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Proposal Requirement 4 Example: OPFRs in Plants

Writer: Swasti Singhai

Instagram: @swasti.singhai

Email: swasti18singhai@gmail.com

Current Solutions and Their Insufficiencies

There are three parts to this research - detecting OPFRs, measuring their potential effects on the plants, and lastly, observing the retention of OPFRs in plants over a period of time. 

The traditional method for detecting OPFRs is liquid chromatography-mass spectrometry (LC-MS), a technique that first separates a sample based on its intermolecular forces with different substances, and then analyzes ions based on their mass to charge ratio, revealing data about the structural information of the sample’s constituents. This method has not only been used in biological samples, but plant biochemistry as well. It’s known for its ease of operation, and reliable results. However, a complication in this technique occurs when multiple compounds are insufficiently separated and/or when multiple compounds have the same mass. Another drawback to this method is its high set-up and running costs [1]. 

Figure 2, workflow for liquid chromatography-mass spectrometry 

Source http://www.mtoz-biolabs.com/plant-metabolomics.html


New methods for the detection of OPFRs are emerging, even though they are all based on the traditional LC-MS method. One of these methods is liquid chromatography-electrospray ionization (+)– tandem mass spectrometry. It essentially separates and quantifies OPFRs using a tandem mass spectrometer (MS/MS) with electrospray ionization, a technique used to produce ions from macromolecules, coupled with a liquid chromatograph. This technique is fast, more sensitive, and is able to detect specific variations of OPFRs (chlorinated, brominated, halogen-free). Lastly, another widely used method for the detection of OPFRs is Microwave-Assisted Extraction Combined with Gel Permeation Chromatography and Silica Gel Clean-Up Followed by Gas Chromatography-Mass Spectrometry. This method avoids the influence of lipids on experimental results, and its use is very efficient. One drawback of this method is that it’s extremely comprehensive, requiring samples to be extracted and maintained with very specific conditions, and hence it’s limiting to the types of samples it can be used on [2]. 

The next step is to analyze the potential effects of OPFR on plant samples. Previous studies have used neuroimaging, bioinformatics, differences in functional behaviors, biomarkers, and statistics to identify the effects of OPFR in animal models, humans, and water/sediment. However, many potential effects of OPFRs in plants are observational. For example, the death of plant tissue is indicative of cell membrane disruptors [3]. All environmental factors in the laboratory must be held constant to ensure that the results are indeed due to the effect of OPFR. Stunted growth or growth abnormalities, leaf discoloration, death of plant tissue, decay, and defoliation all will be closely monitored. This observational technique is known as a plant bioassay. One major insufficiency in this bioassay is in the event that OPFR does not cause any external changes. Once again, the research regarding the effects of OPFR on plants is so limited that this cannot be predicted. 

There are a few additional methods to study and measure toxicity. One of these methods is called LD-50 (lethal dose of a substance that will cause 50% of the population to die). This study will attempt to calculate the LD-50 of OPFRs in plants at the end of all trials if significant tissue damage is caused. Evidently, a drawback to this measure of toxicity is the large sample sizes that must be tested to determine it. 

Figure 3, example workflow of process to identify the specific mechanism of action

Source [3]


Analysis

The author opens up with clearly distinguishing the different parts of the research so each description of the technologies can be sorted into which “phase” of the research it would fall under. Doing this makes it easier for the reader to understand exactly what is happening at each step. The author also makes sure to include insufficiencies as well; while a proposal is often seeking for a grant, by including potential drawbacks to a certain method, you’re being as transparent as possible. There’s always room for improvement, and acknowledging a certain technique’s insufficiencies is important to improve/work around it. Then the author continues to describe the various techniques with detail. One thing the author could have improved on is explaining the equipment necessary for harvesting the sample used for LC-MS. Additionally, one thing to note is the use of graphics throughout. They aren’t repeating the information written; moreover, they’re expanding on a concept/technique used. Overall, this part of the proposal is detailed, but could use some additional clarifications on various parts (ex. what algorithms will be used for the bioinformatics analysis in LC-MS, what will be used to harvest the sample, etc). 


References

[1] Labmate, International. “Pros and Cons of LC-MS.” Labmate Online, www.labmate-online.com/news/news-and-views/5/breaking-news/pros-and-cons-of-lc-ms/31157.

[2] Yang, Jiawen, et al. “A Review of a Class of Emerging Contaminants: The Classification, Distribution, Intensity of Consumption, Synthesis Routes, Environmental Effects and Expectation of Pollution Abatement to Organophosphate Flame Retardants (OPFRs).” International Journal of Molecular Sciences, vol. 20, no. 12, 2019, p. 2874., doi:10.3390/ijms20122874.

[3] Ericson, USFWS/Jenny. “Impacts of Chemical Methods - Chemical Methods: Management Methods - Managing Invasive Plants.” Official Web Page of the U S Fish and Wildlife Service, www.fws.gov/invasives/stafftrainingmodule/methods/chemical/impacts.html.

[4] Ospina, Maria, et al. “Exposure to Organophosphate Flame Retardant Chemicals in the U.S. General Population: Data from the 2013–2014 National Health and Nutrition Examination Survey.” Environment International, vol. 110, 2018, pp. 32–41., doi:10.1016/j.envint.2017.10.001.

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