In the wake of last year’s passage of the US Food and Drug Administration Food Safety Modernization Act, mass spectrometry vendors anticipated increased demand for their instruments as food producers and testing outfits added resources to meet the new regulations.
Such ramping up was expected to impact vendors’ small-molecule detection businesses primarily, as the act’s increased emphasis on prevention would require more upfront testing for chemical adulterants like melamine. However, industry and researchers have also begun exploring protein and proteomic analyses as food safety tools, with the aim of both improving upon existing techniques and anticipating future testing needs.
To an extent, this interest in protein-based food testing is driven by the successes in the small-molecule arena, Paul Zavitsanos, worldwide food business manager at Agilent, told ProteoMonitor. He offered the example of milk safety testing, an area, he said, in which adulterators looking to increase the protein content.of milk are shifting from using chemical contaminants to relying on large-molecule biopolymers.
Milk prices are based largely upon protein content, which has conventionally been determined by measuring nitrogen levels. This nitrogen is “typically protein nitrogen," Zavitsanos said, “but it could be any nitrogen-containing molecule – for instance, melamine.”
As regulators have become more adept at testing milk for small-molecule contaminants like melamine, adulterators have “turned their attention to large-molecule biopolymer contamination of milk," he said. “So waste products from leather factories, hide glue extracts, things from rendering plants all find themselves as protein-containing content that ends up in milk powder.”
Separating these low-abundance non-milk proteins from milk proteins and quantifying them accurately enough for uses such as legal prosecution “is a very difficult thing,” requiring a highly sensitive analysis, Zavitsanos said. “So this is one of the major areas today where proteomics has been identified [for use] in food safety.”
Such work is still in the “embryonic” stage, he said, but, as the crackdown on small-molecule contaminants continues, more attention will be devoted to protein-based testing.
The potential for such contamination “has been on the radar for a long time, but it’s never been prevalent before because it was always easier to just go buy a bag of melamine and adulterate milk that way,” Zavitsanos said. “Now, though, this thing has risen to the surface as other issues have begun to resolve. The game has changed now to, ‘OK, how are you going to detect it if I use ostrich egg protein to contaminate your chicken egg protein? Figure that out.’”
Beyond food contamination, food mislabeling is another issue researchers and mass spec vendors are looking to tackle through proteomics. According to Brent Lefebvre, food and environmental products manager at AB Sciex, studies have suggested that up to 30 percent of fish sold in markets and restaurants are mislabeled.
“That could be a big problem,” Lefebrve told ProteoMonitor, “and one that we could solve through mass spec [using a conventional] extraction, digestion [proteomic] workup.”
Mass spec-based proteomics could also prove useful in tracing food sources, Zavitsanos suggested, likewise citing the example of fish.
“If you want to find out not only what species it is but also: Is it a protected fishery, an illegal fishery? Is it an adult fish? Was it too young to be captured? Those kinds of things are best handled through the proteome rather than the genome,” he said. “So that’s another avenue of proteomic research as applied to food safety.”
Microbial detection is another area where proteomics could prove useful for food safety. The main player on this front currently is Bruker, whose MALDI Biotyper instrument uses protein signatures to ID bacteria for food safety as well as clinical and environmental applications. In May 2010, the company sold a Biotyper to the Austrian Agency for Health and Food Safety for microorganism analysis.
The primary advantages of the Biotyper compared to conventional biochemical tests are lower consumable costs and faster results, George Goedesky, executive director of North American business development at Bruker, told ProteoMonitor. The device can analyze a bacterial colony in roughly five minutes, he said, while “a rapid biochemical test could take four to six hours and conventional biochemicals could take 24 to 48 hours.”
Lefebrve said AB Sciex has done its own proof-of-concept work demonstrating the use of certain of its TOF instruments for bacteria identification, but, he acknowledged, Bruker is the only mass spec vendor currently with a product positioned specifically for that market.
AB Sciex is also investigating use of its instruments for allergen testing, introducing in October its Allergen iMethod Application for determining the presence of egg and milk allergens in baked goods.
The application uses multiple-reaction monitoring mass spec on an AB Sciex 4000 QTRAP to detect egg and milk proteins and is designed to replace current ELISA and PCR-based tests.
Both ELISA and PCR have drawbacks that make a mass spec-based protein test desirable, Lefebrve said. In the case of ELISA, false positives due to interference and false negatives due to denaturing of allergen proteins during cooking can both be problematic. PCR, meanwhile, is limited by the fact that nucleic acid content doesn’t always reflect the content of the corresponding proteins.
“Egg and milk are good examples where there is a ton of protein in there but very few DNA strands that you are able to detect,” Lefebrve said.
He added that the specificity of MRM-MS offers great confidence that the proteins detected are, in fact, the allergens being tested for – something that is key for regulators in the case of product recalls.
“In cases where a positive test result has to go to court [due to a challenge from the manufacturer], the agencies that do the testing are very interested in having legally defensible results,” Lefebrve said.
Zavitsanos agreed that mass spec’s specificity gives it a potential advantage over ELISAs for food safety applications. “Food is a very complex matrix, and so you get a lot of cross-contamination [with ELISAs],” he said. “That generates business risk if you’re a food producer.”
Agilent’s research into mass spec-based allergen testing has, thus far, focused on gluten testing. The company plans to present work on such an application at FDA sometime this year, Zavitsanos said.
Regulation of food safety tests is not as rigorous as it is in clinical diagnostics, but, Lefebrve said, the approval process “is not trivial.”
“We’re currently investigating the levels of approval we need to go through to get the [baked goods allergen] test adopted globally,” he said. “It’s an ongoing process.” He noted that the method would initially appeal most to organizations like contract testing agencies that already own the mass spec equipment required. Such outfits could immediately take advantage of mass spec’s low reagent costs to offer testing at higher margins than is possible using ELISAs.
Several different types of mass spec instruments could prove useful for food safety work. Testing for known analytes, as in allergen testing, will likely make use of triple-quadrupoles, as in the case of AB Sciex’s iMethod Application for baked goods testing. Screening for contamination by unknown proteins, as in milk adulteration testing, will likely be done on high-resolution instruments like high-end Q-TOF or Orbitrap machines.
Use of protein and proteomic analyses for food safety work could add to vendors' revenue streams in a variety of ways, Zavitsanos said. “You have specialized consumables, specialized instrumentation, specialized software, specialized expertise that could be sold as a service.”
“Food safety was small molecule[-based] 99 percent of the time,” he said. “Now the proteomics business is expanding into food, so the mass spec business expands because of the increase in testing from a proteomics angle that never existed before. We all have line items in our strategies that say this is going to be a good opportunity for us financially.”
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