Would you eat a brown apple?

Have you ever cut up an apple to take for lunch, or prepared apples for a fresh fruit tray only to have them turn an unappealing shade of brown? You’re not alone. There’s nothing wrong with brown apple slices, but they certainly don’t look nice, which discourages some people from eating as many apples as they should. Apples are a healthy snack and anything that gets people to eat more fruit could be considered beneficial.

Like it or not, sliced apples that don’t brown are in demand. Many children and some adults have hard time biting into whole apples. In addition, there is much convenience in being able to eat one slice at a time, no matter where you are. Some companies are producing sliced apples treated with a chemical solution to keep them from browning, and you can find them in some schools and in places like McDonald’s and Subway restaurants, but that has its own complications, including what some say is an off-taste and additional plastic waste.

A Canadian company has developed apples that won’t turn brown, which has the potential to solve this problem and get more people eating an apple a day. In this post, I’ll discuss the chemistry behind browning and the science behind non-browning fruits and vegetables.

The enzymatic reaction that turns apples brown within minutes is a major problem for home cooks and professional chefs alike. Just Google how to stop an apple from turning brown and you’ll get 2,770,000 results, including a pretty cool at-home apple browning experiment guide (pdf) by the Australian Institute of Food Science and Technology.

Phenol is the simplest of all the phenolic compounds.

Why do apple, potatoes, avocados, peaches, and many other fruits and vegetables turn brown when cut or bumped?

Phenols are a whole category of compounds found naturally in a lot of foods. Most plant phenols are fine for humans to consume and some even seem to have anti-cancer properties and slow aging, but some are toxins, some may cause cancer, and many do things like reduce the absorption of iron from food. The enzyme polyphenol oxidase catalyzes a variety of reactions among phenols. Polyphenol oxidase oxidizes phenolic compounds into quinones and then links the quinones into pigments that make the surface of light colored produce look brown. There are actually a whole family of polyphenol oxidases that each work on slightly different molecules, and each plant, animal, or bacterium may have many different genes for different types of polyphenol oxidases.

Dr. Anne Marie Helmenstine describes the chemistry behind the prevention of apple browning on About.com:

The reaction can be slowed or prevented by inactivating the enzyme with heat (cooking), reducing the pH on the surface of the fruit (by adding lemon juice or another acid), reducing the amount of available oxygen (by putting cut fruit under water or vacuum packing it), or by adding certain preservative chemicals (like sulfur dioxide). On the other hand, using cutlery that has some corrosion (as is seen with lower quality steel knives) can increase the rate and amount of the browning by making more iron salts available for the reaction.

Frank and a large Pink Lady apple. The apple was beautiful and delicious, but would have browned terribly if it had been prepared in advance for a fruit tray, salad, or similar fresh use.

All of the methods to deter browning have some effect on taste or texture, which is sometimes ok, sometimes not, depending on what you plan to do with the apples. If you’re baking a pie, or putting apples in a salad, a little lemon or salt probably doesn’t matter, but if you’re preparing apples for a fruit tray for guests to savor with cheese and wine, any apple contaminants are unacceptable.

Okanagan Specialty Fruits, a Canadian fruit breeding company in Summerland, British Columbia, has developed a way to keep apples from browning without the need for special heat or chemical treatments. How did they do it? The short story is that they silenced the gene that makes the polyphenol oxidase enzyme so that the enzyme is no longer produced. No enzyme, no browning.

As for the details, we don’t have many. If you’ve read any of the “news” articles about these apples, you know that lots of the stories are short on science and short on facts. The company isn’t telling much on their website*, and hasn’t published any peer-reviewed papers on their process (probably because they don’t want anyone to steal their ideas), so we’ll have to wait until the APHIS risk assessment for petition 10-161-01p is made public.

Until then, the AP article by Shnnon Dininny gives an important clue. USDA asked to approve GMO apple that won’t brown is pretty well researched and includes quotes from Neal Carter, president of Okanagan Specialty Fruits. Ms. Dininny writes: “the company licensed the non-browning technology from Australian researchers who pioneered it in potatoes.” Before I get into the details of how polyphenol oxidase was silenced in potatoes (and apples), there are some things that I apparently have to address, based on comments on this AP story on Grist and elsewhere. Here we go:

THE APPLES HAVE NOTHING TO DO WITH MONSANTO.
THE APPLES HAVE NOTHING TO DO WITH POTATOES.
THE APPLES ARE GOING TO ROT THE SAME AS ALL APPLES ROT.
THE APPLES ARE DIGESTED THE SAME AS ALL APPLES ARE DIGESTED.
THE APPLES HAVE NOTHING TO DO WITH MONSANTO.

Sorry for yelling, but people just aren’t getting it, despite Ms. Dininny’s excellent reporting. Here’s hoping this post helps a little. On to the details.

RNA can bind to itself and form a "hairpin loop", creating double stranded RNA. This structure is key to RNA interference.

I think the Australian researchers that Ms. Dininny referred to are from CSIRO (the Commonwealth Scientific and Industrial Research Organisation, which is Australia’s national science agency), but they haven’t published anything specifically about polyphenol oxidase silencing either. They have published a lot of papers about their efforts to use RNAi, though, which leads me to believe that the gene for the polyphenol oxidase enzyme was silenced in the non-browning apples with RNA interference – RNAi for short.

RNAi is an amazing technology that can be used to shut off genes using the natural mechanisms that exist within a plant (or animal, fungus, etc). Karl has a great explanation of “RNA that Interferes” in his post Cotton like Candy and other excellent explanations can be found elsewhere, such as on the Naked Scientists site, so I won’t go over it again, except to point out that RNAi is used by organisms as a defense against viruses that carry their genetic material as double stranded RNA. RNAi just uses that natural defense mechanism to effectively shut off a gene, and doesn’t require the addition of any new genes.

RNAi can be used to change characteristics in existing plants, such as turning off the genes in onions that make you cryturning off the genes in wheat that make gluten (great for people with celiacs disease!), and turning off other allergens (such as in peanuts and apples). RNAi can also be used to add new characteristics in plants such as nematode resistance or virus resistance (both of which have been done in multiple species). It’s a very versatile tool that I expect we’ll see much more of as researchers and companies figure out new ways to use it, assuming that people can stop freaking out and actually take the time to learn what it’s all about.

Of course, shutting off a gene can cause unintended effects.

For example, a study by Cornell researchers in potato that used RNAi to reduce expression of polyphenol oxidase found that the plants also had reduced disease resistance (Thipyapong, 2004). Polyphenol oxidases seem to play a role in helping plants protect themselves and recover from disease. Note that this experiment reduced the expression of all polyphenol oxidases, not just one, and they used a constitutive promoter that is always on in all tissues. An earlier study, also from Cornell, used a tuber specific promoter so the polyphenol oxidases were turned off only in the potatoes, not in the rest of the plant, and the researchers didn’t find any adverse affects on disease resistance or anything else (Bachem, 1994).

Sometimes the unintended effects of genetic engineering can be very positive. The J. R. Simplot Company has also created reduced browning potatoes using RNAi. In a study that evaluated their potatoes compared to wild-type potatoes, the RNAi potatoes were found to have not only reduced browning but french fries made from the potatoes also tasted better, smelled better, and had greatly reduced accumulation of acrylamide, a toxin naturally produced in potatoes and other foods during high temperature cooking (Rommens, 2004).

Will these non-browning apples have negative unintended effects, positive unintended effects, or both? The truth is, we don’t know yet due to the lack of information coming from Okanagan Specialty Fruits. We’ll just have to wait for that APHIS risk assessment for petition 10-161-01p to see the details of the non-browning apples, but we have a hint in the review Plant Regeneration and Transformation in the Rosaceae (pdf, Rosaceae is the family of plants that includes apples):

Multiple years of field testing of this material confirmed the stability of the non-browning phenotype and have identified no negative impacts on horticultural traits, or on resistance to diseases and insects when grown under field conditions. The non-browning technology developed at [Okanagan Specialty Fruits] has been incorporated into a new enabling platform that: (i) eliminates the selectable marker, (ii) removes all interfering [intellectual property], (ii) uses only plant derived gene sequences and control elements, and (iv) improves the efficiency of gene silencing. Plants arising from this series of transformations are now entering field trials.**

Fruit tray by Tim Inconnu via Flickr.

Remember that fresh fruit tray that this post started with? Which would you prefer – apples treated with chemicals or heat, apples bred to brown a little more slowly, or apples engineered to silence the enzyme that causes browning?

I know what I’d choose for my lunches and for fruit platters that I’d present to my friends and family. Here’s hoping that these apples make it through the regulatory hurdles and lawsuits by activist groups, are planted by a farmer nearby, don’t get uprooted or otherwise destroyed illegally by activists, and make to my table.

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ResearchBlogging.orgAldwinckle H, Malnoy M (2009). Plant Regeneration and Transformation in the Rosaceae Transgenic Plant Journal (3 (Special Issue 1)), 1-39

Bachem C, Speckmann G, van der Linde P, Verheggen F, Hunt M, Steffens J, & Zabeau M (1994). Antisense Expression of Polyphenol Oxidase Genes Inhibits Enzymatic Browning in Potato Tubers Bio/Technology, 12 (11), 1101-1105 DOI: 10.1038/nbt1194-1101

Helmenstine AM (2005). Why Do Cut Apples Turn Brown? About.com Chemistry. Accessed 1 Dec 2010.

Rommens CM, Ye J, Richael C, & Swords K (2006). Improving potato storage and processing characteristics through all-native DNA transformation. Journal of agricultural and food chemistry, 54 (26), 9882-7 PMID: 17177515

Thipyapong P, Hunt MD, & Steffens JC (2004). Antisense downregulation of polyphenol oxidase results in enhanced disease susceptibility. Planta, 220 (1), 105-17 PMID: 15300439

* If anyone from Okanagan Specialty Fruits reads this, it would probably be useful to have a little more info on your website. I know intellectual property is important, but some information is needed. You’re going to have rampant rumor and fear mongering no matter what, but additional info would really help people like me to do a good job of reporting the science. Also, using the trade name Arctic for these apples might not have been the best choice, in my opinion, because it brings to mind anti-freeze genes that we all know get people really freaked out (to anyone else reading this, no, non-browning apples have nothing to do with fish genes, anti-freeze, or anything like that at all).

** This information was from a seminar given at the 1st International Symposium on Biotechnology of Fruit Species, 1-5 September 2008 in Dresden, Germany by J Armstrong and N Carter titled “A new addition to the buffet”. Unfortunately, the text is nowhere to be found. The conference’s website didn’t have any presentation texts and it’s not available on Web of Knowledge either.

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