Lon Wilson

Last year I covered Khodakovskaya et al.’s paper regarding the benefits of growing tomatoes in carbon nanotubes (CNT).^[CB] At the time I was concerned with the potential health risks associated from eating carbon nanotubes, but today in ACS Nano my concerns are alleviated. A paper from Lon Wilson’s and Fathi Moussa’s research groups discusses the effects from administering oral doses of carbon nanotubes (concentrations as high as 1g of CNT per kg body weight) to Swiss mice.^{[ACS Nano]} The authors summarize their work the best.

CNT materials did not induce any abnormalities in the pathological examination. Thus, under these conditions, the lowest lethal dose (LD_Lo) is greater than 1000 mg/kg b.w. in Swiss mice.

So feel free to eat all the CNTs you want in lab, assuming they are not functionalized, you do it only once, and you limit yourself to single walled carbon nanotubes. I think partly because the results of the oral administration of CNTs went without any interesting side effects to present, the authors also looked into what happens when you inject CNTs into the peritoneal cavity of mice.

The image on the left is the control while the image on the right is 14 days after injecting mice with CNTs at a concentration of 1g CNT per kg of mouse. Although it looks sickly, the mice injected with the high concentration of CNTs did not die. Well…, not from the CNTs anyways.

Link to paper: In Vivo Behavior of Large Doses of Ultrashort and Full-Length Single-Walled Carbon Nanotubes after Oral and Intraperitoneal Administration to Swiss Mice (ACS Nano)

Mitch

Mariya Khodakovskaya

Alexandru Biris

There has been a lot of concern over the health effects arising from the burgeoning field of nanotechnology, David Barden covered one such paper focusing on nanotube production in Highlights in Chemical Science earlier this month.^[HCS] What hasn’t been as discussed are the potential health benefits of carbon nanotubes (CNTs). In a paper released yesterday in ACS Nano, Mariya Khodakovskaya & Alexandru Biris (+coauthors) found that tomato seeds grown in a medium of carbon nanotubes germinated and grew more efficiently than their control group brethren.^{[ACS Nano]} This result is spectacularly seen from the image below.

After 27 days of growth.

The tomatoes grown in carbon nanotubes weighed more, grew longer stems, and matured faster. The authors reason this is due to the carbon nanotubes facilitating water intake, however the evidence provided doesn’t prove this beyond a reasonable doubt. Although I wouldn’t recommend eating these tomatoes just yet, one could still use the increase in plant biomass and efficiency for biofuels and related projects.

Link to paper: Carbon Nanotubes Are Able To Penetrate Plant Seed Coat and Dramatically Affect Seed Germination and Plant Growth

Mitch

The chlorinated sugar substitute called sucralose (commercially marketed as Splenda (TM)) was first synthesized in 1976, as part of a collaboration between Queen Elizabeth College in London and the Tate and Lyle Chemical Company. It is manufactured by the selective chlorination of sucrose, in which three of the hydroxyl groups are replaced with chlorine atoms. Supposedly the graduate student, Shashikant Phadnis, working on the synthesis misunderstood his professor’s request to test the chemical as a request to taste the chemical. Just goes to show, sometimes to make a lucrative discovery, a chemist must take the ultimate test!

Whatever happened, it has been found that Sucralose is approximately 600 times sweeter than sucrose, and since being introduced in the USA in 1998, has become one of the leading sweeteners on the market. One of the main reasons for this is that studies have shown that sucralose is highly stable; it doesn’t break down easily due to heat so cooking with it is safe. It also doesn’t dechlorinate over time, photo degrade under visible light, or biodegrade with common bacteria. It is also very insoluble in fat cells, so all of us Americans don’t have to worry about getting a heart attack on the treadmill (at least not from sucralose!). In fact, sucralose is so darn stable, it doesn’t even get broken down in waste treatment plants.

Meet Smitha Ramakrishna, a senior at Corona del Sol High School in Chandler, Arizona, who has been doing research at Arizona State University about sucralose’s inability to be broken down and how this make affect the environment. At only 17 years of age, she has been researching sucralose for nearly 2 years, as part of her greater goal of trying to help with global water issues. She also founded an organization named AWAKE, which is dedicated to increasing her community’s awareness about water-related issues.

She has found that after subjecting sucralose to treatments similar to those used by waste water treatment plants, the sweetener resisted bacterial digestion. Only after a long time and under UV irradiation in the presence of high concentrations of titanium oxide (TiO2) did the sugar break down. Considering that few plants use these methods, the majority of sucralose in wastewater enters the ecosystem. She doesn’t say for sure what effect this will have, but says that preliminary studies suggest high concentrations of sucralose may poison fish.

See more here: That Splenda you’re drinking will be in our water supply for a while

Personally, I think people should use xylitol more. First studied in the 1970’s, almost no negative effects have been found due to ingestion of even 400+ grams a day (imagine 400+ grams of sugar! BLECH!) and many positive health effects have been proven ranging from plaque-reducing effects to boosting your immune system. It is about as sweet as sucrose, and has 2/3 the caloric content.

That said, I am still gonna go get me a coke zero.

Starting from the human estrogen receptor α, the authors employed directed evolution: they changed the residues in proximity of the ligand by mutagenesis, screened the resulting mutants, and selected the best receptor mutants for the next round. After the third round of directed evolution, they came up with an optimized mutant that bound to CV3320 with an EC₅₀ of 55 nM, while the affinity to 17β-estradiol was reduced by a factor of 10 (4 nM).

While the authors admit that the selectivity over 17β-estradiol could still be improved, it still is a nice piece of work that demonstrates the power of directed evolution. This way, a protein receptor for a substrate that does not occur in nature can be made. Such a receptor can be used to make biological switches.

For those of you who don’t know, Dr. Joe Vinson is iconic to the chemical community (believe it or not, even more so than Soderquist).  The American Chemical Society frequently hosts his seminars on some of life’s guilty little pleasures, coffee and chocolate.  I recently had the chance to sit in on his “Science of Chocolate” seminar.  And after and hour of lecturing about the history and chemical make up of chocolate, he took questions from the audience.  When I used to housesit for my aunt, I remember her telling me to be careful not to feed the dog chocolate because it could kill them.  I also recall coming across a warning by the ASPCA about the dangers of cocoa bean fertilizer. 

With my curiosity, I decided to ask the expert.  “Why is chocolate toxic to dogs?”  There was a bit of laughter behind me after I posed the question.  Vinson claimed that the theobromine was responsible.  “You would think that for a 100 pound dog it would be okay to feed them chocolate safely.  But you can’t.”  He then took the next question while I sat there completely unsatisfied with the response. 

So (like my daschund and miniature pinscher) I went digging.  Despite the name, theobromine has nothing to do with halogens.  Theobromine (or more IUPAC-y, 3,7-dimethylxanthine) is a structural derivative of caffeine.  In fact, several species of plants synthesize caffeine by converting xanthosine into theobromine.  The biosynthesis is concluded by N-methylation of theobromine by caffeine synthase (using S-adenosyl-L-methionine or SAM).  Recently, Crozier and co-workers mentioned that several groups have reported identical biosynthetic routes to caffeine (Coffea Arabica – coffee; Camellia sinensis – tea; Theobroma cacao – cacao; see Phytochemistry 2008, 69, 841-856).  At any rate, both theobromine and caffeine are stimulants (caffeine much more so). 

It appears that theobromine metabolism has only been moderately studied in the scientific community; most research has revolved around human metabolism.  Arnaud and Welsch (two research chemists at Nestlé in Switzerland) used ¹⁴C-labeled theobromine to determine the metabolic breakdown of the alkaloid in rats (J. Agric. Food Chem., 1979, 27, 524-527).  They determined that theobromine and methyl uracil were the major radioactive components in the urine (accounting for 85% of total radioactivity).  Other side products included 7-methylxanthine, 7-methyluric acid, 3-methyluric acid and several others.  Interestingly, they noted large similarities in the chemical composition of urine samples in both humans and rats that had been given theobromine.  However, there were quantitative differences between the two species.  Along with their paper, they actually printed pictures of 2D-TLC plates of urine samples of humans and rats.

By comparison, it appears that the canid (or canine) biochemistry for metabolizing theobromine is strangely unique relative to humans (and rats for that matter).  The consensus opinion appears to be that dogs are unable to metabolize and then excrete theobromine efficiently.  Upon ingestion of a theobromine-containing substance, dogs have been reported to excrete “small quantities of an unidentified but apparently unique metabolite” (Drug Metab. Disposition 1984, 12, 154-160).  It also appears that the toxicity associated with the inability to metabolize theobromine causes an increased concentration of intercellular free calcium, which is consistent with significant CNS stimulation and tachycardia (J. Agric. Food Chem., 2005, 53, 4069-4075).  Physiologically, theobromine ingestion in dogs is linked to epileptic seizures, heart attacks and death. 

Bottom line: stick to the peanut butter.  It’s much safer.

Anyway, as the token biologist, I wanted to bring your attention to a commentary in Cell (doi:10.1016/j.cell.2008.06.013) describing the role of Pixar-style computer animations in the future of biology education. Although the article is an interesting read, what I really wanted to show you all is the author’s website (www.molecularmovies.org) which houses a collection of these animations. (Warning: visiting this site can be hazardous to your research productivity)

There are tons of other really great computer animations on the site (though some are not so great in terms of explaining things. Alas, the one video related to organic synthesis falls into the not-so-great category). My personal favorite is the movie on apoptosis (programmed cell death), which features one of my favorite protein complexes, the apoptosome (or as I like to call it, the seven-membered ring of death).

Now, as a biophysicist, I think that these videos are great because they illustrate some very important concepts in biology. The apoptosis video shows how many processes in biology resemble overly complex Rube-Goldberg Machines. Other videos on the site, especially those by Drew Barry, offer a glimpse into an important field of research: protein dynamics. Chemists are used to thinking of catalysts as fairly static entities. Sometimes a catalyst can be as simple as a surface that acts binds a reactant and primes it for subsequent reaction. In contrast, the catalysts in biology, enzymes and ribozymes, are rarely static. The video on DNA replication (video available at the WEHI website) shows the dynamic nature of these biological catalysts. The animation shows the E. coli replisome, a large multienzyme complex, as it copies DNA. The components of this complex have a number of different enzymatic activities that all need to be synchronized and coordinated in order for replication to proceed. Despite all these complicated interactions, the E. coli replisome proceeds at a rate of about 1000 nucleotides per second and with an error rate of about 1 per 10⁹ nucleotides.

Of course, one has to remember that these videos are animations, not realistic simulations. While they are based on empirical results (e.g. crystal structures, biochemical assays, single molecule experiments), the animators do take some creative liberties with the videos. For example, I doubt anyone has observed buzzing and clicking sounds that accompany Brownian motion and enzyme catalysis in many of these videos. Indeed, the animators (with good reason) don’t show two concepts that are becoming increasingly important in understanding biological dynamics: the stochasticity of events in the cell (e.g. polymerases don’t move along at a constant rate) and the very crowded environment of the cell.

Most significant, however, is that while many of these videos depict the dynamics of various enzymes, not much is known about the actual motions of these enzymes and enzyme complexes. When biologists discuss conformational changes, these protein movements are often identified by looking at static “snap-shots” of an enzyme in two different stages of a reaction. Rarely are the kinetics of the transition measured directly, and the techniques that can directly observe conformational changes (e.g. Förster resonance energy transfer) give limited spatial information. Furthermore, the single molecule experiments that give arguably the best kinetic information about enzyme catalysis and protein motion often have limited temporal resolution (it’s hard to go below the millisecond time scale). Computational methods (e.g. molecular dynamics) can give detailed videos of molecular motion with both high spatial and temporal resolution, but modern computers can simulate only tens of nanoseconds, orders of magnitude below the timescale of most large protein motions. NMR spectroscopy has the advantage of being able to access a large range of time scales, but NMR measurements are limited to small systems and can access only dynamics of an enzyme in equilibrium. Being able to somehow synthesize and connect the information from timescales ranging from bond rotations and vibrations to conformational change and allostery is a tough task, but doing so may offer huge insights into the fundamental chemical and physical principles governing enzyme catalysis. Recent attempts to do so (Henzler-Wildman et al. 2007, doi:10.1038/nature06407, doi: 10.1038/nature06410) have been very promising, though there is still much work left for us biophysicists.

The video is an advertisement gimmick for Thermo Fischer Scientific, obviously enough . A link to the original video can be viewed from this web page: http://www.biocompare.com/video/thermo/smi/

[1]: Not saying, I have a background in metabolite identification, but I do have more experience with it than you might expect for a nuclear chemist.

Note 1: Also covered by Closeted Chemistry
Note 2: Made an entertaining game room for the graduate student procrastinators in the audience: http://www.chemicalforums.com/index.php?board=62.0

Mitch

Quinine was originally used by the Incas to treat malaria, and was later used throughout the world by conquerors from the Europe (a couple of other blogs have been discussing the merits of folk remedies versus pharma developed drugs: @The Chem Blog,  @Chemical Musings).  Anyway, this compound has saved countless lives, although now other remedies (e.g., chloroquine) have replaced quinine as a usual malaria treatment for cheapness and synthetic accessibility.  Resistance to chloroquine may put quinine back into the spotlight, however. Beyond its importance as a medicinal compound when isolated from natural sources (the bark of the cinchona tree), this has been a fascinating molecule for synthetic chemistry.  Most of the history of this story can be found in chapter 15 of Classics in Total Synthesis II, possibly the best chapter in either of the Classics books.  I’ll summarize some of this history here. Let’s begin with Hofmann, who decided it might be possible to synthesize quinine from components of coal tar, and he talked his student, Perkin, into trying this.  The idea was to take two equivalents of N-allyltoluidine (C₁₀H₁₃N) and three atoms of oxygen and, since you have the right number of all the atoms you would need for quinine (C₂₀H₂₄N₂O₂), they might spontaneously assemble and make the natural product (with water as a byproduct).

Those of us who are familiar with total synthesis will recognize that this is a low-yielding reaction.  Perkin ended up with a bunch of tar.  When cleaning his glassware with alcohol, he found that a purple compound was extracted from the tar, and this could effectively dye cloth a royal purple color.  The dye, mauveine (actually a mixture of two compounds), led to Perkin becoming a very rich man. Around the same time, Pasteur found that treating natural quinine with H₂SO₄ led to a different compound, now known as quinotoxine.  In 1918, Rabe reported the conversion of quinotoxine back into quinine.  Then some 25 years later, the great R. B. Woodward and his post-doc Doering synthesized quinotoxin, thereby completing a formal synthesis of quinine (details on the route here).

Now it gets interesting.  This synthesis was a landmark for Woodward, and would certainly ensure that he get a tenured faculty position at Harvard.  However, there arose some questions about the validity of the formal synthesis, because the work of Rabe had not been repeated in Woodward’s lab.  There is a fantastic review in Angew. Chem. by Seeman which investigates this debate at length.  I highly recommend reading this article.  It’s 30 some pages, but worth every letter (DOI link also featured in C&EN here).  Seeman ultimately concludes that Rabe did in fact convert quinotoxin to quinine in 1918, but these results may be difficult to reproduce since the experimental details are not very extensive. This is a very interesting example of prominent figures questioning the validity of results reported in chemistry journals.  This of course has been a hotbed of activity recently in light of Sames/Sezen-gate and hexacyclinol-gate.  The difference is that now RBW is not around to explain his actions and decisions.  Seeman did interview Doering (now an emeritus prof. at Harvard) and did get some insights.  It is hard to say with certainty with a 60 year gap in the record. We should all learn from this story.  Chemistry is done by human beings, and that can be a good thing or a bad thing.  Was Woodward knowingly skipping over steps he knew would be difficult to reproduce, if they were reproducible at all?  Was this a situation where the most important factor was publishing in order to get tenure?  We can’t know what was going through his head.  Another point that Seeman makes, which is perhaps the most powerful in the whole debate, is that it is astonishing how quickly opinion turned against the Woodward report.  As we get more and more skeptical of published results, we also run into the danger of becoming too quick to judge something false.  The suggestion that results may be fabricated are certainly not a conviction, and the community must keep that in mind.  Suspicious results are one thing, proving them wrong is quite another. Since the Woodward route was published, several others have appeared, notably one by Stork, who was one of the principle figures in questioning the validity of the Woodward/Rabe route.  Each of these syntheses is a great achievement.  Over the years quinine has touched the fields of medicine, synthetic dyes, politics, and ethics.  See, chemistry and history aren’t all that different after all! By the way, the Stork paper has the greatest abstract of all time. – movies

On to the post. This was inspired by a recent paper by Dickerson and Janda (at Scripps) regarding a catalytic antibody approach to the treatment of addiction (ACS link). Now I am no chemical biologist, so I admittedly have not followed any of the catalytic antibody literature, but here is my understanding: these antibodies are just like the normal antibodies our bodies produce to fight of infection and the like, except that these antibodies do more than just bind the offending molecule, they catalyze a chemical reaction changing the structure and, hopefully, the bioactivity/degradation of that molecule. Catalytic antibodies are great for pharmaceutical applications because they avoid many of the toxicology problems that plague non-natural therapeutics.

This is a powerful concept for addiction therapy since, for many drugs, the specific molecule responsible for the intoxicating effect is well known (e.g., LSD, cocaine). Although this idea is striking, what really caught my eye was the selection of the target in the Dickerson and Janda work: delta⁹-tetrahydrocannibol (THC).

I love to read the intro paragraphs of papers like this because they always talk about the sweeping socioeconomic effects of whatever particular ailment they are targeting. As someone who is in a first-world ivory tower academic institution, most of the time the reality of these diseases and effects are at arms-length; they do not directly touch my life. So this topic really struck me because everybody knows someone who just smokes weed all the time. Say what you want about the way some people choose to live/waste their lives, I have always thought it is not really my place to judge these people too harshly. More annoying to me is that a quick search of the Internet will give you a laundry list of reasons why marijuana should not be a controlled substance because, for one reason or another, some people think it just isn’t that bad, some even saying that it is not addictive (One from each side: DEA (pdf file), NORML). I have read all the arguments for legalizing marijuana, and I have not found one that really convinces me; it is still an intoxicant with demonstrated addictive properties (don’t give me the “so is alcohol” straw-man argument either).

As a scientist, I would really like to see some facts on the matter, and the intro to this paper gives some great scientific data on the subject. To paraphrase: the now famous “cannabis gateway hypothesis” does, in fact, have some real experiments that show that delta⁹-THC specifically changes opiod receptors in the body, perhaps making them more susceptible to other drugs (link to that study). Not proof positive, but a start that is not just based on surveys and statistical manipulation. To summarize the results in the paper, Dickerson and Janda have identified catalytic antibodies which do effect oxidative degradation of delta⁹-THC, mostly to cannabitriol (which is not the predominant metabolite in humans). Some detailed kinetic studies suggest that these antibodies may drop the effective half-life of delta⁹-THC from ~20 hours to less than 40 minutes. They have some preliminary biological data on the effect of cannabitriol (which incidentally is also a naturally occurring compound in cannabis) but they need some more extensive studies to shore up the efficacy of this approach in reality.

I guess my point is that while at one end of the spectrum pervasive drug use has slowly become a more acceptable part of our society, I am very happy to see that there are prominent scientists looking to address these problems as they would any other ailment. Kudos to these researchers for taking on an unpopular problem.

- movies