In the Laboratory
Investigation of the Use of Cucumis sativus for Remediation of Chromium from Contaminated Environmental Matrices An Interdisciplinary Instrumental Analysis Project Lynsey R. Butler,† Michael R. Edwards,†† Russell Farmer, Kathryn J. Greenly, Sherri Hensler,††† Scott E. Jenkins, J. Michael Joyce,†††† Jason A. Mann,# Boone M. Prentice,## Andrew E. Puckette, Christopher M. Shuford,### Sarah E. G. Porter,* and Melissa C. Rhoten Department of Chemistry and Physics, Longwood University, Farmville, VA 23909; *
[email protected] In this interdisciplinary semester-long project, upper-level instrumental analysis students studied the use of the cucumber plant (Cucumis sativus) as a rhizofiltrator and a phytoextractor for the removal of chromium from contaminated water and soil. This project has been completed by three instrumental analysis classes, each of which investigated a different set of experimental variables. The analytical technique used for the project was atomic absorption spectroscopy (AAS), although inductively coupled plasma-atomic emission spectroscopy (ICP-AES) could certainly be used with potentially much lower limits of detection. The goals of this project were to
• Introduce the concepts of phytoremediation
• Introduce research methodology by allowing the students to plan and implement their own experiments
• Introduce AAS as an analytical tool for environmental chemistry
• Compare multiple project results to teach students about the importance of data interpretation (both qualitative and quantitative)
Phytoremediation and metal clean-up labs have been published previously, both in this Journal and elsewhere (1, 2). These publications range from a free-form semester-long research project (1) to laboratory exercises designed for two or three lab periods only (2). The project presented here is a hybrid: the research experience is specifically designed to be carried out in an instrumental analysis class over the course of a semester and interwoven with other experiments to give the students experience with all of the instrumentation available. We have chosen to keep the project focused specifically on remediation of chromium by cucumber plants in order to have a common thread among classes.
Current addresses: †Gorbec Pharmaceutical Services, Durham, NC 27713. ††VA–MD Regional College of Veterinary Medicine, Virginia Tech, Blacksburg, VA 24061. ††† Bernard J. Dunn School of Pharmacy, Winchester, VA 22601. ††††Virginia Commonwealth University, School of Medicine, Richmond, VA 23298. #Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14853. ##Department of Chemistry, Purdue University, West Lafayette, IN 47907. ###Department of Chemistry, North Carolina State University, Raleigh, NC 27695.
Environmental contamination of soil and water has become an increasing concern in industrialized countries. In particular, trace metal elements such as Cr, Cu, Fe, Mn, Mo, and Zn are introduced into our environment through human activities such as mining, traffic, and agriculture. Although some of these elements are essential nutrients at trace levels, at high levels they become toxic and can contribute to a host of human and animal health problems (3). The speciation of metal contamination is also of interest, as chromium(VI) is a well-known toxin while chromium(III) is an essential trace element (4, 5). Chromium is an environmental pollutant that can be introduced via the effluent from the leather tanning and stainless steel industries, where it can pollute streams and sewage that might be used as sources of irrigation for agriculture (6). Remediation strategies are necessary for removing the contaminants in situ before they can be introduced into the food chain. Billions of dollars are spent each year on environmental cleanup of organic and inorganic pollutants in the United States alone (7). Some chemical methods of cleanup for heavy metalcontaminated soil and water include solvent extraction, activated carbon absorption, and large scale ion-exchange chromatography (3). Methods such as solvent extraction create a secondary problem of solvent waste disposal and are costly to implement. Clean-up methods that can help to alleviate this financial and environmental burden have gained popularity in recent years. Phytoremediation has shown promise for the in situ cleanup of contaminated soil or ground water. Broadly defined, phytoremediation refers to the use of plants and their associated microbes for environmental cleanup. Using plants as filters in wetlands or growing them hydroponically to filter polluted water is defined as rhizofiltration. Using plants to clean up pollutants from soil by accumulation in tissues is phytoextraction (7). The use of phytoremediation as a clean-up strategy has been on the rise over the past decade because of the relatively low cost associated with its implementation when compared to more expensive chemical remediation methods. Ultimately these strategies require far fewer raw materials and energy resources than their chemical counterparts, making them attractive alternatives. Various species of cucumbers have been shown in previous studies to effectively remediate organic pollutants such as 1,3,5-trinitro-1,3,5-triazine (RDX, an explosive residue), 1,1,1-trichloro-2,2-bis( p-chlorophenyl)ethane (DDT), and its degradation product 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) (8, 9). Additionally, cucumbers demonstrated phosphorus accumulation in the stems and leaves after 8 weeks of growth in contaminated soil showing that they may function as remediators for inorganic pollutants as well (10). Cucumis sativus was chosen for this project because of its potential as a phytoremediator for inorganic pollutants.
© Division of Chemical Education • www.JCE.DivCHED.org • Vol. 86 No. 9 September 2009 • Journal of Chemical Education
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In the Laboratory
Experimental
Table 1. General Schedule for the Semester-Long Course Week
A general schedule for the semester course is shown in Table 1. This schedule can easily be adjusted to study different growth stages of the plants as indicated by the asterisks in Table 1 or to account for different semester durations. At this university, instrumental analysis lab meets 4 hours per week for 16 weeks. Changing the experimental variables among semesters allowed the students to compare their results to previous classes and draw different conclusions about the ability of the cucumber plant to phytoextract or rhizofilter chromium. It is anticipated that each year’s class will design its own set of variables to study, which increases their interest in the outcome of the project. A PerkinElmer AAnalyst 800 with an air/acetylene flame and a multielement hollow cathode lamp was used to evaluate the quantity of chromium in the plant material. A standard method for analysis of metals in plant tissues was used (11). Calibration curves were prepared by the students in Excel using the average corrected absorbance for the replicate measurements. Other specific details regarding the experimental procedure can be found in the online material.
Task
1
Introduction to lab safety Discussion of project Preparation of experimental design
2
Plant cucumber seeds in vermiculite Prepare “spiked” soil Prepare Hoagland’s solution Uncover seedlings (2 days) Transfer seedlings (4–7 days)
3–4
Preliminary AAS analysis of environmental matrix
***
Harvest plants Collect leaf measurements (if desired) Dry plants and digest for AAS analysis
12
Perform AAS analysis
14
Complete any unfinished instrumental analysis Analyze data
15
All analysis finished
16
Final report due
Note: The asterisks indicate variability for different growth periods of the plants or different semester durations.
Hazards Potassium dichromate is an oxidizer, a potential carcinogen, and can irritate the skin and eyes. Chromium(III) salts are strong oxidizers and irritants. Concentrated hydrochloric acid is corrosive and should always be added to water when diluting (not vice versa). Concentrated solutions of strong bases are caustic and should be handled with gloves. Laboratory coats are recommended and safety goggles are mandatory at all times. Latex or nitrile gloves are strongly recommended owing to the potential carcinogenicity of Cr(VI). All waste must be disposed of in properly labeled waste containers and handled according to local regulations. None of the waste generated in this experiment should be disposed of down the drain. Results Owing to the small sizes of the upper-level classes at this university (usually
