined. Sixteen crickets were purchased from a local pet store to ensure that the insects were all of the same species and grown in the same environment. Each treatment was replicated 4 times (using 16 jars, with 4 as a control). Three turfgrass species were used as sources of gas:

• Bermuda Grass
• Dandelion
• Plantain

The leaves were collected (4.0g) and cut 2 or 3 times, wrapped in cheesecloth, and stapled. Each leaf unit was placed into one of twelve glass jars. One cricket was placed into each of the jars. The experimental control contained one cricket with cheesecloth only. A lid, consisting of doubled cheesecloth and plastic wrap with holes, was placed over the mouth of the jars to seal in the gases as much as possible, while still allowing some air transfer for the insects. The crickets were observed 1 or 2 times a day for 6 days. Each time the distance the crickets were from the cheesecloth-treatment was measured.

Results

Figure 1

 Figure 2

Experiment 1:

• White Snakeroot germination: All turfgrass species, with the exception of Ryegrass, produced a greater cumulative percent germination than the control (Figure 1).
• Dandelion germination: Four species of turfgrass plants induced greater dandelion germination: Ground Ivy , Dandelion, White Clover, and Ryegrass.
• In general, germination was greater in the leaf-gas treatments than in the experimental controls. White Clover leaves netted the greatest amount of White Snakeroot germination while Ryegrass induced the least. The Ground Ivy treatment yielded the greatest amount of Dandelion germination, while Plantain induced the least.

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Figure 3

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Experiment 2:

• Figure 3: The numerical averages indicate that Bermuda Grass and Plantain incurred the greatest cumulative percent germination in White Snakeroot. Dandelion and White Clover produced the lowest cumulative percent germination. Statistical analysis (ANOVA), however, indicated that there were no significant differences for germination between the control and any of the leaf-gas treatments. There was too much variability within the treatments to adequately compare treatments statistically.

Experiment 3:

Table 1:

• Of these 5 standard gases analyzed by GC/MS none were detectable in the experimental flasks (leaf treatments or control).

Table 1.

Table 2 and 3:

• Carbon dioxide (CO₂) was not detected in the empty flask but was in the flasks containing leaf material. Dandelion leaves incubated in the light produced more CO₂ than in the dark, whereas the other 2 species produced more in the dark. This may be because of respiration and/or decay.
• Oxygen (O₂), levels were the same in the empty flask in the light and dark, but were much higher in the flask with leaves that were kept under light. The greatest increase in O₂ was observed in the illuminated flask containing plantain leaves. This increase in O₂ is probably due to photosynthesis.
• Nitrogen (N₂) was the same in all flasks.
• Methane (CH₄) production was higher in the flasks with leaves than the control flasks, and was higher in the flasks kept in the dark (compared to the illumination treatment). This might be due to microbial activity.

Table 2.

Table 3.

Experiment 4:

Table 4:

• On the first 2 days of the study, the Crickets were observed to be closer to the cheesecloth containing leaves than the Crickets in the jars containing cheesecloth only. Therefore, it appeared that the crickets preferred the leafy material. Crickets were even observed to burrow through the cheesecloth into the leaves of the Bermuda Grass treatment.
• By the end of the 3^rd day 4 Crickets had died (3 in leaf treatments, and 1 in a control). On the 4^th day 4 more Crickets died. One more Cricket died on the 5^th day.
• The jar containing plantain leaves appeared to cause the most mortality. Cricket activity and mortality was not noticeably different in the other leaf treatments and controls towards the end of the study.

Table 4:

Cricket distance away from treatment (cm)

Conclusions

• Experiment 1 (Figures 1, 2, 3) provided some evidence of stimulation of germination due to gas emissions from turf leaves, but was not replicated
• Experiment 2 (Figure 4) showed some stimulation of germination with Bermuda Grass and Plantain leaves and some germination inhibition by Dandelion and White Clover leaves. Analysis of Variance, however, showed no statistical significance for germination due to gas emissions, so maybe there’s no effect on the growth of seedlings.
• GC/MS data showed that the levels of methane, carbon dioxide, and oxygen were present in varying abundance (Tables 2 and 3). In the dark, methane and carbon dioxide increased in the presence of leafy material, whereas oxygen decreased, especially in the dark experiment. Oxygen depletion in the dark could be due to decay of the grass and/or lack of photosynthesis.
• No C₆ oxygenates were seen separated in the results from the GC/MS. This could be due to how quickly these gases escape following leaf wounding. Possibly the gases were not collected in time. Also, other researchers on this topic typically detect these gases from drying leaves in a circulating system, whereas in our study the leaves were kept moist by incubation in a closed container.

• Crickets appeared to prefer the leaf treatments as they were generally observed to be near, or on, the cheesecloth containing leaves. So, at the least, the gas emissions from leaves did not repel the insects.

Summary:
We did not detect the type of gases (VOCs) emitted from leaves that have been reported previously. This may be due to our use of enclosed, stationary systems rather than circulating ones. In our studies, neither plant growth (achene germination) nor animal behavior (Cricket movement) was negatively affected by the presence of wounded leaves. In fact, the Crickets appeared to prefer the experimental treatments!

Acknowledgements
We would like to thank the following people for their help with this study:

• Mr. Robert Bernauer (Staff Scientist, Chemistry Department, Bellarmine College)
• Dr. Peter P. Rowell (Professor, Department of Pharmacology and Toxicology, University of Louisville, Louisville, KY)
• Ms. Joann Lau (Graduate Student, Department of Pharmacology and Toxicology, University of Louisville, Louisville, KY)

• Mr. Anton Clemmons (Undergraduate, Biology, Bellarmine College)

Sources

DeGouw, JA, CJ Howard, TG Custer and R Fall. 1999. Emissions of volatile organic compounds from cut grass and clover are enhanced during the drying process. Geophysical Research Letters 26: 811-814.

Kirstine, W, I Galbally, Y Ye and M Hooper. 1998. Emissions of volatile organic compounds (primarily oxygenated species) from pasture. Journal of Geophysical Research 103: 10,605-10,619.