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Andrew J. Price, Craig S. Charron, Arnold M. Saxton and Carl E. Sams

A study was conducted to quantify volatiles generated from Indian mustard (Brassica juncea L. Czerniak) tissue incorporated into soils under controlled conditions. Mustard residues were incorporated into noncovered and covered soils that varied by texture, temperature, moisture, pH, or sterility (autoclaved or nonautoclaved). Sandy loam soil had 38% more allyl isothiocyanate (AITC) than clay loam soil. AITC concentration in 45 °C soil was 81% higher than in soil at 15 °C, and 56% higher in covered compared to noncovered treatments. The microbial catabolism of AITC was suggested by the result that AITC concentration in autoclaved soils was over three times that measured in nonautoclaved soils. The highest AITC level detected (1.71 μmol·L–1) occurred in the autoclaved covered soil. Several factors also influenced CO2 evolution. At 30 or 45 °C, CO2 concentration was at least 64% higher than at 15°C. The covered soil had over twice the CO2 found in the noncovered soil, and the nonautoclaved soil treatment yielded twice the CO2 measured in the autoclaved soil. There were no main effect differences among soil moisture, soil pH, and soil texture treatments for CO2 concentrations. This information could be helpful in defining ideal soil conditions for field scale experiments. Additionally, this study demonstrates a sampling technique for testing fumigation potential of biofumigation and solarization systems that may have the potential to replace methyl bromide.

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Feras Almasri, Husein A. Ajwa, Sanjai J. Parikh and Kassim Al-Khatib

). Research showed that the soil fumigants that are alternatives to MeBr are generally less effective pest control agents. These alternatives to MeBr include chloropicrin (CP), 1,3-dichloropropene (1,3-D), methyl isothiocyanate (MITC) generators such as metam

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Sanjeev K. Bangarwa, Jason K. Norsworthy, Edward E. Gbur and John D. Mattice

bromide. Isothiocyanates (ITC) are a family of compounds composed of nitrogen (N), carbon (C), and sulfur (S), with an alkyl or aryl side chain (R) having the basic structure of R–N=C=S. ITC have shown herbicidal activity on several weed species, including

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Stephanie G. Harvey, Heather N. Hannahan and Carl E. Sams

Allyl isothiocyanate (AITC) is the predominant isothiocyanate produced by damaged tissues of Indian mustard (Brassica juncea (L) Czerniak). This study investigated Indian mustard and AITC mediated suppression of mycelial growth and sclerotial germination of Sclerotium rolfsii Saccardo, a common soilborne pathogen. Indian mustard (IM) treatments of 0, 0.1, 0.2, 0.6, 1.0, 2.0, 4.1, 5.1, 10.2, 20.4, 40.8, 81.6, and 163.3 g·L-1 (weight of reconstituted mustard per liter of air) were evaluated for suppression of mycelial growth. Treatment effect was evaluated by measuring the radial growth of mycelia. Sclerotia were placed in culture tubes containing 18 g autoclaved soil and covered with an additional 5 g soil. AITC at concentrations of 0, 4.0, 16.0, 64.0, 256.0, 1024.0, or 4096.0 μmol·L-1 was injected into the tubes. Treated sclerotia were removed from tubes and plated on potato dextrose agar to determine viability. Mycelial growth was inhibited with IM treatments (P < 0.01). Inhibiting concentrations (IC) of IM for mycelial growth inhibition of 50% and 90% were 0.7 and 1.0 g·L-1, respectively, with death resulting with >2 g·L-1. Inhibition attributable to AITC alone was lower than that achieved by IM producing equivalent amounts of AITC. Germination of sclerotia was negatively correlated with AITC concentration (r = 0.96; P < 0.01). The IC50 and IC90, of AITC were 249.0 and 528.8 μmol·L-1, respectively, at 42 hours. The lethal concentration for sclerotia was not reached; only suppression occurred at the highest treatment concentrations. Sclerotium rolfsii mycelia were sensitive to the IM volatiles and were suppressed at low concentrations. Sclerotia were more resistant than the mycelia and required higher concentrations of AITC to suppress germination.

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Craig S. Charron and Carl E. Sams

The U.S. Clean Air Act bans the use of methyl bromide after 2005. Consequently, the development of alternative methods for control of soilborne pathogens is imperative. One alternative is to exploit the pesticidal properties of Brassica L. species. Macerated leaves (10 g) from `Premium Crop' broccoli [B. oleracea L. (Botrytis Group)], `Charmant' cabbage [B. oleracea L. (Capitata Group)], `Michihili Jade Pagoda' Chinese cabbage [B. rapa L. (Pekinensis Group)], `Blue Scotch Curled' kale [B. oleracea L. (Acephala Group)], Indian mustard [B. juncea (L.) Czerniak, unknown cultivar] or `Florida Broadleaf' mustard [B. juncea (L.) Czerniak] were placed in 500-mL glass jars. Petri dishes with either Pythium ultimum Trow or Rhizoctonia solani Kühn plugs on potato-dextrose agar were placed over the jar mouths. Radial growth of both fungi was suppressed most by Indian mustard. Volatiles were collected by solid-phase microextraction (SPME) and analyzed by gas chromatography-mass spectrometry. Allyl isothiocyanate (AITC) comprised >90% of the volatiles measured from `Florida Broadleaf' mustard and Indian mustard whereas (Z)-3-hexenyl acetate was the predominant compound emitted by the other species. Isothiocyanates were not detected by SPME from `Premium Crop' broccoli and `Blue Scotch Curled' kale although glucosinolates were found in freeze-dried leaves of all species. When exposed to AITC standard, P. ultimum growth was partially suppressed by 1.1 μmol·L-1 (μmol AITC/headspace volume) and completely suppressed by 2.2 μmol·L-1 R. solani was partially suppressed by 1.1, 2.2, and 3.3 μmol·L-1 AITC. Use of Brassica species for control of fungal pathogens is promising; the presence of AITC in both lines of B. juncea suppressed P. ultimum and R. solani but some Brassicas were inhibitory even when isothiocyanates were not detected.

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Stephanie G. Harvey and Carl E. Sams

Isothiocyanates are volatile chemicals produced by damaged tissues of Brassica species. Allyl isothiocyanate (AITC), the predominant isothiocyanate in Indian mustard (B. juncea), has been shown to control pest in laboratory and field experiments. We investigated the effectiveness of AITC against the germination of sclerotia of Sclerotium rolfsii Saccardo, a common soilborne pathogen of tomato. Sclerotium rolfsii was cultured on PDA from a field isolate. Mature sclerotia were collected and placed in polyester mesh bags. Culture tubes (16 × 150 mm) were packed with 18 g clay loam soil. A sclerotia-bag was placed in each tube and covered with an additional 5 g soil. Soil was maintained at 60% field capacity for the duration of the experiment. AITC was injected into each tube through a septum. Treatments consisted of 0, 5.6, 11.2, 22.4, and 44.8 μmol AITC/L of atmosphere and an ethanol control. AITC in each tube was sampled using SPME and analyzed on GC-MS. Tubes remained sealed for 42 h at 30 °C. Sclerotia were then removed from tubes and bags and plated on PDA to determine viability. Radial growth was measured to determine the effects of AITC. Mycelial growth was negatively correlated to AITC concentration (P < 0.01). The highest concentration of AITC resulted in a 40.3% reduction in mycelial growth. Although the AITC concentrations used in this study did not kill sclerotia of S. rolfsii, they did suppress mycelial growth from germinating sclerotia. At higher concentrations, or mixed with other chemicals, AITC may prove to be an affective control for this pathogen.

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Christina L. Pierson, Carl E. Sams, Dennis E. Deyton and Craig S. Charron

Biofumigation is an alternative to traditional methods of soil sterilization such as methyl bromide. Biofumigation utilizes volatile, pesticidal compounds in soil incorporated plant material from various Brassica species. Three experiments were conducted to study the degradation of allyl isothiocyanate (AITC) generated from the breakdown of glucosinolates present in Oriental mustard (Brassica juncea L. Czerniak). Mustard seed meal was incorporated into a sandy clay loam soil in all experiments. In the first experiment, samples were hydrated and then held in an incubator at 20 ± 0.2 °C. Samples were taken periodically for 7 days or until AITC was not detectable. For the second experiment, hydrated samples were removed from the incubator after 4 hours and 5 mL of ethyl acetate was added. The samples were then placed in a refrigerator at 4 ± 0.2 °C and samples were taken periodically over 77 days. For the third experiment, samples were taken from a strawberry plot experiment grown in a randomized complete block design. Samples were taken and 5 mL of ethyl acetate was added. Then samples were placed into a cooler until returning to the laboratory. The incubator experiment was repeated and showed that the highest concentration of AITC occurred between 2 and 8 hours after hydration. The storage experiment showed a stable relationship between time and AITC degradation. AITC was still present after 77 days. The strawberry plot experiment showed rapid AITC degradation similar to the incubator experiment. Future research will be done to confirm the effects of temperature and glucosinolate content on the amount of allyl isothiocyanate present.

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Usha Palaniswamy, Richard McAvoy and Bernard Bible

Watercress (Nasturtium officinale R.Br.) plants were grown in growth chambers at 15 °C or 25 °C and either 8- or 12-h photoperiod (PP). The photosynthetic photon flux (PPF) was 265 μmol·m-2s-1 in all chambers, but beginning 1 week before harvest, half of the plants in each chamber were subjected to a higher PPF (435 μmol·m-2·s-1). At harvest, watercress leaves and stems were analyzed for phenethyl isothiocyanate (PEITC) concentration. Without supplemental PPF, watercress grown at 25 °C and 12-h PP produced higher PEITC concentration in leaves and stems than plants grown at 15 °C and 12-h PP, or plants grown at 8-h PP and either temperature. With one week of supplemental PPF before harvest, plants grown at 15 or 25 °C and the 8-h PP produced PEITC concentrations as high as plants exposed to 12-h PP and similar temperatures. However, a week of supplemental PPF did not alter PEITC concentrations in plants grown at the 12-h PP, regardless of temperature. At 25 °C, plants grown under the low PPF and the 12-h PP produced 62% greater dry mass than plants exposed to a week of high PPF and the 8-h PP, but did not differ in PEITC content. Thus, the effect of one week of high PPF on PEITC concentration depended on photoperiod.

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Heather D. Toler, Craig S. Charron, Carl E. Sams and William R. Randle

decreased risk of cancer ( Gross et al., 2000 ; Hecht, 2000 ; Zhang and Talalay, 1994 ). When consumed, glucosinolates are broken down into their hydrolysis products—isothiocyanates—which in turn stimulate the activities of the anticarcinogenic phase II

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Ryan R.P. Noble, C.S. Charron and C.E. Sams

The development of alternative methods for control of soilborne pathogens is imperative since the U.S. Clean Air Act bans the use of methyl bromide after 2005. One possibility is to exploit the pesticidal properties of compounds released by macerated Brassica tissues. In this study, masked chaffer beetle larvae were placed in sealed 473-mL jars with 335 g of soil amended with 1%, 2%, 4%, or 8% (g·g–1) Brassica tissue. The most prevalent volatile toxic compound of Brassica juncea (PI 458934) is allyl isothiocyanate (AITC). AITC production was measured in the jars at 0.25, 4, 8, 24, and 48 h using a solid-phase microextraction device (SPME) and gas chromatography. After 7 days, larvae mortality was determined. Control treatments included untreated soil, soil amended with 8% tomato plant tissue, soil amended with pure AITC, and untreated soil with an atmosphere of ≈20% O2 and 0% CO2 changing over 48 h to 2% O2 and 20% CO2. AITC levels were positively correlated to larvae mortality. The estimated lethal concentration for 50% kill (LC50) was 3.6 μg AITC/L soil atmosphere. AITC levels may be influenced by Brassica mass added, soil bulk density, and environmental factors including temperature and moisture. B. juncea has a high tissue AITC concentration. However, the mass of Brassica tissue required for insecticidal application against Cyclocephala sp. is also high, between 4% and 8% of soil mass. Development and selection of Brassica species that produce higher concentrations of isothiocyanate would increase the effectiveness of Brassica biofumigation as an alternative to methyl bromide for controlling soilborne insects.