Saturday, February 6, 2016

Petition on Petitions (aka "Leave IUPAC Alone!")

I've already dedicated way more time to 'lemmium' than the movement warrants, and the researchers at JINR, ORNL and LLNL as well as the good folks at IUPAC don't need a proxy without a shred of authority like me to fight their non-existent battles. So go ahead and call me the Chris Crocker for IUPAC. I don't have strong feelings on Lemmy Kilmister or Motörhead one way or the other, so I don't want to sound like I'm trying to besmirch anyone's music or legacy. This is all about a flawed premise and hypocrisy. The most recent effort by is to get teachers and academics to sign a letter to IUPAC asking them to change the guidelines for naming elements to include "Any person by whose honouring the causes of science in education would be furthered." Sound great right? What educator could be against something that promotes education? Like every other educational initiative these days, it's written in STEM Tourette's. As Chemjobber is most adamant about pointing out, STEM shortages are mostly myth.

To their credit, the folks behind lemmium did their research and found out that IUPAC element naming guidelines would invalidate their original petition during the almost 1 month that naming elements has been a cornerstone issue for them. Thus the need to lobby IUPAC to change the rules before petitioning IUPAC to name element 115 'lemmium'. The problem with the premise of the "furthering education" letter, is that never mentioned education before a few days ago. Since it's inception during the first week of January, this movement has always been about the music and Lemmy. No problem being a fan, but the education angle didn't crop up until February 3. Check their timeline. This letter is a misdirection and a means to the actual goal.

In the 21st century, we've come to appreciate the rights of artists (musicians, filmmakers, etc.) to their creative works. There was a lot of debate during the Napster "file sharing" era about whether musicians should be compensated (They should. It was stealing) and artists regularly ban politicians from using their songs. Producers and studios sometimes try to interfere with an artist's art, but that almost never results in a better product. If anyone ever petitioned a band to name one of its songs, would anyone expect the artists to bow to public pressure? This is essentially what petitioning IUPAC (or the researchers) is. The researchers working on elements 113, 115, 117 and 118 aren't mining for an unknown substance in the ground that's already there, they're using ingenuity and creativity to design experiments. I posed this question about creative ownership to @LemmiumMetal after they engaged me on a tweet about their letter. This was part trolling, part serious question, but as you can see, they never answered the question and then admitted the letter was actually about making their original (not actually motivated by education) petition feasible. Within this same conversation, another user asked "shouldn't science belong to people? Or should it stay enclosed in its ivory tower?" The use of the pejorative "ivory tower" insinuates that scientists (academics) believe they own something that rightfully belongs to everyone. This is a false equivalency between science and the procedure for naming elements. The doors of science are wide open to whoever wants to enter. That doesn't mean you get to show up at the 11th hour and claim ownership in an effort that has cost dozens of people years of blood, sweat and tears to achieve. 

So aside from the distortion in motivations, is there reason to believe naming an element for educational purposes would have the desired effect? There was some simultaneous discussion going on with Kat Day about IUPAC's rules for acceptable name sources. To review IUPAC permits "Elements can be named after a mythological concept, a mineral, a place or country, a property or a scientist". We're currently engaged in a game of guessing what names the researchers might choose, and a more detailed commentary on how those predictions were made (sorry, no spoilers). If you look at trends in element naming, all of the elements discovered in the last 60 years were named after either a scientist or a place. Mineral-based names were popular when rare earth elements were being separated from ores. Since that's not the process involved in finding transuranium elements, we can exclude that from the discussion. The last time an element was named for a property was 1940 (astatine from the Greek word 'astatos' meaning 'unstable') and for a mythological concept was 1945 (promethium for the Greek god Prometheus).

IUPAC's selection of categories are a bit ex post facto as all but 32 elements were already named before the organization formally existed in 1919. The actual need for codifying element naming procedures didn't become apparent until the 1940s with the development of the technologies necessary to synthesize short-lived elements that don't occur naturally. In 1947 Friedrich Paneth outlined the ideas that would become IUPAC's guidelines, which have been modified as warranted by various discovery priority controversies. The mythology provision has fallen in popularity as the adherence to Latin and Greek as the languages of science has diminished. So is there room to add a provision to IUPAC rules to use non-scientists in place of minerals and mythology, which are likely to fall into disuse?

The letter states the goal is to use lemmium to "appeal to 11–16 year olds". I question how many 11–16 year olds care about Motörhead right now, let alone in 10, 50 or 100 years. As an astute tweeter pointed out, bieberium would likely be more popular today. Some teenagers like hard rock (how Lemmy actually classified his music, not heavy metal), but a lot more like the Top 40s. Some even like country (something may be wrong with them) or alternative. The appeal of any one artist is not universal, nor is any pop culture possibility. Imagine this debate around the time that element 103 (lawrencium) was being named in 1961. Buddy Holly had died just two years before, so with the proposed changes 'hollium' or 'buddium' would have been valid homages to a musician that would appeal to the youth. You could even make the argument that the name "fits". Buddy Holly was gone too soon at 22, and element 103's longest live isotope has a half-life of 11 hours. Do today's youth know (or really care) enough about Buddy Holly that it would make them think twice about the periodic table? At best, most of them have heard the Weezer song, and there's a good chance that happened on a classic rock station since it was released in 1994. Over 20 years ago. Scary, yes? The problem with lemmium, and this proposed change in general, is that people want to graft their interests onto others under the guise of education. I seriously doubt it would work. Although perhaps not always successful, the motivation behind the IUPAC guidelines is to have names with some universal meaning.

Do we need pop culture to get young people interested in science? Dinosaurs are often a gateway to science for young people, but dinosaurs don't need names derived from music, fiction or video games to draw interest. Space exploration is another area that appeals to the youth, but how many young people would list any of the astronomical bodies named for pop culture icons if they were asked what got them interested in the universe (which has literally has more things in it to name than we ever could)? Our culture is obsessed with celebrity. Do we the entertainment industry to permeate yet another aspect of life in the hopes it will move the education needle a hair? Chemistry does need to do a better job appealing to young people. This has been discussed before. Chemistry needs a personality like Neil deGrasse Tyson to catalyze the process. Academic and industry chemists need to be in schools showing students the amazing things chemistry does. Those are the things that got me, and many other chemists, excited about doing chemistry. It will still work. If a celebrity wants to lend their name and time to the effort, welcome aboard. Just don't expect science return the favor by permanently enshrining you in element form.

Thursday, January 14, 2016

Dr. Bandwagon or How I Learned to Stop Worrying and Love the Cyclotron

Two weeks ago IUPAC announced the Discovery and Assignment of Elements with Atomic Numbers 113, 115, 117 and 118. The term "discovery" is a little misleading with respect to the superheavy elements. It's not like these elements were unexpectedly found at the bottom of a nuclear reactor or by painstakingly searching for new primary constituents of exotic metallic ores. Instead the researchers carefully designed experiments and bombarded nuclei with the hypothesis that certain combinations of lighter elements would fuse into superheavy atoms. So, the discovery is really only whether their synthetic hypothesis was correct or not. This does not diminish the significance of the outcome, but it feels like the connotation of "discovery" downplays the intellectual role of the researchers who did the experiments. Nevertheless, the nature of words and what we call things engenders a lot of strong opinions, especially the names of new elements. IUPAC, as arbiter of element names (and names of things in chemistry in general), is well aware of this especially after the Cold War era Battle Royale between research groups in the United States and Russia over the right to name elements 101 through 106.

The press release by IUPAC produced an immediate fervor in both the scientific community and the popular press. This is certainly a chance for chemistry (and physics) to take center stage in the public eye, so no one should blame anyone for taking full advantage of the opportunity. That goes double for publishers like Chemistry World and Nature, who have the chance to engage an audience that might ignore any number of other science and chemistry stories. In the excitement though, some people are overestimating their role in the process of naming elements.

First, the ridiculous suggestions. There are currently petitions that have received attention in the popular media to call 2 of the new elements 'lemmium' to honor heavy metal musician Ian 'Lemmy' Kilmister of Motörhead, and 'octarine' to honor Terry Prachett's Discworld series. At least the creators of the octarine petition are seeking an audience with the right people (JINR and LLNL) who actually get to suggest a name. The lemmium petition is directed directly toward IUPAC who only sanction/approve names suggested by the discoverers (although there is a clause stating the IUPAC can select a name if an appropriate suggestion is not made, which seems highly unlikely). Neither of these proposals meet IUPAC's guidelines for new element names. The rules state: "Elements can be named after a mythological concept, a mineral, a place or country, a property or a scientist". Unfortunately, "rocking hard" isn't exactly what IUPAC meant by a "property", so 'lemmium' is out. One could try to argue that 'octarine' is a mythological concept, but it's hardly the equivalent of 'promethium' named after the Greek god Prometheus. Check back in a few hundred years. Octarine gets traction now because the Discworld books are very popular, and Pratchett died recently. The periodic table is timeless. We don't know yet how timeless Discworld will be (sorry, folks). 

These suggestions are also counter to any suggestions the researchers might even brainstorm on a whiteboard. The initial reports on these elements starting appearing in the mid-aughts, so they've been waiting nearly, or more than, a decade for the independent confirmations of their experiments and approval by IUPAC needed to suggest an element name. Researchers also had to overcome serious obstacles to even try their experiments such as making and purifying radioactive, fairly shorted lived super heavy starting materials. With all the years, people and resources involved in this effort, does naming an element after a mid-tier musician with a well-documented substance abuse problem and a collection of Nazi paraphernalia even sound reasonable?** Can you even imagine that in the 10 years or so that they've been waiting to suggest an element name that the researchers haven't kicked the tires on a few ideas?

This gets to the central point. The researchers making new elements are doing difficult research. Coming up with a name is easy. They're really smart and creative people who don't need help with names. It's presumptuous to think any one us should have a say in what they chose. When I (Shawn) was interviewed for the ACS Reactions video, I was asked if I had a suggestion for a new element name. I answered no. Never in my wildest dreams did I think anyone would interview me about how elements were named, let alone be in a situation to name an element. I've had a longstanding interest in nuclear chemistry, but since I chose to study a different area, I abdicated any minute chance I had to name an element some time ago. In fairness, we had quite a lot to say about the structure of element names, and the new IUPAC naming guidelines. We specifically suggested modifying the naming conventions for group 17 and group 18 elementsthe halogens and noble elements; however, we never suggested any element names. Okay, we did suggest that 218Rn could beknown as astaton in jest, but no one seems to have listened.

In addition to ludicrous pop culture-inspired names, there have been calls to correct historic gender bias by naming new elements after women and a denouncement of names based on nationalism. We certainly would champion the choice if any of the research groups chose to honor one of the many women who have contributed to the expansion of the periodic table; however, it is unfair for anyone to graft their cause onto researchers who may be equally passionate about other worthy options. There's almost an implication that if researchers don't adopt some "altruistic" name that they will have done a disservice to science and the world. No where is that more apparent than the discussion of nationalistic names. Philip Ball has suggested 'levium' after Primo Levi, a Jewish Italian chemist and Holocaust survivor to send the message that the "periodic table is for all humanity". On the surface, these kinds of ideas sound inclusive, but given the pre-condemnation of a nationalistic Japanese element name from many quarters, even suggestions made in good faith can come off as sanctimonious.

Actually, the periodic table should not be a billboard to promote any cause; though it has unfortunately been exploited in the past. The most nationalistic names are from the late 19th century—germanium (Germany), gallium (France, from the Latin), and polonium (Poland), all named by a scientist from the respective countries. Francium followed this trend in 1939. These four elements stand out today as very nationalistic names.

There are other elements with regional names—scandium reflecting the pan-Scandinavianism enthusiasm of the 1870s. Europium and americium can be taken as referring to the continents rather than any one nation. Any nationalism behind 1828’s naming of ruthenium (Russia) seems weak since it reflected the location of discovery, not the discoverer. There is a bit of vanity in the city and local region element names. Invariably these reflect the location where the element was discovered: lutetium (Paris), holmium (Stockholm), hafnium (Copenhagen), berkelium (Berkeley), dubnium (Dubna), darmstadtium (Darmstadt), rhenium (Rhine River), hassium (Hesse), californium (California), livermorium (Livermore). These don't strike us as nationalistic. In addition, there are yttrium, ytterbium, terbium, and erbium, all named for Ytterby. In defense of this, a lot of elements were discovered in ores found there and ytterbium was named by a Swiss chemist. If the Swedes wanted to be nationalistic, it seems unlikely they'd chose the name of a village on a small island where quartz and feldspar were mined to make porcelain. Hardly the stuff of legend. Besides, how many people realize that hafnium references Copenhagen, Denmark?

There have been rumors that the Riken research group now credited with element 113 might choose the name 'japonium', and some have criticized this as nationalistic. While it's true that japonium would stick out like germanium, gallium, polonium, and francium, what is the deep rationale for criticizing japonium, other than "fairness" to those who had nothing to do with the work behind creating element 113? Japonium is still a rumor remember. In fact, "nationalistic" may not the right way to see any of these elements named for nations, regions, or cities. At worst, they are vanity names, but they celebrate where the work was done, where the element was first found or created. They tell part of the story behind the tablein a way that lemmium or octarine would not.

At the risk of sounding hypocritical, if the Riken group chooses to honor their homeland with an element name, we hope japonum is not the only one being considered. 'Nipponium'*** or 'nihonium' would reflect what the Japanese call their nation not an anglicized exonym; however, IUPAC credited element 113 to Riken, and they don't need our help either.
**Motörhead fans can direct all their hate tweets about this statement to @scburdet as @geochembrett has nothing to do with it.

***Nipponium was once suggested for element 75 (rhenium), but the discovery was later disproved. IUPAC has a restriction on reusing a name that has been "used" before, so 'nipponium' might be rejected. However, this clause has never been invoked. If the name only appears in a paper, but was not adopted or widely utilized, we believe it would still be acceptable.

Tuesday, January 12, 2016

Doomed to repeat history

I still "owe" the world and the EIC of Applied Materials and Interfaces a whistleblower-style disclosure of all the correspondence related to the Fe3+ sensor paper discussed in my previous posts (1, 2, 3). The problem is that I need to carve a few days out of my schedule to document everything properly. In the interim, I came across a couple of related papers recently that bear examination in more than 140 character tweets.

In reverse chronological order, today I saw the ASAP notification of this paper by Magri in RSC Advances. RSC Advances has been a significant contributor to potentially bogus Fe3+ sensor papers in the past, but seems to be aware of the problem now as evidence by this paper that has been in ASAP limbo for >7 months (tangent - there are ASAP papers there from 2013). Magri's paper follows a similar path to my groups study on another popular Fe3+ sensor. Magri's results demonstrate that the earlier study (cited 59 times according to Google Scholar) makes incorrect conclusions about the mechanism of fluorescence signal transduction. In short, the fluorescence quenching is not due to metal ion binding to the sensor's receptor, but rather inner filter effects stemming from Fe3+ inherent incompatibility with water at pH >4. I'd encourage people to read both papers because it will give great insight to the types of problems plaguing the fluorescent sensor field. The original Tetrahedron Letters paper lacks any absorption spectra, which would provide direct evidence of the expected inner filter effects however. Magri does the fluorescent sensor community a great service in publishing a thorough investigation like this.

This leads me to an ASAP Applied Materials and Interfaces paper that ruffles my feathers. The paper claims a signal transduction mechanism that involves forming a 5-coordinate Fe3+ complex with monodentate phenolic ligands in water (unbuffered, deionized). There is a component of the project that involves nanoparticle upconversion of light to excite the Nile Red fluorophore; however, at its core this system is essentially on a small molecule sensor. The fluorphore-ligand dyad (Nile Red-phenol) is reported to bind Fe3+, which quenches the fluorophore emission. Whether the excitation of the fluorophore is direct or from upconversion is not particularly relevant to the sensing mechanism. There are several red flags that suggest this cannot be the actual mechanism:
1. In addition to Fe3+ being incompatible with non-acidic water, monodentate phenols are unlikely to have strong interactions with Fe3+ in the presence of excess water ligands (unlike catechols, which take advantage of the chelate effect). It's even less likely that FeL2, FeL3 and higher order species will form in high concentrations. You will not find many (any) inorganic chemists who will buy into the author's coordination chemistry. Furthermore, evidence for these complexes being reasonable is demonstrated in non-aqueous solution (in DCM, SI Figure S8). Changing the solvent completely changes the  coordination chemistry and hydrolytic stability of the Fe3+ ion.
2. The absorption spectrum (Figure 3) has the hallmarks of particulate formation/light scattering. The individual spectra increasingly deviate from the baseline as Fe3+ is added
3. The procedures for working with Fe3+ in cells do not match the recommended protocols. Adding Fe3+ to the extracellular fluid does not result in an increase in free intracellular Fe3+. I do not believe anyone has studied this in enough detail to know what happens when you do this. I suspect a broad stress response (which could lead to all kinds of intracellular changes) and/or uptake of iron nanoparticles by endocytosis; however, this is speculation.

Clearly, the authors see a change when adding Fe3+, but their mechanism is wrong. There are many possibilities - inner filter effects, formation of iron nanoparticles, pH-induced changes from Fe3+ hydrolyisis. Every published study that fails to account for inorganic and photochemisty just perpetuates the myth that these protocols for handling Fe3+ in aqueous solution and cells provide valid results. This facilitates the publication of the next dubious study. I would alert the journal to this problem before publishing this blog; however, they've made it abundantly clear through our previous correspondence that they do not care whether or not papers published in the journal contain conclusions that are consistent with well-establish chemical concepts. 

Tuesday, September 29, 2015

Stacking the deck with images?

I wish that I could solicit input on this question as an unbiased observer. Anyone who's read my previous posts knows that I am anything but. I considered presenting this data absent the source, but anyone with 1% of Sherlock Holmes' sleuthing ability would track down the paper I am referring to in a nanosecond.  Going back and forth with the EIC at ACS Applied Materials and Interfaces recently (maybe more on this later) forced me to look at aspects of this paper in greater detail than I would have otherwise. As I documented before, one of the reviewers fixated on the cell imaging aspects as a defense of the results, and a rationale to reject my Comment. In preparing an appeal, I looked more carefully at the cell images

Here are cells treated with Fe3+ and probe:

The difference between a) and b) is 1 hr of incubation time. Ref: ACS Appl. Mater. Interfaces20146, 18408–18412

Here is the control of cells treated only with probe:
The difference between d) and e) is 1 hr of incubation time. Ref: ACS Appl. Mater. Interfaces20146, 18408–18412

Notice any differences? I am not very experience with growing cells, but to me it looks like the confluency is much higher and the cell size is larger in the images of the iron-treated cells (Figure a-c), than the sensor-only cells (Figure d-f). This could create the illusion of a greater fluorescence response in the iron-treated cells. There is simply more sensor present per unit area, and there is clearly basal fluorescence. In fact, I might argue that the sensor-only cells (f) are not even healthy compared to the Fe3+-treated cells (c). It also appears that the fluorescence images of the sensor-only cells are out of focus. Even the brighter cells in the middle of the field look fuzzy. Again, this could make the response look more dramatic than the reality. There are other problems with this experiment related to metal transport, but I don't want to conflate imaging with biology. I have a biological interpretation backed by literature for what they observe in the Fe3+-treated cells.

I am interested if anyone concurs, or if I am the victim of confirmation bias. There have been several notorious examples of image manipulation including cut & paste to make nanochopsticks and reusing old data. This isn't that kind of malfeasance, but could it be stacking the deck to fit a preconceived model? How say you?

Friday, September 11, 2015

Fe(III) sensor saga continues

A few months back, I blogged about a paper on iron sensing by Belfield in ACS Applied Materials & Interfaces, and my efforts to bring my concerns about the validity of the conclusions to the attention of the editor. While those efforts were failing to get traction, I brought up similar concerns about a paper in EurJIC. The editor at EurJIC requested that I write a peer-reviewed "Correspondance" on the issues after the author was unable to satisfactorily respond to the criticisms in private emails. Having started the correspondence process with EurJIC, I realized this was also the only recourse I had left at ACS-AMI. A similar mechanism was mentioned in passing by the EIC of ACS-AMI as the only option once a paper was published in the journal, so I decided to submit a "Comment" to ACS-AMI

Obviously, since both papers deal with Fe(III) probes, there are some similarities in the problems as in many other published reports. There are some notable differences as well. In short, the EurJIC paper utilizes PBS buffer and reports a fluorescence quenching mechanism. I indicated evidence of particulate iron and speculated about alternate explanations for the fluorescence changes. The ACS-AMI paper reports fluorescence enhancement in unbuffered water, which I concluded is a clear indication of protonation instead of Fe(III) binding. I very confident in my critique of the ACS-AMI paper because this is exactly the same problem that my group investigated previously with another (very similar) probe. I received a rejection of the comment Friday. Two positive reviews, both suggesting minor revisions, and one rejection. 

Although anonymous, I have suspicions that the rejection was written by an author of the original paper. I conclude this because 1) the authors should have an opportunity to respond in such a situation, and 2) the phrasing of the arguments. I wish that I was surprised that the EIC decided to side with the single "Do not publish" referee and didn't leave an opening for appeal. From the beginning I surmised that the EIC had little interest in criticism of the paper. As a reminder, the lead author of the paper in question is a member of the ACS-AMI editorial advisory board. The dissenting referee made some flimsy arguments against Fe(III) hydrolysis, and focused on the "validity" of the cell imaging studies as a reason to reject the Comment. I specifically did not address cell imaging because sources of false positives in imaging studies are elusive, and more often unrecognized. In the Cu(I)-sensing field, as well as sensing for other metal ions, colloidal aggregates are just recently being recognized as a significant (if not primary) reason why many probes respond  in cells. This is one possible explanation for the imaging results using the Belfield probe, but it is equally likely there is an unknown mechanism that can produce false positive in Fe(III) systems.

The EIC further cited non-disclosure of the Correspondence at EurJIC as an additional rationalization for rejection. This appears to be a thinly veiled insinuation that I behaved unethically. To be clear, no attempt was made to conceal the existence of the Correspondence. I didn't cite the unpublished Correspondence in the Comment due to the lack of a journal citation at the time of submission, as well as the differences in the two systems. I went to great lengths to perform a non-repetitive analysis, even though the underlying issues of Fe(III) hydrolysis are identical (e.g. I needed to cite the same pKa values for hydrated Fe(III) in both articles and it's difficult to express this uniquely). Owing to the different types of experiments performed in each case, I believe that the Comment highlighted a different set of problems that lead to incorrect conclusions. I am not impartial in this case, but I believe people can still write on the same topic more than once in the scientific literature. Many authors, myself included, have been asked to write reviews on the same subject for different journals and therefore reach different audiences. After receiving referee reports from ACS-AMI, I can certainly imagine having revised the Comment with a citation to the EurJIC Correspondence with some additional discussion to address the referee suggestions, although I feel it would confuse the situation to make a close comparison of 2 systems that exhibit different signal transduction pathways.

Furthermore, the EIC suggested that my group should "[carry] out its own investigation on these sensor systems". If one looks at the synthesis of the Belfield probe, you quickly see that this would be an significant commitment of time. The probe requires a 6-step synthesis with a late stage macrocyclization step that proceeds in a reported 20% yield. The only positive outcome of such an effort would be disproving a published paper. A truly comprehensive investigation would require re-synthesizing and re-analyzing dozens of reported Fe(III) probes. This is not usually the kind of study that the community is excited about publishing. This is definitely not deemed fundable research by granting agencies. My aforementioned Inorg. Chem. paper that included a re-investigation of an existing Fe(III) probe as only a small component of a larger study was deemed unimportant by one referee for exactly this reason. Fortunately, that paper was handled by a diligent, thoughtful editor and was published anyway. Again, I am not impartial, but I feel the alternative resolution was proposed by the EIC at least in part because no rationale PI will undertake such an investigation in the current publishing/funding climate. So where does that leave the ACS-AMI situation? I am seeking the input of my scientific friends and colleagues. I have considered the following options:

1. Do nothing. Let this decision stand and move on to bigger and better things. Certainly the most rational course of action from one perspective. The problem with this choice is that the ACS-AMI paper has been cited by authors on multiple occasions as a defense when I critique their experimental protocols/data interpretation in manuscript reviews. This has happened several times in just the last 3 months. This is how these erroneous ideas have been perpetuated in the literature, and it will only get worse.

2. Conduct a study. In the Cu(I) field, it has been proposed that any imaging study should include a control where cells are treated with a structurally similar probe without metal binding ligands. In the case of the Belfield probe, this probably would be some dialkylanilino-BODIPY derivative. That's a pretty easy easy compound to make, but does not create a complete story without the Belfield probe for side-by-side comparison. This returns to the problem of time and resources. Also, what journal publishes such an investigation? It is highly repetitive to say that I am not impartial, but recent history suggest that ACS-AMI won't be particularly interested in this study.

3. A wikileaks-like information dump. No matter what direction I choose, I feel that it would be justified to "self-publish" the Comment here (although the information is already covered in the blog) as well as the referee reports and EIC feedback from ACS-AMI. I have nothing to hide, but what are the ramifications of such an action? Even with full disclosure, I can't escape the feeling that this is such a niche area that the impact will be minimal unlike more prominent issues that have captured the attention of chemists.

What do you think? I'd love to hear your comments and suggestions.

Friday, March 27, 2015

Tilting at Ferric Windmills

As documented last week, I contacted a journal about very questionable methods used to conclude ferric iron could be detected by fluorescence methods in water. I was already aware that this was not the only occurrence of a research group failing to properly account for the aqueous solution chemistry of Fe3+ in their studies. With a reignited hyperawareness about this problem in the fluorescence sensing field, I noticed another study making similar mistakes while catching up on recent literature this week. I alerted the EIC of this journal to the problem as well, and got a completely different response with respect to both the scientific and peer review concerns. Since this is an ongoing dialogue, I will not comment any further until the issue is resolved. This second incident prompted me to conduct a more thorough investigation of the underlying prevalence of these protocols in Fe3+ sensing.

Counts of papers containing questionable ferric sensing methodologies.
*indexed in webofknowledge as of 3/26/205.
Using webofknowledge, I searched the combination of (ferric OR Fe(III) OR Fe3+) AND (sensor OR probe) AND (water OR aqueous OR buffer) AND fluorescence. To decrease the volume of references to analyze, I limited the results to those from journals published by the ACS, the RSC and Wiley since they have a reputation for publishing reliable sensor papers (reducing the number of results from ~500 to ~150). I then went through the search results looking for evidence that the experiments would be at risk for giving spurious results. I am not sure about the exact speciation/stability of Fe3+ in mixed organic/aqueous solvents; however, my experience suggests that any significant amount of water will be problematic. I excluded several studies using 1% aqueous content in solvents like THF and methanol, as that seemed to be a relatively safe protocol. Anything with 20% or more water made the "suspicious list". The overwhelming majority claimed to work in pure water, aqueous pH ~ 7 buffer or with 50% or less organic solvent added to water/buffer. There are many fewer (not counted) that describe protocols for working and handling Fe3+ in acidified aqueous solution, and many others using pure organic solvent (also excluded from consideration). Of the 150 results, 51 contained possible problems. The results broken down by publication year and journal name are shown in the figure above.

The results are quite informative. Before the Rurack paper in 2005, there were no Fe3+ sensing papers in ACS/RSC/Wiley family of journals using water (his paper has been cited >250 times). My paper showing the errors in Rurack's aqueous results was published in 2010. The yearly trends however, suggest that the acceptance of questionable (invalid) Fe3+ titration protocols are increasing rather than decreasing. Presumably, every published paper using similar methods provides unwarranted precedence for adoption in future studies. Whether there is a connection or not is unclear, but Inorg. Chem. where my paper was published, has not published a problematic paper in the Fe3+ sensing field that I can find.

Full disclosure, there are a few papers in this collection that are difficult to analyze, particularly those from the polymer/materials literature. A more thorough investigation would be required to fully evaluate the results in detail. Furthermore, some of the studies have ambiguous or nonexistent experimental protocols in the paper and/or the supporting information, which makes evaluation difficult or impossible. If procedures for measuring/adjusting the pH are not listed, I assume that this was not a consideration. A few papers don't even list the counter ion for the Fe3+. Also, I excluded papers that may have had questionable handling of Fe3+ solutions (selectivity studies), but the Fe3+ response was not a significant component of the paper's discussion/conclusions. The 51 papers all attempt to conclude something specific about Fe3+ detection in aqueous solution. No papers from Elsevier or Springer were examined.

There are some common issues in many of the 51 papers. For example, making stock solutions of FeCl3 in water by just mixing (i.e. without adjusting the pH to <4), and titrating Fe3+ into neutral aqueous solution. Some use phosphate buffer, which would generate Fe(PO4), a water insoluble salt (assuming all the Fe3+ wasn't already precipitated as insoluble Fe(OH)3). A cringe-worthy method used in more than one paper to confirm Fe3+ binds to the sensor in aqueous solution, is mass spectroscopic detection of the ferric complex prepared in methanol. I have no doubt that they detected the complex, but Fe3+ in methanol is completely different than in water. The MS data in methanol does not confirm anything about the species that are (not) present in aqueous solution. 

What does one do in such a situation? Even if I was somehow asked to referee every paper on Fe3+ sensing, there were more papers on the topic published last year than I could possibly handle, even if they were the only kind of requests I received/accepted. A broad search suggests an upper limit of 190 Fe3+ sensor papers were published in 2014. I have no desire to comb the literature and complain to editors/authors every time I find a problematic paper. I've already published a paper in a good journal that includes a cautionary tale about these issues, but it does not seem to have permeated the sensing field zeitgeist. As I mentioned in the previous post, it's discouraging to find out that some individuals need a reminder about the relevant/underlying undergraduate inorganic chemistry. I have no doubt that researchers in other fields can point to similar problems in the literature on other topics. How does one effectively get through to journals, peer reviewers and researchers without wasting time that should be spent on other aspects of academic science? It seems antiquated in the information age that such mistakes should persist and be perpetuated, but the traditional practice of publishing an opposing research study is the only clear recourse.

Friday, March 20, 2015

Metal sensing malarkey: (or the expected virtue of ignorance)

To paraphrase Stephen Colbert, "who's not citing me now?" It's a refrain that many researchers can identify with, but only a small part of why I was motivated to write this post on peer review, journal policy and bad science. A little background first. A couple of months ago, I attended a research presentation that covered several papers on fluorescent sensors. My experience with fluorescent sensors dates back to the late 1990s and early 2000s when I completed my Ph.D. thesis on zinc probes with Stephen Lippard at MIT. At the time we started, fluorescent sensors for metal ions were still a niche area in inorganic/bioinorganic chemistry. Now, it's a widespread topic of research. As many realize, your Ph.D. work will follow (haunt) you long after graduation, even if you no longer actively work in that area. My group published a couple of papers on fluorescent probes for ferric iron 5 years ago, but that was the last time I actively worked in the area. Despite moving on to other topics of inquiry, I am still inundated with referee requests on fluorescent sensor papers, so I remain familiar with the progress and problems in the field.

The presentation focused on two papers published in ACS Applied Materials & Interfaces on fluoride and iron sensing respectively. I questioned the lead author giving the presentation about the validity of the methods and data interpretation, but failed to make any headway or get any concession that there may have been problems with the protocols or conclusions. So-called "post-publication peer review" has become rather controversial over the last few years as social media has facilitated the community's ability to discuss published science in an open forum. The scientific community becomes incensed in cases of fraud or plagiarism, although journals have often reacted more negatively to those who exposed the problems than to those responsible for the actual infractions. After consulting with several other experts on the underlying science and scientific publishing, I sent an email to the EIC of the journal detailing the problems with the papers. To be clear, I did not, and I am not making any accusation of misconduct.* Serious mistakes were made in the research that in part, or in whole invalidate the conclusions of the studies. Furthermore, it is troublesome that these issues were not addressed during the peer review process.

The EIC responded after several days promising to address each paper in a separate message. His response to the fluoride paper was the only course of action would be to submit a peer reviewed comment (providing this as an example) where the author would have a chance to respond, since the paper had been in print for nearly 2 years. I did not find this to be a particularly satisfying response since any publishable comment would need to be supported with new information. Essentially collecting the data and conducting control studies that should have been requested by peer reviewers. With the current state of knowledge, a potential published comment could be summarized as "I think you're wrong" and speculation, which is not any better than what was done in the paper. Data-free speculation does not not usually hold up to peer review, so reluctantly I published my criticism on pubpeer with the hope that anyone seeking to follow-up or use this chemistry would be wary of the authors' conclusions. This is the greatest risk in a case of erroneous research, which makes it similar to some issue encountered in cases of data falsification. Any time spent trying to use flawed science, wastes time and resources that could go toward more productive efforts. Coincidentally, I came across this paper today on fluoride sensing that invokes attack on a positively changed ring (albeit a different heterocycle), which was my initial instinct for an alternative mechanism in the disputed study. I'll have to compare the data and see if it gives me any new insight.

As for the iron paper, I never received the promised response. About 6 weeks after the initial inquiry, I wrote again asking for an update. After receiving nothing during the last +2 weeks, I am once again in the position of not having any particular recourse other than forgetting about it, or providing public post-publication peer review. Of the 2 papers, the iron sensor annoyed me more, because it repeats the same mistake made by another group that my group investigated and published as part of a larger study.

In a 2005 JACS article, Rurack claimed his Fe3+ responsive sensor also worked in aqueous solution. The only significant difference between the Rurack sensor and the new one is the receptor for the metal ion; however, both rely on the same PeT signal transduction mechanism where coordination to the aniline nitrogen atom is the key event. Several years after the Rurack paper was published, my group attempted to use the same receptor for a different application. We spent many months working on the system and eventually had to re-evaluate the Rurack data because our observations did not match what had been reported. We ultimately demonstrated that while Rurack's sensor did bind and respond to Fe3+ in organic solvents that lacked alcohol or carbonyl functional groups, the fluorescence signal in water was due to protonation of the aniline nitrogen atom, which mimics an Fe3+ response. We were fortunate to still be able to develop an interesting story even though we started our investigation with a flawed premise from a published paper (in JACS!). In a similar manner, the new Fe3+ sensor paper provides the opportunity to examine both pre- and post-publication peer review in this post.

The error the authors made in new paper is essentially the same  you absolutely cannot titrate Fe3+ into aqueous solution unless you use use a complexing ligand (e.g. citrate, which has a log K > 10 for Fe3+ and would therefore bind the metal ion more tightly than the sensor) or work at pH ≤ 3. In aqueous solution, Fe3+ rapidly hydrolyzes to Fe(OH)3 plus three equivalents of H+. The pKas for the 1st three deprotonation events are 2.2, 2.9 and ~6. This neglects multi-Fe processes that are also possible and have similarly acidic pKas. This is not an obscure fact, but textbook chemistry that would be covered in a standard undergraduate course. It's a fundamental concept in bioinorganic chemistry used to discuss the importance of siderophores in the acquisition of iron by microorganisms and the acquisition/transport/storage of iron by higher organisms using proteins.

The authors perform their fluorescence assays in 9:1 H2O-CH3CN. The water is not buffered and the authors do not report the pH at the beginning or the end of the fluorescence titration. A back of the envelope calculation shows that at ~150 equiv of Fe3+ added (~1000 μM Fe3+), the pH of the solution should reach ~3 (the approximate point at which Fe3+ hydrolysis no longer occurs and Fe3+ persists in solution). This is also the point at which the fluorescence response levels off for the sensor. The calculation is based on the use of deionized water without accounting for dissolved CO2, not that it would make a huge difference. CH3CN can act as a metal-binding ligand, but CH3CN does very little to stabilize Fe3+ in water. The authors go on to claim that reversibility of the fluorescence response with TPEN (tetrapyridylethylenediamine), a strong metal chelator, demonstrates that the fluorescence response is Fe3+-induced. This is also a flawed conclusion because the pKa of a protonated pyridine is about the same as a protonated aniline (both pKa ~ 5) and the aliphatic amines of TPEN are even more basic. Coordination (protonation) of the aniline nitrogen atom drives the PeT process responsible for the fluorescence response. TPEN can act as a base as well as a chelator, and the authors use a huge excesses of TPEN to demonstrate reversibility (60 equiv or 360 equiv of N bases). There may be some modest equilibrium between [Fe(Sensor)]3+ and [H(Sensor)]+ at pH<3, but the authors are observing a H+-, not an Fe3+-based fluorescence response in their titration. The argument is further supported by the receptor being used. Ethers are notoriously poor metal binding ligands in water. It would be very surprising if a cryptand ligand, even one with as many donor groups as the one in this sensor, could stable an Fe3+ complex with in water. The low affinity for the purported "[Fe(Sensor)]3+ complex" based on the fluorescence data supports this conclusion.

Returning to the Rurack report, the erroneous aqueous chemistry accounted for perhaps 25% of the discussion/conclusions in the paper. The rest of the Rurack paper is correctly interpreted (and an interesting counter-intuitive inorganic story). The flawed methodology accounts for 100% of the discussion/conclusions in the new paper. In my opinion this paper should be withdrawn; however, the EIC's lack of a response suggests that this isn't going to happen. I'd also like to contrast the response I got from the editor at Inorganic Chemistry to the one I got from ACS Applied Materials and Interfaces. Granted, I was trying to published a completed study in Inorg. Chem., but it should not be necessary to conduct months worth of work to prove something that only requires an understanding of basic inorganic chemistry. The Inorg. Chem. editor carefully handled the situation and facilitated a review of the conflicting data by the original author. I recall that I might have sent the EIC a note afterwards commending the editor who handled the situation. To his credit, Rurack completely supported the publication of our paper. The only unfortunate thing is that there is no indication anywhere that the original JACS paper contains a flawed set of experiments 
except in our paper. I still see this paper cited periodically as evidence that Fe3+ sensing can be done in aqueous solution. 

As an aside, our Inorg. Chem. paper was kind of an "end of the innocence" moment for me in scientific publishing. As a recent news story on pubpeer indicates, you can't always believe what you read in scientific journals. As someone who looks at sensor papers regularly, fluorescence might be the most misinterpreted spectroscopic assay used in chemistry. Fluorescence is an easy to execute and readily available technique, but there is a great tendency to interpret the results without acquiring additional supporting data (e.g. absorption spectroscopy, product analysis, etc.). As part of the Inorg. Chem. paper, my postdoc was trying to obtain a crystal structure of the Fe3+ complex of the ligand. After many failed attempts, he tried with Cu2+, and to our surprise got a structure containing only Cu1+. Another colleague in the fluorescent sensing field, pointed us in the direction of a synthetic paper where Cu2+ in CH3CN is used as a 1 electron oxidant (by forming [Cu(CH3CN)4]+). This was clearly what happened in our crystallization as we used CH3CN as the solvent. Despite our attempts to educate the fluorescent sensor community about the dangers of using Cu2+ in CH3CN, this is still a common practice. There are dozens of ring-opening spirolactam probes for Cu2+. These probes are usually based on rhodamine fluorophores, which have 2 embedded aniline groups, and give a terrific fluorescence response in CH3CN; however, everyone attributes the signal transduction mechanism to coordination/Lewis acidity of Cu2+, and ignores the possible contributions of redox chemistry.

There are clearly problems with peer review. In going through my significant backlog of TOC alerts today, I realized that RSC Advances moved to a 100 issues/year publishing schedule in 2014. This is a tremendous number of papers for a single journal, which requires a correspondingly huge number of peer reviewers. Inevitably, the same people who make mistakes in their research will be asked to review papers on similar science. This lets more errors slip through to publication and propagates errors in the literature. So what should editors do when they are alerted to a potentially serious problem? "Nothing" does not seem like an appropriate answer. Is pubpeer the the right pathway? It appears that some journals are very resistant to changing the status quo, and unless the pubpeer comments are linked to the articles, post-publication peer review may go unnoticed. It also remains to be seen if flawed protocols/conclusions will generate the kind of impassioned response that accompanies cases of scientific fraud; however, anyone who has wasted time with someone else's scientific mistakes surely has an opinion they will share without much prompting.

Update 3/22/15:
A note added in proof: the same group has a similar K+ sensor using a very similar receptor. The amides from the Fe3+ system are replaced by anilines, which would make metal interactions stronger. This sensor does not respond to Fe3+; however, the authors used buffered water, which means that protons from the are prevented from interacting with the aniline fluorescence switch. There is almost certainly an impact of having 3 "basic" aniline nitrogen atoms in the receptor as the amount of buffer (5 mM) is low compared to the added metal ions.

*The PI has since move to NJIT as a Dean, which would suggest the research group at UCF is no longer active. He is and was listed as a member of the editorial advisory board for ACS Applied Materials and Interfaces. While it is not uncommon and should not be problematic for board members to publish in the journal for which they serve, this may make some readers more suspicious about the impartiality of the peer review process.