1a. Objectives (from AD-416):
1. Develop, validate, and transfer multiclass, multiresidue methods for pesticides, environmental, and emerging contaminants in FSIS-regulated foods, and conduct a survey of food samples for the contaminants. 1A. Simultaneous analysis method for diverse pesticides, legacy and emerging environmental contaminants in meats. 1B. Multiresidue method for food packaging (FP) contaminants in packaged foods. 1C. Conduct a survey of food samples for emerging environmental and FP contaminants. 2. Develop qualitative screening and identification criteria with automated data processing that meets regulatory needs to minimize/avoid false positives and negatives. 3. Develop sample processing methods for regulatory analysis that improve the ability to detect combinations of veterinary drug residues in the same sample preparation. 3A. Simultaneous analysis method for diverse veterinary drugs residues, including aminoglycoside antibiotics, in meats. 3B. Evaluate cryogenic sample processing to achieve meaningful representative sample size of 100 mg in multiresidue analysis of pesticides and veterinary drug residues in foods. 4. Develop automated high-throughput sample processing, preparation, and analysis methods using flow-injection and/or open probe techniques coupled with mass spectrometry to monitor >500 veterinary drugs, pesticides, and environmental contaminants in foods. 5. Develop novel analytical methods for inorganic and organometallic heavy metals [for example forms of mercury (Hg) and arsenic (As)] in foods. 5A. Develop novel analytical methods for mercury speciation and quantification in foods. 5B. Develop novel analytical methods for arsenic speciation and quantification in foods. 6. Develop and use bioanalytical methods (including mass spectrometry) to monitor for antibiotic resistant organisms and/or their biomarkers in conjunction with antibiotic residues in seafood and meats.
1b. Approach (from AD-416):
The specific approaches for meeting the project’s objectives and milestones are as follows: 1) develop, validate, and transfer multiclass, multiresidue methods for pesticides, environmental, and emerging contaminants in FSIS-regulated foods, and conduct a survey of food samples for the contaminants; 2) develop qualitative screening and identification criteria with automated data processing that meets regulatory needs to minimize/avoid false positives and negatives; 3) develop sample processing methods for regulatory analysis that improve the ability to detect combinations of veterinary drug residues in the same sample preparation; 4) develop automated high-throughput sample processing, preparation, and analysis methods using flow-injection and/or open probe techniques coupled with mass spectrometry to monitor >500 veterinary drugs, pesticides, and environmental contaminants in foods; 5) develop novel analytical methods for inorganic and organometallic heavy metals [for example forms of mercury (Hg) and arsenic (As)] in foods; and 6) develop and use bioanalytical methods (including mass spectrometry) to monitor for antibiotic resistant organisms and/or their biomarkers in conjunction with antibiotic residues in seafood and meats.
3. Progress Report:
Progress was made in all objectives, and the project is progressing according to schedule, and even ahead of schedule in objectives 2, 3, and 4. Objectives 1A and 3A are complete, and for 3B, cryogenic sample processing equipment and techniques were evaluated and compared with room temperature approaches for different food commodities and test portion sizes. The room temperature method and devices are sufficient to meet regulatory needs with smaller sample sizes than commonly used now, which enables further reduction in reagent costs and substantially higher sample throughput. With respect to Objective 5, novel analysis to speciate mercury in foods was achieved by designing a method using a quartz-coil reactor to perform differential photochemical vapor generation following by atomic fluorescence spectroscopy. For analysis of inorganic arsenic, a new approach using a thermoelectric cryogenic trap was successfully validated using rice flour certified reference material. On Objective 6, multiple methods for rapid determination of an aminoglycoside antibiotic modification by resistant bacteria were developed. In addition, a bottom-up proteomics approach combined with computational prediction technologies for targeted antibiotic resistance proteins was developed. Lastly, alternative commercial proteases were explored using antimicrobial resistant bacteria to validate cleavage efficiency.
1. Method for analysis of pesticides and environmental contaminants in meats and poultry. The USDA Food Safety and Inspection Service (FSIS) is responsible for routine monitoring of pesticide residues in meat and poultry products to assure that regulatory tolerances are not exceeded. ARS scientists at Wyndmoor, Pennsylvania developed and validated a fast, efficient high-throughput analytical method for FSIS-priority pesticides and environmental contaminants in meat (swine, cattle) and poultry (chicken) muscle. Satisfactory validation results were achieved for 219 contaminants. The method was transferred to FSIS laboratories, and is currently being implemented for routine monitoring of contaminants. The implementation of the new method is expected to improve regulatory monitoring, providing reduced cost per sample analysis, increased sample throughput, and reliable data for more contaminants of concern.
2. Reduction of inorganic arsenic content in cooked rice. Rice is a staple food for half the world population, but it contains higher levels of toxic inorganic arsenic (iAs) than other common crops. Chronic exposure to iAs through dietary pathway is a worldwide concern for rice consumers. ARS researchers at Wyndmoor, Pennsylvania developed an effective protocol to reduce iAs in cooked rice by presoaking the rice in hot water for 10 minutes, then discarding the water before cooking until the rice becomes dry using a rice cooker. Previous efforts to reduce iAs in cooked rice took longer and/or were less effective. This more rapid protocol achieved similar or better iAs reduction by raising soaking temperature above the gelatinization temperature of rice starch, which causes higher diffusion kinetics to reduce iAs levels. Implementation of this protocol in daily practice would cut cancer risk of rice consumers and improve their long-term health prospects.
3. Proteomic analysis of computationally predicted peptides from antimicrobial resistant bacteria genomes. As antimicrobial resistance continues to become a global issue, better and faster analytical methods need to be devised to slow its progression. Proteomics could be a viable option, but little research has been conducted to fulfill this purpose because large scale proteomics experiments are costly. Instead, open source software platforms are available to streamline proteomics studies without need for extensive empirical measurements. ARS scientists at Wyndmoor, Pennsylvania tested this concept for targeted analysis of enzymes associated with antibiotic resistance using only its genome to predict the tryptic digest peptides generated. This technique facilitates subsequent method development using liquid chromatography - mass spectrometry for implementation. In a feasibility study, comparable digestion efficiencies for two proteases determined by this approach suggests that the more cost-effective method can be used for large scale proteomic investigations in the future.
4. Streamlined extraction for analysis of mercury in seafoods. Traditional open-vessel extraction of mercury from foods for analysis suffers from analyte loss via volatilization, but ARS researchers at Wyndmoor, Pennsylvania designed a better reflux digestion system that consisted of a block heater and multiple Pyrex culture tubes to provide improved extraction efficiency. Unlike closed-vessel systems, this open-vessel setup allows easy dissipation of nitrous oxides and nitrites that normally interfere in the analysis. This protocol was validated using a certified reference material, achieving quantitative recovery with <5% relative errors. This approach only costs $1,000 for materials compared to $35,000 for a traditional closed-vessel microwave-assisted extraction system, including the vessels, and provides equivalent results and potential greater sample throughput.
5. A new analytical method for organophosphate ester plasticizers and flame retardants in meats and fish. ARS scientists at Wyndmoor, Pennsylvania, developed a new method for analysis of 14 organophosphate ester (OPE) plasticizers and flame retardants in meats and fish. OPEs are widely used as additives to plastics, electronics, furniture and textiles, and easily migrate from materials to the environment. Some OPEs are persistent, bioaccumulative, and toxic. The developed method was based on the QuEChERS approach and automated robotic cleanup of the extracts. The final extracts were split and analyzed in parallel by gas and liquid chromatography coupled to tandem mass spectrometry to provide an additional degree of confidence in the results. The new method was successfully applied to the analysis of real-world meat and fish samples form the market, further demonstrating the utility of the method for implementation in regulatory and commercial laboratories.
6. Rapid determination of antimicrobial resistance of aminoglycoside antibiotics. Microbial resistance to antibiotic drugs, such as aminoglycosides, is a major health concern worldwide, and more efficient and effective methods of analysis are needed to monitor and study resistance. ARS researchers in Wyndmoor, Pennsylvania have developed an analytical method capable of determining antibiotic resistance via acetylation and phosphorylation in less than 3 hours following overnight enrichment. This is important because aminoglycosides and their enzyme-modified form are challenging to analyze using conventional HPLC instrumentation. These methods allow their detection/quantification and provides the framework for an all-inclusive antibiotic modification evaluation method. In-house developed software was then used for rapid quantification of the biomarkers.
7. Identification of food packaging chemicals extracted from plastic stretch film. Food packaging is important in protecting food, extending its shelf-life, and providing consumers with food handling convenience. However, during storage and handling, some chemicals from food packaging materials may potentially migrate into packaged food. ARS scientists at Wyndmoor, Pennsylvania, in collaboration with San Diego State University scientists, identified chemicals extracted from food packaging plastic stretch film using comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC×GC/TOF-MS). Common plasticizers, polymer and plastic additives, UV filters, fragrances, flavoring agents, among others, were tentatively identified. These data offer an insight on identity of potential food packaging contaminants, and pave the way for future studies on migration of food packaging chemicals into real foods needed for risk assessment to protect consumer’s health.
8. Determination of contaminants in mussels and oysters from Hong Kong. Aquaculture operations are growing worldwide and will continue to increase to meet global food demands. ARS scientists at Wyndmoor, Pennsylvania, in collaboration with researchers from Baylor University, Texas, and Hong Kong Baptist University, Hong Kong, examined bioaccumulation of contaminants by marine bivalves located near sewage and landfill wastewater discharges in Hong Kong, the fourth most densely populated country in the world. Multiple classes of pharmaceutical residues, pesticides, polycyclic aromatic hydrocarbons (PAHs), and flame retardants were detected at low ng/g levels. Initial estimates of acceptable servings per week indicated low consumption risks for measured contaminants with developed U.S. EPA reference dose. These data shed light on aquaculture product safety, particularly, the products from rapidly urbanizing regions of developing countries with limited infrastructure.
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